University of Southern California Dissertations and Theses

The total synthesis of novel lipid mediators and their role in inflammation

(USC Thesis Other)

The total synthesis of novel lipid mediators and their role in inflammation

PDF

Download

Open document

Flip pages

Download a page range

Download transcript

Copy asset link

Request this asset

Transcript (if available)

Content i

THE TOTAL SYNTHESIS OF NOVEL LIPID MEDIATORS AND THEIR ROLE IN
INFLAMMATION

by

Jeremy Winkler

A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(CHEMISTRY)

December 2013

Copyright 2013 Jeremy Winkler

ii

Dedication

To both my parents and grandparents,
Mary and Dennis Winkler and Ray and Dolores Winkler

iii
Acknowledgments

First and foremost I would like to thank my parents for their tremendous support
along the way. The time and energy they spent on raising me in a good home with all the
necessary and important resources is unparalleled. I would not be where I am today if it
weren’t for the emphasis they placed on my education while exposing me to a wide range
of activities that helped peak my natural curiosity about the way the world works. Along
the lines of family I am very proud and fortunate that my grandparents were able to see
me finish my PhD. Their lives continue to be an inspiration to me each and every day.
Nicos Petasis has given me a dream PhD. I’ve learned along the way that nothing
ever goes exactly how one plans it, but when given a closer look fortune favors the bold
and often times victories and achievements aren’t always the obvious ones. Nicos is an
extremely brave PI constantly looking to take risks and chances. He cultivated a creative
spirit in our lab that allowed one to explore both their natural curiosity towards science as
well as to explore oneself. This has been a very unique and meaningful PhD, something
that I have grown from extremely both as a scientist and as a person.
In addition to Nicos’s mentorship a distinct lineage of inspiring science teachers
fortuitously steered me in the direction of science when my previous inclinations were
guided elsewhere. Ignacio Ocasio is most responsible for guiding me towards a path in
science. He proved to be one of the most inspiring people I have ever met, and after
taking his class in general chemistry I wouldn’t continue on this path for the next several
years. His energy and enthusiasm as well as genuine care for people translated
beautifully into his passion for science and this passion proved to be contagious. Others
iv
who inspired me at a young age included Susan Tozlowski and Caroline Maddocks both
of whom taught me science during my primary and secondary education.
Whilst at USC I have found amazing support along the way from Dr. Surya
Prakash, Robert Aniszfel, Matt Pratt and Travis Williams. They went out of their way to
help me providing invaluable guidance and counsel when I needed it the most regarding
my career, my research and my life goals. Also a big thanks to my two favorite Staff
members at the Loker Hydrocarbon Institute, David sand Carol, for brightening each and
every day.a
My colleagues have been fantastic including Kevin, Kalyan, Jasim and Min. As
well as Gosia, Ken Young, Jamie, Marcos, Dave, Anne Marie, Nikita and Steve. Outside
group members whom I’ve become very close friends with include Somesh Ganesh,
Sujith Chacko and Parag Jog. A special thanks to some good friends who I had an
opportunity to work with during their Undergraduate experience including Eric
Nagengast, Katie Henry and Michael ‘Cubby’ Carlson.
A big shout out to my brother and sister for all of their support and putting up
with me half distracted on the phone when I was preoccupied with something that wasn’t
going well in lab. I’d also like to thank some of my friends along the way including
Michael Prado, David Dennis, Justin Franklin, Tony Aguiar and Erin Kimmel, Jesse and
Chelsea Taylor, Michael Wilson, Jesse Carmichael, Heather Caye Brown, Kimberly
Gordon and Theresa Houston. Last but not least I can’t forget to thank my family away
from home, the Prados, for all their love and support.

v
Table of Contents
Dedication ................................................................................................................................ ii
Acknowledgments .................................................................................................................. iii
List of Figures ....................................................................................................................... vii
List of Schemes ..................................................................................................................... viii
Abstract .................................................................................................................................. ix
CHAPTER 1. Anti-Inflammatory Pro-Resolution Lipid Mediators Derived from
Docosahexaenoic Acid (DHA): ............................................................................................. 11
1.1 Introduction ................................................................................................................. 11
1.2 Diet and Disease .......................................................................................................... 11
1.3 Inflammation – Paradigm Shift ................................................................................... 14
1.4 Structure, Biosynthesis and Biological Role of DHA-Derived Resolvins .................. 16
1.4.1 Structures of D-Series Resolvins and their Aspirin Triggered Analogues .......... 16
1.4.2 Biosynthesis of DHA-derived Resolvins .............................................................. 17
1.4.2.1 Biosynthesis of 17S-series .............................................................................. 17
1.4.2.2 Biosynthesis of 17R-series ............................................................................. 18
1.4.3 Biological Activity of Resolvins D Series ............................................................ 19
1.5 Structue, Biosynthesis and Biological Role of DHA-Derived Protectins ................... 20
1.5.1 Structure of DHA-derived Protectins and their Aspirin Triggered Analogue .... 20
1.5.2 Biosynthesis of DHA-derived Protectin and Its Aspirin Triggered Analogue .... 21
1.5.2.1 Biosynthesis of 17R-series ............................................................................. 21
1.5.2.2 Biosynthesis of 17S-series .............................................................................. 22
1.5.3 Biological Activity of Protectin D Series ............................................................ 22
1.6 Conclusion ................................................................................................................... 23
1.7 References .................................................................................................................. 25
CHAPTER 2. Resolvin D2: Total Synthesis, Stereochemical Assignment, and Key
Intermediates.......................................................................................................................... 29
2.1 Introduction .................................................................................................................. 29
2.2 Results and Discussion ................................................................................................. 30
2.2.1 Synthesis of 17S RvD2 ........................................................................................... 30
2.2.1.1 Retrosynthetic Analysis.................................................................................. 30
2.2.1.2 Synthesis of Vinyl Iodide Intermediate (2.3) .................................................. 32
2.2.1.3 Synthesis of the Wittig Salt ............................................................................ 33
2.2.1.4 Synthesis of Terminal Alkyne ........................................................................ 34
2.2.1.5 Final Construction of 7S RvD2 ...................................................................... 35
2.3 BIOLOGY .................................................................................................................... 37
2.4 CONCLUSION ............................................................................................................ 39
2.5 EXPERIMENTAL........................................................................................................ 40
2.6 References .................................................................................................................... 57
CHAPTER 3. First Total Synthesis and Stereochemical Assignment of Resolvin D3 and
its Aspirin Triggered Analogue (RvD3 and AT-RvD3) ...................................................... 58
3.1 Introduction .................................................................................................................. 58
vi
3.2 Results and Discussion ................................................................................................. 59
3.2.1 Synthesis of RvD3 and AT-RvD3 ............................................................................ 59
3.2.1.1 Retrosynthetic Analysis.................................................................................. 59
3.2.1.2 Synthesis of Alkynyl Ester ............................................................................. 60
3.2.1.3 Synthesis of Vinyl Iodide 3.4 ......................................................................... 61
3.2.1.4 Synthesis of Vinyl Iodide (3.3) ...................................................................... 62
3.2.1.5 Final Construction of Resolvin D3................................................................. 63
3.3 Biological Actions ........................................................................................................ 65
3.4 Conclusion .................................................................................................................... 65
3.5 Experimental Procedure ............................................................................................... 66
3.6 References .................................................................................................................... 88
CHAPTER 4. First Total Synthesis and Stereochemical Assignment of Resolvin D4 and
its Aspirin Triggered Analogue (RvD4 and AT-RvD4) ...................................................... 90
4.1 Introduction .................................................................................................................. 90
4.2 Results and Discussion ................................................................................................. 92
4.2.1 Synthesis of RvD4 and AT-RvD4 ............................................................................ 92
4.2.1.1 Retrosynthetic Analysis.................................................................................. 92
4.2.1.2 Synthesis of Methyl Ester (4.5) ...................................................................... 94
4.2.1.3 Synthesis of Terminal Alkyne (4.4) ............................................................... 95
4.2.1.4 Synthesis of Propargyl Bromide (4.3) ............................................................ 96
4.2.1.5 Final Construction of RvD4 (4.1) .................................................................. 97
4.3 Conclusion .................................................................................................................... 99
4.4 Resolvin D4 Experimental.......................................................................................... 100
4.5 References .................................................................................................................. 111
CHAPTER 5. Total Synthesis and Stereochemical Assignment of Aspirin Triggered
Neuroprotectin D1 and Analogues. .................................................................................... 112
5.1 Introduction ................................................................................................................ 112
5.2 Results and Discussion ............................................................................................... 113
5.2.1 Synthesis of AT-NPD1 and Stereoisomers .......................................................... 113
5.2.1.1 Retrosynthetic Analysis................................................................................ 113
5.2.1.2 Synthesis of propargyl bromide ................................................................... 114
5.2.1.3 Synthesis of Vinyl Iodide Intermediate (5.5) ................................................ 115
5.2.1.4 Synthesis of Terminal Alkyne (5.4) and (5.6) .............................................. 117
5.2.1.5 Final Construction of AT-NPD1 Stereoisomers .......................................... 118
5.3 BIOLOGY .................................................................................................................. 120
5.4 CONCLUSION .......................................................................................................... 122
5.5 Experimental ............................................................................................................... 123
5.6 References .................................................................................................................. 142
BIBLIOGRAPHY ................................................................................................................ 144
APPENDIX.
1
H and
13
C Spectra of Substrates ................................................................ 151
vii

List of Figures
Figure 1. Adapted figure portraying correlation between diet and bioactivity. ............... 13
Figure 2. Acute Inflamation Overview. ........................................................................... 15
Figure 3. Structures of both D Series Resolvins and their Aspirin Triggered Counterparts.
.......................................................................................................................... 16
Figure 4. Biosynthesis of 17S Resolvin D-Series. ........................................................... 17
Figure 5. Structure of 17R Aspirin Triggered Protectin Analogues. ............................... 20
Figure 6. Biosynthesis of AT-Protectin D1/Neuroprotectin D1 (AT-PD1/NPD1). ......... 21
Figure 7. Biosynthetic pathway of Resolvin D2 and AT RvD2. ..................................... 30
Figure 8. Retrosynthetic Analysis of (7S, 16R, 17S) RvD2. ............................................ 31
Figure 9. 400 MHz
1
H-NMR and
1
H-
1
H COSY spectra including olefinic proton
assignments of RvD2 Methyl Ester (RvD2-Me). ............................................. 36
Figure 10. Resolvin D2 PMN reduction and Bioactivity Comparison of E-isomer. ....... 38
Figure 11. RvD2 reduces systemic bacterial burden in both the blood and peritoneum in
Caecal Ligation and Puncture Models (CLP). ............................................... 38
Figure 12. Biosynthesis of RvD3 and AT-RvD3. ............................................................ 58
Figure 13. Retrosynthetic Analysis of RvD3 and AT-RvD3. .......................................... 60
Figure 14. Exudate levels of D Series Resolvins post Zymosan Induced Response. ...... 65
Figure 15. Biosynthesis of RvD4 (4.1) and AT-RvD4. ................................................... 91
Figure 16. Retrosynthetic Analysis of RvD4 and AT-RvD4 ........................................... 93
Figure 17. Biosynthesis of NPD1 / PD1 and AT- NPD1 / PD1. ................................... 112
Figure 18. Retrosynthetic Analysis of (10R / 10S) AT-NPD1 and Isomers. ................. 113
viii
Figure 19. AT NPD1 Analogues Olefinic
1
H Spectra. .................................................. 120
Figure 20. A) Protectin HPLC Profile B) Reduction of PMN Infiltration in a TNF-α
induced peritonitis model. ........................................................................... 121
List of Schemes
Scheme 1. Synthesis of Key Vinyl Iodide Intermediate (2.3) .......................................... 33
Scheme 2. Synthesis of Wittig Intermediate (2.7) ........................................................... 33
Scheme 3. Synthesis of key terminal alkyne intermediate (2.4) ...................................... 34
Scheme 4. Final Coupling for 7S Resolvin D2. ............................................................... 35
Scheme 5. Synthesis of Key Alkynyl Ester Intermediate (3.5). ...................................... 61
Scheme 6. Total Synthesis of Vinyl Iodide Key Intermediate (3.4) ................................. 62
Scheme 7. Total Synthesis of Vinyl Iodide Intermediate (3.3). ....................................... 63
Scheme 8. Final Coupling of RvD3 Methyl Ester (3.27). ............................................... 64
Scheme 9. Synthesis of Methyl Ester Intermediate (4.5). ................................................ 95
Scheme 10. Total Synthesis of Terminal Alkyne (4.4). ................................................... 96
Scheme 11. Construction of Propargyl Bromide Intermediate (4.3). .............................. 96
Scheme 12. Final Coupling of Resolvin D4 (4.1). ........................................................... 98
Scheme 13. Synthesis of Propargyl Bromide Intermediate (5.11). ................................ 115
Scheme 14. Synthesis of Vinyl Iodide Intermediate (5.5 and 5.7). ................................ 116
Scheme 15. Synthesis of Acetylenic Omega Intermediates (5.19 and 5.22). ................. 117
Scheme 16. Final Coupling for AT-PD1 and 10-epi AT-PD1 as well as (13Z, 15E) AT-
PD1 and (13Z, 15E ) 10-epi AT-PD1. ......................................................... 118
Scheme 17. Final Coupling for (11E, 13E, 15E) AT-PD1 and (11E, 13E, 15E) 10-epi AT-
PD1. ............................................................................................................. 119
ix

Abstract

This body of work is about the design, synthesis and activity of a new and exciting class
of compounds made from Docosahexaenoic Acid (DHA) termed resolvins and protectins.
DHA is an abundant omega 3 fatty acid heavily concentrated in the brain and retina. The
reason these compounds have such importance is because they are derived from the
essential fatty acid, omega 3, that plays a crucial role in health. The aim of this work is to
design and synthesize mediators to elucidate and confirm the structures of these
specialized pro-resolving compounds with their synthetic counterparts and determine
their role in the regulation and expression of various cell types such as chemokines,
neutrophils and other enzymes involved in inflammation.
Due to the natural complexity of these bioactive chemical mediators, synthesis requires
great care to ensure absolute stereochemistry and regio-specificity. They are prone to
isomerization, decomposition and lactonization from light, heat and the presence of
transition metals. We therefore carefully utilized a number of mild and key reactions
from the organic chemist toolbox including the carbon-carbon bond Sonogashira
coupling, the Takai olefination and Wittig reactions to cater to their sensitive nature and
establish their olefinic region with complete regio-control. For absolute stereospecifity
naturally chiral precursors were used to assure all chiral hydroxyl groups were in the
correct position. This allowed for all synthetic molecules to be matched to their
biological counterparts with absolute certainty by state of the art LC MS-MS technology
as well as NMR and UV spectroscopy. Despite their seemingly similar structures each
x
mediator exhibits a completely different role in activity as well as a completely different
set of challenges synthetically. Furthermore charectization of each mediators shows a
distinct temporal profile, (via liquid chromatography), absorbance profile, (by UV-Vis
spectroscopy), and unique proton olefinic region by Nuclear Magenetic Resonance
spectroscopy.
Synthesis and characterization of a number of these unique mediators for the first time
provided new information regarding their complex role in regulation and control of a
hosts immune response during injury and infection. This work continues to receive a
good deal of attention for its impact in the field of controlling inflammatory related
diseases such as Alzheimer’s, Cardiovascular disease, asthma, and arthritis just to name a
few. In the future we will continue to report our efforts on the synthesis of novel lipid
mediators.
11
CHAPTER 1. Anti-Inflammatory Pro-Resolution Lipid Mediators Derived from
Docosahexaenoic Acid (DHA):
1.1 Introduction
Resolvin D series and Protectins are two recently discovered subsets of a growing class
of specialized pro-resolving lipid mediators derived from essential fatty acids whose
actions play a crucial role in a hosts immune response. Our group, in conjunction with a
number of collaborators across various fields have spent the last several years
investigating numerous classes of these compounds. In this work I will highlight our
efforts towards the design and total synthesis of two specific families of these autacoid
mediators, resolvins D series and protectins. With their successful completion we were
able to match them with their biological counterparts to better understand their role in
inflammation as well as gain insight into possible drug mimetics and targets for
treatments towards disease states related to chronic inflammation.
1.2 Diet and Disease
It was previously believed fats were not a seminal component of the human diet and were
just another source of energy. This notion was discredited in 1930 when research
published by Burr and coworkers demonstrated the debilitating effects a no fat diet had
on early development of young mice.
1
A number of cultural studies began to emerge
supporting more healthy cohorts on a pescetarian fat rich diet.
Fats and oils are comprised of various long chain fatty acids that serve as the basic
building blocks and an important fuel source in biological organisms. Their basic
12
structure consists of a hydrophilic carboxylic acid group on one end with a long fatty
aliphatic chain running off it. This chain can vary in length as well as have any number
of double bonds within its structure. There are many classes of fatty acids based on their
length and degree of unsaturation. Solid fats are comprised primarily of saturated fatty
acids which consist of no double bonds whereas oils contain primarily mono and poly-
unsaturated fatty acids, (PUFA’s), containing at least one double bond in the cis-
configuration. Since the human body produces plenty of saturated fat, it is recommended
that solid saturated fats be replaced with oils such as mono or poly unsaturated fatty
acids.
2

Essential fatty acids, (EFA’s), are contained within this class of compounds, and are
deemed essential due to their scarcity in the body. It is therefore essential that they are
obtained through diet in order for health and survival. The first EFA’s, were correctly
identified in 1929
3
and two primary classes were identified: the omega 6 and omega 3
fatty acids. This naming convention comes from the position of the first double bond
from the end terminal -CH3 of the aliphatic chain. The n-6 PUFA family can be
converted enzymatically to arachidonic acid, which can then be metabolized to a number
of different eicosanoids, such as leukotrienes and prostaglandins, metabolites that play a
complex role in inflammation. α-Linolenic acid or ALA is the parent fatty acid of the n-3
PUFA family and can be metabolized into eicosapentaenoic acid (EPA; 20:5, n-3) and
docosahexanoic acid (DHA; 22:6, n-3), which are then converted to anti-inflammatory
pro-resolving mediators such as resolvins
14
, protectins
14
and maresins.
4
This body of
work will focus primarily on mediators derived from DHA (Figure 1).

13

Figure 1. Adapted figure portraying correlation between diet and bioactivity.
5

Evidence suggests early man may have evolved consuming a diet favoring omega-3 over
omega-6 fatty acids.
6,7
Over time that ratio has shifted to a western diet favoring the
consumption of ω-6 fatty acids by an estimated 15:1.
8,9
Much has been reported on the
incidence of inflammatory disease related illness and diet across a cultural spectrum.
The most substantial reports relating omega-3 consumption to lower incidence of
cardiovascular disease include the Lyon Diet Heart Study,
10,11,12,13
the GISSI-Prevenzione
trial,
14
the DART study
15
and most recently a staggering report covered by the New York
Times of the first clinical trial measuring the Mediterranean diets impact on heart
disease.
16
A number of other health organizations including the Mayo Clinic, American
14
Health Association and the USDA have published health recommendations to increase
the ratio of ω-3 to ω-6 fatty acid consumption.
171819

1.3 Inflammation – Paradigm Shift
The recent discovery of anti-inflammatory pro-resolving mediators metabolized from
omega 3 fatty acids has given new insight into the elusive resolution process of acute
inflammation. A host’s immune response is a crucial and important process in regulating
homeostasis. It is known that inflammatory pathways initiate a number of these common
and difficult to treat diseases. However there is still a great amount of work to determine
the pathogenesis of such diseases specifically in what gives rise to chronic inflammation
and how the anti-inflammatory pathways are affected. Inflammation begins, (Figure 2),
20

with the production of lipid mediators such as the powerful chemo-attractant, leukotriene
B4, a derivative of arachidonic acid, that recruits neutrophils, (PMNs), to the site of
inflammation. PMNs can engulf microorganisms and cellular debris harmful to the host
through a process known as phagocytosis. Granules and reactive oxygen species, (ROS),
within the PMN can kill and degrade foreign debris however in some cases neutrophil
granules spill into the extra-cellular milieu before phagocytosis is complete. In this case
acute inflammation is amplified as additional inflammatory mediators recruit more
neutrophils to the site of inflammation and in time chronic inflammation ensues. For the
ideal programmed fate of acute inflammation, resolution was previously believed to be a
passive process where signals would slowly burnout.
21
However with the recent
discovery of mediators such as resolvins, protectins and maresins it was found that the
biosynthesis of these signaling mediators at the site of inflammation could compete with
15

Figure 2. Acute Inflamation Overview.
20

the production of their inflammatory counterparts such as prostaglandins and leukotrienes.
Therefore they serve to reduce neutrophil infiltration and further recruit monocytes that
can then form macrophages to devour and remove apoptotic cells.
13
It is within these
cascades and competitive processes we see a paradigm shift in the way we think of
resolution of inflammation as an active process. Herein lies the crux of our efforts to
understand and identify these specialized pro-resolution metabolites to ultimately aid in
the treatment of inflammatory related disease states.
16
1.4 Structure, Biosynthesis and Biological Role of DHA-Derived Resolvins
1.4.1 Structures of D-Series Resolvins and their Aspirin Triggered Analogues
In our efforts to confirm the complete stereochemical assignment of all resolvin D series
we synthesized both the endogenous and aspirin triggered 17S and 17R resolvins (Figure
3). To confirm the structure of the endogenous compounds with complete certainty the
regio and stereo-selectivity must be controlled. Therefore chirally pure starting materials
where utilized to establish the chiral hydroxyl stereocenters. As for the complex tetraene,
triene and diene conjugated double bond moieties we carefully reduced them in the later
steps from their alkyne precursors to avoid any reconfiguration through isomerization.
22

In addition a number of highly efficient carbon-carbon bond coupling reactions were
utilized to build the long allyic chains. RvD3 and RvD4 proved to be particularly
challenging due to the nature of the γ-hydroxy positioned so closely to the ester moiety.
As it pertained to RvD3, the γ-hydroxy was prone to forming the five membered lactone
during deportection and purification. This also presented a problem during the final
reduction of the bis-acetylenic precursor due to the labile nature of the carbinol proton
leaving the possibility of an intramolecular nucleophilic attack of the alkoxide ion to
form the five-membered lactone ring.
23
These factors made the compound uniquely

Figure 3. Structures of both D Series Resolvins and their Aspirin Triggered Counterparts.
17
challenging and especially susceptible to degradation in the presence of mild
nucleophiles. RvD4 was also met with similar challenges.
1.4.2 Biosynthesis of DHA-derived Resolvins
1.4.2.1 Biosynthesis of 17S-series
DHA is converted to resolvins through one of two primary enzymatic pathways. The
first pathway forms the oxygenation product on the 17-carbon position in the S
arrangement as opposed to the alternative pathway which forms the 17R arrangement.
Biosynthesis of the endogenous 17S series begins with the enzymatic oxidation of DHA
by 15-Lipoxygenase to form the 17S-HpDHA peroxide. Peroxidase next converts this
peroxide, 17S-HpDHA, to the 17S-hydroxy-DHA alcohol (Figure 4). Sequential

Figure 4. Biosynthesis of 17S Resolvin D-Series.
14
18
oxygenation by 5-Lipoxygenase can occur at the C4 or the C7 position to form the
respective peroxide. These two peroxides can be reduced once again by peroxidase to
form the diols, RvD5 and RvD6 respectively. Our interest focused on the more potent
and structurally complex resolvins, RvD1-RvD4. These metabolites are presumably
formed by the transformation of the peroxide precursors into the 7S, 8S-Epoxide or the 4S,
5S-Epoxide. The conjugated double bond system left open the possibility of different
routes of formation. Evidence supporting the mechanism in which an epoxide
intermediate forms the conjugated system was confirmed through the use of LC-MS/MS
analysis by the Serhan group. Using 17S-hydroxy(peroxy) intermediate from DHA and
incubating in the presence of human PMNs and acidic methanol the epoxide
intermediates were successfully trapped giving strong evidence to support the claim of an
epoxide intermediate. Enzymatic hydrolysis through a hydrolase enzyme can occur
either from the C8 or the C16 position. Hydrolysis at the C8 position affords the C7, C8
vicinal diol, 7S, 8R, 17S RvD1. Hydrolysis from the other end at the C16 position affords
the C16, C17 vicinal diol, 7S, 16R, 17S RvD2. Regarding the formation of RvD3 and
RvD4 enzymatic hydrolysis of the 4,5-Epoxide, hydrolysis can occur at either the C11
end or the C5 position. Hydrolysis from the C11 position affords the 4S, 11R, 17S RvD3
and hydrolysis from the C5 position provides the C4, C5 vicinal diol, 4S, 5R, 17S RvD4.
1.4.2.2 Biosynthesis of 17R-series
Biosynthesis of these metabolites in the presence of aspirin assumes a completely
different route. Aspirin acetylates a serine residue in the active site of cyclooxygenase
enzymes, (COX), irreversibly blocking cyclooxygenase 1, COX1, however only partially
blocking cyclooxygenase 2, COX2. The partially obstructed COX 2 active site, normally
19
responsible for the production of inflammatory mediators such as thromboxanes and
prostaglandins, is no longer capable of production of the aforementioned prostanoids
while still allowing the conversion of DHA to various pro-resolution metabolites. Under
normal conditions COX2 converts DHA into a 13-hydro(peroxy) product however upon
being acetylated it converts DHA to the 17R position reversing the stereochemistry at the
17 carbon.
24
This initial oxygenation can occur through the acetylated COX-2 as well
Cytochrome P450 pathway. The rest of the biosynthesis follows in suit with the
previously mentioned non-aspirin triggered enzymatic reactions.
1.4.3 Biological Activity of Resolvins D Series
RvD1 and RvD2 have both been studied extensively over the past four years. With the
complete stereo-chemical assignment of resolvin D1 and D2 confirmed, it was shown
that both chemical mediators are potent pro-resolving compounds in adipose tissue and
thus have an effect on obesity.
25
RvD1 and D2 have also been shown to attenuate
histamine release in human lung mast cells, HLMC, by way of Airway epithelial cells,
(AEC), through a G0/Gi receptor mechanism.
26
This places them at an integral role in the
resolution of inflammation found in asthma patients. Resolvins including RvD1 and
RvD2 act as some of the most powerful endogenous inhibitors of TRPV1/TRPA1
known.
27
They have been shown to reduce inflammatory pain related to arthritis by
inhibiting these TRP channels. AT-RvD1 and RvD2 also were shown to “improve
disease activity index in multiple colitis experimental models” associated with
inflammatory bowl disease (IBD). More recently work has uncovered a unique role in
which RvD2 reduces systemic bacterial burden in sepsis that will be highlighted in
greater detail in the following chapter. The role such mediators play in asthma, obesity,
20
arthritis, irritable bowel syndrome, (IBD), and sepsis is compelling and could be useful in
both curative and preventative therapies to treat inflammatory related disease. In our
limited capacity some biological activity of all resolvins D3 and D4 has been confirmed
but cannot be completely explored until the structure is matched and these molecules are
produced in larger quantities. Due to their recent synthesis, there is still much work to be
done to elucidate their role in the body.

1.5 Structue, Biosynthesis and Biological Role of DHA-Derived Protectins
1.5.1 Structure of DHA-derived Protectins and their Aspirin Triggered Analogue
To fully characterize and probe biological activity of neuroprotectin we synthesized a
number of aspirin triggered epimers (Figure 5). It was our aim to improve upon previous
methods while making the synthesis more efficient. We also focused on the challenge of
synthesizing a number of these metabolites while carefully constructing three different
triene backbone configurations.

Figure 5. Structure of 17R Aspirin Triggered Protectin Analogues.
21
1.5.2 Biosynthesis of DHA-derived Protectin and Its Aspirin Triggered Analogue
1.5.2.1 Biosynthesis of 17R-series
The proposed biosynthesis of the aspirin triggered protectin/neuroprotectin, (PD1/NPD1),
begins with the same oxygenation pathway of DHA by the acetylated COX-2 or
Cytochrome P450 pathway as that of its resolvin counterparts at the 17 position however
in the (R) position (Figure 6). It can then be subsequently reduced to 17R-HDHA by
peroxidase to form 17R-HDHA. 17R-HDHA is then oxygenated by 5- LO and reduced
by peroxidase at the C10 position to afford the diol, 10S, 17R-diHDHA. On the other
hand the 17R-HpDHA peroxide can lose a water to afford the 16S, 17R epoxide
intermediate that is hydrolyzed to form the cis, trans, trans triene backbone. This
conjugated backbone is a critical element to the structure as well as the bioactivity of the
potent 10R, 17S- Protectin D1 (PD1/NPD1) lipid mediator.

Figure 6. Biosynthesis of AT-Protectin D1/Neuroprotectin D1 (AT-PD1/NPD1).
28

22
1.5.2.2 Biosynthesis of 17S-series
The biosynthesis of non-aspirin triggered protectin follows the same initial pathway as
resolvins forming the 17S HpDHA intermediate through 15-LO oxygenation.
Subsequent enzymatic transformations similar to the aspirin triggered protectin affords
the protectin/neuroprotectin metabolites that were reported by Rong et al.
1.5.3 Biological Activity of Protectin D Series
The biological role of Protectin/Neuroprotectin PD1/NPD1 and its aspirin triggered
analogues have been well established since its initial discovery in 2004 and yet new
findings are being published every month.
29
In the broadest sense PD1 shortens
inflammation by activating resolution programmes in experimental inflammation.
30

Additionally NPD1 was found to be present in breath condensates among asthmatics.
31
It
has also been shown to reverse damage of ischemia-reperfusion in the kidney. NPD1
also plays a large role in inflammatory pathways in neural systems such as Alzheimers,

32 , 33
Age-related macular degeneration, AMD,
34 , 35
and stroke.
36
Numerous NPD1
analogues both aspirin triggered and non-aspirin triggered were synthesized to determine
rank of potency. NPD1 maintained the highest level of potency amongst all of these
metabolites.
96
Although some metabolites did show activity for example 10S, 17S-
diHDHA demonstrated activity reducing platelet aggregation as well as being active in
peritonitis.
37
These are just a few of the actions PD1 plays in the body and in the future
many more actions will be uncovered.

23
1.6 Conclusion
In conclusion we can get a sense of how valuable these lipid mediators play in our bodies
innate immune response. There have been many reports linking a diet rich in omega 3s
to reduced risk of inflammatory related diseases. Diet rich in such fatty acid precursors,
DHA and EPA, provides these powerfully potent small molecules to the host defense
system that we report the synthesis of herein. These fatty acids are unique in that they
appear to have a baseline level in circulation and this has been observed in edema.
38,39

Therefore they are utilized in a much faster manner than the metabolism of arachidonic
acid which provides pro-inflammatory metabolites such as prostaglandins, thromboxanes
and leukotrienes. This is a paradigm shift from the once believed notion that
inflammation tapers of over time. Once on sight DHA and EPA can be rather quickly
metabolized into resolvins, protectins and or the recently discovered maresins.
40
These
compounds effectively traffic effector cells such as neutrophils and macrophages both of
which are first in line in inflammation and last in line in clearing inflammation
respectively. Excessive accumulation of these cells can advance acute inflammation into
a chronic state that ultimately leads to pathogenesis. With metabolites that operate on a
nanomolar and picomolar scale, homeostasis can quickly return to the host. It is worth
mentioning that these biochemical mediators are primordial in nature in that their
structure is highly conserved from fish, such as the rainbow trout, to humans,
41 , 42

providing the possibility that there is much more they are involved in within the body.
Aspirin also shows unique abilities as a non-steroidal anti-inflammatory drug that can
jump start resolution by shutting down pro-inflammatory pathways and therefore enhance
anti-inflammatory pathways by the biosynthesis of pro-resolving mediators. Herein we
24
report the stereochemical confirmation and first successful synthesis of a number of such
metabolites. These molecules will give insight into new potential therapeutic targets in
inflammation pathways linked with a number of deadly diseases.

