Synthesis of enantiopure structured triacylglycerols

Article abstract of DOI:10.1016/j.tetasy.2013.11.015

The synthesis of nine enantiopure structured triacylglycerols (TAGs) of the AAB type is described, all possessing the (S)-absolute configuration. Six of them possess one saturated and two identical unsaturated fatty acyl chains, whereas the remaining three possess two identical saturated fatty acids along with one unsaturated fatty acid. The former group was synthesized by a five-step chemoenzymatic route involving a highly regioselective immobilized Candida antarctica lipase, starting from enantiopure (R)-solketal. The second group was prepared by a fully chemical five-step approach based on the same chiral precursor. Such enantiopure TAGs are strongly required as standards for the enantiospecific analysis of intact TAGs in fats and oils.

Full text of DOI:10.1016/j.tetasy.2013.11.015

Tetrahedron: Asymmetry 25 (2014) 125–132
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Synthesis of enantiopure structured triacylglycerols
Björn Kristinsson a, Kaisa M. Linderborg b, Heikki Kallio b, Gudmundur G. Haraldsson a,
⇑
a Science Institute, University of Iceland, Dunhaga 3, 107 Reykjavik, Iceland
b Food Chemistry and Food Development, Department of Biochemistry, University of Turku, FI-20014 Turku, Finland
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 3 October 2013
Accepted 21 November 2013
The synthesis of nine enantiopure structured triacylglycerols (TAGs) of the AAB type is described, all
possessing the (S)-absolute configuration. Six of them possess one saturated and two identical unsatu-
rated fatty acyl chains, whereas the remaining three possess two identical saturated fatty acids along
with one unsaturated fatty acid. The former group was synthesized by a five-step chemoenzymatic route
involving a highly regioselective immobilized Candida antarctica lipase, starting from enantiopure (R)-
solketal. The second group was prepared by a fully chemical five-step approach based on the same chiral
precursor. Such enantiopure TAGs are strongly required as standards for the enantiospecific analysis of
intact TAGs in fats and oils.
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
refers to the sn-1 position and the pro-R group refers to the sn-3
position with the remaining stereogenic carbon at the mid-posi-
Triacylglycerols (TAGs) are a major class of nonpolar lipids that
are ubiquitous in the fats and oils of plants and animals.1–4 They
constitute a glycerol backbone to which three fatty acyl groups
are linked through carboxylic ester bonds. The variety of TAG
molecular species is enormous in fats and oils. This relates to the
high number of various fatty acids. The fatty acids differ in the
length of the fatty acyl chain, which is commonly linear and pos-
sesses an even number of carbons from C4 to C22. However,
branched and odd-numbered fatty acids do exist in the fats and oils
of mainly microbial origin. The acyl chains can be saturated or
unsaturated with the number of double bonds ranging from 0 to
6. The position of the double bonds may also vary within the
hydrocarbon chain although they most commonly belong to the
n-9, n-7, n-6 and n-3 fatty acid classes with the cis-configuration
largely dominating, although the trans-configuration is known. In
polyunsaturated fatty acids, the methylene interrupted skipped
double-bond framework dominates although the double bonds
may also be conjugated in the fatty acids and derived lipids.
The location of the fatty acids within the glycerol skeleton adds
further to the great variety in terms of regioisomerism. Moreover,
the glycerol molecule is prochiral, which means that when two dif-
ferent fatty acids are attached to the terminal primary alcohol
groups of the glycerol backbone, a stereogenic centre is generated
at the secondary alcohol mid-position of the glycerol skeleton. A
stereospecific numbering designated by the prefix sn- is used to
distinguish between the two enantiotopic terminal positions of
the glycerol moiety, such that the pro-S hydroxymethyl group
tion referred to as the sn-2 position.1
TAGs are important constituents of the human and animal diet.
They are not only a source of energy but also provide essential fatty
acids for various biological roles.5–7 All the aforementioned varie-
ties largely influence the physical, sensory, nutritional and biolog-
ical properties of the TAGs. The situation is to an extent simplified
by the fact that the fatty acids are not randomly positioned in the
TAGs that are known to differ significantly in animals and plants
from species to species. Classical examples of the non-random
distribution of fatty acids into the stereospecific positions of TAGs
include cocoa butter8 used in chocolate manufacturing and human
milk TAGs9 while there are numerous reports on the stereospecific
positioning of fatty acids in animals and plants.10 On the other
hand this should underline the urgent need for firm analytical
methods to analyse the various regio- and enantiomeric TAGs in
order to better understand their physicochemical properties,
biological roles and the impact of the TAG structures on metabo-
lism, fatty acid bioavailability, digestion, absorption efficiency,
transport and human health.11–13
It is relatively straightforward to determine the total fatty acid
composition of TAGs by GC after the release of their fatty acids and
converting them into volatile methyl esters.14 All information is,
however, lost with regard to their positional distribution within
the glycerol backbone. Atmospheric pressure chemical ionization
mass spectrometry (APCI MS) provides information on the overall
fatty acid composition of the 2-position as well as the 1,
3-positions, but this method does not distinguish between the enan-
tiomeric sn-1 and sn-3 positions.15,16 That type of information is
available by enantiospecific analysis where multi-step procedures
involving 1,3-regioselective lipase17 or Grignard reagents18 to form
⇑
Corresponding author. Tel.: +354 525 4818; fax: +354 552 8911.
E-mail address: gghar@hi.is (G.G. Haraldsson).
0957-4166/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetasy.2013.11.015

126
B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
a mixture of diacylglycerols (DAGs) and monoacylglycerols (MAGs)
undergoing further transformations involving enantioselective
phospholipase A,10,19 chiral derivatization20 or chiral-phase
HPLC21,22 are combined with chromatography. Several drawbacks
related to these processes are due to the tedious multi-step
procedures and acyl-migration23,24 side reactions where acyl groups
migratefrom one position to another within the glycerol framework,
thus affecting the reliability of the stereospecific analysis.
The stereospecific analyses described above provide an insight
into the overall fatty acid composition of the individual stereospe-
cific sn-1, -2 and -3 positions for a large number of TAGs in a
mixture rather than information on individual TAG enantiomers.
The stereospecific analysis of intact TAGs in a natural oil mixture
is far more of a challenge. Non-aqueous reversed phase and
silver-ion HPLC/MS are the most commonly used analytical tech-
niques for the separation and characterization of natural TAG
molecular species,15,25 but these methods cannot be used to sepa-
rate TAG enantiomers. Accordingly, there is an urgent need for a
non-destructive enantiospecific analysis of intact TAGs. This
important challenge has been addressed in a few recent reports
based on chiral HPLC in attempts to separate the TAG enantio-
positions of the TAG glycerol skeleton. Each of the TAGs possesses
two different types of fatty acids, one being saturated and the other
unsaturated, thus allowing the TAGs to be detected by a UV detec-
tor in combination with chiral HPLC.
From a synthetic organic chemistry point of view the TAGs may
be divided into two categories: (a) those possessing one saturated
fatty acyl group located at the sn-3 position and two unsaturated
fatty acyl groups at the remaining sn-1 and sn-2 positions of the
glycerol backbone; (b) those possessing two saturated acyl groups
at the sn-2 and sn-3 positions and one unsaturated acyl group at
the sn-1 position. Of the TAGs synthesized herein, the saturated
fatty acids belonging to the first category included decanoic
(C10:0), hexadecanoic (C16:0), eicosanoic (C20:0) and docosanoic
(C22:0) acids, whereas in the second category, the saturated fatty
acids were limited to hexadecanoic and octadecanoic (C18:0) acids.
