Aqueous two-phase extraction for the purification of alkaline agarases from culture extracts of Pseudomonas aeruginosa AG LSL-11.
The agarases were purified for the first time an using aqueous two-phase system (ATPS) consisting of polyethylene glycol (PEG) and phosphate salt. The three extracellular, alkaline agarases produced by Pseudomonas aeruginosa AG LSL-11 were efficiently extracted into the top PEG-rich layer. The influencing factors on the partition of agarases--molecular weight of the PEG, system pH, system temperature, and NaCl concentration--were investigated. All the factors were found to have a significant effect on the partition of agarases except NaCl. The optimal ATPS parameters for the partitioning and purification of agarases were found to be 12% PEG 600 and 11.9% (w/w) phosphate salt at pH 8.0 and 4°C. All three agarases were concentrated in the top PEG phase with 6.19-fold purity and 71.21% recovery. The ATPS was found to be more convenient and economical than the conventional ion-exchange chromatography (IEC) method for extraction of three agarases and could be significantly employed for the purification of agarases from fermentation broth.
Department of Biochemistry, Gulbarga University, Gulbarga, Karnataka, India.
Basawaraj A
BA
Koti
M
M
Lakshmikanth
S
S
Manohar
J
J
Lalitha
D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases / Q000302:isolation & purification*; D010710:Phosphates / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D012965:Sodium Chloride / Q000737:chemistry; D014867:Water / Q000737:chemistry
D010710:Phosphates; D014867:Water; D011092:Polyethylene Glycols; D012965:Sodium Chloride; D006026:Glycoside Hydrolases; C019624:agarase