Synthesis and physical properties of EOE and EEO, triacylglycerols containing elaidic and oleic fatty acids

Article abstract of DOI:10.1007/s11746-007-1056-2

Symmetrical and non-symmetrical triacylglycerols (TAG) containing oleic (O; 9c-18:1) and elaidic (E; 9t-18:1) acids were required as part of a study relating the physical characteristics and functionality of trans-containing TAG with the mouth feel, taste characteristics and related characteristics desired by consumers in frying oils and pastries. To replace the trans isomers in frying oils-a significant part of frying oils prepared by partial hydrogenation of vegetable oils-without loss of the sensory properties desired by consumers, required the initiation of a study relating the structure of trans-containing TAG with such characteristics as melting range, drop points, and other crystalline properties. Elaidic acid was esterified to trielaidin (EEE), and the EEE partially converted (glycerol/p-toluenesulfonic acid) to a mixture containing ca. 40% DAG (the 1,3- and 1,2-isomers). The DAG fraction was separated by silica gel chromatography, the 1,3-dielaidylglycerol (1,3EE-DAG) isomer isolated (structural purity >99%) by crystallization from acetone and esterified with oleic acid (O) to yield EOE. The 1(3)O-MAG was purchased commercially and esterified with E acid to prepare OEE. Both syntheses yielded multi-gram quantities of EOE and EEO, in 80-85% yields, and with structural purities >99%. Thus, by careful selection of the thermodynamically more-stable MAG or DAG precursors, the symmetrical EOE and non-symmetrical EEO isomers could be readily synthesized, and their drop point and melting point values determined. AOCS 2007.

Full text of DOI:10.1007/s11746-007-1056-2

J Amer Oil Chem Soc (2007) 84:427–431
DOI 10.1007/s11746-007-1056-2
ORIGINAL PAPER
Synthesis and Physical Properties of EOE and EEO,
Triacylglycerols Containing Elaidic and Oleic Fatty Acids
R. O. Adlof Æ G. R. List
Received: 17 October 2006 / Accepted: 20 November 2006 / Published online: 11 April 2007
ꢀ
AOCS 2007
Abstract Symmetrical and non-symmetrical triacylgly-
cerols (TAG) containing oleic (O; 9c-18:1) and elaidic (E;
EOE and non-symmetrical EEO isomers could be readily
synthesized, and their drop point and melting point values
determined.
9
t-18:1) acids were required as part of a study relating the
physical characteristics and functionality of trans-contain-
ing TAG with the mouth feel, taste characteristics and re-
lated characteristics desired by consumers in frying oils
and pastries. To replace the trans isomers in frying oils—a
significant part of frying oils prepared by partial hydroge-
nation of vegetable oils—without loss of the sensory
properties desired by consumers, required the initiation of a
study relating the structure of trans-containing TAG with
such characteristics as melting range, drop points, and
other crystalline properties. Elaidic acid was esterified to
trielaidin (EEE), and the EEE partially converted (glycerol/
p-toluenesulfonic acid) to a mixture containing ca. 40%
DAG (the 1,3- and 1,2-isomers). The DAG fraction was
separated by silica gel chromatography, the 1,3-dielaidyl-
glycerol (1,3EE-DAG) isomer isolated (structural purity
Keywords Triacylglycerols Á DH Á Melting Á Synthesis Á
Oleic Á Elaidic
Abbreviations
TAG
DAG
MAG
P
Triacylglycerol
Diacylglycerol
Monoacylglycerol
Palmitate; 16:0
E
Elaidate; trans-9-18:1
Oleate; cis-9-18:1
O
L
Linoleate; cis-9, cis-12-18:2
Linolenate; cis-9, cis-12, cis-15-18:2
Ln
Ag-HPLC Silver ion high performance liquid
chromatography
>
99%) by crystallization from acetone and esterified with
NMR
DSC
Nuclear magnetic resonance
oleic acid (O) to yield EOE. The 1(3)O-MAG was pur-
chased commercially and esterified with E acid to prepare
OEE. Both syntheses yielded multi-gram quantities of EOE
and EEO, in 80–85% yields, and with structural purities
Differential scanning calorimetry
>99%. Thus, by careful selection of the thermodynamically
Introduction
more-stable MAG or DAG precursors, the symmetrical
The amounts and types of triacylglycerols (TAGs) in the
oil phase of margarines and spreads are considered
responsible for such properties as spreadability, resis-
tance to water/oil loss and melting at body temperature
[
1], with spreadability dependent on the presence of both
lower- and higher-melting TAGs. Spreads which contain
‘high’’ (>50%) levels of TAGs with ‘‘lower’’ (<10 ꢁC)
The 1- and 3- positions on the glycerol backbone of the MAG, DAG
and TAG molecules are assumed to be equivalent.
