Structured estolides: Control of length and sequence

Article abstract of DOI:10.1007/s11746-007-1185-7

Using ester-forming reactions such as carbodiimide coupling and a modified Yamaguchi symmetrical anhydride method, a variety of estolides based on 17-hydroxy oleic and 17-hydroxy stearic acid have been prepared. These hydroxy fatty acids are produced in good yields from hydrolysis of sophorolipids, which are in turn derived from fermentation of fats and oils. Since the estolides are formed one unit, or ester bond, at a time, their length and sequence can be precisely controlled. The key to this control is the use of protecting groups at either the carboxylic or hydroxy end of the starting hydroxy fatty acids. Two mono-protected dimers, for example, when combined in a fragment-condensation approach, give a tetramer with no contamination from estolides of other lengths. This methodology opens the way to functionalized estolides, and several variants were prepared: hybrid estolides, containing non-fatty acid moieties such as amino acids; polymerizable estolides, containing a norbornene unit; and non-linear estolides that extend from a branched core such as glycerol or pentaerythritol. With the benzoyl chloride-mediated symmetrical anhydride method, yields for individual coupling steps ranged from 75 to 93%.

Full text of DOI:10.1007/s11746-007-1185-7

J Am Oil Chem Soc (2008) 85:277–284
DOI 10.1007/s11746-007-1185-7
ORIGINAL PAPER
Structured Estolides: Control of Length and Sequence
Jonathan A. Zerkowski Æ Alberto Nun˜ez Æ
Daniel K. Y. Solaiman
Received: 21 September 2007 / Revised: 20 November 2007 / Accepted: 30 November 2007 / Published online: 19 December 2007
Ó AOCS 2007
Abstract Using ester-forming reactions such as carbo-
diimide coupling and a modified Yamaguchi symmetrical
anhydride method, a variety of estolides based on 17-
hydroxy oleic and 17-hydroxy stearic acid have been pre-
pared. These hydroxy fatty acids are produced in good
yields from hydrolysis of sophorolipids, which are in turn
derived from fermentation of fats and oils. Since the es-
tolides are formed one unit, or ester bond, at a time, their
length and sequence can be precisely controlled. The key to
this control is the use of protecting groups at either the
carboxylic or hydroxy end of the starting hydroxy fatty
acids. Two mono-protected dimers, for example, when
combined in a fragment-condensation approach, give a
tetramer with no ‘‘contamination’’ from estolides of other
lengths. This methodology opens the way to functionalized
estolides, and several variants were prepared: hybrid es-
tolides, containing non-fatty acid moieties such as amino
acids; polymerizable estolides, containing a norbornene
unit; and non-linear estolides that extend from a branched
core such as glycerol or pentaerythritol. With the benzoyl
chloride-mediated symmetrical anhydride method, yields
for individual coupling steps ranged from 75 to 93%.
Introduction
Estolides, which are intermolecular esters of fatty acids,
appear to have significant promise as functional fluids.
They possess good properties for serving as lubricants [1]
and have the benefit of being biodegradable [2]. A more
specialized use is as a lipid-solubilizing auxiliary for drug
delivery [3]. They can be made from a number of vegetable
oils, such as meadowfoam, castor, and cuphea, as well as
from oleic acid [4–6]. Preparation of estolides has been
achieved principally in two ways. The more widely
investigated route uses acid catalysis, either liquid or solid,
to form the ester unit. Lipases have also been used [7].
Both these methods are capable of inexpensively preparing
large amounts of product, but they do have limitations.
With acid catalysis, the harsh conditions (perchloric or
sulfuric acid) will destroy a great many functional groups
that could be of use if inserted into an estolide chain. These
same functional groups might also be incompatible with
enzymatic routes, for example if they hinder access to
active sites or are themselves substrates. Furthermore, both
kinds of routes are in principle subject to equilibria that
regenerate monomer, if water is not removed (although
studies have shown that the back reaction is slow [8]). In
short, the methods used to make estolides to date do not
allow for precise control of the length of the products (they
are not monodisperse) or for control of their sequence, and
many moieties cannot be included.
