Crystal Growth & Design
ARTICLE
gently refluxing in 100 mL of 95% ethanol with Amberlyst 15 (wet) resin.13
The resulting MAG was purified by recrystallization first from acetone and
subsequently from hexane/ethyl ether (4:1, v/v). This yielded 7.56 g of 93%
3-palmitoyl-sn-glycerol containing 7% 2-palmitoyl-glycerol (by GC).
Enantiopure 1,2-Bisdecanoyl-3-palmitoyl-sn-glycerol. Decanoyl
chloride (3.27 g) was dissolved in 30 mL of methylene chloride and
then added dropwise to a solution of 3-palmitoyl-sn-glycerol (2.56 g),
triethylamine (2.90 g), and N,N-dimethylaminopyridine (0.18 g) in
70 mL of methylene chloride stirred in an ice bath. After addition, the
solution was stirred at room temperature (∼22 °C) for 24 h. Solvent was
removed under vacuum, and the residue was taken back up in hexane and
filtered. A total of 3.7 g of pure product (>99% by GC) was obtained by
flash chromatography with hexane/ethyl acetate (10:1, v/v).
Figure 1. Characteristic liquidus lines for phase diagrams of (a) racemic
compounds, (b) conglomerates, and (c) pseudoracemates (ideal behavior).
Binary mixtures of an enantiopure compound (E) and racemic mixture (R)
are used to prepare half of the curve. Data can be extrapolated to include the
opposite enantiomer (E0) if desired.
Racemic 1(3)- Palmitoyl-rac-glycerol. The vinyl ester of palmitic acid
(13.4 g) and racemic isopropylideneglycerol (5.0 g) were stirred in
100 mL of chloroform containing Lipozyme immobilized lipase from
Rhizomucor miehei (1.0 g) at room temperature (∼22 °C) under nitrogen
for 3 days.14 The immobilized enzyme was then removed by filtration, and
free 1(3)-palmitoyl-rac-glycerol was produced as for the enantiopure
3-palmitoyl-sn-glycerol. This yielded 7.56 g of very pure (by TLC) 1(3)-
palmitoyl-rac-glycerol.
Racemic Bisdecanoyl-1(3)-palmitoyl-rac-glycerol. Same method as
with the enantiopure 1,2-bisdecanoyl-3-palmitoyl-sn-glycerol but using
racemic 1(3)-palmitoyl-rac-glycerol as a starting material. Crystallized
from hexane (ꢀ20 °C) followed by flash chromatography with hexane/
ethyl acetate (10:1, v/v) to yield 2.45 g of pure product (>99% by GC).
Gas Chromatography. Prepared compounds were dissolved in
iso-octane and analyzed by GC/FID to determine their purity. These
were eluted on a 25 m ꢁ 0.25 mm polarizable capillary column
(Quadrex, Woodbridge CT).12 The column was housed in a Hewlett-
Packard 5890 (Agilent, Palo Alto CA) GC equipped with FID and on-
column inlet. The inlet pressure for the carrier gas (hydrogen) was set to
15 psi, cool on-column injection was employed, and the detector was
held at 370 °C. After sample injection, the oven was held at 60 °C for 2
min, then the temperature was increased to 250 at 35 °C/min, and finally
the temperature was increased to 350 at 4 °C/min.
Nuclear Magnetic Resonance. Approximately 15 mg of sample
for 1H NMR and 100 mg for 13C NMR dissolved in 750 μL of deuterated
chloroform (containing 0.5% tetramethylsilane (TMS) as internal standard)
was placed in a suitable NMR tube (Wilmad, Buena, NJ). NMR spectra were
obtained using an Avance III 400 MHz instrument (Bruker, Billerica, MA)
(1H 4 scans; 13C 256 scans). Chemical shifts were measured relative to
TMS internal standard. NMR spectra were integrated, analyzed using 1D
NMR Processor, Academic Version (ACD, Toronto), and interpreted using
relevant literature.12,15
Infrared Spectroscopy. Data were collected using a Shimadzu
IRPrestige-12 FTIR (Kyoto) equipped with a Pike MIRacle ATR sample
stage (Madison, WI). Data were analyzed using the Shimadzu IR Solution
1.30 software package.