The unsaturated fatty acids present in the first category included
the mono-unsaturated cis-hexadec-9-enoic (C16:1) and cis-octa-
dec-9-enoic (C18:1) acids and the di-unsaturated cis-octadec-
9,12-dienoic (C18:2) acid with C18:1 and C18:2 belonging to the
second category. The structures of these TAGs (S)-1–9 are shown
in Figure 1 with compounds (S)-1, 2, 3, 4, 5 and 6 belonging to
O
O
O
O
O
O
C8H17
C8H17
C8H17
O
C6H13
C8H17
C8H17
C8H17
O
O
O
C6H13
O
1
O
O
2
O
O
O
O
O
O
C9H19
C15H31
O
O
O
O
C5H11
C5H11
O
3
4
O
O
O
O
C15H31
O
O
C15H31
O
O
C8H17
C8H17
O
6
5
7
O
O
O
O
C21H43
C19H39
O
O
O
C15H31
O
C17H35
O
C15H31
C8H17
O
C8H17
O
C5H11
O
O
O
9
O
O
8
O
O
O
O
O
C15H31
C17H35
C15H31
Figure 1. The enantiopure TAG products (S)-1–9.
mers.26–28 A remaining obstacle here is the lack of appropriate
TAG enantiomers as standards to distinguish between the intact
TAG enantiomers in such analyses.27,28
the first category and (S)-7, 8 and 9 belonging to the second one.
The synthetic route to the TAGs (S)-1–6 belonging to the first
category is illustrated in Scheme 1 for (S)-2. It was based on a
five-step chemoenzymatic process starting from enantiomerically
pure 2,3-isopropylidene-sn-glycerol, (R)-solketal, as a chiral pre-
cursor. In the first four steps, a previously reported approach
towards synthesis of 1-octadecanoyl-sn-glycerol obtained from
(S)-solketal was followed.32 1-Octadecanoyl-sn-glycerol is an
antipodal homologue of the required 3-acyl-sn-glycerols. Herein
enantiomerically pure (R)-solketal was benzylated at the sn-3
position and the resulting benzylated adduct was subsequently
deprotected without purification to remove the isopropylidene
moiety. The enantiomerically pure key intermediate 1-O-benzyl-
sn-glycerol (S)-10 was obtained in 66% overall yield after
purification.
Herein we address that impediment by providing nine enantio-
pure (S)-TAG enantiomers possessing fatty acids commonly pres-
ent in plant oils and fats by chemical and chemoenzymatic
synthesis starting from enantiomerically pure (R)-solketal. Their
synthesis was based on previous reports of the chemoenzymatic
synthesis of regiopure symmetrically structured ABA type
TAGs,29,30 enantiopure similarly structured diacylglycerol ethers
(DAGE)31 and enantiopure ABC type TAGs32 by using immobilized
Candida antarctica lipase (CAL-B), which showed excellent regiose-
lectivity towards the terminal positions of the glycerol moiety.
2. Results and discussion
The benzylated glycerol adduct (S)-10 was acylated with
activated esters (C10:0, C16:0, C20:0 and C22:0) by using highly
regioselective immobilized Candida antarctica lipase (CAL-B from
Novozymes) at rt in dichloromethane. For C10:0 and C16:0,
The enantiopure AAB type structured TAGs addressed herein all
have an (S)-configuration with the term structured TAG29,30
referring to selected fatty acids accommodating predetermined

B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
127
O
Ph
O
Ph
O
OH
Ph
O
c
d
a
b
OH
OH
O
O
OH
O
O
O
(S)-10
C15H31
O
O
C6H13
C6H13
O
OH
e
O
OH
O
O
O
C15H31
O
C15H31
2
(S)-
12
(R)-
Scheme 1. Chemoenzymatic synthesis of the first category TAGs 1–6 [shown for (S)-2]. Reagents and conditions: (a) NaH, THF, then BnBr; (b) 1 M HCl, H2O/EtOH, reflux
30 min; (c) vinyl hexanoate, CAL, CH2Cl2, rt; (d) H2, 10% Pd/C, THF/hexane; (e) cis-hexadec-9-enoic acid, EDAC, DMAP, CH2Cl2, rt.
commercially available vinyl esters were used and the enzymatic
reaction took only 90 min for completion. For C20:0 and C22:0,
activation was needed and it was decided to activate these acids
as acetoxime esters by using acetone oxime and EDAC coupling
agent in the presence of DMAP by a method recently described
for activation of the n-3 polyunsaturated fatty acids EPA and
DHA.33 The acetoxime esters were obtained in 99% and 64% yields,
respectively, for C20:0 and C22:0. In all cases, the acylation
exclusively took place at the sn-3 position as had been anticipated.
Figure 2 illustrates the structure of the vinyl esters (shown for
the structures were firmly established by 1H NMR spectroscopy
at 400 MHz.32
The unsaturated fatty acids C16:1, C18:1 and C18:2 were intro-
duced onto the free hydroxyl groups at the sn-1 and sn-2 positions
of (R)-11–14 using 1-(3-dimethylaminopropyl)-3-ethylcarbodiim-
ide hydrochloride (EDAC) as a chemical coupling agent in the pres-
ence of dimethylaminopyridine (DMAP) following a previously
described procedure.29,30 The resulting first category products
(S)-1–6 were obtained in good to excellent yields (70–99%) after
purification by flash chromatography. No acyl-migration was
observed to take place during this reaction, as had been firmly
established in previous studies.29,30,32 Table 1 shows the type of
products and the yields obtained along with their specific rotation
values. As can be seen, these values are extremely low (vide infra)
which could possibly be an indication of a loss in enantiopurity.
However, that possibility can by ruled out by the absence of acyl
migration and the establishment of the high enantiopurity of all
of the synthesized TAGs by reversed phase chiral-HPLC.
O
O
O
N
O
The second category products (S)-7–9 were synthesized by the
synthetic route shown in Scheme 2. No enzyme was required this
Figure 2. The structure of vinyl hexanoate (top) and eicosanoic acid acetoxime
ester (bottom).
O
C15H31
O
C15H31
vinyl hexanoate) and the oxime esters (shown for eicosanoic acid
acetoxime ester).
OH
Ph
O
Ph
O
a
b
O
O
OH
OH
O
O
O
O
The acylated benzyl ether adducts were not purified but directly
subjected to deprotection of the benzyl ether protecting group by
catalytic hydrogenolysis using Pd/C catalyst in THF-hexane. The
resulting MAGs (R)-11–14 were obtained in moderate to good
overall yields (38–75% in two steps) after crystallization. Table 1
C15H31
C15H31
(R)-15
10
(S)-
O
O
C15H31
C5H11
O
c
O
O
O
Table 1
Yieldsa and specific activity of MAG and DAG intermediates (R)-11–16 obtained in
(S)-8
C15H31
accordance with Schemes 1 and 2
Scheme 2. Chemical synthesis of the second category TAGs 7–9 [shown for (S)-7].
Reagents and conditions: (a) hexanoic acid, EDAC, DMAP, CH2Cl2, rt; (b) H2, 10% Pd/
C, THF/hexane; (c) cis-octadec-9,12-dienoic acid, EDAC, DMAP, CH2Cl2, rt.
½ ꢀ
a 2D0
Entry
sn-1
sn-2
sn-3
Yield (%)
(R)-11
(R)-12
(R)-13
(R)-14
(R)-15
(R)-16
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
C16:0
C18:0
C10:0
C16:0
C20:0
C22:0
C16:0
C18:0
75
56
58
38
74
69
ꢁ3.46
ꢁ2.58
ꢁ1.89
ꢁ1.61
ꢁ4.23
ꢁ3.87
time since the role of the lipase in the first category TAG synthesis
was to secure regiocontrol by introducing a saturated fatty acyl
group exclusively at the sn-3 position of the glycerol skeleton
leaving the sn-2 position free for an unsaturated fatty acid. This
is a three-step chemical route based on the benzylated glycerol ad-
duct (S)-10 as a common precursor, shared with the synthesis of
the first category TAG products. In the first step, two identical
saturated fatty acids, C16:0 or C18:0, were introduced to the free
hydroxyl groups at the sn-2 and sn-3 positions of the glycerol moi-
ety using the EDAC coupling agent in the presence of DMAP in
dichloromethane at rt by the same procedure as described above.