Keywords Hydroxy fatty acids ꢀ Estolides ꢀ
Sophorolipids ꢀ Oligoesters ꢀ Glycerol
Mention of trade names or commercial products in this publication is
solely for the purpose of providing specific information and does not
imply recommendation or endorsement by the US Department of
Agriculture.
The aim of the work presented in this paper is twofold.
First, we wished to apply simple, known esterification
methods and synthetic strategies (Fig. 1) to fatty acids that
would permit a wide range of functionalized estolides, of
precisely defined length and sequence, to be formed. Sec-
ond, we wished to demonstrate the utility of an abundant
hydroxy fatty acid, compound 1, that is obtained from
J. A. Zerkowski (&) ꢀ A. Nun˜ez ꢀ D. K. Y. Solaiman
USDA, ARS, Eastern Regional Research Center,
600 E. Mermaid Lane, Wyndmoor, PA 19038, USA
e-mail: jonathan.zerkowski@ars.usda.gov
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J Am Oil Chem Soc (2008) 85:277–284
Fig. 1 Generalized scheme for the fragment-condensation approach
to estolides of defined length and sequence. ‘‘Chain’’ could be a fatty
acid unit or, in principle, a wide variety of moieties. PG1 and PG2
(protecting groups) need to be removable independently of each other
(‘‘orthogonal’’)
sophorolipids [9]. This compound is an isomer of ricinoleic
acid, without (to the best of our knowledge) any of the
toxicity issues associated with that molecule’s source, and
we expect that it should be useful in as many situations as
ricinoleic acid has been. The position of the hydroxy group
at the far end of the chain (x - 1) from the carboxy group
may, in addition, impart novel properties to these estolides
by contrast to those prepared previously, where the ester
linkage is located mid-chain. We propose that the resulting
estolides will find use in studies on the fundamental
properties of this class of molecules as well as in broad-
ening the scope of applications where they can be
employed, such as in functionalized thin films, coatings, or
macromolecular surfactants. Alternative uses are as GPC
standards or as model compounds for studies of processes
such as crystallization. By analogy with lipids of defined
sequence, we propose the name ‘‘structured estolides’’ for
these compounds.
conditions) on a 2.1 9 150 mm Waters Symmetry C18
3.5 l column, or b) direct injection without an LC column.
Matrix-Assisted Laser Desorption/Ionization mass spectra
with automated tandem time of flight fragmentation of
selected ions (MALDI-TOF/TOF) were acquired with a
4700 Proteomics Analyzer mass spectrometer (Applied
Biosystems, Framingham, MA, USA) in the positive re-
flectron mode. Masses were determined as sodiated adducts
of the products [M + Na]+, using 2,5-dihydroxybenzoic
acid as matrix in a concentration of 10 mg/mL in aceto-
nitrile/water (50:50) with 0.1% TFA. Approximately
0.7 lL of matrix was spotted on the MALDI plate and
allowed to dry. The sample (0.5–1 lL) dissolved in chlo-
roform or acetone at a concentration of 1–2 mg/mL was
spotted on the top of the matrix crystals. Averages of
1,000–2,000 spectra were acquired for optimal signal to
noise ratio.
