A D-carnitine dehydrogenase electrode for the assessment of enantiomeric purity of L-carnitine preparations

The enantiomeric purity of pharmaceutical L-carnitine preparations can be a ssessed within 60 seconds using a highly selective bienzyme electrode. D-ca rnitine dehydrogenase from Agrobaterium is highly specific for the nonphysi ological D-enantiomer and was therefore used as the recognition element. NA DH produced in the primary reaction was oxidized by salicylate hydroxylase (EC 1.14. 13.1) in an oxygen and salicylate dependent reaction. The consump tion of oxygen was monitored with a miniature Clark-electrode. A linear cal ibration graph from 0.01 mM through 0.6 mM D-carnitine was obtained in phos phate buffer pH 8 comprising 0.5 mM concentrations of the cosubstrates NAD and salicylate. The sensitivity for DL-carnitine was exactly 50% of the res pective value for pure D-carnitine, while L-carnitine and ascorbic acid, a common interferent, gave no response at all. Mixtures of both enantiomers c ontaining 1% and 3% D-carnitine, respectively, could be distinguished from each other and from pure (i.e. >98 %) L-carnitine preparations with the new sensor. The biosensor method is faster and less laborious than established HPLC and H-1-NMR methods since it requires no chemical derivatization. The lower detection limit was 10-fold reduced as compared with a recently publ ished enzymatic assay.