Enzymatic Cycling Assay for d-Carnitine Determination

✍ Scribed by J.M. Obón; B. Buendı́a; M. Cánovas; J.L. Iborra

• Publisher: Elsevier Science
• Year: 1999
• Tongue: English
• Weight: 89 KB
• Volume: 274
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.1999.4233

✦ Synopsis

An enzymatic method for D-carnitine determination using the enzyme D-carnitine dehydrogenase is described. The assay is based on the amplified signal produced during NAD ؉ cycling in the presence of a tetrazolium salt and using phenazine methosulfate as electron carrier. Optimum assay conditions were studied with two tetrazolium salt pairs: 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT)/ MTT-formazan and 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyl tetrazolium chloride (INT)/INT-formazan. The first pair (MTT) showed higher sensitivity. The calibration curve was linear from 0.1 to 5 mM D-carni- tine, with a quantification limit of 0.1 mM and a relative standard deviation of 1.51%. The procedure is simple, rapid, accurate, and easily automated. It was satisfactorily applied to following D-carnitine levels during the microbial transformation of D-carnitine into L-carnitine and to determining the D-carnitine content of pharmaceutical preparations.