National Academy of Agricultural Sciences (NAAS)
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PRINT ISSN : 2319-7692
Online ISSN : 2319-7706 Issues : 12 per year Publisher : Excellent Publishers Email : editorijcmas@gmail.com / submit@ijcmas.com Editor-in-chief: Dr.M.Prakash Index Copernicus ICV 2018: 95.39 NAAS RATING 2020: 5.38 |
Ten Lactobacillus cultures were considered for the study. Out of ten Lactobacillus cultures, V3 produced maximum β-galactosidase activity (3.381 O.D), followed by NK2 and NK9 up to 24h than other cultures during β-galactosidase activity. Extraction of β-galactosidase was carried out by disruption of Lactobacillus cells using sonication at 3, 4 and 5 minute (pulse 30 second on / 10 seconds off and 35 to 55 % amplitudes), lysozyme-EDTA (49 mg of lysozyme in 1.5 ml of TE buffer) treatment, toluene acetone (9:1 [toluene: acetone]) treatment, sodium dodecyl sulphate (SDS) - chloroform (250 µl chloroform and 100 ml of 0.1 % SDS) treatment. Out of 4 different treatments [sonication, lysozyme-EDTA, toluene-acetone and sodium dodecyl sulphate (SDS)-chloroform treatment] for β-galactosidase production, sonication treatment [V3 (1.093), NK2 (1.084) and NK9 (1.091)] was found to be best for production of β-galactosidase with optimized sonication period [5 min (pulse 15 sec off / 30 sec on at 55% amplitudes)]. Purification of β-galactosidase enzyme was carried out employing various t-butanol ratios (crude extract : t-butanol = 1:0.25, 1.0:0.5, 1.0:1.0, 1.0:1.5, 1.0:2.0) with a different ammonium sulfate saturation i.e. 30, 60 and 90%.After purification by treating with ratio of crude enzyme: t-butanol (R) and ammonium sulphate saturation (AS), 2 % of R (0.396 O.D) and 60 % of AS (0.292 O.D) showed maximum β-galactosidase activity for three selected Lactobacillus cultures i.e. V3, NK2 and NK9. The molecular mass of partially purified β-galactosidase (60% Ammonium sulphate saturation) from selected Lactobacillus cultures i.e. V3, NK2 and NK9 as determined by SDS-PAGE was approx. 69 kDa. Therefore, the study entailed to conclude that production and purification of β-galactosidase enzyme from safe Lactobacillus cultures is an alternative source of biological enzyme for the preparation of lactose hydrolysed milk for lactose intolerant population.
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