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Solution and method to release L-asparaginase inside cell

A technology of asparaginase and cells, which is applied in the field of biochemistry, can solve undiscovered problems, achieve low cost, high release rate, and reduce the cost of raw materials

Inactive Publication Date: 2003-09-03
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After searching the literature of the prior art, it has not been found so far that the Escherichia coli cells are treated with phosphate and Triton non-ionic surfactants, so that the intracellular L-asparaginase is released to the extracellular solution and method

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0010] Escherichia coli ATCC 11303 was cultured in a shaker flask for 15 hours, and the cells were obtained by centrifugation. Wash the cells with distilled water and centrifuge again. The obtained bacteria were suspended in a solution prepared with distilled water. The total volume of the liquid is 20ml, containing 1g of wet cells, 10% of dipotassium hydrogen phosphate, and 2% of Triton X100, and placed in a 250ml shake flask. Shake at 25°C for 20 hours and centrifuge. Cells pellet at the bottom of the centrifuge tube. Take the supernatant, which contains L-asparaginase, and the release rate is 61%.

Embodiment 2

[0012] Escherichia coli ATCC 11303, the process of cultivating and washing the thalline was the same as Example 1. The obtained bacteria were suspended in a solution prepared with distilled water. The total volume of the liquid is 20ml, containing 1g of wet cells, 15% of dipotassium hydrogen phosphate, and 2% of TritonX100, and placed in a 250ml shake flask. Shake at 25°C for 20 hours and centrifuge. Cells pellet at the bottom of the centrifuge tube. Take the supernatant, which contains L-asparaginase, and the release rate is 65%.

Embodiment 3

[0014] Escherichia coli ATCC 11303, the process of cultivating and washing the thalline was the same as Example 1. The obtained bacteria were suspended in a solution prepared with distilled water. The total volume of the liquid is 20ml, containing 1g of wet cells, 12.5% ​​of dipotassium hydrogen phosphate, and 2% of Triton X100, and placed in a 250ml shake flask. Shake at 25°C for 20 hours and centrifuge. Cells pellet at the bottom of the centrifuge tube. Take the supernatant, which contains L-asparaginase, and the release rate is 70%.

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PUM

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Abstract

Colon bacillus culture liquid is centrifugally settled or filtered to eliminate liquid and obtain colon bacillus cell. Phosphate 10-15%, Triton surfactant 1-45% and water are compounded into a solution, and colon bacillus cell is added into the solution, which is then stirred in a stirrer or shaked in a shake flash for same period so that L-asparaginase is released from cell to liquid at normal temperature 4-37 deg.C for preferably 4-10 hr. After centrifugal settlement or filtration to eliminate the cell, the L-asparaginase in solution is further purified.

Description

technical field [0001] The invention relates to a solution and a method for releasing intracellular enzymes, in particular to a solution and a method for releasing intracellular L-asparaginase (EC 3.5.1.1), belonging to the field of biochemistry. Background technique [0002] Many microbial metabolites are present intracellularly. Usually, the cells need to be broken to release the intracellular products to the outside of the cells for the next step of extraction. Commonly used methods for disrupting cells include mechanical disruption, ultrasonic treatment, osmotic pressure shock, chemical reagent treatment, and lysozyme treatment. L-asparaginase (EC 3.5.1.1) produced by Escherichia coli is a drug for the treatment of leukemia and belongs to intracellular enzymes. The method of making acetone powder is generally used at home and abroad to destroy the cell wall. That is, E. coli cells were added to acetone to disrupt the cells. Acetone was removed by filtration to obtain...

Claims

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Application Information

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IPC IPC(8): C12N9/82
Inventor 赵凤生
Owner SHANGHAI JIAO TONG UNIV