Early alterations induced by carbon tetrachloride in the lipids of the membranes of the endoplasmic reticulum of the liver cell I. Separation and partial characterization of altered lipids

Alterations induced by carbon tetrachloride poisoning in fatty acids of liver microsomal lipids were studied. Thin layer chromatography of fatty acid methyl esters prepared from liver microsomal lipids, revealed, in the CCl₄-treated rats, the presence of a component (the “D” spot) with an R_f value lower than that of the methyl esters. The lipids recovered from this component showed a marked diene conjugation absorption when examined spectrophotometrically over the UV range, while the lipids recovered from the spot of the methyl esters showed no absorption of conjugated dienes.Studies carried out with labelled carbon tetrachloride indicated that compounds present in the “D” spot contained 28% of ¹⁴C applied to the chromatoplate. The spot of the methyl esters (the “M” spot) contained 42% of ¹⁴C applied to the chromatoplate. However, specific activity of the “D” spot was about 1000 times greater than specific activity of the “M” spot.The lipids recovered from either the “D” spot or the spot of the methyl esters were analyzed separately by gas-liquid chromatography (GLC) with an electron capture detector (ECD). It was found that the lipids recovered from the “D” spot showed no response, while those recovered from the spot of the methyl esters exhibited the response of the ECD, which was similar to that observed with the unfractionated fatty acid methyl esters. The lack of the response of the ECD for compounds in the “D” spot appears to be due to the fact that they cannot be eluted from the column.On the basis of the analytical results, it can be postulated that the “D” spot contains compounds formed by a chain termination addition reaction of free radicals derived from CCl₄ (probably trichloromethyl free radicals) to fatty acid free radicals containing conjugated dienes. On the other hand, the spot of the methyl esters appears to contain also, together with unmodified fatty acids, the fatty acids in which a simple addition of CCl₄ free radicals to double bonds has occurred.