A method for measuring the lifetime of chemically and metabolically generated electrophilic species.
A method is presented for the characterization of the reaction order and rate constant for chemically and metabolically generated reactive electrophilic intermediates. The procedure employs flow kinetics and trapping of the electrophilic species. Reactive agents or metabolic intermediates are passed through a column containing a bound nucleophilic reagent. The electrophilic species reacts at a characteristic rate while moving through the column at a fixed pH, temperature and rate of flow. The alkylation products formed during this reaction are quantified in individual column segments which correspond to time intervals. This provides data for the calculation of the lifetime of the short-lived species. The method may be used for agents having half-lives of 1 s to 30 min. Metabolic intermediates need not be isolated. Data is presented for the reaction rates of 1-methyl-1-nitrosourea (MNU), 1-methyl-3-nitro-1-nitrosoguanidine (MNNG) and iodomethane. A metabolic activation system is described for the conversion of acetoxy-methylmethylnitrosamine (DMN-OAc) to hydroxymethylmethylnitrosamine (DMN-OH) and measurement of the stability of that intermediate. Hydroxymethylmethylnitrosamine was found to have a half-life of 28s at pH 7.4, 37 degrees C.
D000477:Alkylating Agents / Q000378:metabolism / Q000633:toxicity*; D000478:Alkylation; D000818:Animals; D001711:Biotransformation; D002273:Carcinogens / Q000378:metabolism / Q000633:toxicity*; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods; D008401:Gas Chromatography-Mass Spectrometry; D007700:Kinetics; D008745:Methylation; D008862:Microsomes, Liver / Q000378:metabolism; D013997:Time Factors
D000477:Alkylating Agents; D002273:Carcinogens