Activity, stereospecificity, and stereoselectivity of microsomal enzymes in dependence on storage and freezing of rat liver samples.
Liver microsomes are now one of the most widely used in vitro test systems for biotransformation studies of drugs, toxins, and other xenobiotics. The standard procedure of preparation of microsomes from fresh liver taken immediately after death of the animal is impossible in experiments with liver samples from human or wild animals and the choice of a relatively optimal way of liver storage is necessary in these cases. We studied the possibility of using the stereoselectivity and stereospecificity of biotransformation enzymes for evaluation of the changes in enzyme function dependent on tissue handling. Activity, stereospecificity, and stereoselectivity of several enzymes in microsomes prepared from fresh liver, frozen liver in liquid nitrogen, or ice-cooled liver were compared. The effect of storage period (2, 3, 5 h) on these parameters were also tested. Both freezing and cooling of liver change the native function of enzyme systems and could result in incorrect stereospecificity data for the microsomal metabolism. All parameters observed also differ in their dependence on period of ice cooled storage. As it is difficult to hold strictly to the same storage period, we recommend freezing liver in liquid nitrogen if the storage of liver is necessary. In projects comparing enzyme activities in human and laboratory animals the same freezing procedure of liver should be maintained before preparation of microsomes from all species.
Skálová
L
L
Department of Biochemical Sciences, Faculty of Pharmacy, Charles University, Hradec Králové, Czech Republic. SKALOVAL@FAF.CUNI.CZ
Szotáková
B
B
Wsól
V
V
Král
R
R
Baliharová
V
V
Lamka
J
J
D000429:Alcohol Oxidoreductases / Q000378:metabolism; D000818:Animals; D001711:Biotransformation; D015925:Cryopreservation; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004983:Ethanolamines / Q000737:chemistry / Q000378:metabolism; D005260:Female; D006801:Humans; D007546:Isoquinolines / Q000737:chemistry / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology*; D010084:Oxidation-Reduction; D051381:Rats; D017208:Rats, Wistar; D013237:Stereoisomerism; D013379:Substrate Specificity; D013997:Time Factors
D004983:Ethanolamines; D007546:Isoquinolines; C103439:oracine; D003577:Cytochrome P-450 Enzyme System; D000429:Alcohol Oxidoreductases; C119935:oracin reductase