The Identification of Vegetable Oils
Richard E. Cover St. John's University Jamaica, N e w York 11432
A gas chromatographic experiment
M a n y vegetable and animal oils can be identified if tlrcir fatty acid compositions are known. This fact provides the basis for a challenging gas chromatographic experiment which has been used by the author in a graduate instrumental analysis course over a period of three years. The student is given a sample of oil which he saponifies and then esterifies to form the methyl esters of the fatty acids. The esters are then extracted and concentrated by evaporation of solvent. The preparation procedure is simple and rapid. The esters are injected into a gas chromatograph, the various ester peaks are identified, and the fatty acid composition calculated. The oil is then identified by comparison with known fatty acid compositions of various oils. Table 1 contains some of these known data. The Experiment
Two reagents are required: a 3% solution of sodium methoxide in absolute methanol and a 10yo solution of boron trifluoride in absolute methanol (Ewtman 3706). A gas chromatograph equipped Table 1.
Fotty Acid Compositions of Vegetable Oils (Weight Percent)
Acid Myristic Palmitic Palmitaleic Stearic Oleic Linoleic Linolenic Araohidic
120
/
Corn
... ...
12.5 2.5 29.0 55.0 0.5 0.5
Cottonseed 1.0 26.0 1.0 3.0 17.5 51.5
...
...
-
OilLinseed
...
...
6.0
13.0 1.0 2.5 74.0 9.0 0.5
...
3.5 20.0 14.5 56.0
...
Journol of Chemical Education
...
Palm 1.5 42.0
...
4.0 43.0 9.5
...
...
~ d both h a thermal conductivity detector and a flameionization detector should be used; a Perkin-Elmer 154-B chromatograph is satisfactory. A 2.0-m diethylene glycol succinate column run a t 190°C provides adequate resolution of all components. Procedure
A sample of 0 . 1 0 . 2 0 g of oil is weighed into a 2-02 screv-cap vial. Three milliliters of sodium methoxide reagent are added; the vial is capped tightly and then suspended in boiling water for 3 min. The vial is cooled and 3 ml of the boron trifluoride reagent are added. The vial is capped tightly and again suspended in boiling water for 3 rnin. The vial is then cooled and its contents are poured into a 50-ml volumetric flask. One milliliter of pentane and 2 ml of ethyl ether are added, and the flask is shaken while stoppered. Tu-enty-five milliliters of water is then added to the flask which is again shaken thoroughly. Water is then added until the solvent layer is in the neck of the flask. The solvent layer is transferred to a small, dry, scre~v-capvial, and the esters are concentrated by warming and passing a gentle air stream into thevial. Each ester sample is run twice. Using the concentrated esters, a chromatogram is run using the thermal conductivity detector. Two to three milliliters of pentane are then added to the esters and they are thoroughly mixed. A second chromatogram is then run with these diluted esters using the flame-ionization detector. A chromatogram of a known mixture of fatty acid esters, run with the flame-ionization detector under identical experimental conditions, is supplied to each student. Such standard mixtures are available from Applied Science Laboratories, Inc., State College, Pa.
Discussion
The reliability of this identification procedure is attested to by years of successful use by chemists in industry as well as by our experience with the 26 students who have done the experiment. The students have successfully identified these oils: corn, cottonseed, olive, and palm. The few errors in identification were traceable to poor data analysis. Typical student analyses are given in Table 2. It Table 2.
Acid Palmitio Palmitaleic Steario Oleic Linoleic Linolenic
Typical Student Analysis (Weight Percent)
-
~
Corns
Olive
58.8
These data were obtained by different students on the same oil samples.
should be pointed out that the reproducibility of the results listed for corn oil is considerably better than that found in the interlaboratory study' of A.S.T.M. Committee E-19. The experimental data can well be used to sharpen the students' understanding of other areas of chromatographic theory and practice. If the standard ester mixture is properly selected, the standard data can be used to calibrate the detector to increase analytical accuracy. Similarly, the student can evaluate the nature of detector response, i.e., its relationship to weight percent, mole percent, or carbon content of the various compounds2. Finally, from the standard data, the student can demonstrate the validity of the well-known linear relationship between the logarithm of the retention time and carbon number for an homologous series. This is possible for both the saturated ester and the monoeoate series. 1 Annual Meeting, Committee E19, American Society for Testing Materiels, October, 1964, Houston, Texas. a DM, NOGARB, S., AND JUVET, R. S. JR., "Ga~LiquidChromatography," John Wiley & Sons, Inc., New York, 1962, pp. 197,
220.
The unknowr~sample is run twice with two different detectors and, actually, with two different sample sizes. The student can calculate the number of theoretical plates and the retention time for each component on each chromatogram. When the data obtained for each component with the two detectors are compared, the number of theoretical plates calculated for a given component are significantly higher (by a factor of two or three) with the flame-ionizationdetector as compared with the thermal conductivity detector. The retention time for a given component is significantly l o ~ ~ ewith r the flame-ionization detector. Decreases of the order of 20% are not uncommon. These differences are, of course, due to the difference in sample sizes and reflect the limitations of our present ability to deal theoretically with finite samples. Current chromatographic theory is based on the assumption that the sample is completely and instantaneously injected onto the first theoretical plate. This assumpth techtion cannot be satisfied in practice ~ ~ i existing niques. Experimental chromatographic peaks therefore do not obey the theoretical predictions. Real peak geometry and behavior are not solely functions of column parameters and the chemical properties of the solute; they are related to sample size as ~vell. Finally, intercomparison of theoretical plate estimates from a single chromatogram d l reveal significant differences in this parameter as calculated for the various compounds. The number of theoretical plates of a column, as evaluated experimentally from the peak behavior of a given con~ponent,is very much affected by the chemical nature of that component. Evaluation of these data, therefore, prevents a common misconception from arising, namely, that the theoretical plate parameter is a property of the column alone and can be used indiscriminately as an absolute and precise index of column efficiency. Acknowledgment
The author wishes to thank Humko Products, Sterick Building, Memphis, Tenn. 31801, for permission to publish the data in Table 1. Similar information on other oils can be obtained from the same source.
Volume 45, Number 2, February 1968
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