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Solid fermentation method of bilirubin oxidase

A bilirubin oxidase and solid fermentation technology, applied in the field of bilirubin oxidase solid fermentation, can solve the problems of long stirring and aeration culture time, large sewage discharge, large steam consumption, etc., and achieve low production cost, The production process is energy-saving and environmentally friendly, and the effect of abundant sources

Inactive Publication Date: 2011-02-16
XINXIANG MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method consumes a lot of steam, takes a long time for stirring and aeration, has high energy consumption, and has a large amount of sewage discharge after fermentation.
So far, there is no report about the solid fermentation method of bilirubin oxidase

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0007] Embodiment 1: Bran passes through 40 mesh sieves after purchasing from the market. Weigh 3g of bran and place it in a 250ml Erlenmeyer flask, then add 13ml of distilled water, stir the bran and distilled water evenly with a glass rod, sterilize under high pressure at 121°C for 20 minutes, after cooling, add 1ml of it to a concentration of 1×10 8 ~10×10 8 Individual / ml spore suspension, cultured statically at 22°C for 11 days. Take out the Erlenmeyer flask after cultivating for 11 days, add 50ml of distilled water into the Erlenmeyer flask, shake and extract at 180r / min for 2.5 hours, filter with qualitative filter paper to obtain the crude enzyme liquid of bilirubin oxidase, and its enzyme activity can reach 25.4U / g .

Embodiment 2

[0008] Embodiment 2: Bran passes through 40 mesh sieves after purchasing from the market. Weigh 7g of bran and place it in a 250ml Erlenmeyer flask, then add 17ml of distilled water, stir the bran and distilled water evenly with a glass rod, autoclave at 125°C for 25 minutes, and after cooling, add 5ml of it to a concentration of 1×10 8 ~10×10 8 Individual / ml spore suspension, cultured statically at 31°C for 7 days. Take out the Erlenmeyer flask after culturing for 7 days, add 65ml of distilled water into the Erlenmeyer flask, oscillate at 180r / min for 3.5 hours, filter with qualitative filter paper to obtain the crude enzyme liquid of bilirubin oxidase, and its enzyme activity can reach 26.2U / g .

Embodiment 3

[0009] Embodiment 3: Bran passes through 40 mesh sieves after purchasing from the market. Weigh 5g of bran and place it in a 250ml Erlenmeyer flask, then add 15ml of distilled water, stir the bran and distilled water evenly with a glass rod, sterilize under high pressure at 130°C for 30 minutes, after cooling, add 1.5ml of it to a concentration of 1×10 8 ~10×10 8 Individual / ml spore suspension, cultured at 26°C for 9 days. Take out the Erlenmeyer flask after cultivating for 9 days, add 60ml of distilled water into the Erlenmeyer flask, shake and extract for 3.5 hours at 180r / min, filter with qualitative filter paper to obtain the crude enzyme liquid of bilirubin oxidase, and its enzyme activity can reach 27.6U / g .

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Abstract

The invention discloses a solid fermentation method of bilirubin oxidase, which has the purposes of low cost, simple production process and convenient enzyme extraction. The solid fermentation method comprises the following steps of: (1) separating and screening myrothecium roridum from soil, and preparing a myrothecium roridum spore suspension; and (2) weighing 3-7 parts by weight of bran, placing the bran in a triangular flask, adding 13-17 parts by weight of distilled water, uniformly agitating the bran and the distilled water by using a glass tick, sterilizing under high pressure at 121-130 DEG C for 20-30 min, cooling, inoculating with 1-5 parts by weight of spore suspension prepared in the step (1), cultivating at 22-31 DEG C for 7-11 days, taking out the triangular flask, adding 50-70 parts by weight of distilled water into the triangular flask, leaching for 2-4 h through table concentrator oscillation, and filtering by using qualitative filter paper to obtain the bilirubin oxidase crude liquid. The invention has the advantages of abundant resources and easy acquisition of the raw materials, low production cost, simple, convenient and feasible enzyme extraction process and energy-saving and environmental-friendly production process.

Description

Technical field: [0001] The invention relates to the fermentation of bilirubin oxidase, in particular to a solid fermentation method of bilirubin oxidase. Background technique: [0002] Bilirubin oxidase (Bilirubin oxidase, EC 1.3.3.5) is a copper-containing polyphenol oxidase that can catalyze the oxidation of bilirubin, and oxygen is its electron acceptor. The reaction process is that bilirubin oxidase catalyzes bilirubin to generate biliverdin, which can be further oxidized to finally generate a colorless compound whose properties are still unclear. Utilizing this reaction, the decrease of the absorbance value at the wavelength of 440nm or 450nm can be measured, which can be used for the determination of serum bilirubin. Since the discovery by Japanese scholars in 1981 that Myrotheca genus can produce high-yielding bilirubin oxidase, this enzyme has been used in clinical medicine to measure bilirubin. This method has the advantages of specificity, high sensitivit...

Claims

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Application Information

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IPC IPC(8): C12N9/02C12R1/645
Inventor 刘友勋黄娟石如玲张煜赵春澎
Owner XINXIANG MEDICAL UNIV
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