This resulted in formation of the corresponding 1-O-benzyl-2,3-
diacyl-sn-glycerols that were, without purification, subjected to
a
Overall yields for two steps from (S)-10 in accordance with Schemes 1 and 2.
shows the yields obtained for these intermediate adducts along
with their specific optical rotation values. The lower yields for
the long-chain C20:0 (R)-13 and particularly C22:0 (R)-14 adducts
were attributed to the hydrogenolysis step that required the
addition of a drop of perchloric acid34,35 to speed-up the reaction
that would otherwise not have taken place at all. These intermedi-
ates were all obtained in excellent chemical and regiopurity, and

128
B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
catalytic hydrogenolysis deprotection of the benzyl protecting
group. This resulted in formation of the DAG intermediates
(R)-15 (C16:0) and (R)-16 (C18:0), which were obtained in high
overall yields (74% and 69%, respectively) after recrystallization
(see Table 1). Their excellent regio- and enantiopurity was estab-
lished by the fact that no acyl-migration was observed to take
place during these reactions.
intermediates that essentially need to be saturated. TAGs possess-
ing one saturated and two identical unsaturated fatty acids or two
identical saturated and one unsaturated fatty acids of the type
addressed herein as well as TAGs possessing two different types
of saturated fatty acids belong to this ABB type structured TAG cat-
egory that are readily accessible by the current synthetic approach.
The 2,3-sn-DAG intermediates were then acylated with the
unsaturated fatty acids using the same chemical coupling method
as before, (R)-15 with 18:1 and 18:2 to obtain the corresponding
TAGs (S)-7 and (S)-8, respectively, and (R)-16 with 18:1 to obtain
(S)-9. These products were obtained in high to excellent yields
after recrystallization, as can be seen in Table 2. The specific
3. Conclusion
One of the main obstacles in the enantiospecific analysis of
intact TAGs in fats and oils of plant and animal origin is the lack
of enantiopure structured TAG enantiomers as standards for such
analysis. That impediment has been addressed to an extent here
where we have described the synthesis of nine enantiopure AAB
type structured (S)-TAGs. Six of them possess one saturated and
two identical unsaturated fatty acyl chains and were synthesized
Table 2
Yieldsa and specific activity of TAG products (S)-1–9 obtained in accordance with
by
a five-step chemoenzymatic route involving immobilized
Schemes 1 and 2
Candida antarctica lipase, and starting from enantiopure (R)-solk-
etal. The three remaining TAGs possess two identical unsaturated
fatty acids along with one saturated fatty acid and were prepared
by a similar, but fully chemical, five-step approach based on the
same chiral precursor. This highly efficient synthetic strategy
should be useful in the synthesis of a variety of related enantiopure
AAB type structured TAGs to be applied as standards in the stereo-
specific analysis of intact TAGs in fats and oils.
½ ꢀ
a 2D0
Entry
sn-1
sn-2
sn-3
Yield (%)
(S)-1
(S)-2
(S)-3
(S)-4
(S)-5
(S)-6
(S)-7
(S)-8
(S)-9
C18:1
C16:1
C18:1
C18:2
C18:1
C18:1
C18:1
C18:2
C18:1
C18:1
C16:1
C18:1
C18:2
C18:1
C18:1
C16:0
C16:0
C18:0
C10:0
C16:0
C16:0
C16:0
C20:0
C22:0
C16:0
C16:0
C18:0
95
73
70
99
70
78
86
98
79
ꢁ0.111
ꢁ0.055
ꢁ0.041
ꢁ0.045
ꢁ0.028
ꢁ0.027
ꢁ0.035
ꢁ0.053
ꢁ0.044
4. Experimental
4.1. General
a
The yields are based on the acylation in the final step in Schemes 1 and 2.
1H and 13C nuclear magnetic resonance spectra were recorded
on a Bruker Avance 400 spectrometer in deuterated chloroform
as a solvent at 400.12 and 100.61 MHz, respectively. Chemical
shifts (d) are quoted in parts per million (ppm) and the coupling
constants (J) in Hertz (Hz). The following abbreviations are used
to describe the multiplicity: s, singlet; d, doublet; t, triplet; dd,
doublet of doublets; m, multiplet. The number of carbon nuclei
behind each 13C signal is indicated in parentheses after each chem-
ical shift value, when there is more than one carbon responsible for
the peak. Infrared spectra were conducted on a Thermo Nicolet FT-
IR iS10 Spectrophotometer on a KBr pellet (crystalline material) or
as a neat liquid (oils). Melting points were determined on a Büchi
520 melting point apparatus and are uncorrected. The high-resolu-
tion mass spectra (HRMS) were acquired on a Bruker micrOTOF-Q
mass spectrometer. All data analyses were done on Bruker
software. Optical activity measurements were performed on an
AutopolR V Automatic Polarimeter from Rudolph Research Analyt-
ical using a 40T-2.5-100-0.7 Temp Trol™ polarimetric cell with
2.5 mm inside diameter, 100 mm optical path length and 0.7 ml
volume, with c referring to g sample/100 ml.
rotation values for these enantiopure (as established by reversed
phase chiral-HPLC) TAGs remained low (see Table 2).
The specific rotation values are extremely low and warrant a
special comment. Enantiopure TAGs are known for their extremely
low specific rotation values as was reported by Baer and Fischer in
the late 1930s.36,37 Such compounds are known for their cryptochi-
rality38 or cryptoactivity,39,40 terms which refer to their specific
rotation remaining close to zero, and which are hardly measure-
able. As can be seen from the results in Table 2, the specific rotation
values for all of the enantiopure TAGs ranged from ꢁ0.027 to
ꢁ0.055 except for (S)-1, which possessed the shortest saturated
C10:0 chain, where the absolute value was higher although still
low at ꢁ0.111. The specific rotation values for the intermediates
(R)-MAGs and (R)-DAGs displayed in Table 1 are significantly high-
er, or roughly two orders of magnitude higher. The trend towards
lower activity values when the acyl chain gets shorter is
noteworthy.
All intermediate adducts including the sn-3-MAGs and
sn-2,3-DAGs and the final TAG products were obtained in high
chemical purity and were all fully characterized by synthetic
organic chemistry methods including high-resolution 1H
(400 MHz) and 13C NMR and IR spectroscopy methods as well as
satisfactory high-resolution accurate mass spectrometry analyses.
The specific rotation was determined for all of the chiral com-
pounds involved. Full regiocontrol and therefore enantiocontrol
in all of the reactions was firmly established as based on detailed
studies by high-resolution 1H and 13C NMR spectroscopy in accor-
dance with previous reports29,30,32 which was fully confirmed by
chiral HPLC measurements.
The immobilized Candida antarctica lipase (Novozym 435; CAL-
B) was supplied as
a gift from Novozymes A/S (Bagsvaerd,
Denmark). All chemicals and solvents were used without further
purification unless otherwise stated. (R)-(ꢁ)-2,2-Dimethyl-1,
3-dioxolane-4-methanol [(R)-solketal, 98% purity and 99% ee]
was purchased from Sigma–Aldrich (Steinheim, Germany). Vinyl
capriate (>99%) and vinyl palmitate (>96%) were purchased from
TCI Europe (Zwindrecht, Belgium. Eicosanoic (arachidic), docosa-
noic (behenic), octadecanoic (stearic), hexadecanoic (palmitic),
cis-hexadec-9-enoic (palmitoleic), cis-octadec-9-enoic (oleic) and
cis-octadec-9,12-dienoic (linoleic) acids were all obtained in high
(>99%) purity from Larodan Fine Chemicals (Malmö, Sweden).
