Lipoic acid analogs and L-tryptophan do not interfere with the assay of biotin by HPLC/avidin-binding assay

J. Zempleni, P. M. Mock, S. L. Stratton, D. B. McCormick

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Abstract

There are structural similarities in the side chain and heterocyclic ring among biotin, lipoic acid analogs (LAA), and L-tryptophan (TRP). The large association constant for the avidin - lipoic acid complex and the cleavage of lipoyllysine by biotinidase further indicate a close relationship of LAA and biotin. We assessed the potential interference of LAA and TRP with the measurement of biotin by the HPLC/avidin-binding assay. In this assay, compounds are separated by HPLC, followed by assay of each HPLC fraction based on binding to avidin-horseradish peroxidase. Compounds that interfere can be separated from biotin and its metabolites by chromatography. Using physiologic concentrations, LAA (d-lipoate, 1-lipoale, d.l-lipoamide, bisnor-, β-hydroxybisnor-, and tetranorlipoates) did not compete for avidin-binding in our assay, whereas TRP showed minor avidin binding (0.0003% compared to biotin, 100%). Spectrophotometric determination of HPLC retention times showed that none of the LAA co-eluted with any of 16 available naturally occurring biotin metabolites, whereas TRP co-eluted (22 min) with bisnorbiotin methyl ketone. Because avidin affinity of TRP was 5 orders of magnitude smaller than that of bisnorbiotin methyl ketone, we conclude that neither LAA nor TRP interfere with the measurement of biotin or its metabolites by the HPLC/avidin-binding assay.

Original languageEnglish (US)
Pages (from-to)A802
JournalFASEB Journal
Volume10
Issue number3
StatePublished - Dec 1 1996

Fingerprint

lipoic acid
Thioctic Acid
avidin
Avidin
biotin
Biotin
Tryptophan
tryptophan
Assays
high performance liquid chromatography
High Pressure Liquid Chromatography
assays
Metabolites
metabolites
ketones
Biotinidase
Horseradish Peroxidase
Chromatography
Ac-Nal(1)-Cpa(2)-Pal(3,6)-Arg(5)-Ala(10)-lHRH
chromatography

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

Cite this

Lipoic acid analogs and L-tryptophan do not interfere with the assay of biotin by HPLC/avidin-binding assay. / Zempleni, J.; Mock, P. M.; Stratton, S. L.; McCormick, D. B.

In: FASEB Journal, Vol. 10, No. 3, 01.12.1996, p. A802.

Research output: Contribution to journalArticle

Zempleni, J. ; Mock, P. M. ; Stratton, S. L. ; McCormick, D. B. / Lipoic acid analogs and L-tryptophan do not interfere with the assay of biotin by HPLC/avidin-binding assay. In: FASEB Journal. 1996 ; Vol. 10, No. 3. pp. A802.
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