PURIFICATION AND CHARACTERIZATION OF AMYLASE PRODUCED FROM PROBIOTIC LACTOBACILLUS PLANTARUM CS FOR INDUSTRIAL APPLICATIONS

Ujunwa Felicia Nwachukwu; Uzoamaka Ogechi George-Okafor; Kelechi Nkechinyere Mba-Omeje; Amara Chioma Ezeme-Nwafor; Ifeoma Agatha Onah; Ifeanyi Jude Victor Egbuji

doi:10.34016/pjbt.2024.21.02.890

Authors

Ujunwa Felicia Nwachukwu Enugu State University of Science and Technology, Faculty of Applied Natural Sciences, Department of Applied Microbiology and Brewing.
Uzoamaka Ogechi George-Okafor Enugu State University of Science and Technology, Faculty of Applied Natural Sciences, Department of Applied Microbiology and Brewing.
Kelechi Nkechinyere Mba-Omeje Enugu State University of Science and Technology, Faculty of Applied Natural Sciences, Department of Applied Microbiology and Brewing.
Amara Chioma Ezeme-Nwafor Caritas University Amorji-Nike Enugu State, Faculty of Natural Sciences, Department of Microbiology.
Ifeoma Agatha Onah Caritas University Amorji-Nike Enugu State, Faculty of Natural Sciences, Department of Microbiology.
Ifeanyi Jude Victor Egbuji Caritas University Amorji-Nike Enugu State, Faculty of Natural Sciences, Department of Microbiology.

DOI:

https://doi.org/10.34016/pjbt.2024.21.02.890

Keywords:

amylase, characterization, Lactobacillus plantarum CS, purification.

Abstract

Previous studies have demonstrated that probiotic Lactobacillus plantarum CS was able to generate an appreciable amount of extracellular amylase, hence the need to purify and characterize it. The aim of the study was to purify and characterize crude amylase from probiotic Lactobacillus plantarum CS for its industrial applications Three purification steps including ammonium sulphate precipitation, ion exchange chromatography on carboxymethyl sephadex and gel filtration on Sephadex G-75 were utilized. The homogeneity of the purified enzyme was confirmed using sodium deodocyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The purified amylase was characterized on different parameters including substrates hydrolyses, pH and temperature activity and stability profiles. The general purification elution profile revealed two different peaks of amylase activities with outstanding one having a molecular weight of 59.7kDa. Its purification fold was 4.0 with specific activity of 16.44U/mg protein and enzyme yield of 3%. Temperature optimal activity and stability was at 40⁰C and 7.5 for pH activity and stability. Mangenese (Mn²⁺) (135.17%), tween 80 (128.30%) and some food condiments garlic, thyme, ginger, and tumeric) significantly (p> 0.05) enhanced amylase activity (≥262.40%). However, selenium (Se⁴⁺) and hydrogen peroxide (H₂O₂) were observed to have greatest inhibiting effect (≥30.9%) on the enzyme. Substrate hydrolysis profiles showed that the amylase hydrolyzed all the test starchy substrates with the highest hydrolytic potential on indigenous sweet potato starch (Km value/ Vmax of 1.33mg/ml/ 7.89ml). The rate of hydrolysis of other test substrates had yam> rice>cassava>corn with km values ≤ 4.0mg/ml and Vmax ≤ 25ml. The obtained results gave an insight that amylase produced from Lactobacillus plantarum CS met with the possessed properties suitable for any industrial application especially in food

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