25
1.7 References

1. Burr, G. O.; Burr, M. M. Journal of Biological Chemistry 1929, 82, 345.
2. Krauss, R. M.; Eckel, R. H.; Howard, B.; Appel, L. J.; Daniels, S. R.; Deckelbaum, R.
J.; Erdman, J. W.; Kris-Etherton, P.; Goldberg, I. J.; Kotchen, T. A.; Lichtenstein, A.
H.; Mitch, W. E.; Mullis, R.; Robinson, K.; Wylie-Rosett, J.; Jeor, S. S.; Suttie, J.;
Tribble, D. L.; Bazzarre, T. L. Circulation 2000, 102, 2284.
3. Burr, G. O.; Burr, M. M. Journal of Biological Chemistry 1930, 86, 0587.
4. Vance, D.E.; Vance, J.E. Biochemistry of lipids, lipoproteins, and membranes. 2008,
(5th ed.) Amsterdam, Elsevier.
5. Niemoller, T. D.; Bazan, N. G. Prostaglandins & Other Lipid Mediators 2010, 91, 85.
6. Simopoulos, A. P. American Journal of Clinical Nutrition 1991, 54, 438.
7. Weber, P.C.; Fish Fats and Your Health, 1989, Report No. 4, 9-18.
8 . Bazan, N. G.; Molina, M. F.; Gordon, W. C. Annual Review of Nutrition, Vol
31, 2011, 31, 321.
9. Nordoy, A.; Goodnight, S. Arteriosclerosis. 1990, 10, 149-163.
10 . de Lorgeril, M.; Salen, P.; Martin, J. L.; Monjaud, I.; Delaye, J.; Mamelle,
N. Circulation 1999, 99, 779.
11. de Lorgeril, M.; Renaud, S.; Mamelle, N.; Salen, P.; Martin, J. L.; Monjaud, I.;
Guidollet, J.; Touboul, P.; Delaye, J. Lancet 1994, 343, 1454.
12. Trichopoulou, A.; Costacou, T.; Bamia, C.; Trichopoulos, D. New England Journal
of Medicine 2003, 348, 2599.
13 . Kris-Etherton, P.; Eckel, R. H.; Howard, B. V.; St Jeor, S.; Bazzarre, T.
L. Circulation 2001, 103, 1823.
26

14. Valagussa, F.; Franzosi, M. G.; Geraci, E.; Mininni, N.; Nicolosi, G. L.; Santini, M.;
Tavazzi, L.; Vecchio, C.; Marchioli, R.; Bomba, E.; Chieffo, C.; Maggioni, A. P.;
Schweiger, C.; Tognoni, G.; Barzi, F.; Flamminio, A. V.; Marfisi, R. M.; Olivieri, M.;
Pera, C.; Polidoro, A.; Santoro, E.; Zama, R.; Pagliaro, L.; Correale, E.; Del Favero,
A.; Loi, U.; Marubini, E.; Campolo, L.; Casari, A.; Di Minno, G.; Donati, M. B.; Galli,
M.; Gattone, M.; Garattini, S.; Mancini, M.; Marino, P.; Santoro, G. M.; Scardulla, C.;
Specchia, G.; Cericola, A.; Di Gregorio, D.; Di Mascio, R.; Levantesi, G.; Mantini, L.;
Mastrogiuseppe, G.; Tucci, C.; Mocarelli, P.; Baldinelli, R.; Ceriotti, F.; Colonna, A.;
Cortese, C.; Fortunato, G.; Franzini, C.; Gonano, F.; Graziani, M. S. Lancet, 1999, 354,
447.
15. Burr, M. L.; Gilbert, J. F.; Holliday, R. M.; Elwood, P. C.; Fehily, A. M.; Rogers, S.;
Sweetnam, P. M.; Deadman, N. M. Lancet 1989, 2, 757.
16. Estruch, R.; Ros, E.; Salas-Salvado, J.; Covas, M.-I.; Corella, D.; Aros, F.; Gomez-
Gracia, E.; Ruiz-Gutierrez, V.; Fiol, M.; Lapetra, J.; Maria Lamuela-Raventos, R.;
Serra-Majem, L.; Pinto, X.; Basora, J.; Angel Munoz, M.; Sorli, J. V.; Alfredo
Martinez, J.; Angel Martinez-Gonzalez, M.; Investigators, P. S. New England Journal
of Medicine 2013, 368, 1279.
17. Ottawa: Minister of National Health and Welfare, Nutrition Recommendations. 1990,
H-49-42, 1990E.
18. U.S. Department of Agriculture and U.S. Department of Health and Human Services.
Dietary Guidelines for Americans, 2010. 7th Edition, Washington, DC: U.S.
Government Printing Office, Dec. 2010.
27

19. Lichtenstein, A. H.; Appel, L. J.; Brands, M.; Carnethon, M.; Daniels, S.; Franch, H.
A.; Franklin, B.; Kris-Etherton, P.; Harris, W. S.; Howard, B.; Karanja, N.; Lefevre,
M.; Rudel, L.; Sacks, F.; Van Horn, L.; Winston, M.; Wylie-Rosett,
J. Circulation 2006, 114, 82.
20. Serhan, C. N.; Chiang, N.; Van Dyke, T. E. Nature Reviews Immunology 2008, 8,
349.
21. Serhan, C. N.; Petasis, N. A. Chemical Reviews 2011, 111, 5922.
22. Schroer, N., Sieler, C., Bottmuhle, A. Der, & Feigel, M. Helv. Chim. Acta, 1987, 70,
1025–1040.
23. Bundgaard, H.; Falch, E.; Larsen, C.; Mosher, G. L.; Mikkelson, T. J. Journal of
Medicinal Chemistry 1985, 28, 979.
24. Serhan, C. N. Prostaglandins Leukotrienes and Essential Fatty Acids 2005, 73, 141.
25 Claria, J.; Dalli, J.; Yacoubian, S.; Gao, F.; Serhan, C. N. Journal of
Immunology 2012, 189, 2597.
26. Martin, N.; Ruddick, A.; Arthur, G. K.; Wan, H.; Woodman, L.; Brightling, C. E.;
Jones, D. J. L.; Pavord, I. D.; Bradding, P. Plos One 2012, 7.
27 . Park, C.-K.; Xu, Z.-Z.; Liu, T.; Lu, N.; Serhan, C. N.; Ji, R.-R. Journal of
Neuroscience 2011, 31, 18433.
28. Serhan, C. N. Annual Review of Immunology 2007, 25, 101.
29. Mukherjee, P. K.; Marcheselli, V. L.; Serhan, C. N.; Bazan, N. G. Proceedings of the
National Academy of Sciences of the United States of America 2004, 101, 8491.
30. Schwab, J. M.; Chiang, N.; Arita, M.; Serhan, C. N. Nature 2007, 447, 869.
28

31. Levy, B. D.; Kohli, P.; Gotlinger, K.; Haworth, O.; Hong, S.; Kazani, S.; Israel, E.;
Haley, K. J.; Serhan, C. N. Journal of Immunology 2007, 178, 496.
32. Lukiw, W. J.; Cui, J. G.; Marcheselli, V. L.; Bodker, M.; Botkjaer, A.; Gotlinger, K.;
Serhan, C. N.; Bazan, N. G. Journal of Clinical Investigation 2005, 115, 2774.
33. Stark, D. T.; Bazan, N. G. Molecular Neurobiology 2011, 43, 131.
34. Bazan, N. G. Brain Pathology 2005, 15, 159.
35. Antony, R.; Lukiw, W. J.; Bazan, N. G. Journal of Biological Chemistry 2010, 285,
18301.
36. Belayev, L.; Khoutorova, L.; Atkins, K. D.; Eady, T. N.; Hong, S.; Lu, Y.; Obenaus,
A.; Bazan, N. G. Translational Stroke Research 2011, 2, 33.
37. Chen, P.; Fenet, B.; Michaud, S.; Tomczyk, N.; Vericel, E.; Lagarde, M.; Guichardant,
M. Febs Letters 2009, 583, 3478.
38. Bannenberg, G. L.; Chiang, N.; Ariel, A.; Arita, M.; Tjonahen, E.; Gotlinger, K. H.;
Hong, S.; Serhan, C. N. Journal of Immunology 2005, 174, 4345.
39. Chiang, N.; Schwab, J. M.; Fredman, G.; Kasuga, K.; Gelman, S.; Serhan, C. N. Plos
One 2008, 3.
40. Serhan, C. N.; Yang, R.; Martinod, K.; Kasuga, K.; Pillai, P. S.; Porter, T. F.; Oh, S.
F.; Spite, M. Journal of Experimental Medicine 2009, 206, 15.
41. Rowley, A. F.; Lloydevans, P.; Barrow, S. E.; Serhan, C. N. Biochemistry 1994, 33,
856.
42 . Hong, S.; Tjonahen, E.; Morgan, E. L.; Lu, Y.; Serhan, C. N.; Rowley, A.
F. Prostaglandins & Other Lipid Mediators 2005, 78, 107.

29

CHAPTER 2. Resolvin D2: Total Synthesis, Stereochemical Assignment, and Key
Intermediates.
2.1 Introduction
Through the investigation of inflammatory pathways and the role of inflammation on
disease states, a novel family of DHA derivied metabolites termed resolvins D series
were discovered as potent anti-inflammatory pro-resolution mediators. Since the
inception of their discovery it has been our focus to better understand these autacoids and
their behavior as active regulators in the process of resolving inflammation in the body
through total synthesis and biological matching. Resolvin D2 was initially identified
among these resolution phase exudates however its specific role remained unclear.
20
In
addition to its biological ambiguity the exact stereochemistry of this lipid metabolite was
unknown more specifically the geometry of its tetraene backbone as well as its
stereochemistry at its three chiral centers. Knowing based on its sister compounds and
their potency in resolving inflammation we set out to synthesize Resolvin D2 to learn its
role as a potent regulator of inflammation pathways. Figure 1 illustrates the biosynthesis
in which DHA is converted towards both resolvin D2 and its aspirin triggered Resolvin
D2 epimer through polymorphonucleocytes, (PMN), and eosinophil transcellular
30

Figure 7. Biosynthetic pathway of Resolvin D2 and AT RvD2.
13

biosynthesis.
43
Due to the sensitive nature of these metabolites it was of crucial
importance to chemically synthesize these compounds from chirally pure starting
materials and in a completely stereo-controlled manner for biological confirmation.
Through our efforts its presence in biological systems was confirmed both in human
tissue
44
as well as salmon.
45

2.2 Results and Discussion
2.2.1 Synthesis of 17S RvD2
2.2.1.1 Retrosynthetic Analysis
Resolvin D2 contained two distinct structural components similar to its resolvin D1
46

counterpart that we incorporated into our synthetic strategy to build this compound. The
C4 cis-olefin and the tetraene backbone both are key features of the resolvin D1 synthesis
previously reported by our lab. During the synthesis we relied on natural chiral
precursors (2.5 and 2.8) to make key building blocks (2.3 and 2.4) with the C7, C16 and
C17 chiral hydroxyl groups fixed in the appropriate configuration. To begin our
retrosynthetic strategy, (Figure 8), we decided to conserve the nature of the C10 cis
double bond compromising part of the tetraene backbone by reducing it at the final step
31

Figure 8. Retrosynthetic Analysis of (7S, 16R, 17S) RvD2.

with a mild Zn/Cu/Ag reducing agent that we have adapted to the synthesis for a number
of similarly delicate metabolites. To form this tetraene backbone we carried out a highly
efficient Pd
0
/Cu
1
sonogashira coupling with alkyne (2.4) and vinyl iodide (2.3). Vinyl
iodide (2.3) was synthesized by the ring opening of chrial glycidol (2.5) with the OBO
protected pentynoic ester (2.6). Through a number of deprotections and protections the
primary alcohol was isolated subsequently allowing for conversion to the vinyl iodide
(2.3). The construction of terminal alkyne (2.4) utilized the wittig coupling of propyl
wittig ylide (2.9) with the masked aldehyde and chirally pure sugar (2.8) to install C16
and C17 hydroxyl groups as well as afford the exclusive C4 Z-alkene product. Oxidation
of the primary alcohol set up a second wittig coupling with ylide (2.7) that afforded the E,
E protected alkyne which could be desilyated to afford terminal alkyne (2.4). With this
in hand we were able to carry out an efficient coupling, deprotection and reduction to
32
afford 7S, 16R, 17S Resolvin D2 in appreciable yields and match it successfully with its
endogenous counterpart. An alternative total synthesis of Resolvin D2 was also reported.
2.2.1.2 Synthesis of Vinyl Iodide Intermediate (2.3)
To build the vinyl iodide methyl ester (2.3) we needed to install the C7 hydroxyl group in
the S position using the chirally pure precursor R-gylcidol (Scheme 1). Due to the nature
of this base catalyzed coupling, the ester was susceptible to degradation and therefore had
to be protected using Corey’s OBO ortho-ester protecting group.
47,48
To start this piece
we began by coupling pentynoic acid (2.10) with oxetane (2.11) using DCC and DMAP
to afford the oxetane product (2.6). The oxetane ring was rearranged into the OBO ester
in the by coupling pentynoic acid (2.10) with oxetane (2.11) using DCC and DMAP to
afford the oxetane product (2.6). The oxetane ring was rearranged into the OBO ester in
thepresence of BF3•OEt2 and Et3N. With the base stable OBO ester in place R-glycidol
was coupled to the alkyne in the presence of n-BuLi to afford the crude ester which was
hydrolysed without purification in the presence of HCl to afford the protected diol (2.13).
The diol (2.13) was then converted to the methyl ester by hydrolyzing with lithium
hydroxide and then reacted with diazomethane. The free secondary alcohol was
protected with TBDPS using standard protection conditions and the primary alcohol was
deprotected at 0°C with camphor sulfonic acid to afford the hydroxyl ester (2.14). Using
Lindlar catalyst and quinolone the internal alkyne was reduced establishing the cis-olefin
at the C4 position. The primary alcohol group was converted to the aldehyde (2.15)
using the mild Swern-conditions. Finally the aldehyde was converted to the vinyl halide
(2.3) through takai olefination in the presence of CrCl2 and CHI3.
49

33

Scheme 1. Synthesis of Key Vinyl Iodide Intermediate (2.3)
2.2.1.3 Synthesis of the Wittig Salt
Next we focused our efforts on the synthesis of the conjugated tetraene backbone by
building the Wittig Intermediate (Scheme 2). We began by the silyation of alkyne (2.16)
with TMS-Cl in the presence of base to afford the protected alkyne (2.17). Bromination
of the primary alcohol using NBS and PPh3 afforded the allylic bromide (2.18) in good
yield which was then converted to the Wittig salt (2.7) by heating in the presence of tri-
phenyl phosphine in toluene overnight. With the Wittig salt in hand we next focused on
building its aldehyde coupling partner to establish the 16 and 17 hydroxyl positions.

Scheme 2. Synthesis of Wittig Intermediate (2.7)
34
2.2.1.4 Synthesis of Terminal Alkyne
To assemble the terminal omega piece shown in Scheme 3, deoxy sugar (2.8) was
coupled with the propyl wittig salt (2.9) in the presence of base to give the cis-olefin
product (2.19).
50
By carrying out this reaction in our first step from the pyranose
precursor we were able to set the stereochemistry for the 16R and 17S chiral hydroxyl
positions for the final molecule. Oxidation of the free alcohol using Dess-Martin
conditions afforded the aldehyde (2.20). Our next step was to establish the conjugated
backbone. We accomplished this by coupling a second bromo phosphonate Wittig salt
with our aldehyde to afford a cis-transmixture of the omega dihydroxyl diene (2.21). The
acetal group was removed from protecting the diol in the presence of 1M HCl and
protected with TBS-OTf in the presence of 2,6-lutidine to afford (2.22). Next the cis-

Scheme 3. Synthesis of key terminal alkyne intermediate (2.4)

35
trans mixture was isomerized utilizing I2 and light to afford the all trans compound.
7

Lastly to prepare the terminal alkyne the tri-methyl silyl protecting group was removed in
the presence of sodium carbonate to afford the E, E terminal alkyne (2.4).
2.2.1.5 Final Construction of 7S RvD2
With our two key building blocks in place, (2.3 and 2.4), we carried out the formation of
the final carbon-carbon bond coupling of resolvin D2 using previously reported Pd
0
/Cu
1

sonogashira conditions. In Scheme 4 we show the coupling step to form the C10-C11
acetylenic protected precursor (2.23). We carried out this sonogashira coupling in
excellent yield. The subsequent TBAF deprotection and final reduction of the acetylenic
triol were both also accomplished in high yields allowing us to procure enough material
to fully characterize resolvin D2 as well as confirm its stereochemistry by matching it to
its endogenous counterpart and probe its biological role in inflammation.
51

Scheme 4. Final Coupling for 7S Resolvin D2.

36
H-9 H-12 H-14 H-13
H-11 H-10
H-15 H-8
H-20
H-4
H-5
H-19
In Figure 9 we see the olefinic region of the NMR spectra of resolvin D2 as well as a
1
H-
1
H 2-D COSY experiment allowing us to fully characterize all protons in the lipid
mediator as well as give us some insight into its structural backbone.

Figure 9. 400 MHz
1
H-NMR and
1
H-
1
H COSY spectra including olefinic proton assignments of
RvD2 Methyl Ester (RvD2-Me).

37
2.3 BIOLOGY
Through the discovery and structural confirmation of resolvin D2, it was discovered that
this pro-resolution metabolite plays a multi-level role in reducing neutrophil trafficking
while also increasing macrophage phagocytosis and peritoneal mononuclear cells. RvD2
reduced polymorpho nucleocyte inflatration up to 70% in microbial peritonitis at doses as
low as 10 picograms (Figure 10A). The geometrical structure of the tetraene backbone is
crucial for bioactivity as the 10E regioisomer showed no biological activity (Figure 10B).
Resolvin D2 did not solely alter PMN adhesion directly but also by stimulating rapid
NOx production. The production of NOx by resolvin D2 however was not at levels
considered to be pro-inflammatory such as those exerted by LTB4. For the first time, this
resolvin autacoid reduced both local and systemic bacterial infection in the blood and
peritoneum through reduction of leukocytes and PMN infiltration into the peritoneum as
well as reducing levels of a number of pro-inflammatory cytokines including IL-17 and
IL-10 both detrimental to survival in sepsis. Despite not having direct antimicrobial
properties similar to ampicillin, resolvin D2 was extremely effective in clearing bacteria
during microbial sepsis through promoting both macrophage and PMN directed
38

Figure 10. Resolvin D2 PMN reduction and Bioactivity Comparison of E-isomer.
9

phagocytosis.
32
This offers unique insight into novel treatments in fighting microbial
sepsis, (Figure 11), whose previous shortcomings typically cause sustained
immunosuppression and danger to the host.
52

Figure 11. RvD2 reduces systemic bacterial burden in both the blood and peritoneum in Caecal
Ligation and Puncture Models (CLP).
9

39
2.4 CONCLUSION
In conclusion the total synthesis of the natural Resolvin D2 isomer was accomplished.
Through this chirally controlled strategy the absolute stereochemistry was confirmed
through physically matching the endogenous compound with its synthetic counterpart by
UV-absorbance, retention time, and tandem mass spec derivatization as well. Matching
served as an extremely crucial role in elucidating its biological activity. Through our
efforts and since its confirmation it has demonstrated pro-resolving actions in colitis
53
as
well as reducing pain.
54
Our efforts utilized a concise and convergent strategy allowing us
to use building blocks for multiple metabolites. In our future work regarding resolvin D2
we will report the concise synthesis and full characterization of its aspirin triggered
analogue as well as biological activity in the body. These efforts could help develop new
approaches to inflammatory related diseases with possible resolvin-like drug mimetics.

40
2.5 EXPERIMENTAL

Unless otherwise noted, all reactions were carried out in a flame-dried flask with stir bar
under argon routed through a three-necked valve. Reactions were carried out at room
temp using DriSolv solvents purchased commercially from VWR. All reagents used
were purchased without further purification from Sigma Aldrich, Strem, and Alfa Aesar.

Progress was monitored and recorded using EMD analytical thin layer chromatography
plates, Silica Gel 60 F254. TLC plates were visualized through UV absorbance, (254 nm),
or staining such as vanillin, phosphomolybdic acid, potassium-permanganate, or
ninhydrin followed by heating. Unless otherwise stated, purification was carried out by
flash column chromatography manually using Silica Gel (100-200 mesh) or automatically
using the Biotage Isolera One.

Characterization was carried out using LC-MS, NMR and UV-VIS instrumentation. All -
1
H and
13
C spectra were procured on the Departments Varian 400, 500 and 600 MHz
NMR instruments in the solvent indicated.
1
H and
13
C chemical shifts, (δ), are recorded
in parts per million, (ppm), and referenced to the residual solvent converted by the TMS
scale (CDCl3,
1
H = 7.26 ppm). Splitting patterns are denoted by s, d, t, dd, td, ddd, and
m and refer to the respective multiplicities; singlet, doublet, triplet, doublet of doublets,
triplet of doublets, doublet of doublet of doublet and multiplet. Mass spectra was
recorded on an Agilent 1260 LC-MS. UV-Vis spectra was obtained by a Hewlett-
Packard 8350 instrument.
41

(2-methyloxetan-2-yl)methyl pent-4-ynoate (2.12). To a flask was added 4-pentynoic
acid (5.0g, 51.0 mmol) 3-Methyl-3-oxetanemethanol (10.1 mL, 102 mmol) and DMAP
(0.3g, 2.6 mmol) in dry MeOH (60 mL). Dicyclohexylcarbodiimide (DCC 10.5g, 51.0
mmol) was cannulated into the reaction mixture at room temperature and stirred
overnight. The reaction was filtered and dried and with no further workup the crude was
purified on silica gel using EtOAc-hexanes mixture of (1:3) as the eluent to afford the
product 2.12 (9.0g, 97%).
1
H NMR (400 MHz, CDCl3) δ 4.46 (d, J = 6.2 Hz, 2H), 4.32
(d, J = 6.2 Hz, 2H), 4.14 (s, 2H), 2.59 – 2.50 (m, 2H), 2.50 – 2.42 (m, 2H), 1.95 (t, J =
2.5 Hz, 1H), 1.28 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 171.74, 82.29, 79.43, 79.43,
69.22, 68.84, 38.99, 33.24, 21.12, 14.37.

1-(3-Butynyl)-4-methyl-2,6,7-trioxabicyclo[2.2.2]octane (2.6). To a solution of the
half-ester 2.12 (3.5g, 19.2 mmol) in dry DCM (55 mL) was added BF3•OEt2 ( 0.6 mL, 4.8
mmol) dropwise at room temperature. After stirring for 1 h, 5 mL of Et 3N was added to
the reaction mixture. The reaction was diluted with ether, filtered and dried. With no
further workup the crude was purified on silica gel using EtOAc-hexanes- Et3N(1%)
mixture of (1:3) as the eluent to afford the pure product 2.6 (2.4g, 68%).
1
H NMR (400
MHz, CDCl3) δ 3.86 (s, 6H), 2.36 – 2.23 (m, 2H), 1.98 – 1.85 (m, 2H), 0.78 (s, 3H).
13
C
NMR (400 MHz, CDCl3) δ 108.05, 84.18, 72.64, 67.97, 35.86, 30.36, 14.58, 12.99.
O
O O
O
O
O
42

(7S)-8-(tert-butyldimethylsilyloxy)-3’-hydroxy-2’-(hydroxymethyl)-2’-methylpropyl-
7-hydroxyoct-4-ynoate (2.13). To a solution of OBO-ester 2.6 (2.0 g, 11.0 mmol) in
dry THF (50 mL) was added 2.5 M n-BuLi (4.4 mL, 11.0 mmol) at -78°C. After 0.25 h
BF3•Et2O (0.64 mL, 11.0 mmol) was added drop wise at -78°C. To the reaction mixture
was added protected glycidol (0.8 mL, 11.1 mmol) and stirred for 3 h at -78°C. The
reaction mixture was warmed to room temperature, quenched with saturated aqueous
NH4Cl (40 mL) and extracted with Et2O (3 x 60 mL). The organic layer solvent was
removed in vacuo and the crude mixture was suspended in 10 mL THF-H2O (1:1) and 1
M HCl was added at 0°C (2.5 mL, 2.5 mmol). The reaction mixture was stirred for 1h at
0°C quenched with NaHCO3 and extracted with Et2O (3 x 60 mL). The solvent was
removed in vacuo and the crude mixture was purified on silica gel using EtOAc-hexanes
(1:1) as the eluent to afford compound 2.13 (2.6 g, 63%) as a clear oil.
1
H NMR (400
MHz, CDCl3) δ 3.70 (q, J = 5.4 Hz, 1H), 3.65 (s, 3H), 3.64 – 3.61 (m, 1H), 3.55 (dd, J =
9.9, 5.7 Hz, 1H), 2.60 (d, J = 5.3 Hz, 1H), 2.44 (tt, J = 10.1, 4.6 Hz, 4H), 2.36 – 2.30 (m,
2H), 0.86 (s, 9H), 0.04 (s, 6H).
(S) (S)
TBSO
HO
O
O
HO
OH
43

Methyl (7S)-8-(tert-butyldimethylsilyloxy)-7-hydroxyoct-4-ynoate (2.13b). To a
solution of the dihydroxyl ester 2.13 (1.0 g, 2.7 mmol) in 10 mL THF-H2O (4:1) was
added Lithium hydroxide at room temperature and stirred for 1h. The reaction was
slowly neutralized using a 1M HCl solution (2.5 mL) and worked up with saturated
aqueous NH4Cl (20 mL) and extracted with Et2O (3 x 20 mL). The organic layer was
then concentrated and freshly prepared CH2N2 was added to convert the acid to the
Methyl ester. The solvent was completely removed in vacuo and the compound was
purified on silica gel using EtOAc-hexanes (1:20) as the eluent to afford compound 2.13b
(770 mg, 95%).
1
H NMR (400 MHz, CDCl3) δ 3.70 (q, J = 5.4 Hz, 1H), 3.65 (s, 3H),
3.64 – 3.61 (m, 1H), 3.55 (dd, J = 9.9, 5.7 Hz, 1H), 2.60 (d, J = 5.3 Hz, 1H), 2.44 (tt, J =
10.1, 4.6 Hz, 4H), 2.36 – 2.30 (m, 2H), 0.86 (s, 9H), 0.04 (s, 6H).
13
C NMR (400 MHz,
CDCl3) δ 172.61, 80.44, 76.93, 70.41, 65.60, 51.82, 33.68, 25.91, 23.41, 18.35, 14.78, -
5.36.

Methyl (7S)-8-(tert-butyldimethylsilyloxy)-7-(tert-butyldiphenylsilyloxy)oct-4-ynoate
(2.13c). To a flask with imidazole (176 mg, 2.6 mmol) and DMAP (11 mg, 0.09 mmol)
COOMe
(S) (S)
TBSO
HO
COOMe
(S) (S)
TBSO
TBDPSO
44
in CH2Cl2 (25 mL total volume) was added TBDPS-Cl (0.67 mL, 2.6 mmol) dropwise at
0°C. The alcohol 2.13b (520 mg, 1.7 mmol) was cannulated to the flask and stirred
overnight at room temperature. The reaction mixture was quenched with saturated
aqueous NH4Cl (30 mL) and extracted with Et2O (3 x 30 mL). The organic layer was
dried with MgSO4, filtered and the solvent removed in vacuo. The crude reaction
mixture was purified on silica gel using EtOAc-hexanes (1:9) as the eluent to afford
compound 2.13c (750 mg, 82%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ
7.82 – 7.69 (m, 4H), 7.41 (dt, J = 14.1, 6.9 Hz, 6H), 3.85 (p, J = 5.2 Hz, 1H), 3.69 (s, 3H),
3.65 – 3.55 (m, 2H), 2.68 – 2.04 (m, 6H), 1.10 (s, 9H), 0.87 (s, 9H), -0.00 (s, 3H), -0.03
(s, 3H).
13
C NMR (400 MHz, CDCl3) δ 172.69, 136.01, 135.95, 134.21, 134.16, 129.71,
129.68, 127.79, 127.76, 79.69, 77.99, 72.47, 65.62, 51.81, 33.76, 27.02, 26.00, 24.00,
19.47, 18.40, 14.86, -5.38.

Methyl (7S)-7-(tert-butyldiphenylsilyloxy)-8-hydroxyoct-4-ynoate (2.14). To a
solution of protected diol ester 2.13b (0.35 g, 0.65 mmol) in CH2Cl2 (12mL) was added
camphor sulfonic acid (120 mg, 0.52 mmol) at room temperature and monitored for 1 h.
The reaction was quenched with Et3N (0.45 mL, 0.62 mmol) and the solvent was
evaporated in vacuo without workup. The crude reaction mixture was purified on silica
gel using 15% EtOAc-hexanes as the eluent to afford compound 2.14 (200 mg, 70%) as a
clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 7.75 – 7.60 (m, 4H), 7.53 – 7.36 (m,
COOMe
(S) (S)
HO
TBDPSO
45
6H), 4.02 – 3.82 (m, 1H), 3.66 (s, 3H), 3.65 – 3.61 (m, 2H), 2.61 – 2.16 (m, 6H), 1.08 (s,
9H).
13
C NMR (400 MHz, CDCl3) δ 172.60, 135.91, 135.77, 133.62, 133.59, 130.01,
129.97, 127.90, 127.78, 80.43, 77.09, 72.51, 65.58, 51.80, 33.65, 27.06, 23.88, 19.39,
14.80.

Methyl (7S, 4Z)-7-(tert-butyldiphenylsilyloxy)-8-hydroxyoct-4-enoate (2.14b). To a
solution of alcohol 2.14 (0.15 g, 0.35 mmol) in EtOAc (50 mL) was added Lindlar
catalyst (10 mg) and 1 drop of quinoline. The reaction mixture was placed under a H2
atmosphere and stirred for 1 h. The reaction was filtered through celite and the solvent
was removed in vacuo. The crude product was purified on silica gel using 14% EtOAc-
hexanes as an eluent to afford the Z alkene (0.14 g, 94%) as a clear colorless oil.
1
H
NMR (400 MHz, CDCl3) δ 7.71 (td, J = 7.9, 1.6 Hz, 4H), 7.53 – 7.37 (m, 6H), 5.55 –
5.16 (m, 2H), 3.82 (dq, J = 9.3, 4.7 Hz, 1H), 3.65 (s, 3H), 3.51 (dtd, J = 18.2, 11.4, 5.3
Hz, 2H), 2.45 – 2.02 (m, 6H), 1.10 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ 173.65,
135.92, 135.76, 133.84, 133.73, 130.00, 129.86, 127.81, 127.72, 126.12, 77.49, 77.17,
76.85, 73.52, 65.35, 51.60, 33.78, 31.46, 27.08, 22.69, 19.37.

Methyl (7S, 4Z)-7-(tert-butyldiphenylsilyloxy)-8-oxooct-4-enoate (2.15). To a -78°C
solution of DMSO (0.1 mL, 1.06 mmol) in anhydrous CH2Cl2 (25 mL) was added oxalyl
chloride (0.06 mL, 11.4 mmol) dropwise. After 0.25 h alcohol 2.14a (0.14 g, 0.33 mmol)
7
COOMe
(S) (S)
HO
TBDPSO
7
COOMe
(S) (S)
O
TBDPSO
46
was added and stirred for 1 h at -78°C. To the reaction mixture was added Et3N (0.25
mL, 1.76 mmol) and stirred for another 3 h at -78°C. The reaction mixture was allowed
to warm to room temperature and quenched with saturated aqueous NH4Cl (25 mL) and
extracted with Et2O (3 x 25 mL). The combined extract was dried with Na2SO4 and
evaporated to give a crude clear oil which was then chromatographed on silica gel using
EtOAc-hexanes mixture of (1:5) as the eluent to afford the product 2.15 (0.14, 98%) as a
viscous and colorless oil.
1
H NMR (400 MHz, CDCl3) δ 9.57 (d, J = 1.3 Hz, 1H), 7.69 –
7.60 (m, 4H), 7.41 (dtd, J = 24.7, 7.0, 4.2 Hz, 6H), 5.61 – 5.25 (m, 2H), 4.08 (td, J = 6.0,
1.6 Hz, 1H), 3.65 (s, 3H), 2.53 – 2.09 (m, 6H), 1.12 (s, 9H).

Methyl (7S, 4Z, 8E)-7-(tert-butyldiphenylsilyloxy)-9-iodonona-4,8-dienoate (2.3). To
a solution of CrCl2 (0.321 g, 2.61 mmol) dissolved in THF (3 mL total volume) was
cannulated a mixture of compound 2.15 (0.13 g, 0.3 mmol) and CHI3 (0.34 g, 0.87
mmol) dissolved in anhydrous THF (4 mL) under Argon at 0°C. The reaction was stirred
at 0°C for 3 h and an additional 1 h at room temperature. The reaction mixture was
quenched with water (15 mL) extracted with Et2O (3 x 15 mL) rinsed with brine and
dried over MgSO4. The organic phase was filtered and the solvent was removed in vacuo
to afford a crude oil which was purified on silica gel using first pure pentanes and then
EtOAc-hexanes (1:24) as the eluent to afford vinyl iodide 2.13 (84 mg, 53%) as a clear
colorless oil.
1
H NMR (400 MHz, CDCl3) δ 7.66 (ddd, J = 16.9, 8.0, 1.6 Hz, 4H), 7.42
(dq, J = 14.2, 7.0, 6.5 Hz, 6H), 6.49 (dd, J = 14.5, 6.7 Hz, 1H), 6.00 (dd, J = 14.6, 1.2 Hz,
1H), 5.50 – 5.25 (m, 2H), 4.12 (q, J = 6.9, 6.5 Hz, 1H), 3.67 (s, 3H), 2.32 – 2.14 (m, 6H),
7
COOMe
(S) (S)
TBDPSO
I
47
1.08 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ 173.53, 147.75, 135.97, 135.93, 133.80,
133.46, 130.24, 129.90, 129.86, 127.72, 127.69, 125.48, 77.16, 75.62, 51.67, 35.17, 27.08,
22.90, 19.43.