‘
R. O. Adlof (&) Á G. R. List
melting values tend to be too soft, while those with
higher-melting (20–30 ꢁC) TAGs tend to be too hard for
good spreadability directly out of the refrigerator.
USDA, ARS, NCAUR, 1815 N. University St,
Peoria, IL 61604, USA
e-mail: Richard.Adlof@ars.usda.gov
1
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J Amer Oil Chem Soc (2007) 84:427–431
Hydrogenated vegetable oils, containing lower levels of
unsaturated fats and elevated levels of trans and satu-
rated fats, have traditionally been used to prepare base
feedstocks with the melting point ranges required [2, 3]
for structure and spreadability at refrigerator tempera-
tures (i.e. 50 ꢁF, 10 ꢁC). During the catalytic hydroge-
nation of edible oils, major TAG such as trilinolein and
trilinolenin are converted to elaidic acid-containing
TAGs such as EEO and EOE, higher-melting TAG
which contribute to the desired functional properties
(Phillipsburg, NJ, USA). All solvents were either HPLC
Grade (acetone, acetonitrile, methanol) or ACS Grade
[benzene, carbon tetrachloride, ethyl ether (EE), petroleum
ether (PE)], and were used as received.
Methods
1. Ag-HPLC: A Spectra-Physics P2000 solvent delivery
system (Spectra-Physics Analytical, San Jose, CA,
USA/now Thermo-Finnigan),
a Rheodyne 7125
(
spreadability) of the resultant spreads.
injector (Rheodyne, Inc., Cotati, CA, USA) with a
20 lL injection loop, and an ISCO V4 Absorbance
Detector (ISCO, Inc., Lincoln, Nebraska, USA) at a
wavelength of 206 nm was used. (A representative Ag-
HPLC chromatogram of a TAG fraction containing
Health, consumer-driven and recent FDA labeling
mandates have stimulated research aimed at reducing the
levels of trans and long-chain saturated fatty acids in
shortening and frying oils by such procedures as inter-
esterification, the blending of tropical and liquid vegeta-
ble oils, low-temperature fractionation and, more
recently, by development of structurally modified oils by
transgenic or conventional plant breeding methods [4].
After interesterification, as much as 50% of the sym-
metrical TAGs of structure ABA are converted to non-
symmetrical TAGs of structure AAB, a change which,
when coupled with lower levels of saturated fatty acids
both the EEO and EOE isomers is shown in Fig. 1.)
ꢀ
The ChromSpher
Lipids columns (4.6 mm
I.D. · 250 mm stainless steel; 5 micron particle size;
silver ion impregnated) were purchased from Varian-
Chrompack Int., Middelburg, The Netherlands, and
used as received.
2. Thin-Layer Chromatography (TLC): Formation of the
mono- or dielaidoyl and oleoylglycerol intermediates
and the final TAGs were followed by TLC [3].
Samples (5–10 mg) were dissolved in 1.0 mL hex-
ane and applied (ca.10 lL) to 1 in. · 3 in. silica gel
TLC [K6] plates (Whatman, Inc., Clifton, NJ, USA).
Eluting solvent: 80:20—Hexane: Ethyl Ether (v/v;
(
FAs) and the elimination of trans isomers, could account
for the observed changes in melting point ranges, solid
fat content profiles and other parameters contributing to
the poor ‘‘mouth feel’’ and loss of flavor observed in
commercially-available ‘‘low-trans’’ (<5%) TAG formu-
lations [1]. By careful selection of mono- or diacyl-
glycerol substrates, both the symmetrical (EOE) and
nonsymmetrical (EEO) TAG isomers can be chemically
synthesized [5, 6], their purities readily determined by
silver-ion HPLC (Ag-HPLC), and their drop point [7] and
heats of fusion values measured. These values, currently
not available, will be utilized as part of a study corre-
lating the sensory properties of TAG formulations with
TAG structures (ABA vs. AAB, etc.) and TAG FA
[8]); visualization by I vapor or by spraying with
2
10% CuSO in 8% H PO solution/ heating to120 ꢁC
4
3
4
(hot plate).
3. Gas Chromatography: Chemical purities of the 1(3)-
MAG or 1,3-DAG precursors and of the synthesized
TAGs were determined by gas chromatography after
conversion (5% HCl in methanol) to FAMEs [9]. A
Varian 3400 Gas Chromatograph (GC; Varian
Instruments, Palo Alto, CA, USA) equipped with a
100 m · 0.32 mm SP2380 (Supelco, Inc., Bellefonte,
PA, USA) capillary column, flame ionization detec-
tor (FID) and He as carrier gas (operating condi-
tions: injector, 240 ꢁC; split ratio, 100:1; oven
temperature programmed from 155 to 220 ꢁC at
3 ꢁC/min with an initial hold of 15 min; detector,
(
saturated vs. cis vs. trans) compositions.