X-ray Powder Diffraction. Solid samples were ground into fine
powders and applied to sample slides with a 0.2 mm deep depression
(Rigaku, Tokyo, JP). An aftermarket temperature controller utilizing Peltier
cooling (Electron Dynamics, Southampton UK) maintained the sample
stage at 20 °C. Powder diffraction data was recorded using a Rigaku Multiflex
Powder X-ray Diffractometer with a Cu KR radiation (λ = 1.5406 Å)16
source and a scintillation detector. Samples were scanned from 2° to 30° at
2°/min. Results were analyzed and major peaks were identified using Jade
software (Materials Data, Livermore CA).
Figure 2. Stereochemistry of (a) (S)-1,2-isopropylidene glycerol and
(b) (R)-1,2-bisdecanoyl-3-palmitoyl-sn-glycerol. Chiral carbon at the sn-
2 position of glycerol is marked with an asterisk. Hydrogen atom displays
stereochemistry at the sn-2 position.
mixture. The crystallization and polymorphism of enantiopure TAG
and their binary mixtures has not previously been published.
The main purpose of this work is to compare the physical
chemistry of an enantiopure TAG with its racemic counterpart. This
will improve understanding of the role stereochemistry plays in
crystallization and perhaps lend some insight into the polymorphism
of TAG. 1,2-Bisdecanoyl-3-palmitoyl-sn-glycerol and bisdecanoyl-1
(3)-palmitoyl-rac-glycerol were chosen as models for study because
the difference in chain length was regarded as sufficient for the
potential effects of chirality to be detectable.
’ MATERIALS AND METHODS
Unless noted otherwise, reagents, chemicals, and enzyme were pur-
chased from Sigma-Aldrich (Mississauga, ON) and were of the highest
practical grade; solvents were purchased from Fisher Scientific (Ottawa ON)
and were HPLC grade. Decanoyl chloride (10:0), palmitoyl chloride, and iso-
propylideneglycerol all had a declared purity g98%. Enantiopure 1,2-iso-
propylideneglycerol was 99.96% pure (by GC) and had an optical purity of
99.8% according to the certificate of analysis (available from Sigma Aldrich;
product number 237744; lot number 1328499). The vinyl ester of palmitic
acid donated by Japan VAM & POVAL (Osaka, JP) was g96% purity. Prior
to analysis, pure TAG were crystallized and tempered by holding isothermally
at 25 °C for more than 28 days to ensure formation of the highest-melting
form. These tempered samples were used for gas chromatographic analysis,
infrared and X-ray spectroscopy, and differential scanning calorimetry
experiments. Binary mixtures of TAG were stored at 22 °C for 3 days prior
to analysis.
Synthesis. Enantiopure 3-Palmitoyl-sn-glycerol. Palmitoyl chlor-
ide (12.5 g) was dissolved in 30 mL of methylene chloride and then added
dropwise to a solution of 1,2-isopropylidineglycerol (5.0 g) (Figure 2),
triethylamine (7.66 g), and N,N-dimethylaminopyridine (0.46 g) in 70 mL
of methylene chloride stirred in an ice bath. After addition, the solution was
stirred at room temperature (∼22 °C) for an additional 3 h. The solution
was then washed with brine, dried over sodium sulfate, and filtered, and the
solvent was evaporated.11,12 Free 3-palmitoyl-sn-glycerol was produced by
Small Angle X-ray Scattering. Samples were ground into a fine
powder and sealed between sheets of Kapton. Samples were held at
20 °C in a temperature-controlled stage while data was measured from
0.7° to 4.3° for 1 h using a Nanostar SAXS (Bruker AXS, Madison WI)
with Cu KR radiation and a High Star wire-type detector.16 Results were
analyzed using Bruker AXS software version 4.1.26.
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dx.doi.org/10.1021/cg101654c |Cryst. Growth Des. 2011, 11, 1723–1732