Chemoenzymatic synthesis of a focused library of enantiopure structured 1-O-alkyl-2,3-diacyl-sn-glycerol type ether lipids

Article abstract of DOI:10.1016/j.tet.2011.01.032

A highly efficient two-step chemoenzymatic synthesis of enantiopure structured ether lipids of the 1-O-alkyl-2,3-diacyl-sn-glycerol type has been developed. Chimyl, batyl and selachyl alcohols possessing pure saturated fatty acid (SFA) attached to the sn-3 position and pure EPA and DHA attached to the sn-2 position were obtained under full regiocontrol. This was offered by mild conditions and a highly efficient lipase that operated at room temperature. High-resolution ¹H NMR spectroscopy was used to monitor the progress of the reactions and to evaluate the full regiocontrol of the reactions involved by keeping track of all prospective adducts involved in these reactions. This was extended to preparation of a focused library of eight monoacyl intermediate adducts for all even-numbered SFA ranging from C₂-C₁₆ and the corresponding EPA and DHA structured diacyl glycerol ethers (DAGE) products for chimyl, batyl and selachyl alcohols, the total of 72 compounds.

Full text of DOI:10.1016/j.tet.2011.01.032

Tetrahedron 67 (2011) 1821e1836
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Chemoenzymatic synthesis of a focused library of enantiopure structured
1
-O-alkyl-2,3-diacyl-sn-glycerol type ether lipids
Carlos D. Magnusson, Anna V. Gudmundsdottir, Gudmundur G. Haraldsson *
Science Institute, University of Iceland, Dunhaga 3, 107 Reykjavik, Iceland
a r t i c l e i n f o
a b s t r a c t
Article history:
A highly efficient two-step chemoenzymatic synthesis of enantiopure structured ether lipids of the 1-O-
alkyl-2,3-diacyl-sn-glycerol type has been developed. Chimyl, batyl and selachyl alcohols possessing pure
saturated fatty acid (SFA) attached to the sn-3 position and pure EPA and DHA attached to the sn-2
position were obtained under full regiocontrol. This was offered by mild conditions and a highly efficient
lipase that operated at room temperature. High-resolution 1H NMR spectroscopy was used to monitor
the progress of the reactions and to evaluate the full regiocontrol of the reactions involved by keeping
track of all prospective adducts involved in these reactions. This was extended to preparation of a focused
library of eight monoacyl intermediate adducts for all even-numbered SFA ranging from C eC16 and the
2
corresponding EPA and DHA structured diacyl glycerol ethers (DAGE) products for chimyl, batyl and
selachyl alcohols, the total of 72 compounds.
Received 27 October 2010
Received in revised form 15 December 2010
Accepted 11 January 2011
Available online 15 January 2011
Keywords:
1-O-Alkyl-sn-glycerols
Structured ether lipids
n-3 PUFA
Lipase
Focused library
Ó 2011 Elsevier Ltd. All rights reserved.
1. Introduction
rac-glycerols have been exploited to facilitate the delivery of anti-
biotics and antineoplastic drugs to the brain by increasing the
8
The naturally occurring 1-O-alkyl-sn-glycerol based ether lipids
permeability of the bloodebrain barrier.
9
(
EL) are ubiquitously found as 1-O-alkyl-2,3-diacyl-sn-glycerols in
The sn-(stereospecific numbering) terminology implies that the
absolute configuration of the natural 1-O-alkyl-sn-glycerols is S. The
most prevalent fatty hydrocarbon chains representing the O-alkyl
1
the non-polar lipid fractions of aquatic and terrestrial animals.
Such ether lipids are also known as diacyl glycerol ethers (DAGE).
In humans, they are widely found in various tissues, usually as
minor lipid components. They are present in particularly high
amounts in the liver oils of various cartilaginous fish species in-
cluding sharks, rays and chimaeras. For instance, the liver oil of
deep-sea sharks of the Squaliformes order usually contains about
2
moiety found in nature emerge from palmityl (C16:0), stearyl (C18:0
)
2
and oleyl (C18:1) alcohols. The corresponding 1-O-alkyl-sn-glycerols
1e3 are known by their trivial names as chimyl, batyl and selachyl
alcohols, named after the cartilaginous fish species they were first
1
isolated from, i.e., sharks, chimaeras and rays, respectively.
3
5% DAGE, but up to 89% DAGE has been reported.
O
The 1-O-alkyl-sn-glycerols exhibit multiple biological activities
1
a,4
1
OH
OH
and are considered to have high therapeutic potential.
They have
been demonstrated to possess antineoplastic, immune stimulant
and adjuvant properties. 1-O-Alkyl-sn-glycerols have been shown
to prevent leucopenia and thrombocytopenia in patients un-
O
2
OH
OH
1
a,5
dergoing cancer radiation therapy.
They have been shown to
enhance antibody production and stimulate macrophages in vivo,
a fact that may explain their relatively high amounts found in hu-
man colostrum and milk, presumably to compensate the immature
immune system in newborns. Furthermore, studies have shown
that 1-O-alkyl-sn-glycerols reduce tumour growth and metastasis
O
OH
OH
3
6
7
rate of tumour cells in vivo. Besides these biological activities, the
amphiphilic and surface active properties of synthetic 1-O-alkyl-
The long-chain n-3 polyunsaturated fatty acids (PUFA) are
10
characteristic of marine fat and their numerous beneficial effects
*
Corresponding author. Tel.: þ345 525 4818; fax: þ354 552 8911; e-mail
11
on human health have been firmly established. These effects are
address: gghar@hi.is (G.G. Haraldsson).
0
040-4020/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2011.01.032

1822
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
almost solely attributed to eicosapentaenoic acid (EPA) and doco-
to act at the terminal primary alcohol positions of the glycerol
9
sahexaenoic acid (DHA), the two most prevalent n-3 PUFA in fish.
backbone lipases are perfectly suited as biocatalysts to prepare
12
18,23
The strongest evidence relates to their cardioprotective effects,
structured TAG and DAGE of the type at issue here.
The low
13
various inflammatory and autoimmune disorders, brain function
and cognitive health.14 More recently, EPA and DHA have emerged
as anti-carcinogenic nutrients, especially regarding treatment of
tumour of the upper digestive tract and pancreas.15 This has lead to
high demands for EPA and DHA concentrates by the health and
dietary food industry as dietary supplements as well as the phar-
temperature and mild conditions under which they act are also
crucial for maintaining the regiocontrol by avoiding acyl-migration
21,24
side-reactions
that are common problems when dealing with
synthesis involving partially acylated polyols and carbohydrates.
The immobilized Candida antarctica lipase B (CAL-B) was observed
to display a superb regioselectivity by acting exclusively at the 1,3-
positions of glycerol when using activated vinyl esters of the saturated
16
maceutical industry for drugs.
Structured lipids17 usually refer to lipids offering a pre-
determined composition and distribution of fatty acids within the
glycerol framework. Structured triacylglycerols (TAG) possessing
bioactive long-chain PUFA attached to the 2-position and medium-
chain fatty acids (MCFA) located at the end-positions have acquired
fatty acids in dichloromethane at 0e4 C. The vinyl esters enabled the
ꢀ
lipase to act efficiently at low temperature as well as rendering irre-
versibility to the transesterification reaction by ketoeenol tautomer-
ism undergone by the enolic leaving group. The reaction was
completed in 3e5 h and there were no signs of any acyl-migration side
reaction taking place. In all cases the 1,3-diacyl glycerol (1,3-DAG)
intermediate adducts were accomplished in excellent yields after
purification, and were regioisomerically and chemically pure.
growing attention of scientists as human dietary and health sup-
plements.1
7,18
This pertains to MCFA when located at the end-po-
rapid hydrolysis by 1,3-regioselective
sitions undergoing
a
pancreatic lipase in the digestive tract and a fast delivery into the
liver where they are consumed as a quick source of energy. The
remaining 2-monoacylglycerols (2-MAG) become a source of es-
Pure EPA and DHA were subsequently introduced to the 2-po-
sition of the 1,3-DAG intermediates using EDAC (1-(3-dimethyla-
minopropyl)-ethylcarbodiimide hydrochloride) as a coupling agent
in the presence of dimethylaminopyridine (DMAP). The reaction
was completed in only 3e4 h with stoichiometric amounts of EPA
and DHA to accomplish the regiopure structured TAG products in
excellent yields after purification. There were no indications of any
losses in regiocontrol during the coupling reaction as was firmly
19
sential fatty acids. Structured MLM (medium-long-medium) type
TAG constituting EPA or DHA located at the 2-position with MCFA at
18,20
the end-positions are strongly desired.
The work described in this report aims at possibly combining
within the same molecule the beneficial effects of enantiopure
ether lipids of the 1-O-alkyl-sn-glycerol type, the MLM type
structured TAG described above, and the long-chain n-3 PUFA. With
that in mind a chemoenzymatic synthesis of structured DAGE of the
ALM type (alkyl-long-medium) enantiopure ether lipids comprised
1
13
21
established by high-resolution H and C NMR spectroscopy.
The current report describes the synthesis of enantiopure ALM
type structured DAGE possessing a pure SFA (C , C , C , C
14 and C16) at the sn-3 position with a pure bioactive PUFA (EPA or
2
4
6
8
, C10, C12,
C
of pure EPA or DHA at the sn-2 position and a pure MCFA (C
6
, C
8
, C10
DHA) located at the sn-2 position of the 1-O-alkyl-sn-glycerol
backbone of chimyl, batyl and selachyl alcohols. A two-step che-
moenzymatic approach similar to that described above for the
MLM type structured TAG was followed to introduce the SFA into
the sn-3 position of the 1-O-alkyl-sn-glycerol moiety and pure EPA
and DHA subsequently to the sn-2 position (see Scheme 2). It was
anticipated that the CAL would retain its excellent regioselectivity
towards the 1-O-alkyl-sn-glycerols.
and C12) at the sn-3 position was addressed (Fig. 1). It was decided
to base this approach on each of the chimyl, batyl and selachyl type
alcohols, optically pure possessing the natural S configuration.
Furthermore, this work was extended to cover short-chain fatty
acids (C
2 4
and C ) and longer-chain saturated fatty acids (C14 and
C
16) to complete the task to prepare a focused library of such
structured DAGE and all their monoacyl intermediate adducts.
O
O
O
O
O
O
O
O
O
O
Fig. 1. ALM type structured chimyl alcohol possessing EPA and capric acid (top) and selachyl alcohol possessing DHA and caprylic acid (bottom).
2
. Results and discussion
Previous reports describe the synthesis of symmetrically struc-
2.1. Preparation of chimyl, batyl and selachyl alcohols
Optically pure chimyl, batyl and selachyl alcohols 1e3 were
prepared in two steps starting from enantiopure (R)-solketal (2,3-
O-isopropylidene-sn-glycerol) as a chiral precursor (see Scheme 1).
The ether moiety was introduced by alkylation with the corre-
tured TAG of the MLM and the more general ABA type by a highly
efficient two-step chemoenzymatic approach starting from glyc-
21
erol. The synthesis by traditional synthetic organic chemistry
methods requires a full regiocontrol that can hardly be accom-
plished without multi-step protectionedeprotection processes.
Lipases are well established as powerful bioacatalysts in synthetic
organic chemistry.22 Owing to their regioselectivity and preference
ꢀ
sponding alkyl bromides at 35e40 C in the absence of a solvent
using grounded potassium hydroxide as a base in the presence of
tetra-n-butylammonium bromide serving as a phase-transfer cat-
25
alyst. The resulting 1-O-alkyl-2,3-O-isopropylidene-sn-glycerol

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1823
H
temperature without any deterioration of results. At that tempera-
ture only 3 h wererequiredfor the reaction toproceed tocompletion
when using 1.25 equiv of the vinyl esters as based on mole EL. This
became the condition of choice for all cases addressed in the current
report. All monoacyl EL adducts 4aeh, 5aeh and 6aeh were
obtained enantio- and regio-pure in excellent yields (90e96%) as
can be noticed in Tables 1e3 also showing the specific optical rota-
tion of these compounds, respectively, for chimyl, batyl and selachyl
alcohols. Yields are based on highly pure material as was established
by high-resolution H and C NMR spectroscopy analysis after
crystallization from petroleum ether (short-column silica gel treat-
ment for the monounsaturated shortest chain selachyl alcohol ad-
ducts 6aec after distilling off the volatile excessive vinyl ester and
by-produced carboxylic acid in vacuo).
R
R
O
O
O
a
O
O
b
O
O
OH
OH
(
(
(
S)-1: R = C16H33
S)-2: R = C18H37
S)-3: R = C18H35
ꢀ
Scheme 1. Reagents and conditions: (a) KOH, n-Bu
b) p-TsOH, THF/water (15:5), reflux, 15 h.
4
NBr, R-Br, 35e40 C, 15 h;
1
13
(
intermediate adducts were usually not purified. A subsequent
deprotection of the isopropylidene moiety under acidic conditions
afforded the enantiopure 1-O-alkyl-sn-glycerols 1e3, respectively,
for chimyl, batyl and selachyl alcohols, in excellent yields.
2
.3. The coupling reaction
The subsequent chemical coupling reaction to introduce pure
EPA and DHA into the sn-2 position of the 3-acyl-1-O-alkyl-sn-
glycerol adducts 4aeh, 5aeh and 6aeh (see Scheme 2) was per-
formed at room temperature in dichloromethane under similar
conditions as described for the TAG synthesis. EDAC (1.4 equiv) was
used as a coupling agent in the presence of DMAP (80% as based on
mole) with an exact stoichiometric amount of EPA or DHA as based
on the 1-O-alkyl-sn-glycerol adducts. The reactions were com-
pleted within 15 h, significantly slower than the 3e4 h required for
the TAG synthesis where lower amounts of EDAC (1.2 equiv) and
2
.2. The enzymatic step
When (S)-chimyl, batyl and selachyl alcohols 1e3 were treated
under similar conditions as described for TAG using vinyl esters of
8
SFA (a, b, c, d, e, f, g and h, respectively, for C , C , C , C , C10, C12, C14
2
4
6
and C16) and the CAL-B the excellent degree of regioselectivity was
indeed preserved. This resulted in the 3-acyl-1-O-alkyl-sn-glycerol
adducts 4aeh, 5aeh and 6aeh, respectively, for chimyl, batyl and
selachyl alcohols (see Scheme 2), that like before were obtained in
virtually quantitative yields.
21
DMAP (20%) were needed. Chemically and regioisomerically pure
structured DAGE 7aeh, 8aeh and 9aeh (for EPA) and 10aeh,
R
R
R
O
PUFA
O
O
O
O
a
OH
O
b
OH
OH
O
O
O
SFA
SFA
(
(
(
S)-1
S)-2
S)-3
(R)-4a-h
(R)-5a-h
(R)-6a-h
(R)-7a-h
(R)-8a-h
(R)-9a-h
(R)-10a-h
(R)-11a-h
(R)-12a-h
SFA = CH3 (a); C3H7 (b); C5H11 (c); C7H15 (d); C9H19 (e); C11H23 (f); C13H27 (g); C15H31 (h)
1, 4, 7, 10: R = C16H33
2, 5, 8, 11: R = C18H37
3, 6, 9, 12: R = C18H35
7, 8, 9: PUFA = EPA
10, 11, 12: PUFA = DHA
2 2 2 2
Scheme 2. Reagents and conditions: (a) C. antarctica lipase, SFA as vinyl esters, CH Cl , rt, 3 h; (b) PUFA, EDAC, DMAP, CH Cl , rt 15 h.
The highly regioselective acylation of the 1-O-alkyl-sn-glycerols
11aeh and 12aeh (for DHA), respectively, for chimyl, batyl and
selachyl alcohols, were afforded as yellowish oils in excellent yields
(90e95%) after purification by short-column chromatography
treatment on silica gel. The yields are revealed in Tables 1e3 along
with their specific optical rotation.
was initially performed under conditions similar to those described
ꢀ
for glycerol in dichloromethane at 0e4 C. More solvent was
needed this time to dissolve the 1-O-alkyl-sn-glycerols. This slowed
down the reaction rate so that more vinyl ester was required for
obtaining rates comparable to those obtained before. With
2
M equiv the reactions were completed after 4e5 h reaction time
2.4. Regiocontrol and investigation of acyl-migration
and there were no indications of any acyl-migration products and
the lipase was observed to disregard the sn-2 position. Comparable
results were accomplished when using 1.25 equiv, but the reaction
required 15e17 h as was demonstrated for chimyl and batyl alco-
hols using vinyl caprylate (C ). A need for modification to lower the
8
excessive use of vinyl esters as well as shortening the reaction time
was clearly apparent.
Acyl-migration24 is a major problem in regioselective acylations
of polyhydroxy compounds, such as glycerols and carbohydrates.
This is a spontaneous intramolecular rearrangement, independent
of the lipase, that competes against regioselectivity and regiocon-
trol. The acyl-migration processes are accelerated by various pa-
rameters such as temperature, the most prominent one in the
21b
Ether lipids of the DAGE type have been observed to be less prone
than TAG to undergo acyl-migration under transesterification con-
current case,
the presence of acid and base, type of solvent and
2
4
immobilized enzyme carrier material. Obviously, the increased
reaction rate offered by the activated vinyl esters and their irre-
versibility is of importance in terms of shorter reaction time. The
2
6
ditions when using lipase. Accordingly, the current lipase pro-
moted acylation reaction was observed to tolerate room