Carbodiimide-Mediated Ester Formation
Experimental Procedures
The general procedure is to dissolve roughly equimolar
amounts of the hydroxy component and the carboxylic
acid component with approximately 1.5 equiv of N-(3-
dimethylaminopropyl)-N0-ethyl carbodiimide hydrochlo-
ride (EDC) and 0.2–0.5 equiv 4-dimethylaminopyridine
(DMAP) in CH2Cl2. A specific example follows: The
benzyl (Bn) ester of the dimer of 1a (‘‘Bn-oleic-oleic-
OH’’, 216 mg, 0.32 mmol), the dimer of 1b, protected as
its tert-butyl diphenylsilyl (tBDPS) ether (‘‘HO2C-stearic-
stearic-tBDPS’’, 225 mg, 0.27 mmol), EDC (80 mg,
0.4 mmol), and DMAP (20 mg, 0.16 mmol) were dis-
solved in 15 mL CH2Cl2 and stirred under an N2
atmosphere at room temperature. After 24 h, another
40 mg of EDC and 20 mg DMAP were added, and the
reaction continued. After 36 h, the solvent was removed
on the rotary evaporator and the material chromatographed
on silica gel with 3:1 hexane/ethyl acetate to afford the
tetramer, Bn-oleic-oleic-stearic-stearic-tBDPS, 310 mg,
78%. Yields for individual coupling steps ranged from 66
to 90%, except for formation of the octamer 2 (see
‘‘Results and Discussion’’).
General
Compound 1 was prepared as previously reported from
sophorolipids [9]. Simple esters of 1 were prepared using
the esterification methods described below, and silyl ethers
of 1 were prepared as previously described [9]. Chemicals
and reagents were obtained commercially from Sigma-
Aldrich (St Louis, MO, USA) and Lancaster Synthesis
(Alfa Aesar, Ward Hill, MA, USA). All solvents and
˚
reagents were used as received. Silica gel (Grade 60 A,
Mesh 230–400, particle size 40–63 lm) used for column
chromatography was obtained from Fisher Scientific
(Fairlawn, NJ, USA). NMR spectra were recorded at room
temperature in CDCl3 on either a Varian Associates
(Walnut Creek, CA, USA) Gemini 200 MHz or Inova
400 MHZ instrument and are referenced to Si(CH3)4.
LCMS data were recorded on a Waters/Micromass (Mil-
ford, MA, USA) ZMD instrument with APCI, using a) an
elution gradient of 40:60 water/acetonitrile to 100% ace-
tonitrile over 30 min (or minor variations on those
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J Am Oil Chem Soc (2008) 85:277–284
279
Benzoyl Chloride-Mediated Ester Formation
propionic), 2.56 (m, 2H, C(O)CH2 3-hydroxybutyrate
fragment), 3.50–3.78 (m, 2H, hydroxyproline N-CH2), 3.67
(s, 3H, Me ester), 4.43 (q, 7.7 Hz, 1H, hydroxyproline
C(a)H), 4.90 (hex, 6.4 Hz, 7H, C(17)H and hydroxyproline
CH-O), 5.15 (m, 2H, benzylic), 5.22–5.44 (m, 7H, olefinic
H and 3-hydroxybutyrate CH-O), 7.35 (m, 5H, Ar). 13C:
9.3 (terminal propionic CH3); 19.7, 19.9, and 20.1 (C18’s);
24.9, 25.0, 25.2, 25.4, 25.5, 27.3, 28.1; 29.2–29.8
(numerous peaks); 34.2, 34.3, 34.6, and 34.9 (C2’s); 35.8,
36.0, 36.1 (C16’s); 41.4 (C2 of 3-hydroxybutyrate); 51.5
(Me ester), 52.3 (hydroxyproline C(5)-N), 58.0 (hydroxy-
proline Ca); 67.3 and 67.4 (benzyl CH2 and C3 of 3-
hydroxybutyrate); 70.8, 70.9, 71.5, 71.7, and 72.4 (C17’s);
127.9, 128.1, 128.5, 128.6, 129.9, 136.5, 154.8 (hydroxy-
proline N-C(O)), 170.0, 171.7, 171.9, 173.0, 173.3, 173.7,
174.2, 174.4. MALDI: calculated 2131.7 for
The general procedure [10] is to dissolve equimolar
amounts (although other ratios can be used) of the alcohol
and carboxylic acid in THF with at least 2 equiv of
diisopropylethylamine (DIEA), then add 1 equiv benzoyl
chloride, then 20 mol% DMAP. A specific example fol-
lows: The 2,2,2-trichloroethyl ester of 1a (600 mg,
1.4 mmol), the tert-butyl dimethylsilyl (tBDMS) ether of
1a (free acid, 495 mg, 1.2 mmol), and DIEA (522 lL,
3.0 mmol) were dissolved in 15 mL THF. Benzoyl chlo-
ride (139 lL, 1.2 mmol) was then added dropwise. After
5 min, DMAP (30 mg, 0.2 mmol) was added. The reaction
was stirred overnight. Solvent was removed on the rotary
evaporator, the residue applied to a silica gel column, and
eluted with 3:1 hexane/ethyl acetate to afford the tBDMS
trichloroethyl dimer (923 mg, 1.1 mmol, 93%). Unreacted
trichloroethyl ester was also recovered. Yields for indi-
vidual coupling steps ranged from 75 to 93%.
