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Purification of Porcine Pancreatic Lipase by Aqueous Two-Phase Systems of Polyethylene Glycol and Potassium Phosphate

Purification of Porcine Pancreatic Lipase by Aqueous Two-Phase Systems of Polyethylene Glycol and Potassium Phosphate

Yu-Jie Zhou, Cheng-Li Hu, Na Wang, Wei-Wei Zhang, Xiao-Qi Yu

J Chromatogr B Analyt Technol Biomed Life Sci. 2013 May 1;926:77-82.

PMID: 23562905

Abstract:

An aqueous two-phase system (ATPS) was applied for the purification of porcine pancreatic lipase (PPL) from crude PPL using polyethylene glycol (PEG) and potassium phosphate. Phase diagrams for polyethylene glycol (PEG) and potassium phosphate dibasic were determined at room temperature to find an operating region to first form the ATPS. The PPL was preferentially partitioned into the PEG-rich phase in systems with molecular weights of 1000 and 1500 and concentrated in the phosphate-rich phase in systems with PEG of 4000. Moreover, instead of tie line length (TLL), we used a stability ratio without NaCl in the system, and we first applied fluorescence spectroscopy for the protein conformational analysis of the ATPS. The molecular weight of the purified lipase was determined to be approximately 52 kDa by SDS-PAGE. The enzyme was efficiently purified in PEG 1500/potassium phosphate (17/13, %) at a pH of 7.0 at 4 °C. This system obtained an enzyme partition coefficient of 12.7, an extraction efficiency of 94.7% and a purification factor of approximately 4. These results demonstrate that the aqueous two-phase system is a highly efficient method for PPL purification.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP7758114-A	Potassium phosphate dibasic		7758-11-4	Price

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