1824
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
Table 1
Table 3
Type of intermediates (4aeh) and structured DAGE products constituting pure SFA
and EPA (7aeh) or DHA (10aeh) for the chimyl alcohol series
Type of intermediates (6aeh) and structured DAGE products constituting pure SFA
and EPA (9aeh) or DHA (12aeh) for the selachyl alcohol series
Compound
SFA
PUFA
Yield (%)
[a
]
D
Compound
SFA
PUFA
Yield (%)
[a]
D
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
R)-4a
R)-4b
R)-4c
R)-4d
R)-4e
R)-4f
R)-4g
R)-4h
R)-7a
R)-7b
R)-7c
R)-7d
R)-7e
R)-7f
R)-7g
R)-7h
R)-10a
R)-10b
R)-10c
R)-10d
R)-10e
R)-10f
R)-10g
R)-10h
eCH
3
d
95
92
91
90
92
95
92
91
90
90
91
90
90
90
91
91
91
92
92
91
92
91
93
91
ꢁ3.7
ꢁ3.5
ꢁ3.8
ꢁ3.3
ꢁ3.5
ꢁ3.7
ꢁ4.0
ꢁ2.5
ꢁ5.0
ꢁ5.4
ꢁ4.8
ꢁ3.9
ꢁ2.9
ꢁ4.1
ꢁ3.7
ꢁ4.5
ꢁ5.8
ꢁ5.2
ꢁ4.7
ꢁ3.3
ꢁ3.0
ꢁ3.1
ꢁ3.7
ꢁ3.4
(R)-6a
(R)-6b
(R)-6c
(R)-6d
(R)-6e
(R)-6f
(R)-6g
(R)-6h
(R)-9a
(R)-9b
(R)-9c
(R)-9d
(R)-9e
(R)-9f
(R)-9g
(R)-9h
(R)-12a
(R)-12b
(R)-12c
(R)-12d
(R)-12e
(R)-12f
(R)-12g
(R)-12h
eCH
3
d
96
95
93
90
90
90
90
95
90
90
91
91
90
90
91
92
90
90
90
90
91
90
90
91
ꢁ3.5
ꢁ3.6
ꢁ3.6
ꢁ3.3
ꢁ2.8
ꢁ3.1
ꢁ2.7
ꢁ2.3
ꢁ5.8
ꢁ5.1
ꢁ4.7
ꢁ4.6
ꢁ4.5
ꢁ4.4
ꢁ3.5
ꢁ4.8
ꢁ5.5
ꢁ4.7
ꢁ4.3
ꢁ4.1
ꢁ3.7
ꢁ3.8
ꢁ3.5
ꢁ3.8
eC
3
eC
5
eC
7
eC
9
H
H
H
H
7
d
eC
3
eC
5
eC
7
eC
9
H
H
H
H
7
d
11
15
19
d
11
15
19
d
d
d
d
d
eC11
eC13
eC15
H
H
H
23
27
31
d
eC11
eC13
eC15
H
H
H
23
27
31
d
d
d
d
d
eCH
3
EPA
EPA
EPA
EPA
EPA
EPA
EPA
EPA
DHA
DHA
DHA
DHA
DHA
DHA
DHA
DHA
eCH
3
EPA
EPA
EPA
EPA
EPA
EPA
EPA
EPA
DHA
DHA
DHA
DHA
DHA
DHA
DHA
DHA
eC
3
eC
5
eC
7
eC
9
H
7
eC
3
eC
5
eC
7
eC
9
H
7
H
H
H
11
15
19
H
H
H
11
15
19
eC11
eC13
eC15
H
H
H
23
27
31
eC11
eC13
eC15
H
H
H
23
27
31
eCH
3
eCH
3
eC
3
eC
5
eC
7
eC
9
H
7
eC
3
eC
5
eC
7
eC
9
H
7
H
H
H
11
15
19
H
H
H
11
15
19
eC11
eC13
eC15
H
H
H
23
27
31
eC11
eC13
eC15
H
H
H
23
27
31
Table 2
Type of intermediates (5aeh) and structured DAGE products constituting pure SFA
and EPA (8aeh) or DHA (11aeh) for the batyl alcohol series
the reverse regioisomeric structured DAGE product 15d through
the coupling step.
There were no indications of any losses in regiocontrol during
the enzymatic and coupling reactions (Scheme 2) as was estab-
lished after careful collection of samples from the progressing re-
Compound
SFA
PUFA
Yield (%)
[a]
D
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
(
R)-5a
R)-5b
R)-5c
R)-5d
R)-5e
R)-5f
R)-5g
R)-5h
R)-8a
R)-8b
R)-8c
R)-8d
R)-8e
R)-8f
R)-8g
R)-8h
R)-11a
R)-11b
R)-11c
R)-11d
R)-11e
R)-11f
R)-11g
R)-11h
eCH
3
d
91
90
91
94
95
90
91
90
90
91
90
92
92
90
92
90
90
92
92
90
95
90
93
90
ꢁ3.6
ꢁ3.2
ꢁ4.3
ꢁ3.4
ꢁ3.5
ꢁ3.0
ꢁ2.5
ꢁ2.3
ꢁ6.1
ꢁ5.6
ꢁ4.7
ꢁ5.0
ꢁ4.5
ꢁ4.3
ꢁ3.7
ꢁ4.7
ꢁ6.0
ꢁ4.0
ꢁ3.7
ꢁ3.5
ꢁ2.8
ꢁ3.0
ꢁ3.8
ꢁ4.1
eC
3
eC
5
eC
7
eC
9
H
H
H
H
7
d
1
13
11
15
19
d
actions and their analysis by high-resolution H (400 MHz) and
C
d
NMR spectroscopy. This prompted further investigations on the
stability of the 3-acyl-1-O-alkyl-sn-glycerol adducts under the
conditions of the coupling reaction and to what extent a possible
acyl-migration to the corresponding 2-acyl-1-O-alkyl-sn-glycerol
might occur. Investigations involving 3-acyl-1-O-alkyl-sn-glycerol
d
eC11
eC13
eC15
H
H
H
23
27
31
d
d
d
eCH
3
EPA
EPA
EPA
EPA
EPA
EPA
EPA
EPA
DHA
DHA
DHA
DHA
DHA
DHA
DHA
DHA
eC
3
eC
5
eC
7
eC
9
H
7
8
(batyl alcohol with C ) revealed only traces of the acyl-migration
H
11
H
15
H
19
adduct 13d after 48 h stirring at room temperature under the exact
background conditions of the coupling reaction in dichloromethane
in the presence of EDAC and DMAP with the 3-acyl-1-O-alkyl-sn-
glycerol initial concentration thus remaining virtually constant
throughout the experiment. After 120 h there were indications of
eC11
eC13
eC15
H
H
H
23
27
31
eCH
3
0.6% presence of the 2-acyl adduct.
eC
3
eC
5
eC
7
eC
9
H
7
H
11
H
15
H
19
This compares to less than 0.5% acyl-migration observed in
analogous investigations for 1,3-DAG comprising C10 under similar
21b
condition after 24 h. In that case the coupling reaction proceeded
eC11
eC13
eC15
H
H
H
23
27
31
to completion in less than 4 h, whereas the current case required
15 h. All this supports a full regiocontrol of these reactions. It should
be added that the equilibrium level of 2-acyl-1-O-alkyl-sn-glycerol
was determined to remain at approximately11% in dichloromethane
at room temperature as obtained after 7 days by the use of catalytic
amount of Amberlyst 15 when starting with pure 3-acyl-1-O-alkyl-
sn-glycerol. This is comparable to the corresponding equilibrium
results from previous and current studies also indicate that longer-
chain acyl groups are less prone to undergo acyl-migration than the
shorter-chain.
Scheme 3 covers all possible interrelated reactions involved in
the enzymatic and coupling steps, including those of the lipase
acting directly at the sn-2 position and the acyl-migration side re-
2
4d,e
levels of 1-MAG and 2-MAG with 1-MAG dominating.
1
2.5. Regiocontrol evaluation by H NMR spectroscopic
analysis
8
actions, as illustrated for batyl alcohol 2, caprylic acid C and EPA.
The desired two-step pathway consists of the regioselective lipase
promoted conversion of 2 into 5d and the subsequent chemical
coupling of EPA to the sn-2 position by EDAC to form 8d. Unwanted
acyl-migration is possible in both reaction steps resulting in the
wrong regioisomeric adduct 13d. Once formed that adduct may
undergo a fast lipase promoted acylation to form the unwanted
DAGE 14d possessing two MCFA under the enzymatic conditions or
All four types of compounds potentially involved in the current
structured EL synthesis displayed characteristic resonance pattern
in the glyceryl moiety proton region of their H NMR spectra. The
spectra may be easily assigned thus making it quite straightforward
to track down all individual EL constituents present in a reaction
mixture at a time and quantifying them by reasonable or good
1

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1825
C18H37
O
OH
Lipase
slow)
OH
Lipase
fast)
(
(
(
S)-2
C18H37
O
(
C7H15
O
C18H37
O
C7H15
C18H37
C18H37
O
EPA
O
O
O
O
O
OH
O
O
O
O
EPA
EDAC
O
Acyl-migration
EPA
EDAC
O
EPA
OH
O
C7H15
C7H15
R)-15d
(S)-13d
(R)-5d
(R)-8d
Lipase
fast)
Lipase
(slow)
(
C18H37
O
C7H15
O
O
O
O
C7H15
(R)-14d
Scheme 3. A pathway showing all reactions potentially involved in the enzymatic and coupling reactions.
accuracy by integration of appropriate signals. This became an
excellent tool to monitor in detail the progress of the reactions as
they proceeded and evaluating their regiocontrol in a similar
manner as was described in the previous structured TAG
synthesis.2
involved include the methylene proton pair of the sn-1 position as
the first category, the single proton that belongs to the sn-2
methine group as the second one, and the two methylene protons
representing the sn-3 position as the third one. The protons that
belong to the methylene group next to oxygen in the alkoxy moiety
1a,27
2
8
The adducts involved include 1-O-alkyl-sn-glycerol starting
material, both 3-acyl- and 2-acyl-1-O-alkyl-sn-glycerol monoacyl
intermediates and the DAGE product. Fig. 2 presents the glyceryl
resonating as two very close ABX
2
-type doublets of triplets at
d
3.50e3.40 ppm interfere with the sn-1 methylene protons in the
up-field part of that region. This was particularly prominent in the
cases of 1-O-alkyl-sn-glycerol and 3-acyl-1-O-alkyl-sn-glycerol,
where the sn-2 position remained non-acylated.
1
proton segment of their H NMR spectra that were acquired from
reference samples obtained from the current synthesis (batyl al-
cohol, C
8
and EPA). The 2-acyl-1-O-alkyl-sn-glycerol adduct (13d in
The chemical shift of the glyceryl protons was much dependent
upon whether the attached hydroxyl groups were free or acylated.
This was particularly evident for the proton at the sn-2 position.
Acylation was also observed to result in a down-field shift of the
protons of adjacent carbons. Very similar trends were also observed
accordance with Scheme 3) is the prospective acyl-migration
product of 3-acyl-1-O-alkyl-sn-glycerol. It was generated from 5d
as the minor fraction (11%) after inducing acyl-migration equilib-
rium under acidic conditions (Amberlyst 15 or p-TsOH as a catalyst)
in dichloromethane at room temperature and a subsequent sepa-
ration and purification by preparative TLC on silica gel.
21a
for individual acylglycerols in the structured TAG synthesis.
For the 1-O-alkyl-sn-glycerol there is multiplet at
3.89e3.83 ppm corresponding to the proton located at the sn-2
a
d
position. The non-substituted methylene group protons of the sn-3
position resonated as two distinct multiplets at 3.75e3.69 and
3.67e3.62 ppm. Each of the methylene protons of the ether
substituted glyceryl sn-1 carbon resonated as an ABX-type doublet of
doublets at 3.54 and 3.48 ppm in close vicinity to the protons be-
longing to the methylene group next to the oxygen atom of the alkyl
moiety.
Substantial changes were observed in the glyceryl proton region
when an acyl group was introduced to the sn-3 hydroxyl group of 1-
O-alkyl-sn-glycerol. Upon acylation, the methylene protons be-
longing to the sn-3 position underwent nearly 0.5 ppm down-field
shift, now resonating as two distinguishable ABX-type doublets of
doublets at
d 4.17 and 4.12 ppm. The proton belonging to the sn-2
position also underwent a significant shift as a result of acylation of
the vicinal methylene group, this time resonating as a multiplet at
4.03e3.95 ppm. The remaining sn-1 methylene protons resonated
at chemical shift values close to those obtained for the 1-O-alkyl-
sn-glycerol as two distinguishable ABX-type doublets of doublets at
3.49 and 3.42 ppm with the methylene protons next to oxygen in
the alkoxy moiety still strongly interfering.
Fig. 2. The glyceryl proton segment of the 1H NMR spectra of all 1-O-alkyl-sn-glycerol
type adducts involved in the synthesis of structured DAGE.
More dramatic changes occurred in the glyceryl proton region
upon the second acylation as can be noticed from the corresponding
spectrum for 1-O-alkyl-2,3-diacyl-sn-glycerol. Now, all the protons
are very well separated and distinct and easily assigned. This time the
As may be noticed from Fig. 2 there are three main categories of
signals present in the glyceryl proton region that are readily as-
signable for each of the individual adducts. The five protons

1826
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
proton located at the sn-2 position resonated as a multiplet at
5.22e5.17 ppm. This down-field shift of more than 1.3 ppm upon
acylation is comparable to that observed in the synthesis of the cor-
responding triacylglycerols bystepwise acylationof glycerol. There
wasalsoa furtherdown-fieldshiftof the vicinal O-acyl substitutedsn-
firm regiocontrol. The resulting focused libraries of structured TAG
and EL and their intermediate adducts may be exploited for various
purposes, such as to compare the biological effects of these com-
pounds and individual fatty acids by screening, as standards for
analysis, as various drug supplements, to develop prodrugs as well
as serving as potential drugs.
d
21a
3
methylene group protons, now appearing as two well separated
ABX-type doublets of doublets at 4.33 and 4.16 ppm. It is of interest to
notice changes occurring to the 1-O-alkyl substituted sn-1 methylene
group protons now resonating as two close ABX-type doublets of
doublets at 3.57e3.50 ppm, this time showing stronger second order
behaviour.
4. Experimental
4.1. General
In the glyceryl proton region of the 2-acyl-1-O-alkyl-sn-glycerol
adduct the proton at the sn-2 position resonated as a quintet at
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
5.00 ppm. The protons belonging to the non-acylated sn-3
methylene group resonated as a multiplet at 3.83e3.80 ppm. There
are two doublets of doublets representing each of the two sn-1
methylene protons at 3.64 and 3.60 ppm, well apart from the two
(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; dt, doublet of triplets; m, multiplet. The number of car-
2
ABX -type doublets of triplets belonging to the alkoxy group.
1
13
It is evident that the high-resolution H NMR spectroscopy is of
great importance for monitoring the reactions involved in the
chemoenzymatic synthesis of structured ether lipids, both in terms
of their progress as well as evaluating the chemical and regioiso-
meric purity of the compounds and the regiocontrol in these re-
bon nuclei behind each C signal is indicated in parentheses after
each chemical shift value, when there is more than one carbon
1
1
responsible for the peak. Two dimensional He H COSY and HET-
COR correlation analysis were utilized for unequivocally assigning
13
C NMR peaks. All Infrared spectra were conducted on a Nicolet
1
actions. The limit of quantification as detected by 400 MHz H NMR
Avatar 360 FT-IR (E.S.P.) Spectrophotometer on a ZnSe plate and on
KBr pellets for neat liquid and solid compounds, respectively. The
optical activities were measured on an Autopol V from Rudolph
Research Analytical, New Jersey USA. Melting points were de-
termined on a B u€ chi 520 melting point apparatus and are un-
corrected. The high-resolution mass spectra (HRMS) were acquired
on a Bruker micrOTOF-Q mass spectrometer equipped with an
atmospheric pressure chemical ionization chamber (APCI) or an
E-spray atmospheric pressure ionization chamber (ESI). All data
analyses were done on Bruker software.
spectroscopy has been determined for a possible acyl-migration of
1
,3-DAG to 1,2-DAG as well as 1-MAG to 2-MAG in previous
21b
work.
The results indicate that the levels of 2-MAG and 1,2-DAG
acyl-migration products can be accurately quantified down to
.25 mol % for practical sample concentration levels. Obviously, the
0
limit of detection is well below that. The detection degree in the
current case involving the structured EL synthesis is estimated to be
close to that in the above acylglycerol studies and regiopurity levels
of the minimum of 99.9% can be confidently asserted.
The immobilized C. antarctica lipase (Novozym 435; CAL-B) was
supplied as a gift from Novozyme A/S (Bagsvaerd, Denmark). All
chemicals and solvents were used without further purification unless
otherwise stated. (R)-2,3-O-Isopropylidene-sn-glycerol (98%, 99% ee)
and EDAC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydro-
chloride) were obtained from SigmaeAldrich (Steinheim, Germany).
1-bromohexadecane (97%), 1-bromooctadecane (98%), cis-1-bromo-
hexadec-9-ene (ꢂ99%) and tetrabutyl ammonium bromide were
purchased from Sigma Chemicals (St. Louis, Missouri, USA). Vinyl
butanoate (>97%), vinyl hexanoate (>99%), vinyl octanoate (>99%),
vinyl decanoate (>99%), vinyl tetradecanoate (>99%) and vinyl hex-
adecanoate (96%) were obtained from TCI Europe (Zwijndrecht, Bel-
gium), vinyl acetate (ꢃ99%), and vinyl dodecanoate (ꢃ99%) from
Fluka Chemie GmbH (Buchs, Switzerland) and 4-dimethylamino-
pyridine (99%) from Acros Organics (Geel, Belgium). EPA (98%) and
DHA (ꢃ95%) were obtained as ethyl esters from Pronova Biocare
(Sandefjord, Norway) and were hydrolysed to their corresponding
2
.6. 13C NMR spectroscopy
13C NMR spectroscopy was also useful to aid evaluating the
regiocontrol of the reactions, although offering lower accuracy than
1
21a,29
H NMR spectroscopy.
This relates to the carbonyl group car-
bons of each of the three types of fatty acids involved (MCFA and
longer-chain SFA, EPA and DHA) displaying two distinct resonance
peaks depending upon whether the fatty acid is located at the sn-3
position or the sn-2 position of the glyceryl backbone. The carbonyl
carbons of all four MCFA types and two longer-chain fatty acids
involved in the current work were observed to resonate at
d
173.41 ppm when located at the sn-3 position of the DAGE. The
corresponding resonance for EPA located at the sn-2 position
remained at 172.84 ppm and that for DHA at 172.37 ppm.
3
. Conclusion
free acids. Dichloromethane (dried over CaH
2
) chloroform, diethyl
Previous work was aimed at preparing libraries of MLM type
ether, benzene and tetrahydrofuran were all obtained HPLC grade
from SigmaeAldrich (Steinheim, Germany). Silica gel (Silica gel 60),
potassium hydroxide and p-toluenesulfonic acid monohydrate were
purchased from Merck (Darmstadt, Germany) and preparative TLC
structured TAG possessing pure MCFA, EPA and DHA at the pre-
determined positions. This has more recently been extended to cover
the shortest chain fatty acids as well as the longer-chain saturated
fatty acids.21 This means that synthesis of such ABA type structured
TAG possessing all even number saturated fatty acids ranging from
plates (250 mm, F-254) from Silicycle (Quebec, Canada).
C
2
eC16 has been completed for both EPA and DHA. Work towards
4.1.1. Synthesis of (S)-1-O-hexadecyl-sn-glycerol (1), (S)-1-O-octa-
decyl-sn-glycerol (2) and (S)-1-O-cis-octadec-9-enyl-sn-glycerol
(3). Chimyl (1), batyl (2) and selachyl (3) alcohols were prepared by
synthesis of asymmetrically structured TAG of the ABC type pos-
sessing different saturated fatty acids at the sn-1 and sn-3 positions
and EPA or DHA at the sn-2 position has also been initiated.
The current work presents a similar generation of a focused li-
brary of enantiopure structured EL of the ALM (alkyl-long-medium)
3
0
25
a previously published procedure by Halldorsson et al. The exis-
31
tence of all these compounds is manifested in the literature. Full
experimental details are provided in Supplementary data.
type for chimyl, batyl and selachyl alcohols possessing pure SFA (C
, C , C , C10, C12, C14 and C16) attached to the sn-3 position and
both EPA and DHA to the sn-2 position, highly efficiently and under
2
,
C
4
6
8
4.1.2. Synthesis of (R)-1-O-hexadecyl-3-acetyl-sn-glycerol (4a). To
a mixture of 1-O-hexadecyl-sn-glycerol 1 (500 mg, 1.58 mmol) and

