Improved enzymatic assay of chloramphenicol.
We partly purified R-factor-encoded chloramphenicol acetyltransferase (EC 2.3.1.28) from a highly choloramphenicol-resistant mutant derived from Escherichia coli W677/R5. The preparation permitted rapid quanitation of chloramphenicol by use of [14C]acetylcoenzyme A, removing the diacetylated product by selective adsorption onto micropore filters. Succinyl and glucuronyl 3-hydroxyl esters of chloramphenicol were not active as substrates for the preparation, nor were they active as inhibitors. The enzyme was free of chloramphenicol reductase activity, and utilizes other biologically active chloramphenicol analogs. Other antibiotics, at concentrations commonly found in human sera, and blood preservatives, at concentrations 10-fold that found in blood-collection tubes, did not interfere with the enzymic quantitation of chloramphenicol. We conclude that this enzyme preparation permits rapid clinical quantitation of chloramphenicol.
D000123:Acetyltransferases / Q000378:metabolism; D002250:Carbon Radioisotopes; D002701:Chloramphenicol / Q000031:analogs & derivatives / Q000032:analysis*; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000201:enzymology; D005966:Glucuronidase; D006801:Humans; D007700:Kinetics; D008722:Methods; D013379:Substrate Specificity; D001616:beta-Galactosidase
D002250:Carbon Radioisotopes; D002701:Chloramphenicol; D000123:Acetyltransferases; D001616:beta-Galactosidase; D005966:Glucuronidase