BIOTIN DETERMINATION BY 3 DIFFERENT METHODS - SPECIFICITY AND APPLICATION TO URINE AND PLASMA ULTRAFILTRATES OF PATIENTS WITH AND WITHOUT DISORDERS IN BIOTIN METABOLISM

A microbiological, an avidin-binding and a streptavidin-binding method for biotin determination were compared. all three methods detected bi otin equally well but they exhibit different specificities for derivat ives of biotin. The microbiological assay has the highest specificity and is the method of choice for biotin determination in biotinidase-de ficient patients. The specificity of streptavidin-binding has not been investigated so far. Application of the three methods to urine sample s of patients with and without biotin therapy indicated that only 50% of biotin equivalents measured with the avidin method correspond to au thentic biotin as previously shown. The other 50% comprise mainly bisn orbiotin and biotin-d-sulfoxide. HPLC-separation of urine samples prio r to assay confirmed this finding and revealed a bisnorbiotin oxidatio n product and an unknown compound as further biotin metabolites. The l atter was measurable by all three methods and not detectable in plasma ultrafiltrate. this was th only metabolite which was able to restor d eficient 3-methylcrotonyl-CoA carboxylase activity in biotin-deficient fibroblasts. The combination of the three methods together with HPLC- separation proved to be a valuable analytical tool for the identificat ion of the main biotin metabolites in biological fluids.