5-Trimethylsilyl-2E-penten-4-yn-1-ol (2.17). To a solution of (E)-pent-2-en-4-yn-1-ol
(3.0 g, 36.5 mmol) in dry THF (40 mL) was added 2.5 M n-BuLi (29.2 mL, 73.0 mmol)
dropwise at -78°C. After 1 h TMS-Cl (9.2 g, 73.0 mmol) was added and stirred at room
temperature. After 4 h acetic acid (7 mL) was added to the reaction mixture and
neutralized with saturated aqueous NaHCO3 (50 mL). The reaction mixture was
extracted with Et2O (3 x 50 mL), washed with brine, dried with MgSO4, filtered and the
solvent removed in vacuo. The crude reaction mixture was purified on silica gel using
EtOAc-hexanes (10%) as the eluent to afford the protected pentenynol 2.17 (2.6 g, 87%)
as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 6.29 (dt, J = 16.0, 5.1 Hz, 1H),
5.76 (dt, J = 15.9, 1.8 Hz, 1H), 4.19 (dd, J = 5.1, 1.8 Hz, 2H), 1.92 (s, 1H), 0.18 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ 143.08, 110.43, 103.13, 95.39, 62.91, 0.01.

(3E)-(5-bromopent-3-en-1-ynyl) trimethylsilane (2.18). To a flask with PPh3 (1.9 g,
7.3 mmol) and N-bromosuccinimide (1.3 g, 7.3 mmol) in dry CH 2Cl2 (20 mL) was added
pentenynol 2.17 (1.0 g, 6.5 mmol) at 0°C. The reaction was stirred for 30 minutes at
room temperature, quenched with saturated aqueous NH4Cl (25 mL) and extracted with
48
Et2O (3 x 25 mL). The organic layer was dried with MgSO4, filtered and the solvent
removed in vacuo. The crude reaction mixture was purified on silica gel using EtOAc-
hexanes (1%) as the eluent to afford the allylic bromide 2.18 (1.2 g, 86%) as a clear
colorless oil.
1
H NMR (400 MHz, CDCl3) δ 6.30 (dt, J = 15.6, 7.8 Hz, 1H), 5.75 (d, J =
15.6 Hz, 1H), 3.96 (d, J = 7.8 Hz, 2H), 0.19 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ
139.04 , 114.54 , 102.08 , 97.77 , 31.54 , -0.07.

(2E)-5-(trimethylsilyl)pent-2-en-4-ynyl triphenylphosphonium bromide (2.7). To a
flask with PPh3 (1.5 g, 5.5 mmol) in dry C6H6 (10 mL) was added allylic bromide 2.18
(1.0 g, 4.6 mmol) in the dark. The reaction mixture was stirred overnight, filtered,
washed with Et2O (3 x 15 mL) and dried in vacuum to afford the wittig ylide 2.7 (2.0 g,
90%) as a white solid.
1
H NMR (400 MHz, CDCl3) δ 7.95 – 7.56 (m, 15H), 6.19 (dd, J =
15.7, 5.4 Hz, 1H), 5.99 – 5.80 (m, 1H), 5.03 (dd, J = 16.1, 7.5 Hz, 2H), 0.12 (s, 9H).
13
C
NMR (400 MHz, CDCl3) δ 135.24, 135.21, 134.16, 134.07, 130.58, 130.45, 120.99,
120.84, 118.36, 117.50, 110.11, -0.14.

(4S, 5R, 2Z)-1,3-Dioxolane-4-methanol-2,2-dimethyl-5-pent-2-en-1-yl (2.19). To a
suspension of propyl wittig salt 2.9 (0.57 g, 15 mmol) in THF (10 mL) was added
(S) (S)
(R) (R)
O O
OH
49
dropwise a 1 M solution of Sodium hexamethyldisilizane (NaHMDS; 13.5 mL, 13.5
mmol) at -78°C. The reaction was stirred for 0.5 h at room temperature. The sugar 2.8
(1.3 g, 7.5 mmol) was cannulated to the flask at -78°C for 3 h at -78°C. The reaction
was allowed to warm to room temperature and stir for 2 h. The reaction mixture was
quenched with saturated aqueous NH4Cl (15 mL) and extracted with Et2O (3 x 15 mL).
The organic layer was dried with MgSO4, filtered and the solvent removed in vacuo. The
crude reaction mixture was purified on silica gel using EtOAc-hexanes (1:4) as the eluent
to afford the alcohol 2.16 (700 mg, 47%) as a clear colorless oil.
1
H NMR (400 MHz,
CDCl3) δ 5.60 – 5.43 (m, 1H), 5.40 – 5.27 (m, 1H), 4.18 (dq, J = 11.3, 6.0 Hz, 2H), 3.78
– 3.41 (m, 2H), 2.32 (ddt, J = 41.4, 14.1, 6.7 Hz, 2H), 2.14 – 1.97 (m, 2H), 1.47 (s, 3H),
1.36 (s, 3H), 0.97 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ 134.49, 123.93,
108.30, 77.96, 76.89, 61.79, 28.25, 27.43, 25.55, 20.94, 14.16.

(4S, 5R, 2Z)-1,3-Dioxolane-4-carboxaldehyde-2,2-dimethyl-5-pent-2-en-1-yl (2.20).
To a solution of alcohol 2.16 (0.5 g, 2.5 mmol) in anhydrous CH2Cl2 (30 mL) was added
NaHCO3 (4.2 g, 50 mmol) dropwise. Dess-Martin periodinane (2.12 g, 5 mmol)
dissolved in CH2Cl2 (5 mL) was cannulated into the reaction mixture and stirred. After 2
h the reaction mixture was quenched with saturated aqueous Na2S2O3 (6 mL) and
extracted with Et2O (3 x 35 mL). The combined extract was dried with Na2SO4 and
evaporated to give a crude clear oil which was then chromatographed on silica gel using
(R) (R)
(R) (R)
O O
O
50
EtOAc-hexanes mixture of (1:4) as the eluent to afford the product 2.17 (0.3 g, 62%) as a
viscous and colorless oil.
1
H NMR (400 MHz, CDCl3) δ 9.63 (d, J = 3.2 Hz, 1H), 5.56 –
5.42 (m, 1H), 5.39 – 5.26 (m, 1H), 4.36 (q, J = 6.8 Hz, 1H), 4.28 (dd, J = 7.1, 3.1 Hz, 1H),
2.29 (t, J = 6.7 Hz, 2H), 2.11 – 1.89 (m, 2H), 1.56 (s, 3H), 1.37 (s, 3H), 0.92 (t, J = 7.5
Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ 201.63, 135.03, 123.01, 110.60, 81.99, 78.49,
27.85, 27.50, 27.46, 25.26, 20.85, 13.98.

(6S, 7R)-1,3-Dioxolane-13-(trimethylsilyl)-trideca-3,8,10-triene-12-ynyl (2.21). To a
solution of wittig salt 2.7 (1.1 g, 2.32 mmol) in THF (20 mL) was added 2.5 M n-BuLi
(0.7 mL, 1.7 mmol) at -78°C and stirred for 0.5 h at 0°C. Aldehyde 2.20 (0.23 g, 1.16
mmol) was cannulated to the reaction mixture at -78°C. The reaction was then allowed to
stir at room temperature for 3 h. After 3 h the reaction was quenched with saturated
aqueous NH4Cl (15 mL) and extracted with Et2O (3 x 20 mL). The organic layer was
dried with MgSO4, filtered and the solvent removed in vacuo. The crude mixture was
taken onto the next step without any further purification (0.25 g, 68%) as a brown oil.

(R) (R)
(S) (S)
O O
TMS
(R) (R)
(S) (S)
OTBS
TBSO
TMS
51
(6S, 7R, 3Z, 8E, 10E)-6,7-bis(tert-butyldimethylsilyloxy)-13-(trimethylsilyl)-trideca-
3,8,10-triene-12-ynyl (2.22). To a solution of compound 2.21 (170 mg, 0.53 mmol) in
MeOH-DCM (1:1, 3 mL) was added 1M HCl (2 mL) at 0°C and stirred for 1 h at room
temperature. The reaction mixture was quenched with saturated NaHCO3 (5 mL) and
extracted with Et2O (3 x 15 mL). The solvent was removed in vacuo and the crude
mixture was purified on silica gel using MeOH-CH2Cl2 (1:20) as the eluent to afford a
mixture (40:60) E, E / Z, E as a colorless oil. The product mixture was then dissolved in
CH2Cl2 (15 mL) and stirred with TBS-OTf (0.6 mL, 2.7 mmol) and Lutidine (0.6 mL, 5.3
mmol) overnight. Upon workup and extraction the crude mixture was purified with
EtOAc-hexanes (1:49) as the eluent to afford compound 2.22 (193 mg, 72%) over two
steps as a colorless oil.
1
H NMR (400 MHz, CDCl3) δ 6.64 (dd, J = 15.5, 11.1 Hz, 1H),
6.13 (dd, J = 15.3, 10.7 Hz, 1H), 5.79 (dd, J = 15.3, 7.3 Hz, 1H), 5.57 (d, J = 15.6 Hz,
1H), 5.45 – 5.35 (m, 2H), 4.00 (ddd, J = 6.8, 4.0, 0.9 Hz, 1H), 3.61 (td, J = 6.0, 4.3 Hz,
1H), 2.30 – 2.12 (m, 2H), 2.01 (p, J = 7.3 Hz, 2H), 0.95 (t, J = 7.5 Hz, 3H), 0.87 (s, 9H),
0.85 (s, 9H), 0.19 (s, 6H), 0.02 (s Hz, 3H), 0.01 (s, 3H), -0.02 (s, 3H). (6S, 7R, 3Z, 8E,
10Z)-6,7-bis(tert-butyldimethylsilyloxy)-13-(trimethylsilyl)-trideca-3,8,10-triene-12-
ynyl.
1
H NMR (400 MHz, CDCl3) δ 6.82 (ddd, J = 15.4, 11.5, 1.1 Hz, 1H), 6.05 (ddt, J =
11.7, 11.0, 0.9 Hz, 1H), 5.59 (dd, J = 15.6, 13.0 Hz, 1H), 5.49 (ddt, J = 11.0, 8.9, 0.9 Hz,
1H), 5.45 – 5.35 (m, 2H), 4.40 (ddd, J = 8.9, 4.2, 1.1 Hz, 1H), 3.66 (td, J = 6.0, 4.2 Hz,
1H), 2.27 – 2.16 (m, 2H), 2.06 – 1.97 (m, 2H), 0.97 (t, J = 7.5 Hz, 3H), 0.87 (s, 9H), 0.86
(s, 9H), 0.20 (s, 9H), 0.04 (s, 6H), 0.02 (s, 3H), -0.00 (s, 3H).
52
2.22b
(6S, 7R, 3Z, 8E, 10E)-6,7-bis(tert-butyldimethylsilyloxy)-13-(trimethylsilyl)-trideca-
3,8,10-triene-12-ynyl (2.22b). To a solution of compound 2.22 (85 mg, 0.17 mmol) in
CH2Cl2 (30 mL) was added I2 (5 mg) and stirred overnight at room temperature with hv.
The reaction mixture was quenched with saturated Na2S2O5 (30 mL) and exctracted with
Et2O (30 mL). The solvent was removed in vacuo and the crude oil was purified on silica
gel using EtOAc-hexanes (1:49) as the eluent to afford compound 2.22b (83 mg, 98%) as
a colorless oil.
1
H NMR (400 MHz, CDCl3) δ 6.64 (dd, J = 15.5, 11.1 Hz, 1H), 6.13 (dd,
J = 15.3, 10.7 Hz, 1H), 5.79 (dd, J = 15.3, 7.3 Hz, 1H), 5.57 (d, J = 15.6 Hz, 1H), 5.45 –
5.35 (m, 2H), 4.00 (ddd, J = 6.8, 4.0, 0.9 Hz, 1H), 3.61 (td, J = 6.0, 4.3 Hz, 1H), 2.30 –
2.12 (m, 2H), 2.01 (p, J = 7.3 Hz, 2H), 0.95 (t, J = 7.5 Hz, 3H), 0.87 (s, 9H), 0.85 (s, 9H),
0.19 (s, 6H), 0.02 (s Hz, 3H), 0.01 (s, 3H), -0.02 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ
143.28, 138.09, 133.44, 130.24, 125.20, 109.55, 83.06, 79.51, 77.48, 77.16, 76.84, 76.66,
76.47, 31.77, 26.08, 20.94, 18.39, 18.28, 14.32, -4.02, -4.02, -4.24, -4.52.

(6S, 7R, 3Z, 8E, 10E)-6,7-bis(tert-butyldimethylsilyloxy)-trideca-3,8,10-triene-12-
ynyl (2.4). To a solution of compound 2.22b (63 mg, 0.124 mmol) in a one to one ratio
of MeOH-CH2Cl2 (4 mL) was added Na2CO3 (10 mg) and stirred overnight at room
temperature. The solvent was removed in vacuo and the crude was dissolved in water (5
(R) (R)
(S) (S)
OTBS
TBSO
TMS
(R) (R)
(S) (S)
OTBS
TBSO
53
mL) and extracted with Et2O (3 x 5 mL). The oil was purified on silica gel using EtOAc-
hexanes (1:49) as the eluent to afford compound 2.4 (52 mg, 96%) as a colorless oil.
1
H
NMR (400 MHz, CDCl3) δ 6.67 (dd, J = 15.6, 10.7 Hz, 1H), 6.16 (dd, J = 15.3, 10.7 Hz,
1H), 5.82 (dd, J = 15.3, 7.0 Hz, 1H), 5.53 (dd, J = 15.8, 2.3 Hz, 1H), 5.46 – 5.29 (m, 2H),
4.02 (ddd, J = 6.9, 4.3, 0.8 Hz, 2H), 3.62 (td, J = 6.0, 4.4 Hz, 1H), 3.02 (d, J = 2.1 Hz,
1H), 2.28 – 2.14 (m, 3H), 2.02 (p, J = 7.4 Hz, 2H), 0.95 (t, J = 7.6 Hz, 3H), 0.89 (s, 8H),
0.86 (s, 7H), 0.04 (s, 3H), 0.03 (s, 3H), 0.02 (s, 3H), -0.00 (s, 3H).
13
C NMR (400 MHz,
CDCl3) δ 143.28, 138.09, 133.44, 130.24, 125.20, 109.55, 83.06, 79.51, 76.66, 76.47,
31.77, 26.08, 26.08, 20.94, 18.39, 18.28, 14.32, -4.02, -4.02, -4.24, -4.52.

Methyl (7S, 16R, 17S, 4Z, 8E, 12E, 14E, 19Z)-tris-(tert-butyldimethylsilyloxy)
docosa-4,8,12,14,19-pentaen-10-ynoate (2.23). To the arm of a three-necked flask was
charged Pd(PPh3)4 (20 mg, .018 mmol) and CuI (7 mg, .036 mmol) under Argon. A
solution of iodide 2.3 (45 mg, 0.09 mmol), alkyne 2.4 (40 mg, 0.09 mmol) and Et3N (0.13
mL, 0.9 mmol) in C6H6 (5 mL) was cannulated into the reaction vessel. The reaction
flask was then freeze-thawed with liquid nitrogen three times to remove an oxygen.
After removing any oxygen from the reaction flask the Pd(PPh3)4 and CuI was added and
the reaction mixture was stirred overnight at room temperature. The reaction was worked
up with aqueous saturated NH4Cl (5 mL) and extracted with Et2O (3 x 5 mL). The
solvent was evaporated and the mixture was purified on silica gel using 3% EtOAc-
COOMe
(S) (S)
(R) (R)
(S) (S)
TBSO
OTBS
TBSO
54
hexanes as the eluent to afford compound 2.23 (60 mg, 77%) as a clear oil.
1
H NMR
(400 MHz, CDCl3) δ 7.70 – 7.58 (m, 4H), 7.44 – 7.34 (m, 6H), 6.57 (dd, J = 15.4, 10.8
Hz, 1H), 6.16 (dd, J = 15.3, 10.9 Hz, 1H), 6.08 (dd, J = 15.7, 5.7 Hz, 1H), 5.78 (dd, J =
15.3, 7.1 Hz, 1H), 5.75 – 5.62 (m, 2H), 5.45 – 5.26 (m, 4H), 4.29 – 4.20 (m, 1H), 4.01
(dd, J = 7.8, 4.4 Hz, 1H), 3.64 (s, 3H), 3.63 – 3.57 (m, 1H), 2.23 (q, J = 8.1, 7.4 Hz, 6H),
2.13 (dd, J = 13.0, 6.9 Hz, 2H), 2.05 – 1.98 (m, 2H), 1.07 (s, 9H), 0.95 (t, J = 7.5 Hz, 3H),
0.88 (s, 9H), 0.86 (s, 9H), 0.03 (s, 3H), 0.03 (s, 3H), 0.02 (s, 3H), -0.01 (s, 3H).

Methyl (7S, 16R, 17S)-Trihydroxydocosa-(4Z, 8E, 12E, 14E, 19Z)-pentaen-10-ynoate
(2.2). To a solution of compound 3 (60 mg, 0.07 mmol) in THF (4 mL) was added 1.0 M
solution of TBAF (0.42 mL, 0.42 mmol) at 0°C and stirred overnight. The reaction was
quenched with water (3 mL) and extracted with Et2O (5 x 3 mL), rinsed with brine, dried
over MgSO4 and filtered. The solvent was then concentrated and freshly prepared CH2N2
was added to convert any acid to the ester. The solvent was completely removed in
vacuo and the compound was purified on silica gel using MeOH-CH2Cl2 (3%) as the
eluent to afford the ester product 2.2 as a light yellow oil (17.2 mg, 86%).
1
H NMR (400
MHz, CDCl3) δ 6.59 (dd, J = 15.6, 11.0 Hz, 1H), 6.35 (dd, J = 15.1, 10.9 Hz, 1H), 6.16
(dd, J = 15.8, 5.4 Hz, 1H), 5.95 – 5.81 (m, 2H), 5.74 (dd, J = 15.4, 1.9 Hz, 1H), 5.61 –
5.38 (m, 3H), 5.43 – 5.28 (m, 1H), 4.23 (p, J = 5.3, 4.5 Hz, 2H), 3.78 – 3.70 (m, 1H),
3.66 (s, 3H), 2.42 – 2.32 (m, 6H), 2.26 (dt, J = 15.4, 8.2 Hz, 1H), 2.20 – 2.11 (m, 1H),
COOMe
(S) (S)
(R) (R)
(S) (S)
HO
OH
HO
55
2.09 – 1.98 (m, 2H), 0.95 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ 173.95,
145.04, 140.67, 135.48, 133.35, 132.06, 131.50, 125.90, 124.03, 90.82, 89.56, 77.48,
77.16, 76.84, 74.71, 74.03, 71.46, 65.98, 51.85, 35.09, 33.67, 30.04, 29.82, 29.69, 22.83,
20.85, 14.33.

Methyl (7S, 16R, 17S)-Trihydroxydocosa-(4Z, 8E, 10Z, 12E, 14E, 19Z)-pentaenoate,
or RvD2 Methyl Ester (2.1). A flame dried flask was charged with a freshly prepared
Zn/Cu/Ag amalgam (500 mg, excess) and suspended in H2O-MeOH mixture (1:1, 1 mL).
To the reaction slurry was added compound 2.2 (9.1 mg, 0.023 mmol) and stirred
overnight while monitoring. The reaction was quenched with a few drops of quinolone,
filtered, dried and purified via HPLC at H2O-MeOH mixture (32%) to afford the clean
Resolvin D2 Methyl Ester 2.1 (8.45 mg, 92%).
1
H NMR (400 MHz, CDCl3) δ 6.74 (dd, J
= 15.0, 8.6 Hz, 1H), 6.71 (dd, J = 14.9, 8.5 Hz, 1H), 6.40 (dd, J = 14.7, 10.7 Hz, 1H),
6.26 (dd, J = 14.7, 10.7 Hz, 1H), 6.04 (dd, J = 10.2, 8.5 Hz, 1H), 6.01 (dd, J = 10.2, 8.6
Hz, 1H), 5.81 (dd, J = 14.7, 8.4 Hz, 1H), 5.77 (dd, J = 15.0, 7.2 Hz, 1H), 5.57 (qt, 1H),
5.52 – 5.44 (m, 2H), 5.37 (dd, J = 10.8, 1.3 Hz, 1H), 4.32 – 4.26 (m, 1H), 4.26 – 4.21 (m,
1H), 3.76 – 3.69 (m, 1H), 3.67 (s, 3H), 2.43 – 2.34 (m, 5H), 2.32 – 2.25 (m, 1H), 2.22 –
2.15 (m, 1H), 2.11 – 2.02 (m, 3H), 0.96 (t, J = 7.5 Hz, 3H).
13
C NMR (500 MHz, CDCl3)
δ 137.05, 135.52, 133.32, 132.87, 131.42, 131.31, 129.58, 129.55, 128.90, 126.29, 125.57,
7
17
COOMe
(S) (S)
(R) (R) (S) (S)
HO
OH
HO
16
56
124.14, 77.41, 77.16, 76.91, 75.10, 74.09, 71.86, 51.82, 35.51, 33.78, 30.10, 22.92, 20.88,
14.37.

57
2.6 References

43. Hong, S.; Gronert, K.; Devchand, P. R.; Moussignac, R. L.; Serhan, C. N. Journal of
Biological Chemistry 2003, 278, 14677.
44. Mas, E.; Croft, K. D.; Zahra, P.; Barden, A.; Mori, T. A. Clinical Chemistry 2012, 58,
1476.
45. Raatz, S. K.; Golovko, M. Y.; Brose, S. A.; Rosenberger, T. A.; Burr, G. S.; Wolters,
W. R.; Picklo, M. J., Sr. Journal of Agricultural and Food Chemistry 2011, 59,
11278.
46. Sun, Y.-P.; Oh, S. F.; Uddin, J.; Yang, R.; Gotlinger, K.; Campbell, E.; Colgan, S. P.;
Petasis, N. A.; Serhan, C. N. Journal of Biological Chemistry 2007, 282, 9323.
47. Corey, E. J.; Raju, N. Tetrahedron Letters 1983, 24, 5571.
48. Wipf, P.; Tsuchimoto, T.; Takahashi, H. Pure and Applied Chemistry 1999, 71, 415.
49. Takai, K.; Nitta, K.; Utimoto, K. Journal of the American Chemical
Society 1986, 108, 7408.
50. Maryanoff, B. E.; Reitz, A. B. Chemical Reviews 1989, 89, 863.
51. Spite, M.; Norling, L. V.; Summers, L.; Yang, R.; Cooper, D.; Petasis, N. A.; Flower,
R. J.; Perretti, M.; Serhan, C. N. Nature 2009, 461, 1287.
52. Hotchkiss, R. S.; Karl, I. E. New England Journal of Medicine 2003, 348, 138.
53. Bento, A. F.; Claudino, R. F.; Dutra, R. C.; Marcon, R.; Calixto, J. B. Journal of
Immunology 2011, 187, 1957.
54. Ji, R.-R.; Xu, Z.-Z.; Strichartz, G.; Serhan, C. N. Trends in Neurosciences 2011, 34,
599.
58
CHAPTER 3. First Total Synthesis and Stereochemical Assignment of
Resolvin D3 and its Aspirin Triggered Analogue (RvD3 and AT-RvD3)
3.1 Introduction
Recent investigations have shown biochemical mediators are part of a complex active
immune response involving a number of receptor-signaling pathways that respond to
inflammation.
55,56 ,57
In 2002, Serhan and coworkers isolated a novel family of such
mediators derived from endogenous exudates termed resolvins.
58
It was known that these
lipid metabolites were formed enzymatically from docosahexanoic acid (DHA) to act in
the resolution of inflammation, however it remained unclear their exact role and in what
specific pathway.
59,60
Recently it has been shown that inflammation is not a passive
process but rather is actively regulated by many of these lipid metabolites.
61
The resolvin
D series is biosynthesized in two adjacent lipoxygenase trasformations, (Figure 12).

Figure 12. Biosynthesis of RvD3 and AT-RvD3.
59

First, DHA is converted to 17S-HpDHA by 15-Lipoxygenase. Then, 5-Lipoxygenase
forms the peroxide at the C-4 position, ultimately forming metabolites RvD3 and RvD4
through enzymatic epoxidation. Alternatively DHA can undergo a COX-2 pathway
producing the aspirin triggered analogue of resolvin D3. Resolvin D3 is known to
mitigate acute kidney injury,
62
however it is still unknown what other roles it has in the
immune response.
63
We have focused our most recent efforts on this compound and its
analogues to establish its complete stereochemistry as well as understand its role in
inflammation. It was with this goal in mind we report the total synthesis of (4S, 11R,
17S, 5Z, 7E, 9E, 13Z, 15E, 19Z)-trihydroxy docosahexanoic acid, (1) and its aspirin
triggered analogue, (2). To the best of our knowledge, no total synthesis of resolvin D3
has been reported and recently we reported our total synthesis and stereochemical
assignment of this molecule and its aspirin triggered epimer.
64

3.2 Results and Discussion
3.2.1 Synthesis of RvD3 and AT-RvD3
3.2.1.1 Retrosynthetic Analysis
The 22 carbon chain lipid has a distinct structure containing two conjugated systems, the
C5-C10 Z, E, E triene and the C13-16 Z, E diene as well as a trihydroxyl structure in the
4S, 11R, 17S position (Figure 13). Our aim was to follow a synthetic route allowing for
our light sensitive triene and diene moieties to be formed in the final steps of our
synthesis. By taking advantage of a mild Zn/Cu/Ag hydrogenation at the final stage we
envisioned a simple reduction of the bis- acetylinic precursor to its cis olefinic lipid.
Retrosynthetically, and with our internal alkynes, in mind we utilized the extremely
60

Figure 13. Retrosynthetic Analysis of RvD3 and AT-RvD3.

efficient Sonogashira to couple the highly reactive E, E vinyl iodide (3.4) with the
terminal alkyne (3.5). The final carbon carbon bond was made as well using sonogashira
conditions coupling vinyl halide (3.3) with the ester piece. Enantiomerically pure
building blocks could be attained through the use of natural chiral compounds such as R,
S, glycidol (9, 10) as well as the gamma carboxy butryolactone (11).

3.2.1.2 Synthesis of Alkynyl Ester
Efficient access of the C(1) C(6) fragment 3.5 was synthesized on large-scale beginning
from the ring opening Fisher esterification of (3.8) in methanol and in the presence of a
catalytic amount of HCl (Scheme 5). Selective reduction to afford the diol (3.9) was
carried out employing borane dimethyl sulfide complex and a catalytic amount of sodium
61

Scheme 5. Synthesis of Key Alkynyl Ester Intermediate (3.5).

tetrahydroborate.
65,66,67
Silylation followed by selective removal of the primary alkoxy
and further oxidation afforded the silyl ether aldehyde (3.13) in appreciable yields.
Aldehyde was transformed to the alkyne using Corey-Fuchs homologation
68
with mild
base, lithium diisopropylamide,
69
to preserve the integrity of the ester in modest yield,
(62%). Alternatively, employing the Bestmann Ohira homologation
70,71,72
afforded the
product in 43% yield with the added disadvantage of multistep preparation of reagent.
3.2.1.3 Synthesis of Vinyl Iodide 3.4
To synthesize vinyl halide (3.4), Scheme 6, we carried out the lithiation of silyated
acetylene (3.15) followed by the addition of protected S-Glycidol (3.7) to afford the diol,
62

Scheme 6. Total Synthesis of Vinyl Iodide Key Intermediate (3.4)

which was then silylated to form intermediate (3.16). The protected diol was selectively
desilyated (3.17) and oxidized using Swern conditions to form (3.18). Wittig olefination
using (triphenylphosphoranylidene) acetaldehyde affords α, β – unsaturated aldehyde
(3.19) which was then converted to the vinyl iodide
73
(3.4) using Takai olefination

to
afford a mixture of the trans – cis olefin (9:1).
74
3.2.1.4 Synthesis of Vinyl Iodide (3.3)
Next we synthesized building block (3.3), pictured in Scheme 7, for the final Sonogashira
coupling of our omega end piece. Lithiation of 1-butyne (3.20) and the ring opening of
protected R Glycidol (3.6) afforded the diol, which was subsequently silyated with tert-
butyldiphenylsilyl (TBDPS) to afford the silyl ether (3.21). Selective deprotection
afforded the primary alcohol (3.22), which was then hydrogenated using Lindlar
63

Scheme 7. Total Synthesis of Vinyl Iodide Intermediate (3.3).

conditions to give the all cis alcohol (3.23). Oxidation using swern conditions gave
compound (3.24) in good yield which was then olefinated using the takai reaction. De-
silylation allowed clean purification and then protection afforded compound (3.3).
3.2.1.5 Final Construction of Resolvin D3
With alkynyl ester (3.5) in hand we coupled building block (3.4) under Sonogashira
conditions to afford the C1-C14 chain (3.25). Next we desilyated the terminal alkyne to
setup a second and final Sonogashira coupling with vinyl iodide (3.3). We carried out the
Sonogashira, Scheme 8, to afford the protected bis-acetylenic methyl ester in good yield,
(3.26), however when deprotecting using TBAF we encountered some difficulties. The
reaction gave a very low yield as a mixture of the 5 membered lactone and ester.
75
It is
64
well known that in the slightest presence of acid or base inter-conversion of gamma
butyrolactone to the open hydroxyl carboxylic acid is thermodynamically favorable.
76,77

It was also observed that during the purification of the pure bis-acetylenic methyl ester on
C-18 reverse phased HPLC the ester would undergo lactonization to form a 60:40 ratio of
ester to lactone. It was determined that any slightly acidic media or potentially
nucleophilic anion such as the fluoride could potentially cause side reactions leading to
the formation of side products and making for inefficient deprotection and reduction to
form the final product.
78
Further experimentation is being carried out to optimize these
conditions and better determine a lactone pathway. Finally the bis-acetylenic triol was
hydrogenated using a mild Zn/Cu/Ag amalgam
79,80
to afford a mixture of the reduced
product as a lactone and ester. The product mixture was purified by HPLC and
hydrolyzed to form a single stereoisomer of RvD3 as the sodium salt.
81

Scheme 8. Final Coupling of RvD3 Methyl Ester (3.27).

65
3.3 Biological Actions
Recently we reported our findings elucidating the structure of Resolvin D3 and AT-
Resolvin D3. In this work the temporal profile of RvD3 and AT-RvD3 was shown to be
very unique in that it takes of late, (24 hours), peaks at 48 hours and is still abundant after
72 hours.
82 83
It also reduces levels of pro-inflammatory eicosanoids such as
thromboxanes, prostaglandins and and leukotrienes while stimulating resolution through
macrophage phagocytosis.
3.4 Conclusion
In conclusion we report the total synthesis of RvD3 (1) and its aspirin triggered
stereoisomer (2). Using a convergent strategy we assembled crucial building blocks that
afforded complete stereochemistry. These compounds will be further tested in
collaborative studies to explore their role against inflammation and ultimately help make
profound advances in therapies to treat a number of disease processes related to chronic
inflammation.

Figure 14. Exudate levels of D Series Resolvins post Zymosan Induced Response.

66
3.5 Experimental Procedure
Unless otherwise noted, all reactions were carried out in a flame-dried flask with stir bar
under argon routed through a three-necked valve. Reactions were carried out at room
temp using DriSolv solvents purchased commercially from VWR. All reagents used
were purchased without further purification from Sigma Aldrich, Strem, and Alfa Aesar.

Progress was monitored and recorded using EMD analytical thin layer chromatography
plates, Silica Gel 60 F254. TLC plates were visualized through UV absorbance, (254 nm),
or staining such as vanillin, phosphomolybdic acid, potassium-permanganate, or
ninhydrin followed by heating. Unless otherwise stated, purification was carried out by
flash column chromatography manually using Silica Gel (100-200 mesh) or automatically
using the Biotage Isolera One.

Characterization was carried out using LC-MS, NMR and UV-VIS instrumentation. All -
1
H and
13
C spectra were procured on the Departments Varian 400, 500 and 600 MHz
NMR instruments in the solvent indicated.
1
H and
13
C chemical shifts, (δ), are recorded
in parts per million, (ppm), and referenced to the residual solvent converted by the TMS
scale (CDCl3,
1
H = 7.26 ppm). Splitting patterns are denoted by s, d, t, dd, td, ddd, and
m and refer to the respective multiplicities; singlet, doublet, triplet, doublet of doublets,
triplet of doublets, doublet of doublet of doublet and multiplet. Mass spectra was
recorded on an Agilent 1260 LC-MS. UV-Vis spectra was obtained by a Hewlett-
Packard 8350 instrument.

