A spectrophotometric assay for allicin, alliin, and alliinase (alliin lyase) with a chromogenic thiol: Reaction of 4-mercaptopyridine with thiosulfinates

Talia Miron, Irina Shin, Guy Feigenblat, Lev Weiner, David Mirelman, Meir Wilchek, Aharon Rabinkov

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103 Scopus citations

Abstract

Allicin (diallylthiosulfinate) is the best known active compound of garlic. It is generated upon the interaction of the nonprotein amino acid alliin with the enzyme alliinase (alliin lyase, EC 4.4.1.4). Previously, we described a simple spectrophotometric assay for the determination of allicin and alliinase activity, based on the reaction between 2-nitro-5-thiobenzoate (NTB) and allicin. This reagent is not commercially available and must be synthesized. In this paper we describe the quantitative analysis of alliin and allicin, as well as of alliinase activity with 4-mercaptopyridine (4-MP), a commercially available chromogenic thiol. The assay is based on the reaction of 4-MP (λmax = 324 nm) with the activated disulfide bond of thiosulfinates -S(O)-S-, forming the mixed disulfide, 4-allylmercaptothiopyridine, which has no absorbance at this region. The structure of 4-allylmercaptothiopyridine was confirmed by mass spectrometry. The method was used for the determination of alliin and allicin concentrations in their pure form as well as of alliin and total thiosulfinates concentrations in crude garlic preparations and garlic-derived products, at micromolar concentrations. The 4-MP assay is an easy, sensitive, fast, noncostly, and highly efficient throughput assay of allicin, alliin, and alliinase in garlic preparations.

Original languageEnglish
Pages (from-to)76-83
Number of pages8
JournalAnalytical Biochemistry
Volume307
Issue number1
DOIs
StatePublished - 1 Aug 2002
Externally publishedYes

Bibliographical note

Funding Information:
This research was supported by a grant from the Levine Center and from Yeda Co. at the Weizmann Institute of Science.

Funding

This research was supported by a grant from the Levine Center and from Yeda Co. at the Weizmann Institute of Science.

FundersFunder number
Levine Center
Weizmann Institute of Science

    Keywords

    • 2-Nitro-5-thiobenzoic acid
    • 4-Mercaptopyridine
    • Allicin
    • Alliin
    • Alliinase
    • Garlic

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