Three phase partitioning for simultaneous purification of aloe polysaccharide and protein using a single-step extraction

AbstractIn this study, for the first time Bacillus methylotrophicus Y37 cellulase was purified and recovered in a single step by three-phase partitioning (TPP). The optimal purification parameters for TPP were 40% ammonium sulfate saturation (m/v) with a 1.0:1.0 (v/v) ratio of crude extract:t-butanol, which gave 5.8-fold purification with 155% recovery of cellulase. Non-covalent immobilization of the partitioned cellulase was performed using bentonite as a support material. The activity observed in the 20th experiment was 100%. The optimal pH values and temperatures determined for the free enzyme and the immobilized enzyme were 5.0 and 6.0 and 45°C and 50°C, respectively. The Arrhenius activation energy (Ea) of the immobilized enzyme was lower than that of the free enzyme, whereas the Michaelis–Menten constant (Km) and maximum velocity (Vm) of the immobilized enzyme increased. The turnover number (kcat) and the catalytic performance (kcat/Km) demonstrated the improved catalytic properties of the immobilized enzyme compared to the free enzyme. Immobilization of cellulase is thermodynamically preferred.

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Purification, recovery, and characterization of chick pea (Cicer arietinum) β-galactosidase in single step by three phase partitioning as a rapid and easy technique

Protein Expression and Purification ◽

10.1016/j.pep.2013.08.003 ◽

2013 ◽

Vol 91 (2) ◽

pp. 155-160 ◽

Cited By ~ 11

Author(s):

Yonca (Avci) Duman ◽

Erdem Kaya

Keyword(s):

Cicer Arietinum ◽

Single Step ◽

Chick Pea ◽

Phase Partitioning ◽

Three Phase

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Three phase partitioning as a rapid and efficient method for purification of plant-esterase from wheat flour

Polish Journal of Chemical Technology ◽

10.2478/pjct-2020-0015 ◽

2020 ◽

Vol 22 (2) ◽

pp. 42-49

Author(s):

Liang Dong ◽

Linxin He ◽

Danqun Huo

Keyword(s):

Wheat Flour ◽

Single Step ◽

Phase Partitioning ◽

Surface Method ◽

Three Phase ◽

High Efficient ◽

New Type ◽

Purification Plant ◽

Very High

AbstractThree-phase partitioning (TPP) was used to purify plant-esterase from wheat flour. Effect of various process parameters has been evaluated and plant-esterase was purified to 11.35-fold by optimized single step TPP system (50%, (w/v) (NH4)2SO4 saturation, 1:1 (v/v) ratio of crude extract: t-butanol at pH 4).The enzyme was found to be exclusively partitioned in the aqueous phase. Using TPP system, plant-esterase quickly purified to homogeneity with very high purity and activity. On the basis of single factor research, purification process was optimized by using response surface method, established a new type of high efficient purification plant-esterase method. To the best of our knowledge, this is the first report for purification and characterization of plant-esterase by using three phase partitioning (TPP). The results indicated that, TPP is a simple, quick, economical and very attractive process for purification of plant-esterase compared to conventional chromatographic protocols.

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Quality Analysis of Fresh Cheese Prepared Using Wrightia tinctoria Proteases

Current Nutrition & Food Science ◽

10.2174/1573401316666200214102828 ◽

2020 ◽

Vol 16 (9) ◽

pp. 1309-1317

Author(s):

Anusha Rajagopalan ◽

Bindhu O. Sukumaran

Keyword(s):

Textural Properties ◽

Artemia Salina ◽

Quality Analysis ◽

Human Consumption ◽

Cheese Making ◽

Phase Partitioning ◽

Milk Clotting ◽

Three Phase ◽

Fresh Cheese ◽

Wrightia Tinctoria

Background: Aqueous leaf extract of Wrightia tinctoria has been in use for artisanal cheese preparation in diverse parts of India. Scientific validation behind the milk clotting potential of W. tinctoria stem proteases purified through three-phase partitioning (TPP) has been attempted. However, its contribution to cheese quality standards has not been tried yet. Objective: To evaluate the suitability of three-phase partitioned W. tinctoria (WT) stem proteases in fresh cheese making. Methods: Fresh cheese was prepared using W. tinctoria TPP proteases and its characteristics were compared with that of commercial plant milk coagulant, Enzeco® (CEz) and rennin (CRn). Yield, organoleptic and textural properties, total fat, protein and moisture content of all cheese were determined according to FSSAI standards. Toxicity of the TPP proteases was assessed on the shrimp model (Artemia salina egg and Litopenaeus vannamei post-larval shrimps). Results: TPP proteases were found to be non-toxic and safe for human consumption with no change in egg hatchability and survival of the shrimps in comparison to that of control. Cheese analysis results indicated perceptible resemblance in nutritional characteristics of WT cheese with control cheese. Yield, textural properties and organoleptic acceptance of WT cheese resembled more closely with that of CEz cheese. Conclusion: Observation from the study paves the way for the acceptance of W. tinctoria proteases as a suitable vegetable rennet for fresh cheese making.

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