H. H. Draper

Method for the assay of free and esterified tocopherols

Abstract A method for estimating the eight naturally occurring isomers of vitamin E and their esters is described. The prime feature of this method is the removal of the bulk of the lipids by low-temperature crystallization instead of saponification. The technique concomitantly affords a separation of tocopherol esters from free tocopherols. The recovery of tocopherols was superior to that obtained by the conventional saponification method, which was shown to cause extensive destruction of tocotrienols. Reduction with LiAlH 4 provided the highest yield of free tocopherols from tocopherol esters. The conjugated forms were found only in latex lipids. Analytical values are given for the tocopherols in several plant oils.

Metabolism of 1-14C-methyl linoleate hydroperoxide in the rabbit

The metabolism of 1-14C-methyl linoleate hydroperoxide (1-14C-MLHP) by the rabbit was investigated. Administration of 1.1–1.9 mg of 1-14C-MLHP by ear vein injection proved lethal to four of the nine experimental animals. After 2 hr the lungs and liver contained 3.3% and 7.2%, respectively, of the dose. This radioactivity was found to be associated primarily with intact 1-14C-MLHP. The triglycerides from these tissues also contained14C-trienoic and14C-dienoic fatty acids. Of the dose, 68% was recovered as14CO2 in 2 hr compared to 39% after 1-14C-methyl linoleate injection. The triglycerides from kidney adipose tissue contained a small amount of14C-hydroxy fatty acid, providing confirmation of previous evidence for the presence of a fatty acid hydroperoxide reductase in animal tissues.

Oxidation products of α-tocopherol formed in autoxidizing methyl linoleate

The oxidation products of14C-α-tocopherol formed by heating with methyl linoleate in an air atmosphere at 60 C or 100 C were investigated. The products included a dimer, trimer and dihydroxy dimer, α-tocopherol quinone and unidentified degradation compounds. The dimer and trimer constituted the major products present after heating for 70 hr at 60 C. After 70 hr at 100 C most of the14C-α-tocopherol had been converted to degradation products; part of the14C originallt present in the 5-methyl group was recovered as14CO2 and14CH3OH.

Absorption and metabolism of 1-14C-hydroxy octadecadienoate in the rat

The metabolism of 1-14C-9(13)-hydroxy octadecadienoic acid methyl ester (1-14C-HAME) by the rat was investigated in vivo and in liver slices. A 1.5 mg dose of 1-14C-HAME administered by stomach tube was efficiently hydrolyzed and absorbed from the intestinal tract. In comparison with 1-14C-methyl linoleate (1-14C-ML), 1-14C-HMAE was more extensively oxidized to14CO2 in vivo and in vitro. After 1-14C-HAME administration as much as 50% of the radioactivity in the adipose tissue triglycerides was associated with14C-hydroxy fatty acids. The remaining activity was present in randomly labeled normal fatty acids. No evidence was obtained for the incorporation of14C-hydroxy acids into liver lipids; most of the radioactivity from 1-14C-HAME in this organ was recovered in saturated and monoenoic fatty acids. About 10% of the radioactivity 24 hr after 1-14C-HAME administration was associated with triglyceride trienoic acids, indicating that at least a portion of this acid was dehydrated in the liver. An unidentified polar acid was detected in the urine of the 1-14C-HAME-treated animals.

Glutathione peroxidase activity and glutathione concentration in genetically dystrophic mice.

Summary The present studies were conducted to determine whether inherited muscular dystrophy in the 129/ReJ-dy mouse was associated with differences in specific activity, substrate availability, or apparent K m of glutathione peroxidase. The results indicate that glutathione peroxidase is elevated in skeletal muscle of mice with genetic muscular dystrophy when the activity is expressed on a protein basis. This elevation precedes the development of severe paralysis since muscles from the fore legs showed increased enzyme activity as early as the more severely affected hind legs. There was no difference in glutathione peroxidase activity in tissues other than skeletal muscle. GSH concentration was elevated in muscle and normal in other tissues of dystrophic mice, showing that adequate substrate was available to the enzyme. The apparent K m for cumene hydroperoxide was also similar for muscle of normal and dystrophic mice. This report provides further evidence that mice with dystrophia muscularis have a functional glutathione peroxidase system in all tissues including skeletal muscle, and that a defect in this in vivo protective system is apparently not a contributing factor in the pathology of the disease.