C
129H225NO20 ꢀ Na+, found 2131.8 [M + Na]+.
5. 1H: 0.9 (t, 6.5 Hz, 3H, terminal CH3), 1.21 (d, 6.2 Hz,
6H, internal C(18)H3), 1.30 (br s, 68H), 1.41–1.52 (m, 4H),
1.52–1.63 (m, 7H), 1.92–2.11 (m, 5H, allylic and nor-
bornene), 2.21–2.42 (m, 6H, C(O)CH2), 2.95 (m, 1H,
norbornene), 3.00–3.17 (m, 2H, norbornene), 3.46 (m, 2H,
piperazine), 3.54–3.73 (m, 6H, piperazine), 4.91 (hex,
6.2 Hz, 2H, C(17)H), 5.36 (m, 2H olefin), 6.06 (dd, 2.9 and
5.5 Hz, 1H) and 6.21 (dd, 3.0 and 5.5 Hz, 1H) both nor-
bornene olefinic. 13C: 14.2 (terminal C18), 20.1 (internal
C18), 22.8, 25.2, 25.4, 25.5, 27.3, 29.2–29.8 (numerous
peaks), 31.1, 32.0, 33.4, 34.8, 36.0 (C16), 41.8 (broad,
piperazine), 42.2 and 42.7 (norbornene), 45.5 (broad,
piperazine), 45.8 and 49.6 (norbornene), 70.8 (C17), 129.9
(oleic C = C), 133.0 and 136.9 (norbornene C = C), 172.1,
173.4, 173.6. APCI: calculated 1035.9 for C66H116N2O6
H+, found 1035.2 (MH+).
6. 1H: 1.14–1.23 (m, 9H, C(18)H3), 1.23–1.44 (m, 48H),
1.51 (s, 9H, Boc) 1.53–1.74 (m, 15H), 1.80–2.15 (m, 13H,
allylic and norbornene), 2.22–2.39 (m, 6H, C(O)CH2),
2.87–3.00 (m, 2H, norbornene), 3.22 (m, 1H, norbornene),
3.37–3.51 (m, 6H, piperazine), 3.55–3.66 (m, 2H, pipera-
zine), 4.75–5.00 (m, 3H, C(17)H), 5.34 (m, 6H, olefinic H),
5.93 (m, 1H) and 6.19 (m, 1H), both norbornene olefinic.