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1827
vinyl ethanoate (170 mg, 1.97 mmol) in dichloromethane (5 ml), C.
(quintet (br), J¼7.4 Hz, 2H, OCH
(br t, J¼6.8 Hz, 6H, CH
NMR (CDCl 173.94 (C]O), 71.76 (OCH
68.88 (CH CHCH ), 65.41 (CH OCO), 34.18, 31.92, 31.85, 29.69 (3),
29.68, 29.66 (2), 29.62, 29.59, 29.58, 29.46, 29.41, 29.36, 29.25 (2),
29.13, 26.07, 24.94, 22.69, 22.66,14.11,14.10 ppm; HRMS (APCI): m/z
2
CH
2
), 1.37e1.19 (m, 38H, CH
2
), 0.88
1
3
antarctica lipase (67 mg) was added and the resulting mixture
stirred for 3 h at room temperature. After filtration of the lipase the
reaction solution was concentrated to dryness in vacuo and the
3
in decanoic acid and 1-O-alkyl) ppm;
C
3
)
d
2
CH ), 71.37 (CHCH O),
2
2
2
2
2
ꢀ
residue introduced to crystallization in petroleum ether at ꢁ18 C,
2
0
affording the product as white crystals (538 mg, 95% yield); [
a
]
D
ꢀ
ꢁ
3.7 (c 1.10, benzene); mp 49.5e50.0 C; IR (KBr) 3404 (OeH), 2917
58 4
calcd for C29H O /OH: 453.4302; found 453.4300 amu.
ꢁ
1 1
(
CeH), 1702 (C]O), 1123 (CeOeC ether) cm ; H NMR (400 MHz,
CDCl OCO), 4.11 (dd, J¼11.5,
4.17 (dd, J¼11.5, 4.3 Hz, 1H, CH
.2 Hz, 1H, CH OCO), 4.03e3.97 (m, 1H, CH CHCH
), 3.50 (dd, J¼9.7,
.2 Hz, 1H, CHCH CH ),
O), 3.50e3.42 (2ꢄdt, J¼9.8, 6.2 Hz, 2H, OCH
.42 (dd, J¼9.7, 6.3 Hz, 1H, CHCH O), 2.47 (d (br), J¼4.3 Hz, 1H,
COO), 1.57 (quintet (br), J¼6.9 Hz, 2H,
), 1.36e1.21 (m, 26H, CH in
), 0.88 (br t, J¼6.9 Hz, 3H, CH
171.11 (C]O), 71.78 (OCH CH ),
), 65.67 (CH OCO), 31.92, 29.69
3), 29.67, 29.66 (2), 29.61, 29.58, 29.57, 29.46, 29.36, 26.07, 22.69,
3
)
d
2
4.1.7. Synthesis of (R)-1-O-hexadecyl-3-dodecanoyl-sn-glycerol (4f).
Full details are provided in Supplementary data. The product 4f was
6
4
3
2
2
2
2
0
2
2
2
afforded as white crystals (95% yield); [
a
]
D
ꢁ3.7 (c 1.01, benzene);
ꢀ
2
mp 57.0e57.5 C; HRMS (APCI): m/z calcd for
C
31
H
62
O
4
/OH:
CHOH), 2.10 (s, 3H, CH
OCH CH
-O-alkyl) ppm; C NMR (CDCl
1.33 (CHCH O), 68.80 (CH CHCH
3
481.4615; found 481.4594 amu.
2
2
2
d
3
13
1
7
3
)
2
2
4.1.8. Synthesis of (R)-1-O-hexadecyl-3-tetradecanoyl-sn-glycerol
(4g). A procedure identical to the one described above for 4a was
followed using 1-O-hexadecyl-sn-glycerol 1 (500 mg, 1.58 mmol),
vinyl tetradecanoate (501 mg, 1.97 mmol), dichloromethane
(10 ml) and C. antarctica lipase (100 mg). The product was afforded
2
2
2
2
(
2
42 4
0.87, 14.11 ppm; HRMS (APCI): m/z calcd for C21H O /OH:
3
41.3050; found 341.3050 amu.
2
0
as white crystals (766 mg, 92% yield); [
a]
D
ꢁ4.0 (c 0.63, benzene);
ꢀ
4.1.3. Synthesisof(R)-1-O-hexadecyl-3-butanoyl-sn-glycerol(4b). Full
mp 62.6e63.1 C; IR (KBr) 3391 (OeH), 2917 (CeH), 1730 (C]O),
ꢁ
1 1
details are provided in Supplementary data. The product 4b was
1135e1113 (CeOeC ether) cm ; H NMR (400 MHz, CDCl
(dd, J¼11.5, 4.4 Hz, 1H, CH OCO), 4.12 (dd, J¼11.5, 6.1 Hz, 1H,
CH OCO), 4.03e3.96 (m, 1H, CH CHCH
), 3.49 (dd, J¼9.7, 4.3 Hz, 1H,
CHCH CH ), 3.42 (dd,
O), 3.50e3.42 (2ꢄdt, J¼9.3, 6.2 Hz, 2H, OCH
J¼9.7, 6.3 Hz, 1H, CHCH O), 2.48 (d (br), J¼4.6 Hz, 1H, CHOH), 2.34
(t, J¼7.6 Hz, 2H, CH COO), 1.63 (quintet (br), J¼7.5 Hz, 2H,
CH CH CH ), 1.35e1.18
COO), 1.57 (quintet (br), J¼7.3 Hz, 2H, OCH
(m, 46H, CH in tetradecanoic acid and
), 0.88 (br t, J¼6.9 Hz, 6H, CH
1-O-alkyl) ppm; C NMR (CDCl 173.94 (C]O), 71.76 (OCH CH ),
71.36 (CHCH O), 68.88 (CH CHCH ), 65.41 (CH OCO), 34.18, 31.92
3
) d 4.17
2
0
afforded as white crystals (92% yield); [
a
]
D
ꢁ3.5 (c 0.92, benzene);
2
ꢀ
mp 31.7e32.2 C; HRMS (APCI): m/z calcd for
C
23
H
46
O
4
/OH:
2
2
2
3
69.3363; found 369.3379 amu.
2
2
2
2
4.1.4. Synthesis of (R)-1-O-hexadecyl-3-hexanoyl-sn-glycerol (4c). A
2
procedure identical to the one described above for 4a was followed
using 1-O-hexadecyl-sn-glycerol 1 (500 mg, 1.58 mmol), vinyl
hexanoate (280 mg, 1.97 mmol), dichloromethane (5 ml) and C.
antarctica lipase (78 mg). The product was afforded as white crys-
2
2
2
2
2
3
13
3
)
d
2
2
2
2
2
2
2
0
tals (593 mg, 91% yield); [
a
]
D
ꢁ3.8 (c 1.08, benzene); mp
(2), 29.69 (3), 29.67 (2), 29.66, 29.64 (2), 29.62, 29.60 (3), 29.58,
29.46 (2), 29.35 (2), 29.26, 29.13, 26.07, 24.94, 22.69 (2), 14.11 (2)
ppm; HRMS (APCI): m/z calcd for C35H O /OH: 509.4928; found
66 4
ꢀ
3
3.6e33.9 C; IR (KBr) 3439 (OeH), 2917 (CeH), 1731 and 1717 (C]
ꢁ1 1
O), 1132 (CeOeC ether) cm ; H NMR (400 MHz, CDCl
3
)
d
4.17 (dd,
OCO), 4.12 (dd, J¼11.5, 6.1 Hz, 1H, CH OCO),
CHCH O),
), 3.49 (dd, J¼9.6, 4.3 Hz, 1H, CHCH
.50e3.42 (2ꢄdt, J¼9.3, 6.1 Hz, 2H, OCH CH ), 3.42 (dd, J¼9.6,
.3 Hz, 1H, CHCH
O), 2.47 (d (br), J¼4.6 Hz, 1H, CHOH), 2.34 (t,
COO), 1.64 (quintet, J¼7.5 Hz, 2H, CH CH COO),
CH ), 1.38e1.20 (m, 30H, CH ),
in hexanoic acid), 0.88 (br t, J¼6.9 Hz,
J¼11.5, 4.4 Hz, 1H, CH
2
2
509.4919 amu.
4
3
6
.03e3.96 (m, 1H, CH
2
2
2
2
2
4.1.9. Synthesis of (R)-1-O-hexadecyl-3-hexadecanoyl-sn-glycerol
2
(4h). Full details are provided in Supplementary data. The product
2
0
J¼7.6 Hz, 2H, CH
2
2
2
4h was afforded as white crystals (91% yield); [
a]
D
ꢁ2.5 (c 0.91,
ꢀ
1
0
3
.57 (quintet (br), J¼6.9 Hz, 2H, OCH
2
2
2
benzene); mp 69.0e69.3 C; HRMS (APCI): m/z calcd for C35
H
70
O
4
/
.90 (br t, J¼6.9 Hz, 3H, CH
3
OH: 537.5241; found 537.5238 amu.
1
3
H, CH
3
in 1-O-alkyl) ppm; C NMR (CDCl
), 71.36 (CHCH O), 68.87 (CH CHCH
3
)
d
173.93 (C]O), 71.76
), 65.41 (CH OCO),
(
OCH
2
CH
2
2
2
2
2
4.1.10. Synthesis of (R)-1-O-octadecyl-3-acetyl-sn-glycerol (5a). Full
details are provided in Supplementary data. The product 5a was
3
2
4.14, 31.92, 31.28, 29.69 (3), 29.67, 29.65 (2), 29.61, 29.58, 29.57,
9.46, 29.35, 26.07, 24.61, 22.68, 22.30, 14.11, 13.89 ppm; HRMS
2
0
afforded as white crystals (91% yield); [
a
]
D
ꢁ3.6 (c 1.05, benzene);
ꢀ
(
ESI): m/z calcd for C25
H
50
O
4
/OH: 432.4047; found 432.4068 amu.
mp 56.6e57.0 C; HRMS (APCI): m/z calcd for C23
H
46
O
4
/OH:
369.3363; found 369.3376 amu.
4.1.5. Synthesis of (R)-1-O-hexadecyl-3-octanoyl-sn-glycerol (4d). Full
details are provided in Supplementary data. The product 4d was
4.1.11. Synthesis of (R)-1-O-octadecyl-3-butanoyl-sn-glycerol (5b). A
procedure identical to the one described above for 4a was followed
using 1-O-octadecyl-sn-glycerol 2 (500 mg, 1.45 mmol), vinyl
2
0
afforded as white crystals (90% yield); [
a]
D
ꢁ3.3 (c 0.92, benzene);
ꢀ
mp 38.5e39.0 C; HRMS (APCI): m/z calcd for C27
54 4
H O /OH:
4
25.3989; found 425.4005 amu.
butanoate (207 mg, 1.81 mmol), dichloromethan (5 ml) and C.
antarctica lipase (70 mg). The product was afforded as white crys-
2
0
4.1.6. Synthesis of (R)-1-O-hexadecyl-3-decanoyl-sn-glycerol (4e). A
tals (542 mg, 90% yield); [
a
]
D
ꢁ3.2 (c 0.99, benzene); mp
ꢀ
procedure identical to the one described above for 4a was followed
using 1-O-hexadecyl-sn-glycerol 1 (500 mg, 1.58 mmol), vinyl
decanoate (391 mg, 1.97 mmol), dichloromethane (5 ml) and C.
antarctica lipase (89 mg). The product was afforded as white crys-
tals (684 mg, 92% yield); [
4
40.2e40.7 C; IR (KBr) 3445 (OeH), 2917 (CeH), 1735 and 1718 (C]
ꢁ1 1
O), 1132 (CeOeC ether) cm ; H NMR (400 MHz, CDCl
J¼11.6, 4.4 Hz, 1H, CH OCO), 4.12 (dd, J¼11.6, 6.1 Hz, 1H, CH
4.03e3.97 (m, 1H, CH CHCH
), 3.49 (dd, J¼9.6, 4.3 Hz, 1H, CHCH
3.50e3.42 (2ꢄdt, J¼9.3, 6.1 Hz, 2H, OCH CH ), 3.42 (dd, J¼9.6,
6.3 Hz, 1H, CHCH
O), 2.46 (d, J¼4.8 Hz, 1H, CHOH), 2.33 (t, J¼7.5 Hz,
2H, CH CH COO) 1.57 (quintet
COO), 1.67 (sextet, J¼7.4 Hz, 2H, CH
(br), J¼6.9 Hz, 2H, OCH CH ), 1.36e1.21 (m, 30H, CH ), 0.96 (t,
J¼7.3 Hz, 3H in butanoic acid), 0.88 (br t, J¼6.9 Hz, 3H, CH in 1-O-
173.75 (C]O), 71.77 (OCH CH ),
CHCH ), 65.40 (CH OCO), 36.04, 31.93,
29.70 (6), 29.67 (2), 29.62, 29.59, 29.58, 29.46, 29.36, 26.08, 22.69,
3
)
d
4.18 (dd,
OCO),
O),
2
2
2
2
2
2
0
a
]
D
ꢁ3.5 (c 1.13, benzene); mp
2
2
ꢀ
4.8e45.5 C; IR (KBr) 3391 (OeH), 2918 (CeH), 1730 (C]O), 1128
2
ꢁ
1 1
(
CeOeC ether) cm ; H NMR (400 MHz, CDCl
.4 Hz, 1H, CH
3
)
d
4.17 (dd, J¼11.5,
OCO), 4.12 (dd, J¼11.5, 6.1 Hz, 1H, CH OCO),
.03e3.96 (m, 1H, CH CHCH O),
), 3.49 (dd, J¼9.7, 4.3 Hz, 1H, CHCH
.50e3.42 (2ꢄdt, J¼9.3, 6.2 Hz, 2H, OCH CH ), 3.42 (dd, J¼9.7,
.3 Hz, 1H, CHCH
O), 2.47 (d, J¼4.7 Hz, 1H, CHOH), 2.34 (t, J¼7.6 Hz,
H, CH CH COO) 1.57
COO), 1.63 (quintet (br), J¼7.6 Hz, 2H, CH
2
2
2
4
4
3
6
2
2
2
2
2
2
2
2
2
3
1
3
2
2
alkyl) ppm; C NMR (CDCl
71.36 (CHCH O), 68.89 (CH
3
)
d
2
2
2
2
2
2
2
2
2
2

1828
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
ꢀ
18.42, 14.12, 13.65 ppm; HRMS (APCI): m/z calcd for C25
50
H O
4
/OH:
74 4
benzene); mp 69.0e69.3 C; HRMS (APCI): m/z calcd for C37H O /
3
97.3676; found 397.3680 amu.
OH: 565.5554; found 565.5549 amu.
4
.1.12. Synthesis of (R)-1-O-octadecyl-3-hexanoyl-sn-glycerol (5c).
4.1.18. Synthesis of (R)-1-O-cis-octadec-9-enyl-3-acetyl-sn-glycerol
(6a). To a mixture of (S)-1-O-cis-octadec-9-enyl-sn-glycerol 3
(350 mg, 1.02 mmol), vinyl ethanoate (110 mg, 1.28 mmol) in dry
dichloromethane (3 ml), immobilized C. antarctica lipase (46 mg)
was added and the reaction mixture stirred for 3 h at room tem-
perature. After filtration of the lipase the volatile components were
first removed on a rotary evaporator and then on an oil vacuum
pump system at 10 mm Hg. The purified residue was obtained as
clear oil that, if needed, could be further purified by passing it
through a short silica gel column using petroleum ether/ethyl ac-
Full details are provided in Supplementary data. The product 5c was
afforded as white crystals (91% yield); [
mp 41.2e41.5 C; HRMS (APCI): m/z calcd for
4
2
0
a]
D
ꢁ4.3 (c 1.04, benzene);
ꢀ
27 54 4
C H O
/OH:
25.3989; found 425.3993 amu.
4
.1.13. Synthesis of (R)-1-O-octadecyl-3-octanoyl-sn-glycerol (5d). A
ꢁ
2
procedure identical to the one described above for 4a was followed
using 1-O-octadecyl-sn-glycerol (500 mg, 1.45 mmol), vinyl
octanoate (309 mg, 1.81 mmol), dichloromethane (5 ml) and C.
antarctica lipase (81 mg). The product was afforded as white crys-
tals (642 mg, 94% yield); [
4
2
2
0
etate (80:20) as eluent (375 mg, 96% yield); [
a]
D
ꢁ3.5 (c 1.33,
2
0
a
]
D
ꢁ3.4 (c 0.94, benzene); mp
benzene); IR (ZnSe) 3457 (OeH), 3003 (]CeH cis), 2922 (CeH),
ꢀ
ꢁ1
1
5.2e45.7 C; IR (KBr) 3444 (OeH), 2918 (CeH), 1731 and 1717 (C]
1743 (C]O), 1655 (C]C), 1117 (CeOeC ether) cm
(400 MHz, CDCl
5.39e5.31 (m, 2H, ]CH) 4.18 (dd, J¼11.6, 4.3 Hz,
1H, CH OCO), 4.03e3.97 (m,
OCO), 4.12 (dd, J¼11.6, 6.3 Hz, 1H, CH
1H, CH CHCH O), 3.50e3.42
), 3.50 (dd, J¼9.6, 4.3 Hz, 1H, CHCH
(2ꢄdt, J¼9.7, 6.2 Hz, 2H, OCH CH ), 3.42 (dd, J¼9.6, 6.3 Hz, 1H,
CHCH COO),
O), 2.46 (d (br), J¼4.5 Hz, 1H, CHOH), 2.10 (s, 3H, CH
2.01 (quartet (br), J¼6.4 Hz, 4H, ]CHCH ), 1.57 (quintet (br),
J¼6.9 Hz, 2H, OCH CH ), 1.38e1.22 (m, 22H, CH ), 0.88 (br t,
J¼6.8 Hz, 3H, CH in 1-O-alkyl) ppm; C NMR (CDCl 171.10 (C]
O), 129.95, 129.81, 71.76 (OCH CH ), 71.33 (CHCH O), 68.80
(CH CHCH ), 65.67 (CH OCO), 31.90, 29.77, 29.75, 29.57, 29.52,
29.48, 29.43, 29.32 (2), 29.25, 27.21, 27.19, 26.06, 22.68, 20.87,
; H NMR
ꢁ
1 1
O), 1133 (CeOeC ether) cm ; H NMR (400 MHz, CDCl
3
)
d
4.18 (dd,
OCO), 4.12 (dd, J¼11.5, 6.2 Hz, 1H, CH OCO),
CHCH O),
), 3.49 (dd, J¼9.6, 4.3 Hz, 1H, CHCH
.50e3.42 (2ꢄdt, J¼9.3, 5.9 Hz, 2H, OCH CH ), 3.42 (dd, J¼9.6,
.4 Hz, 1H, CHCH
O), 2.46 (d, J¼4.8 Hz, 1H, CHOH), 2.34 (t, J¼7.6 Hz,
H, CH CH COO) 1.57
COO), 1.63 (quintet (br), J¼7.4 Hz, 2H, CH
CH ), 1.37e1.19 (m, 38H, CH ), 0.88
in octanoic acid and 1-O-alkyl) ppm;
173.94 (C]O), 71.77 (OCH CH ), 71.36 (CHCH
CHCH ), 65.41 (CH OCO), 34.18, 31.92, 31.65, 29.70 (6),
3
) d
J¼11.5, 4.4 Hz, 1H, CH
2
2
2
2
4
3
6
2
.03e3.96 (m, 1H, CH
2
2
2
2
2
2
2
2
2
2
2
2
3
2
2
2
2
(
quintet (br), J¼7.0 Hz, 2H, OCH
2
2
2
2
2
2
13
13
(
br t, J¼6.9 Hz, 6H, CH
3
C
3
3
) d
NMR (CDCl
3
)
d
2
2
2
O),
2
2
2
6
2
2
C
8.88 (CH
2
2
2
2
2
2
9.66 (2), 29.62, 29.59, 29.58, 29.47, 29.36, 29.08, 28.91, 26.08,
4.93, 22.69, 22.59, 14.11, 14.05 ppm; HRMS (APCI): m/z calcd for
44 4
14.11 ppm; HRMS (APCI): m/z calcd for C23H O /OH: 367.3207;
found 367.3192 amu.
H
29 58
O
4
/OH: 453.4302; found 453.4322 amu.
4.1.14. Synthesis of (R)-1-O-octadecyl-3-decanoyl-sn-glycerol (5e).
4.1.19. Synthesis of (R)-1-O-cis-octadec-9-enyl-3-butanoyl-sn-glyc-
Full details are provided in Supplementary data. The product 5e
erol (6b). Full details are provided in Supplementary data. The
2
0
20
was afforded as white crystals (95% yield); [
zene); mp 50.8e51.5 C; HRMS (APCI): m/z calcd for C31
a
]
D
ꢁ3.5 (c 1.12, ben-
/OH:
product 6b was obtained as clear oil (95% yield); [
a
]
D
ꢁ3.6 (c 1.34,
ꢀ
H
62
O
4
benzene); HRMS (APCI): m/z calcd for C25
H
48
O
4
þH: 413.3625;
4
81.4615; found 481.4604 amu.
found: 413.3637 amu.
4
.1.15. Synthesis of (R)-1-O-octadecyl-3-dodecanoyl-sn-glycerol (5f).
4.1.20. Synthesis of (R)-1-O-cis-octadec-9-enyl-3-hexanoyl-sn-glyc-
erol (6c). A procedure identical to the one described above for 6awas
followed using (S)-1-O-cis-octadec-9-enyl-sn-glycerol 3 (350 mg,
1.02 mmol), vinyl hexanoate (181 mg, 1.28 mmol), dry dichloro-
methane (3 ml) and immobilized C. antarctica lipase (53 mg). The
volatile components were first removed on a rotary evaporator and
A procedure identical to the one described above for 4a was followed
using 1-O-octadecyl-sn-glycerol (500 mg, 1.45 mmol), vinyl
dodecanoate (410 mg, 1.81 mmol), dichloromethan (5 ml) and C.
antarctica lipase (91 mg). The product was afforded as white crystals
2
2
0
ꢀ
(
688 mg, 90% yield); [
a]
D
ꢁ3.0 (c 0.94, benzene); mp 55.1e55.6 C;
ꢀ
ꢁ2
IR(KBr) 3444 (OeH), 2918 (CeH), 1730 and 1717 (C]O), 1133
then on a Kugelrohr apparatus (70 C, 10 mm Hg). The purified
residue was obtained as clear oil that, if needed, could be further
purified by passing it through a short silica gel column using petro-
ꢁ
1 1
(
4
(
(
CeOeC ether) cm ; H NMR (400 MHz, CDCl
.4 Hz,1H, CH
OCO), 4.12 (dd, J¼11.5, 6.1 Hz,1H, CH
m, 1H, CH CHCH
2ꢄdt, J¼9.3, 6.1 Hz, 2H, OCH
O), 2.47 (d, J¼4.7 Hz, 1H, CHOH), 2.34 (t, J¼7.6 Hz, 2H,
COO), 1.62 (quintet (br), J¼7.5 Hz, 2H, CH CH COO) 1.57 (quintet
br), J¼7.3 Hz, 2H, OCH CH ), 1.37e1.19 (m, 46H, CH ), 0.88 (br t,
J¼6.9 Hz, 6H, CH in dodecanoic acid and 1-O-alkyl) ppm; C NMR
100 MHz, CDCl 173.94 (C]O), 71.77 (OCH CH ), 71.36 (CHCH O),
), 65.41 (CH OCO), 34.18, 31.92, 31.91, 29.70 (6),
3
)
d
4.17 (dd, J¼11.5,
2
2
OCO), 4.03e3.96
2
0
2
2
), 3.49 (dd, J¼9.7, 4.3 Hz, 1H, CHCH
2
O), 3.50e3.42
), 3.42 (dd, J¼9.7, 6.3 Hz, 1H,
leum ether/ethyl acetate (80:20) as eluent (418 mg, 93% yield); [
a]
D
2
CH
2
ꢁ3.6 (c 1.32, benzene); IR (ZnSe) 3446 (OeH), 3004 (]CeH cis), 2923
ꢁ1 1
CHCH
CH
(
2
(CeH), 1739 (C]O), 1655 (C]C), 1111 (CeOeC ether) cm ; H NMR
(400 MHz, CDCl
5.39e5.31 (m, 2H, ]CH), 4.18 (dd, J¼11.5, 4.4 Hz,
1H, CH OCO), 4.02e3.97 (m,1H,
OCO), 4.12 (dd, J¼11.5, 6.2 Hz,1H, CH
CH CHCH O), 3.49e3.40 (2ꢄdt,
), 3.49 (dd, J¼9.6, 4.2 Hz, 1H, CHCH
J¼9.3, 6.1 Hz, 2H, OCH CH ), 3.42 (dd, J¼9.6, 6.3 Hz, 1H, CHCH O),
2.53e2.45 (s (br), 1H, CHOH), 2.34 (t, J¼7.6 Hz, 2H, CH COO), 2.01
(quartet (br), J¼6.4 Hz, 4H, ]CHCH ), 1.64 (quintet (br), J¼7.5 Hz, 2H,
CH CH CH ), 1.38e1.21
COO), 1.57 (quintet (br), J¼6.8 Hz, 2H, OCH
(m, 26H, CH inhexanoicacid), 0.88(br t,
), 0.90(br t, J¼6.9 Hz, 3H, CH
J¼6.9 Hz, 3H, CH
173.93 (C]O), 129.94, 129.80, 71.75 (OCH
68.87 (CH CHCH ), 65.41 (CH OCO), 34.13, 31.90, 31.28, 29.76, 29.75,
29.57, 29.52, 29.48, 29.43, 29.31 (2), 29.25, 27.20, 27.19, 26.06, 24.61,
2
2
2
3
) d
2
2
2
2
2
13
3
2
2
2
(
3
)
d
2
2
2
2
2
2
6
2
2
C
8.88 (CH
2
CHCH
2
2
2
9.66 (2), 29.62, 29.60 (3), 29.58, 29.46 (2), 29.36, 29.33, 29.26, 29.14,
6.08, 24.94, 22.69 (2), 14.11 (2) ppm; HRMS (APCI): m/z calcd for
2
2
2
2
2
33 66 4
H O /OH: 509.4928; found 509.4916 amu.
2
3
13
3
in 1-O-alkyl) ppm; C NMR (100 MHz, CDCl
3
)
4.1.16. Synthesis of (R)-1-O-octadecyl-3-tetradecanoyl-sn-glycerol
d
2
CH ), 71.37 (CHCH O),
2
2
(5g). Full details are provided in Supplementary data. The product
2
2
2
2
0
5
g was afforded as white crystals (91% yield); [
a
]
D
ꢁ2.5 (c 0.67,
ꢀ
benzene); mp 64.2e64.8 C; HRMS (APCI): m/z calcd for C35
H
70
O
4
/
52 4
22.67, 22.30, 14.10, 13.89 ppm; HRMS (APCI): m/z calcd for C27H O /
OH: 537.5241; found 537.5249 amu.
OH: 423.3833; found: 423.3842 amu.
4
.1.17. Synthesis of (R)-1-O-octadecyl-3-hexadecanoyl-sn-glycerol
4.1.21. Synthesis of (R)-1-O-cis-octadec-9-enyl-3-octanoyl-sn-glyc-
erol (6d). To a mixture of (S)-1-O-cis-octadec-9-enyl-sn-glycerol 3
(350 mg, 1.02 mmol), vinyl octanoate (218 mg, 1.28 mmol) in dry
(5h). Full details are provided in Supplementary data. The product
2
0
5
h was afforded as white crystals (90% yield); [
a]
D
ꢁ2.3 (c 0.26,

