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Method for preparing t-carrageenase

A technology of carrageenase and seeds, applied in the field of biological enzymes, can solve problems such as less research on ι-carrageenase, and achieve the effect of good application value

Active Publication Date: 2016-09-21
JIMEI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There are more than a dozen reported κ-carrageenases, all of which belong to the glycoside hydrolase 16 family, but ι-carrageenase has been less studied, and only Alteromonas fortis 、 Cellulophaga 、 Zobellia galactanovorans 、 Microbubifer thermotolerans The ι-carrageenase has been studied on the enzymatic properties and the recombinant expression of the enzyme

Method used

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  • Method for preparing t-carrageenase

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preparation example Construction

[0029] 2 Preparation of iota-carrageenase

[0030] (1) Activation of the strain: the carrageenase-producing strain Pseudoalteromonas carrageenovora ASY5 was inoculated in the seed medium, and activated at 23°C for 24 h.

[0031] (2) Fermentation culture: The strains were activated and cultured, then transferred to the fermentation medium, and fermented at 20°C for 40 h.

[0032] (3) Centrifuge the above fermentation broth at 4°C and 10,000 r / min for 20 min, collect the supernatant, and obtain the crude iota-carrageenase enzyme liquid.

[0033] 3. Single factor optimization of ι-carrageenase fermentation process

[0034] The medium composition and culture conditions of the carrageenase-producing strains were optimized by single factor method, and the effects of different factors on the cell density and enzyme production of the strain ASY5 were analyzed. On the basis of shake flask fermentation, the culture time, culture The effects of temperature, inoculum size, liquid volume...

Embodiment 1

[0040] Single factor optimization of ι-carrageenase fermentation process

[0041] (1) Effect of culture time on strain growth and enzyme production

[0042] The activated carrageenase-producing strain Pseudoalteromonas carrageenonovora ASY5 was inoculated into the fermentation medium, and cultured at 18°C ​​for 0 h, 8 h, 16 h, 24 h, 32 h, 40 h, 48 h, 56 h, After 64 hours, the cell density was measured respectively, the enzyme solution was collected by refrigerated centrifugation, and the enzyme activity was measured by DNS method to determine the optimal culture time of the medium.

[0043] Depend on figure 2 It can be seen that with the increase of culture time, the enzyme activity showed an upward trend. Between 32 h and 40 h, the enzyme activity of ι-carrageenan tended to be stable, reached the highest value at 40 h, and the enzyme activity began to decline after 40 h. Therefore, it was determined that the optimal culture time for the fermentation and enzyme production o...

Embodiment 2

[0066] The ι-carrageenase fermentation optimization process is contrasted with the initial fermentation process, and according to the optimization parameter of embodiment 1, carries out following experiment:

[0067] 1. Preparation of optimized culture medium

[0068] Artificial seawater: including sodium chloride, potassium chloride, calcium chloride dihydrate, magnesium chloride hexahydrate, sodium bicarbonate, magnesium sulfate heptahydrate, the mass fractions (g / 100 mL) are 2.8, 0.08, 0.16, 0.48, 0.01, 0.35.

[0069] Seed medium: 10 g of beef extract, 10 g of tryptone, add 250 mL of distilled water, adjust the pH to 7.8, bathe in water for 10 min, adjust the pH to 7.3, then add 750 mL of artificial sea water, mix well, sterilize at 121 °C for 15 min, Store at 4°C after cooling.

[0070] Fermentation medium: sodium chloride 15 g, yeast powder 15 g, calcium chloride dihydrate 0.2 g, disodium hydrogen phosphate dodecahydrate 3.82 g, sodium dihydrogen phosphate dihydrate 1.3...

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Abstract

The invention discloses a method for preparing t-carrageenase. A seed medium is inoculated with a strain Pseudoalteromonas carrageenovora ASY5 for producing carrageenase, the strain is transferred into a fermentation medium in an inoculation mode for induced enzyme production after activation culture, and t-carrageenase containing fermentation liquor is separated to obtain the t-carrageenase. T-carrageenan is adopted as an inductive agent for fermentation production of the t-carrageenase, a fermentation production technology of the t-carrageenase is established, and the method has good application value for industrial production of carrageenan degrading enzymes and carrageenan oligosaccharide.

Description

technical field [0001] The invention relates to the technical field of biological enzymes, in particular to a preparation method of iota-carrageenase. Background technique [0002] Carrageenan, also known as carrageenan and staghorn alginate, is a polysaccharide of great economic value extracted from certain red algae (such as Eucheuma genus, Cedarium genus, Carrageenan genus, etc.), and is composed of 1,3- A linear sulfated polysaccharide formed by alternating links of β-D-galactopyranose and 1,4-α-D-galactopyranose. There are only three types with extensive economic value: mainly κ-type, ι-type and λ-type, and their disaccharide units contain 1, 2 and 3 sulfate groups respectively. The gel formed by ι-type carrageenan is rich in elasticity, softness, thixotropy, no syneresis, and has high salt resistance properties, which are obviously different from κ-type and λ-type colloids, making it suitable for use in food , cosmetics, medicine and other industries have a wide rang...

Claims

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Application Information

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IPC IPC(8): C12N9/38C12R1/01
CPCC12N9/2468C12Y302/01157
Inventor 肖安风李佳佳肖琼倪辉蔡慧农吴昌正朱艳冰
Owner JIMEI UNIV
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