Anhydrous magnesium sulfate was obtained from Merck (Darms-
tadt, Germany), benzyl bromide (98%) and sodium hydride (60%
dispersion in mineral oil) from Sigma-Aldrich (Steinheim) and
The synthetic strategy used is suitable for preparing enantio-
pure TAGs of the AAB type possessing two different fatty acids with
the prerequisite that at least one of them is saturated. This relates
to the use of catalytic hydrogenolysis to remove the benzyl pro-
tecting group from the mono- and diacylated 1-O-benzylglycerol

B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
129
10% palladium on carbon catalyst from Merck (Munich, Germany).
4.4. Eicosanoic acid acetoxime ester
Dichloromethane and ethyl acetate were obtained HPLC grade
from Sigma–Aldrich (Steinheim, Germany), n-hexane as p.a. from
Merck (Darmstadt, Germany), pet. ether, boiling range 40–60 °C,
and diethyl ether as p.a. from Riedel-de Haën (Seelze, Germany).
Dichloromethane was dried over calcium hydride under a dry
nitrogen atmosphere. Tetrahydrofuran for analysis was obtained
from Acros Organics (Geel, Belgium) and dried over Na wire in
the presence of benzophenone under a dry nitrogen atmosphere.
Silica gel (Silica gel 60) and analytical TLC plates (DC Alufolien Kie-
selgel 60 F254) were obtained from Merck (Darmstadt, Germany).
EDAC was obtained of commercial grade from Sigma–Aldrich
(Steinheim, Germany) and DMAP (99%) and acetone oxime (98%)
from Acros Organics (Geel, Belgium).
To a solution of eicosanoic acid (710 mg, 2.27 mmol), DMAP
(20 mg, 0.16 mmol) and EDAC (463 mg, 2.42 mmol) in dried
dichloromethane (10 ml), acetone oxime (193 mg, 2.64 mmol)
was added and the resulting solution was stirred overnight at rt.
The reaction solution was passed through a short silica gel column
with ethyl acetate/petroleum ether (60:40) as an eluent, affording
the product as white crystalline material after evaporation of the
solvents and recrystallization from petroleum ether (826 mg,
2.25 mmol) in 99% yield. Mp = 60.6–62.0 °C. 1H NMR d 2.39 (t,
J = 7.6 Hz, 2H, CH2COO), 2.04 (s, 3H, N@C(CH3)2), 2.00 (s, 3H,
N@C(CH3)2), 1.68 (quin, J = 7.6 Hz, 2H, CH2CH2COO), 1.38–1.18
(m, 32H, CH2), and 0.87 (t, J = 6.8 Hz, 3H, –CH3) ppm. 13C d 171.2
(C@O), 163.6 (C@N), 33.0, 31.9, 29.7 (10), 29.6, 29.3, 29.2, 29.1,
25.0, 22.7, 22.0, 16.9 and 14.1 ppm. IR mmax 2917 (vs, C–H), 2848
4.2. Determination of the enantiopurity by HPLC
(vs, C–H) and 1757 (vs, C@O) cmꢁ1
23H45NO2Na+ m/z 390.3343; found 390.3358 amu.
. HRMS (ESI): calcd for
The retained enantiopurity (99% ee) of all the synthetic TAG
C
enantiomers (S)-1–9 was confirmed on reversed phase chiral HPLC
by using two identical CHIRALCEL OD-RH (150 ꢂ 4.6 mm, 5
lm)
4.5. Docosanoic acid acetoxime ester
columns obtained from Chiral Technologies Europe (Illkirch,
France), which were connected in a series. The nine corresponding
racemic TAGs 1–9 purchased from Larodan Fine Chemicals
(Malmö, Sweden) were used as standards and recycled until
adequate separation was obtained as detected by UV, where the
(S)-enantiomers were observed to elute faster than the (R)-enanti-
omers. The elution order of the enantiomers was determined by
the co-injection of enantiopure and racemic TAGs. All details of
the relevant methodology that benefited from the current TAGs
(S)-1–9 being used as enantiopure standards will be published
separately.41
A procedure identical to that described above for eicosanoic
acid acetoxime ester was followed using docosanoic acid
(564 mg, 1.66 mmol), DMAP (20 mg, 0.16 mmol), EDAC (400 mg,
2.09 mmol), dichloromethane (10 ml) and acetoxime (186 mg,
2.54 mmol). The product was afforded as white crystals after
recrystallization from petroleum ether (420 mg, 1.06 mmol) in
64% yield. Mp = 69.0–70.0 °C. 1H NMR d 2.41 (t, J = 7.6 Hz, 2H,
CH2COO), 2.06 (s, 3H, N@C(CH3)2), 2.00 (s, 3H, N@C(CH3)2), 1.70
(quin, J = 7.6 Hz, 2H, CH2CH2COO), 1.38–1.18 (m, 36H, CH2), and
0.89 (t, J = 6.8 Hz, 3H, –CH3) ppm. 13C d 171.3 (C@O), 163.6
(C@N), 33.1, 32.0, 29.7 (11), 29.6, 29.5, 29.4, 29.3, 29.2, 25.0,
22.0, 22.1, 17.0 and 14.2 ppm. IR mmax 2923 (vs, C–H), 2849 (vs,
C–H) and 1758 (vs, C@O) cmꢁ1. HRMS (ESI): calcd for C25H49NO2
Na+ m/z 418.3656; found 418.3670 amu.
4.3. 1-O-Benzyl-sn-glycerol (S)-10
Prior to use, all glassware was flame-dried and allowed to cool
under dry nitrogen atmosphere. Next, NaH (60% mineral oil disper-
sion; 1.00 g, 25.0 mmol) was added to a three-necked r.b. flask and
treated several times with dry pet. ether using a syringe to remove
the mineral oil prior to the addition of freshly dried THF (20 ml).
(R)-Solketal (2.20 g, 16.7 mmol) dissolved in THF was added via
syringe at a rate appropriate to keep the heat formation under con-
trol. The resulting suspension was stirred for a few minutes before
benzyl bromide (3.00 g, 17.5 mmol) dissolved in dry THF was
added via syringe. The mixture was refluxed for 4 h after which
all of the solketal had been consumed. The reaction mixture was
allowed to cool to rt and 1 M HCl (30 ml) was added together
with ethanol sufficient to make the solution homogeneous. The
resulting mixture was refluxed for 30 min or until all of 1-O-ben-
zyl-2,3-ispropylidene-sn-glycerol intermediate had reacted. After
cooling to rt the reaction mixture was extracted with diethyl ether
and the organic phase was treated several times with 14% NaHCO3
until pH >7 and finally dried over MgSO4. The solvent was removed
in vacuo on a rotary evaporator and the residue was introduced to
a kugelrohr distillation at 2.8 Torr. A first fraction was collected at
50 °C and discarded. The desired product (S)-10 was collected as a
colourless liquid (2.00 g, 11.1 mmol) of high purity as indicated by
4.6. 3-Decanoyl-sn-glycerol (R)-11
To
a solution of 1-O-benzyl-sn-glycerol (S)-10 (305 mg,
1.67 mmol) and vinyl decanoate (408 mg, 2.06 mmol) in dichloro-
methane (3 ml) was added immobilized CAL (30 mg). The resulting
suspension was stirred at rt for approx. 90 min when TLC monitor-
ing (1:1 ethyl acetate/pet. ether) indicated a complete reaction. The
lipase preparation was separated by filtration and the solvent was
removed in vacuo on a rotary evaporator. The resulting residue was
dissolved in THF (30 ml) without further purification, followed by
the addition of n-hexane (70 ml) and Pd/C catalyst (15 mg). The
reaction was performed in a PARR reactor under hydrogen pressure
(5 bar) during which time the product precipitated. When the reac-
tion had proceeded to completion (approximately 2 h), THF was
added until all of the product had been dissolved. The catalyst
was separated off by filtration with the aid of Celite and the solvent
was removed in vacuo on a rotary evaporator. The residue was
redissolved in the minimum amount of THF and a four-fold volume
of n-hexane was added. The resulting mixture was allowed to
stand at rt overnight to afford product (R)-11 as white crystals
(307 mg, 1.25 mmol) in 75% overall yields. Mp = 43.5–44.7 °C.