13C: 20.1 (C18), 25.2, 25.4, 25.5, 27.3, 28.5, 29.1–29.7
(numerous peaks), 33.5, 34.8, 36.0, 41.4 (piperazine), 42.7
and 43.6 (norbornene), 43.8 (broad, piperazine), 45.5
(piperazine), 45.9 and 49.8 (norbornene), 70.7 and 70.8
(C17), 80.3 (Boc quaternary C), 129.9 (oleic C = C), 132.3
and 137.7 (norbornene C = C), 154.7 (Boc C = O), 172.0,
Spectroscopic and Physical Data
2. 1H: 1.09 (s, 9H, Si–t-Bu), 1.21 (d, 6.5 Hz, 24H,
C(18)H3), 1.24–1.42 (br s, 176H), 1.49–1.75 (m, 32H),
1.83–2.11 (m, 8H, allylic H), 2.22–2.42 (m, 16H,
C(O)CH2), 3.84 (hex, 6.1 Hz, 1H, terminal C(17)H-OSi),
4.91 (hex, 6.4 Hz, 7H, internal C(17)H), 5.13 (s, 2H,
benzylic H), 5.35 (m, 4H, olefinic H), 7.33–7.51 (m, 11H,
SiPh2 and benzyl Ar), 7.70 (m, 4H, SiPh2). 13C: 20.1 (C18),
25.2, 25.5, 27.1, 27.3, 29.2–29.8 (numerous peaks), 34.9
(C2), 36.1 (C16), 66.1 (C17-OSi), 69.7 (benzyl CH2), 70.8
(C17 esterified), 127.4, 128.2, 128.3, 128.6, 129.6, 129.9,
132.7, 135.9, 173.6. MALDI: calculated 2623.2 for
C
167H294O17Si ꢀ Na+, found 2623.4 [M + Na]+.
3. 1H: 0.05 (s, 24H, SiCH3), 0.89 (m, 39H, Si(t-Bu) and
propionamide CH3), 1.13 (d, 6.0 Hz, 12H, C(18)H3 adja-
cent to OtBDMS), 1.20 (d, 6.5 Hz, 9H, C(18)H3 adjacent
to esters), 1.19–1.44 (m, 152H), 1.44–1.72 (m, 40H), 2.18
(t, 6.5 Hz, 2H, N-C(O)CH2), 2.22–2.45 (m, 12H, O-
C(O)CH2), 3.23 (q, 6.9 Hz, 2H, CH2N), 3.71–3.85 (m, 4H,
C(17)H-OSi), 4.79–4.97 (m, 6H, esterified C(8)H, C(11)H,
C(17)H), 5.56 (br s, NH). 13C: -4.5 and -4.2 (SiMe2),
14.4 (propionamide CH3), 18.4 (C18 adjacent to OSi), 20.2
(C18 adjacent to esterified C17), 22.9, 23.1, 24.0, 25.4,
25.5, 26.0, 26.1, 29.4–29.9 (numerous peaks), 34.3, 34.4,
34.9, 36.1, 40.0, 41.4 (CH2-NH), 60.6 (C17-OSi), 68.8,
70.8, 74.1, 173.4, 173.9. MALDI: calculated 2562.2 for
173.6,
174.4.
APCI:
calculated
1147.9
for
C71H122N2O9ꢀH+, found 1147.5 (MH+).
C
153H303NO17Si4 ꢀ Na+, found 2562.3 [M + Na]+.
1
7. H: 0.95 (d, 6.5 Hz, 18H, Leu CH3), 1.20 (d, 6.2 Hz,
18H, C(18)H3), 1.25–1.41 (br s, 96H), 1.48 (s, 27 H, Boc),
1.50–1.81 (m, 33H), 1.89–2.12 (m, 24H, allylic H), 2.21–
2.48 (m, 12H, C(O)CH2), 4.03–4.41 (m, 7H, glycerol CH2
and Leu C(a)H), 4.72–5.01 (m, 9H, C(17)H and NH), 5.22–
1
4. H: 1.04–1.22 (m, 21H, C(18)H3 and terminal pro-
pionic CH3), 1.22–1.44 (br s, 123H), 1.44–1.76 (m, 24H),
1.89–2.11 (m, 14H, allylic H and C(3)H2 from hydroxy-
proline), 2.20–2.37 (m, 14H, C(O)CH2 including
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J Am Oil Chem Soc (2008) 85:277–284
5.47 (m, 13H, olefinic H and glycerol CH). 13C: 20.0 and
20.1 (C18), 24.9, 25.1, 25.4, 25.5, 27.2, 28.4, 29.2–29.7
(numerous peaks), 34.1, 34.8, 35.9, 36.0, 42.1, 52.4 (Leu
Ca), 62.2 (glycerol CH), 69.0 (glycerol CH2), 70.7 and
72.2 (C17), 79.6 (Boc quaternary), 128.5, 129.9, 133.4,
155.4 (Boc C = O), 172.9, 173.2, 173.3, 173.5. MALDI:
calculated 2435.9 for C144H257N3O24ꢀNa+, found 2435.8
[M + Na]+.
