Identification of glucosides in green beans of Vanilla planifolia Andrews and kinetics of vanilla β-glucosidase

Natural vanilla is extracted from the fruits of Vanilla planifolia. In the overall vanilla aroma, minor compounds p-cresol, creosol, guaiacol and 2-phenylethanol have a high impact. This is shown by GC-Olfactometry analysis of cured vanilla beans. The presence of β-D-glucosides of these compounds was investigated, in order to determine if these compounds are derived from glucosides or if they are formed during the curing process via different pathways. Glucosides of vanillin, vanillic acid, p-hydroxy benzaldehyde, vanillyl alcohol, p-cresol, creosol and bis[4-(β-d-glucopyranosyloxy)-benzyl]-2-isopropyltartrate and bis[4-(β-d-glucopyranosyloxy)-benzyl]-2-(2-butyl)tartrate have been identified in a green bean extract. The kinetics of the β-glucosidase activity from green vanilla beans towards eight glucosides naturally occurring in vanilla and towards p-nitrophenol were investigated. For glucosides of p-nitrophenol, vanillin and ferulic acid the enzyme had a Km of about 5 mM. For other glucosides (vanillic acid, guaiacol and creosol) the Km-values were higher (>20 mM). The Vmax was between 5 and 10 IU mg−1 protein for all glucosides tested. Glucosides of 2-phenylethanol and p-cresol were not hydrolysed. β-Glucosidase does not have a high substrate specificity for the naturally occurring glucosides compared to the synthetic p-nitrophenol glucoside (Km 3.3 mM, Vmax 11.5 IU mg−1 protein).