TLC (1:1 ethyl acetate/pet. ether) in 66% overall yield. ½a D20
¼ ꢁ5:48
ꢀ
(c 20, chloroform); ꢁ2.42 (c 5.0, THF). 1H NMR d 7.38–7.30 (m, 5H,
Ph–H), 4.55 (s, 2H, Ph–CH2), 3.92–3.87 (m, 1H, –CH–OH), 3.70 (dd,
J = 11.6 Hz, J = 4.0 Hz, 1H, –CH2–OH), 3.62 (dd, J = 11.6 Hz,
J = 5.6 Hz, 1H, –CH2–OH), 3.58 (dd, J = 9.6 Hz, J = 4.4 Hz, 1H,
–CH2–OCH2Ph), 3.53 (dd, J = 9.6 Hz, J = 6.4 Hz, 1H, –CH2–OCH2Ph),
2.81 (br s, 1H, –OH), 2.40 (br s, 1H, OH) ppm. 13C d 137.6, 128.5
(2), 127.9, 127.8 (2), 73.5, 71.7, 70.7 and 64.0 ppm. IR mmax 3150-
3600 (br, O-H), 2926 (vs, C–H) and 2868 (vs, C–H) cmꢁ1. HRMS
(ESI): calcd for C10H14O3Na+ m/z 205.0835; found 205.0841 amu.
½
a 2D0
ꢀ
¼ ꢁ3:46 (c 5.1, THF). 1H NMR
d 4.22 (dd, J = 11.6 Hz,
J = 4.8 Hz, 1H, –CH2–OCO), 4.16 (dd, J = 11.6 Hz, J = 6.0 Hz, 1H,
–CH2–OCO), 3.96–3.90 (m, 1H, –CH–OH), 3.72–3.67 (m, 1H,
–CH2–OH), 3.63–3.57 (m, 1H, –CH2–OH), 2.52 (br d, 1H, –CH–OH),
2.36 (t, J = 7.6 Hz, 2H, CH2COO), 2.08 (br s, 1H, –CH2–OH), 1.64
(quin, J = 7.2 Hz, 2H, CH2CH2COO), 1.37–1.18 (m, 12H, CH2), and
0.88 (t, J = 6.8 Hz, 3H, –CH3) ppm. 13C d 174.4, 70.3, 65.1, 63.3,
34.1, 31.8, 29.4, 29.2 (2), 29.1, 24.9, 22.6 and 14.1 ppm. IR mmax
3150-3600 (br, O–H), 2918 (vs, C–H), 2850 (vs, C–H) and 1733

130
B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
(vs, C@O) cmꢁ1. HRMS (ESI): calcd for C13H26O4Na+ m/z 269.1732;
found 269.1719 amu.
4.10. 2,3-Dihexadecanoyl-sn-glycerol (R)-15
To solution of 1-O-benzyl-sn-glycerol (S)-10 (300 mg,
a
4.7. 3-Hexadecanoyl-sn-glycerol (R)-12
1.65 mmol) and hexanoic acid (928 mg, 3.62 mmol) in dried
dichloromethane (4.0 ml) were added DMAP (95 mg, 0.78 mmol)
and EDAC (695 mg, 3.63 mmol). The resulting solution was stirred
magnetically at rt until TLC monitoring (80:20:1 pet. ether/ether/
acetic acid) indicated complete reaction. The reaction was stopped
by passing the reaction mixture through a short column packed
with silica gel using petroleum ether as an eluent. The solvent
was removed in vacuo and the resulting residue without further
purification was dissolved in n-hexane (50 ml) and transferred to
a hydrogenation flask followed by the addition of Pd/C catalyst
(25 mg). The reaction was performed at rt in a PARR reactor under
hydrogen pressure (5 bar). When the reaction was complete (2 h),
the catalyst was separated off by filtration with the aid of Celite
and the solvent was removed in vacuo on a rotary evaporator.
Recrystallization from n-hexane or petroleum ether afforded the
product (R)-15 as white crystals (695 mg, 1.22 mmol) in 74% over-
The same procedure was followed as described for (R)-11 using
1-O-benzyl-sn-glycerol (S)-10 (505 mg, 2.77 mmol), vinyl hexade-
canoate (1028 mg, 3.64 mmol), dichloromethane (5 ml), immobi-
lized CAL (60 mg) and Pd/C catalyst (30 mg). The product (R)-12
(510 mg, 0.212 mmol) was afforded as white crystals in 56% overall
yield. Mp = 70.8–72.0 °C. ½a D20
ꢀ
¼ ꢁ2:58 (c 5.2, THF). 1H NMR d 4.21
(dd, J = 11.6 Hz, J = 4.4 Hz, 1H, –CH2–OCO), 4.15 (dd, J = 11.6 Hz,
J = 6.0 Hz, 1H, –CH2–OCO), 3.96–3.90 (m, 1H, –CH–OH), 3.70 (dd,
J = 11.6 Hz, J = 4.0 Hz, 1H, –CH2–OH), 3.60 (dd, J = 11.6 Hz,
J = 6.0 Hz, 1H, –CH2–OH), 2.54 (br s, 1H, –CH–OH), 2.35
(t, J = 7.6 Hz, 2H, CH2COO), 2.10 (br s, 1H, –CH2–OH), 1.63 (quin,
J = 7.2 Hz, 2H, CH2CH2COO), 1.37–1.18 (m, 24H, CH2), and 0.88 (t,
J = 6.8 Hz, 3H, –CH3) ppm. 13C d 174.4, 70.3, 65.1, 63.3, 34.1, 31.9,
29.7 (3), 29.6 (3), 29.4, 29.3, 29.2, 29.1, 24.9, 22.7 and 14.1 ppm.
IR mmax 3150-3600 (br, O-H), 2919 (vs, C–H), 2849 (vs, C–H) and
1735 (vs, C@O) cmꢁ1. HRMS (ESI): calcd for C19H38O4Na+ m/z
353.2662; found 353.2672 amu.
all yield. Mp = 71.0–72.2 °C. ½a D20
ꢀ
¼ ꢁ4:23 (c 5.1, THF). 1H NMR d
5.11–5.06 (m, 1H, CH–O–CO), 4.32 (dd, J = 12.0 Hz, J = 4.8 Hz, 1H,
–CH2–OCO), 4.23 (dd, J = 12.0 Hz, J = 5.6 Hz, 1H, –CH2–OCO),
3.75–3.72 (m, 2H, –CH2–OH), 2.34 (t, J = 7.6 Hz, 2H, CH2COO),
2.33 (t, J = 7.6 Hz, 2H, CH2COO), 2.10 (t, J = 6.4 Hz, 1H, –CH2–OH),
1.64–1.58 (m, 4H, CH2CH2COO), 1.38–1.17 (m, 48H, CH2), and
4.8. 3-Eicosanoyl-sn-glycerol (R)-13
0.87 (t, J = 6.8 Hz, 6H, –CH3) ppm. 13C d 173.8 (
a C@O), 173.4 (b
The same procedure was followed as descibed for (R)-11 using
1-O-benzyl-sn-glycerol (S)-10 (250 mg, 1.37 mmol), eicosanoic
acid acetoxime ester (550 mg, 1.50 mmol), dichloromethane
(4 ml), immobilized CAL (35 mg) and Pd/C catalyst (15 mg). The
hydrogenolysis reaction required a drop of perchloric acid to be
added to the reaction mixture as a promoter.34,35 The product
(R)-13 (306 mg, 0.791 mmol) was afforded as white crystals in
C@O), 72.1, 62.0, 61.6, 34.3, 34.1, 31.9 (2), 29.7 (10), 29.6 (2),
29.5 (2), 29.4 (2), 29.3 (2), 29.1 (2), 24.9 (2), 22.7 (2) and 14.1
(2) ppm. IR mmax 3300–3600 (br, O–H), 2957 (vs, C–H), 2918 (vs,
C–H), 2850 (vs, C–H), 1732 (vs, C@O) and 1709 (vs, C@O) cmꢁ1
HRMS (ESI): calcd for
591.4962 amu.
