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A lactase-producing psychrophilic bacillus strain and method for preparing low-temperature lactase using the strain

A psychrophilic bacteria and lactase technology, applied in the field of microorganisms, can solve the problem that the performance of the strain cannot meet the low temperature lactase industrial production technology and the like

Inactive Publication Date: 2020-01-10
SHIHEZI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0018] The performance of strains producing low-temperature lactase at home and abroad can not meet the requirements of industrial production technology of low-temperature lactase

Method used

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  • A lactase-producing psychrophilic bacillus strain and method for preparing low-temperature lactase using the strain
  • A lactase-producing psychrophilic bacillus strain and method for preparing low-temperature lactase using the strain
  • A lactase-producing psychrophilic bacillus strain and method for preparing low-temperature lactase using the strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Embodiment 1: the cultivation of low-temperature lactase-producing psychrophilic bacillus (Cryobacterium sp.) bacterial strain LW097

[0057] The low-temperature lactase-producing bacteria LW097 of the present invention was inoculated in 1 / 4TSB plate medium supplemented with 0.01% X-Gal, 0.1mM IPTG, wherein soybean broth hydrolyzate 1.25g, casein hydrolyzate 4.25g, sodium chloride 0.5g, glucose 0.75g, disodium hydrogen phosphate 0.425g, distilled water 1000ml, 1.0kg / cm2 steam sterilization for 15 minutes. After being cultured at 16° C. for 72 hours, the colonies of the low-temperature lactase-producing bacteria of the present invention are blue.

Embodiment 2

[0058] Example 2: Fermentation of low-temperature lactase-producing psychrophilic bacillus (Cryobacterium sp.) bacterial strain LW097

[0059]Insert the low-temperature lactase-producing psychrophilic bacillus (Cryobacterium sp.) bacterial strain LW097 of the present invention into the seed culture medium: lactose 10g, yeast extract 10g, NaCl 4g, tryptone 5g, distilled water 1000ml, pH 7.0, 16 ℃ culture 4~ 5 days; then, inoculate in the following fermentation medium with a 5% inoculum amount. Lactose 40g, potassium dihydrogen phosphate 0.05g, tryptone 15g, calcium chloride 0.11g, yeast extract 5g, manganese sulfate 0.001g, magnesium sulfate 0.3g, ferrous sulfate 0.03g, pH 7.0, in a 250ml Erlenmeyer flask, each Bottled in 50ml, 1.0kg / cm2 steam sterilization for 15 minutes. Shake culture at 16°C and 200 rpm for 56 hours. After cultivation, the bacterial cells are collected by centrifugation, and after repeated freezing and thawing at ~70°C and 50°C, the supernatant is collecte...

Embodiment 3

[0060] Embodiment 3: the assay method of low temperature lactase enzyme activity

[0061] Low-temperature lactase activity was determined using a colorimetric assay. With o-nitrobenzene~β~D~galactoside (oNPG) as the substrate, the enzymatic activity was measured by the amount of enzymatic hydrolysis product o-nitrophenol (oNP) produced per unit volume of unit enzyme solution per unit time. calculate.

[0062] Definition of enzyme activity: Under certain conditions, 1 μmol oNP generated by hydrolyzing oNPG in 1 minute is defined as 1 enzyme activity unit. The oNP content was calculated from the standard curve and the OD value determined at 420 nm. Under the above conditions, the hydrolysis of oNPG in 1 min to generate 1 μmoloNP was defined as 1 enzyme activity unit. The specific method is as follows:

[0063] A: Drawing of oNP standard curve: take oNP139.0mg (o-nitrophenol) standard substance and put it into a 100ml volumetric flask, add 10ml of 95% ethanol to dissolve it, ...

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Abstract

The invention discloses a new strain of psychrophilic bacillus (Cryobacterium sp. LW097) and a method for producing low-temperature lactase by utilizing the strain, and provides the low-temperature lactase at the same time. By using the new strain of psychrophilic bacteria Cryobacterium sp.LW097 (preservation number is CCTCC M 2017015), the method of strain selection, preservation and rejuvenation was established, and a stable fermentation method was established after optimization of the medium formula and optimization of the fermentation process conditions , the produced lactase has high enzymatic activity at low temperature, and has application value in the processing of low-lactose dairy products.

Description

technical field [0001] The invention relates to a low-temperature lactase, a bacterium producing the enzyme and a preparation method thereof. Specifically, the present invention relates to a lactase produced by a new strain LW097 of psychrophilic bacteria (Cryobacterium sp.), which has higher activity at a lower temperature, and the production of low-temperature lactase produced by using the microbial strain The method belongs to the technical field of microorganisms. Background technique [0002] Lactose is present in the breast milk of most mammals, including cow's milk. After humans consume lactose in dairy products, it is mainly hydrolyzed by the lactase phlorizin hydrolase distributed on the brush border cell membrane of the small intestine. However, due to genetic background and dietary differences, more than 90% of the Asian population is deficient in lactase. Lactose cannot be digested and absorbed after consuming dairy products, resulting in symptoms of lactose in...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N9/24C12N15/56C12R1/01
CPCC12N9/2402C12Y302/01108C12N1/205C12R2001/01
Inventor 关波胡有贞倪永清李梦王丽军
Owner SHIHEZI UNIVERSITY