An Enzymatic Assay of Chloramphenicol Coupled with Fluorescence Reaction

Abstract

An alternative method for the assay of chloramphenicol using an enzymatic reaction coupled with a fluorescence detection system has been developed. Chloramphenicol was enzymatically acetylated by chloramphenicol acetyl-transferase in the presence of acetyl-coenzyme A (acetyl-CoA) as the acetyl-donor, after which the liberated CoA-SH was derivatized with a fluorogenic reagent, 4-(aminosulfonyl)7-fluoro-2, 1, 3-benzoxadiazole. The assay was linear over the range of 2.5-40 μg/ml. Analytical recoveries of chloramphenicol at concentrations of 7.5 and 22.5 μg/ml, added to human serum, plasma, or a slightly hemolyzed serum, were in the range of 93.3 to 106.2%. The enzymatic assay was not affected by the presence of ten other antibiotics tested.