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. 2016 Jan 20:5:57.
doi: 10.1186/s40064-015-1621-3. eCollection 2016.

Cocoa pod husk, a new source of hydrolase enzymes for preparation of cross-linked enzyme aggregate

Faridah Yusof  1 , Soofia Khanahmadi  1 , Azura Amid  1 , Safa Senan Mahmod  1
Affiliations

Cocoa pod husk, a new source of hydrolase enzymes for preparation of cross-linked enzyme aggregate

Faridah Yusof et al. Springerplus. .

Abstract

Cocoa pod husk (CPH) is a by-product of cocoa production obtained after removing the beans from the fruit. The analysis of CPH has shown that it contains high amounts of protein. This study is aimed to utilize this protein source in hydrolase enzyme production. In this study, seven hydrolase enzymes (amylase, fructosyltransferase, mannanase, glucosidase, glucanase, lipase and protease) were screened from CPH for the first time for feasible industrial production. Among these hydrolases, lipase was chosen for the next steps of experiments as it has a lot of applications in different industries. The extraction of high active lipase from CPH has been done under optimum conditions. The condition that was optimum for the three major factors was achieved using Face centered central composite design (FCCCD) with response surface methodology (RSM) to obtain the highest enzyme activity of crude lipase from CPH. The optimum condition of extraction is used for preparation of cross-linked enzyme aggregate (CLEA). For the production of immobilized biocatalyst, the technique of CLEA is considered as an effective technique for its industrially attractive advantages. Referring to the results of OFAT, CLEA-lipase was prepared in the best condition at the presence of 30 mM ammonium sulphate, 70 mM glutaraldehyde with 0.23 mM Bovine serum albumin as an additive. Immobilization effectively improved the stability of lipase against various organic solvents.

Keywords: CLEA-lipase; Cocoa pod husk; Face centered central composite design; Hydrolase enzymes; Response surface methodology.

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Figures

Fig. 1
Fig. 1
Measuring the activity of various hydrolases from CPH
Fig. 2
Fig. 2
SDS-PAGE of lipase extracted from CPH. Lane 1 molecular weight marker, Lane 2 molecular weight of lipase
Fig. 3
Fig. 3
Schematic picture of CLEA preparation with addition of BSA as additive
Fig. 4
Fig. 4
Effect of different concentration of ammonium sulphate (10–50 %w/v) on CLEA-lipase activity from CPH
Fig. 5
Fig. 5
Effect of different concentration of glutaraldehyde (40–80 mM) on CLEA- lipase activity from CPH
Fig. 6
Fig. 6
Effect of different concentration of BSA (0–0.37 mM) on CLEA-lipase activity from CPH
Fig. 7
Fig. 7
Comparison in the resistance of both the free lipase and its CLEA against hydrophilic solvents. Free lipase and CLEA were incubated at 45 and 60 °C, respectively, for 30 min in various buffer/solvent mixtures: methanol (a), dioxane (b), and acetone (c), and their residual activities were determined. A value of 100 % refers to the activity obtained by the enzyme in the solvent-free buffer solution
Fig. 8
Fig. 8
FE-SEM picture of CLEA-lipase from CPH
See this image and copyright information in PMC

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