An agar degrading bacterium Pseudomonas aeruginosa AG LSL-11 was acclimatized to alginate for the production of alginase. Production parameters such as pH, temperature, influence of simple carbohydrates and nitrogen sources, and effect of NaCl on growth and alginase production were carried out. Maximum growth was observed at pH 9.0 and 35 °C, while alginase was produced optimally at pH 9.0 and 30 °C. The alginase produced by Pseudomonas aeruginosa AG LSL-11 was inducible by alginate, and repressed by other simple sugar when supplemented along with alginate in the medium. The bacterium did not require NaCl for growth and production of alginase. The activity staining of partially purified culture supernatant after native PAGE revealed the presence of a single alginase.
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.
An agar degrading bacterium Pseudomonas aeruginosa AG LSL-11 was acclimatized to alginate for the production of alginase. Production parameters such as pH, temperature, influence of simple carbohydrates and nitrogen sources, and effect of NaCl on growth and alginase production were carried out. Maximum growth was observed at pH 9.0 and 35°C, while alginase was produced optimally at pH 9.0 and 30°C. The alginase produced by Pseudomonas aeruginosa AG LSL-11 was inducible by alginate, and repressed by other simple sugar when supplemented along with alginate in the medium. The bacterium did not require NaCl for growth and production of alginase. The activity staining of partially purified culture supernatant after native PAGE revealed the presence of a single alginase.
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