8. 1H: 0.89 (t, 6.9 Hz, 9H, terminal CH3), 1.21 (d,
6.4 Hz, 9H, C(18)H3), 1.24–1.42 (br s, 132H), 1.48 (s,
18H, Boc and OtBu), 1.50–1.73 (m, 20H), 1.85–2.16 (m,
12H, allylic H), 2.22–2.49 (m, 14H, C(O)CH2 including
Glu side chain), 4.14 (s, 6H, pentaerythritol CH2 esterified
with estolide), 4.21 (s, 2H, pentaerythritol CH2 esterified
with Glu), 4.23–4.38 (m, 1H, C(a)H), 4.91 (hex, 6.3 Hz,
3H, C(17)H), 5.11 (d, 8.3 Hz, 1H, NH), 5.36 (m, 6H,
olefinic H). 13C: 14.2 (terminal C18), 20.1 (internal C18),
22.8, 24.9, 25.2, 25.5, 27.3, 28.1, 28.4, 29.2–29.8
(numerous peaks), 32.0, 34.2, 34.9, 36.1 (C16), 53.3 (Glu
Ca), 62.0 (pentaerythritol), 70.8 (C17), 80.9 (Boc quater-
nary), 129.9, 130.1, 155.4 (Boc C = O), 172.0, 173.3,
173.7. MALDI: calculated 2083.7 for C127H233NO18 ꢀ Na+,
found 2083.6 [M + Na]+.
tetramer, which was in turn used to prepare octamer 2.
While we chose to use ‘‘symmetrical’’ fragment conden-
sations, e.g. dimer plus dimer to give tetramer, then
tetramer plus tetramer to give octamer, this method should
be perfectly applicable to other combinations, such as
dimer plus trimer to give pentamer. Although we have not
prepared odd-numbered linear oligomers through fragment
condensation in this work, the next example demonstrates a
3 + 3 + 1 fragment condensation to give a heptamer.
Using dihydroxy fatty acids with this reaction scheme
should lead to branched estolides. We used the allylically
hydroxylated versions of 1a reported previously [9], after
separating the C8, C17 and C11, C17 dihydroxy isomers
and hydrogenating the olefin. First, a trimer was prepared
from the benzyl ester of the C11, C17 diol and two
equivalents of the tert-butyl dimethylsilyl (tBDMS) ether
of 1b (see Fig. 2). Meanwhile the propyl amide of the C8,
C17 diol was prepared—this amide serves as a convenient
NMR tag. Coupling of two equivalents of the free acid
(after deprotective hydrogenation) of the trimer with the
propyl amide diol led to the desired heptamer 3. Two tet-
ramers, monoacylated at either the C8 or C17 hydroxy
groups, were also isolated. Interestingly, these isomers
could be separated from each other with silica gel
chromatography.
Concerning the physical appearances of the products,
compounds 2, 3, 6, and 7 were thick syrups or glasses at
room temperature. Compounds 4 and 8 were sticky waxes
that melted in the range of 35–40 °C, while compound 5
was a more friable solid that melted at 50–55 °C.