67

(S)-O-t-Butyldimethylsilyl glycidol (3.9). To a solution of (S)-γ-carboxy butyrolactone
[S-5-oxo-2-tetrahydrofurancarboxylic acid] (5.0g, 38.5 mmol) in dry MeOH (40 mL) was
added 4 drops of concentrated HCl and heated to reflux overnight. The reaction was
cooled to 0°C and quenched with NaHCO3 and filtered. The solvent was then evaporated
with no further workup and the crude was purified on silica gel using EtOAc-hexanes
mixture of (1:1) as the eluent to afford the product 3.9 as a viscous and colorless oil (6.5g,
97%).
1
H NMR (400 MHz, CDCl3) δ 4.23 (dd, J = 7.9, 4.2 Hz, 1H), 3.78 (s, 3H), 3.66 (s,
3H), 2.94 (s, 1H), 2.60 – 2.33 (m, 2H), 2.24 – 2.05 (m, 1H), 1.99 – 1.83 (m, 1H). 13C
NMR (400 MHz, CDCl3) δ 175.14, 173.67, 69.58, 52.76, 51.84, 29.53, 29.39.

Methyl (4S)-4,5-dihydroxypentanoate (3.10). To a solution of compound 3.9 (6.5g, 37
mmol) in dry THF (55 mL) was added BH3.DMS dropwise maintaining a temperature of
10- 15°C. After stirring at this temperature for 1h, a catalytic amount of NaBH4 (70mg,
1.9 mmol) was added and stirred for an additional 1h maintaining the temperature around
10°C. The reaction mixture was quenched with dry MeOH (20 mL) and stirred for an
additional .5 h. The solvent was removed in vacuo with no further workup and the crude
was purified on silica gel using EtOAc as the eluent to afford the diol 3.10 as viscous
colorless oil (4.7g, 86%).
1
H NMR (400 MHz, CDCl3) δ 3.73 – 3.68 (m, 1H), 3.67 (s, 3H),
COOMe
(S) (S)
HO
COOMe
68
3.62 (dd, J = 11.1, 3.1 Hz, 1H), 3.44 (dd, J = 11.2, 7.1 Hz, 1H), 3.20 (s, 1H), 2.88 (s, 1H),
2.47 (td, J = 7.2, 2.9 Hz, 2H), 1.84 – 1.67 (m, 2H).
13
C NMR (400 MHz, CDCl3) δ 174.74,
71.53, 66.61, 51.92, 30.38, 28.06.

Methyl (4S)-4,5-bis(t-butyldimethylsilyloxy)pentanoate (3.11). To a flask with stir bar
was added TBS-Cl (8.2g, 54.5 mmol), imidazole (3.7g, 54.5 mmol) and DMAP (110 mg,
0.9 mmol) and dissolved in 10 mL dry DMF at 0°C. Diol 3.10 (2.5g, 18 mmol) was
suspended in 5 mL dry DMF and cannulated into the reaction flask at 0°C. The reaction
was allowed to stir overnight at room temperature. It was then quenched with saturated
aqueous NH4Cl (20 mL) and extracted with Et2O (3 x 20 mL). The combined extract was
dried with Na2SO4 and evaporated to give a crude clear oil which was then
chromatographed on silica gel using EtOAc-hexanes mixture of (1:24) as the eluent to
afford the product 3.11 (6.3g, 92%) as a viscous and colorless oil.
1
H NMR (400 MHz,
CDCl3) δ 3.74 – 3.68 (m, 1H), 3.66 (s, 3H), 3.53 (dd, J = 10.0, 5.2 Hz, 1H), 3.39 (dd, J =
10.0, 6.8 Hz, 1H), 2.52 – 2.28 (m, 2H), 2.00 – 1.86 (m, 1H), 1.79 – 1.64 (m, 1H), 0.89 (s,
9H), 0.88 (s, 9H), 0.05 (s, 3H), 0.05 (s, 3H), 0.05 (s, 3H), 0.04 (s, 3H).
13
C NMR (400
MHz, CDCl3) δ 174.47, 72.05, 67.14, 51.61, 29.75, 29.47, 26.10, 26.01, 18.50, 18.25, -
4.18, -4.70, -5.20, -5.24.

69

Methyl (4S)-4-(t-butyldimethylsilyloxy)-5-hydroxypentanoate (3.12).

To a solution of
protected diol 3.11 (7.7g, 20.5mmol) in a 1:1 mixture of dry CH2Cl2/MeOH (50 mL) was
added camphor sulfonic acid (3.8g, 16.38 mmol) at 0°C. The reaction was allowed to stir
for 0.5 h and quenched with Et3N (2.85mL, 20.5 mmol). Avoiding workup, the solvent
was removed in vacuo and purified on silica gel using EtOAc-hexanes mixture (3:7) as
the eluent to afford the product 3.12 (3.1g, 58%) as a viscous and colorless oil.
1
H NMR
(400 MHz, CDCl3) δ 3.85 – 3.77 (m, 1H), 3.67 (s, 3H), 3.59 – 3.48 (m, 1H), 3.52 – 3.41
(m, 1H), 2.47 – 2.29 (m, 2H), 1.94 (t, J = 6.3 Hz, 1H), 1.92 – 1.77 (m, 2H), 0.90 (s, 9H),
0.08 (s, 6H).
13
C NMR (400 MHz, CDCl3) δ 174.25, 71.59, 66.01, 51.78, 29.56, 28.70,
25.95, 18.20, -4.44, -4.55. HRMS (ESI) m/z calcd for C12H27O4Si : [M+H]
+
: 263.1673
found: 263.1673.

Methyl (4S)-4-(tert-butyldimethylsilyloxy)-5-oxopentanoate (3.13). To a -78°C
solution of DMSO (1.32 mL, 17.1 mmol) in anhydrous CH2Cl2 (30 mL) was added
oxalyl chloride (1.01 mL, 11.4 mmol) dropwise. After 0.25 h alcohol 3.12 (1.5g, 5.7
mmol) was added and stirred for 1 h at -78°C. To the reaction mixture was added Et3N
(3.9 mL, 28.5 mmol) and stirred for another 3 h at -78°C. The reaction mixture was
allowed to warm to room temperature and quenched with saturated aqueous NH4Cl (35
70
mL) and extracted with Et2O (3 x 35 mL). The combined extract was dried with Na2SO4
and evaporated to give a crude clear oil which was then chromatographed on silica gel
using EtOAc-hexanes mixture of (1:5) as the eluent to afford the aldehyde 3.13 (1.4g,
98%) as a viscous and colorless oil. 1H NMR (400 MHz, CDCl3) δ 9.59 (d, J = 1.3 Hz,
1H), 4.11 – 4.02 (m, 1H), 3.67 (s, 3H), 2.47 – 2.38 (m, 2H), 2.09 – 1.96 (m, 1H), 1.91 (td,
J = 14.1, 7.6 Hz, 1H), 0.92 (s, 9H), 0.09 (s, 3H), 0.07 (s, 3H).
13
C NMR (400 MHz,
CDCl3) δ 203.46, 173.42, 76.48, 51.81, 28.91, 27.58, 25.85, 18.30, -4.53, -4.93.

Methyl (4S)-4-(t-butyldimethylsilyloxy)-6,6-dibromohex-5-enoate (3.14).

To a
solution of CBr4 (385 mg, 1.16 mmol) at 0°C in anhydrous CH2Cl2 (25 mL total volume)
was cannulated PPh3 (607 mg, 2.32 mmol) to give a clear yellow solution. To the
reaction mixture at 0°C was added aldehyde 3.13 (150 mg, 0.58 mmol). The reaction
was run for 1 h to assure completion. Without workup the solvent was evaporated in
vacuo and the crude mixture was purified on silica gel using EtOAc-hexanes mixture of
(1:9) as the eluent to afford the dibromo ester 3.14 (193 mg, 80%) as a viscous and
colorless oil. 1H NMR (400 MHz, CDCl3) δ 6.37 (d, J = 8.0 Hz, 1H), 4.36 (dt, J = 8.0,
6.4 Hz, 1H), 3.68 (s, 3H), 2.44 – 2.35 (m, 2H), 1.90 – 1.80 (m, 2H), 0.88 (s, 9H), 0.07 (s,
3H), 0.06 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 173.70, 141.43, 89.11, 72.62, 51.79,
31.70, 29.57, 25.89, 18.18, -4.39, -4.91.

71

4S-Methyl-4-(t-butyldimethylsilyloxy)-hex-5-ynoate (3.5). To a solution of dibromo
ester 3.14 (548 mg, 1.32 mmol) at -78°C in anhydrous THF (25 mL) was added 2.0 M
solution of LDA (1.98 mL, 4.0 mmol) drop wise and stirred for 0.5 h. The reaction was
quenched with water (30 mL) and extracted with Et2O (3 x 30 mL), dried using MgSO4,
filtered and concentrated. The crude was then purified using silica gel with a EtOAc-
hexanes eluent of (3:47) to afford the alkyne product 3.5 (233 mg, 69%).
1
H NMR (400
MHz, CDCl3) δ 4.45 (td, J = 6.1, 2.1 Hz, 1H), 3.67 (s, 3H), 2.58 – 2.45 (m, 2H), 2.39 (d,
J = 2.1 Hz, 1H), 2.05 – 1.94 (m, 2H), 0.90 (s, 9H), 0.13 (s, 3H), 0.10 (s, 3H).
13
C NMR
(400 MHz, CDCl3) δ 173.83, 84.78, 72.74, 61.71, 51.73, 33.50, 29.55, 25.87, 18.31, -4.52,
-5.02; HRMS (ESI) m/z calcd for C13H25O3Si : [M+H]
+
: 257.1568 found: 257.1566.

TBS-protected S-gylcidol (3.7). To a solution of TBS-Cl (30.6 g, 203 mmol),
imidazole (13.8 g, 203 mmol) and DMAP (825 mg, 6.8 mmol) dissolved in 250 mL dry
CH2Cl2 at 0°C was added S-glycidol (10.0 g, 135 mmol) at 0°C. The reaction was
allowed to stir overnight at room temperature. It was then quenched with saturated
aqueous NH4Cl (250 mL) and extracted with Et2O (3 x 250 mL). The combined extract
was dried with Na2SO4 and evaporated to give a crude clear oil which was then
chromatographed on silica gel using EtOAc-hexanes mixture of (1:20) as the eluent to
72
afford the product 3.7 (24.3 g, 96%) as a viscous and colorless oil.
1
H NMR (400 MHz,
CDCl3) δ 3.84 (dd, J = 11.9, 3.2 Hz, 1H), 3.65 (dd, J = 12.0, 4.8 Hz, 1H), 3.08 (tt, J = 4.3,
2.8 Hz, 1H), 2.76 (dd, J = 5.2, 4.1 Hz, 1H), 2.63 (dd, J = 5.2, 2.7 Hz, 1H), 0.90 (s, 9H),
0.08 (s, 3H), 0.07 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 79.50 – 75.65 (m), 63.88,
52.56, 44.61, 26.01, -5.17, -5.21.

(R)-1, 2-di-(t-butyldimethylsilyloxy)-5-trimethylsilyl-pent-4-yne (3.16). Part 1: To a
solution of TMS acetylene (5.6 mL, 39.8 mmol) in THF (40 mL) was added 2.5 M n-
BuLi (15.9 mL, 39.8 mmol) at -78°C. After 0.25 h BF3•Et2O (5.0 mL, 39.8 mmol) was
added drop wise at -78°C. To the reaction mixture was added protected glycidol (5.0 g,
26.5 mmol) and stirred for 3 h at -78°C. The reaction mixture was warmed to room
temperature, quenched with saturated aqueous NH4Cl (45 mL) and extracted with Et2O (3
x 45 mL). The organic layer was dried with MgSO4, filtered and the solvent removed in
vacuo. The crude reaction mixture was purified on silica gel using EtOAc-hexanes (4%)
as the eluent to afford 2R,1, (t-butyldimethylsilyloxy)-5-trimethylsilyl-pent-4-yn-2-ol (5.5
g, 73%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 3.78 (dt, J = 11.1, 5.9 Hz,
1H), 3.72 (dd, J = 10.0, 4.1 Hz, 1H), 3.63 (dd, J = 10.0, 5.6 Hz, 1H), 2.51 – 2.41 (m, 2H),
1.64 (s, 1H), 0.91 (s, 11H), 0.14 (s, 2H), 0.09 (s, 5H), 0.08 (s, 6H).
13
C NMR (400 MHz,
CDCl3) δ 102.96 , 87.19 , 70.32, 65.60, 26.03, 24.67, 18.48, 0.17, -5.24, -5.28. Part 2:
To a solution of the product of Part 1 (5.7g, 19.9 mmol) in anhydrous CH2Cl2 (30 mL)
was added TBS-Cl (3.6 g, 23.9 mmol ), imidazole (1.6g, 23.9 mmol), and DMAP (243
73
mg, 1.99 mmol) at 0°C. The reaction was allowed to stir at room temperature overnight.
It was then quenched with saturated aqueous NH4Cl (30 mL) and extracted with Et2O (3
x 30 mL). The combined extract was dried with MgSO4, filtered and evaporated to give a
crude clear oil which was then chromatographed on silica gel using EtOAc-hexanes
mixture of (1:24) as the eluent to afford the product 3.16 (7.5g, 94%) as a viscous and
colorless oil.
1
H NMR (400 MHz, CDCl3) δ 3.86 – 3.75 (m, 1H), 3.60 – 3.47 (m, 2H),
2.50 (dd, J = 16.8, 5.3 Hz, 1H), 2.30 (dd, J = 16.8, 6.5 Hz, 1H), 0.89 (s, 18H), 0.14 (s,
9H), 0.11 (s, 3H), 0.08 (s, 3H), 0.05 (s, 3H), 0.02 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ
104.86, 85.95, 72.25, 66.83, 26.13, 26.01, 18.53, 18.29, 0.23, -2.78, -4.29, -4.44, -5.18, -
5.24.

(R)-2-(t-butyldimethylsilyloxy)-5-trimethylsilyl-pent-4-yn-1-ol (3.17).

To a solution of
protected diol 3.16 (3.4 g, 8.5 mmol) in a 1:1 mixture of dry CH2Cl2/MeOH (20 mL) was
added camphor sulfonic acid (1.0 g, 4.2 mmol) at 0°C. The reaction was allowed to stir
for 0.5 h and quenched with Et3N (1.2 mL, 8.5 mmol). The solvent was removed in
vacuo without work-up and purified on silica gel using EtOAc-hexanes mixture (7%) as
the eluent to afford the product 3.17 (1.9 g, 77%) as a viscous and colorless oil.
1
H NMR
(400 MHz, CDCl3) δ 4.13 – 3.84 (m, 1H), 3.67 (ddd, J = 11.2, 5.8, 3.8 Hz, 1H), 3.57 (ddd,
J = 11.2, 5.8, 3.8 Hz, 1H), 2.51 – 2.33 (m, 3H), 1.88 (dd, J = 7.1, 6.0 Hz, 2H), 0.90 (s,
9H), 0.14 (s, 9H), 0.13 (s, 4H), 0.11 (s, 4H).
13
C NMR (400 MHz, CDCl3) δ 103.47,
86.91, 71.64, 66.04, 25.94, 25.52, 18.20, 0.16, -4.34, -4.59.

74

(R)-2-(t-butyldimethylsilyloxy)-5-trimethylsilyl-pent-4-ynal (3.18). To a -78°C
solution of DMSO (1.3 mL, 18.5 mmol) in anhydrous CH2Cl2 (40 mL) was added oxalyl
chloride (1.1 mL, 12.3 mmol) drop wise. After 0.25 h alcohol 3.17 (1.58 g, 6.2 mmol)
was added and stirred for 1 h at -78°C. To the reaction mixture was added Et3N (4.3 mL,
30.8 mmol) and stirred for another 3 h at -78°C. The reaction mixture was allowed to
warm to room temperature and quenched with saturated aqueous NH4Cl (45 mL) and
extracted with Et2O (3 x 45 mL). The combined extract was dried with Na2SO4 and
evaporated to give a crude clear oil which was then chromatographed on silica gel using
EtOAc-hexanes mixture of (1:20) as the eluent to afford the aldehyde product 3.18 (1.6g,
92%) as a viscous and colorless oil.
1
H NMR (400 MHz, CDCl3) δ 9.63 (d, J = 1.3 Hz,
1H), 4.17 – 4.05 (m, 1H), 2.64 (dd, J = 17.0, 5.0 Hz, 1H), 2.48 (dd, J = 17.0, 7.8 Hz, 1H),
0.94 (s, 9H), 0.14 (s, 12H), 0.13 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 202.25, 101.99,
87.54, 76.25, 31.08, 25.87, 24.65, 18.39, 0.08, -4.57.

(2E, 4R)-4-(t-butyldimethylsilyloxy)-7trimethylsilyl-hept-2-en-6-ynal (3.19).

To a
solution of aldehyde 3.18 (700 mg, 2.46 mmol) dissolved in Toluene (15 mL) was added
(Triphenylphosphoranylidene)acetaldehyde (750 mg, 2.46 mmol) and heated to reflux
75
overnight. The solvent was removed in vacuo without work up and the crude mixture
was purified on silica gel using a EtOAc-hexanes mixture of (1:20) as the eluent to afford
the product 3.19 (634 mg, 83%) as a clear viscous oil.
1
H NMR (400 MHz, CDCl3) δ
9.60 (d, J = 8.0 Hz, 1H), 6.90 (dd, J = 15.5, 4.3 Hz, 1H), 6.31 (ddd, J = 15.5, 8.0, 1.4 Hz,
1H), 4.54 (ddd, J = 11.8, 6.1, 1.4 Hz, 1H), 2.55 (dd, J = 16.6, 6.2 Hz, 1H), 2.44 (dd, J =
16.6, 7.4 Hz, 1H), 0.91 (s, 9H), 0.15 (s, 9H), 0.11 (s, 3H), 0.07 (s, 3H).
13
C NMR (400
MHz, CDCl3) δ 193.56, 157.92, 131.40, 102.21, 70.66, 29.11, 25.87, 18.30, 0.11, -4.63, -
4.69.

(3S, 1E, 4Z)-3-(t-butyldimethylsilyloxy)-1-iodoocta-1,5-diene (3.4). To a solution of
CrCl2 (2.0 g, 16.1 mmol) dissolved in THF (65 mL total volume) was cannulated a
mixture of aldehyde 3.19 (560 mg, 1.8 mmol) and CHI3 (3.2 g, 8.05 mmol) dissolved in
anhydrous THF (10 mL) under Argon at 0°C. The reaction was stirred at 0°C for 3 h and
an additional 1 h at room temperature. The reaction mixture was quenched with water
(80 mL) extracted with Et2O (3 x 80 mL) rinsed with brine and dried over MgSO4. The
organic phase was filtered and the solvent was removed in vacuo to afford a crude oil
which was purified on silica gel using first pure pentanes and then EtOAc-hexanes (1:24)
as the eluent to afford a 9:1 mixture of compound 3.4 and its E, Z stereoisomer (see
inset) as a clear colorless oil (660 mg, 84%).
1
H NMR (400 MHz, CDCl3) δ 7.02 (dd, J =
14.5, 10.7 Hz, 1H), 6.32 (d, J = 14.5 Hz, 1H), 6.15 (ddd, J = 15.2, 10.8, 1.2 Hz, 1H), 5.78
(dd, J = 15.2, 5.7 Hz, 1H), 4.27 (d, J = 6.6 Hz, 1H), 2.49 – 2.29 (m, 2H), 0.90 (s, 9H),
76
0.14 (s, 9H), 0.09 (s, 3H), 0.05 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 144.72, 136.49,
129.69, 103.74, 86.78, 79.23, 71.48, 29.90, 25.97, 18.39, 0.20, -4.42, -4.60.

TBS Protected R-Glycidol (3.6). To a solution of TBS-Cl (15.3 g, 101.5 mmol),
imidazole (6.9 g, 101.5 mmol) and DMAP (412 mg, 3.4 mmol) dissolved in 125 mL dry
CH2Cl2 at 0°C was added R-glycidol (5.0 g, 67.5 mmol) at 0°C. The reaction was
allowed to stir overnight at room temperature. It was then quenched with saturated
aqueous NH4Cl (125 mL) and extracted with Et2O (3 x 125 mL). The combined extract
was dried with Na2SO4 and evaporated to give a crude clear oil which was then
chromatographed on silica gel using EtOAc-hexanes mixture of (1:20) as the eluent to
afford the protected glycidol 3.6 (12.2 g, 96%) as a viscous and colorless oil.
1
H NMR
(400 MHz, CDCl3) δ 3.84 (dd, J = 11.9, 3.2 Hz, 1H), 3.65 (dd, J = 11.9, 4.8 Hz, 1H), 3.13
– 3.01 (m, 1H), 2.76 (dd, J = 5.2, 4.0 Hz, 1H), 2.63 (dd, J = 5.2, 2.7 Hz, 1H), 0.90 (s, 9H),
0.08 (s, 3H), 0.07 (s, 3H).
13
C NMR (400 MHz, CDCl3) 63.88 52.56, 44.60, 26.01,
18.50, -5.17, -5.21 .

2S, 1-(t-butyldiphenylsilyloxy)hept-4-yn-2-ol (3.21a). To a flask at -78°C was added 1-
butyne (0.29 g, 5.3 mmol). n-BuLi (2.12 mL, 5.3 mmol) was added next, dropwise at -
78°C. After 0.25 h BF3 Et2O (0.64 mL, 5.3 mmol) was added drop wise at -78°C. To
77
the reaction mixture was added protected glycidol (0.5 g, 2.65 mmol) and stirred for 3 h
at -78°C. The reaction mixture was warmed to room temperature, quenched with
saturated aqueous NH4Cl (15 mL) and extracted with Et2O (3 x 15 mL). The organic
layer was dried with MgSO4, filtered and the solvent removed in vacuo. The crude
reaction mixture was purified on silica gel using EtOAc-hexanes (4%) as the eluent to
afford compound 3.21a (600 mg, 94%) as a clear colorless oil.
1
H NMR (400 MHz,
CDCl3) δ 3.77 – 3.72 (m, 1H), 3.70 (dd, J = 9.9, 4.2 Hz, 1H), 3.60 (dd, J = 9.8, 5.8 Hz,
1H), 2.46 (d, J = 4.8 Hz, 1H), 2.42 – 2.33 (m, 2H), 2.22 – 2.10 (m, 2H), 1.11 (t, 3H), 0.90
(s, 9H), 0.08 (s, 6H).
13
C NMR (400 MHz, CDCl3) δ 84.15, 75.21, 70.63, 65.81, 26.03,
23.55, 18.46, 14.33, 12.55, -5.23, -5.26.

(S), 1-(t-butyldiphenylsilyloxy)-2-(t-butyldiphenylsilyloxy)hept-4-yne (3.21). To a
flask with imidazole (95 mg, 1.39 mmol) and DMAP (8 mg, 0.06 mmol) in CH2Cl2 (5
mL total volume) was added TBDPS-Cl (0.36 mL, 1.39 mmol) dropwise at 0°C. The
alcohol 3.21a (280 mg, 1.15 mmol) was cannulated to the flask and stirred overnight at
room temperature. The reaction mixture was quenched with saturated aqueous NH4Cl (7
mL) and extracted with Et2O (3 x 7 mL). The organic layer was dried with MgSO4,
filtered and the solvent removed in vacuo. The crude reaction mixture was purified on
silica gel using EtOAc-hexanes (1%) as the eluent to afford the protected diol 3.21 (525
mg, 9%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 7.85 – 7.65 (m, 4H), 7.55
– 7.29 (m, 6H), 3.83 (p, J = 5.5 Hz, 1H), 3.55 (dd, J = 5.4, 3.3 Hz, 2H), 2.44 – 2.34 (m,
78
1H), 2.32 – 2.23 (m, 1H), 2.12 (qt, J = 7.5, 2.4 Hz, 2H), 1.10 (t, 3H), 1.07 (s, 9H), 0.84 (s,
9H), -0.03 (s, 3H), -0.06 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 136.10, 136.03, 134.47,
134.34, 129.69, 129.66, 127.62, 83.29, 76.47, 72.78, 65.80, 27.09, 26.05, 24.12, 19.54,
18.46, 14.34, 12.62, -5.34, -5.35.

(S), 2-(t-butyldiphenylsilyloxy)hept-4-yn-1-ol (3.22). To a solution of protected diol
3.21 (0.5 g, 1.04 mmol) in CH2Cl2 (5 mL) was added camphor sulfonic acid (144 mg,
0.62 mmol) at room temperature and monitored for 1 h. The reaction was quenched with
Et3N (0.09 mL, 0.62 mmol) and the solvent was evaporated in vacuo without workup.
The crude reaction mixture was purified on silica gel using EtOAc-hexanes (1:8) as the
eluent to afford alcohol 3.22 (370 mg, 97%) as a clear colorless oil.
1
H NMR (400 MHz,
CDCl3) δ 7.82 – 7.72 (m, 4H), 7.57 – 7.37 (m, 6H), 4.06 – 3.87 (m, 1H), 3.70 (d, J = 4.6
Hz, 2H), 2.47 (ddt, J = 16.4, 7.7, 2.5 Hz, 1H), 2.36 (ddt, J = 16.4, 4.9, 2.4 Hz, 1H), 2.18 –
2.06 (m, 3H), 1.16 (s, 9H), 1.10 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ
135.86, 135.71, 133.64, 133.62, 129.89, 129.85, 127.80, 127.68, 83.83, 75.43, 72.62,
65.56, 27.01, 23.91, 19.33, 14.10, 12.41.

(2S, 4Z)-2-(t-butyldiphenylsilyloxy)hept-4-en-1-ol (3.23). To a solution of alcohol 3.22
(3.9 g, 10.2 mmol) in EtOAc (200 mL) was added Lindlar catalyst (200 mg) and 5 drops
of quinoline. The reaction mixture was placed under a H2 atmosphere and stirred for 2 h.
79
The reaction was filtered through celite and the solvent was removed in vacuo. The
crude product was purified on silica gel using EtOAc-hexanes (1:8) as an eluent to afford
the alcohol 3.23 (3.5 g, 94%) as a clear colorless oil. 1H NMR (400 MHz, CDCl 3) δ 7.89
– 7.56 (m, 4H), 7.51 – 7.30 (m, 6H), 5.43 – 5.28 (m, 1H), 5.17 (dtt, J = 10.7, 7.6, 1.5 Hz,
1H), 3.79 (dtd, J = 8.5, 4.8, 3.6 Hz, 1H), 3.63 – 3.43 (m, 2H), 2.36 – 2.23 (m, 1H), 2.22 –
2.10 (m, 1H), 1.91 – 1.73 (m, 2H), 1.09 (s, 9H), 0.84 (t, J = 7.5 Hz, 3H).
13
C NMR (400
MHz, CDCl3) δ 136.03, 135.85, 134.42, 133.97, 133.93, 129.97, 129.93, 127.91, 127.80,
123.77, 74.00, 65.74, 31.69, 27.19, 20.61, 19.49, 14.24.

2S, 4Z, 2-(t-butyldiphenylsilyloxy)-1-oxohept-4-enal (3.24). To a solution of DMSO
(0.43 mL, 3.7 mmol) in CH2Cl2 (15 mL) was added oxalyl chloride (0.33 mL, 3.7 mmol)
at -78°C. To the reaction mixture was cannulated alcohol 3.23 (0.5 g, 1.87 mmol) and
stirred for 1 h. Et3N was added to the reaction at -78°C and stirred for 3 h. The reaction
mixture was quenched with saturated aqueous NH4Cl (20 mL) and extracted with Et2O (3
x 20 mL). The Et2O was washed with brine, dried with MgSO4 and filtered. The solvent
was removed in vacuo and the crude product was purified on silica gel using EtOAc-
Hexanes (1:10) as the eluent to afford the aldehyde 3.24 (0.44 g, 90%) as a colorless oil.
1H NMR (400 MHz, CDCl3) δ 9.57 (d, J = 1.7 Hz, 2H), 7.71 – 7.61 (m, 4H), 7.49 – 7.33
(m, 6H), 5.52 – 5.41 (m, 1H), 5.40 – 5.27 (m, 1H), 4.06 (td, J = 6.5, 1.7 Hz, 1H), 2.44 (dt,
J = 14.1, 6.7 Hz, 1H), 2.34 (dt, J = 13.9, 6.5 Hz, 1H), 1.99 – 1.87 (m, 2H), 1.12 (s, 9H),
0.90 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ 203.52, 135.95, 135.95, 135.11,
80
133.24, 133.12, 130.18, 130.13, 127.95, 127.89, 122.17, 77.94, 31.15, 27.08, 20.74, 19.49,
14.14.

3S, 1E, 4Z, 3-(t-butyldimethylsilyloxy)-1-iodoocta-1,5-diene (3.3). Part 1: To a
solution of CrCl2 (3.4 g, 27.3 mmol) dissolved in THF (40 mL total volume) was
cannulated a mixture of aldehyde 3.24 (1.0 g, 2.7 mmol) and CHI3 (5.4 g, 13.7 mmol)
dissolved in anhydrous THF (10 mL) under Argon at 0°C. The reaction was stirred at
0°C for 3 h and an additional 1 h at room temperature. The reaction mixture was
quenched with water (50 mL) extracted with Et2O (3 x 50 mL) rinsed with brine and
dried over MgSO4. The organic phase was filtered and the solvent was removed in vacuo
to afford a crude oil which was purified on silica gel using first pure pentanes and then
EtOAc-hexanes (1:24) as the eluent to afford TBDPS protected vinyl iodide (3S, 1E, 5Z)-
3-(t-Butyldiphenylsilyloxy)-1-iodoocta-1,5-diene (780 mg, 59%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 7.76 – 7.54 (m, 4H), 7.51 – 7.32 (m, 6H), 6.49 (dd, J =
14.4, 6.5 Hz, 1H), 5.98 (dd, J = 14.4, 1.1 Hz, 1H), 5.49 – 5.32 (m, 1H), 5.32 – 5.13 (m,
1H), 4.17 – 3.98 (m, 0H), 2.32 – 2.07 (m, 2H), 1.84 (dtd, J = 9.2, 7.4, 5.6 Hz, 2H), 1.07 (s,
9H), 0.87 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ 148.04, 136.03, 136.01,
134.54, 133.97, 133.65, 129.90, 129.87, 127.74, 127.72, 123.18, 76.88, 75.93, 35.31,
27.14, 20.75, 19.48, 14.33. Part 2: To a solution of the TBDPS-protected vinyl iodide 19
(0.8 g, 1.63 mmol) in THF (5 mL) was added 1.0 M solution of TBAF (1.63 mL, 1.63
mmol) at 0°C and stirred for 2 h. The reaction was quenched with water (10 mL) and
81
extracted with Et2O (5 x 10 mL), rinsed with brine, dried over MgSO4 and filtered. The
crude reaction mixture was purified on silica gel using EtOAc-hexanes (10%) as the
eluent to afford the alcohol (3S, 1E, 5Z)-1-iodoocta-1,5-diene-3-ol (0.38 g, 92%) as a
clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 6.60 (dd, J = 14.4, 5.8 Hz, 1H), 6.37
(dd, J = 14.5, 1.3 Hz, 1H), 5.73 – 5.52 (m, 1H), 5.45 – 5.25 (m, 1H), 4.23 – 3.98 (m, 1H),
2.31 (ddd, J = 7.7, 6.5, 1.6 Hz, 2H), 2.14 – 1.95 (m, 2H), 0.98 (t, J = 7.5 Hz, 3H).
13
C
NMR (400 MHz, CDCl3) δ 147.92, 134.93, 122.82, 77.48, 77.38, 77.16, 76.84, 74.01,
34.68, 20.89, 14.35. Part 3: The alcohol (0.38 g, 1.5 mmol) was protected using TBS-
OTf (0.54 mL, 3.0 mmol) and 2,6-lutidine (0.52 mL, 4.5 mmol) in anhydrous CH2Cl2 (20
mL). The reaction mixture was quenched after 2 h with saturated aqueous NH 4Cl (20
mL) and extracted with Et2O (3 x 20 mL). The Et2O was rinsed with brine, dried with
MgSO4 and filtered. The solvent was removed in vacuo and the crude product was
purified on silica gel using EtOAc-Hexanes (1:49) as the eluent to afford the silylated
vinyl iodide 5 as a colorless oil (0.54 g, 98%).
1
H NMR (400 MHz, CDCl3) δ 6.54 (dd, J
= 14.3, 5.7 Hz, 1H), 6.21 (dd, J = 14.3, 1.5 Hz, 1H), 5.54 – 5.42 (m, 1H), 5.37 – 5.24 (m,
1H), 4.17 – 4.00 (m, 1H), 2.33 – 2.18 (m, 2H), 2.03 (dt, J = 14.9, 7.4 Hz, 1H), 0.96 (t, J =
7.5 Hz, 4H), 0.89 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ 148.90, 134.39, 123.75, 75.86,
75.20, 35.70, 25.96, 20.90, 18.37, 14.37, -4.48, -4.68.

