THE APPLICATION NOTEBOOK — August 2002
Food and Beverage
ADVERTISING SUPPLEMENT
21
Determination of Triglyceride
Composition of Vegetable Oils
Using High Performance Liquid
Chromatography and Evaporative
Light Scattering Detection
Romulus Gaita, Alltech Associates, Inc., Deerfield, Illinois
This application demonstrates a fast HPLC method for
the separation of several triglycerides in various edible
oils with evaporative light scattering detection.
F
ats and oils are extremely important food products and
accordingly, much attention has been given to their analysis.
There are a number of detection methods for analyzing fats and oils
that use HPLC. The poor solubility and long retention times of the
higher saturated triglycerides make gradient elution desirable, but
not possible with RI detection.
The direct gas chromatographic analysis of triglycerides is exceedingly difficult because of the low volatility of these compounds.
Most often this requires derivatization such as transesterification of
the triglycerides, followed by gas chromatographic analysis of the
resulting methyl esters. The transesterification of triglycerides is not
always quantitative and, more importantly, the method does not
provide any information about the actual composition of the
triglycerides.
UV detection is acceptable, but limited to a very small selection
of non–UV absorbing solvents. The high molecular weight triglycerides have limited solubility in non–UV absorbing solvents, which
make the use of these solvents undesirable for triglyceride separation.
The ELSD provides essentially flat baselines with gradient elution, operates with many UV absorbing solvents, and successfully
detects triglycerides in oils without derivatization.
This application outlines a single, fast HPLC method for the separation and identification of several triglycerides in various edible
oils using an ELSD and a nonaqueous, reversed-phase gradient
method. Examples of triglyceride composition in extra virgin olive
oil, vegetable oil, and coconut oil are shown in Figures 1–3.
Conclusion
A fast (less than 15-min) reversed-phase HPLC method for the
analysis of triglycerides in edible oils has been developed using the
ELSD 2000. An Alltima™ C18, 150 4.6 mm, 3-m column
offered the best separation of triglycerides in edible oils.
Table I: Symbols used for the
various fatty acids
Number ˜
of
Carbons Compound
10
C Capric acid
12
La Lauric acid
14
M Myristic acid
16
P Palmitic acid
Po Palmitoleic acid
18
S Stearic acid
O Oleic acid (cis, 9)
L Linoleic acid (9,12)
Ln Linolenic acid
(9,12,15)
1 LLL
2 LLO
3 LLP
4 OOL
5 POL
6 PPO
7 OOO
8 OOP
1 LLO
2 OOL
3 POL
4 OOO
5 OOP
6 PPO
7 OOS
4
5
2
1 3 67
0 2 4 6 8 10 12 14 16
Time (min)
Figure 1: Triglycerides in extra
virgin olive oil.
1
1 CCC
2 LnLnLn
3 LaLaLa
4 LLL
5 LLO
6 LLP
7 OOO
23
2
3
45
7
68
1
6
4 5
7
0 2 4 6 8 10 12 14 16
Apparatus and HPLC Conditions
An LC system equipped with an Alltech Model 2000 ELSD and an
Alltech Alltima™ C18, 3-m, 150 4.6 mm column was used.
Gradient Conditions
A: Acetonitrile
B: Dichloromethane
Time:
0
% B:
30
Flow rate: 1.5 mL/min
10
45
18
30
20
30
Time (min)
0 2 4 6 8 10 12 14 16
Time (min)
Figure 2: Triglycerides in vegetable oil.
Figure 3: Triglycerides in coconut
oil.
Alltech Associates, Inc.
2051 Waukegan Road, Deerfield, IL 60015
tel. (847) 948-8600, fax (847) 948-1078
e-mail [email protected]
http://www.alltechWEB.com
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