.
C
35H68O5Na+ m/z 591.4959 found
58% overall yield. Mp = 78.2–79.7 °C. ½a D20
¼ ꢁ1:89 (c 5.2, THF).
ꢀ
4.11. 2,3-Dioctadecanoyl-sn-glycerol (R)-16
1H NMR d 4.21 (dd, J = 11.6 Hz, J = 4.8 Hz, 1H, OCO–CH2–), 4.15
(dd, J = 11.6 Hz, J = 6.0 Hz, 1H, OCO–CH2–), 3.96–3.90 (m, 1H,
–CH–OH), 3.70 (dd, J = 11.6 Hz, J = 4.0 Hz, 1H, –CH2–OH), 3.60 (dd,
J = 11.6 Hz, J = 6.0 Hz, 1H, –CH2–OH), 2.57 (br s, 1H, –CH–OH),
2.35 (t, J = 7.6 Hz, 2H, CH2COO), 2.14 (br s, 1H, –CH2–OH), 1.63
(quin, J = 7.2 Hz, 2H, CH2CH2COO), 1.37–1.18 (m, 32H, CH2), and
0.88 (t, J = 6.8 Hz, 3H, –CH3) ppm. 13C d 174.3, 70.3, 65.2, 63.3,
34.2, 31.9, 29.7 (10), 29.4 (2), 29.2, 29.1, 24.9, 22.7 and 14.1 ppm.
IR mmax 3150–3600 (br, O–H), 2919 (vs, C–H), 2849 (vs, C–H) and
1735 (vs, C@O) cmꢁ1. HRMS (ESI): calcd for C23H46O4Na+ m/z
409.3288; found 409.3299 amu.
The same procedure was followed as described for (R)-15 using
1-O-benzyl-sn-glycerol (S)-10 (150 mg, 0.823 mmol), octadecanoic
acid (490 mg, 1.72 mmol), dichloromethane (4 ml), DMAP (60 mg,
0.49 mmol), EDAC (347 mg, 1.81 mmol) and Pd/C (12 mg). The
product (R)-16 (354 mg, 0.566 mmol) was afforded as white crys-
tals in 69% overall yield. Mp = 76.3–77.0 °C. ½a D20
¼ ꢁ3:87 (c 5.1,
ꢀ
THF). 1H NMR
d 5.11–5.06 (m, 1H, CH–O–CO), 4.32 (dd,
J = 12.0 Hz, J = 4.4 Hz, 1H, –CH2–OCO), 4.24 (dd, J = 12.0 Hz,
J = 5.6 Hz, 1H, –CH2–OCO), 3.75–3.71 (m, 2H, –CH2–OH), 2.34 (t,
J = 7.6 Hz, 2H, CH2COO), 2.32 (t, J = 7.6 Hz, 2H, CH2COO), 2.06 (t,
J = 6.4 Hz, 1H, –CH2–OH), 1.66–1.57 (m, 4H, CH2CH2COO),
1.38–1.17 (m, 56H, CH2), and 0.87 (t, J = 6.8 Hz, 6H, –CH3) ppm.
4.9. 3-Docosanoyl-sn-glycerol (R)-14
13C d 173.8 (
a C@O), 173.4 (b C@O), 72.1, 62.0, 61.6, 34.3, 34.1,
31.9 (2), 29.7 (14), 29.6 (2), 29.5 (2), 29.4 (2), 29.3 (2), 29.1 (2),
24.9 (2), 22.7 (2) and 14.1 (2) ppm. IR mmax 3300–3600 (br, O-H),
2957 (vs, C–H), 2918 (vs, C–H), 2849 (vs, C–H), 1732 (vs, C@O)
and 1709 (vs, C@O) cmꢁ1. HRMS (ESI): calcd for C39H76O5Na+ m/z
647.5585; found 647.5576 amu.
The same procedure was followed as described for (R)-13 using
1-O-benzyl-sn-glycerol (S)-10 (149 mg, 0.818 mmol), docosanoic
acid acetoxime ester (330 mg, 0.834 mmol), dichloromethane
(4 ml), immobilized CAL (18 mg) and Pd/C catalyst (10 mg). The
product (R)-14 (130 mg, 0.314 mmol) was afforded as white crys-
tals in 38% overall yield. Mp = 79.2–80.0 °C. ½a D20
¼ ꢁ1:61 (c 5.1,
ꢀ
THF). 1H NMR d 4.21 (dd, J = 11.6 Hz, J = 4.8 Hz, 1H, OCO–CH2–),
4.15 (dd, J = 11.6 Hz, J = 6.0 Hz, 1H, OCO–CH2–), 3.96–3.90 (m, 1H,
–CH–OH), 3.72–3.67 (m, 1H, –CH2–OH), 3.63–3.57 (m, 1H,
–CH2–OH), 2.50 (br s, 1H, –CH–OH), 2.35 (t, J = 7.6 Hz, 2H, CH2COO),
2.04 (br s, 1H, –CH2–OH), 1.63 (quin, J = 7.2 Hz, 2H, CH2CH2COO),
1.37–1.18 (m, 36H, CH2), and 0.88 (t, J = 6.8 Hz, 3H, –CH3) ppm.
13C d 174.3, 70.3, 65.2, 63.3, 34.2, 31.9, 29.7 (11), 29.6, 29.4 (2),
29.2, 29.1, 24.9, 22.7 and 14.1 ppm. IR mmax 3150–3600 (br,
4.12. 3-Decanoyl-1,2-di(cis-octadec-9-enoyl)-sn-glycerol (S)-1
To
a solution of 3-decanoyl-sn-glycerol (R)-11 (103 mg,
0.418 mmol) and cis-octadec-9-enoic acid (295 mg, 1.04 mmol) in
dried dichloromethane (4.0 ml) were added DMAP (40 mg,
0.33 mmol) and EDAC (200 mg, 1.04 mmol). The resulting solution
was stirred magnetically at rt until TLC monitoring (80:20:1 pet.
ether/ether/acetic acid) indicated complete reaction. The reaction
was stopped by passing the reaction mixture through a short col-
umn packed with silica gel using petroleum ether as an eluent
and the solvent was removed in vacuo on a rotary evaporator.
O–H), 2918 (vs, C–H), 2849 (vs, C–H) and 1735 (vs, C@O) cmꢁ1
HRMS (ESI): calcd for
437.3619 amu.
.