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1829
ꢀ
ꢀ
dichloromethane (3 ml), immobilized C. antarctica lipase (57 mg) was
added and the reaction mixture stirred for 3 h at room temperature.
After filtration of the lipase, the reaction solution was concentrated
crystallization in petroleum ether at ꢁ18 C (564 mg, 95% yield);
2
0
[a
]
D
ꢁ2.3 (c 1.05, benzene); mp 37.8e38.3 C; IR(KBr) 3404 (OeH),
2998 (]CeH cis), 2918 (CeH), 1742 (C]O), 1660 (C]C), 1119
ꢀ
ꢁ1 1
and introduced to crystallization in petroleum ether at ꢁ40 C (ace-
(CeOeC ether) cm ; H NMR (400 MHz, CDCl
]CH), 4.17 (dd, J¼11.5, 4.4 Hz, 1H, CH OCO), 4.12 (dd, J¼11.5, 6.2 Hz,
1H, CH OCO), 4.02e3.97 (m, 1H, CH CHCH
), 3.49 (dd, J¼9.8, 4.3 Hz,
1H, CHCH CH ), 3.42
O), 3.50e3.42 (2ꢄdt, J¼9.2, 6.1 Hz, 2H, OCH
(dd, J¼9.8, 6.3 Hz,1H, CHCH O), 2.52e2.43 (s (br),1H, CHOH), 2.34 (t,
J¼7.6 Hz, 2H, CH COO), 2.01 (quartet (br), J¼6.3 Hz, 4H, ]CHCH ),
1.62 (quintet (br), J¼7.3 Hz, 2H, CH CH COO), 1.57 (quintet (br),
J¼7.1 Hz, 2H, OCH CH ), 1.38e1.18 (m, 46H, CH ), 0.88 (t, J¼6.9 Hz,
) ppm; C NMR (100 MHz, CDCl 173.95 (C]O), 129.94,
129.80, 71.75 (OCH CH ), 71.38 (CHCH O), 68.88 (CH CHCH ), 65.41
(CH OCO), 34.18, 31.92, 31.91, 29.77, 29.76, 29.69 (2), 29.68, 29.65 (2),
3
) d 5.39e5.31 (m, 2H,
tonitrilecoolingbath)affordingtheproductaswhitesolidthatmelted
2
2
0
into clear oil at room temperature (430 mg, 90% yield); [
a
]
D
ꢁ3.3 (c
2
2
2
1
.16, benzene); IR (ZnSe) 3454 (OeH), 3004 (]CeH cis), 2923 (CeH),
2
2
2
ꢁ1
1
1740 (C]O), 1655 (C]C), 1113 (CeOeC ether) cm
;
H NMR
2
(
400 MHz, CDCl
3
)
d
5.39e5.31 (m, 2H, ]CH), 4.17 (dd, J¼11.6, 4.4 Hz,
OCO), 4.12 (dd, J¼11.6, 6.0 Hz,1H, CH OCO), 4.03e3.95(m(br),
CHCH O), 3.50e3.41
), 3.49 (dd, J¼9.6, 4.3 Hz, 1H, CHCH
CH
), 3.42 (dd, J¼9.6, 6.3 Hz, 1H,
O), 2.47 (d (br), J¼3.8 Hz, 1H, CHOH), 2.34 (t, J¼7.6 Hz, 2H,
COO), 2.01 (quartet (br), J¼6.4 Hz, 4H, ]CHCH ), 1.63 (quintet
br), J¼7.4 Hz, 2H, CH CH COO), 1.57 (quintet (br), J¼7.0 Hz, 2H,
OCH CH ), 1.38e1.20 (m, 30H, CH in
), 0.88 (br t, J¼6.8 Hz, 6H, CH
octanoic acid and 1-O-alkyl) ppm; C NMR (CDCl 173.94 (C]O),
29.94,129.80, 71.75 (OCH CH ), 71.38 (CHCH O), 68.88 (CH CHCH ),
5.41 (CH OCO), 34.18, 31.90, 31.65, 29.77, 29.75, 29.58, 29.52, 29.48,
9.43, 29.32 (2), 29.25, 29.08, 28.91, 27.21, 27.20, 26.07, 24.93, 22.68,
2
2
1
1
H, CH
H, CH
2
2
2
2
2
2
2
2
2
2
13
(
2ꢄdt, J¼9.3, 6.1 Hz, 2H, OCH
CHCH
CH
2
2
6H, CH
3
3
) d
2
2
2
2
2
2
2
2
2
(
2
2
29.60, 29.58, 29.52, 29.49, 29.47, 29.44, 29.36, 29.32 (2), 29.26 (2),
29.14, 27.22, 27.20, 26.07, 24.94, 22.69 (2), 14.11 (2) ppm; HRMS
(APCI): m/z calcd for C37H O /OH: 563.5398; found: 563.5396 amu.
72 4
2
2
2
3
13
3
)
d
1
2
2
2
2
2
6
2
2
4
2
4.1.26. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-acetyl-
sn-glycerol (7a). To a solution of (R)-1-O-hexadecyl-3-acetyl-sn-
glycerol 4a (100 mg, 0.279 mmol), DMAP (27 mg, 0.223 mmol) and
EDAC (75 mg, 0.391 mmol) in dry dichloromethane (2.5 ml) was
added EPA as a free acid (84 mg, 0.279 mmol) and the resulting
solution stirred at room temperature for 15 h under nitrogen at-
mosphere. The reaction mixture was passed through a short silica
gel column with ether/dichloromethane (10:90) as eluent, afford-
56 4
2.59, 14.11, 14.05 ppm; HRMS (APCI): m/z calcd for C29H O /OH:
51.4146; found: 451.4153 amu.
0
4
.1.22. Synthesis of (R)-1-O-cis-octadec-9 -enyl-3-decanoyl-sn-glyc-
erol (6e). Full details are provided in Supplementary data. The
product 6e was afforded as white solid that melted into clear oil at
ꢀ
room temperature after crystallization in petroleum ether at ꢁ18 C
ing the product as yellowish oil (161 mg, 90% yield) after concen-
2
0
20
(
90% yield); [
a]
D
ꢁ2.8 (c 1.35, benzene); HRMS (APCI): m/z calcd for
tration in vacuo; [
a
]
D
ꢁ5.0 (c 1.43, chloroform); IR (ZnSe) 3012
ꢁ
1 1
C
31
60
H O
4
/OH: 479.4459; found: 479.4441 amu.
(]CeH cis), 2922 (CeH), 1743 (C]O), 1653 (C]C) cm ; H NMR
400 MHz, CDCl 5.44e5.28 (m, 10H, ]CH), 5.22e5.17 (m, 1H,
CH CHCH OCO), 4.15 (dd, J¼11.9,
), 4.33 (dd, J¼11.9, 3.6 Hz, 1H, CH
6.5 Hz, 1H, CH
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H,
CHCH CH ), 2.88e2.77
O), 3.47e3.37 (2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH
(m, 8H, ]CCH COO), 2.14e2.04 (m,
C]), 2.34 (t, J¼7.6 Hz, 2H, CH
4H, ]CCH CH and ]CCH CH ), 2.05 (s, 3H, CH COO),1.71 (quintet,
J¼7.5 Hz, 2H, CH CH COO), 1.53 (quintet (br), J¼6.8 Hz, 2H,
OCH CH ), 1.35e1.20 (m, 26H, CH in
EPA), 0.88 (br t, J¼6.9 Hz, 3H, CH
(CDCl 172.87 ( , C]O), 170.62 (
(
3
) d
4
.1.23. Synthesis of (R)-1-O-cis-octadec-9-enyl-3-dodecanoyl-sn-
glycerol (6f). A procedure identical to the one described above for
d was followed using (S)-1-O-cis-octadec-9-enyl-sn-glycerol 3
350 mg, 1.02 mmol), vinyl dodecanoate (290 mg, 1.28 mmol), dry
dichloromethane (3 ml) and immobilized C. antarctica lipase
64 mg). The product was afforded as a white solid after crystalli-
2
2
2
2
6
(
2
2
2
2
2
2
2
2
3
3
(
2
2
ꢀ
20
zation in petroleum ether at ꢁ18 C (481 mg, 90% yield); [
a]
D
ꢁ3.1
2
2
2
), 0.97 (t, J¼7.5 Hz, 3H, CH
3
ꢀ
13
(
(
c 0.91, benzene); mp 28.7e29.1 C; IR (KBr) 3511 (OeH), 3004
]CeH cis), 2920 (CeH), 1725 (C]O), 1654 (C]C), 1123 (CeOeC
3
in 1-O-alkyl) ppm; C NMR
a, C]O), 132.01, 128.88, 128.85,
3
)
d
b
ꢁ1 1
ether) cm ; H NMR (400 MHz, CDCl
.17 (dd, J¼11.5, 4.4 Hz, 1H, CH OCO), 4.12 (dd, J¼11.5, 6.0 Hz, 1H,
CH OCO), 4.03e3.96 (m, 1H, CH CHCH
), 3.49 (dd, J¼9.6, 4.3 Hz, 1H,
CHCH CH ), 3.42 (dd,
O), 3.50e3.42 (2ꢄdt, J¼9.2, 6.1 Hz, 2H, OCH
J¼9.6, 6.5 Hz, 1H, CHCH O), 2.46 (d (br), J¼4.5 Hz, 1H, CHOH), 2.34
t, J¼7.6 Hz, 2H, CH COO), 2.01 (quartet, J¼6.4 Hz, 4H, ]CHCH ),
.63 (quintet (br), J¼7.4 Hz, 2H, CH CH COO), 1.57 (quintet (br),
J¼7.1 Hz, 2H, OCH CH ), 1.38e1.19 (m, 38H, CH ), 0.88 (br t,
J¼6.8 Hz, 6H, CH ) ppm; C NMR (CDCl 173.95 (C]O), 129.95,
29.80, 71.75 (OCH CH ), 71.37 (CHCH O), 68.89 (CH CHCH ), 65.42
CH OCO), 34.18, 31.90 (2), 29.77, 29.75, 29.60 (2), 29.58, 29.52,
9.48, 29.46, 29.44, 29.32 (3), 29.26 (2), 29.14, 27.21, 27.20, 26.07,
4.94, 22.68 (2), 14.11 (2) ppm; HRMS (APCI): m/z calcd for
/OH: 507.4772; found: 507.4748 amu.
3
)
d
5.39e5.31 (m, 2H, ]CH),
128.55, 128.25, 128.18, 128.15, 128.06, 127.84, 126.99, 71.75
(OCH CH ), 70.10 (CH CHCH ), 68.88 (CHCH O), 63.04 (CH OCO),
33.70, 31.91, 29.69 (4), 29.67, 29.65, 29.62, 29.60, 29.53, 29.45,
29.35, 26.48, 26.02, 25.61 (2), 25.59, 25.53, 24.80, 22.68, 20.75,
4
2
2
2
2
2
2
2
2
2
2
2
2
2
2
20.54, 14.26, 14.10 ppm; HRMS (APCI): m/z calcd for C41
70
H O
5
þH:
(
1
2
2
643.5296; found 643.5285 amu.
2
2
2
2
2
4.1.27. Synthesis of (R)-1-O-hexadecyl-2-eicospentaenoyl-3-buta-
noyl-sn-glycerol (7b). Full details are provided in Supplementary
data. The product 7b was afforded as yellowish oil (90% yield);
13
3
3
) d
1
(
2
2
C
2
2
2
2
2
2
0
2
[a
]
D
ꢁ5.4 (c 1.08, chloroform); HRMS (APCI): m/z calcd for
43 74 5
C H O
þH: 671.5609; found 671.5637 amu.
H
33 64
O
4
4.1.28. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-hex-
anoyl-sn-glycerol (7c). A procedure identical to the one described
above for 7a was followed using (R)-1-O-hexadecyl-3-hexanoyl-sn-
glycerol 4c (100 mg, 0.241 mmol), DMAP (24 mg, 0.193 mmol),
EDAC (65 mg, 0.337 mmol), dry dichloromethane (2.5 ml) and EPA
as a free acid (73 mg, 0.241 mmol). The product was afforded as
4.1.24. Synthesis of (R)-1-O-cis-octadec-9-enyl-3-tetradecanoyl-sn-
glycerol (6g). Full details are provided in Supplementary data. The
2
0
product 6g was afforded as a white solid (90% yield); [
1
a]
D
ꢁ2.7 (c
ꢀ
.06, benzene); mp 32.6e33.1 C; HRMS (APCI): m/z calcd for
2
0
C
H
35 68
O
4
/OH: 535.5085; found: 535.5083 amu.
yellowish oil after concentration (153 mg, 91% yield); [
.21, chloroform); IR (ZnSe) 3013 (]CeH cis), 2923 (CeH), 1740
a]
D
ꢁ4.8 (c
1
0
1
4
.1.25. Synthesis of (R)-1-O-cis-octadec-9 -enyl-3-hexadecanoyl-sn-
(C]O), 1654 (C]C); H NMR (400 MHz, CDCl
10H, ]CH), 5.22e5.17 (m, 1H, CH CHCH
1H, CH
2
3
) d 5.43e5.28 (m,
glycerol (6h). A procedure identical to the one described above for
d was followed using (S)-1-O-cis-octadec-9-enyl-sn-glycerol 3
350 mg,1.02 mmol), vinyl hexadecanoate (360 mg,1.28 mmol), dry
dichloromethane (3 ml) and immobilized C. antarctica lipase
71 mg). The product was afforded as white solid after
2
2
), 4.33 (dd, J¼11.9, 3.7 Hz,
OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H, CH OCO), 3.57e3.50
(2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH O), 3.47e3.37 (2ꢄdt, J¼9.4, 7.5 Hz,
2H, OCH CH ), 2.88e2.76 (m, 8H, ]CCH
C]), 2.34 (t, J¼7.8 Hz, 2H,
CH COO in hexanoic acid),
COO in EPA), 2.30 (t, J¼7.4 Hz, 2H, CH
6
(
2
2
2
2
2
(
a
2
2

