Analytical controls in drug metabolic studies. II. Artifact formation during chloroform extraction of drugs and metabolites with amine substituents.

Chemical artifacts are readily produced during chloroform extractions of drugs and metabolites with amine substituents by the reaction of chloroform contaminants, phosgene and ethyl chloroformate, to form carbamoyl chlorides and carbamates. Chloroform obtained from three different suppliers was examined with regard to the effects of preservatives and exposure to air on the quantity and nature of these contaminants. Extractions with chloroform obtained from a supplier's sealed bottle and containing ethanol preservative produced the least amount of artifact. With previously opened bottles (prior exposure to air, open), the amount of artifact formation rose substantially. Chloroform without preservative or with a nonpolar hydrocarbon preservative likewise produced large amounts of artifacts during extraction. Attempts to purify chloroform showed that distillation was ineffective for the removal of phosgene or ethyl chloroformate, whereas their effective removal was accomplished by solvent elution through a column of activated alumina or upon standing over calcium hydroxide powder. Possible artifact formation must be considered in drug-metabolism studies and appropriate controls included for the detection and elimination of chemical artifacts.