C
25H50O4Na+ m/z 437.3601; found

B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
131
The pure product (S)-1 was afforded as a colourless oil (311 mg,
J = 7.6 Hz, 4H, CH2COO), 2.07–1.99 (m, 8H, ACH2ACH@), 1.66–1.57
(m, 6H, CH2CH2COO), 1.38–1.18 (m, 52H, CH2), and 0.88 (t,
J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.3 (2), 172.9, 130.3 (2), 130.0
(2), 128.1 (2), 127.9 (2), 68.9, 62.1 (2), 34.2, 34.1 (2) 32.0, 31.6
(2), 29.7–22.6 (32) and 14.1 (3) ppm. IR mmax 3009 (s, C–H), 2925
(vs, C–H), 2854 (vs, C–H) and 1747 (vs, C@O) cmꢁ1. HRMS (ESI):
calcd for C55H98O6Na+ m/z 877.7256; found 877.7316 amu.
0.401 mmol) in 95% yield. ½a D20
ꢀ
¼ ꢁ0:111 (c 9.0, benzene). 1H
NMR d 5.39–5.30 (m, 4H, HAC@C), 5.29–5.23 (m, 1H, CH–OCO),
4.29 (dd, J = 12.0 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.14 (dd,
J = 12.0 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.31 (t, J = 7.6 Hz, 2H,
CH2COO), 2.31 (t, J = 7.6 Hz, 4H, CH2COO), 2.05–1.97 (m, 8H,
ACH2ACH@), 1.66–1.57 (m, 6H, CH2CH2COO), 1.38–1.18 (m, 32H,
CH2), and 0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.2 (2),
172.8, 130.0 (2), 129.7 (2), 68.9, 62.1 (2), 34.2, 34.0 (2) 31.9 (3),
29.8–29.1 (21), 27.2 (3), 24.9 (3), 22.7 (3) and 14.1 (3) ppm. IR mmax
2925 (vs, C–H), 2854 (vs, C–H) and 1747 (vs, C@O) cmꢁ1. HRMS
(ESI): calcd for C49H90O6Na+ m/z 797.6630; found 797.6667 amu.
4.16. 3-Eicosanoyl-1,2-di(cis-octadec-9-enoyl)-sn-glycerol (S)-5
The same procedure was followed as described for (S)-1 using
3-eicosanoyl-sn-glycerol (R)-13 (200 mg, 0.517 mmol), cis-octa-
dec-9-enoic acid (162 mg, 0.573 mmol), dichloromethane
(4.0 ml), DMAP (40 mg, 0.33 mmol) and EDAC (190 mg, 0.991
mmol). The pure product (S)-5 was afforded as a white wax
(186 mg, 0.203 mmol) in 70% yield (as based on the fatty acid).
4.13. 3-Hexadecanoyl-1,2-di(cis-hexadec-9-enoyl)-sn-glycerol (S)-2
The same procedure was followed as described for (S)-1 using
3-hexadecanoyl-sn-glycerol (R)-12 (100 mg, 0.303 mmol), cis-
hexadec-9-enoic acid (169 mg, 0.664 mmol), dichloromethane
(4.0 ml), DMAP (20 mg, 0.16 mmol) and EDAC (128 mg,
0.668 mmol). The pure product (S)-2 was afforded as a colourless
½
a 2D0
ꢀ
¼ ꢁ0:028 (c 7.2, benzene). 1H NMR d 5.39–5.29 (m, 4H,
HAC@C), 5.29–5.23 (m, 1H, CH–OCO), 4.29 (dd, J = 12.0 Hz,
J = 4.4 Hz, 2H, –CH2–OCO), 4.14 (dd, J = 12.0 Hz, J = 6.0 Hz, 2H,
–CH2–OCO), 2.31 (t, J = 7.6 Hz, 2H, CH2COO), 2.31 (t, J = 7.6 Hz,
4H, CH2COO), 2.06–1.95 (m, 8H, ACH2ACH@), 1.66–1.57 (m, 6H,
CH2CH2COO), 1.38–1.18 (m, 72H, CH2), and 0.88 (t, J = 6.8 Hz, 9H,
–CH3) ppm. 13C d 173.2 (2), 172.8, 130.0 (2), 129.7 (2), 68.9, 62.1
(2), 34.2, 34.0 (2) 31.9 (3), 29.7–29.1 (31), 27.2 (3), 24.9 (3), 22.7
(3) and 14.1 (3) ppm. IR mmax 2924 (vs, C–H), 2853 (vs, C–H) and
1747 (vs, C@O) cmꢁ1. HRMS (ESI): calcd for C59H110O6Na+ m/z
937.8195; found 937.8196 amu.
oil (174 mg, 0.217 mmol) in 73% yield. ½a D20
¼ ꢁ0:055 (c 9.1, ben-
ꢀ
zene). 1H NMR d 5.40–5.31 (m, 4H, HAC@C), 5.30–5.24 (m, 1H,
CH–OCO), 4.30 (dd, J = 11.6 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.15
(dd, J = 11.6 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.32 (t, J = 7.6 Hz, 2H,
CH2COO), 2.32 (t, J = 7.6 Hz, 4H, CH2COO), 2.08–1.99 (m, 8H,
ACH2ACH@), 1.65–1.57 (m, 6H, CH2CH2COO), 1.38–1.18 (m, 56H,
CH2), and 0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.2 (2), 172.8,
130.0 (2), 129.7 (2), 68.9, 62.1 (2), 34.2, 34.0 (2) 31.9 (2), 31.8,
29.7–29.0 (23), 27.2 (3), 24.9 (3), 22.7 (3) and 14.1 (3) ppm. IR mmax
3004 (s, C–H), 2925 (vs, C–H), 2854 (vs, C–H) and 1746 (vs,
C@O) cmꢁ1. HRMS (ESI): calcd for C51H94O6Na+ m/z 825.6943;
found 825.6924 amu.
4.17. 3-Docosanoyl-1,2-di(cis-octadec-9-enoyl)-sn-glycerol (S)-6
The same procedure was followed as described for (S)-1 using
3-docosanoyl-sn-glycerol (R)-14 (104 mg, 0.251 mmol), cis-octa-
dec-9-enoic acid (168 mg, 0.595 mmol), dichloromethane
(4.0 ml), DMAP (20 mg, 0.16 mmol) and EDAC (105 mg,
0.548 mmol). The pure product (S)-6 was afforded as a white
4.14. 3-Hexadecanoyl-1,2-di(cis-octadec-9-enoyl)-sn-glycerol (S)-3
The same procedure was followed as described for (S)-1 using
3-hexadecanoyl-sn-glycerol (R)-12 (100 mg, 0.303 mmol), cis-octa-
dec-9-enoic acid (188 mg, 0.666 mmol), dichloromethane (4.0 ml),
DMAP (20 mg, 0.16 mmol) and EDAC (128 mg, 0.668 mmol). The
pure product (S)-3 was afforded as a colourless oil (181 mg,
wax (186 mg, 0.197 mmol) in 78% yield. ½a D20
¼ ꢁ0:027 (c 5.1,
ꢀ
benzene). 1H NMR d 5.39–5.29 (m, 4H, HAC@C), 5.29–5.23 (m,
1H, CH–OCO), 4.29 (dd, J = 12.0 Hz, J = 4.4 Hz, 2H, –CH2–OCO),
4.14 (dd, J = 12.0 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.31 (t, J = 7.6 Hz,
2H, CH2COO), 2.31 (t, J = 7.6 Hz, 4H, CH2COO), 2.06–1.96 (m, 8H,
ACH2ACH@), 1.66–1.57 (m, 6H, CH2CH2COO), 1.38–1.18 (m, 76H,
CH2), and 0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.2 (2),
172.8, 130.0 (2), 129.7 (2), 68.9, 62.1 (2), 34.2, 34.1 (2) 31.9 (3),
30.0–29.1 (33), 27.2 (3), 24.9 (3), 22.7 (3) and 14.1 (3) ppm. IR mmax
3004 (s, C–H), 2922 (vs, C–H), 2852 (vs, C–H) and 1743 (vs,
C@O) cmꢁ1. HRMS (ESI): calcd for C61H114O6Na+ m/z 965.8508;
found 965.8528 amu.
