LIPOIC ACID (THIOCTIC ACID) ANALOGS, TRYPTOPHAN ANALOGS, AND UREA DO NOT INTERFERE WITH THE ASSAY OF BIOTIN AND BIOTIN METABOLITES BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY AVIDIN-BINDING ASSAY

Lipoic acid, urea, and tryptophan show structural similarities to the vitamin biotin. If these compounds can successfully compete with bioti n or a biotinylated protein for binding to avidin, their presence in s erum and urine would cause artifacts in avidin-binding assays for biot in. We assessed the ability of lipoic acid and certain analogs, urea, and L-tryptophan and tryptophan derivatives to interfere with the meas urement of 16 biotin analogs by a high-performance liquid chromatograp hy (HPLC)/avidin-binding assay. In this assay, compounds are separated by reversed-phase HPLC, followed by assay of each fraction based on b inding to avidin-horseradish peroxidase. At physiologic concentrations , neither lipoic acid analogs (d-lipoate, 1-lipoate, d,1-lipoamide, bi snorlipoate, beta-hydroxybisnorlipoate, and tetranorlipoate) nor trypt ophan derivatives exhibited detectable avidin-binding. Minor avidin-bi nding was seen for urea and L-tryptophan; binding ratios relative to b iotin were 1 x 10(-9) and 3 x 10(-6) respectively. Urea did not co-elu te on HPLC with any of the 16 biotin analogs bus L-tryptophan did co-e lute with bisnorbiotin methyl ketone. However, because the relative av idin affinity of L-tryptophan is five orders of magnitude smaller than that of bisnorbiotin methyl ketone, we conclude that none of the test ed compounds are likely to interfere with the measurement of biotin or its metabolites by an HPLC/avidin-binding assay.