Methyl 4S, 11R, 7E, 9E, 4,11-bis(t-butyldimethylsilyloxy)-14-(trimethylsilyl)-
tetradeca-7,9-dien-5,13-diynoate (3.25). To the arm of a three-necked flask was
82
charged Pd(PPh3)4 (45 mg, 0.039 mmol) and CuI (10 mg, 0.078 mmol) under Argon. A
solution of alkyne 3.5 (100 mg, 0.39 mmol) and the dienyl iodide 3.4 mixture (191mg,
0.43 mmol) and Et3N (0.64 mL, 3.9 mmol) in C6H6 (3 mL) was cannulated into the
reaction vessel. The reaction flask was then freeze thawed with liquid nitrogen three
times to remove an oxygen. After removing any oxygen from the reaction flask the
Pd(PPh3)4 and CuI was added and the reaction mixture was stirred overnight at room
temperature. The reaction was worked up with aqueous saturated NH4Cl (5 mL) and
extracted with Et2O (3 x 5 mL). The solvent was evaporated and the mixture was
purified on silica gel using EtOAc-hexanes (1:24) as the eluent to afford compound 3.25
as a clear oil (202 mg, 92%). The E, Z stereoisomer could be isolated during the
separation.
1
H NMR (400 MHz, CDCl3) δ 6.53 (dd, J = 15.6, 10.9 Hz, 1H), 6.23 (dd, J =
15.2, 10.9 Hz, 1H), 5.83 (dd, J = 15.3, 5.9 Hz, 1H), 5.58 (d, J = 15.5 Hz, 1H), 4.58 (td, J
= 6.2, 1.8 Hz, 1H), 4.32 (q, J = 6.5 Hz, 1H), 3.67 (s, 3H), 2.49 (td, J = 7.6, 1.4 Hz, 2H),
2.48 – 2.29 (m, 2H), 2.07 – 1.94 (m, 2H), 0.90 (s, 18H), 0.14 (s, 9H), 0.13 (s, 3H), 0.10 (s,
3H), 0.09 (s, 3H), 0.05 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 173.94, 141.17, 138.13,
129.15, 110.92, 103.84, 92.70, 86.72, 83.92, 71.78, 62.50, 51.70, 33.65, 30.04, 29.75,
25.97, 25.95, 18.39, 18.36, 0.20, -4.37, -4.40, -4.60, -4.92.

Methyl (4S, 11R, 7E, 9E)-4,11-bis(t-butyldimethylsilyloxy-tetradeca-7,9-dien-5,13-
diynoate (3.25b). To a solution of compound 3.25 (80 mg, 0.14 mmol) in MeOH (3 mL)
was added Na2CO3 (20 mg) and stirred overnight at room temperature. The solvent was
83
removed in vacuo and the crude was dissolved in water (5 mL) and extracted with Et 2O
(3 x 5 mL). The oil was purified on silica gel using EtOAc-hexanes (1:49) as the eluent
to afford compound 3.25b (63 mg, 91%) as a colorless oil.
1
H NMR (500 MHz, CDCl3)
δ 6.54 (dd, J = 15.4, 11.0 Hz, 1H), 6.26 (dd, J = 15.1, 11.1 Hz, 1H), 5.86 (dd, J = 15.3,
5.7 Hz, 1H), 5.60 (dd, J = 15.6, 1.6 Hz, 1H), 4.58 (td, J = 6.0, 1.6 Hz, 1H), 4.42 – 4.30 (m,
1H), 3.67 (s, 3H), 2.49 (td, J = 7.2, 1.8 Hz, 2H), 2.48 – 2.38 (m, 1H), 2.33 (ddd, J = 16.4,
7.1, 2.7 Hz, 1H), 2.06 – 1.94 (m, 2H), 1.99 (t, J = 2.7 Hz, 1H), 0.90 (s, 18H), 0.13 (s, 3H),
0.10 (s, 3H), 0.08 (s, 3H), 0.05 (s, 3H).
13
C NMR (500 MHz, CDCl3) δ 173.93, 141.08,
137.75, 129.35, 111.12, 92.78, 83.87, 81.04, 71.49, 70.37, 62.48, 51.72, 33.66, 29.75,
28.59, 25.95, 25.94, 18.39, 18.36, -4.35, -4.44, -4.68, -4.92; HRMS (ESI) m/z calcd for
C27H46NaO4Si2 : [M+Na]
+
: 513.2827 found: 513.2838.

Methyl (4S, 11R, 17S, 7E, 9E, 15E, 19Z)-tris-(tert-butyldimethylsilyloxy)-docosa-
7,9,15,19-tetraen-5,13-diynoate (3.26). To the arm of a three-necked flask was charged
Pd(PPh3)4 (5 mg, .0039 mmol) and CuI (1 mg, .078 mmol) under Argon. A solution of
compound alkyne 3.25b (19 mg, 0.039 mmol), vinyl iodide 3.3 (21 mg, 0.058 mmol) and
Et3N (0.05 mL, .39 mmol) in C6H6 (1 mL) was cannulated into the reaction vessel. The
reaction flask was then freeze thawed with liquid nitrogen three times to remove an
oxygen. After removing any oxygen from the reaction flask the Pd(PPh3)4 and CuI was
84
added and the reaction mixture was stirred overnight at room temperature. The reaction
was worked up with aqueous saturated NH4Cl (3 mL) and extracted with Et2O (3 x 3 mL).
The solvent was evaporated and the mixture was purified on silica gel using EtOAc-
hexanes (3:97) as the eluent to afford the protected triol 3.26 (26 mg, 92%) as a clear oil.
1
H NMR (500 MHz, CDCl3) δ 6.54 (dd, J = 15.5, 10.9 Hz, 1H), 6.25 (dd, J = 15.3, 10.9
Hz, 1H), 6.04 (dd, J = 15.7, 5.4 Hz, 1H), 5.86 (dd, J = 15.3, 5.7 Hz, 1H), 5.73 – 5.55 (m,
2H), 5.51 – 5.40 (m, 1H), 5.40 – 5.26 (m, 1H), 4.58 (td, J = 6.0, 1.6 Hz, 1H), 4.33 (q, J =
5.5 Hz, 1H), 4.24 – 4.11 (m, 1H), 3.67 (s, 3H), 2.61 – 2.35 (m, 4H), 2.23 (dp, J = 20.8,
7.0 Hz, 2H), 2.11 – 1.95 (m, 4H), 0.95 (t, J = 7.6 Hz, 3H), 0.90 (s, 18H), 0.89 (s, 9H),
0.13 (s, 3H), 0.10 (s, 3H), 0.08 (s, 3H), 0.05 (s, 3H), 0.03 (s, 3H).
13
C NMR (500 MHz,
CDCl3) δ 173.92, 145.23, 141.21, 138.25, 133.99, 129.05, 124.28, 110.89, 109.15, 92.68,
87.10, 83.92, 80.71, 72.75, 71.83, 62.49, 51.71, 36.07, 33.67, 29.77, 29.75, 29.62, 26.00,
25.95, 25.94, 20.89, 18.41, 18.39, 14.34, -4.35, -4.41, -4.46, -4.65, -4.65, -4.92.

(4S, 11R, 17S)-Trihydroxydocosa-(7E, 9E, 15E, 19Z)-tetraene-5,13-diynoic acid
(3.26b Acid). To a solution of compound 3.26 (10 mg, 0.014 mmol) in THF (2 mL) was
added 1.0 M solution of TBAF (0.084 mL, 0.084 mmol) at 0°C and stirred for 2 h. The
reaction was quenched with water (3 mL) and extracted with Et2O (5 x 3 mL), rinsed
with brine, dried over MgSO4 and filtered. The solvent was the concentrated and freshly
prepared CH2N2 was added to convert any acid to the ester. The solvent was completely
OH
(S) (S)
(R) (R)
HO
(S) (S)
HO
COOH
4
17
11
85
removed in vacuo and the compound was purified on silica gel using MeOH-CH2Cl2
(3%) as the eluent to afford an ester/lactone mixture (3:1 ratio see HPLC and NMR data
attached). The product was then suspended in a H2O-MeOH mixture (1:1, 1 mL) and
NaOH (1 mg, 2.5 x 10
-2
mmol) was added. After 3 h the reaction mixture was dried and
purified via C-18 reversed Phase HPLC using H2O-MeOH mixture (41%) to afford
compound 3.26b (1.5 mg, 27%) as colorless oil.
1
H NMR (400 MHz, CD3OD) δ 6.59
(dd, J = 15.5, 10.8 Hz, 1H), 6.36 (dd, J = 15.4, 10.9 Hz, 1H), 6.03 (dd, J = 15.8, 6.1 Hz,
1H), 5.89 (dd, J = 15.1, 6.1 Hz, 1H), 5.68 (t, J = 14.9 Hz, 2H), 5.55 – 5.45 (m, 1H), 5.42
– 5.33 (m, 1H), 4.52 (dd, J = 6.7, 5.0 Hz, 1H), 4.30 – 4.21 (m, 1H), 4.08 (q, J = 5.9 Hz,
1H), 2.57 – 2.48 (m, 1H), 2.39 (td, J = 15.6, 7.5 Hz, 3H), 2.28 (q, J = 7.5 Hz, 2H), 2.06
(dq, J = 12.3, 6.8, 6.0 Hz, 2H), 1.97 (q, J = 7.2 Hz, 2H), 0.98 (t, J = 7.5 Hz, 3H).
13
C
NMR (400 MHz, CD3OD) δ 180.57, 144.28, 139.62, 136.98, 133.44, 123.68, 110.86,
109.50, 92.69, 85.94, 82.80, 79.96, 71.34, 70.10, 61.98, 61.94, 34.44, 34.20, 33.56, 27.53,
20.22, 13.08; HRMS (ESI) m/z calcd for C22H27O5
-
: [M
-
]: 371.1864 found: 371.1881.

(4S, 11R, 17S)-Trihydroxydocosa-(5Z, 7E, 9E, 13Z, 15E, 19Z)-hexaenoic acid, or
Resolvin D3 (3.1). A flame dried flask was charged with a freshly prepared Zn/Cu/Ag
amalgam (300 mg, excess) and suspended in H2O-MeOH mixture (1:1, 1 mL). To the
reaction slurry was added compound 3.26b (1.5 mg, 3.8 x 10
-3
mmol) and stirred for 13 h
OH
(S) (S)
(R) (R)
HO
(S) (S)
HO
COOH
86
while monitoring. The reaction was filtered dried and purified via HPLC at H 2O-MeOH
mixture (41%) to afford compound 3.1 (0.33 mg, 22%).
1
H NMR (600 MHz, CD3OD) δ
6.60 (dd, J = 14.6, 11.5 Hz, 1H), 6.52 (dd, J = 15.4, 11.1 Hz, 1H), 6.34 (dd, J = 14.9, 10.8
Hz, 1H), 6.24 (dd, J = 14.6, 10.7 Hz, 1H), 6.13 – 6.02 (m, 2H), 5.75 (dd, J = 15.2, 6.7 Hz,
1H), 5.70 (dd, J = 15.4, 6.5 Hz, 1H), 5.55 – 5.31 (m, 4H), 4.92 (s, 24H), 4.23 – 4.08 (m,
2H), 2.45 (q, J = 7.9 Hz, 2H), 2.38 – 2.22 (m, 4H), 2.07 (q, J = 7.3 Hz, 2H), 1.88 (dd, J =
14.1, 7.2 Hz, 1H), 1.82 – 1.71 (m, 1H), 0.98 (t, J = 7.5 Hz, 3H).
13
C NMR (600 MHz,
CD3OD) δ 182.46, 137.54, 137.49, 135.48, 134.75, 134.58, 131.75, 131.04, 130.31,
129.15, 128.17, 126.58, 125.52, 73.20, 73.07, 68.86, 36.68, 36.23, 35.43, 35.28, 21.68,
14.54; HRMS (ESI) m/z calcd for C22H31O5 : [M
-
]: 375.2177 found: 375.2182.

(4S, 11R, 17R)-Trihydroxydocosa-(5Z, 7E, 9E, 13Z, 15E, 19Z)-hexaenoic acid, or AT
Resolvin D3 (3.2). This compound was prepared similarly to its 17S-epimer, Resolvin
D3 (3.1). The removal of the silyl protective groups led to a similar mixture of hydrolysis
products, which were similarly converted to 2.
1
H NMR (600 MHz, CD3OD) δ 6.59 (dd,
J = 14.7, 11.5 Hz, 1H), 6.52 (dd, J = 15.2, 11.2 Hz, 1H), 6.33 (dd, J = 15.1, 10.7 Hz, 1H),
6.24 (dd, J = 14.7, 10.9 Hz, 1H), 6.08 (td, J = 11.1, 4.3 Hz, 2H), 5.75 (dd, J = 15.1, 6.6 Hz,
1H), 5.70 (dd, J = 15.1, 6.6 Hz, 1H), 5.48 (td, J = 11.1, 5.8 Hz, 2H), 5.45 – 5.37 (m, 2H),
4.61 (q, J = 7.2 Hz, 1H), 4.19 – 4.16 (m, 1H), 4.14 (q, J = 6.7 Hz, 1H), 2.52 – 2.38 (m,
OH
(R) (R)
(R) (R)
HO
(S) (S)
HO
COOH
4
17
11
87
2H), 2.30 (dq, J = 29.2, 7.3 Hz, 4H), 2.08 (dt, J = 14.4, 7.1 Hz, 2H), 1.87 (dt, J = 15.0, 7.3
Hz, 1H), 1.77 (dt, J = 13.7, 6.7 Hz, 1H), 0.98 (t, J = 7.5 Hz, 3H).
13
C NMR (600 MHz,
CD3OD) δ 182.45, 137.54, 137.49, 135.48, 134.75, 134.58, 131.76, 131.04, 130.31,
129.15, 128.17, 126.58, 125.52, 73.20, 73.07, 68.86, 49.51, 49.34, 49.17, 49.00, 48.83,
48.66, 48.49, 36.68, 36.23, 35.43, 35.28, 21.68, 14.54; HRMS (ESI) m/z calcd for
C22H31O5 : [M
-
]: 375.2177 found: 375.2185.

88
3.6 References

55. Serhan, C. N.; Petasis, N. A. Chemical Reviews 2011, 111, 5922.
56. Seki, H.; Sasaki, T.; Ueda, T.; Arita, M. The Scientific World Journal 2010, 10, 818.
57. Serhan, C. N.; Clish, C. B.; Brannon, J.; Colgan, S. P.; Chiang, N.; Gronert,
K. Journal of Experimental Medicine 2000, 192, 1197.
58. Serhan, C. N.; Hong, S.; Gronert, K.; Colgan, S. P.; Devchand, P. R.; Mirick, G.;
Moussignac, R. L. Journal of Experimental Medicine 2002, 196, 1025.
59. Serhan, C. N.; Savill, J. Nature Immunology 2005, 6, 1191.
60. Serhan, C. N.; Brain, S. D.; Buckley, C. D.; Gilroy, D. W.; Haslett, C.; O'Neill, L. A.
J.; Perretti, M.; Rossi, A. G.; Wallace, J. L. Faseb Journal 2007, 21, 325.
61. Serhan, C. N.; Krishnamoorthy, S.; Recchiuti, A.; Chiang, N. Current Topics in
Medicinal Chemistry 2011, 11, 629.
62. Duffield, J. S.; Hong, S.; Vaidya, V. S.; Lu, Y.; Fredman, G.; Serhan, C. N.;
Bonventre, J. V. Journal of Immunology 2006, 177, 5902.
63. Petasis, N. A.; Winkler, J.; Nagengast, E. S.; Uddin, J.; Serhan, C. N. Abstracts of
Papers of the American Chemical Society, 2009, 237.
64. Winkler, J. W.; Uddin, J.; Serhan, C. N.; Petasis, N. A. Organic Letters 2013, 15,
1424.
65. Saito, S.; Ishikawa, T.; Kuroda, A.; Koga, K.; Moriwake, T. Tetrahedron 1992, 48,
4067.
66. Ishiiwa, T.; Kuroda, A. Tetrahedron, 1992, 48, 4067–4086.
67. Fox, M. E.; Jackson, M.; Lennon, I. C.; McCague, R. Journal of Organic
Chemistry 2005, 70, 1227.
89

68. Corey, E. J.; Fuchs, P. L. Tetrahedron Letters 1972, 3769.
69. Lu, W.; Zheng, G. R.; Gao, D. X.; Cai, J. C. Tetrahedron 1999, 55, 7157.
70. Ohira, S. Synthetic Communications 1989, 19, 561.
71. Muller, S.; Liepold, B.; Roth, G. J.; Bestmann, H. J. Synlett 1996, 521.
72. Roth, G. J.; Liepold, B.; Muller, S. G.; Bestmann, H. J. Synthesis-Stuttgart 2004, 59.
73. Unpublished observations by the Petasis lab.
74. Takai olefination would afford a 9:1 mixture of E E to E Z , however storing
compound (3.4) for longer than a day would result in isomerization to a 60:40 E, Z
mixture.
75. See data section page for NMR and HPLC data showing 60:40 mixture of
Ester/Lactone.
76. March J. Advanced organic chemistry. 4th ed. New York: John Wiley & Sons.
77. Frost, A.A.; Pearson R.G. Kinetics and mechanism. A study of homogeneous
chemical reactions. 2nd ed. New York: John Wiley and Sons, 1961; 317–34.
78. Hennessy, S. A.; Moane, S. M.; McDermott, S. D. Journal of Forensic
Sciences 2004, 49, 1220.
79. Boland, W.; Sieler, C.; Feigel, M. Helvetica Chimica Acta 1987, 70, 1025.
80. Chemin, D.; Linstrumelle, G. Tetrahedron 1992, 48, 1943.
81. Complete Stereochemsitry of compound (1) was confirmed by2-D COSY NMR
studies on a 600 MhZ Magnet.
82. Dalli, J.; Winkler, J. W.; Colas, R. A.; Arnardottir, H.; Cheng, C.-Y. C.; Chiang, N.;
Petasis, N. A.; Serhan, C. N. Chemistry & Biology 2013, 20, 188.
83. Haeggstrom, J. Z.; Hamberg, M. Chemistry & Biology 2013, 20, 138.
90
CHAPTER 4. First Total Synthesis and Stereochemical Assignment of Resolvin D4
and its Aspirin Triggered Analogue (RvD4 and AT-RvD4)
4.1 Introduction
The ever increasing body of work on omega 3’s and their endogenous metabolites
demonstrates their wider reaching role in inflammation and disease than previously
thought before.
84
Of the class of D series resolvin compounds discovered by Serhan and
coworkers in this chapter we detail our latest efforts towards the total synthesis of
resolvin D4. Despite the similarities in structure the D-series resolvins each exhibit a
unique temporal profile in different stages and types of inflammation. As a result some
are seemingly more abundant than others. Due to the nature of these compounds being
detectable at only sub nano-molar concentration, there was a good deal of effort to
synthesize the entire family in order to confirm their absolute stereochemistry as well as
differentiate between their structure and various inflammatory pathways. RvD3 and
RvD4 were initially found to be much less abundant than RvD1 and RvD2.
85
After a
tremendous effort was made to synthesize these unique compounds with their gamma-
hydroxy group adjacent to the free acid we found that the nature of these compounds
could have been overlooked to a large extent due to their ability to lactonize into a five-
membered gamma-butyrolactone. This leaves to question the extent of their role in
biological systems and how they may have similar or different effects in comparison to
their better studied counterparts, resolvin D1 and resolvin D2. Of this family of
compounds, thus far RvD4 has been the least recognizable as well as the littlest known.
Despite its limited detectability RvD4 has shown some promising results regarding it’s
91
biological activity. In addition to its role in mitigating acute kidney injury recent
investigations have shown it may potentially help mediate an inflammatory response in
Alzheimers, similarly to its resolvin counterparts (RvE1 which reduces the
proinflammatory gene expression of IL-12/p40
86
and RvD2 which reduces levels of
proinflammatory cytokines). Resolvin D4 also likely plays a role in the acute local
impact and long range responses triggered in the resolution response to abdominal aortic
aneurysm surgery.
87
Resolvin D4 is formed similarly to Resolvin D3 through subsequent oxygenation
pathways by Lipoxygenase enzymes (Figure 15). After the second oxygenation forms
4S-Hydroperoxy-17S-hydroxy-DHA enzymatic epoxidation converts this di-hydroxyl
DHA intermediate to the believed 4S,5S-Epoxy-17S-hydroxy-DHA. Their divergence

Figure 15. Biosynthesis of RvD4 (4.1) and AT-RvD4 (4.2).
92

occurs at this point in which the epoxy ring is opened through enzymatic hydrolysis.
Most likely this transformation occurs from the C5 position due to the positively charged
intermediate being stabilized by the conjugated triene system. This can also occur
through an alternative pathway involving COX-2 in the presence of aspirin forming the
17R aspirin triggered analogue of resolvin D4. It is our aim to establish the absolute
stereochemistry through our controlled synthetic strategy and herein we report the total
synthesis of Resolvin D4 (4S, 5R, 17S, 6E, 8E, 10Z, 13Z, 15E, 19Z)-trihydroxy
docosahexanoic acid, (4.1) and its aspirin triggered analogue, (4.2). To the best of our
knowledge, this is the first total synthesis of Resolvin D4 and our biological findings will
be reported in due course.
4.2 Results and Discussion
4.2.1 Synthesis of RvD4 and AT-RvD4
4.2.1.1 Retrosynthetic Analysis
Resolvin D4 contains two distinct conjugated systems as well, similar to RvD3 however
its C1-C11 arrangement is completely different. Specifically the C6-C11 E, E, Z triene
moiety takes on a completely different spatial arrangement and structure having a
different affect on binding affinity. The C12 through C22 portion of the molecule
however, remains similar to Resolvin D3 with a C13-16 Z, E diene moiety. It contains
three hydroxyl groups, two of which are positioned at the 4S, 5R carbons posing a
problematic complication in which the compound can rearrange to form a 5 or 6
membered lactone, (6 membered being slightly more strained), that is thermodynamically
favored. The third hydroxyl group retains its position at the 17 carbon in the S or R
configuration depending on the presence of aspirin (Figure 16). Our goal once again was
93

Figure 16. Retrosynthetic Analysis of RvD4 and AT-RvD4

to conserve the nature of the conjugated systems by reducing the cis-olefins in the final
step of our synthesis scheme. The difficulty herein lied not only in controlling
lactonization or polymerization that was thermodynamically favorable but also carefully
monitoring the final deprotection in the presence of the rather acidic methylene protons
situated between the two alkynes making up the triene and diene conjugated systems
prior to the final reduction. With these aforementioned challenges in mind we began our
synthesis with the ring opening of chirally pure d-erythrose (4.6) to set the 4S, 5R
hydroxyl groups in place (4.5). Next we took advantage of a Wittig coupling to form the
carbon-carbon bond at the 6-position between the wittig salt (2.7) and aldehyde (4.5) and
94
through additional manipulation built the conjugated triene system. The 17-hydroxy
position was once again set in place through a ring opening of chiral glycidol. The final
carbon carbon bond was formed through a copper mediated cross coupling of an allylic-
bromide (4.3) with our terminal alkyne (4.4) to afford the protected bis-acetylenic
precursor. Due to the unique nature of each metabolite RvD4 at this stage required
careful manipulation. The bis-acetylenic precursor was reduced prior to the deprotection
however Zn/Cu/Ag hydrogenation was too mild in the presence of bulky protecting
groups. Therefore, Lindlar catalyst was employed monitoring the reaction very closely
and subsequent desilyation allowed for the synthesis of the final compound (4.1) without
decomposition.
4.2.1.2 Synthesis of Methyl Ester (4.5)
Efficient access to C(1) C(6) fragment (4.5) was achieved by two-carbon homologation
of the chirally pure precursor, D-erythrose (4.6), with the Methyl
(triphenylphosphoranylidene) acetate stabilized ylide (4.7) to afford the α,β-unsaturated
ester (4.8) (Scheme 9).
88 89
It is well known that the 4-OH at an aldehyde affords
primarily the Z olefin in the wittig reaction. Protection of the triol moieties with TBS-Cl
afforded the protected triol (4.9) in good yields. The α,β-unsaturated ester was reduced in
the presence of Lindlar catalyst to afford compound (4.10) in excellent yield. Selective
de-silylation afforded the primary alcohol (4.11) which was then oxidized using Swern
conditions
90
to afford the silyl ether aldehyde (4.5).
95

Scheme 9. Synthesis of Methyl Ester Intermediate (4.5).

4.2.1.3 Synthesis of Terminal Alkyne (4.4)
The triene backbone was assembled by coupling the same wittig piece utilized in the
synthesis RvD2, (2.7), with aldehyde (4.5) in the presence of base, Scheme 10. This
afforded a mixture of the E, Z and E, E isomers (4.12) (9:1 ratio favoring E:Z). To assure
pure E, E stereochemistry the mixture was efficiently isomerized in the presence of I2 to
afford the all trans terminal alkyne (4.13). This was desilyated to expose the terminal
alkyne (4.4) allowing for the next stage of final coupling.
96

Scheme 10. Total Synthesis of Terminal Alkyne (4.4).

4.2.1.4 Synthesis of Propargyl Bromide (4.3)
The synthesis of propargyl bromide (4.3) was synthesized rather straight forward from
the omega vinyl iodide building block precursor (3.3) used in the synthesis of resolvin D3.
Palladium mediated cross coupling of propargyl alcohol with vinyl halide (3.3) using
Sonogashira conditions afforded the propargyl alcohol (4.14). The alcohol was then
converted to the bromide (4.3) in good yield once again utilizing N-bromo Succinimide
and tri-phenyl phosphine. Both the coupling and Bromination were achieved in good
yields. The alternative approach coupling the propargyl alcohol to the terminal alkynyl
piece (4.4) gave unfavorable results.

Scheme 11. Construction of Propargyl Bromide Intermediate (4.3).

97
4.2.1.5 Final Construction of RvD4 (4.1)
Copper mediated coupling of the terminal alkyne (4.4) and propargyl bromide building
blocks (4.3) afforded the protected bisacetylenic RvD4 precursor (4.14) in excellent yield,
Scheme 12.
91
However the subsequent reduction and deprotection steps required a great
deal of caution due to the unique nature of RvD4. After attempting the TBAF assisted
deprotection of the three hydroxyl groups under various conditions it was determined that
the methylene protons sandwiched between the two conjugated triple bonds were causing
the compound to decompose. Further investigation found that side reactions could occur
due to the acidic nature of C-12 methylene protons between the E, E, Z triene and the Z,
E diene moieties. The labile nature of this methylene unit made the compound
susceptible to degradation and double bond rearrangement in the presence of the highly
basic fluoride anion during TBAF deprotection.
92
To bypass this we hypothesized that we
could reduce the triple bonds first thus decreasing the labile nature of the acidic
methylene protons allowing us to carry out the de-silylation as the final step.
Unfortunately the reduction using Zn/Cu/Ag amalgam is too mild and therefore we were
required to reduce the protected bisacetylenic precursor in the presence of a more active
reducing agent such as Lindlar catalyst. This was no simple task since Lindlar catalyst
can easily over hydrogenate these conjugated alkyne bonds. Monitoring very closely and
controlling the amount of H2 introduced into the flask as well as running the reaction in a
limited amount of solvent we found that we could successfully reduce both bonds and
subsequently deprotect using TBAF to afford a mixture of the product as well as some
starting material and a small amount of over hydrogenated side product.
93
This requires a
great deal of skill and was only successful after carrying out multiple reactions that failed.
98

Scheme 12. Final Coupling of Resolvin D4 (4.1).

Fortunately we were able to run the reaction on rather small scales, roughly 1 mg, and
monitor with the help of our new Agilent 1260 LC/MS system. In summary not only did
RvD4 require great caution during the deprotection due to the nature of the C-12 protons
but it also encountered similar difficulties with lactonization of the C4 and C5 hydroxyl
groups with the free acid. And once again we now know that terminal Hydroxy acids can
lactonize or polymerize due to concentration, temperature as well as the presence of any
nucleophiles in the reaction media.
94
As a result the deprotection step that utilizes the
fluoride anion as a strong nucleophile can cause side reactions with these sensitive lipid
metabolites. Similarly to the work carried out on resolvin D3 further experimentation
will aid in a better and more efficient synthesis of this potent mediator.

99
4.3 Conclusion

In conclusion we report the successful total synthesis of RvD4 (4.1) and its aspirin
triggered stereoisomer (4.2). This was accomplished through trial and error using a
number of well-employed organic reactions and a convergent synthetic strategy that has
allowed our group to build a number of structurally related but very different biological
mediators. Currently work is underway to test these compounds to better understand
their role against inflammation and will be reported in the near future. Ideally this work
will help further advance applications in treatment and of a number of inflammatory
related diseases.

100
4.4 Resolvin D4 Experimental
Unless otherwise noted, all reactions were carried out in a flame-dried flask with stir bar
under argon routed through a three-necked valve. Reactions were carried out at room
temp using DriSolv solvents purchased commercially from VWR. All reagents used
were purchased without further purification from Sigma Aldrich, Strem, and Alfa Aesar.

Progress was monitored and recorded using EMD analytical thin layer chromatography
plates, Silica Gel 60 F254. TLC plates were visualized through UV absorbance, (254 nm),
or staining such as vanillin, phosphomolybdic acid, potassium-permanganate, or
ninhydrin followed by heating. Unless otherwise stated, purification was carried out by
flash column chromatography manually using Silica Gel (100-200 mesh) or automatically
using the Biotage Isolera One.

Characterization was carried out using LC-MS, NMR and UV-VIS instrumentation. All -
1
H and
13
C spectra were procured on the Departments Varian 400, 500 and 600 MHz
NMR instruments in the solvent indicated.
1
H and
13
C chemical shifts, (δ), are recorded
in parts per million, (ppm), and referenced to the residual solvent converted by the TMS
scale (CDCl3,
1
H = 7.26 ppm). Splitting patterns are denoted by s, d, t, dd, td, ddd, and
m and refer to the respective multiplicities; singlet, doublet, triplet, doublet of doublets,
triplet of doublets, doublet of doublet of doublet and multiplet. Mass spectra was
recorded on an Agilent 1260 LC-MS. UV-Vis spectra was obtained by a Hewlett-
Packard 8350 instrument.

101

(4S,5R,E)-methyl 4,5,6-trihydroxyhex-2-enoate (4.8).

To a solution of D-erythrose 4.6
(0.85 g, 7.1 mmol) in dry THF (10 mL) was added methyl (triphenylphosphoranylidene)
acetate 4.7 (2.4 g, 7.1 mmol). The reaction mixture was stirred at 65°C overnight.
Without workup the solvent was removed in vacuo and the crude mixture was purified on
silica gel using MeOH/CH2Cl2 (10%) as the eluent to afford the triol ester 4.8 (1.1 g,
90%) as a clear colorless oil.
1
H NMR (400 MHz, CD3OD) δ 7.16 (dd, J = 15.7, 4.8 Hz,
1H), 6.13 (dd, J = 15.7, 1.3 Hz, 1H), 4.27 (t, J = 6.4 Hz, 1H), 3.76 (s, 3H), 3.72 – 3.67 (m,
1H), 3.65 – 3.60 (m, 1H), 3.62 – 3.54 (m, 1H).
13
C NMR (400 MHz, CD3OD) δ 168.61,
150.16, 121.67, 75.79, 72.67, 64.35, 52.01.

(4S,5R,E)-methyl 4,5,6-tris(tert-butyldimethylsilyloxy)hex-2-enoate (4.9). To a flask
with imidazole (310 mg, 3.75 mmol) and DMAP (7 mg, 0.12 mmol) in DMF (5 mL total
volume) was added TBS-Cl (680 mg, 3.75 mmol) dropwise at 0°C. The triol 4.8 (220 mg,
1.25 mmol) was cannulated to the flask and stirred overnight at room temperature. The
reaction mixture was quenched with saturated aqueous NH4Cl (7 mL) and extracted with
Et2O (3 x 7 mL). The organic layer was dried with MgSO4, filtered and the solvent
removed in vacuo. The crude reaction mixture was purified on silica gel using EtOAc-
102
hexanes (2%) as the eluent to afford the protected triol ester 4.9 (580 mg, 90%) as a clear
colorless oil.
1
H NMR (500 MHz, CDCl3) δ 7.00 (dd, J = 15.7, 5.4 Hz, 1H), 5.97 (dd, J =
15.7, 1.5 Hz, 1H), 4.41 (ddd, J = 5.5, 3.8, 1.7 Hz, 1H), 3.74 (s, 1H), 3.69 (td, J = 5.8, 3.6
Hz, 1H), 3.61 – 3.49 (m, 2H), 0.91 (s, 8H), 0.89 (s, 8H), 0.86 (s, 9H), 0.06 (s, 3H), 0.05 (s,
8H), 0.05 (s, 5H), 0.04 (s, 3H), 0.03 (s, 3H).
13
C NMR (500 MHz, CDCl3) δ 167.03,
148.86, 121.12, 77.53, 73.43, 64.52, 51.60, 26.10, 26.07, 26.02, 18.45, 18.40, 18.29, -
4.30, -4.41, -4.46, -4.64, -5.22, -5.27.