Experimental
Materials
280 ꢁC) was utilized.
Oleic (O) and elaidic (E) acids, and 1-monoolein were
purchased from Nu-Chek-Prep (Elysian, MN, USA).
Sodium methoxide was obtained from Harshaw Chem. Co.
4. Acylglycerol Structures: The structural purities of the
isolated or purchased MAG and DAG starting mate-
rials (after conversion to di- or mono-acetate(s),
respectively) and of the synthesized TAGs were
determined by Ag-HPLC [10, 11].
(
Cleveland, OH, USA), N,N¢-dicyclohexylcarbodiimide,
p-toluenesulfonic acid and triolein from Sigma-Aldrich (St.
Louis, MO, USA), 4-dimethylaminopyridine from Eastman
Fine Chemicals (Rochester, NY, USA), Silica gel (60/200)
5. Drop Point: Drop points were determined by AOCS
Official Method Cc 18-80. The values listed in Table 1
represent the means of duplicate determinations.
ꢀ
and Florisil (100/200) from J.T. Baker Chemical Co.
1
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J Amer Oil Chem Soc (2007) 84:427–431
429
Fig. 1 Analysis of EOE and
EEO triacylglycerol mixture by
dual-column Ag-HPLC. Sample
size: 10 lg. Flow rate: 1.0 mL/
min 0.5% ACN in hexane. UV
detection at 206 nm. Peaks:
A = EOE, B = EEO
Table 1 Physical and chemical
properties of MAG, DAG and
TAG
Precursor/product Chem. (%) Struct. (%) Onset of Melting points (ꢁC)
Heats of fusion (DH)
melt (ꢁC)
Mettler
DSC
cal/g
J/g
EEE
99.9
99.9
99.9
99.7
99.9
99.9
99.9
>99
39.4
–
43.5
–
43.4
–
36.1
–
151.1
1
1
,3EE-DAG
(3)E-MAG
98
–
12.5
15.9
14.4
6.1
–
–
EOE
EEO
OOO
>98
11.0
4.7
2.4
14.9
12.2
6.1
22.5
26.8
25.6
94.2
112.2
107.2
99.6
99.9
6
.
Differential Scanning Calorimetry (DSC): Heats of
fusion (DH) and melting points were determined
according to AOCS Official Method Cj 1-94.
25 mL r.b. flask equipped with reflux condenser and
argon inlet. Sodium methoxide (0.1 g) was added and
the mixture was heated to 115 ꢁC with stirred magnet-
ically. DAG formation was followed by TLC, and 0.1 g
portions of glycerol and sodium methoxide added after
7
h. After 24 h, the mixture was cooled to room
Syntheses
temperature, dissolved in 20 mL benzene and eluted
through a 2.54-cm glass column packed with 50 g silica
gel. The TAGs (4.8 g) were eluted with 400 mL
benzene, the DAGs (5.4 g; ca. 50% yield) with 90:10
benzene: EE, and the MAGs and FAs (1.0 g) with 100%
EE. (Toluene may be substituted for benzene.) After
solvent removal by rotary evaporation, the DAG fraction
was dissolved in 500 mL acetone, and the solution was
slowly cooled (crystal formation noted) to/stored over-
night at 5 ꢁC. The crystals (4.8 g) were removed by
filtration through a cold Buchner funnel, and dried in a
vacuum oven at 40 ꢁC. The DAG structural purity,
determined by conversion to the mono-acetate/Ag-
HPLC [Method (4), above], was found to be >99%
1,3-dielaidylglycerol.
See Table 1 for drop point and DSC values, as well as
chemical and geometrical purities of the MAG and DAG
precursors and of the TAG isomers.
1. Preparation of EEE: Elaidic acid (66.8 g, 0.25 mol)
was combined with glycerol (7.3 g, 0.08 mol) and 0.7 g
p-toluenesulfonic acid in a 250 mL, 3-necked round
bottom (r.b.) flask fitted with an argon inlet and
thermometer. The contents were stirred magnetically
and heated by oil bath to 115 ꢁC. Water which
condensed on the flask walls during the reaction was
removed by use of a heat gun. Reaction progress
was measured by TLC. After 5 h the reaction mixture
was cooled, the solid residue dissolved in 150 mL
petroleum ether and eluted through a glass column
packed with 500 g Florisil. The EEE was isolated by
elution with 1,500 mL 90/10 PE/EE, and the solvents
removed to yield 59.2 g (85% yield) of product.
