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eight. Consequently, it would appear that the limiting sensitivity of the technique for measuring
anabolic materials is likely to be well below 1 ng per ml.

Analysis of Waxes and Lipid Type Materials

The metabolism of fatty acids in cell cultures is important, particularly with regard to cancer cells. The
possible conversion of the fatty acids to peroxides and the role of these compounds in carcinogenesis is
still not completely understood. As a tool for this type of research Wallace and Coleman [10] developed
a procedure employing a GC/MS tandem instrument for the fatty acid assay of cell tissue. Two human
colon carcinoma lines, HT29/219 and HY115 together with a human breast cancer cell line ZR-75-1
were studied. The cells were grown in Dulbeccos's modified Eagles medium, augmented with 10% v/v
foetal calf serum (DFC )or horse serum (DH ). They were seeded at a density of 1.9 x 104 cells per
10
10
cm and maintained at 37°C in a humidified atmosphere of 5% carbon dioxide and 95% air. The cells
2
were harvested into a phosphate buffered saline at specific intervals up to 120 hours. Lipids were
extracted with chloroform/methanol (2+1) containing 2, 6-di-tert-butyl-4-methylphenol as an
antioxidant. The extract was separated on a thin layer plate using n-hexane-diethyl ether-acetic acid
(70+30+1) as a solvent. The phospholipid, triglyceride and free fatty acid spots were scraped off the
plates and the lipids removed by eluting with chloroform methanol (2+1). The phospholipids and
triglycerides were trans-esterified [11] using sodium methoxide in methanol and the free fatty acids
were methylated with diazomethane.

The authors found that a column that had a very low level of stationary phase bleed was essential for the
GC/MS system to operate. Although the use of polymer coated capillary columns appear to be ideal,
they were found to give inadequate separation and the optimum stationary phase was found to be a
Carbowax polymer column (polyethylene glycol). It was found that the combination of the retention
data with the mass spectrum gave virtually certain fatty acid identification. The authors used the
technique to determine the fatty acid profiles of different cancer cell