1830
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
2
.14e2.04 (m, 4H, ]CCH
J¼7.5 Hz, 2H, CH CH COO in EPA), 1.61 (quintet, J¼7.5 Hz, 2H,
CH CH
COO in hexanoic acid), 1.54 (quintet (br), J¼6.9 Hz, 2H,
OCH CH ), 1.37e1.18 (m, 30H, CH in
), 0.97 (t, J¼7.5 Hz, 3H, CH
EPA), 0.89 (br t, J¼6.2 Hz, 3H, CH in hexanoic acid), 0.88 (br t,
J¼6.3 Hz, 3H, CH in 1-O-alkyl) ppm; C NMR (CDCl
C]O), 172.84 ( , C]O), 132.02, 128.89, 128.84, 128.55, 128.25,
28.19, 128.15, 128.06, 127.85, 127.00, 71.75 (OCH CH ), 70.16
CH CHCH ), 68.90 (CHCH O), 62.75 (CH OCO), 34.08, 33.72, 31.92,
1.25, 29.69 (5), 29.65, 29.63, 29.61, 29.55, 29.46, 29.35, 26.50,
2
CH
2
and ]CCH
2
CH
3
), 1.70 (quintet,
2.34 (t, J¼7.6 Hz, 2H, CH
in tetradecanoic acid), 2.14e2.04 (m, 4H, ]CCH
CCH CH CH COO in EPA), 1.60
), 1.70 (quintet, J¼7.5 Hz, 2H, CH
(quintet (br), J¼7.3 Hz, 2H, CH CH COO in tetradecanoic acid), 1.54
(quintet (br), J¼6.9 Hz, 2H, OCH CH ), 1.35e1.15 (m, 46H, CH ), 0.98
(t, J¼7.5 Hz, 3H, CH in EPA), 0.88 (br t, J¼6.9 Hz, 6H, CH in tetra-
decanoic acid and 1-O-alkyl) ppm; C NMR (CDCl 173.43 ( , C]
O), 172.85 ( , C]O), 132.03, 128.90, 128.85, 128.57, 128.26, 128.20,
128.16, 128.07, 127.86, 127.00, 71.76 (OCH CH ), 70.17 (CH CHCH ),
68.91 (CHCH O), 62.74 (CH OCO), 34.14, 33.73, 31.92 (2), 29.70 (4),
2
COO in EPA), 2.30 (t, J¼7.6 Hz, 2H, CH
2
COO
CH and ]
2
2
2
2
2
2
2
3
2
2
2
2
2
3
2
2
3
2
2
2
13
3
b
3
)
d
173.41 (
a
,
3
3
13
3
)
d
a
1
(
2
2
b
2
2
2
2
2
2
2
2
3
2
2
2
6.03, 25.61 (2), 25.60, 25.53, 24.81, 24.57, 22.68, 22.30, 20.55,
þNH
29.69 (3), 29.66 (3), 29.63 (2), 29.55, 29.49, 29.47, 29.36 (2), 29.29,
29.13, 26.50, 26.04, 25.62 (2), 25.61, 25.54, 24.91, 24.82, 22.69 (2),
20.56, 14.27, 14.12 (2) ppm; HRMS (APCI): m/z calcd for
14.26, 14.11, 13.89 ppm; HRMS (APCI): m/z calcd for C45
H
78
O
5
4
:
716.6188; found 716.6185 amu.
C
53
94
H O
5
þNH
4
: 828.7440; found 828.7459 amu.
4.1.29. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-octa-
noyl-sn-glycerol (7d). Full details are provided in Supplementary
data. The product 7d was afforded as yellowish oil (90% yield);
4.1.33. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-hex-
adecanoyl-sn-glycerol (7h). Full details are provided in
Supplementary data. The product 7h was afforded as yellowish oil
2
0
[
C
a
]
D
H
ꢁ3.9 (c 1.58, chloroform); HRMS (APCI): m/z calcd for
2
0
47
82
O
5
þH: 727.6235; found 727.6250 amu.
(91% yield); [
a]
D
ꢁ4.5 (c 1.04, chloroform); HRMS (APCI): m/z calcd
for C55H
98
O
5
þNH
4
: 856.7753; found 856.7784 amu.
4.1.30. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-dec-
anoyl-sn-glycerol (7e). A procedure identical to the one described
above for 7a was followed using (R)-1-O-hexadecyl-3-decanoyl-sn-
glycerol 4e (100 mg, 0.212 mmol), DMAP (21 mg, 0.170 mmol), EDAC
4.1.34. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-acetyl-
sn-glycerol (8a). Full details are provided in Supplementary data.
The product 8a was afforded as yellowish semisolid oil (90% yield);
2
0
(
57 mg, 0.297 mmol), dry dichloromethane (2.5 ml) and EPA as
[
C
a
]
D
ꢁ6.1 (c 0.85, chloroform); HRMS (APCI): m/z calcd for
a free acid (64 mg, 0.212 mmol). The product was afforded as yel-
43
H
74
O
5
þNH : 688.5875; found 688.5857 amu.
4
2
0
lowish oil after concentration (145 mg, 90% yield); [
a
]
D
ꢁ2.9 (c 1.03,
chloroform); IR (ZnSe) 3013 (]CeH cis), 2921 (CeH), 1741 (C]O),
4.1.35. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-buta-
noyl-sn-glycerol (8b). A procedure identical to the one described
above for 7a was followed using (R)-1-O-octadecyl-3-butanoyl-sn-
glycerol 5b (100 mg, 0.241 mmol), DMAP (24 mg, 0.193 mmol),
EDAC (65 mg, 0.337 mmol), dry dichloromethane (2.5 ml) and EPA
as a free acid (73 mg, 0.241). The product was afforded as yellowish
ꢁ1 1
1
]
653 (C]C) cm ; H NMR (400 MHz, CDCl
CH), 5.22e5.17 (m, 1H, CH CHCH
CH
3
) d 5.43e5.28 (m, 10H,
2
2
), 4.33 (dd, J¼11.9, 3.7 Hz, 1H,
OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H, CH OCO), 3.57e3.50 (2ꢄdd,
J¼10.6, 5.3 Hz, 2H, CHCH O), 3.47e3.37 (2ꢄdt, J¼9.4, 7.5 Hz, 2H,
OCH CH ), 2.88e2.76 (m, 8H, ]CCH
C]), 2.34 (t, J¼7.6 Hz, 2H,
CH COO in decanoic acid),
COO in EPA), 2.30 (t, J¼7.6 Hz, 2H, CH
.14e2.04 (m, 4H, ]CCH CH and ]CCH CH ), 1.70 (quintet,
J¼7.5 Hz, 2H, CH CH COO in EPA), 1.60 (quintet (br), J¼7.3 Hz, 2H,
CH CH
COO in decanoic acid), 1.53 (quintet (br), J¼6.4 Hz, 2H,
OCH CH ),1.35e1.18 (m, 38H, CH in EPA),
), 0.98 (t, J¼7.5 Hz, 3H, CH
.88 (t, J¼6.8 Hz, 6H, CH in decanoic acid and 1-O-alkyl) ppm;
NMR (CDCl 173.42 ( , C]O), 172.85 ( , C]O), 132.02, 128.89,
28.85, 128.56, 128.26, 128.19, 128.16, 128.07, 127.85, 127.00, 71.76
OCH CH ), 70.17 (CH CHCH ), 68.91 (CHCH O), 62.74 (CH OCO),
2
2
2
2
2
2
2
0
2
2
semisolid oil after concentration (153 mg, 91% yield); [
a
]
D
ꢁ5.6 (c
2
2
2
2
3
1.14, chloroform); IR(ZnSe) 3012 (]CeH cis), 2922 (CeH), 1740 (C]
ꢁ
1 1
2
2
O), 1653 (C]C) cm
10H, ]CH), 5.23e5.18 (m, 1H, CH
1H, CH
;
H NMR (400 MHz, CDCl
3
) d 5.44e5.28 (m,
2
2
2
CHCH
2
), 4.34 (dd, J¼11.9, 3.7 Hz,
OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H, CH OCO), 3.57e3.50
(2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH O), 3.47e3.38 (2ꢄdt, J¼9.4, 7.5 Hz,
2H, OCH CH ), 2.88e2.77 (m, 8H, ]CCH
C]), 2.34 (t, J¼7.6 Hz, 2H,
CH COO in butanoic acid),
COO in EPA), 2.29 (t, J¼7.4 Hz, 2H, CH
2.14e2.04 (m, 4H, ]CCH CH and ]CCH CH ), 1.71 (quintet,
J¼7.1 Hz, 2H, CH CH COO in EPA), 1.64 (sextet, J¼7.1 Hz, 2H,
CH CH
COO in butanoic acid), 1.54 (quintet (br), J¼6.8 Hz, 2H,
OCH CH ),1.35e1.20 (m, 30H, CH in EPA),
), 0.98 (t, J¼7.5 Hz, 3H, CH
0.94 (t, J¼7.4 Hz, 3H, CH in butanoic acid), 0.88 (br t, J¼6.9 Hz, 3H,
2
2
2
3
2
2
13
0
3
C
2
3
)
d
a
b
2
2
2
1
2
2
(
2
2
2
2
2
2
2
2
2
3
3
2
2
4.13, 33.73, 31.92, 31.86, 29.70 (5), 29.66, 29.64, 29.62, 29.55, 29.47,
9.43, 29.36, 29.28 (2), 29.12, 26.51, 26.04, 25.62 (2), 25.61, 25.54,
4.90, 24.82, 22.69, 22.66, 20.55, 14.27, 14.11, 14.10 ppm; HRMS
2
2
2
2
2
2
2
3
(
APCI): m/z calcd for C49
H
86
O
5
þH: 755.6548; found 755.6567 amu.
3
1
3
CH
C]O), 132.03, 128.90, 128.85, 128.57, 128.26, 128.20, 128.16, 128.07,
127.86,127.00, 71.76 (OCH CH ), 70.17 (CH CHCH ), 68.91 (CHCH O),
62.75 (CH OCO), 36.00, 33.73, 31.92, 29.70 (7), 29.66, 29.64, 29.61,
3 3
in 1-O-alkyl) ppm; C NMR (CDCl ) d 173.23 (a, C]O),172.86 (b,
4.1.31. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-dodec-
anoyl-sn-glycerol (7f). Full details are provided in Supplementary
data. The product 7f was afforded as yellowish oil (90% yield);
2
2
2
2
2
2
2
0
[
C
a
]
D
ꢁ4.1 (c 1.44, chloroform); HRMS (APCI): m/z calcd for
29.55, 29.47, 29.36, 26.51, 26.04, 25.62 (2), 25.61, 25.54, 24.82, 22.69,
20.56, 18.38, 14.27, 14.12, 13.62 ppm; HRMS (APCI): m/z calcd for
51
H
90
O
5
þH: 783.6861; found 783.6900 amu.
C
45
78
H O
5
þNH
4
: 716.6188; found 716.6201 amu.
4.1.32. Synthesis of (R)-1-O-hexadecyl-2-eicosapentaenoyl-3-tetra-
decanoyl-sn-glycerol (7g). A procedure identical to the one
described above for 7a was followed using (R)-1-O-hexadecyl-3-
tetradecanoyl-sn-glycerol 4 g (100 mg, 0.190 mmol), DMAP (19 mg,
4.1.36. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-hex-
anoyl-sn-glycerol (8c). Full details are provided in Supplementary
data. The product 8c was afforded as yellowish semisolid oil (90%
2
0
0
(
.152 mmol), EDAC (51 mg, 0.266 mmol), dry dichloromethane
2.5 ml) and EPA as a free acid (57 mg, 0.190 mmol). The product
was afforded as yellowish oil after concentration (140 mg, 91%
yield); [
a
]
D
ꢁ4.7 (c 1.16, chloroform); HRMS (APCI): m/z calcd for
C
47
H
82
O
5
þNH : 744.6501; found 744.6473 amu.
4
20
yield); [
922 (CeH), 1741 (C]O), 1654 (C]C) cm
CDCl 5.44e5.28 (m, 10H, ]CH), 5.22e5.17 (m, 1H, CH
.33 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H,
CH O), 3.47e3.37
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
2ꢄdt, J¼9.4, 7.5 Hz, 2H, OCH CH ), 2.88e2.77 (m, 8H, ]CCH C]),
a
]
D
ꢁ3.7 (c 1.13, chloroform); IR (ZnSe) 3013 (]CeH cis),
4.1.37. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-octa-
noyl-sn-glycerol (8d). A procedure identical to the one described
above for 7a was followed using (R)-1-O-octadecyl-3-octanoyl-sn-
glycerol 5d (100 mg, 0.212 mmol), DMAP (21 mg, 0.170 mmol),
EDAC (57 mg, 0.297 mmol), dry dichloromethane (2.5 ml) and EPA
as a free acid (64 mg, 0.212). The product was afforded as yellowish
ꢁ
1
1
2
;
H NMR (400 MHz,
CHCH ),
3
)
d
2
2
4
2
2
2
(
2
2
2

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1831
2
0
20
oil after concentration (147 mg, 92% yield); [
a
]
D
ꢁ5.0 (c 1.02,
after concentration (134 mg, 90% yield); [
a
]
D
ꢁ4.7 (c 0.91, chloro-
chloroform); IR (ZnSe) 3013 (]CeH cis), 2923 (CeH), 1740 (C]O),
form); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH),1742 (C]O),1655 (C]
ꢁ
1
1
ꢁ1
1
1
]
653 (C]C) cm ; H NMR (400 MHz, CDCl
CH), 5.22e5.17 (m, 1H, CH CHCH
CH
3
)
d
5.44e5.28 (m, 10H,
C) cm
5.22e5.17 (m, 1H, CH
4.15 (dd, J¼11.9, 6.4 Hz, 1H, CH
2H, CHCH
O), 3.49e3.39 (2ꢄdt, J¼9.4, 7.5 Hz, 2H, OCH
2.88e2.77 (m, 8H, ]CCH COO in EPA),
C]), 2.34 (t, J¼7.6 Hz, 2H, CH
2.30 (t, J¼7.6 Hz, 2H, CH COO in hexadecanoic acid), 2.14e2.04 (m, 4H,
]CCH CH and ]CCH CH CH COO
), 1.70 (quintet, J¼7.5 Hz, 2H, CH
in EPA), 1.60 (quintet (br) J¼7.2 Hz, 2H, CH CH COO in hexadecanoic
acid), 1.53 (quintet (br), J¼6.8 Hz, 2H, OCH CH ), 1.35e1.17 (m, 54H,
CH in EPA), 0.88 (br t, J¼6.9 Hz, 6H, CH in
), 0.97 (t, J¼7.5 Hz, 3H, CH
hexadecanoic acid and 1-O-alkyl) ppm; C NMR (CDCl 173.41 (
C]O), 172.83 ( , C]O), 132.01, 128.89, 128.84, 128.55, 128.26, 128.19,
128.15, 128.06, 127.85, 127.00, 71.75 (OCH CH ), 70.16 (CH CHCH ),
68.90 (CHCH O), 62.73 (CH OCO), 34.13, 33.72, 31.92 (2), 29.70 (8),
;
H NMR (400 MHz, CDCl
CHCH
), 4.33 (dd, J¼11.9, 3.7 Hz, 1H, CH
OCO), 3.58e3.51 (2ꢄdd, J¼10.6, 5.3 Hz,
CH ),
3
)
d
5.43e5.28 (m, 10H, ]CH),
2
2
), 4.33 (dd, J¼11.9, 3.7 Hz, 1H,
OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H, CH OCO), 3.57e3.50 (2ꢄdd,
J¼10.6, 5.3 Hz, 2H, CHCH O), 3.47e3.38 (2ꢄdt, J¼9.4, 7.5 Hz, 2H,
OCH CH ), 2.88e2.77 (m, 8H, ]CCH
C]), 2.34 (t, J¼7.6 Hz, 2H,
CH COO in octanoic acid),
COO in EPA), 2.30 (t, J¼7.6 Hz, 2H, CH
.14e2.04 (m, 4H, ]CCH CH and ]CCH CH ), 1.71 (quintet,
J¼7.5 Hz, 2H, CH CH COO in EPA), 1.61 (quintet (br), J¼7.4 Hz, 2H,
CH CH
COO in octanoic acid), 1.54 (quintet (br), J¼6.6 Hz, 2H,
OCH CH ), 1.37e1.18 (m, 38H, CH in
), 0.98 (t, J¼7.5 Hz, 3H, CH
EPA), 0.88 (br t, J¼6.9 Hz, 6H, CH in octanoic acid and 1-O-alkyl)
ppm; 13C NMR (CDCl
173.43 ( , C]O), 172.86 ( , C]O), 132.01,
28.88, 128.83, 128.54, 128.24, 128.18, 128.14, 128.05, 127.84, 126.98,
1.73 (OCH CH ), 70.13 (CH CHCH ), 68.88 (CHCH O), 62.72
CH OCO), 34.11, 33.70, 31.91, 31.64, 29.69 (7), 29.64 (2), 29.60,
9.53, 29.46, 29.35, 29.05, 28.92, 26.48, 26.02, 25.60 (3), 25.52,
4.88, 24.79, 22.68, 22.59, 20.54, 14.26, 14.11, 14.06 ppm; HRMS
þH: 755.6548; found 755.6550 amu.
2
2
2
OCO),
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
3
2
2
2
3
2
2
2
2
2
2
2
2
2
2
2
2
2
3
2
3
3
13
3
3
)
d
a,
3
)
d
a
b
b
1
2
2
2
2
7
2
2
2
2
2
2
2
(
2
29.69 (2), 29.66 (3), 29.64, 29.63, 29.62, 29.55, 29.49, 29.47, 29.36 (2),
29.29, 29.13, 26.51, 26.04, 25.62 (2), 25.60, 25.53, 24.91, 24.82, 22.69
(2), 20.55, 14.26, 14.11 (2) ppm; HRMS (APCI): m/z calcd for
2
2
(
APCI): m/z calcd for C49
86
H O
5
57 102 5
C H O þNH
4
: 884.8066; found 884.8066 amu.
4.1.38. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-dec-
4.1.42. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-3-
anoyl-sn-glycerol (8e). Full details are provided in Supplementary
data. The product 8e was afforded as yellowish oil (92% yield);
acetyl-sn-glycerol (9a). Full details are provided in Supplementary
data. The product 9a was afforded as yellowish oil (90% yield); [a]
D
20
2
0
[
a]
D
ꢁ4.5 (c 0.82, chloroform); HRMS (APCI): m/z calcd for
ꢁ5.8 (c 1.58, chloroform); HRMS (APCI): m/z calcd for C43
72 5 4
H O þNH :
C
H
51 90
O
5
þNH : 800.7127; found 800.7100 amu.
4
686.5718; found: 686.5715 amu.
4
.1.39. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-dodec-
4.1.43. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-
3-butanoyl-sn-glycerol (9b). A procedure identical to the one de-
scribed above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-butanoyl-sn-glycerol 6b (120 mg, 0.291 mmol), DMAP
(25 mg, 0.236 mmol), EDAC (78 mg, 0.407 mmol), dry dichloro-
methane (2 ml) and EPA as a free acid (88 mg, 0.291). The product
was afforded as yellowish oil after concentration (183 mg, 90%
anoyl-sn-glycerol (8f). A procedure identical to the one described
above for 7a was followed using (R)-1-O-octadecyl-3-dodecanoyl-
sn-glycerol 5f (100 mg, 0.190 mmol), DMAP (19 mg, 0.152 mmol),
EDAC (51 mg, 0.266 mmol), dry dichloromethane (2.5 ml) and EPA
as a free acid (57 mg, 0.190). The product was afforded as yellowish
2
0
oil after concentration (139 mg, 90% yield); [
roform); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH), 1741 (C]O), 1655
a
]
D
ꢁ4.3 (c 1.17, chlo-
2
0
yield). [
2924 (CeH), 1742 (C]O), 1655 (C]C) cm
CDCl 5.44e5.28 (m, 12H, ]CH), 5.23e5.18 (m, 1H, CH
4.34 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H,
CH O), 3.47e3.38
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
(2ꢄdt, J¼9.4, 7.5 Hz, 2H, OCH CH ), 2.88e2.77 (m, 8H, ]CCH C]),
2.34 (t, J¼7.6 Hz, 2H, CH COO in EPA), 2.29 (t, J¼7.4 Hz, 2H, CH COO
in butanoic acid), 2.14e2.03 (m, 4H, ]CCH CH and ]CCH CH ),
2.01 (quartet (br), J¼6.3 Hz, 4H, ]CHCH in 1-O-alkyl), 1.71 (quin-
tet, J¼7.1 Hz, 2H, CH CH COO in EPA), 1.64 (sextet, J¼7.1 Hz, 2H,
CH CH
COO in butanoic acid), 1.53 (quintet (br), J¼6.6 Hz, 2H,
OCH CH ), 1.38e1.19 (m, 22H, CH in
), 0.97 (t, J¼7.5 Hz, 3H, CH
EPA), 0.94 (t, J¼7.4 Hz, 3H, CH in butanoic acid), 0.88 (br t, J¼6.8 Hz,
a
]
D
ꢁ5.1 (c 1.61, chloroform); IR (ZnSe) 3012 (]CeH cis),
ꢁ1
1
ꢁ1
1
(
C]C) cm ; H NMR (400 MHz, CDCl
3
)
d
5.43e5.28 (m,10H, ]CH),
;
H NMR (400 MHz,
CHCH ),
5
4
5
2
2
.22e5.17 (m,1H, CH
2
CHCH
2
), 4.33 (dd, J¼11.9, 3.7 Hz,1H, CH OCO),
.15 (dd, J¼11.9, 6.4 Hz, 1H, CH OCO), 3.58e3.51 (2ꢄdd, J¼10.6,
.3 Hz, 2H, CHCH CH ),
O), 3.48e3.38 (2ꢄdt, J¼9.4, 7.5 Hz, 2H, OCH
.88e2.77 (m, 8H, ]CCH COO in EPA),
C]), 2.34 (t, J¼7.6 Hz, 2H, CH
.30 (t, J¼7.6 Hz, 2H, CH COO in dodecanoic acid), 2.14e2.04 (m, 4H,
CCH CH and ]CCH CH CH COO
),1.70 (quintet, J¼7.5 Hz, 2H, CH
in EPA), 1.60 (quintet (br), J¼7.1 Hz, 2H, CH CH COO in dodecanoic
acid), 1.53 (quintet (br), J¼6.5 Hz, 2H, OCH CH ), 1.35e1.20 (m, 46H,
CH in EPA), 0.88 (br t, J¼6.9 Hz, 6H, CH in
), 0.97 (t, J¼7.5 Hz, 3H, CH
dodecanoic acid and 1-O-alkyl) ppm; C NMR (CDCl 173.41 (
C]O),172.84 ( , C]O),132.02,128.89,128.84,128.56,128.26,128.19,
28.16, 128.07, 127.85, 127.00, 71.75 (OCH CH ), 70.16 (CH CHCH ),
8.90 (CHCH O), 62.73 (CH OCO), 34.13, 33.72, 31.92, 31.91, 29.70
7), 29.66, 29.65, 29.62 (3), 29.55, 29.48 (2), 29.36, 29.34, 29.29,
2
3
)
d
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
]
2
2
2
3
2
2
2
2
2
3
2
2
2
2
2
2
2
2
3
3
2
2
13
3
)
d
a
,
2
2
2
3
b
3
1
3
1
6
(
2
2
2
2
3H, CH
172.84 (
128.25, 128.19, 128.15, 128.06, 127.85, 127.00, 71.74 (OCH
70.15 (CH CHCH ), 68.91 (CHCH O), 62.74 (CH OCO), 35.99, 33.71,
3
in 1-O-alkyl) ppm; C NMR (CDCl
, C]O), 132.01, 129.92, 129.81, 128.89, 128.84, 128.56,
CH ),
3
) d 173.22 (a, C]O),
2
2
b
2
2
2
2
C
9.13, 26.51, 26.04, 25.62 (2), 25.61, 25.54, 24.91, 24.82, 22.69 (2),
0.55, 14.27, 14.11 (2) ppm; HRMS (APCI): m/z calcd for
2
2
2
2
31.90, 29.776 (2), 29.54, 29.52, 29.50, 29.43, 29.31 (2), 29.27, 27.20
(2), 26.50, 26.02, 25.62 (2), 25.60, 25.53, 24.81, 22.67, 20.55, 18.37,
H
53 94
O
5
4
þNH : 828.7440; found 828.7437 amu.
1
4.26, 14.10, 13.62 ppm; HRMS (APCI): m/z calcd for C45
H
76
O
5
þH:
4
.1.40. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-tetrade-
697.5779; found: 697.5782 amu.
canoyl-sn-glycerol (8g). Full details are provided in Supplementary
data. The product 8g was afforded as yellowish oil (92% yield);
4.1.44. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-3-
2
0
[
C
a
]
D
ꢁ3.7 (c 0.95, chloroform); HRMS (APCI): m/z calcd for
hexanoyl-sn-glycerol (9c). Full details are provided in Supplementary
2
0
55
H
98
O
5
þNH : 856.7753; found 856.7751 amu.
4
data. The product 9c was afforded as yellowish oil (91% yield); [
4.7 (c 1.54, chloroform); HRMS (APCI): m/z calcd for C47
þH:
725.6079: found: 725.6062 amu.
a]
D
ꢁ
80 5
H O
4.1.41. Synthesis of (R)-1-O-octadecyl-2-eicosapentaenoyl-3-hexa-
decanoyl-sn-glycerol (8h). A procedure identical to the one described
above for 7a was followed using (R)-1-O-octadecyl-3-hexadecanoyl-
sn-glycerol 5h (100 mg, 0.172 mmol), DMAP (17 mg, 0.137 mmol),
EDCI (46 mg, 0.241 mmol), dry dichloromethane (2.5 ml) and EPA as
a free acid (52 mg, 0.172). The product was afforded as yellowish oil
4.1.45. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-
3-octanoyl-sn-glycerol (9d). A procedure identical to the one
described above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-octanoyl-sn-glycerol 6d (100 mg, 0.213 mmol), DMAP