(4S,5R)-methyl 4,5,6-tris(tert-butyldimethylsilyloxy)hexanoate (4.10). To a solution
of protected triol 4.9 (450 mg, 0.87 mmol) in EtOAc (6 mL) was added one scoop of 5%
palladium on charcoal. The reaction was stirred under H 2 overnight. The reaction
mixture was filtered through celite and with no workup the solvent evaporated. The
crude mixture was purified on silica gel using EtOAc-hexanes (3%) as the eluent to
afford the protected triol ester 4.10 (440 mg, 97%) as a clear colorless oil.
1
H NMR (400
MHz, CDCl3) δ 3.78 (dt, J = 7.2, 3.4 Hz, 1H), 3.66 (s, 3H), 3.67 – 3.60 (m, 1H), 3.58 (dd,
J = 10.2, 6.1 Hz, 1H), 3.45 (dd, J = 10.1, 5.8 Hz, 1H), 2.46 – 2.34 (m, 2H), 1.90 – 1.79
(m, 2H), 0.89 (s, 18H), 0.88 (s, 9H), 0.10 (s, 3H), 0.08 (s, 3H), 0.07 (s, 6H), 0.04 (s, 6H).
13
C NMR (400 MHz, CDCl3) δ 174.54, 77.26, 72.90, 65.00, 51.58, 30.27, 27.35, 26.14,
26.13, 26.09, 18.47, 18.37, 18.29, -3.95, -4.21, -4.52, -4.72, -5.22, -5.28.
103

(4S,5R)-methyl 4,5-bis(tert-butyldimethylsilyloxy)-6-hydroxyhexanoate (4.11). To a
solution of protected triol 4.10 (440 mg, 0.84 mmol) in a 1:1 mixture of CH2Cl2/MeOH
(5 mL) was added camphorsulfonic acid (157 mg, 0.68 mmol) at 0°C. The reaction was
quenched after 50 min with Et3N (0.12 mL, 0.84 mmol) and the solvent was removed in
vacuo. The crude mixture was purified on silica gel using EtOAc-hexanes (12%) as the
eluent to afford the protected triol ester 4.11 (200 mg, 65%) as a clear colorless oil.
1
H
NMR (400 MHz, CDCl3) δ 3.80 (q, J = 5.0 Hz, 1H), 3.66 (s, 3H), 3.72 – 3.62 (m, 1H),
3.60 (dd, J = 8.3, 4.4 Hz, 2H), 2.39 (td, J = 7.8, 5.6 Hz, 2H), 1.87 (td, J = 7.9, 5.1 Hz, 2H),
0.90 (s, 8H), 0.89 (s, 8H), 0.10 (s, 5H), 0.10 (s, 3H), 0.07 (s, 2H).
13
C NMR (400 MHz,
CDCl3) δ 174.27, 75.15, 72.86, 63.87, 51.70, 29.34, 28.53, 26.05, 26.01, 18.26, 18.24, -
4.29, -4.36, -4.43, -4.51.

(4S,5S)-methyl 4,5-bis((tert-butyldimethylsilyl)oxy)-6-oxohexanoate (4.5). To a -
78°C solution of DMSO (0.11 mL, 1.5 mmol) in anhydrous CH 2Cl2 (10 mL) was added
oxalyl chloride (0.1 mL, 1.0 mmol) drop wise. After 0.25 h alcohol 4.11 (200 mg, 0.5
mmol) was added and stirred for 1 h at -78°C. To the reaction mixture was added Et3N
(0.35 mL, 2.5 mmol) and stirred for another 3 h at -78°C. The reaction mixture was
104
allowed to warm to room temperature and quenched with saturated aqueous NH4Cl (10
mL) and extracted with Et2O (3 x 10 mL). The combined extract was dried with Na 2SO4
and evaporated to give a crude clear oil which was then chromatographed on silica gel
using EtOAc-hexanes mixture of (10%) as the eluent to afford the aldehyde 4.5 as a
viscous and colorless oil (190 mg, 94%).
1
H NMR (400 MHz, CDCl3) δ 9.59 (d, J = 1.9
Hz, 1H), 3.99 – 3.93 (m, 1H), 3.88 (dd, J = 3.5, 1.9 Hz, 1H), 3.66 (s, 3H), 2.37 (dd, J =
8.4, 7.1 Hz, 2H), 2.01 – 1.73 (m, 2H), 0.91 (s, 9H), 0.87 (s, 9H), 0.08 (s, 3H), 0.08 (s, 6H),
0.06 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 203.43, 173.75, 81.02, 74.19, 51.74, 29.69,
28.57, 25.97, 25.93, 18.40, 18.22, -4.24, -4.52, -4.70, -4.72.

(4S,5R)-methyl 4,5-bis(tert-butyldimethylsilyloxy)-11-(trimethylsilyl)undeca-6,8-
dien-10-ynoate (4.12). To a solution of wittig salt 2.7 (0.38 g, 0.79 mmol) previously
used in the synthesis of Resolvin D2 in THF (10 mL) was added 2.5 M n-BuLi (1.5 mL,
0.6 mmol) at -78°C. The reaction was stirred for 0.5 h at 0°C before Aldehyde 4.5 (0.16
g, 0.40 mmol) was cannulated to the mixture once again at -78°C. The mixture was
stirred at room temperature for 3 h where it was then quenched with saturated aqueous
NH4Cl (10 mL) and extracted with Et2O (3 x 15 mL). The organic layer was dried with
MgSO4, filtered and the solvent removed in vacuo. The crude mixture was purified on
silica gel using EtOAc-hexanes mixture of (1:49) as the eluent to afford a 9:1 ratio of Z, E
105
and E, E olefin 4.12 (172 mg, 83%).
1
H NMR (400 MHz, CDCl3) δ 6.81 (ddd, J = 15.5,
11.5, 1.1 Hz, 1H), 6.05 (t, J = 11.3 Hz, 1H), 5.61 (dt, J = 15.4, 0.8 Hz, 1H), 5.44 (ddt, J =
11.0, 8.9, 1.0 Hz, 1H), 4.38 (ddd, J = 8.8, 5.0, 1.1 Hz, 1H), 3.67 (s, 3H), 3.68 – 3.60 (m,
1H), 2.42 (td, J = 7.9, 1.6 Hz, 2H), 1.90 – 1.78 (m, 2H), 0.87 (s, 9H), 0.87 (s, 9H), 0.20 (s,
9H), 0.05 (s, 3H), 0.04 (s, 6H), 0.01 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 174.41,
137.95, 135.58, 129.11, 129.13, 112.88, 104.42, 97.97, 75.28, 72.18, 51.70, 29.45, 28.37,
26.11, 26.03, 18.28, 0.07, -3.79, -3.80, -3.87, -3.89, -4.61.

(4S,5R,6E,8E)-methyl4,5-bis((tert-butyldimethylsilyl)oxy)-11-(trimethylsilyl)undeca-
6,8-dien-10-ynoate (4.13). To a solution of compound mixture 4.12 (140 mg, 0.27
mmol) in CH2Cl2 (50 mL) was added I2 (7 mg) and stirred for 24 h in the presence of
natural light. The reaction was quenched with saturated Na2S2O5 (40 mL) and exctracted
with Et2O (50 mL). The solvent was removed in vacuo and the crude oil was purified on
silica gel using EtOAc-hexanes (1:49) as the eluent to afford only the E, E isomer 4.13
(130 mg, 92%) as a colorless oil.
1
H NMR (400 MHz, CDCl3) δ 6.81 (ddd, J = 15.5, 11.5,
1.1 Hz, 1H), 6.11 – 5.97 (m, 1H), 5.61 (dt, J = 15.5, 0.8 Hz, 1H), 5.44 (ddt, J = 11.0, 8.9,
1.0 Hz, 1H), 4.38 (ddd, J = 8.8, 5.0, 1.1 Hz, 1H), 3.67 (s, 3H), 3.69 – 3.60 (m, 1H), 2.42
(td, J = 7.9, 1.6 Hz, 2H), 1.90 – 1.76 (m, 2H), 0.87 (s, 9H), 0.87 (s, 9H), 0.20 (s, 9H),
0.05 (s, 3H), 0.04 (s, 6H), 0.01 (s, 3H).
106

(4S,5R,6E,8E)-methyl 4,5-bis((tert-butyldimethylsilyl)oxy)undeca-6,8-dien-10-ynoate
(4.4). To a solution of TMS protected alkyne 4.13 (48 mg, 0.09 mmol) in MeOH (2 mL)
was added Na2CO3 (9 mg) and stirred overnight at room temperature. The solvent was
removed in vacuo and the crude was dissolved in water (3 mL) and extracted with Et2O
(3 x 5 mL). The oil was purified on silica gel using EtOAc-hexanes (1:49) as the eluent
to afford compound 4.4 (38 mg, 94%) as a colorless oil.
1
H NMR (400 MHz, CDCl3) δ
6.55 (dd, J = 15.7, 10.9 Hz, 1H), 6.08 (ddt, J = 15.3, 10.9, 0.9 Hz, 1H), 5.67 (dd, J = 15.3,
7.0 Hz, 1H), 5.45 (dd, J = 15.7, 2.4 Hz, 1H), 3.89 (ddd, J = 7.0, 4.7, 1.1 Hz, 1H), 3.56 (s,
3H), 3.53 (q, J = 5.1 Hz, 1H), 2.92 (d, J = 2.4 Hz, 1H), 2.28 (dd, J = 9.3, 7.0 Hz, 2H),
1.79 – 1.61 (m, 2H), 0.78 (s, 9H), 0.77 (s, 9H), -0.05 (s, 3H), -0.07 (s, 3H), -0.08 (s, 3H),
-0.10 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 174.37, 142.98, 137.98, 130.48, 109.98,
82.98, 79.65, 76.65, 75.20, 51.65, 29.62, 28.38, 26.08, 26.05, 18.31, 18.33, -3.89, -3.98, -
4.54, -4.60.

(6S,4E,8Z)-6-(t-butyldimethylsilyloxy)-undeca-4,8-dien-2-yn-1-ol (4.14). To a
solution of vinyl iodide 3.3 (80 mg, 0.218 mmol) in Piperidine (0.5 mL) was added
(S) (S)
OTBS
OH
107
Pd(OAc)2/PPh3 (10 mg, 0.013 mmol) and propargyl alcohol (0.13 mL, 2.18 mmol). The
reaction was quenched after 1 h with NH4Cl (5 mL) and extracted with Et2O (3 x 5 mL).
The solvent was removed in vacuo and the crude mixture was purified on silica gel using
EtOAc-hexanes (7%) as the eluent to afford compound 4.14 as a clear colorless oil (60
mg, 92%).
1
H NMR (400 MHz, CDCl3) δ 6.15 (dd, J = 15.8, 5.1 Hz, 1H), 5.69 (dq, J =
15.8, 2.0 Hz, 1H), 5.53 – 5.38 (m, 1H), 5.38 – 5.23 (m, 1H), 4.38 (d, J = 2.0 Hz, 2H),
4.25 – 4.11 (m, 1H), 2.32 – 2.17 (m, 2H), 2.02 (p, J = 7.6 Hz, 2H), 1.25 (s, 0H), 0.95 (t, J
= 7.6 Hz, 3H), 0.89 (s, 4H), 0.05 (s, 3H), 0.04 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ
146.86, 134.18, 124.03, 108.16, 87.56, 84.26, 72.52, 51.81, 35.95, 25.98, 20.89, 18.37,
14.30, -4.48, -4.66.

(6S, 3Z, 7E)-6-(t-butyldimethylsilyloxy)-1-bromoundeca-3,7-dien-9-ynyl (4.3). To a
solution of propargyl alcohol 4.14 (60 mg, 0.2 mmol) in dry CH2Cl2 (5 mL) was added
PPh3 (48 mg, 0.18 mmol) and N-bromosuccinimide (33 mg, 0.184 mmol) at 0°C. After
30 mins the reaction was quenched with NaHCO3 (5 mL) and extracted with Et2O (3 x 5
mL). The solvent was removed in vacuo and the crude mixture was purified on silica gel
using EtOAc-hexanes (2%) as the eluent to afford compound 4.3 as a clear colorless oil
(59 mg, 83%).
1
H NMR (400 MHz, CDCl3) δ 6.20 (dd, J = 15.8, 4.9 Hz, 1H), 5.71 (dq, J
= 15.8, 2.2 Hz, 1H), 5.54 – 5.41 (m, 1H), 5.39 – 5.23 (m, 1H), 4.24 – 4.10 (m, 1H), 4.06
(d, J = 2.3 Hz, 2H), 2.35 – 2.15 (m, 2H), 2.09 – 1.93 (m, 2H), 0.96 (t, J = 7.5 Hz, 3H),
(S) (S)
OTBS
Br
108
0.89 (s, 9H), 0.05 (s, 3H), 0.03 (s, 3H).
13
C NMR (400 MHz, CDCl3) δ 147.90, 134.27,
123.92, 107.91, 85.39, 84.43, 72.41, 35.90, 25.97, 20.89, 18.35, 15.69, 14.29, -4.50, -4.66.

Methyl-(4S, 5R, 17S, 6E, 8E, 15E, 19Z)-tris-(tert-butyldimethylsilyloxy)-docosa-
6,8,15,19-tetraen-10,13-diynoate (4.15). To a flame dried flask with CuI (16 mg, 0.084
mmol), NaI (13 mg, 0.084 mmol), and K2CO3 (12 mg, 0.084 mmol) in dry DMF (3 mL
total volume) was cannulated alkyne 4.4 (20 mg, 0.042 mmol) and allylic bromide 4.3
(30 mg, 0.084 mmol). The reaction was stirred for 24 h and quenched with saturated
NH4Cl (5 mL). The mixture was extracted with Et2O (3 x 5 mL), rinsed with water to
remove any DMF, and the organic layer was dried with MgSO4, filtered and the solvent
removed in vacuo. The crude reaction mixture was purified on silica gel using EtOAc-
hexanes (1%) as the eluent to afford compound 4.15 as a clear colorless oil (29 mg, 95%).
1
H NMR (600 MHz, CDCl3) δ 6.55 (dd, J = 15.6, 10.8 Hz, 1H), 6.15 (dd, J = 15.3, 10.9
Hz, 1H), 6.17 – 6.07 (m, 2H), 5.71 (dd, J = 15.3, 7.2 Hz, 1H), 5.64 (dq, J = 15.8, 2.0 Hz,
1H), 5.55 (dt, J = 15.5, 2.4 Hz, 1H), 5.52 – 5.40 (m, 1H), 5.37 – 5.25 (m, 0H), 4.20 – 4.11
(m, 1H), 3.96 (dd, J = 7.1, 4.8 Hz, 1H), 3.66 (s, 3H), 3.62 (q, J = 5.1 Hz, 1H), 3.44 (t, J =
2.2 Hz, 2H), 2.42 – 2.33 (m, 2H), 2.31 – 2.17 (m, 2H), 2.02 (p, J = 7.4 Hz, 2H), 1.87 –
1.76 (m, 2H), 0.95 (t, J = 7.6 Hz, 3H), 0.89 (s, 9H), 0.88 (s, 9H), 0.86 (s, 9H), 0.05 (s,
3H), 0.04 (s, 3H), 0.03 (s, 3H), 0.03 (s, 3H), 0.02 (s, 3H), -0.01 (s, 3H).
13
C NMR (600
(R) (R) (S) (S)
COOMe
OTBS TBSO
(S) (S)
OTBS
109
MHz, CDCl3) δ 174.40, 146.17, 141.27, 136.98, 134.07, 130.88, 124.18, 111.01, 108.60,
86.07, 83.46, 80.03, 79.24 , 76.71, 75.17, 72.60, 51.63, 36.00, 29.59, 28.40, 26.08, 26.05,
25.99, 20.89, 18.37, 18.36, 18.26, 14.30, 11.46, -3.87, -3.95, -4.45, -4.54, -4.61, -4.66.

(4S, 5R, 17S)-Trihydroxydocosa-(6E,8E,10Z,13Z,15E)-hexaenoic acid, or RvD4 (4.1).
Part 1: To a solution of compound 4.15 (35 mg, 0.048 mmol) in EtOAc, (1 mL), Octene,
(0.1 mL), and Pyridine (0.1 mL). Next was added a small scoop of Lindlar catalyst and
stirred for 12 hours under a Hydrogen atmosphere before checking the reaction. When
the reaction was monitored closely via LC-MS it ran several days however when
checking only every 12 to 24 hours the reaction went to completion. After two days the
reaction was filtered over celite and the solvent removed under reduced pressure. The
crude reaction mixture was quickly run over a silica plug to afford a mixture of
hydrogenated products (27 mg, 77%). Part 2: To the hydrogenated crude mixture
dissolved in THF (1 mL) was added dropwise ~ 10 equivalents of 1M TBAF (0.4 mL, 0.4
mmol) at 0°C. The reaction was monitored closely via thin layer chromatography and
after 4 h the reaction was quenched with saturated NH4Cl (15 mL) and extracted with
Et2O (5 x 15 mL). The organic layer was rinsed with brine, dried over MgSO4 and
filtered. The solvent was then concentrated and freshly prepared CH2N2 was added to
convert any acid to the ester-lactone mixture. The solvent was completely removed in
(R) (R)(S) (S)
COOH
OH HO
(S) (S)
OH
110
vacuo and the compound was purified on silica gel using MeOH-CH2Cl2 (1%) as the
eluent to afford an ester/lactone mixture. The product was then suspended in a H2O-
MeOH mixture (1:1, 1 mL) and 10 equivalents of LiOH (1 mg, 2.5 x 10
-2
mmol) was
added. After 3 h the reaction mixture was dried and purified via C-18 reversed Phase
HPLC using H2O-MeOH mixture (37%) to afford compound 4.1 as colorless oil (0.31
mg, 2%).
1
H NMR (600 MHz, MeOD) δ 6.51 – 6.43 (dd, J = 15.0, 11.1 Hz, 2H), 6.27 (dd,
J = 15.3, 10.7 Hz, 1H), 6.15 (dd, J = 14.7, 10.7 Hz, 1H), 5.94 (t, J = 10.9 Hz, 1H), 5.90 (t,
J = 10.9 Hz, 1H), 5.74 (dd, J = 15.2, 6.8 Hz, 1H), 5.59 (dd, J = 15.0, 6.6 Hz, 2H), 5.41 –
5.34 (m, 1H), 5.33 – 5.23 (m, 3H), 4.04 (q, J = 6.5 Hz, 1H), 3.89 (t, J = 6.0 Hz, 1H), 3.45
– 3.39 (m, 1H), 3.00 (t, J = 7.7 Hz, 2H), 2.32 – 2.13 (m, 4H), 1.95 (q, J = 7.6 Hz, 2H),
1.75 (ddd, J = 14.7, 7.1, 3.2 Hz, 1H), 1.58 (dq, J = 15.1, 7.5 Hz, 1H), 0.86 (t, J = 7.5 Hz,
3H).

111
4.5 References

84. Larjava, H. (Ed.). (2012). Oral Wound Healing: Cell Biology and Clinical
Management (1st ed., pp. 57–78). West Sussex, UK.
85. Serhan, C. N.; Arita, M.; Hong, S.; Gotlinger, K. Lipids. 2004, 39, 1125–1132.
86. Arita, M.; Yoshida, M.; Hong, S.; Tjonahen, E.; Glickman, J. N.; Petasis, N. A.;
Blumberg, R. S.; Serhan, C. N. Proceedings of the National Academy of Sciences of
the United States of America 2005, 102, 7671.
87. Pillai, P. S.; Leeson, S.; Porter, T. F.; Owens, C. D.; Kim, J. M.; Conte, M. S.;
Serhan, C. N.; Gelman, S. Inflammation 2012, 35, 98.
88. Webb, T. H.; Thomasco, L. M.; Schlachter, S. T.; Gaudino, J. J.; Wilcox, C.
S. Tetrahedron Letters 1988, 29, 6823.
89. Gallos, J. K.; Koumbis, A. E.; Xiraphaki, V. P.; Dellios, C. C.; Coutouli-
Argyropoulou, E. Tetrahedron 1999, 55, 15167.
90. Tidwell, T. T. Org. React. (N.Y.) 1990, 39, 297.
91. Lapitskaya, M. A.; Vasiljeva, L. L.; Pivnitsky, K. K. Synthesis-Stuttgart 1993, 65.
92. Ogawa, N.; Kobayashi, Y. Tetrahedron Letters 2011, 52, 3001.
93. Oger, C.; Balas, L.; Durand, T.; Galano, J.-M. Chemical Reviews 2013, 113, 1313.
94. Petasis, N. A.; Akritopoulou-Zanze, I.; Fokin, V. V.; Bernasconi, G.; Keledjian, R.;
Yang, R.; Uddin, J.; Nagulapalli, K. C.; Serhan, C. N. Prostaglandins Leukotrienes
and Essential Fatty Acids 2005, 73, 301.

112
CHAPTER 5. Total Synthesis and Stereochemical Assignment of Aspirin Triggered
Neuroprotectin D1 and Analogues.

5.1 Introduction
Protectin D1 and its analogues are a series of PUFA-derived oxygenated lipid metabolites
formed enzymatically from docosahexanoic acid (DHA) through processes of
inflammation and resolution. Much of our labs work over the past several years has
focused on the study of these complex DHA-based derivatives and their unnatural
stereoisomers. In 2006 a number of NPD1/PD1 analogues were synthesized in our lab to
firmly establish the complete stereochemical assignment of this new family of bioactive
metabolites (Fig. 17).
95, 96
In the biosynthesis of protectins it is known that two forms
exist: the aspirin triggered pathway
97
and the lipoxygenase-initiated pathway.
98
In fact it
has been known for some time that aspirin triggers the biosynthesis of metabolites
through an alternative enzymatic pathway by acetylating COX-1 and COX-2.
99

Figure 17. Biosynthesis of NPD1 / PD1 and AT- NPD1 / PD1.
2
113
Acetylation of COX-2 blocks the endoperoxide pathway however the enzyme continues
to remain active through a 15-HETE pathway generating products from DHA and AA
with the 17 position of the alcohol in the R configuration. In our efforts to bring the
study of Neuroprotectin/Protectin full circle we investigated an additional family of
aspirin-triggered protectin analogues and their stereoisomers, (Figure 18), to not only
confirm the stereoassignment of AT-NPD1/PD1 but also understand its role in the
resolution of a number inflammatory related diseases including Alzheimer’s,
100

stroke,
101,102
and age related macular degeneration (AMD).
103

5.2 Results and Discussion
5.2.1 Synthesis of AT-NPD1 and Stereoisomers
5.2.1.1 Retrosynthetic Analysis
In our retrosynthetic analysis of this class of compounds we required three different
double bond configurations of the triene moiety which serve as a critical component in
bioactivity of such metabolites (Figure 18). These six synthetic materials, (5.1, 5.2 and

Figure 18. Retrosynthetic Analysis of (10R / 10S) AT-NPD1 and Isomers (5.1, 5.2, 5.3).
114
5.3 both the 10S and 10R stereoisomers), were made for the direct matching of AT-
NPD1/PD1 biosynthesized metabolites. In order to successfully build the E, E, Z, (5.1),
and E, Z, E, (5.3) metabolites, we utilized the highly efficient Sonogashira coupling using
Palladium (0) and Copper (1).
104
This reaction provided the product in 95 and 98% yield
respectively when carried out under inert conditions. The E, E, E, (5.2), arrangement was
successfully obtained by Negishi coupling utilizing a Zr/Zn complex in the presence of
Palladium (0).
105
To obtain the omega 3 cis olefin we employed a Lindlar hydrogenation
in excellent yield. The methyl ester piece required a much more careful synthesis due to
the nature of the allylic protons located between the cis cis olefins as well as the
isomerization that could occur by the E, E vinyl iodide (5.5). In part a copper mediated
coupling of an allylic bromide with our alkynyl ester afforded (5.5) in appreciable yields
and the chiral alcohol was obtained through an epoxide ring opening of the
enantiomerically pure glycidol starting material (5.7). In order to devise a more efficient
synthetic strategy we optimized some reactions by refining conditions or eliminating
steps all together.
5.2.1.2 Synthesis of propargyl bromide
The title propargyl bromide (5.11) was prepared, Scheme 13, in good yields initially by
the epoxide opening of protected chiral glycidol with a metallated 1-alkyne, or in this
case protected propargyl alcohol (5.8). Without purification the crude reaction mixture
was worked up, dried over magnesium sulfate and and dried in vacuo. The crude mixture
was placed and placed in a flame dry flask and protected (5.9) as the tert-
butyldiphenylsilyl ether (TBDPS). Treatment with camphor sulfonic acid (CSA)
afforded the diol (5.10) in appreciable yields. The diol could then be selectively
115

Scheme 13. Synthesis of Propargyl Bromide Intermediate (5.11).

brominated affording the propargyl bromide alcohol (5.11). This selective bromination
most likely caused by the steric

interference of the TBDPS protecting group, avoided an
additional protection and deprotection step or a finicky selective deprotection step in
addition to a latter deprotection step.
5.2.1.3 Synthesis of Vinyl Iodide Intermediate (5.5)
The synthesis of the vinyl iodide intermediates, (5.5 and 5.7), proved challenging on
multiple levels (Scheme 14). It began with the copper mediated coupling of propargyl
bromide (5.11) and alkyne (5.12). The propargyl bromide is substituted by an excess of
NaI allowing for the more reactive halogen to couple to the copper coordinated terminal
alkyne in the presence of base.
106
Unfortunately this made purification challenging when
the propargyl bromide alcohol is protected. With it deprotected however, purification of
the coupled bis-acetylenic product (5.13) was purified with greater ease. Lindlar
hydrogenation of the bis-acetylenic precursor required great caution and skill. The
116

Scheme 14. Synthesis of Vinyl Iodide Intermediate (5.5 and 5.7).

Lindlar hydrogenation was susceptible to over hydrogenation largely in part to the lability
of the allylic methylene hydrogens located between the two cis double bonds. This
created a mixture of difficult to separate mono-hydrogenated, bis-hydrogenated as well as
over hydrogenated products. As a result this reaction was monitored with great care to
procure the diene only and thus avoid over-hydrogenation or any other side products.
Oxidation of the diene alcohol afforded the aldehyde (5.15) which could then be taken to
either the vinyl iodide (5.7) utilizing the takai olefination or to the α,β-unsaturated
aldehyde (5.16) by wittig homologation. Subsequent takai olefination of the unsaturated
117
aldehyde afforded the E, E vinyl iodide (5.5) in good yields and with good regio-
selectivity.
5.2.1.4 Synthesis of Terminal Alkyne (5.4) and (5.6)

The synthesis of the bottom terminal alkyne piece (Scheme 15, 5.4 and 5.6) with R-
stereochemistry began with the title compound (3.24’). To obtain terminal alkyne (5.4),
the aldehyde was converted first to the geminal di-bromo olefin (5.17) using the Corey-
Fuchs method and then reduced in the presence of strong base. However, to construct the
omega alkyne for the E, Z, E olefin (5.3) aldehyde (3.24’) was converted to the α,β-
unsaturated aldehyde (5.18) by wittig homologation and then converted to the E alkyne
(5.6) using Corey-Fuchs method once again.

Scheme 15. Synthesis of Acetylenic Omega Intermediates (5.19 and 5.22).

118
5.2.1.5 Final Construction of AT-NPD1 Stereoisomers
With key building blocks in hand, (5.4, 5.6, 5.5, and 5.7), we focused our efforts on the
formation of the final carbon-carbon bond of the aspirin triggered stereoisomers. In
Scheme 16 we coupled the vinyl iodide (5.5) with the terminal alkyne (5.4) as reported
previously for NPD1 to construct the protected acetylenic precursor (5.20). Yields were
as expected both of the coupling, fluoride deprotection and final hydrogenation. For the

Scheme 16. Final Coupling for AT-PD1 and 10-epi AT-PD1 as well as (13Z, 15E) AT-PD1 and (13Z,
15E ) 10-epi AT-PD1.
119
purpose of our biological studies mostly the free acid was compared in bioassays.
Therefore samples were prepared as the sodium salt (5.1) in ethanol and converted to the
free acid through a very careful and mild workup using a buffer solution and a dilute
acetic acid solution. As for the construction of (11E, 13Z, 15E) AT-PD1 (5.3) and (11E,
13E, 15Z) 10-epi AT-PD1 (5.3) we coupled vinyl iodide, (5.7), with terminal alkyne (5.6)
following the exact same procedure for AT- (NPD1/PD1). In this case however there
were notably improved results in yield for both the coupling, the fluoride deprotection
and the final reduction using the Zn (Cu/Ag) amalgam in a water/methanol mixture.
Finally in Scheme 17 we constructed the (11E, 13E, 15E) AT-PD1 (5.2) and the (11E,
13E, 15E) 10-epi AT-PD1 (5.2) under Negishi conditions using vinyl iodide, (5.5), with
terminal alkyne (5.4). We achieved modest yields however TBAF deprotection was our
final step therefore we had no need to carry out the at times troublesome final reduction
since the Negishi coupling put the E, E, E triene system in place in a single step.

Scheme 17. Final Coupling for (11E, 13E, 15E) AT-PD1 and (11E, 13E, 15E) 10-epi AT-PD1.

120
In Figure 19 we see the olefinic overlay of the NMR spectra of AT-NPD1 stereoisomers
in the olefinic region between 7 and 5 ppm. The correct geometry was confirmed by
NMR analysis utilizing both one dimensional experiments as well as 2-D COSY
experiments. Also since all AT- NPD1/PD1 isomers were synthesized from natural
chirally pure starting materials the correct R/S configuration was known.
5.3 BIOLOGY

Protectin AT-NPD1/PD1 has been distinguished for its unique anti-inflammatory and
pro-resolving actions. In reporting the aspirin triggered protectin pathway from mice we
see that AT-NPD1/PD1 displays protective bioactions comparable and in some cases
more favorable than NPD1. We see 50% and 40% PMN reduction respectively at
10ng/mouse of NPD1 and AT-NPD1. It was also found that despite the ability of AT-

Figure 19. AT NPD1 Analogues Olefinic
1
H Spectra.
81

121
NPD1 to act as a potent regulator of PMN infiltration in vivo, it is not an
immunosuppressive and therefore does not completely block PMN recruitment.
81
In
Figure 4A we see the HPLC profile of lipid mediators in response to zymozan-induced
peritonitis in mice supplemented with DHA and aspirin. Confirmation of biogenic AT-
NPD1/PD1 was achieved through matching HPLC retention times with its synthetic
counterpart. Figure 4B shows a TNF-α induced peritonitis model correlating
concentration of metabolite to reduction of PMN infiltration. As was mentioned earlier
the double bond configuration is crucial and we see that in comparison to the AT-
NPD1/PD1 and NPD1/PD1 metabolites, the all-trans isomer is for the most part inactive.
AT-NPD1 has also been shown to attenuate cerebral ischemic injury both through a novel
biosynthetic pathway as well as administered in its synthetic form. Aspirin triggered
neuroprotectin formed through the acetylated cyclooxygenase pathway could have
potential therapeutic implications on inflammatory related diseases such as stroke and
other conditions.

Figure 20. A) Protectin HPLC Profile B) Reduction of PMN Infiltration in a TNF-α induced
peritonitis model.
81
122
5.4 CONCLUSION
In summary we accomplished the total synthesis of a number of AT-NPD1/PD1
stereoisomers related to those produced endogenously through anti-inflammatory
pathways.
107
We used a highly convergent approach our lab has developed over the years,
while taking further steps to make the synthesis more efficient. We found that we could
selectively brominate one of the primary alcohols thus eliminating an additional
protection and deprotection step while at the same time efficiently coupling the allylic
bromide with the alkynyl ester and purify with much better separation. This project will
continue to provide new insights into pathways of inflammation and the role of
endogenous lipid metabolites in inflammatory related diseases.

123
5.5 Experimental
Unless otherwise noted, all reactions were carried out in a flame-dried flask with stir bar
under argon routed through a three-necked valve. Reactions were carried out at room
temp using DriSolv solvents purchased commercially from VWR. All reagents used
were purchased without further purification from Sigma Aldrich, Strem, and Alfa Aesar.

