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Brevibacillus parabrevis producing keratinase and application thereof

A technology of Bacillus brevis and keratinase, applied in the field of microorganisms, can solve problems such as polluting the environment

Inactive Publication Date: 2015-12-02
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, gray-alkali method is basically used for hair removal in tanning industry, but the depilatory waste liquid contains high concentration of sulfide, which seriously pollutes the environment, and the enzymatic method instead of ash-alkali method can completely eliminate the pollution of sulfide. The enzymatic method has a clean Non-toxicity, environmental friendliness and other characteristics, and at the same time, it can improve the quality of the product compared with the chemical tanning process, so it has been more and more widely used in tanning

Method used

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  • Brevibacillus parabrevis producing keratinase and application thereof
  • Brevibacillus parabrevis producing keratinase and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Example 1 Screening and identification of keratinase producing strains

[0016] (1) Medium formula

[0017] ①Preliminary screening medium: wool powder 0.5%, K 2 HPO 4 0.1%, NaCl0.05%, agar powder 2%.

[0018] ②Seed liquid culture medium

[0019] Gao's medium: soluble starch 2%, KNO 3 0.1%, NaCl0.05%, K 2 HPO 4 0.05%, MgSO 4 . 7H 2 O0.05%, FeSO 4 ·7H 2 0.001%.

[0020] LB medium: peptone 1%, yeast powder 0.5%, sodium chloride 1%.

[0021] Cha's medium: 3% sucrose, NaNO 3 0.3%, K 2 HPO 4 0.1%, MgSO 4 . 7H 2 O0.05%, KCl0.05%, FeSO 4 . 7H 2 0.001%.

[0022] ③Initial fermentation medium: MgCl 2 ·6H 2 O0.01%, K 2 HPO 4 0.03%, KH 2 PO 4 0.04%, NaCl0.05%, NH 4 Cl0.05%, wool 1%.

[0023] (2) Plate initial screening and determination of keratin activity

[0024] Soil samples were collected from rural sheep pens in Wuxi, and 1g of soil samples were added to a 50mL Erlenmeyer flask containing 10mL of normal saline and glass beads. The shaker was shake...

Embodiment 2

[0032] Example 2 Preliminary optimization of strain fermentation conditions

[0033] The fermentation medium and fermentation conditions of the strain were optimized by single factor and orthogonal experiments, and the optimal medium formula was wool 1%, soluble starch 1%, corn flour 1%, NaCl 0.15%, NH 4 Cl0.05%, K 2 HPO 4 0.03%, KH 2 PO 4 0.04%, MgCl 2 ·6H 2 O0.01%. Shaker culture conditions are as follows: initial pH value is 8.0, inoculum size is 2%, culture temperature is 40°C, rotation speed is 220r / min, liquid volume is 30mL / 250mL, fermentation period is 16h, and keratinase activity reaches 412U / mL.

Embodiment 3

[0034] Embodiment 3 enzymatic property research

[0035] The enzymatic properties of the crude enzyme produced by the keratinase were preliminarily studied, and the effects of the action temperature (30°C-70°C) and the action pH (6.0-10.0) on the enzyme activity were studied respectively. The applicable temperature is 40°C ( figure 1 ), the optimum pH is 7.0 ( figure 2 ).

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Abstract

The invention discloses a keratinase producing strain and culture conditions and enzymatic properties thereof, and belongs to the technical field of microorganisms. As identified, the strain is Brevibacillus parabrevis, is named as Brevibacillus parabrevis R-7 and now is preserved in China General Microbiological Culture Collection Center, with preservation number CGMCC No.10798. After preliminary optimization, the culture medium of the strain comprises 1 percent of wool, 1 percent of soluble starch, 1 percent of corn meal, 0.15 percent of NaCl, 0.05 percent of NH4Cl, 0.03 percent of K2HPO4, 0.04 percent of KH2PO4 and 0.01 percent of MgCl2 6H2O. Shaker culture conditions include initial pH 8.0, inoculation amount 2 percent, culture temperature 40 DEG C, revolving speed 220r / min, liquid volume 30mL / 250mL and fermentation period 16h, wherein the activity of the produced keratinase reaches 412U / mL. The optimum acting temperature of the keratinase is 40 DEG C and the optimum pH is 7.0. The enzyme has important application in fields such as tanning and medicines.

Description

technical field [0001] The invention belongs to the technical field of microorganisms, and in particular relates to a keratinase-producing Bacillus parabrevius and properties of crude enzymes thereof. Background technique [0002] Keratinase ( Keratinases ) is a proteolytic enzyme that can specifically degrade natural keratin. Keratin has a large number of disulfide bonds, hydrogen bonds and hydrophobic interactions, and is not easily hydrolyzed by common proteases, so most of them are not used, and some even cause environmental pollution. [0003] Keratinase can be used in the leather industry. At present, gray-alkali method is basically used for hair removal in tanning industry, but the depilatory waste liquid contains high concentration of sulfide, which seriously pollutes the environment, and the enzymatic method instead of ash-alkali method can completely eliminate the pollution of sulfide. The enzymatic method has a clean Non-toxicity, environmental friendliness and...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/48C12R1/01
Inventor 龚劲松侯璎朔罗阳翟靖鑫张楠朱佳能史劲松许正宏李恒
Owner JIANGNAN UNIV
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