ABSTRACT
Autoxidation of para-aminophenol (PAP) has been proposed to account for the selective nephrotoxicity of this compound. However, other studies suggest that hepatic metabolites of PAP rather than the parent compound may be responsible for renal damage. These studies were designed to investigate PAP metabolism in isolated hepatocytes. We synthesized several proposed metabolites for analysis by HPLC/mass spectrometry and compared those results with HPLC/mass spectrometric analyses of metabolites found after incubating hepatocytes with PAP. Hepatocytes prepared from male Sprague-Dawley rats were incubated in Krebs-Henseleit buffer at 37 degrees C for 5 h with 2.3 mM PAP under an atmosphere of 5% CO2/95% O2. Aliquots were withdrawn at 0.1 h of incubation and then hourly through 5 h of incubation. Reactions were terminated by the addition of acetonitrile. Hepatocyte viability was unaltered with PAP present in the incubation medium. We found that hepatocytes converted PAP to two major metabolites (PAP-GSH conjugates and PAP-N-acetylcysteine conjugates) and several minor metabolites [PAP-O-glucuronide, acetaminophen (APAP), APAP-O-glucuronide, APAP-GSH conjugates, and 4-hydroxyformanilide]. Preincubating hepatoyctes with 1-aminobenzotriazole, an inhibitor of cytochromes P450, did not alter the pattern of PAP metabolism. In conclusion, we found that PAP was metabolized in hepatocytes predominantly to PAP-GSH conjugates and PAP-N-acetylcysteine conjugates in sufficient quantities to account for the nephrotoxicity of PAP.
Subject(s)
Aminophenols/metabolism , Liver/metabolism , Acetylcysteine/metabolism , Aminophenols/pharmacology , Animals , Cell Survival/drug effects , Chromatography, High Pressure Liquid , Gas Chromatography-Mass Spectrometry , Glutathione/metabolism , In Vitro Techniques , Kidney/drug effects , Liver/cytology , Liver/drug effects , Male , Rats , Rats, Sprague-DawleyABSTRACT
Ethanol in certain beverages and in similar solutions may be determined by reversed-phase liquid chromatography (LC) using the UV detector. The mobile phase in this indirect photometric detection technique contains a low concentration of a UV-absorbing compound, such as acetone, that coelutes with the ethanol peak. Several variables such as the choice and concentration of the UV-detection agent are examined regarding their effects on the retention time, magnitude and linearity of peak area, and other aspects of quantitation. Except for filtering to remove particulate matter, samples can be injected without pretreatment. The concentration of ethanol in several types of beverages can be determined with 2% relative standard deviation, calibration is linear to 40% ethanol, and the minimum detectable concentration is 0.1%.
