A rapid method for detection of alliinase activity in Allium, especially in the subgenus Melanocrommyum

In the genus Allium about 800 species are currently known belonging to several subgenera [1]. Wild Allium species can be found on the northern hemisphere with a main habitat in Central Asia. The enzyme alliinase occurs in Allium plants catalyzing the cleavage of cysteine sulphoxides leading to typical odourous compounds [2]. Whereas the alliinases of common Allium species like garlic and onion [3] are well analyzed, the properties of other alliinases occurring in further Allium species have not been examined yet. Especially the species of subgenus Melanocrommyum do not express an alliinase like garlic, as can be shown on SDS-PAGE. The separation of alliinase from other proteins is sometimes difficult because of similar protein properties due to their size and probably also due to glycosilation. Therefore, a new method for direct alliinase activity detection has been developed.

A crude alliinase preparation can be separated on a basic-native polyacrylamide gel. This method results in functional enzymes, separated into different spots. These protein spots can be cut out of the gel and screened for their alliinase activity. The L-(+)-S-(3-pyrrolyl)cysteine sulphoxide used as indicator turns into a red dye after the enzymatic cleavage by alliinase (Figure 1) [4]. Although the alliinase is still incorporated inside the gel matrix, a positive reaction can be detected after a few minutes (Figure 2). This new test allows an easy and quick detection of alliinase-like enzymes. Furthermore, the amount of sample needed is very small, allowing tests out of a single bulb.

Figure 1: positive reaction b) negative reaction

Figure 2: indicator reaction

Keywords: Allium, Melanocrommyum, alliinase, basic-native polyacrylamide gel

Acknowledgement: Institute of Pharmaceutical Chemistry, Regina Gerlach-Riehl

References: 1 Fritsch R M. et al. (2010) Phyton (Horn, Austria) 49(2):145–220

2 Stoll A, Seebeck E (1949) Helvet Chim Acta 32(1):197–205

3 Nock L P, Mazelis M (1987) Plant Physiol 85(4):1079–1083

4 Jedelská J et al. (2008)J Agric Food Chem 56(4):1465–1470