Fermentation preparation and extraction method for bacillus subtilis debitterized aminopeptidase

A technology of Bacillus subtilis and aminopeptidase, which is applied in the field of enzyme preparations and food additives, can solve problems such as unpleasantness, affecting food flavor, and restricting promotion and application.

Active Publication Date: 2009-07-29
江苏博立生物制品有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Different degrees of bitterness of soybean polypeptide products will directly affect the flavor of food, and it will be "unpleasant" when eaten by people, which also limits its promotion and application in the food industry to a certain extent

Method used

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  • Fermentation preparation and extraction method for bacillus subtilis debitterized aminopeptidase

Examples

Experimental program
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Effect test

Embodiment 1

[0032] The slant of Bacillus subtilis Zj016 was inserted into the seed solution, and after expanded cultivation, it was then inserted into 4L fermentation medium according to the inoculum size of 4%. The initial pH is 7.0-8.0, 35-40°C, 400-1000rpm, ventilation rate 2:1, minimum DO is controlled at 60%, DO is coupled with rotational speed, and cultivated for 15 hours. Centrifuge the fermented liquid at 4° C. at 10,000 rpm for 10 min to collect 3.28 L fermented liquid, and add 15% (NH 4 ) 2 SO 4 , 4 DEG C, 10000rpm centrifugation 10min collects clarified liquid, selects the ultrafiltration membrane of 30kDa to carry out concentrating appropriate times, adds mass fraction again in concentrated liquid and is 25% (NH 4 ) 2 SO 4 Salt out, centrifuge at 10,000 rpm at 4°C for 10 minutes to collect the precipitate, and freeze-dry the precipitate to obtain solid aminopeptidase.

Embodiment 2

[0034] The enzyme system and substrate specificity of the aminopeptidase prepared in Example 1 were studied, and its acidic, neutral and alkaline protease activities were measured respectively, and the results showed no related endoprotein activity. Further studies have found that it has exoaminopeptidase activity, and its activity is the strongest for the substrate L-leucine-p-nitroaniline (Leu-pNA), which can reach about 3900U / mL, and for the substrate L-glutamine The activity of amino acid p-nitroaniline (Glu-pNA) is only 391U / mL, while the substrate is L-glycine-proline p-nitroaniline (Gly-Pro-pNA) has no hydrolysis ability, indicating that its opposite end is The more hydrophobic amino acids are more effective.

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Abstract

The invention discloses a method for preparing and extracting Bacillus subtilis debittering aminopeptidase by fermentation, which belongs to the technical field of enzyme preparation and food additive. The invention uses the fermentation cylinder production condition of Bacillus subtilis Zj016 to collect fermentation liquor and obtain the product of debittering aminopeptidase by carrying out clarifying, ultrafiltration concentration and salting out on the fermentation liquor. The aminopeptidase produced by the selected Bacillus subtilis Zj016 is exopeptidase which can dissociate amino acid from a polypeptide chain amino terminal. Researched by the laboratory, the aminopeptidase cooperates with alkali protease to hydrolyze soybean protein isolate; the prepared soybean peptide does not have bitterness, and the contents of oligopeptide like dipeptide and tripeptide are higher. Besides, researches on the enzyme lines produced by the Bacillus subtilis show that the Bacillus subtilis only produces circumscribed prolease and does not have the activity of endo protease; moreover, the circumscribed prolease has a stronger hydrolysis capacity to the terminal amino acid with strong hydrophobicity. The research further explains that the invention can reduce or eliminate the bitterness generated during hydrolysis of the soybean protein.

Description

technical field [0001] A method for large-scale fermentation preparation and extraction of Bacillus subtilis depicteric aminopeptidase, specifically using Bacillus subtilis to ferment, collecting the fermentation broth, and obtaining depicteric aminopeptidase products through clarification, ultrafiltration concentration and salting out The invention belongs to the technical field of enzyme preparation and food additives. Background technique [0002] Protein hydrolysis produces peptides and a small amount of amino acids, whose solubility, thermal stability and nutritional properties are significantly improved, and are important raw materials in modern food processing. However, bitter bitter peptides are often produced in the hydrolyzate. This bitter peptide is generally a polypeptide chain with a single or multiple hydrophobic amino acids at the amino end, and sometimes it may be a hydrophobic amino acid at the carboxyl end. The bitter taste limits the application of protein...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/56C12R1/125
Inventor 田亚平王俊吕广林
Owner 江苏博立生物制品有限公司
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