Progress was monitored and recorded using EMD analytical thin layer chromatography
plates, Silica Gel 60 F254. TLC plates were visualized through UV absorbance, (254 nm),
or staining such as vanillin, phosphomolybdic acid, potassium-permanganate, or
ninhydrin followed by heating. Unless otherwise stated, purification was carried out by
flash column chromatography manually using Silica Gel (100-200 mesh) or automatically
using the Biotage Isolera One.

Characterization was carried out using LC-MS, NMR and UV-VIS instrumentation. All -
1
H and
13
C spectra were procured on the Departments Varian 400, 500 and 600 MHz
NMR instruments in the solvent indicated.
1
H and
13
C chemical shifts, (δ), are recorded
in parts per million, (ppm), and referenced to the residual solvent converted by the TMS
scale (CDCl3,
1
H = 7.26 ppm). Splitting patterns are denoted by s, d, t, dd, td, ddd, and
m and refer to the respective multiplicities; singlet, doublet, triplet, doublet of doublets,
triplet of doublets, doublet of doublet of doublet and multiplet. Mass spectra was
recorded on an Agilent 1260 LC-MS. UV-Vis spectra was obtained by a Hewlett-
Packard 8350 instrument.
124

2S-1,6-Bis-(tert-butyl-dimethylsilanyloxy)-2-(tert-Butyl-diphenylsilanyloxy)-hex-4-
yn-ol (5.9). To a solution of protected propargyl alcohol (2.0 g, 11.8 mmol) in dry THF
(25 mL) was added 1.6 M n-BuLi (7.4 mL, 11.7 mmol) at -78°C. After 0.25 h BF3•Et2O
(1.5 mL, 11.7 mmol) was added drop wise at -78°C. To the reaction mixture was added
protected glycidol (1.7 g, 7.8 mmol) and stirred for 3 h at -78°C. The reaction mixture
was warmed to room temperature, quenched with saturated aqueous NH4Cl (25 mL) and
extracted with Et2O (3 x 25 mL). The organic layer was dried with MgSO 4, filtered and
the solvent removed in vacuo. The crude reaction mixture was canulated at 0°C into a
reaction flask with TBDPS-Cl (2.6 g, 9.41 mmol), imidazole (640 mg, 9.41 mmol) and
DMAP (57 mg, 0.47 mmol) in 25 mL CH 2Cl2 and stirred overnight. The reaction
mixture was quenched with saturated aqueous NH4Cl (25 mL) and extracted with Et2O (3
x 25 mL). The combined extract was dried with MgSO4, filtered and evaporated to give a
crude clear oil which was then chromatographed on silica gel using EtOAc-hexanes
mixture of (1:9) as the eluent to afford the product 5.9 (3.1 g, 66%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 7.72 (dd, J = 7.8, 1.8 Hz, 4H), 7.43 – 7.35 (m, 6H), 4.30 (t,
J = 2.2 Hz, 1H), 3.78 (q, J = 5.4 Hz, 1H), 3.71 (dd, J = 9.9, 4.2 Hz, 1H), 3.60 (dd, J =
10.0, 5.9 Hz, 1H), 1.07 (s, 9H), 0.91 (s, 9H), 0.91 (s, 9H), 0.12 (s, 3H), 0.09 (s, 3H).
13
C
NMR (400 MHz, CDCl3) δ 135.32, 134.94, 129.79, 127.89, 127.86, 81.15, 80.99, 70.36,
65.76, 52.04, 26.71, 26.02, 25.99, 23.59, 19.16, 18.47, 18.44, -4.98, -5.23, -5.25.
(S) (S)
OTBS
TBSO
OTBDPS
125

5S-(tert-Butyl-diphenyl-silanyloxy)-hex-2-yne-1,6-diol (5.10). To a solution of
compound 5.9 (3.1 g, 5.2 mmol) in a 1:1 mixture of dry CH2Cl2/MeOH (50 mL) was
added camphor sulfonic acid (1.2 g, 5.0 mmol). The reaction was allowed to stir for 1 h
and quenched with Et3N (0.72 mL, 8.5 mmol). The solvent was removed in vacuo
without work-up and purified on silica gel using EtOAc-hexanes mixture (7%) as the
eluent to afford the compound 5.10 (2.0 g, 81%) as a viscous and colorless oil. The
mono-protected alcohol can also be recovered cleanly if the reaction was monitored
closely using 0.6 eq of CSA while maintaining a 0°C temperature.

5S-1-bromo-5-(tert-Butyl-diphenyl-silanyloxy)-hex-2-yne-6-ol (5.11). To a solution of
mono-protected diol 5.10 (2.4 g, 6.5 mmol) in dry CH2Cl2 (35 mL) was added tri-phenyl
phosphine (1.88 g, 7.16 mmol) at 0°C. After 0.25 h NBS (1.27 g, 7.16 mmol) was added
at 0°C. The reaction was allowed to stir for 0.5 h and quenched with Et 3N (0.72 mL, 8.5
mmol). The solvent was removed in vacuo without work-up and purified on silica gel
using EtOAc-hexanes mixture (7%) as the eluent to afford the bromo alcohol 5.11 (1.69 g,
60%) as a viscous and colorless oil. Note: The protected diol could be selectively
brominated in good yield to afford the bromo – alcohol which could then be coupled in
good yield avoiding a deprotection step as well as making it easier to separate product
during the copper mediated coupling reaction.
1
H NMR (500 MHz, CDCl3) δ 7.77 –
(S) (S)
OH
HO
OTBDPS
(S) (S)
OH
Br
OTBDPS
126
7.64 (m, 4H), 7.48 – 7.36 (m, 6H), 3.99 – 3.88 (m, 1H), 3.80 (td, J = 2.4, 0.8 Hz, 2H),
3.62 (dd, J = 6.6, 4.2 Hz, 2H), 2.49 (ddt, J = 16.8, 7.5, 2.5 Hz, 1H), 2.38 (ddt, J = 16.8,
4.9, 2.3 Hz, 1H), 1.08 (s, 9H).
13
C NMR (500 MHz, CDCl3) δ 135.98, 135.83, 133.48,
133.46, 130.15, 130.10, 128.02, 127.89, 84.21, 72.19, 65.61, 27.11, 24.11, 19.45, 15.30.

Methyl (10S)-10-(tert-butyldiphenylsilyloxy)-11-hydroxy-undeca-4,7-diynoate (5.13).
To a solution of the bromo alcohol 5.11 (3.5 g, 8.14 mmol) was added the alkynyl ester
(1.0 g, 8.95 mmol) in DMF (60 mL). To the mixture was added NaI (2.44 g, 16.28
mmol) CuI (3.1 g, 16.28 mmol) and K2CO3 (2.25 g, 16.28) and stirred over night. The
reaction was worked up with aqueous saturated NH4Cl (50 mL) and extracted with Et2O
(3 x 60mL) and the Et2O layer was washed several times with water. The solvent was
evaporated and the mixture was purified on silica gel using EtOAc-hexanes (1:5) as the
eluent to afford compound 5.13 (2.8 g, 75%) as a clear oil.
1
H NMR (500 MHz, CDCl3)
δ 7.58 (td, J = 7.9, 1.5 Hz, 4H), 7.36 – 7.23 (m, 6H), 3.86 – 3.76 (m, 1H), 3.58 (s, 3H),
3.53 (t, J = 4.8 Hz, 2H), 2.90 (p, J = 2.3 Hz, 2H), 2.44 – 2.32 (m, 4H), 2.32 – 2.15 (m,
2H), 0.98 (s, 9H).
13
C NMR (500 MHz, CDCl3) δ 172.57, 171.28, 135.99, 135.83,
133.62, 133.59, 130.08, 130.03, 127.97, 127.84, 78.60, 76.71, 76.57, 75.11, 72.46, 65.69,
51.90, 33.51, 27.12, 23.90, 19.44, 14.77, 9.85.
COOMe
(S) (S)
HO OTBDPS
127

Methyl (10S, 4Z, 7Z)-10-(tert-Butyl-diphenyl-silanyloxy)-11-hydroxy-undeca-4,7-
dienoate (5.14). To a solution of alcohol 5.13 (0.8 g, 1.73 mmol) in EtOAc (45 mL) was
added Lindlar catalyst (500 mg). Next 12 drops of quinolone were added to the mixture.
The reaction mixture was placed under a H2 atmosphere and stirred for 2 h. The reaction
was filtered through celite and the solvent was removed in vacuo. The crude product was
purified on silica gel using EtOAc-hexanes (1:5) as an eluent to afford the product 5.14
(0.6 g, 75%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ 7.74 – 7.63 (m, 1H),
7.47 – 7.34 (m, 2H), 5.42 – 5.12 (m, 1H), 3.79 (tt, J = 8.6, 4.4 Hz, 1H), 3.67 (s, 1H), 3.57
– 3.39 (m, 1H), 2.38 – 2.12 (m, 2H), 1.93 – 1.73 (m, 1H), 1.08 (s, 2H).
13
C NMR (400
MHz, CDCl3) δ 173.71, 136.03, 135.84, 133.96, 133.89, 130.31, 129.96, 129.94, 129.30,
127.92, 127.82, 127.80, 125.16, 73.81, 65.70, 51.71, 34.18, 34.14, 31.75, 29.22, 28.84,
27.22, 27.19, 27.10, 25.69, 24.98, 22.91, 19.48.

Methyl (10S, 4Z, 7Z)-10-(tert-Butyl-diphenyl-silanyloxy)-5-oxoundeca-4,7-dienoate
(5.15). To a solution of DMSO (0.17 mL, 2.43 mmol) in CH2Cl2 (10 mL) was added
oxalyl chloride (0.17 mL, 1.94 mmol) at -78°C. To the reaction mixture was cannulated
COOMe
(S) (S)
HO OTBDPS
COOMe
(S) (S)
O OTBDPS
128
alcohol 5.14 (0.44 g, 0.97 mmol) and stirred for 1 h. Et3N (0.68 mL, 4.85 mmol) was
added to the reaction at -78°C and stirred for 3 h. The reaction mixture was quenched
with saturated aqueous NH4Cl (10 mL) and extracted with Et2O (3 x 15 mL). The Et2O
layer was washed with brine, dried with MgSO4 and filtered. The solvent was removed
in vacuo and the crude product was purified on silica gel using EtOAc-Hexanes (3:22) as
the eluent to afford the aldehyde 5.15 (0.44 g, 90%) as a colorless oil.
1
H NMR (400
MHz, CDCl3) δ 9.57 (d, J = 1.6 Hz, 1H), 7.71 – 7.59 (m, 3H), 7.47 – 7.30 (m, 5H), 5.46 –
5.37 (m, 2H), 5.38 – 5.27 (m, 1H), 4.08 (td, J = 6.0, 1.6 Hz, 1H), 3.67 (s, 4H), 2.70 (td, J
= 5.3, 2.1 Hz, 2H), 2.39 – 2.31 (m, 6H), 1.12 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ
203.47, 173.63, 135.95, 135.94, 133.17, 133.06, 130.21, 130.17, 129.02, 127.97, 127.92,
123.55, 77.79, 77.48, 77.16, 76.84, 51.69, 34.11, 31.21, 27.08, 26.86, 25.77, 22.93, 19.49.

Methyl (10S, 4Z, 7Z, 11E)-10-(tert-Butyl-diphenyl-silanyloxy)-12-iodododeca-4,7,11-
trienoate (5.7). To a solution of CrCl2 (0.54 g, 4.38 mmol) dissolved in THF (10 mL
total volume) was cannulated a mixture of compound 5.15 (0.34 g, 0.73 mmol) and CHI3
(0.57 g, 1.46 mmol) dissolved in anhydrous THF (5 mL) under Argon at 0°C. The
reaction was stirred at 0°C for 3 h and an additional 1 h at room temperature. The
reaction mixture was quenched with water (15 mL) extracted with Et 2O (3 x 15 mL)
rinsed with brine and dried over MgSO4. The organic phase was filtered and the solvent
was removed in vacuo to afford a crude oil which was purified on silica gel using first
COOMe
(S) (S)
OTBDPS I
129
pure pentanes and then EtOAc-hexanes (1:24) as the eluent to afford compound 5.7 (171
mg, 67%) as a clear colorless oil.

Methyl (10S, 4Z, 7Z, 11E)-10-(tert-Butyl-diphenyl-silanyloxy)-13-oxotrideca-4,7,11-
trienoate (5.16). To a solution of compound 5.15 (250 mg, 0.54 mmol) dissolved in
Toluene (5 mL) was added (Triphenylphosphoranylidene)acetaldehyde (365 mg, 0.65
mmol) and heated at reflux overnight. The solvent was removed in vacuo without work
up and the crude mixture was purified on silica gel using a EtOAc-hexanes mixture of
(1:9) as the eluent to afford the product 5.16 (205 mg, 80%) as a clear viscous oil.
1
H
NMR (500 MHz, CDCl3) δ 9.46 (d, J = 7.9 Hz, 1H), 7.64 (dd, J = 36.2, 6.3 Hz, 4H), 7.47
– 7.30 (m, 6H), 6.70 (ddd, J = 15.5, 4.9, 1.1 Hz, 1H), 6.21 (ddd, J = 15.5, 8.1, 1.6 Hz,
1H), 5.45 – 5.17 (m, 4H), 4.56 – 4.41 (m, 1H), 3.66 (s, 3H), 2.62 – 2.55 (m, 1H), 2.40 –
2.18 (m, 6H), 1.80 (q, J = 7.5 Hz, 1H), 1.09 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ
193.46, 173.42, 158.61, 135.80, 135.74, 133.43, 133.10, 131.03, 130.85, 129.96, 129.95,
128.79, 128.04, 127.70, 123.65, 77.48, 77.30, 77.18, 76.98, 76.84, 76.66, 72.14, 51.54,
34.95, 33.89, 31.57, 26.95, 25.59, 22.75, 22.63, 19.29, 14.10.

COOMe
(S) (S)
OTBDPS
O
130

Methyl (10S, 4Z, 7Z, 11E, 13E)-10-(tert-Butyl-diphenyl-silanyloxy)-14-
iodotetradeca-4,7,11,13-tetraenoate (5.5). To a solution of CrCl2 (600 mg, 4.9 mmol)
dissolved in THF (10 mL total volume) was cannulated a mixture of compound 5.16 (400
mg, 0.82 mmol) and CHI3 (1.0 g, 2.46 mmol) dissolved in anhydrous THF (10 mL) under
Argon at 0°C. The reaction was stirred at 0°C for 3 h and an additional 1 h at room
temperature. The reaction mixture was quenched with water (20 mL) extracted with Et2O
(3 x 20 mL) rinsed with brine and dried over MgSO4. The organic phase was filtered and
the solvent was removed in vacuo to afford a crude oil which was purified on silica gel
using first pure pentanes and then EtOAc-hexanes (1:24) as the eluent to afford
compound 5.5 (206 mg, 68%) as a clear colorless oil.
1
H NMR (400 MHz, CDCl3) δ
7.74 – 7.56 (m, 4H), 7.45 – 7.27 (m, 6H), 6.91 (dd, J = 14.4, 10.6 Hz, 1H), 6.16 (d, J =
14.3 Hz, 1H), 5.88 (dd, J = 15.4, 10.7 Hz, 1H), 5.65 (dd, J = 15.3, 6.4 Hz, 1H), 5.35 –
5.23 (m, 4H), 4.25 – 4.10 (m, 1H), 3.67 (s, 3H), 2.67 – 2.51 (m, 2H), 2.38 – 2.14 (m, 6H),
1.06 (s, 9H).
13
C NMR (400 MHz, CDCl3) δ 175.21, 144.81, 136.77, 136.02, 130.00,
129.87, 129.83, 129.77, 129.38, 127.92, 127.69, 127.64, 127.59, 125.04, 78.86, 77.48,
77.16, 76.84, 73.19, 35.79, 34.14, 27.15, 27.10, 25.80, 22.90.

COOMe
(S) (S)
OTBDPS
I
(R) (R)
TBDPSO
Br
Br
131
(3R, 1E, 4Z)-3-(t-butyldimethylsilyloxy)-1-dibromoocta-1,5-diene (5.17). To a
solution of CBr4 (2.14 g, 8.16 mmol) at 0°C in anhydrous CH2Cl2 (15 mL total volume)
was cannulated PPh3 (1.35 g, 4.08 mmol) to give a clear yellow solution. To the reaction
mixture at 0°C was added aldehyde 3.X (1.0 g, 2.72 mmol). The reaction was run for 1 h
at 0°C to assure completion. Without workup the solvent was evaporated in vacuo and
the crude mixture was purified on silica gel using EtOAc-hexanes mixture of (1:24) as
the eluent to afford the product 5.17 (1.13 g, 80%) as a viscous and colorless oil.
1
H
NMR (400 MHz, CDCl3) δ 7.66 (ddd, J = 13.1, 7.9, 1.6 Hz, 4H), 7.46 – 7.33 (m, 6H),
6.41 (d, J = 8.2 Hz, 1H), 5.52 – 5.40 (m, 1H), 5.39 – 5.27 (m, 1H), 4.35 (dd, J = 14.3, 6.1
Hz, 1H), 2.28 (ddt, J = 26.3, 12.5, 6.6 Hz, 2H), 1.93 (td, J = 7.5, 1.5 Hz, 2H), 1.07 (s, 9H),
0.90 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3) δ 141.01, 135.89, 135.87, 134.72,
133.59, 133.51, 129.76, 127.64, 122.75, 89.05, 73.94, 34.55, 26.96, 20.66, 19.29, 14.21.

(6R, 3Z)-6-(tert-butyldiphenylsilyloxy)-7-ynyl (5.4). To a solution of di-bromo olefin
5.17 (1.0 g, 1.91 mmol) at -78°C in anhydrous Et2O (10 mL) was added 2.5 M solution of
n-BuLi (2.29 mL, 5.74 mmol) drop wise and stirred for 1 h. The reaction was quenched
with water (30 mL) and extracted with Et2O (3 x 10 mL), dried using MgSO4, filtered and
concentrated. The crude was then purified using silica gel with a EtOAc-hexanes eluent
of (1:24) to afford the product 5.4 (0.67 g, 98%).
1
H NMR (500 MHz, CDCl3) δ 7.70 –
7.57 (m, 4H), 7.38 – 7.25 (m, 6H), 5.43 – 5.34 (m, 1H), 5.35 – 5.26 (m, 1H), 4.25 (td, J =
6.5, 2.1 Hz, 1H), 2.33 (t, J = 6.9 Hz, 2H), 2.24 (d, J = 2.1 Hz, 1H), 1.87 – 1.76 (m, 2H),
(R) (R)
TBDPSO
132
1.01 (s, 9H), 0.80 (t, J = 7.5 Hz, 3H).
13
C NMR (500 MHz, CDCl3) δ 136.06, 136.05,
136.04, 135.87, 135.86, 134.59, 133.54, 133.37, 129.77, 129.68, 127.60, 127.59, 127.55,
127.44, 123.16, 84.84, 77.27, 77.01, 76.76, 72.63, 63.63, 36.17, 26.90, 20.67, 19.29,
14.20.

(4R, 2E, 6Z)-4-(tert-butyldiphenylsilyloxy)-1-oxonona-2,6-dienal (5.18). To a solution
of compound 3.X (460 mg, 1.25 mmol) dissolved in Toluene (10 mL) was added
(Triphenylphosphoranylidene)acetaldehyde (380 mg, 1.25 mmol) and heated to reflux
overnight. The solvent was removed in vacuo without work up and the crude mixture
was purified on silica gel using a EtOAc-hexanes mixture of (1:12) as the eluent to afford
the product 5.18 (392 mg, 80%) as a clear viscous oil.

(5R, 1E, 3E, 6Z)-5-(tert-butyldiphenylsilyloxy)-1,1-dibromodoca-1,3,7-trienal (5.19).
To a solution of CBr4 (318 mg, 0.96 mmol) at 0°C in anhydrous CH2Cl2 (20 mL total
volume) was cannulated PPh3 (503 mg, 1.92 mmol) to give a clear yellow solution. To
the reaction mixture at 0°C was added aldehyde 5.18 (190 mg, 0.48 mmol). The reaction
was run for 1 h to assure completion. Without workup the solvent was evaporated in
vacuo and the crude mixture was purified on silica gel using EtOAc-hexanes mixture of
(R) (R)
TBDPSO
O
(R) (R)
TBDPSO
Br
Br
133
(1:20) as the eluent to afford the product 5.19 (189 mg, 72%) as a viscous and colorless
oil.

(6R, 3Z, 7E)-6-(tert-butyldiphenylsilyloxy)-doca-3,7-dien-9-yn-al (5.6). To a solution
of di-bromo olefin 5.19 (200 mg, 0.37 mmol) at -78°C in anhydrous THF (5 mL) was
added 2.5 M solution of LDA (0.44 mL, 1.1 mmol) drop wise and stirred for 0.5 h. The
reaction was warmed to room temperature and stirred for an additional 0.5 h. Upon
completion the reaction mixture was quenched with water (10 mL) and extracted with
Et2O (3 x 10 mL), dried using MgSO4, filtered and concentrated. The crude was then
purified using silica gel with a EtOAc-hexanes eluent of (1:25) to afford the product 5.6
(104 mg, 72%).
1
H NMR (400 MHz, CDCl3) δ 7.75 – 7.60 (m, 4H), 7.48 – 7.33 (m, 6H),
6.23 (ddd, J = 15.9, 5.6, 0.7 Hz, 1H), 5.60 (ddd, J = 16.0, 2.3, 1.5 Hz, 1H), 5.44 – 5.33 (m,
1H), 5.26 – 5.15 (m, 1H), 4.27 – 4.17 (m, 1H), 2.85 (dt, J = 2.3, 0.5 Hz, 1H), 2.30 – 2.09
(m, 2H), 1.79 (p, J = 7.4 Hz, 2H), 1.09 (s, 9H), 0.85 (t, J = 7.5 Hz, 3H).
13
C NMR (400
MHz, CDCl3) δ 147.38, 136.03, 136.01, 134.42, 134.15, 133.68, 129.88, 129.86, 127.73,
123.22, 108.36, 82.23, 77.51, 73.17, 35.52, 27.17, 20.71, 19.51, 14.22.

(R) (R)
TBDPSO
COOMe
(R) (R)
(R) (R)
OTBDPS
OTBDPS
134
Methyl (10R, 17R, 4Z, 7Z, 11E, 13E, 19Z)-10,17-bis-(tert-butyldiphenylsilyloxy)-
docosa-4,7,11,13,19-heptaene-15-ynoate, (5.20). To the arm of a three-necked flask
was charged Pd(PPh3)4 (6 mg, .006 mmol) and CuI (2 mg, .011 mmol) under Argon. A
solution of compound 5.5 (65 mg, 0.11 mmol) compound 5.6 (38 mg, 0.11 mmol) and
Et3N (0.07 mL, .55 mmol) in C6H6 (3 mL) was cannulated into the reaction vessel. The
reaction flask was then freeze thawed with liquid nitrogen three times to remove an
oxygen. After removing any oxygen from the reaction flask the Pd(PPh 3)4 and CuI was
added and the reaction mixture was stirred overnight at room temperature. The reaction
was worked up with aqueous saturated NH4Cl (5 mL) and extracted with Et2O (3 x 10
mL). The solvent was evaporated and the mixture was purified on silica gel using
EtOAc-hexanes (1:50) as the eluent to afford compound 5.20 (75 mg, 80%) as a clear oil.
1
H NMR (500 MHz, CDCl3) δ 7.84 – 7.56 (m, 8H), 7.53 – 7.30 (m, 12H), 6.20 (dd, J =
15.6, 10.8 Hz, 1H), 5.89 (dd, J = 15.2, 10.9 Hz, 1H), 5.67 (dd, J = 15.2, 6.6 Hz, 1H), 5.52
– 5.43 (m, 1H), 5.44 – 5.35 (m, 1H), 5.36 – 5.26 (m, 4H), 4.46 (td, J = 6.5, 1.8 Hz, 1H),
4.25 – 4.18 (m, 1H), 3.66 (d, J = 1.9 Hz, 3H), 2.69 – 2.59 (m, 2H), 2.44 (t, J = 6.9 Hz,
2H), 2.39 – 2.19 (m, 6H), 1.09 (s, 9H), 1.07 (s, 9H), 0.90 (t, J = 7.5 Hz, 3H).
13
C NMR
(500 MHz, CDCl3) δ 173.65, 141.07, 138.04, 136.21, 136.07, 136.05, 136.03, 134.51,
134.28, 134.03, 133.90, 133.83, 129.89, 129.81, 129.79, 129.74, 129.64, 129.53, 129.43,
127.90, 127.69, 127.67, 127.60, 127.50, 125.23, 123.72, 110.73, 93.15, 84.38, 73.56,
64.51, 64.49, 51.69, 36.48, 35.98, 34.16, 29.85, 27.17, 27.08, 25.81, 22.92, 20.85, 19.49,
19.45, 14.38, 14.35.
135

Methyl (10S, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 13E, 19Z) heptaene-15-ynoate
(5.21). To a solution of compound 5.20 (44 mg, 0.05 mmol) in THF (5 mL) was added
1.0 M solution of TBAF (0.25 mL, 0.25 mmol) slowly at 0°C and stirred for 2 h. The
reaction was quenched with water (10 mL) and extracted with Et2O (5 x 10 mL), rinsed
with brine, dried over MgSO4 and filtered. The solvent was the concentrated and freshly
prepared CH2N2 was added to convert any acid to the ester. The solvent was completely
removed in vacuo and the compound was purified via C-18 reversed Phase HPLC using
H2O-MeOH mixture (30%) to afford compound 5.21 (9.3 mg, 50%) as colorless oil.
1
H
NMR (400 MHz, CDCl3) δ 6.57 (dd, J = 15.5, 10.9 Hz, 1H), 6.30 (dd, J = 15.2, 10.9 Hz,
1H), 5.83 (dd, J = 15.2, 6.0 Hz, 1H), 5.67 – 5.59 (m, 2H), 5.55 (dd, J = 17.7, 7.5 Hz, 1H),
5.49 – 5.32 (m, 4H), 4.53 (q, J = 5.7 Hz, 1H), 4.29 – 4.18 (m, 1H), 3.67 (s, 3H), 2.89 –
2.75 (m, 2H), 2.51 (t, J = 6.8 Hz, 2H), 2.43 – 2.32 (m, 6H), 2.10 (p, J = 7.3 Hz, 2H), 1.90
(d, J = 6.0 Hz, 1H), 1.81 (d, J = 4.1 Hz, 1H), 0.98 (t, J = 7.5 Hz, 3H).
13
C NMR (400
MHz, CDCl3) δ 173.72, 141.44, 138.20, 136.11, 131.59, 129.37, 129.05, 128.20, 127.23,
124.73, 122.75, 110.89, 77.42, 77.10, 76.78, 72.86, 71.56, 62.64, 54.49, 51.71, 46.99,
35.71, 35.41, 34.03, 31.14, 25.89, 22.93, 20.91, 14.32.
COOMe
(R) (R)
(R) (R)
OH
OH
136

Methyl (10R, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 13E, 15Z, 19Z)-hexaenoate, AT-
NPD1/PD1 Methyl Ester (5.1). A flame dried flask was charged with a freshly prepared
Zn/Cu/Ag amalgam (490 mg, excess) and suspended in H2O-MeOH mixture (1:1, 3 mL).
To the reaction slurry was added compound 5.21 (5.5 mg, 0.015 mmol) and stirred for 3.5
h while monitoring. The reaction was filtered dried and purified via HPLC at H2O-
MeOH mixture (30%) to afford compound 5.1 (3.2 mg, 56%).
1
H NMR (400 MHz,
CDCl3): δ 6.51 (dd, J = 14.0 Hz and 11.4 Hz, 1H), 6.26 (m, 2H), 6.09 (dd, J = 11.0 and
11.0 Hz, 1H), 5.62–5.30 (m, 7H), 4.60 (m, 1H), 4.23 (m, 1H), 3.67 (s, 3H), 2.83 (m, 2H),
2.44–2.22 (m, 8H), 2.07 (m, 2H), 0.96 (t, J = 7.5 Hz, 3H).
13
C NMR (400 MHz, CDCl3):
δ 173.64, 136.48, 135.32, 133.78, 133.46, 131.21, 130.32, 130.03, 129.02, 128.02, 127.78,
124.85, 123.49, 71.81, 67.70, 35.37, 33.94, 31.58, 25.79, 22.82, 22.64, 14.10.

Methyl (10S, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 13E, 15Z, 19Z)-hexaenoate, 10-
epi AT-NPD1/PD1 Methyl Ester (5.1’)
Same As, AT-PD1
COOMe
(R) (R)
(R) (R)
OH
OH
137

Methyl (10R, 17R, 4Z, 7Z, 11E, 13E, 19Z)-10,17-bis-(tert-butyldiphenylsilyloxy)-
docosa-4,7,11,15,19-heptaene-13-ynoate, (5.23). To the arm of a three-necked flask
was charged Pd(PPh3)4 (3 mg, .0025 mmol) and CuI (1 mg, .005 mmol) under Argon. A
solution of compound 5.7 (29 mg, 0.049 mmol) compound 5.6 (21 mg, 0.054 mmol) and
Et3N (0.06 mL, .49 mmol) in C6H6 (1 mL) was cannulated into the reaction vessel. The
reaction flask was then freeze thawed with liquid nitrogen three times to remove an
oxygen. After removing any oxygen from the reaction flask the Pd(PPh 3)4 and CuI was
added and the reaction mixture was stirred overnight at room temperature. The reaction
was worked up with aqueous saturated NH4Cl (3 mL) and extracted with Et2O (3 x 3 mL).
The solvent was evaporated and the mixture was purified on silica gel using EtOAc-
hexanes (1:50) as the eluent to afford compound 5.23 (39 mg, 96%) as a clear oil.

COOMe
(R) (R)
OTBDPS
(R) (R) OTBDPS
138

Methyl (10S, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 15E, 19Z) heptaene-13-ynoate,
(5.24). To a solution of compound 5.23 (30 mg, 0.035 mmol) in THF (3 mL) was added
1.0 M solution of TBAF (0.142 mL, 0.142 mmol) at 0°C and stirred for 2 h. The reaction
was quenched with water (5 mL) and extracted with Et2O (5 x 5 mL), rinsed with brine,
dried over MgSO4 and filtered. The solvent was the concentrated and freshly prepared
CH2N2 was added to convert any acid to the ester. The solvent was completely removed
in vacuo and the compound was purified via C-18 reversed Phase HPLC using H2O-
MeOH mixture (30%) to afford compound 5.24 (8.5 mg, 65%) as colorless oil.
1
H NMR
(400 MHz, CDCl3) δ 6.15 (ddd, J = 16.2, 5.6, 1.6 Hz, 1H), 5.85 (d, J = 15.2 Hz, 0H), 5.65
– 5.49 (m, 0H), 5.47 – 5.26 (m, 1H), 4.30 – 4.13 (m, 1H), 3.67 (s, 1H), 2.83 (q, J = 5.7,
5.1 Hz, 0H), 2.42 – 2.24 (m, 2H), 2.13 – 1.98 (m, 0H), 0.97 (t, J = 7.5 Hz, 2H).
13
C
NMR (400 MHz, CDCl3) δ 173.63, 144.76, 144.68, 135.84, 131.60, 128.90, 128.09,
124.40, 123.04, 110.01, 109.95, 88.19, 88.13, 71.52, 71.47, 51.60, 35.05, 34.89, 33.92,
25.78, 22.81, 20.72, 14.15.
COOMe
(R) (R)
OH
(R) (R) OH
139

Methyl (10R, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 13Z, 15E, 19Z)-hexaenoate, Δ
13
-
Cis, Δ
15
-trans AT-NPD1/PD1 Methyl Ester (5.3). A flame dried flask was charged
with a freshly prepared Zn/Cu/Ag amalgam (300 mg, excess) and suspended in H2O-
MeOH mixture (1:1, 3 mL). To the reaction slurry was added compound 5.24 (2.75 mg,
7.0 x 10
-3
mmol) and stirred for 4 h while monitoring. The reaction was filtered dried
and purified via HPLC at H2O-MeOH mixture (30%) to afford compound 5.3 (1.5 mg,
60%).
1
H NMR (600 MHz, CDCl3) δ 6.77 – 6.67 (m, 2H), 6.09 – 5.89 (m, 2H), 5.76
(ddd, J = 15.0, 6.2, 1.2 Hz, 2H), 5.63 – 5.49 (m, 2H), 5.46 – 5.32 (m, 4H), 4.25 (ddd, J =
16.9, 11.3, 5.7 Hz, 2H), 3.66 (s, 3H), 2.83 (q, J = 6.9 Hz, 2H), 2.44 – 2.30 (m, 6H), 2.10 –
2.02 (m, 2H), 1.87 (d, J = 3.3 Hz, 1H), 1.81 (d, J = 3.5 Hz, 1H), 0.96 (t, J = 7.5 Hz, 3H).
13
C NMR (600 MHz, CDCl3) δ 173.78, 136.89, 136.72, 135.54, 131.35, 129.18, 129.13,
129.02, 128.17, 125.62, 125.53, 125.07, 123.73, 72.02, 71.98, 58.59, 53.56, 51.74, 35.57,
35.42, 34.10, 25.95, 22.98, 20.89, 14.34.