3. Preparation of EOE: EOE was prepared according to
the method of Kodali et al. [12]. 1,3-dielaidylglycerol
–
3
(2.2 g, 3.5 · 10 mol) in 80 mL carbon tetrachloride
was transferred to a heat-dried, 3-necked, 500 mL r.b.
flask equipped with a mechanical stirrer, thermometer
2. Preparation of 1,3-Dielaidin: EEE (10.0 g, 0.01 mol)
and glycerol (1.0 g, 0.01 mol) were combined in a
–
3
and argon inlet. Oleic acid (1.1 g, 3.5 · 10 mol, 10%
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ꢀ
excess) in 10 mL of carbon tetrachloride was added via
ChromSpher Lipids columns connected in series resulted
in improved sample capacity and improved peak-to-peak
resolution. Ag-HPLC is easy to use and yields reproducible
results within 25–50 min. UV detection at 206 nm was
used to determine structural purities of the EEO and EOE
isomers and, as noted previously [9], the results were
comparable to those achieved by lipolysis/gas chromatog-
raphy.
syringe. 4-Dimethylaminopyridine (0.5 g in 5 mL CCl4)
was added in one batch and 1.34 g N,N¢-dicyclohexyl-
carbodiimide dropwise over a 30-min period. The
reaction was stirred at 28 ꢁC for 2 h (precipitate noted).
The precipitate was removed by vacuum filtration and
the solvents by rotary evaporator. The EOE residue
(4.2 g) was purified by elution (5% EE in PE) through a
2
.54 cm diameter glass column packed with 30 g silica
The chemical and physical properties of the structured
TAGs and their precursors are shown in Table 1. Included
are Mettler dropping points (MDP) and melting points/
point ranges obtained by differential scanning calorimetry.
Heats of fusion, also determined by DSC, are also included.
The difference (by DSC) between the onset of melt and the
melting points indicates that EEE, EOE, and EEO TAG are
sharply-melting materials, since their melting points are
about 4–7 ꢁC above the onset of melt. As would be ex-
pected, EEE shows a higher heat of fusion than EOE or
EEO. On the other hand, EEO and EOE have heats of
fusion similar to OOO. By comparison, completely satu-
rated TAGs such as tristearin and tripalmitin show heats of
fusion of 45.0 and 39.6 cal/g, respectively [22]. Thus, EEE
and SSS have similar heats of fusion, and the introduction
of oleic acid into the molecule lowers DH by approximately
a factor of 2 (i.e. from 36.1 cal/g for EEE to 22.5 and
26.8 cal/g for EOE and EEO, respectively).
gel to yield 2.7 g (86% yield) of product. Analysis of the
EOE by Ag-HPLC and, after conversion to FAME, by
GC yielded structural and chemical purities of 98 and
>
98%, respectively. (see ‘‘Materials’’ and ‘‘Methods’’
above and Table 1).
. Preparation of OEE: Commercially-available 1(3)-
4
–
3
monoolein (3.0 g, 8.4 · 10 mol) was esterified with
–
2
elaidic acid (5.4 g, 1.9 · 10 mol) to prepare OEE
6.5 g; 87% yield) as described in ‘‘Preparation of
EOE’’, above.
(
Results and Discussion
Multi-gram (4–6 g) quantities of highly-pure symmetrical
or non-symmetrical TAG could be prepared by esterifica-
tion of the appropriate MAG or DAG precursors (also
commercially available), as described above. Silica gel
chromatography was utilized to isolate the DAG and MAG
fractions, with the thermodynamically more stable 1-
MAGs and 1,3-DAGs predominating (thermodynamic ra-
tios of 80/20 1(3)- vs. 2-MAGs and 1,3- vs. 1,2-DAGs),
and could be readily isolated by low-temperature crystal-
lization/filtration from acetone with structural purities
exceeding 98%. (The percentage of MAG or DAG present
in the final mixture can be adjusted by the amount of
ethylene glycol added, and the thermodynamically more-
stable MAG or DAG isomers increased by addition of 1%
of an acid such as HCl [8].) The thermodynamically less-
stable 1,2-DAGs, present at only 20–25% in the original
DAG fraction, could also be utilized for preparation of non-
symmetrical TAG isomers, but only mg quantities of
highly-pure (>99%) samples could be isolated by Ag-
HPLC [13]. The commercially-available (and less-expen-
sive) 1O-MAG was therefore esterified with elaidic acid to
yield the non-symmetrical OEE isomer. This flexible syn-
thetic approach [5–7, 10, 12] offers a viable alternative to
enzymatic synthesis [14–20] for preparation of TAGs
present in chemically-interesterified commercial spreads
and formulations.
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