Overall, while the EDC route did give estolide products,
it had the shortcoming of being sluggish: especially with
larger molecules such as tetramers, even after 3 days
starting material remained. Octamer 2 was formed in only
25% yield. Steric hindrance is presumably partially
responsible for this poor reactivity. Adding more EDC did
not solve the problem. Nonetheless, EDC-based prepara-
tion of short estolides is simple to perform, and column
chromatography suffices to remove impurities.
Results and Discussion
Carbodiimide Couplings
Our initial efforts at forming estolides from 1 focused on
the well-known reagents for forming ester or amide bonds,
the carbodiimides [11, 12, and references therein]. To form
estolides of a specific length, the strategy was to use pro-
tecting groups on either the carboxy or hydroxy ends to
limit reaction to one ester unit, as outlined in Fig. 1. These
procedures are totally analogous to those used commonly
in peptide chemistry, and they have in fact been applied to
oligoesters [13]. In principle, any of a number of pairs of
protecting groups could be employed; for these initial
experiments we chose benzyl esters and tBDPS ethers.
Coupling of the benzyl ester of 1 and the tBDPS ether of 1
led to the diestolide, protected at both ends. Splitting this
material into two portions and treating them with selective
deprotecting reagents (hydrogenation over Pd/C to remove
the Bn ester, or fluoride to remove the silyl ether) gave (a)
the benzyl diestolide with a free hydroxy group and (b) the
tBDPS diestolide with a free carboxy group. These two
dimers could then be recoupled with EDC to give the
Benzoyl Chloride Activation
Our second attempt at finding a coupling method led to
better results. We opted for a variation on the Yamaguchi
coupling, where a carboxylate anion initially couples with
benzoyl chloride (see Fig. 3), then with another equivalent
to give the symmetrical anhydride [10]. This route was
more successful for forming estolides (Fig. 4). By way of
comparison to the EDC method, we repeated the synthesis
of 2 using the recovered starting materials from the EDC
run. With the benzoyl chloride route, the reaction gave a
yield of 75% after 3 days of reaction. More significantly,
we chose to explore the flexibility of this reaction by
forming a ‘‘hybrid’’ estolide, the reaction sequence for
which is sketched in Fig. 5. In principle, any hydroxy
carboxylic acid can be used with this route to build a chain.
We therefore chose to incorporate two non-fatty hydroxy
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J Am Oil Chem Soc (2008) 85:277–284
281
Fig. 2 Estolides prepared using
the EDC coupling method from
starting material 1a/b. The
shorthand ‘‘oleic/stearic’’
nomenclature is shown in the
upper right corner: a residue
runs from the carbonyl group at
one terminus to the hydroxyl at
the other. Whenever this
shorthand is used, the carbonyl
is positioned at the left and the
hydroxyl at the right. In
heptamer 3, a trimer unit is
boxed. See text for
abbreviations
Fig. 3 Outline of the proposed
mechanism for benzoyl
chloride-mediated ester
formation (see [10]). Side
products are shown
parenthetically; B is a
trialkylamine base
Fig. 4 Functionalized and branched estolides prepared using the benzoyl chloride route. The eight residues of 4 are numbered, and the
pentaerythritol core of 8 is in bold
acids—3-hydroxybutyrate and 4-hydroxyproline—into a
chain otherwise composed of 17-hydroxy C18 chains. In
addition, we planned to segregate the C18 units by type
along the chain, so that oleic residues were clustered on
one half and stearic on the other. The protecting groups had
to be chosen carefully, therefore, since benzyl esters would
be incompatible with oleic acid. Instead, the trichloroethyl
ester (removed with Zn dust [14]) was used for the oleic
congener, and benzyl for the stearic. These esters of 1 were
also prepared using the same benzoyl chloride methodol-
ogy. For the hydroxy protecting group, tBDMS removed
with trifluoroacetic acid was used throughout. A similar
fragment condensation strategy as for the homooctamer 2
was used, that is, dimer plus dimer gives tetramer (Fig. 5),
and tetramer plus tetramer (Fig. 6) gives the hybrid oct-
amer 4. We noted a marked improvement over the EDC
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J Am Oil Chem Soc (2008) 85:277–284
Fig. 5 Early and middle steps of the synthetic route to heterooctamer
estolide 4. To conserve space, the explicit molecular formula is drawn
out for monomers (at left) and a condensed nomenclature (see Fig. 2)
is employed for products. D and T refer to dimer and trimer,
respectively. Reagents and conditions. a Benzoyl chloride, DIEA,
DMAP; b 9:1 CF3CO2H:H2O; b Zn dust, pH 7.2 phosphate buffer;
d H2 over Pd/C in MeOH
1
Fig. 6 Comparison of the H-
NMR spectra of the two
precursor tetramers (see Fig. 5)
and their coupling product 4.
