A method for assay of chloramphenicol acetyltransferase from crude cell extract

This is a method for detection and assay of CAT from Chloramphenicol resistant bacterial isolates. The procedure involved quantitative separation of both chloramphenicol acetate (CmA) and Chloramphenicol (Cm) from assay mixtures by adsorption chromatography on a neutral alumina column. The solvent system--2.5% methanol in benzene--was used for elution of CmA and methanol for Cm. This was followed by either colorimetric estimation of CmA by coupling with N--(1-Naphthyl)--ethylene diamine HCL after zinc hydrocholoric acid reduction or by microbiological assay. In the latter method CmA was deacetylated by enzyme from chicken liver acetone powder. As low as 1 microgram of CmA could be determined by the described microbiological turbidimetric assay method.