Substrates have often been observed to protect enzymes against denaturation by heat and other agents (Joly, 1965; Bernhard & Rossi, 1968). On the other hand, yeast alcohol dehydrogenase activity can be decreased by preincubation at 43°C with NAD+ (Grisolia & Joyce, 1959). Liver alcohol dehydrogenase, however, when heated at 750C (at pH 7.0) with NAD+ at low concentration (0.04mM) shows a small stabilization effect compared with no addition of NAD+ (Yonetani & Theorell, 1962).We find 35-30% recovery of yeast alcohol dehydrogenase activity after preincubation in 10nimsodium pyrophosphate buffer, pH8.8, at 430C for 20min, within the range 0-1.5mM-NAD+ or -NADP+. This recovery decreased rapidly with increasing concentration of NAD+ or NADP+ above 1.5mM, reaching zero at 3.5mm-NAD+ or -NADP+. With liver alcohol dehydrogenase 35-30% recovery was obtained after preincubation at 430C for 50min within the range 0-2.0mM-NADW. The recovery reached zero at 4.3mM-NAD+ or 3.3mm-NADP+. No stabilization was observed at 0.04mM-NAD+ under our conditions of preincubation.Cysteine or bovine serum albumin (most effective at 0.8 and 0.03mg/ml respectively) included in the preincubation medium had a fairly constant protective action throughout the concentration range of NAD+ used, in addition to a small activator effect in the enzyme assay itself. By using 3.5mM-NAD+ with yeast enzyme (normally zero recovery here), 13% recovery was obtained with cysteine and 6% recovery with bovine serum albumin. For liver alcohol dehydrogenase (with 4.3mM-NAD+) recoveries were 7 % with cysteine and 14% with bovine serum albumin.Only the liver enzyme can use NADP+ as cofactor, yet both of these enzyme preparations are inactivated at high concentrations of NADP+. Accelerated thermal denaturation of these enzymes by NAD+ or NADP+ could be due to promotion of oxidation ofessential thiol groups or conformational change, including dissociation into subunits. Our findings with stabilizers suggest that both of these changes may occur. NIEVEL. (BritishIndu8trial Biological Re8earch A88ociation, Woodmansterne Road, Car8halton, Surrey, U.K.) Nitrosamines are strongly hepatotoxic compounds and after chronic administration produce tumours in liver and other organs (Magee & Bames, 1956; Druckrey, Preussmann, Ivankovic & Schmiihl, 1967). The formation of toxic metabolites in vivo was assumed to be responsible for the toxic effect of several dialkyl derivatives. However, it has been suggested by Mirvish (1969) that the metabolic activation of diethylnitrosamine as directed by alkylation of macromolecules is not carried out by an inducible drug-metabolizing enzyme system. We investigated the induction of diethylnitrosamine metabolism by studying de-ethylation in vitro by liver microsomal fraction and its relation to the overall toxicity.Female Wistar rats were injected intraperitoneally with 80mg of phenobarbitone/kg and 20mg of 3-methylcholanthrene/kg and orally with 200mg of butylated hydroxytoluene/kg and 160mg of DDT [1,1,1-trichloro-2,2-bis-(p-chlorophenyl)et...