Methyl (10S, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 13Z, 15E, 19Z)-hexaenoate, Δ
13
-
Cis, Δ
15
-trans, 10-epi AT-NPD1/PD1 Methyl Ester
Same As Δ
13
-cis Δ
15
-trans, AT-PD1
COOMe
(R) (R)
OH
(R) (R) OH
140

Methyl (10R, 17R, 4Z, 7Z, 11E, 13E, 15E, 19Z)-10,17-bis-(tert-
butyldiphenylsilyloxy)-docosa-4,7,11,15,19-hexaenoate, (5.23a). To a flask of iodide
5.7 (50 mg, 0.085 mmol) was added Pd(Ph3)4 (10 mg, 0.0085 mmol) and stirred at room
temperature. In a separate flask Schwartz reagent was suspended in THF (1 mL) and
alkyne 5.6 (50 mg, 0.128 mmol) was cannulated and stirred at 50°C for 1 h. The reaction
was allowed to cool to room temperature before adding fresh 0.5M ZnCl2 in THF (0.34
mL, 0.17 mmol). After stirring for 10 min the Zn / alkyne mixture was transferred to the
Pd / vinyl iodide and stirred overnight at room temperature. The reaction mixture was
quenched with saturated NH4Cl (5 mL) and extracted with diethyl ether (3 x 5 mL). The
organic phase was rinsed with brine (10 mL) and the organic layer was concentrated to
afford a crude oil purified quickly by silica gel using EtOAc-hexanes (1:50) as the eluent
to afford compound 5.23a (28 mg, 38%) as a clear colorless oil.

Methyl (10R, 17R)-Dihydroxydocosa-(4Z, 7Z, 11E, 13E, 15E, 19Z)-hexaenoate, Δ
15
-
trans AT-NPD1/PD1 Methyl Ester (5.2) To a solution of compound 5.23a (20 mg,
0.024 mmol) in THF (2 mL) was added 1.0 M solution of TBAF (0.12 mL, 0.12 mmol) at
0°C and stirred for 2 h. The reaction was quenched with water (4 mL) and extracted with
Et2O (5 x 5 mL), rinsed with brine, dried over MgSO4 and filtered. The solvent was the
(R) (R)
(R) (R)
OTBDPS
OTBDPS
COOMe
(R) (R)
(R) (R)
OH
OH
COOMe
141
concentrated and freshly prepared CH2N2 was added to convert any acid to the ester. The
solvent was completely removed in vacuo and the compound was purified via C-18
reversed Phase HPLC using H2O-MeOH mixture (30%) to afford compound 5.2 (5.8 mg,
65%) as colorless oil.
1
H NMR (600 MHz, CDCl3) δ 6.31 – 6.23 (m, 2H), 6.23 – 6.18 (m,
2H), 5.74 (ddd, J = 14.8, 6.3, 1.5 Hz, 2H), 5.60 – 5.53 (m, 2H), 5.55 – 5.49 (m, 2H), 5.46
– 5.38 (m, 2H), 5.40 – 5.31 (m, 2H), 4.21 (dt, J = 10.5, 5.6 Hz, 2H), 3.67 (s, 3H), 2.83 (q,
J = 7.0 Hz, 2H), 2.44 – 2.25 (m, 8H), 2.09 – 2.02 (m, 2H), 0.97 (t, 3H).
13
C NMR (600
MHz, CDCl3) δ 173.78, 135.99, 135.86, 135.55, 132.44, 132.35, 131.32, 130.55, 130.47,
129.20, 128.19, 125.08, 123.72, 72.07, 72.02, 35.55, 35.40, 34.11, 25.95, 22.98, 14.35.

142
5.6 References

95. Serhan, C. N.; Gotlinger, K.; Hong, S.; Lu, Y.; Siegelman, J.; Baer, T.; Yang, R.;
Colgan, S. P.; Petasis, N. A. Journal of Immunology 2006, 176, 1848.
96. Yang, R. Synthesis of Study of Novel Lipid Mediators 2006, PhD thesis.
97. Serhan, C. N.; Fredman, G.; Yang, R.; Karamnov, S.; Belayev, L. S.; Bazan, N. G.;
Zhu, M.; Winkler, J. W.; Petasis, N. A. Chemistry & Biology 2011, 18, 976.
98. Bazan, N.G.; Trends Neurosci. 2006, 29, 263-271.
99. Claria, J.; Serhan, C. N. Proceedings of the National Academy of Sciences of the
United States of America 1995, 92, 9475.
100. Zhao, Y.; Calon, F.; Julien, C.; Winkler, J. W.; Petasis, N. A.; Lukiw, W. J.; Bazan,
N. G. Plos One 2011, 6.
101. Calandria, J. M.; Marcheselli, V. L.; Mukherjee, P. K.; Uddin, J.; Winkler, J. W.;
Petasis, N. A.; Bazan, N. G. Journal of Biological Chemistry 2009, 284, 17877.
102. Marcheselli, V. L.; Mukherjee, P. K.; Arita, M.; Hong, S.; Antony, R.; Sheets, K.;
Winkler, J. W.; Petasis, N. A.; Serhan, C. N.; Bazan, N. G. Prostaglandins
Leukotrienes and Essential Fatty Acids 2010, 82, 27.
103. Bazan, N. G.; Eady, T. N.; Khoutorova, L.; Atkins, K. D.; Hong, S.; Lu, Y.; Zhang,
C.; Jun, B.; Obenaus, A.; Fredman, G.; Zhu, M.; Winkler, J. W.; Petasis, N. A.;
Serhan, C. N.; Belayev, L.Experimental Neurology 2012, 236, 122.
104. Petasis, N. A.; Yang, R.; Winkler, J. W.; Zhu, M.; Uddin, J.; Bazan, N. G.; Serhan,
C. N. Tetrahedron Letters 2012, 53, 1695.
105. Negishi, E.; Alimardanov, A.; Xu, C. D. Organic Letters 2000, 2, 65.
106. Evano, G.; Blanchard, N.; Toumi, M. Chemical Reviews 2008, 108, 3054.
143

107. Petasis, N. A.; Winkler, J. W.; Zhu, M.; Bazan, N. G.; Serhan, C. N. Abstracts of
Papers of the American Chemical Society 2012, 243.

144
BIBLIOGRAPHY
1. Antony, R.; Lukiw, W. J.; Bazan, N. G. Journal of Biological
Chemistry 2010, 285, 18301.
2. Arita, M.; Yoshida, M.; Hong, S.; Tjonahen, E.; Glickman, J. N.; Petasis, N. A.;
Blumberg, R. S.; Serhan, C. N. Proceedings of the National Academy of Sciences
of the United States of America 2005, 102, 7671.
3. Bannenberg, G. L.; Chiang, N.; Ariel, A.; Arita, M.; Tjonahen, E.; Gotlinger, K.
H.; Hong, S.; Serhan, C. N. Journal of Immunology 2005, 174, 4345.
4. Bazan, N. G. Brain Pathology 2005, 15, 159.
5. Bazan, N. G.; Eady, T. N.; Khoutorova, L.; Atkins, K. D.; Hong, S.; Lu, Y.;
Zhang, C.; Jun, B.; Obenaus, A.; Fredman, G.; Zhu, M.; Winkler, J. W.; Petasis,
N. A.; Serhan, C. N.; Belayev, L.Experimental Neurology 2012, 236, 122.
6. Bazan, N. G.; Molina, M. F.; Gordon, W. C. Annual Review of Nutrition, Vol
31 2011, 31, 321.
7. Belayev, L.; Khoutorova, L.; Atkins, K. D.; Eady, T. N.; Hong, S.; Lu, Y.;
Obenaus, A.; Bazan, N. G. Translational Stroke Research 2011, 2, 33.
8. Bento, A. F.; Claudino, R. F.; Dutra, R. C.; Marcon, R.; Calixto, J. B. Journal of
Immunology 2011, 187, 1957.
9. Boland, W.; Sieler, C.; Feigel, M. Helvetica Chimica Acta 1987, 70, 1025.
10. Bundgaard, H.; Falch, E.; Larsen, C.; Mosher, G. L.; Mikkelson, T. J. Journal of
Medicinal Chemistry 1985, 28, 979.
11. Burr, G. O.; Burr, M. M. Journal of Biological Chemistry 1929, 82, 345.
12. Burr, G. O.; Burr, M. M. Journal of Biological Chemistry 1930, 86, 0587.
13. Burr, M. L.; Gilbert, J. F.; Holliday, R. M.; Elwood, P. C.; Fehily, A. M.; Rogers,
S.; Sweetnam, P. M.; Deadman, N. M. Lancet 1989, 2, 757.
14. Calandria, J. M.; Marcheselli, V. L.; Mukherjee, P. K.; Uddin, J.; Winkler, J. W.;
Petasis, N. A.; Bazan, N. G. Journal of Biological Chemistry 2009, 284, 17877.
15. Chemin, D.; Linstrumelle, G. Tetrahedron 1992, 48, 1943.
16. Chen, P.; Fenet, B.; Michaud, S.; Tomczyk, N.; Vericel, E.; Lagarde, M.;
Guichardant, M. Febs Letters 2009, 583, 3478.
145
17. Chiang, N.; Schwab, J. M.; Fredman, G.; Kasuga, K.; Gelman, S.; Serhan, C.
N. Plos One 2008, 3.
18. Claria, J.; Serhan, C. N. Proceedings of the National Academy of Sciences of the
United States of America 1995, 92, 9475.
19. Claria, J.; Dalli, J.; Yacoubian, S.; Gao, F.; Serhan, C. N. Journal of
Immunology 2012, 189, 2597.
20. Corey, E. J.; Fuchs, P. L. Tetrahedron Letters 1972, 3769.
21. Corey, E. J.; Raju, N. Tetrahedron Letters 1983, 24, 5571.
22. Dalli, J.; Winkler, J. W.; Colas, R. A.; Arnardottir, H.; Cheng, C.-Y. C.; Chiang,
N.; Petasis, N. A.; Serhan, C. N. Chemistry & Biology 2013, 20, 188.
23. de Lorgeril, M.; Salen, P.; Martin, J. L.; Monjaud, I.; Delaye, J.; Mamelle,
N. Circulation 1999, 99, 779.
24. de Lorgeril, M.; Renaud, S.; Mamelle, N.; Salen, P.; Martin, J. L.; Monjaud, I.;
Guidollet, J.; Touboul, P.; Delaye, J. Lancet 1994, 343, 1454.
25. Duffield, J. S.; Hong, S.; Vaidya, V. S.; Lu, Y.; Fredman, G.; Serhan, C. N.;
Bonventre, J. V. Journal of Immunology 2006, 177, 5902.
26. Estruch, R.; Ros, E.; Salas-Salvado, J.; Covas, M.-I.; Corella, D.; Aros, F.;
Gomez-Gracia, E.; Ruiz-Gutierrez, V.; Fiol, M.; Lapetra, J.; Maria Lamuela-
Raventos, R.; Serra-Majem, L.; Pinto, X.; Basora, J.; Angel Munoz, M.; Sorli, J.
V.; Alfredo Martinez, J.; Angel Martinez-Gonzalez, M.; Investigators, P. S. New
England Journal of Medicine 2013, 368, 1279.
27. Evano, G.; Blanchard, N.; Toumi, M. Chemical Reviews 2008, 108, 3054.
28. Fox, M. E.; Jackson, M.; Lennon, I. C.; McCague, R. Journal of Organic
Chemistry 2005, 70, 1227.
29. Frost, A.A.; Pearson R.G. Kinetics and mechanism. A study of homogeneous
chemical reactions. 2nd ed. New York: John Wiley and Sons, 1961; 317–34.
30. Gallos, J. K.; Koumbis, A. E.; Xiraphaki, V. P.; Dellios, C. C.; Coutouli-
Argyropoulou, E. Tetrahedron 1999, 55, 15167.
31. Gonzalez-Periz, A.; Claria, J. The Scientific World Journal 2010, 10, 832.
32. Haeggstrom, J. Z.; Hamberg, M. Chemistry & Biology 2013, 20, 138.
33. Hennessy, S. A.; Moane, S. M.; McDermott, S. D. Journal of Forensic
Sciences, 2004, 49, 1220.
146
34. Hong, S.; Gronert, K.; Devchand, P. R.; Moussignac, R. L.; Serhan, C. N. Journal
of Biological Chemistry 2003, 278, 14677.
35. Hong, S.; Tjonahen, E.; Morgan, E. L.; Lu, Y.; Serhan, C. N.; Rowley, A.
F. Prostaglandins & Other Lipid Mediators 2005, 78, 107.
36. Hotchkiss, R. S.; Karl, I. E. New England Journal of Medicine 2003, 348, 138.
37. Ji, R.-R.; Xu, Z.-Z.; Strichartz, G.; Serhan, C. N. Trends in
Neurosciences 2011, 34, 599.
38. Jin, Y.; Arita, M.; Zhang, Q.; Saban, D. R.; Chauhan, S. K.; Chiang, N.; Serhan, C.
N.; Dana, R. Investigative Ophthalmology & Visual Science 2009, 50, 4743.
39. Krauss, R. M.; Eckel, R. H.; Howard, B.; Appel, L. J.; Daniels, S. R.;
Deckelbaum, R. J.; Erdman, J. W.; Kris-Etherton, P.; Goldberg, I. J.; Kotchen, T.
A.; Lichtenstein, A. H.; Mitch, W. E.; Mullis, R.; Robinson, K.; Wylie-Rosett, J.;
Jeor, S. S.; Suttie, J.; Tribble, D. L.; Bazzarre, T. L. Circulation 2000, 102, 2284.
40. Kris-Etherton, P.; Eckel, R. H.; Howard, B. V.; St Jeor, S.; Bazzarre, T.
L. Circulation 2001, 103, 1823.
41. Lapitskaya, M. A.; Vasiljeva, L. L.; Pivnitsky, K. K. Synthesis-Stuttgart 1993, 65.
42. Larjava, H. (Ed.). (2012). Oral Wound Healing: Cell Biology and Clinical
Management (1st ed., pp. 57–78). West Sussex, UK
43. Levy, B. D.; Kohli, P.; Gotlinger, K.; Haworth, O.; Hong, S.; Kazani, S.; Israel,
E.; Haley, K. J.; Serhan, C. N. Journal of Immunology 2007, 178, 496.
44. Lichtenstein, A. H.; Appel, L. J.; Brands, M.; Carnethon, M.; Daniels, S.; Franch,
H. A.; Franklin, B.; Kris-Etherton, P.; Harris, W. S.; Howard, B.; Karanja, N.;
Lefevre, M.; Rudel, L.; Sacks, F.; Van Horn, L.; Winston, M.; Wylie-Rosett,
J. Circulation 2006, 114, 82.
45. Lu, W.; Zheng, G. R.; Gao, D. X.; Cai, J. C. Tetrahedron 1999, 55, 7157.
46. Lukiw, W. J.; Cui, J. G.; Marcheselli, V. L.; Bodker, M.; Botkjaer, A.; Gotlinger,
K.; Serhan, C. N.; Bazan, N. G. Journal of Clinical Investigation 2005, 115, 2774.
47. March J. Advanced organic chemistry. 4th ed. New York: John Wiley & Sons.
48. Marcheselli, V. L.; Mukherjee, P. K.; Arita, M.; Hong, S.; Antony, R.; Sheets, K.;
Winkler, J. W.; Petasis, N. A.; Serhan, C. N.; Bazan, N. G. Prostaglandins
Leukotrienes and Essential Fatty Acids 2010, 82, 27.
49. Martin, N.; Ruddick, A.; Arthur, G. K.; Wan, H.; Woodman, L.; Brightling, C. E.;
Jones, D. J. L.; Pavord, I. D.; Bradding, P. Plos One 2012, 7.
147
50. Maryanoff, B. E.; Reitz, A. B. Chemical Reviews 1989, 89, 863.
51. Mas, E.; Croft, K. D.; Zahra, P.; Barden, A.; Mori, T. A. Clinical
Chemistry 2012, 58, 1476.
52. Mukherjee, P. K.; Marcheselli, V. L.; Serhan, C. N.; Bazan, N. G. Proceedings of
the National Academy of Sciences of the United States of America 2004, 101,
8491.
53. Muller, S.; Liepold, B.; Roth, G. J.; Bestmann, H. J. Synlett 1996, 521.
54. Negishi, E.; Alimardanov, A.; Xu, C. D. Organic Letters 2000, 2, 65.
55. Niemoller, T. D.; Bazan, N. G. Prostaglandins & Other Lipid Mediators 2010, 91,
85.
56. Nordoy, A.; Goodnight, S. Arteriosclerosis. 1990, 10, 149-163.
57. Ogawa, N.; Kobayashi, Y. Tetrahedron Letters 2011, 52, 3001.
58. Oger, C.; Balas, L.; Durand, T.; Galano, J.-M. Chemical Reviews 2013, 113, 1313.
59. Ohira, S. Synthetic Communications 1989, 19, 561.
60. Ottawa: Minister of National Health and Welfare, Nutrition Recommendations.
1990, H-49-42, 1990E.
61. Park, C.-K.; Xu, Z.-Z.; Liu, T.; Lu, N.; Serhan, C. N.; Ji, R.-R. Journal of
Neuroscience 2011, 31, 18433.
62. Petasis, N. A.; Akritopoulou-Zanze, I.; Fokin, V. V.; Bernasconi, G.; Keledjian,
R.; Yang, R.; Uddin, J.; Nagulapalli, K. C.; Serhan, C. N. Prostaglandins
Leukotrienes and Essential Fatty Acids 2005, 73, 301.
63. Petasis, N. A.; Winkler, J. W.; Zhu, M.; Bazan, N. G.; Serhan, C. N. Abstracts of
Papers of the American Chemical Society 2012, 243.
64. Petasis, N. A.; Winkler, J.; Nagengast, E. S.; Uddin, J.; Serhan, C. N. Abstracts of
Papers of the American Chemical Society 2009, 237.
65. Petasis, N. A.; Yang, R.; Winkler, J. W.; Zhu, M.; Uddin, J.; Bazan, N. G.; Serhan,
C. N. Tetrahedron Letters 2012, 53, 1695.
66. Pillai, P. S.; Leeson, S.; Porter, T. F.; Owens, C. D.; Kim, J. M.; Conte, M. S.;
Serhan, C. N.; Gelman, S. Inflammation 2012, 35, 98.
67. Raatz, S. K.; Golovko, M. Y.; Brose, S. A.; Rosenberger, T. A.; Burr, G. S.;
Wolters, W. R.; Picklo, M. J., Sr. Journal of Agricultural and Food
Chemistry 2011, 59, 11278.
148
68. Roth, G. J.; Liepold, B.; Muller, S. G.; Bestmann, H. J. Synthesis-Stuttgart 2004,
59.
69. Rowley, A. F.; Lloydevans, P.; Barrow, S. E.; Serhan, C.
N. Biochemistry 1994, 33, 856.
70. Saito, S.; Hasegawa, T.; Inaba, M.; Nishida, R.; Fujii, T.; Nomizu, S.; Moriwake,
T. Chemistry Letters 1984, 1389.
71. Saito, S.; Ishikawa, T.; Kuroda, A.; Koga, K.; Moriwake,
T. Tetrahedron 1992, 48, 4067.
72. Schroer, N., Sieler, C., Bottmuhle, A. Der, & Feigel, M. Helv. Chim. Acta, 1987,
70, 1025–1040.
73. Schwab, J. M.; Chiang, N.; Arita, M.; Serhan, C. N. Nature 2007, 447, 869.
74. Seki, H.; Sasaki, T.; Ueda, T.; Arita, M. The Scientific World Journal 2010, 10,
818.
75. Serhan, C. N. Prostaglandins Leukotrienes and Essential Fatty Acids 2005, 73,
141.
76. Serhan, C. N.; Clish, C. B.; Brannon, J.; Colgan, S. P.; Chiang, N.; Gronert,
K. Journal of Experimental Medicine 2000, 192, 1197.
77. Serhan, C. N.; Gotlinger, K.; Hong, S.; Lu, Y.; Siegelman, J.; Baer, T.; Yang, R.;
Colgan, S. P.; Petasis, N. A. Journal of Immunology 2006, 176, 1848.
78. Serhan, C. N.; Hong, S.; Gronert, K.; Colgan, S. P.; Devchand, P. R.; Mirick, G.;
Moussignac, R. L. Journal of Experimental Medicine 2002, 196, 1025.
79. Serhan, C. N.; Savill, J. Nature Immunology 2005, 6, 1191.
80. Serhan, C. N. Annual Review of Immunology 2007, 25, 101.
81. Serhan, C. N.; Brain, S. D.; Buckley, C. D.; Gilroy, D. W.; Haslett, C.; O'Neill, L.
A. J.; Perretti, M.; Rossi, A. G.; Wallace, J. L. Faseb Journal 2007, 21, 325.
82. Serhan, C. N.; Chiang, N.; Van Dyke, T. E. Nature Reviews Immunology 2008, 8,
349.
83. Serhan, C. N.; Fredman, G.; Yang, R.; Karamnov, S.; Belayev, L. S.; Bazan, N.
G.; Zhu, M.; Winkler, J. W.; Petasis, N. A. Chemistry & Biology 2011, 18, 976.
84. Serhan, C. N.; Krishnamoorthy, S.; Recchiuti, A.; Chiang, N. Current Topics in
Medicinal Chemistry 2011, 11, 629.
85. Serhan, C. N.; Petasis, N. A. Chemical Reviews 2011, 111, 5922.
149
86. Serhan, C. N.; Yang, R.; Martinod, K.; Kasuga, K.; Pillai, P. S.; Porter, T. F.; Oh,
S. F.; Spite, M. Journal of Experimental Medicine 2009, 206, 15.
87. Simopoulos, A. P. American Journal of Clinical Nutrition 1991, 54, 438.
88. Spite, M.; Norling, L. V.; Summers, L.; Yang, R.; Cooper, D.; Petasis, N. A.;
Flower, R. J.; Perretti, M.; Serhan, C. N. Nature 2009, 461, 1287.
89. Stark, D. T.; Bazan, N. G. Molecular Neurobiology 2011, 43, 131.
90. Sun, Y.-P.; Oh, S. F.; Uddin, J.; Yang, R.; Gotlinger, K.; Campbell, E.; Colgan, S.
P.; Petasis, N. A.; Serhan, C. N. Journal of Biological Chemistry 2007, 282, 9323.
91. Takai, K.; Nitta, K.; Utimoto, K. Journal of the American Chemical
Society 1986, 108, 7408.
92. Tidwell, T. T. Org. React. (N.Y.) 1990, 39, 297.
93. Trichopoulou, A.; Costacou, T.; Bamia, C.; Trichopoulos, D. New England
Journal of Medicine 2003, 348, 2599.
94. U.S. Department of Agriculture and U.S. Department of Health and Human
Services. Dietary Guidelines for Americans, 2010. 7th Edition, Washington,
DC: U.S. Government Printing Office, Dec. 2010.
95. Valagussa, F.; Franzosi, M. G.; Geraci, E.; Mininni, N.; Nicolosi, G. L.; Santini,
M.; Tavazzi, L.; Vecchio, C.; Marchioli, R.; Bomba, E.; Chieffo, C.; Maggioni, A.
P.; Schweiger, C.; Tognoni, G.; Barzi, F.; Flamminio, A. V.; Marfisi, R. M.;
Olivieri, M.; Pera, C.; Polidoro, A.; Santoro, E.; Zama, R.; Pagliaro, L.; Correale,
E.; Del Favero, A.; Loi, U.; Marubini, E.; Campolo, L.; Casari, A.; Di Minno, G.;
Donati, M. B.; Galli, M.; Gattone, M.; Garattini, S.; Mancini, M.; Marino, P.;
Santoro, G. M.; Scardulla, C.; Specchia, G.; Cericola, A.; Di Gregorio, D.; Di
Mascio, R.; Levantesi, G.; Mantini, L.; Mastrogiuseppe, G.; Tucci, C.; Mocarelli,
P.; Baldinelli, R.; Ceriotti, F.; Colonna, A.; Cortese, C.; Fortunato, G.; Franzini,
C.; Gonano, F.; Graziani, M. S. Lancet, 1999, 354, 447.
96. Vance, D.E.; Vance, J.E. Biochemistry of lipids, lipoproteins, and membranes.
2008, (5th ed.) Amsterdam, Elsevier.
97. Weber, P.C.; Fish Fats and Your Health, 1989, Report No. 4, 9-18.
98. Webb, T. H.; Thomasco, L. M.; Schlachter, S. T.; Gaudino, J. J.; Wilcox, C.
S. Tetrahedron Letters 1988, 29, 6823.
99. Winkler, J. W.; Uddin, J.; Serhan, C. N.; Petasis, N. A. Organic Letters 2013, 15,
1424.
150
100. Wipf, P.; Tsuchimoto, T.; Takahashi, H. Pure and Applied Chemistry 1999, 71,
415.
101. Yang, R. Synthesis of Study of Novel Lipid Mediators 2006, PhD thesis.
102. Zhao, Y.; Calon, F.; Julien, C.; Winkler, J. W.; Petasis, N. A.; Lukiw, W. J.;
Bazan, N. G. Plos One 2011, 6.

151
APPENDIX.
1
H and
13
C Spectra of Substrates

2.12
O
O O
152

2.6
O
O
O
153

2.13a
COOMe
(S) (S)
TBSO
HO
154

2.13b
COOMe
(S) (S)
TBSO
TBDPSO
155

2.14 COOMe
(S) (S)
HO
TBDPSO
156

2.14a
7
COOMe
(S) (S)
HO
TBDPSO
157

2.15
7
COOMe
(S) (S)
O
TBDPSO
158

2.3
7
COOMe
(S) (S)
TBDPSO
I
159

160

161

162

2.19
(S) (S)
(R) (R)
O O
OH
163

2.20
(R) (R)
(R) (R)
O O
O
164

165

2.4
(R) (R)
(S) (S)
OTBS
TBSO
166

2.23
COOMe
(S) (S)
(R) (R)
(S) (S)
TBSO
OTBS
TBSO
167

2.2
COOMe
(S) (S)
(R) (R)
(S) (S)
HO
OH
HO
168

COOMe
(S) (S)
(R) (R) (S) (S)
HO
OH
HO
2.1
169

COOMe
(S) (S)
HO
COOMe
3.9
170

(S) (S)
HO
COOMe
HO
3.10
171

3.11
(S) (S)
TBSO
COOMe
TBSO
172

3.12
(S) (S)
TBSO
COOMe
HO
173

(S) (S)
TBSO
COOMe
O 3.13
174

(S) (S)
TBSO
COOMe
3.14
Br
Br
175

(S) (S)
TBSO
COOMe
3.5
176

(R) (R)
O
OTBS
3.7
177

OH
OTBS
TMS
3.16a
178

OTBS
OTBS
TMS
3.16
179

OTBS
OH
TMS
3.17
180

OTBS
O
TMS
3.18
181

OTBS
O
TMS
3.19
182

OTBS
(R) (R)
I
TMS
3.4
183

OTBS
(R) (R)
I
TMS
184

(S) (S)
OH
TBSO
3.21a
185

(S) (S)
TBSO
3.21
OTBDPS
186

(S) (S)
HO
3.22
OTBDPS
187

HO
(S) (S)
TBDPSO
3.23
188

O
(S) (S)
TBDPSO
3.24
189

(S) (S)
TBSO
3.3
I
190

OTBS
(R) (R)
(S) (S)
TBSO
COOMe
3.25
TMS
191

OTBS
(R) (R)
(S) (S)
TBSO
COOMe
3.25b
192

OTBS
(S) (S)
(R) (R)
TBSO
(S) (S)
TBSO
COOMe
4
17
11
3.26
193

OH
(S) (S)
(R) (R)
HO
(S) (S)
HO
COOH
4
17
11
3.26b Acid
194

OH
(S) (S)
(R) (R)
HO
(S) (S)
HO
COOH
4
17
11
RvD3 (3.1)
195

OH
(R) (R)
(R) (R)
HO
(S) (S)
HO
COOH
4
17
11
AT-RvD3 (3.2)
196
HPLC and
1
H and
13
C NMR spectra of Ester Lactone Mix

Methyl Ester Lactone
197

198

199

200

201

202

203

204

205

206

(R) (R) (S) (S)
COOMe
OTBS TBSO
(S) (S)
OTBS
207

H 6 H 7
H 8
H 9
H 10
H 11
H 16
H 15
H 14
H 13
H 20
H 19
208
LCMS data of new compounds

209

210

211

212

213

214

215

216

217

218

219

220

221

222

223

224

Abstract (if available)

Abstract This body of work is about the design, synthesis and activity of a new and exciting class of compounds made from Docosahexaenoic Acid (DHA) termed resolvins and protectins. DHA is an abundant omega 3 fatty acid heavily concentrated in the brain and retina. The reason these compounds have such importance is because they are derived from the essential fatty acid, omega 3, that plays a crucial role in health. The aim of this work is to design and synthesize mediators to elucidate and confirm the structures of these specialized pro-resolving compounds with their biological counterparts and determine their role in the regulation and expression of various cell types such as chemokines, neutrophils and other enzymes involved in inflammation.

Linked assets

University of Southern California Dissertations and Theses

Conceptually similar

PDF

Analytical investigation of the proteasome inhibitor Bortezomib and the total synthesis of specialized pro-resolving lipid mediators

PDF

Total synthesis of specialized pro-resolving lipid mediators and their analogs

PDF

Studies on lipid mediators, and on potential modulators of GRP78

PDF

Total synthesis of specialized pro-resolving lipid mediators and their analogs

PDF

Design and synthesis of novel anti-inflammatory lipid mediators and anticancer small molecules

PDF

Design, synthesis, and biological evaluation of novel therapeutics for cancer

PDF

Design and synthesis of novel lipid mediators

PDF

Palladium mediated methods for the total synthesis of lipid mediators and organoboron compounds

PDF

Adventures in medicinal chemistry: design and synthesis of small molecule biological modulators

PDF

Design and synthesis of novel heterocycles and peptidomimetics from organoboronic acids, amines and carbonyl compounds

PDF

New organoboron based multicomponent methodologies for the synthesis of novel heterocycles

PDF

Multicomponent synthesis of optically pure aminodicarboxylic acids in water and total synthesis of 15-EPI-benzo-lipoxin A4 and aspirin-triggered neuroproctectin D1/protectin D1

PDF

Design, optimization, and synthesis of novel therapeutics

PDF

Stereo controlled synthesis of substituted cyclic ethers

PDF

Synthesis of multifunctional heterocycles, amino phosphontes using boronic acids

PDF

Progess towards the total synthesis of the antifungal natural product (-)-pramanicin

PDF

Synthesis of protein-protein interaction inhibitors and development of new catalytic methods

PDF

Design and synthesis of novel eicosanoids

PDF

Synthesis of novel nucleotide analogues based on the traditional and nontraditional bioisosteres

PDF

Multicomponent reactions of allenyl and alkynyl boron derivatives with amines and aldehydes and their use in the synthesis of novel multifunctional amines and heterocycles

Asset Metadata

Creator Winkler, Jeremy W. (author)

Core Title The total synthesis of novel lipid mediators and their role in inflammation

School College of Letters, Arts and Sciences

Degree Doctor of Philosophy

Degree Program Chemistry

Publication Date 12/04/2014

Defense Date 12/04/2013

Publisher University of Southern California (original), University of Southern California. Libraries (digital)

Tag chemistry,inflammation,lipid,neuroprotectin,OAI-PMH Harvest,omega-3,organic chemistry,resolvin

Format application/pdf (imt)

Language English

Contributor Electronically uploaded by the author (provenance)

Advisor Petasis, Nicos A. (committee chair), Louie, Stan G. (committee member), Prakash, G. K. Surya (committee member)

Creator Email winkler.jeremy@gmail.com

Permanent Link (DOI) https://doi.org/10.25549/usctheses-c3-356236

Unique identifier UC11297510

Identifier etd-WinklerJer-2203.pdf (filename),usctheses-c3-356236 (legacy record id)

Legacy Identifier etd-WinklerJer-2203.pdf

Dmrecord 356236

Document Type Dissertation

Format application/pdf (imt)

Rights Winkler, Jeremy W.

Type texts

Source University of Southern California (contributing entity), University of Southern California Dissertations and Theses (collection)

Access Conditions The author retains rights to his/her dissertation, thesis or other graduate work according to U.S. copyright law. Electronic access is being provided by the USC Libraries in agreement with the a...

Repository Name University of Southern California Digital Library

Repository Location USC Digital Library, University of Southern California, University Park Campus MC 2810, 3434 South Grand Avenue, 2nd Floor, Los Angeles, California 90089-2810, USA