1832
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
2
0
(
21 mg, 0.170 mmol), EDAC (57 mg, 0.298 mmol), dry dichloro-
(91% yield); [
a
]
D
ꢁ3.5 (c 1.06, chloroform); HRMS (ESI): m/z calcd
4
: 854.7596; found: 854.7573 amu.
methane (2 ml) and EPA as a free acid (65 mg, 0.213). The product
was afforded as yellowish oil after concentration (146 mg, 91%
for C55H
96
O
5
þNH
2
0
yield); [
924 (CeH), 1740 (C]O), 1653 (C]C) cm
CDCl 5.44e5.28 (m, 12H, ]CH), 5.22e5.17 (m, 1H, CH
.33 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H,
CH O),
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
.47e3.37 (2ꢄdt, J¼9.4, 7.5 Hz, 2H, OCH CH ), 2.87e2.77 (m, 8H,
CCH COO in EPA), 2.30
C]), 2.34 (t, J¼7.6 Hz, 2H, CH
COO in octanoic acid), 2.14e2.04 (m, 4H,
and ]CCH CH
), 2.01 (quartet (br), J¼6.4 Hz, 4H,
in 1-O-alkyl), 1.70 (quintet, J¼7.5 Hz, 2H, CH CH COO in
CH COO in octanoic
CH ), 1.39e1.18 (m,
in EPA), 0.88 (br t, J¼6.9 Hz,
a
]
D
ꢁ4.6 (c 1.58, chloroform); IR (ZnSe) 3012 (]CeH cis),
4.1.49. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-
3-hexadecanoyl-sn-glycerol (9h). A procedure identical to the one
described above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-hexadecanoyl-sn-glycerol 6h (100 mg, 0.172 mmol), DMAP
(17 mg, 0.138 mmol), EDCI (46 mg, 0.241 mmol), dry dichloro-
methane (2 ml) and EPA as a free acid (52 mg, 0.172 mmol). The
product was afforded as yellowish oil after concentration (137 mg,
ꢁ
1
1
2
;
H NMR (400 MHz,
CHCH ),
3
)
d
2
2
4
2
2
2
3
]
2
2
2
2
2
0
(
]
t, J¼7.6 Hz, 2H, CH
2
92% yield); [
cis), 2923 (CeH),1742 (C]O),1655 (C]C) cm ; H NMR (400 MHz,
CDCl 5.43e5.28 (m, 12H, ]CH), 5.22e5.17 (m, 1H, CH CHCH ),
4.33 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.15 (dd, J¼11.9, 6.4 Hz, 1H,
CH O), 3.48e3.38
OCO), 3.58e3.51 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
(2ꢄdt, J¼9.3, 7.5 Hz, 2H, OCH CH ), 2.88e2.77 (m, 8H, ]CCH C]),
2.34 (t, J¼7.6 Hz, 2H, CH COO in EPA), 2.29 (t, J¼7.6 Hz, 2H,
CH COO in hexadecanoic acid), 2.14e2.04 (m, 4H, ]CCH CH and
]CCH CH in 1-O-alkyl),
), 2.01 (quartet (br), J¼6.4 Hz, 4H, ]CHCH
1.70 (quintet, J¼7.5 Hz, 2H, CH CH COO in EPA), 1.60 (quintet (br),
J¼7.2 Hz, 2H, CH CH COO in hexadecanoic acid), 1.53 (quintet (br),
J¼6.8 Hz, 2H, OCH CH ), 1.38e1.20 (m, 46H, CH
3H, CH
1-O-alkyl) ppm; C NMR (CDCl
a
]
D
ꢁ4.8 (c 0.95, chloroform); IR (ZnSe) 3012 (]CeH
ꢁ1 1
CCH
2
CH
2
2
3
]
CHCH
2
2
2
3
)
d
2
2
EPA), 1.61 (quintet (br), J¼7.4 Hz, 2H, CH
2
2
2
acid), 1.54 (quintet (br), J¼6.8 Hz, 2H, OCH
2
2
2
2
3
6
d
0H, CH
H, CH
173.42 (
28.85, 128.56, 128.26, 128.19, 128.16, 128.07, 127.86, 127,00, 71.75
OCH CH ), 70.16 (CH CHCH ), 68.92 (CHCH O), 62.75 (CH OCO),
4.13, 33.73, 31.90, 31.66, 29.77 (2), 29.55, 29.52, 29.51, 29.44,
2
), 0.98 (t, J¼7.5 Hz, 3H, CH
in octanoic acid and 1-O-alkyl) ppm; C NMR (CDCl
, C]O), 172.84 ( , C]O), 132.03, 129.93, 129.81, 128.89,
3
2
2
2
13
3
3
)
2
a
b
2
2
2
1
(
3
2
2
2
3
2
2
2
2
2
2
2
2
2
2
2
9.32 (2), 29.28, 29.07, 28.93, 27.21 (2), 26.51, 26.03, 25.62 (2),
5.61, 25.54, 24.90, 24.82, 22.68, 22.60, 20.56, 14.27, 14.11,
2
2
2
), 0.97 (t, J¼7.5 Hz,
in hexadecanoic acid and
, C]O),172.82 ( , C]O),
3
in EPA), 0.88 (t, J¼6.9 Hz, 6H, CH
3
13
1
4.06 ppm; HRMS (APCI): m/z calcd for C49
H
84
O
5
þNH
4
: 770.6657;
3
)
d
173.39 (
a
b
found: 770.6647 amu.
132.00, 129.91, 129.79, 128.88, 128.84, 128.55, 128.25, 128.18, 128.15,
28.05, 127.84, 126.99, 71.73 (OCH CH ), 70.16 (CH CHCH ), 68.91
(CHCH O), 62.73 (CH OCO), 34.12, 33.71, 31.91, 31.90, 29.76 (2), 29.69
1
2
2
2
2
0
4
.1.46. Synthesis of (R)-1-O-cis-octadec-9 -enyl-2-eicosapentaenoyl-3-
2
2
decanoyl-sn-glycerol (9e). Full details are provided in Supplementary
(3), 29.65 (2), 29.62, 29.54, 29.51, 29.50, 29.48, 29.43, 29.35, 29.31
(2), 29.28 (2), 29.12, 27.20 (2), 26.50, 26.02, 25.61 (2), 25.60, 25.53,
24.90, 24.81, 22.68 (2), 20.54, 14.26, 14.10 (2) ppm; HRMS (ESI): m/z
20
data. The product 9e was afforded as yellowish oil (90% yield); [
4.5 (c 1.52, chloroform); HRMS (APCI): m/z calcd for C51
98.6970; found: 798.6976 amu.
a]
D
4
:
ꢁ
88 5
H O þNH
7
calcd for C57
H
100
O
5
4
þNH : 882.7909; found 882.7923 amu.
4
3
.1.47. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-
-dodecanoyl-sn-glycerol (9f). A procedure identical to the one
4.1.50. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-acetyl-
sn-glycerol (10a). Full details are provided in Supplementary data.
2
0
described above for 7a was followed using (R)-1-O-cis-octadec-
-enyl-3-dodecanoyl-sn-glycerol 6f (100 mg, 0.191 mmol), DMAP
19 mg, 0.153 mmol), EDAC (52 mg, 0.267 mmol), dry dichloro-
The product 10a was afforded as yellowish oil (91% yield); [
a
]
D
ꢁ5.8
9
(
(c 1.46, chloroform); HRMS (APCI): m/z calcd for C43
72
H O
5
þNH
4
:
686.5718; found 686.5720 amu.
methane (2 ml) and EPA as a free acid (58 mg, 0.191 mmol). The
product was afforded as yellowish oil after concentration
4
.1.51. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-buta-
2
0
noyl-sn-glycerol (10b). A procedure identical to the one described
above for 7a was followed using (R)-1-O-hexadecyl-3-butanoyl-sn-
glycerol 4b (100 mg, 0.259 mmol), DMAP (25 mg, 0.207 mmol), EDAC
(
139 mg, 90% yield); [
012 (]CeH cis), 2922 (CeH), 1741 (C]O), 1655 (C]C) cm
H
a
]
D
ꢁ4.4 (c 1.17, chloroform); IR (ZnSe)
ꢁ1
3
;
1
NMR (400 MHz, CDCl
.22e5.17 (m, 1H, CH CHCH
CH
OCO), 4.15 (dd, J¼11.9, 6.4 Hz, 1H, CH
2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH O), 3.48e3.38 (2ꢄdt, J¼9.4,
.5 Hz, 2H, OCH CH ), 2.88e2.77 (m, 8H, ]CCH C]), 2.34 (t,
J¼7.6 Hz, 2H, CH COO in EPA), 2.30 (t, J¼7.6 Hz, 2H, CH COO in
dodecanoic acid), 2.14e2.04 (m, 4H, ]CCH CH and ]CCH CH ),
in 1-O-alkyl), 1.70
COO in EPA), 1.60 (quintet (br),
COO in dodecanoic acid), 1.53 (quintet (br),
CH ), 1.38e1.19 (m, 38H, CH ), 0.98 (t,
in EPA), 0.88 (t, J¼6.8 Hz, 6H, CH in dodec-
173.42 ( , C]
, C]O), 132.02, 129.92, 129.81, 128.89, 128.85,
28.56, 128.26, 128.19, 128.16, 128.07, 127.85, 127.00, 71.75
OCH CH ), 70.16 (CH CHCH ), 68.92 (CHCH O), 62.74 (CH OCO),
4.13, 33.72, 31.91 (2), 29.77 (2), 29.62 (2), 29.55, 29.52, 29.51,
3
)
d
5.44e5.28 (m, 12H, ]CH),
), 4.33 (dd, J¼11.9, 3.7 Hz, 1H,
OCO), 3.58e3.51
(70 mg, 0.363 mmol), dry dichloromethane (2.5 ml) and DHA as a free
5
2
2
acid (85 mg, 0.259 mmol). The product was afforded as yellowish oil
after concentration (166 mg, 92% yield); [
form); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH),1741 (C]O),1653 (C]
C) cm
2
2
2
0
a]
D
ꢁ5.2 (c 1.16, chloro-
(
7
2
2
2
2
ꢁ
1
1
;
3
H NMR (400 MHz, CDCl ) d 5.43e5.28 (m, 12H, ]CH),
2
2
5
4
2
2
.23e5.18 (m, 1H, CH CHCH
2
2
), 4.34 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO),
.16 (dd, J¼11.9, 6.4 Hz,1H, CH OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz,
H, CHCH CH ),
O), 3.47e3.38 (2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH
.90e2.78 (m, 10H, ]CCH C]), 2.41e2.37 (m, 4H, CH CH COO in
COO in butanoic acid), 2.08 (quintet
), 1.64 (sextet, J¼7.4 Hz, 2H, CH CH COO
in butanoic acid),1.52 (quintet (br), J¼6.8 Hz, 2H, OCH CH ),1.35e1.19
m, 26H, CH in DHA), 0.94 (t, J¼7.4 Hz, 3H,
), 0.97 (t, J¼7.5 Hz, 3H, CH
CH
butanoic acid), 0.88 (br t, J¼6.8 Hz, 3H, CH
NMR (CDCl 173.22 ( , C]O), 172.37 ( , C]O), 132.02, 129.29,
28.56, 128.27 (2), 128.23, 128.09, 128.06, 128.03, 127.86, 127.79,
27.00, 71.76 (OCH CH ), 70.28 (CH CHCH ), 68.87 (CHCH O), 62.71
CH OCO), 36.00, 34.16, 31.92, 29.70 (5), 29.66, 29.64, 29.61, 29.55,
9.46, 29.36, 26.03, 25.62 (3), 25.59, 25.54, 22.72, 22.69, 20.55, 18.38,
þNH
2
2
2
2
3
2
(
.01 (quartet (br), J¼6.4 Hz, 4H, ]CHCH
2
2
quintet, J¼7.5 Hz, 2H, CH
2
CH
2
2
2
2
J¼7.3 Hz, 2H, CH
2 2
CH
2
2
2
DHA), 2.29 (t, J¼7.4 Hz, 2H, CH
2
J¼6.7 Hz, 2H, OCH
2
2
2
(br), J¼7.4 Hz, 2H ,]CCH CH
2
3
2
2
J¼7.5 Hz, 3H, CH
anoic acid and 1-O-alkyl) ppm; C NMR (CDCl
O), 172.84 (
3
3
13
3
)
d
a
2
2
(
2
3
b
13
3
3
in 1-O-alkyl) ppm; C
1
(
3
2
2
3
)
d
a
b
2
2
2
2
2
2
1
1
(
2
9.48, 29.44, 29.34, 29.32 (2), 29.28 (2), 29.13, 27.21 (2), 26.51,
6.03, 25.62 (2), 25.61, 25.54, 24.91, 24.82, 22.68 (2), 20.56,
2
2
2
2
2
2
1
8
4.27, 14.11 (2) ppm; HRMS (APCI): m/z calcd for C53
26.7283; found: 826.7280 amu.
H
92
O
5
4
þNH :
14.27, 14.11, 13.63 ppm; HRMS (APCI): m/z calcd for C45
H O
76 5
4
:
714.6031; found 714.6044 amu.
4
3
.1.48. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-eicosapentaenoyl-
-tetradecanoyl-sn-glycerol (9g). Full details are provided in
4
.1.52. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-hex-
anoyl-sn-glycerol (10c). Full details are provided in Supplementary
Supplementary data. The product 9g was afforded as yellowish oil