0.211 mmol) in 70% yield. ½a D20
ꢀ
¼ ꢁ0:041 (c 8.7, benzene). 1H
NMR d 5.39–5.30 (m, 4H, HAC@C), 5.30–5.24 (m, 1H, CH–OCO),
4.29 (dd, J = 11.6 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.14 (dd,
J = 11.6 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.31 (t, J = 7.6 Hz, 2H,
CH2COO), 2.31 (t, J = 7.6 Hz, 4H, CH2COO), 2.05–1.97 (m, 8H,
ACH2ACH@), 1.66–1.57 (m, 6H, CH2CH2COO), 1.38–1.18 (m, 64H,
CH2), and 0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.2 (2),
172.8, 130.0 (2), 129.7 (2), 68.9, 62.1 (2), 34.2, 34.0 (2) 31.9 (3),
29.8–29.1 (27), 27.2 (3), 24.9 (3), 22.7 (3) and 14.1 (3) ppm. IR mmax
2924 (vs, C–H), 2854 (vs, C–H) and 1747 (vs, C@O) cmꢁ1. HRMS
(ESI): calcd for C55H102O6Na+ m/z 881.7569; found 881.7567 amu.
4.18. 2,3-Dihexadecanoyl-1-cis-octadec-9-enoyl-sn-glycerol (S)-7
To a solution of 2,3-dihexadecanoyl-sn-glycerol (R)-15 (151 mg,
0.265 mmol) and cis-octadec-9-enoic acid (104 mg, 0.368 mmol) in
dried dichloromethane (4.0 ml) were added DMAP (24 mg,
0.20 mmol) and EDAC (62 mg, 0.32 mmol). The resulting solution
was stirred magnetically at rt until TLC monitoring (80:20:1 pet.
ether/ether/acetic acid) indicated complete reaction. The reaction
was stopped by passing the reaction mixture through a short col-
umn packed with silica gel using petroleum ether as an eluent
and the solvent was removed in vacuo on a rotary evaporator.
The pure product (S)-7 was afforded as a white wax (190 mg,
4.15. 3-Hexadecanoyl-1,2-di(cis-octadec-9,12-dienoyl)-sn-glyce-
rol (S)-4
The same procedure was followed as described for (S)-1 using
3-hexadecanoyl-sn-glycerol (R)-12 (100 mg, 0.303 mmol), cis-octa-
dec-9,12-dienoic acid (190 mg, 0.677 mmol), dichloromethane
(4.0 ml), DMAP (20 mg, 0.16 mmol) and EDAC (128 mg,
0.668 mmol). The pure product (S)-4 was afforded as a colourless
oil (257 mg, 0.300 mmol) in 99% yield. ½a D20
ꢀ
¼ ꢁ0:045 (c 9.0, ben-
0.228 mmol) in 86% yield. ½a D20
ꢀ
¼ ꢁ0:035 (c 8.7, benzene). 1H
zene). 1H NMR d 5.42–5.29 (m, 8H, HAC@C), 5.28–5.24 (m, 1H,
CH–OCO), 4.29 (dd, J = 11.6 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.14
(dd, J = 11.6 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.77 (t, J = 6.4 Hz, 4H,
@CHACH2ACH@), 2.31 (t, J = 7.6 Hz, 2H, CH2COO), 2.31 (t,
NMR d 5.39–5.30 (m, 2H, HAC@C), 5.29–5.24 (m, 1H, CH–OCO),
4.29 (dd, J = 11.6 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.14 (dd,
J = 11.6 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.31 (t, J = 7.6 Hz, 2H,
CH2COO), 2.31 (t, J = 7.6 Hz, 4H, CH2COO), 2.06–1.99 (m, 4H,

132
B. Kristinsson et al. / Tetrahedron: Asymmetry 25 (2014) 125–132
ACH2ACH@), 1.65–1.57 (m, 6H, CH2CH2COO), 1.39–1.19 (m, 68H,
CH2), and 0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.2 (2),
172.8, 130.0, 129.7, 68.9, 62.1 (2), 34.2, 34.0 (2) 31.9 (3), 29.7–
24.9 (30), 24.9 (3), 22.7 (3) and 14.1 (3) ppm. IR mmax 2954 (s, C–
H), 2918 (vs, C–H), 2850 (vs, C–H) and 1734 (vs, C@O) cmꢁ1. HRMS
(ESI): calcd for C53H100O6Na+ m/z 855.7412; found 855.7436 amu.
measurements. Funding from the Academy of Finland (grant
251623) is acknowledged.
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The same procedure was followed as described for (S)-7 using
2,3-dihexadecanoyl-sn-glycerol (R)-15 (155 mg, 0.272 mmol),
cis-octadec-9,12-dienoic acid (106 mg, 0.378 mmol), dichloro-
methane (4.0 ml), DMAP (21 mg, 0.17 mmol) and EDAC (60 mg,
0.31 mmol). The pure product (S)-8 was afforded as a white wax
(211 mg, 0.266 mmol) in 98% yield. ½a D20
¼ ꢁ0:053 (c 9.5, benzene).
ꢀ
1H NMR d 5.42–5.29 (m, 4H, HAC@C), 5.28–5.23 (m, 1H, CH–OCO),
4.29 (dd, J = 12.0 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.14 (dd,
J = 12.0 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.77 (t, J = 6.4 Hz, 2H,
@CHACH2ACH@), 2.31 (t, J = 7.6 Hz, 2H, CH2COO), 2.31 (t,
J = 7.6 Hz, 4H, CH2COO), 2.07–2.00 (m, 4H, ACH2ACH@),
1.66–1.57 (m, 6H, CH2CH2COO), 1.40–1.19 (m, 62H, CH2), and
0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.3 (2), 172.9, 130.2,
130.0, 128.1, 127.9, 68.9, 62.1 (2), 34.2, 34.1 (2) 32.0 (2), 31.5,
29.7–24.9 (34) and 14.1 (3) ppm. IR mmax 3009 (s, C–H), 2918 (vs,
C–H), 2850 (vs, C–H) and 1735 (vs, C@O) cmꢁ1. HRMS (ESI): calcd
for C53H98O6Na+ m/z 853.7256; found 853.7212 amu.
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ˇ
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NMR d 5.40–5.30 (m, 2H, HAC@C), 5.29–5.23 (m, 1H, CH–OCO),
4.29 (dd, J = 12.0 Hz, J = 4.4 Hz, 2H, –CH2–OCO), 4.14 (dd,
J = 12.0 Hz, J = 6.0 Hz, 2H, –CH2–OCO), 2.31 (t, J = 7.6 Hz, 2H,
CH2COO), 2.31 (t, J = 7.6 Hz, 4H, CH2COO), 2.05–1.97 (m, 4H,
ACH2ACH@), 1.65–1.57 (m, 6H, CH2CH2COO), 1.39–1.18 (m, 76H,
CH2), and 0.88 (t, J = 6.8 Hz, 9H, –CH3) ppm. 13C d 173.2 (2),
172.9, 130.0, 129.7, 68.9, 62.1 (2), 34.2, 34.0 (2) 31.9 (3),
29.7–24.9 (34), 24.9 (3), 22.7 (3) and 14.1 (3) ppm. IR mmax 2923
(vs, C–H), 2853 (vs, C–H) and 1747 (vs, C@O) cmꢁ1. HRMS (ESI):
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Acknowledgements
Novozymes AS in Denmark is acknowledged for the lipase and
Dr. Sigridur Jonsdottir at University of Iceland for accurate MS