Degree Type


Date of Award


Degree Name

Doctor of Philosophy


Biochemistry, Biophysics and Molecular Biology


Hydrolysis of vitamin A esters by liver is important during their uptake as part of chylomicron remnants and for their mobilization from storage in vitamin A-containing globules. The regulation and properties of the enzyme thought to catalyze these reactions, retinyl ester hydrolase (REH), were studied by the use of a sensitive in vitro assay developed for this enzyme in pig and rat liver. Hydrolysis of retinyl palmitate was maximal in 3- (3-cholamidopropyl) dimethylammonio -1-propanesulfonate (CHAPS) and Triton X-100, and the production of retinol was quantified by HPLC;Regulation of hepatic REH was studied by determining the relationship between its activity and physiological indicators of vitamin A nutritional status. Maximally stimulated hydrolase activity was not strongly influenced by serum or liver vitamin A concentrations in rats or pigs. However, in reduced detergent and substrate concentrations, REH activity was 3-fold higher (p < 0.01) in rats with depleted liver reserves (<10 (mu)g/g). Thus, REH may play a role in the homeostatically controlled release of vitamin A from liver, by mechanisms that are masked in vitro by concentrated detergent;Bile salts also strongly affected the level and interanimal variability of REH activity. Notably, rat REH activity in individual livers was 10-fold higher in CHAPS compared to sodium cholate, and less variable (5- compared to 37-fold). Therefore, the variability others have reported may be due to the use of sodium cholate to stimulate REH;13-cis, 9-cis and 9,13-cis Retinyl palmitate were excellent substrates for pig liver REH, being hydrolyzed at rates 2- to 7-times greater than the all-trans isomer with similar apparent K(,m) values and pH dependencies. However, by salt fractionation and column chromatography, three enzyme preparations were separated, which differed in their sensitivities to inhibitors and in their detergent-dependent substrate specificities. Thus, retinyl ester hydrolysis in the liver may well involve more than one enzyme.



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Dale Alan Cooper



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134 pages

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Biochemistry Commons