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1833
data. The product 10c was afforded as yellowish oil (92% yield);
ꢁ3.7 (c 1.25, chloroform); HRMS (APCI): m/z calcd for
þNH : 854.7596; found 854.7598 amu.
2
0
[
a]
D
ꢁ4.7 (c 1.44, chloroform); HRMS (APCI): m/z calcd for
C H O
55 96 5
4
47 80 5
C H O
þH: 725.6079; found 725.6091 amu.
4.1.57. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-hexa-
4
.1.53. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-octa-
decanoyl-sn-glycerol (10h). A procedure identical to the one de-
scribed above for 7a was followed using (R)-1-O-hexadecyl-3-
hexadecanoyl-sn-glycerol 4h (100 mg, 0.180 mmol), DMAP (18 mg,
0.144 mmol), EDAC (48 mg, 0.252 mmol), dry dichloromethane
(2.5 ml) and DHA as a free acid (59 mg, 0.180 mmol). The product
noyl-sn-glycerol (10d). A procedure identical to the one described
above for 7a was followed using (R)-1-O-hexadecyl-3-octanoyl-sn-
glycerol 4d (100 mg, 0.226 mmol), DMAP (22 mg, 0.181 mmol), EDAC
(
61 mg, 0.316 mmol), dry dichloromethane (2.5 ml) and DHA as a free
acid (74 mg, 0.226 mmol). The product was afforded as yellowish oil
was afforded as yellowish oil after concentration (142 mg, 91%
2
0
20
after concentration (155 mg, 91% yield); [
a
]
D
ꢁ3.3 (c 1.35, chloro-
yield); [
a
]
D
ꢁ3.4 (c 0.93, chloroform); IR (ZnSe) 3013 (]CeH cis),
ꢁ
1 1
form); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH),1742 (C]O),1653 (C]
2922 (CeH), 1742 (C]O), 1656 (C]C) cm
CDCl 5.43e5.28 (m, 12H, ]CH), 5.22e5.17 (m, 1H, CH
4.33 (dd, J¼11.9, 3.8 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.3 Hz, 1H,
CH O), 3.47e3.38
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
(2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH CH ), 2.91e2.77 (m, 10H, ]CCH C]),
2.42e2.37 (m, 4H, CH CH
COO in DHA), 2.30 (t, J¼7.5 Hz, 2H,
CH
COO in hexadecanoic acid), 2.08 (quintet (br), J¼7.4 Hz, 2H,
]CCH CH CH COO in hex-
), 1.61 (quintet (br), J¼7.3 Hz, 2H, CH
adecanoic acid),1.54 (quintet (br), J¼6.9 Hz, 2H, OCH CH ),1.37e1.17
(m, 50H, CH in DHA), 0.88 (br t, J¼6.9 Hz,
), 0.97 (t, J¼7.5 Hz, 3H, CH
6H, CH in hexadecanoic acid and 1-O-alkyl) ppm; C NMR (CDCl
173.40 ( , C]O),172.35( , C]O),132.00,129.29,128.55,128.26 (2),
128.23, 128.09, 128.06, 128.03, 127.85, 127.79, 127.00, 71.76
(OCH CH ), 70.28 (CH CHCH ), 68.87 (CHCH O), 62.69 (CH OCO),
;
H NMR (400 MHz,
ꢁ
1
1
C) cm
;
H NMR (400 MHz, CDCl
3
)
d
5.43e5.28 (m, 12H ,]CH),
3
)
d
2
CHCH ),
2
5
4
2
.22e5.17 (m, 1H, CH CHCH
2
2
), 4.33 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO),
.16 (dd, J¼11.9, 6.3 Hz, 1H, CH OCO), 3.57e3.50 (2ꢄdd, 10.7, 5.3 Hz,
H, CHCH CH ), 2.91e2.77
O), 3.47e3.38 (2ꢄdt, 9.4, 7.4 Hz, 2H, OCH
C]), 2.42e2.37 (m, 4H, CH CH COO in DHA), 2.30 (t,
J¼7.5 Hz, 2H, CH COO in octanoic acid), 2.08 (quintet (br), J¼7.3 Hz,
H, ]CCH CH CH COO in
), 1.61 (quintet (br), J¼7.4 Hz, 2H, CH
octanoic acid), 1.54 (quintet (br), J¼6.6 Hz, 2H, OCH CH ), 1.36e1.18
m, 34H, CH in DHA), 0.88 (br t, J¼6.9 Hz,
), 0.97 (t, J¼7.5 Hz, 3H, CH
H, CH inoctanoicacidand1-O-alkyl)ppm; CNMR(CDCl
, C]O),172.37 ( , C]O),132.02,129.29,128.56,128.27 (2C),128.23,
28.09,128.06, 128.03,127.86, 127.79,127.00, 71.76 (OCH CH ), 70.29
CH CHCH ), 68.87 (CHCH O), 62.70 (CH OCO), 34.16, 34.13, 31.92,
1.66, 29.70 (5C), 29.66, 29.64, 29.61, 29.55, 29.47, 29.36, 29.07, 28.93,
2
2
2
2
2
2
2
2
2
2
2
(
m, 10H, ]CCH
2
2
2
2
2
2
2
2
2
3
2
2
2
3
2
2
2
2
2
2
(
6
2
3
2
3
13
13
3
3
)
d
173.41
3
3
)
(a
b
d
a
b
1
(
3
2
2
2
2
2
2
2
2
2
2
2
2
2
34.16, 34.13, 31.92 (2), 29.70 (8), 29.66 (3), 29.64, 29.63, 29.61, 29.55,
29.49, 29.47, 29.36 (2), 29.29, 29.13, 26.03, 25.62 (3), 25.59, 25.53,
24.90, 22.72, 22.69 (2), 20.55,14.26,14.11 (2) ppm; HRMS (APCI): m/z
6.03, 25.62 (3C), 25.59, 25.53, 24.90, 22.72, 22.69, 22.60, 20.55,14.26,
1
4.11,14.06 ppm; HRMS (APCI): m/z calcd for C49
84 5
H O þH: 753.6392;
found 753.6388 amu.
calcd for C57
H
100
O
5
4
þNH : 882.7909; found 882.7906 amu.
4
.1.54. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-dec-
4.1.58. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-acetyl-
sn-glycerol (11a). A procedure identical to the one described above
for 7a was followed using (R)-1-O-octadecyl-3-acetyl-sn-glycerol 5a
(100 mg, 0.259 mmol), DMAP (25 mg, 0.207 mmol), EDAC (70 mg,
0.363 mmol), dry dichloromethane (2.5 ml) and DHA as a free acid
anoyl-sn-glycerol (10e). Full details are provided in Supplementary
data. The product 10e was afforded as yellowish oil (92% yield);
2
0
[
a
]
D
ꢁ3.0 (c 1.16, chloroform); HRMS (ESI): m/z calcd for
C
51
H
88
O
5
þNH : 798.6970; found 798.6967 amu.
4
(
85 mg, 0.259). The product was afforded as yellowish semisolid oil
2
0
4.1.55. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-dodec-
after concentration (162 mg, 90% yield); [
form); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH), 1744 (C]O), 1656
a]
D
ꢁ6.0 (c 0.80, chloro-
anoyl-sn-glycerol (10f). A procedure identical to the one described
above for 7a was followed using (R)-1-O-hexadecyl-3-dodecanoyl-
sn-glycerol 4f (100 mg, 0.200 mmol), DMAP (20 mg, 0.160 mmol),
EDAC (54 mg, 0.280 mmol), dry dichloromethane (2.5 ml) and DHA
as a free acid (66 mg, 0.200 mmol). The product was afforded as
ꢁ
1 1
(C]C) cm ; H NMR (400 MHz, CDCl
5.23e5.17 (m,1H, CH CHCH
), 4.33 (dd, J¼11.9, 3.7 Hz,1H, CH
4.16 (dd, J¼11.9, 6.5 Hz, 1H, CH OCO), 3.57e3.50 (2ꢄdd, J¼10.6,
5.3 Hz, 2H, CHCH CH ),
O), 3.47e3.38 (2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH
2.88e2.77 (m, 10H, ]CCH C]), 2.42e2.36 (m, 4H, CH CH COO),
2.11e2.04 (m, 2H, ]CCH CH ), 2.05 (s, 3H, CH COO), 1.54 (quintet
(br), J¼7.0 Hz, 2H, OCH CH ), 1.35e1.19 (m, 30H, CH ), 0.97 (t,
J¼7.5 Hz, 3H, CH in DHA), 0.88 (br t, J¼6.9 Hz, 3H, CH in 1-O-alkyl)
ppm; C NMR (100 MHz, CDCl 172.40 ( , C]O),170.62 ( , C]O),
132.01, 129.30, 128.55, 128.26 (2), 128.23, 128.08, 128.06, 128.02,
127.85, 127.77, 127.00, 71.76 (OCH CH ), 70.23 (CH CHCH ), 68.85
(CHCH O), 63.00 (CH OCO), 34.16, 31.92, 29.69 (7), 29.65, 29.63,
3
)
d
5.43e5.28 (m,12H, ]CH),
2
2
2
OCO),
2
2
2
2
2
0
yellowish oil after concentration (147 mg, 91% yield); [
a
]
D
ꢁ3.1 (c
2
2
2
1.43, chloroform); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH), 1742
2
3
3
ꢁ1 1
(
C]O), 1653 (C]C) cm ; H NMR (400 MHz, CDCl
m, 12H, ]CH), 5.22e5.17 (m, 1H, CH CHCH
3
)
d
5.43e5.28
2
2
2
(
2
2
), 4.33 (dd, J¼11.9,
OCO), 4.16 (dd, J¼11.9, 6.3 Hz, 1H, CH OCO),
.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH O), 3.47e3.38 (2ꢄdt,
CH ), 2.91e2.77 (m, 10H, ]CCH C]),
CH
COO in DHA), 2.30 (t, J¼7.5 Hz, 2H,
COO in dodecanoic acid), 2.08 (quintet (br), J¼7.3 Hz, 2H,
CCH CH CH COO in dodecanoic
), 1.60 (quintet, J¼7.3 Hz, 2H, CH
acid), 1.53 (quintet (br), J¼6.8 Hz, 2H, OCH CH ), 1.37e1.18 (m, 42H,
CH
), 0.97 (t, J¼7.5 Hz, 3H, CH
in dodecanoic acid and 1-O-alkyl) ppm; C NMR (CDCl
, C]O), 172.37 (
28.09, 128.07, 128.04, 127.86, 127.79, 127.00, 71.76 (OCH
0.29 (CH CHCH ), 68.87 (CHCH O), 62.70 (CH OCO), 34.16, 34.14,
1.93, 31.91, 29.70 (5), 29.66, 29.64, 29.62 (3), 29.55, 29.48 (2),
3
3
13
3
3
.7 Hz, 1H, CH
2
2
3
)
d
b
a
2
J¼9.4, 7.4 Hz, 2H, OCH
.44e2.36 (m, 4H, CH
CH
2
2
2
2
2
2
2
2
2
2
2
2
2
29.60, 29.54, 29.46, 29.35, 26.02, 25.62 (3), 25.59, 25.53, 22.72, 22.68,
20.76, 20.55, 14.26, 14.11 ppm; HRMS (APCI): m/z calcd for
]
2
3
2
2
2
2
C
45
H
76
O
5
4
þNH : 714.6031; found 714.6008 amu.
2
3
in DHA), 0.88 (br t, J¼6.9 Hz, 6H, CH
3
13
3
)
d
173.42
4.1.59. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-buta-
noyl-sn-glycerol (11b). Full details are provided in Supplementary
data. The product 11b was afforded as yellowish semisolid oil
(a
b
, C]O), 132.02, 129.29, 128.56, 128.27(2), 128.24,
CH ),
1
7
3
2
2
2
0
2
2
2
2
(92% yield); [
a
]
D
ꢁ4.0 (c 1.07, chloroform); HRMS (APCI): m/z calcd
for C47
80
H O
5
þNH
4
: 742.6344; found 742.6324 amu.
2
2
9.36, 29.34, 29.29, 29.13, 26.04, 25.63 (3), 25.60, 25.54, 24.91,
2.72, 22.69 (2), 20.56, 14.27, 14.11 (2) ppm; HRMS (APCI): m/z calcd
4.1.60. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-hex-
anoyl-sn-glycerol (11c). A procedure identical to the one described
above for 7a was followed using (R)-1-O-octadecyl-3-hexanoyl-sn-
glycerol 5c (100 mg, 0.226 mmol), DMAP (22 mg, 0.181 mmol),
EDAC (61 mg, 0.316 mmol), dry dichloromethane (2.5 ml) and EPA
as a free acid (74 mg, 0.226). The product was afforded as yellowish
for C53
92
H O
5
þNH
4
: 826.7283; found 826.7285 amu.
4
.1.56. Synthesis of (R)-1-O-hexadecyl-2-docosahexaenoyl-3-tetrade-
canoyl-sn-glycerol (10g). Full details are provided in Supplementary
data. The product 10g was afforded as yellowish oil (93% yield); [a]
D
2
0

1834
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
2
0
20
semisolid oil after concentration (157 mg, 92% yield); [
a
]
D
ꢁ3.7 (c
yield); [
2922 (CeH), 1742 (C]O), 1655 (C]C) cm
CDCl 5.43e5.28 (m, 12H, ]CH), 5.22e5.17 (m, 1H, CH
4.33 (dd, J¼11.9, 3.8 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.3 Hz, 1H,
CH O), 3.47e3.38
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
(2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH CH ), 2.88e2.78 (m, 10H, ]CCH C]),
2.42e2.37 (m, 4H, CH CH
COO in DHA), 2.30 (t, J¼7.6 Hz, 2H,
CH
COO in tetradecanoic acid), 2.07 (quintet (br), J¼7.3 Hz, 2H, ]
CCH CH CH COO in tetradecanoic
), 1.60 (quintet, J¼7.3 Hz, 2H, CH
acid), 1.53 (quintet (br), J¼6.9 Hz, 2H, OCH CH ), 1.35e1.20 (m, 50H,
CH
), 0.97 (t, J¼7.5 Hz, 3H, CH
in tetradecanoic acid and 1-O-alkyl) ppm; C NMR (CDCl
, C]O),172.38 (
128.10,128.07,128.04,127.87,127.80,127.01, 71.77 (OCH
(CH CHCH ), 68.88 (CHCH O), 62.70 (CH OCO), 34.17, 34.14, 31.93
a
]
D
ꢁ3.8 (c 1.04, chloroform); IR (ZnSe) 3013 (]CeH cis),
ꢁ
1 1
1
(
.36, chloroform); IR (ZnSe) 3013 (]CeH cis), 2922 (CeH), 1742
;
H NMR (400 MHz,
CHCH ),
ꢁ
1 1
C]O), 1654 (C]C) cm ; H NMR (400 MHz, CDCl
m, 12H, ]CH), 5.23e5.18 (m, 1H, CH CHCH
OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H, CH
.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH O), 3.47e3.38 (2ꢄdt,
CH ), 2.88e2.78 (m, 10H, ]CCH C]),
CH
COO in DHA), 2.30 (t, J¼7.5 Hz, 2H,
CH ),
COO in hexanoic acid), 1.54
3
)
d
5.43e5.28
3
)
d
2
2
(
2
2
), 4.34 (dd, J¼11.9,
2
3
3
.7 Hz, 1H, CH
2
2
OCO),
2
2
2
2
2
2
J¼9.4, 7.4 Hz, 2H, OCH
.42e2.37 (m, 4H, CH
CH
2
2
2
2
2
2
2
2
2
2
COO in hexanoic acid), 2.08 (quintet, J¼7.3 Hz, 2H, ]CCH
2
3
2
3
2
2
1
(
(
.62 (quintet, J¼7.5 Hz, 2H, CH
quintet (br), J¼6.9 Hz, 2H, OCH CH
t, J¼7.5 Hz, 3H, CH
2
CH
2
2
2
2
2
), 1.35e1.20 (m, 34H, CH
2
), 0.97
2
3
in DHA), 0.88 (br t, J¼6.9 Hz, 6H, CH
3
13
3
in DHA), 0.89 (br t, J¼6.2 Hz, 3H, CH
3
hexanoic
in 1-O-alkyl) ppm; C NMR
3
)
d
173.41
1
3
acid), 0.88 (br t, J¼6.2 Hz, 3H, CH
CDCl 173.42 ( , C]O), 172.37 (
28.27 (2), 128.24, 128.10, 128.07, 128.04, 127.87, 127.80, 127.00,
1.77 (OCH CH ), 70.29 (CH CHCH ), 68.88 (CHCH O), 62.72
CH OCO), 34.17, 34.10, 31.93, 31.26, 29.71 (7), 29.66, 29.65, 29.62,
9.56, 29.47, 29.36, 26.04, 25.63 (3), 25.60, 25.54, 24.58, 22.72,
2.69, 22.31, 20.56, 14.27, 14.12, 13.90 ppm; HRMS (APCI): m/z calcd
þNH : 770.6657; found 770.6676 amu.
3
(
a
b
, C]O),132.02,129.30,128.56,128.27 (2),128.24,
CH ), 70.30
(
3
)
d
a
b
, C]O), 132.03, 129.30, 128.56,
2
2
1
2
2
2
2
7
2
2
2
2
2
(2), 29.71 (6), 29.69 (2), 29.66 (4), 29.63 (2C), 29.56, 29.50, 29.48,
29.36 (2), 29.30, 29.14, 26.04, 25.63 (3), 25.60, 25.54, 24.91, 22.73,
22.69 (2), 20.56, 14.27, 14.12 (2) ppm; HRMS (APCI): m/z calcd for
(
2
2
2
C
57
H
100
O
5
þH: 865.7644; found 865.7647 amu.
for C49
84
H O
5
4
4
.1.65. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-hexa-
4
.1.61. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-octa-
decanoyl-sn-glycerol (11h). Full details are provided in
Supplementary data. The product 11h was afforded as yellowish oil
noyl-sn-glycerol (11d). Full details are provided in Supplementary
data. The product 11d was afforded as yellowish oil (, 90% yield);
2
0
(90% yield); [
for C59
a
]
D
ꢁ4.1 (c 1.02, chloroform); HRMS (APCI): m/z calcd
2
0
[
a
]
D
ꢁ3.5 (c 1.11, chloroform); HRMS (APCI): m/z calcd for
H
104
O
5
þNH : 910.8222; found 910.8239 amu.
4
C
H
51 88
O
5
þNH : 798.6970; found 798.6961 amu.
4
4.1.66. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-
4
.1.62. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-dec-
3-acetyl-sn-glycerol (12a). A procedure identical to the one de-
scribed above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-acetyl-sn-glycerol 6a (100 mg, 0.260 mmol), DMAP (25 mg,
0.208 mmol), EDAC (70 mg, 0.364 mmol), dry dichloromethane
(2 ml) and DHA as a free acid (85 mg, 0.260 mmol). The product
anoyl-sn-glycerol (11e). A procedure identical to the one described
above for 7a was followed using (R)-1-O-octadecyl-3-decanoyl-sn-
glycerol 5e (100 mg, 0.200 mmol), DMAP (20 mg, 0.160 mmol), EDCI
(
54 mg, 0.280 mmol), dry dichloromethane (2.5 ml)and DHA asa free
acid (66 mg, 0.200). The product was afforded as yellowish oil after
was afforded as yellowish oil after concentration (163 mg, 90%
2
0
20
concentration (155 mg, 95% yield); [
a
]
D
ꢁ2.8 (c 0.76, chloroform); IR
yield); [
a
]
D
ꢁ5.5 (c 1.68, chloroform); IR (ZnSe) 3012 (]CeH cis),
ꢁ
1
ꢁ1
1
(
ZnSe) 3013 (]CeH cis), 2922 (CeH),1742 (C]O),1655 (C]C) cm
;
2924 (CeH), 1744 (C]O), 1654 (C]C) cm
CDCl 5.43e5.28 (m, 14H, ]CH), 5.23e5.18 (m, 1H, CH
4.33 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.5 Hz, 1H,
CH O), 3.49e3.39
(2ꢄdt, J¼9.3, 7.3 Hz, 2H, OCH ), 2.88e2.80 (m, 10H, ]CCH C]),
2.42e2.39 (m, 4H, CH CH COO in DHA), 2.11e2.03 (m, 2H,
]CCH CH ), 2.05 (s, 3H, CH
]CHCH
in 1-O-alkyl), 1.54 (quintet (br), J¼6.8 Hz, 2H, OCH
1.38e1.21 (m, 22H, CH in DHA), 0.88 (br
), 0.97 (t, J¼7.5 Hz, 3H, CH
t, J¼6.9 Hz, 3H, CH in 1-O-alkyl) ppm; C NMR (CDCl 172.41 (
C]O), 170.64 ( , C]O), 132.03, 129.93, 129.82, 129.32, 128.57,
128.28 (2), 128.24, 128.10, 128.07, 128.04, 127.87, 127.78, 127.01,
71.77 (OCH CH ), 70.23 (CH CHCH ), 68.87 (CHCH O), 63.01
(CH OCO), 34.17, 31.91, 29.77 (2), 29.55, 29.52, 29.51, 29.44, 29.32
;
H NMR (400 MHz,
1
H NMR (400 MHz, CDCl
3
)
d
5.43e5.28 (m,12H, ]CH), 5.22e5.17 (m,
), 4.33 (dd, J¼11.9, 3.7 Hz,1H, CH OCO), 4.16 (dd, J¼11.9,
OCO), 3.57e3.50 (2ꢄdd, J¼10.7, 5.3 Hz, 2H, CHCH O),
.47e3.38 (2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH CH ), 2.89e2.78 (m, 10H,
CCH C]), 2.42e2.36 (m, 4H, CH CH
COO in DHA), 2.30 (t, J¼7.5 Hz,
H, CH
COO in decanoic acid), 2.08 (quintet (br), J¼7.4 Hz, 2H,
CCH CH CH COO in decanoic
), 1.61 (quintet (br), J¼7.4 Hz, 2H, CH
acid), 1.54 (quintet (br), J¼6.5 Hz, 2H, OCH CH ), 1.36e1.18 (m, 42H,
CH in DHA), 0.88 (br t, J¼6.8 Hz, 6H, CH in
), 0.97 (t, J¼7.5 Hz, 3H, CH
decanoicacidand1-O-alkyl)ppm; CNMR(CDCl , C]O),
72.37 ( , C]O), 132.02, 129.29, 128.56, 128.27 (2), 128.24, 128.10,
28.07, 128.04, 127.86, 127.80, 127.01, 71.77 (OCH CH ), 70.29
CH CHCH ), 68.88 (CHCH O), 62.70 (CH OCO), 34.17, 34.14, 31.93,
1.87, 29.71 (7), 29.66, 29.65, 29.62, 29.55, 29.48, 29.44, 29.36, 29.28
2), 29.13, 26.04, 25.63 (3), 25.60, 25.54, 24.91, 22.72, 22.69, 22.67,
3
)
d
2
CHCH ),
2
1
6
3
H, CH
.3 Hz, 1H, CH
2
CHCH
2
2
2
2
2
2
OCO), 3.58e3.51 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
2
2
2
2
CH
2
2
]
2
2
2
2
2
2
2
2
3
3
COO), 2.01 (quartet (br), J¼6.5 Hz, 4H,
]
2
3
2
2
2
2
CH ),
2
2
2
2
3
13
2
3
3
3
3
)
d
b,
13
3
)
d
173.42(
a
a
1
b
1
2
2
2
2
2
2
2
(
2
2
2
2
2
3
(
(2), 29.28, 27.21 (2), 26.03, 25.63 (3), 25.60, 25.54, 22.73, 22.68,
20.78, 20.56, 14.27, 14.11 ppm; HRMS (APCI): m/z calcd for
2
C
0.56, 14.27, 14.12, 14.10 ppm; HRMS (APCI): m/z calcd for
þNH : 826.7283; found 826.7294 amu.
C
45
H
74
O
5
þ NH
4
: 712.5875; found: 712.5870 amu.
H O
53 92 5
4
4.1.67. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-
4.1.63. Synthesis of (R)-1-O-octadecyl-2-docosaenoyl-3-dodecanoyl-
3-butanoyl-sn-glycerol (12b). Full details are provided in
sn-glycerol (11f). Full details are provided in Supplementary data.
Supplementary data. The product 12b was afforded as yellowish oil
2
0
20
The product 11f was afforded as yellowish oil (90% yield); [
a
]
D
ꢁ3.0
(90% yield); [
for C47
a]
D
ꢁ4.7 (c 1.54, chloroform); HRMS (APCI): m/z calcd
(
c 1.26, chloroform); HRMS (APCI): m/z calcd for C55
H
96
O
5
þNH
4
:
78
H O
5
þNH
4
: 740.6188; found: 740.6190 amu.
8
54.7596; found 854.7566 amu.
4.1.68. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-
4
.1.64. Synthesis of (R)-1-O-octadecyl-2-docosahexaenoyl-3-tetra-
3-hexanoyl-sn-glycerol (12c). A procedure identical to the one
described above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-hexanoyl-sn-glycerol 6c (120 mg, 0.272 mmol), DMAP
(26 mg, 0.218 mmol), EDAC (73 mg, 0.380 mmol), dry dichloro-
methane (2 ml) and DHA as a free acid (89 mg, 0.272). The product
was afforded as yellowish oil after concentration (184 mg, 90%
decanoyl-sn-glycerol (11g). A procedure identical to the one de-
scribed above for 7a was followed using (R)-1-O-octadecyl-3-
tetradecanoyl-sn-glycerol 5g (100 mg, 0.180 mmol), DMAP (18 mg,
0
(
.144 mmol), EDAC (48 mg, 0.252 mmol), dry dichloromethane
2.5 ml) was added DHA as a free acid (59 mg, 0.180). The product
was afforded as yellowish oil after concentration (145 mg, 93%
2
0
yield); [
a
]
D
ꢁ4.3 (c 1.26, chloroform); IR (ZnSe) 3012 (]CeH cis),

