Abstract
Introduction. The traditional curing of vanilla pods includes "killing" and sweating steps when pods are exposed to heat (35-65 degrees C) for various lengths of time. Although it is known that liberation of vanillin and other phenolics from their non- aromatic glucosides is due to the action of an endogenous beta-glucosidase, its in vivo kinetics remained unknown. Materials and methods. Mature green vanilla pods were pretreated for 2 h at 50 degrees C, 55 degrees C and 60 degrees C, then stored for 118 days at 27 degrees C. Phenolic glucosides and their aglycons were extracted at regular intervals during the storage period and analyzed by HPLC. Results and discussion. All phenolic beta-glucosides were slowly hydrolyzed during the storage period with production of vanillin, p-hydroxybenzaldehyde, vanillic acid, and other unknown aglycons. Most of the beta-glucosidase was heat-denatured by the pretreatment, and analysis of its kinetic parameters showed that it adopts, in vivo, an allosteric mode of functioning with a lower affinity for glucovanillin than in vitro, where it behaves as a Michaelian enzyme. Conclusion. Extensive research is needed to confirm the allosteric mechanism of the vanilla beta-glucosidase in vivo.