Joseph MacGee

Preparation of methyl esters from the saponifiable fatty acids in small biological specimens for gas-liquid chromatographic analysis.

Abstract Tissue (3–5 mg) or serum (5–100 μl) is hydrolyzed with alkali, acidified and extracted with hexane. The fatty acids are extracted from the hexane into a small volume of trimethyl(α,α,α-trifluoro- m -tolyl)ammonium hydroxide. Injection of the resultant quaternary ammonium salts together with methyl propionate into a gas chromatograph yields the fatty acid methyl esters. Preparation of the sample requires less than 45 min and less than 10 min of the analysts time. The results are comparable with those obtained by conventional methods with larger samples.

Identification of 5-meta-Hydroxyphenyl-5-phenylhydantoin as a metabolite of diphenylhydantoin☆

Abstract 5-meta-Hydroxyphenyl-5-phenylhydantoin has been identified in conjugated form in the urine of dogs and patients treated with diphenylhydantoin (DPH). This metabolite appears to be the major product of the hydroxylation of DPH in dogs, but it accounts for relatively little of the DPH hydroxylated by patients.

Prepartion of sphingolipid fatty acid methyl esters for determination by gas—liquid chromatography

Sphingolipid faty acids are first convrted to a mixture of free acids and their n-butyl esters by heating the specimen at 82°C in aqueous butanolic hydrogen chloride; the butyl esters are then saponified with methanolic potassium hydroxide. After acidification and extraction into hexane, the fatty acids are extracted into a very small volume of aqueous trimethyl(m-trifluorotoylyl)ammonium hydroxide (TMTFTH) injection of an aliquot of the TMTFTH extract into the gas chromatograph yields the fatty acid methyl esters by pyrolytic methylation of the quartenary ammonium salts of the fatty acids. The preparation of a specimen ready for gas—liquid chromatographic(GLC) analysis with quantitative recovery of teh sphingolipid fatty acids can be accoomplished in less than 2 h. By comparison, none of a number of well-accepted techniques for the release of sphingomyelin fatty acids by hydrolysis or methanolysis released the fatty acids quantitatively in less than 3 h, and all required additional manipulations before GLC analysis.