C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
1835
ꢁ
2
924 (CeH), 1741 (C]O), 1653 (C]C) cm 1; 1H NMR (400 MHz,
CDCl 5.43e5.28 (m, 14H, ]CH), 5.23e5.18 (m, 1H, CH CHCH ),
.34 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.4 Hz, 1H,
CH O), 3.47e3.38
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH CH ), 2.88e2.78 (m, 10H, ]CCH C]),
.42e2.36 (m, 4H, CH CH
COO in DHA), 2.30 (t, J¼7.5 Hz, 2H,
CH CH ),
COO in hexanoic acid), 2.07 (quintet, J¼7.5 Hz, 2H, ]CCH
.01 (quartet (br), J¼6.4 Hz, 4H, ]CHCH in 1-O-alkyl),1.61 (quintet,
J¼7.5 Hz, 2H, CH CH COO in hexanoic acid), 1.54 (quintet (br),
J¼7.2 Hz, 2H, OCH CH ), 1.38e1.19 (m, 26H, CH ), 0.97 (t, J¼7.5 Hz,
H, CH hexanoic acid), 0.88 (br
in DHA), 0.89 (br t, J¼6.9 Hz, 3H, CH
t, J¼6.1 Hz, 3H, CH in 1-O-alkyl) ppm; C NMR (CDCl 173.43 (
C]O),172.38 ( , C]O),132.01,129.91,129.80,129.28,128.54,128.25
2), 128.22, 128.07, 128.05, 128.02, 127.84, 127.77, 126.99, 71.73
OCH CH ), 70.25 (CH CHCH ), 68.85 (CHCH O), 62.70 (CH OCO),
4.13, 34.07, 31.89, 31.23, 29.75 (2), 29.51 (2), 29.49, 29.42, 29.30 (2),
product was afforded as yellowish oil after concentration (140 mg,
2
0
3
)
d
2
2
90% yield); [
cis), 2923 (CeH), 1742 (C]O), 1654 (C]C) cm
(400 MHz, CDCl 5.43e5.28 (m, 14H, ]CH), 5.22e5.17 (m, 1H,
CH CHCH OCO), 4.16 (dd, J¼11.9,
), 4.33 (dd, J¼11.9, 3.7 Hz, 1H, CH
6.3 Hz, 1H, CH
OCO), 3.57e3.50 (2ꢄdd, J¼10.6, 5.3 Hz, 2H,
CHCH CH ), 2.88e2.77
O), 3.47e3.38 (2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH
(m, 10H, ]CCH C]), 2.42e2.36 (m, 4H, CH CH COO in DHA), 2.30
(t, J¼7.5 Hz, 2H, CH COO in tetradecanoic acid), 2.08 (quintet (br),
J¼7.4 Hz, 2H, ]CCH CH ), 2.01 (quartet (br), J¼6.4 Hz, 4H, ]CHCH
in 1-O-alkyl), 1.60 (quintet (br), J¼7.3 Hz, 2H, CH CH COO in tet-
radecanoic acid), 1.54 (quintet (br), J¼6.7 Hz, 2H, OCH CH ),
1.38e1.20 (m, 42H, CH in DHA), 0.88 (br
), 0.97 (t, J¼7.5 Hz, 3H, CH
t, J¼6.9 Hz, 6H, CH in tetradecanoic acid and 1-O-alkyl) ppm;
NMR (CDCl 173.42 ( , C]O), 172.37 ( , C]O), 132.02, 129.93,
129.81, 129.30, 128.56, 128.27 (2), 128.24, 128.10, 128.07, 128.04,
127.86, 127.79, 127.01, 71.76 (OCH CH ), 70.29 (CH CHCH ), 68.89
(CHCH O), 62.70 (CH OCO), 34.16, 34.14, 31.92, 31.91, 29.77 (2),
29.69, 29.66, 29.65, 29.63, 29.56, 29.53, 29.51, 29.49, 29.45, 29.34,
9.32 (2), 29.29 (2), 29.14, 27.22 (2), 26.04, 25.63 (3), 25.60, 25.54,
24.91, 22.72, 22.69 (2), 20.56, 14.27, 14.12 (2) ppm; HRMS (ESI): m/z
calcd for C57 : 880.7753; found: 880.7755 amu.
þNH
a
]
D
ꢁ3.5 (c 1.29, chloroform); IR (ZnSe) 3013 (]CeH
ꢁ
1 1
4
2
;
H NMR
2
2
3
) d
(
2
2
2
2
2
2
2
2
2
2
2
2
3
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
3
2
3
3
3
2
2
13
3
3
)
d
a
,
2
2
b
2
3
13
(
(
3
2
2
C
3
C
2
2
2
2
2
2
3
)
d
a
b
9.26, 27.19 (2), 26.01, 25.60 (3), 25.57, 25.51, 24.56, 22.69, 22.67,
2.29, 20.54, 14.26, 14.11, 13.89 ppm; HRMS (APCI): m/z calcd for
2
2
2
2
2
2
H
49 82
O
5
þH: 751.6235; found: 751.6230 amu.
2
4
.1.69. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-3-
octanoyl-sn-glycerol (12d). Full details are provided in Supplementary
data. The product 12d was afforded as yellowish oil (90% yield); [
98
H O
5
4
2
0
a]
D
ꢁ
4.1 (c 1.25, chloroform); HRMS (APCI): m/z calcd for C51
79.6548; found: 779.6552 amu.
H
86
O
5
þH:
4.1.73. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-
3-hexadecanoyl-sn-glycerol (12h). Full details are provided in
Supplementary data. The product 12h was afforded as yellowish oil
7
2
0
4
3
.1.70. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-dodecahexaenoyl-
-decanoyl-sn-glycerol (12e). A procedure identical to the one
(91% yield); [
a
]
D
ꢁ3.8 (c 0.90, chloroform); HRMS (ESI): m/z calcd
for C59H
102
O
5
þNH : 908.8066; found 908.8102 amu.
4
described above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-decanoyl-sn-glycerol 6e (100 mg, 0.201 mmol), DMAP
4.1.74. Calculation of the equilibrium level of 2-monoacyl-1-O-alkyl-
(
20 mg, 0.161 mmol), EDAC (54 mg, 0.281 mmol), dry dichloro-
sn-glycerol. To a solution of pure 1-O-octadecyl-3-octanoyl-sn-
methane (2 ml) and DHA as a free acid (66 mg, 0.201). The product
was afforded as yellowish oil after concentration (148 mg, 91%
2 2
glycerol 5d (100 mg, 0.212 mmol) in dry CH Cl (3 ml), Amberlyst
15 (10 mg) was added and the resulting mixture stirred at room
temperature for 7 days. Aliquots (0.8 ml) were withdrawn from the
reaction mixture, with 56 h intervals, passed through a Pasteur
pipette packed with a plug of cotton wool and concentrated in
20
yield); [
923 (CeH), 1741 (C]O), 1654 (C]C) cm
CDCl 5.43e5.28 (m, 14H, ]CH), 5.22e5.17 (m, 1H, CH
.33 (dd, J¼11.9, 3.7 Hz, 1H, CH OCO), 4.16 (dd, J¼11.9, 6.3 Hz, 1H,
CH O), 3.48e3.39
OCO), 3.58e3.51 (2ꢄdd, J¼10.6, 5.3 Hz, 2H, CHCH
2ꢄdt, J¼9.4, 7.4 Hz, 2H, OCH CH ), 2.88e2.78 (m, 10H, ]CCH C]),
.42e2.36 (m, 4H, CH CH
COO in DHA), 2.30 (t, J¼7.5 Hz, 2H,
CH
COO in decanoic acid), 2.07 (quintet (br), J¼7.4 Hz, 2H,
CCH CH in 1-O-alkyl),
), 2.01 (quartet (br), J¼6.5 Hz, 4H, ]CHCH
.60 (quintet (br), J¼7.3 Hz, 2H, CH CH COO in decanoic acid), 1.54
quintet (br), J¼6.8 Hz, 2H, OCH CH ), 1.38e1.18 (m, 34H, CH ), 0.97
t, J¼7.5 Hz, 3H, CH in DHA), 0.88 (br t, J¼7.2 Hz, 6H, CH
a
]
D
ꢁ3.7 (c 1.55, chloroform); IR (ZnSe) 3013 (]CeH cis),
ꢁ
1 1
2
;
H NMR (400 MHz,
CHCH ),
3
)
d
2
2
4
2
vacuo. The afforded white crystals were subsequently dissolved in
1
2
2
0.7 ml of CDCl
3
and submitted to H NMR measurements.
(
2
2
2
2
2
2
4.1.75. Preparation
of
(S)-1-O-octadecyl-2-octanoyl-sn-glycerol
2
(13d). To a 10 ml round-bottomed flask containing (R)-1-octadecyl-
3-octanoyl-sn-glycerol 5d (100 mg, 0.212 mmol) dissolved in chlo-
roform (5 ml), p-toluenesulfonic acid monohydrate (4 mg,
0.0212 mmol) was added and the solution stirred for 1 h at room
temperature. After evaporation of the solvent the crude mixture was
applied to preparative TLC and eluted with petroleum ether/ether/
]
2
3
2
1
2
2
(
(
2
2
2
3
3
in dec-
, C]O),
1
3
anoic acid and 1-O-alkyl) ppm; C NMR (CDCl
72.37 (
28.24, 128.09, 128.06, 128.03, 127.86, 127.79, 127.01, 71.75
OCH CH ), 70.29 (CH CHCH ), 68.89 (CHCH O), 62.70 (CH OCO),
4.16, 34.14, 31.90, 31.86, 29.77 (2), 29.56, 29.52, 29.51, 29.44 (2),
3
)
d
173.42 (
a
1
1
(
3
2
2
b
, C]O), 132.02, 129.92, 129.81, 129.29, 128.56, 128.27 (2),
acetic acid (80:20:2). The product (R
f
¼0.45) was finally afforded as
white crystals. H NMR (400 MHz, CDCl
5.00 (quintet, J¼4.8, 1H,
CH CHCH ), 3.83e3.80 (m, 2H, CH
OH), 3.64 (dd, J¼10.5, 4.9 Hz, 1H,
CHCH O), 3.50e3.40 (dt, J¼9.3,
O), 3.60 (dd, J¼10.5, 5.1 Hz,1H, CHCH
7.1 Hz, 2H, OCH CH COO), 2.18 (t,
), 2.36 (t, J¼7.5 Hz, 2H, CH
J¼6.2 Hz, 1H, CH OH), 1.64 (quintet (br), J¼7.4 Hz, 2H, CH CH COO),
1.57e1.52 (m, 2H, OCH CH ), 1.37e1.18 (m, 38H, CH ), 0.88 (br t,
J¼6.9 Hz, 6H, CH in octanoic acid and 1-O-alkyl) ppm; C NMR
(CDCl 173.70 (C]O), 72.81, 71.92, 70.04, 63.08, 34.40, 31.93, 31.66,
1
3
) d
2
2
2
2
2
2
2
2
2
2
2
9.32 (2), 29.28 (3), 29.13, 27.21 (2), 26.04, 25.63 (3), 25.60, 25.54,
4.91, 22.72, 22.68, 22.66, 20.56, 14.27, 14.11 (2) ppm; HRMS (APCI):
2
2
2
2
2
2
m/z calcd for C53
H
90
O
5
þH: 807.6861; found: 807.6888 amu.
2
2
2
13
3
4
3
.1.71. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-
-dodecanoyl-sn-glycerol (12f). Full details are provided in
3
) d
29.70 (6), 29.66 (2), 29.63, 29.60, 29.55, 29.45, 29.36, 29.05, 28.92,
26.05, 24.99, 22.69, 22.60, 14.12, 14.06 ppm.
Supplementary data. The product 12f was afforded as yellowish oil
2
0
(
90% yield); [
a
]
D
ꢁ3.8 (c 1.64, chloroform); HRMS (APCI): m/z calcd
for C55
94
H O
5
þNH
4
: 852.7440; found: 852.7423 amu.
Acknowledgements
4
3
.1.72. Synthesis of (R)-1-O-cis-octadec-9-enyl-2-docosahexaenoyl-
-tetradecanoyl-sn-glycerol (12g). A procedure identical to the one
The University of Iceland Research Fund and Lysi ehf in Iceland
are acknowledged for financial support. Dr. Sigridur Jonsdottir
and Dr. Sigurdur V. Smarason at University of Iceland are
acknowledged for high-resolution 2D NMR and accurate MS
measurements.
described above for 7a was followed using (R)-1-O-cis-octadec-9-
enyl-3-tetradecanoyl-sn-glycerol 6g (100 mg, 0.181 mmol), DMAP
(
18 mg, 0.145 mmol), EDAC (49 mg, 0.253 mmol), dry dichloro-
methane (2 ml) and DHA as a free acid (59 mg, 0.181 mmol). The

1836
C.D. Magnusson et al. / Tetrahedron 67 (2011) 1821e1836
Supplementary data
pp 336e368; (b) Hjaltason, B.; Haraldsson, G. G. In Modifying Lipids for Use in
Food; Gunstone, F. D., Ed.; Woodhead Publishing: Cambridge UK, 2006; Chapter
4, pp 587e602.
7. (a) Gunstone, F. D. Structured and Modified Lipids; Marcel Decker: New York, NY,
2001; (b) Akoh, C. C. In Structured Lipids; Akoh, C. C., Min, D. B., Eds.; Marcel
Dekker: New York, NY, 1998; Chapter 26, pp 699e727; (c) Christophe, A. B.
Structural Modified Food Fats: Synthesis, Biochemistry, and Use; AOCS: Cham-
paign, IL, 1998.
2
Supplementary data associated with this article can be found in
online version at doi:10.1016/j.tet.2011.01.032.
1
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7
2
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1
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6
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