Protease prospecion and determination of its isoenzymes activity in cocoa cultivars (Theobroma cacao L.)

Abstract

Dur objective was to characterize the protease enzymatic activity and its isoenzymes on cacao cultivars PH 16 and TSH 1188, produced in southern Bahia, linking it to the conditions of the fermentation process. Proteases were extracted and semi-purified, their activities determined changing substrate, pH and temperature, and the values compared with the parameters of fermentation (pH and temperature), and yet determined the kinetic parameters and the activity of its isoenzymes: aminopeptidase, carboxypeptidase and endoprotease. In the experimental conditions, differences in protease activities were shown as to different cocoa cultivars on various conditions. The albumin stands out as the preferred enzyme substrate, in a pH range between and 6 and temperatures between 29 ° C and 50 ° C. As for the isoenzymes activity, an increased activity in these seeds and in the cultivar TSH 1188 was observed. When correlated with the fermentative parameters, the conditions for enzymatic activity are not the best determined, with emphasis on farming pH 16 presenting its fermentation conditions far from those found as optimal, especially in pH evaluation, since temperature varies very little between the two cultivars.Keywords: aminopeptidase; carboxypeptidase; endoprotease; peptide bonds; amino acids. Practical Application:The work characterizes proteases and determines its isoenzymes activity, highlighting and discussing its importance in the formation of precursors of the chocolate flavor. The knowledge on the enzyme characteristics opens possibilities for future technological interventions in the cocoa fermentation process, to improve the quality of the raw material and, consequently, of monovarietal chocolate.

Protein reservoirs of seeds are amyloid composites employed differentially for germination and seedling emergence

“…At fixed time points, the proteins were precipitated with 1% tri‐chloro acetic acid and centrifuged at 6000 rpm (2000 g ) for 15 min. The supernatant was checked for amino acid content using ninhydrin assay, by measuring absorbance at 570 nm using Perkin Elmer micro‐plate reader (Silveira et al., 2017).…”

“…At fixed time points, the proteins were precipitated with 1% tri-chloro acetic acid and centrifuged at 6000 rpm (2000 g) for 15 min. The supernatant was checked for amino acid content using ninhydrin assay, by measuring absorbance at 570 nm using Perkin Elmer micro-plate reader (Silveira et al, 2017). For sustained release assay, each substrate (albumin, globulin, their dialyzed aggregates, SSPB, and SSPB without membrane) and the endopeptidase fraction (0.5 mg ml À1 ) were incubated together in the upper chamber of a 200-nm transwell filter, as commonly employed for checking sustained degradation of drugs and other biomolecules (Rohrschneider et al, 2015).…”

Application of a Mix of Vegetables Residues as Inhibitor for Carbon Steel

“…In the cocoa productive chain, the fruit is not fully utilized, generating large amounts of waste, which estimations indicate the disposal of 80%. Most of the cocoa barks are left in the plantations in the form of "casqueiros", which can provoke the degradation of the soil structure as a source of inoculum of pathogenic fungi toward the plants, harming the environment 57 .…”

Multivariate analysis of chemical parameters of cocoa: a contribution to origin designation / Análise multivariada de parâmetros químicos do cacau: uma contribuição para a denominação de origem

“…Variation in protein content can occur for a number of factors. During fermentation, for example, biochemical reactions occur that lead to protein reduction due to the action of enzymes or complexation with phenolic compounds (Silveira et al, 2017;D'Souza et al, 2018). However, there are studies that attribute variations in protein content to the total nitrogen content of the soil where the cocoa was grown, as well as to the genetic variety of cocoa (Adewole et al, 2011).…”