Method for stabilizing nitrilase activity and preserving microbial cells
A technology of microbial cells and nitrilase, applied in the direction of microorganism-based methods, preserved microorganisms, biochemical equipment and methods, etc.
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Embodiment 1
[0068] Storage Stability of Nitrilase Activity of Unimmobilized Acidovorax facilis Cells
[0069] Acidovorax facilis 72W cells were separated from the culture medium by centrifugation, and then suspended in potassium phosphate solution (0.10 or 1.00M), sodium acetate solution (0.10 or 1.00M) or sodium carbonate solution (0.10 or 0.30M) at pH 7.3 ), the concentration is 1-5% of the dry weight of the cells, and the final suspension is stored in a capped glass bottle at 5° C. After a long time, samples are taken from the cell suspension to analyze the nitrilase activity.
[0070] Table 1 illustrates the relative nitrilase activity of the stored cell suspensions, the nitrilase activity of the suspension stored for 0 days was defined as 100%. Cells stored in 0.10M or 0.30M sodium carbonate solution retained a substantially higher percentage of initial nitrilase activity at 92 days compared to cells stored in 0.10M or 1.0M sodium acetate or potassium phosphate solution.
[007...
Embodiment 2
[0073] Immobilization of Acidovorax facilis 72W in carrageenan particles
[0074] Acidovorax facilis 72W wet cell pellet (45 g) was mixed with 45 ml of 0.88% NaCl and the final suspension was heated at 50°C for 1 hour to inactivate the undesired nitrile hydratase activity. Then the cell suspension at 50° C. was added to 135.0 g of 5% by weight Pronova ISAGE L300 carrageenan solution at 55° C. with mixing. The final suspension was immediately cooled to 5°C ice / water bath for 1 hour to gel. The final gel was cut into particles with a diameter of about 2 mm, and the immobilized cell particles were kept in 450 ml of 0.30 M potassium chloride, 20 mM potassium dihydrogen phosphate (adjusted to pH 7.0 with potassium hydroxide) solution at 5 ° C for 18 hours And hardened. The sclerosing solution was removed by washing the immobilized cell pellet three times with 5 mM potassium chloride, 20 mM potassium dihydrogen phosphate (pH 7.0) at 5°C.
Embodiment 3
[0076] Immobilization of Acidovorax facilis 72W in Polyacrylamide Gel Particles
[0077] A solution of 13.8 g of acrylamide and 1.20 g of methylenebisacrylamide dissolved in 15 ml of water at 25 ° C was added with stirring to 10 ° C 30 g (wet weight of cells) Acidovorax facilis 72W was added with stirring 82 ml of 0.10 M potassium dihydrogen phosphate (pH7 .0) in suspension. The final mixture was added to diaminobutane (0.45ml) and 7.5ml of a 5% (w / v) potassium persulfate solution at 25°C. The final polymer gel was kept in an ice bath at 10° C. for 1 hour, and then cut into particles with a diameter of about 2 mm. The immobilized cell pellet was washed twice with 50 mM potassium dihydrogen phosphate (pH 7.0) solution at 5°C.
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