Prior to coupling, T-1 is
deprotected at its hydroxy
terminus with aqueous
CF3CO2H, T-2 is deprotected at
its carboxy terminus by
hydrogenation over Pd/C, then
the deprotected tetramers are
coupled using benzoyl chloride
as described in the text. The
asterisk indicates BHT or ethyl
acetate impurities
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J Am Oil Chem Soc (2008) 85:277–284
283
couplings: most of the reactions were essentially complete
in about 2 h. On the other hand, the reactions did appear to
be slower for coupling of longer chains, just as for the EDC
route, although we have not studied the kinetics in a sys-
tematic way. The cleanliness of the reaction was a further
benefit: product, hydroxy starting material, and carboxylic
starting material were readily separable from each other by
column chromatography on silica gel, using hexane/ethyl
acetate mixtures (Rf’s high, mid, and low, respectively).
This feature allowed the reaction to be driven to complete
usage of one component if desired, such as by using an
excess of the hydroxy component to be sure to consume all
of a valuable carboxylic component, while still being able
to recover the unreacted hydroxy component by chroma-
tography. Yields of the individual coupling steps for all the
benzoyl chloride reactions ranged from 75 to 93%.
core was pentaerythritol, which was first monoacylated
with BocGlu(OtBu) (this reaction required use of N,N-
dimethylformamide as solvent for the pentaerythritol), then
reacted with excess stearic-oleic diestolide. This 3:1
ratio of substituents in 8 was chosen to aid in NMR
identification, as well as to demonstrate the range of
functionalizability, but the fully symmetric quadruply
substituted variant should also be accessible. In both these
examples, as well as 4, there is nothing privileged about
protected amino acids, but they simply are representative
of the kinds of reactive groups that can be incorporated into
non-traditional estolide structures.
Acknowledgments We gratefully acknowledge the technical
assistance of Mr. Bun-Hong Lai and Ms. Krista Sirois (chromatog-
raphy and fermentations), Mr. Marshall Reed (microbiology), and
Dr. Gary Strahan (NMR).
With the successful synthesis of heterooctamer 4, we
turned out attention to other sorts of estolides. A desirable
feature to include in an estolide is the possibility for further
reactivity, such as polymerization. The norbornene unit has
been used extensively with ring-opening metathesis poly-
merization [15], so we chose to incorporate this building
block in estolides. Molecules 5 and 6, both trimers, were
synthesized analogously to 4, using trichloroethyl ester
protection. Molecule 6 shows that, with the polymerizable
unit at the hydroxy terminus, the carboxy terminus can be
used to append other functionality of interest; here, the Boc
group could be removed to afford a basic site. Polymeri-
zation of 5 or 6 would afford a comb-type polymer, where
the length and functionality of the side chains could be
controlled by altering the structure of 5 or 6.
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Brief mention should be made of the concept of ‘‘esto-
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characterize estolides. For oligomers of the sort monomer-
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percentage of the bulk samples, which have an average EN
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ambiguous to assign a value to compound 3 (or 7 or 8, see
below). Alternative definitions, for example the number of
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functionalized heterooligomers the question of which ester
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