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Application and method of manganese peroxidase for degrading citrinin

A technology of manganese peroxidase and citrinin, which is applied in the application field of degrading citrinin, can solve the problems that there are no reports on the degradation of other mycotoxins by manganese peroxidase, and achieve low cost and wide application range Effect

Active Publication Date: 2021-12-24
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Degradation of aflatoxin by manganese peroxidase has been reported, but no other mycotoxins have been reported by manganese peroxidase

Method used

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  • Application and method of manganese peroxidase for degrading citrinin
  • Application and method of manganese peroxidase for degrading citrinin
  • Application and method of manganese peroxidase for degrading citrinin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Embodiment 1 Preparation of recombinant manganese peroxidase MrMnP

[0019] The MrMnP gene sequence from Moniliophthora roreri was fully synthesized from Jinweizhi Company. The X33 / MrMnP Pichia pastoris engineering strain containing the recombinant plasmid was inoculated in 50 mL of YPD medium, shaken at 30 °C and 220 rpm for 48 h, and then transferred to 300 mL of BMGY medium at a ratio of 2%, at 30 °C. , 220 rpm shaking culture for 48 h, the BMGY yeast culture solution was centrifuged at 5,500 rpm for 5 min, and the supernatant was discarded. Add 200 mL of BMMY medium (with the addition of heme at a final concentration of 100 µM) to the fermentation flask. 30°C, 200 rpm shaker culture for 48 hours, 5,500 rpm centrifugation for 5 minutes, the fermentation broth was collected to obtain the recombinant manganese peroxidase MrMnP.

Embodiment 2

[0020] Example 2 Degradation of citrinin by manganese peroxidase

[0021] Dissolve citrinin in methanol to prepare a 5 g / L stock solution, and follow the following reaction system: 250 μL malonic acid buffer (0.2 M, pH 5.0), 100 μL citrinin solution (100 mg / L), 250 μL manganese sulfate (40 mM), 250 μL manganese peroxidase (5000 U / L) prepared in Example 1, 200 μL hydrogen peroxide (5 mM). The system without adding manganese peroxidase was used as a control, and the reaction system was repeated three times. The reaction was carried out at 30°C. After 96 h, three times the volume of methanol was added to terminate the reaction. The degradation rate of citrinin was analyzed by high performance liquid chromatography (HPLC). The liquid chromatography is Shimadzu Nexera UHPLC high-performance liquid chromatography analysis system, the chromatographic separation column is Zorbax SB-C18 (4.6×250 mm, 5 μm), mobile phase A (water with 0.1% acetic acid), mobile phase B (acetonitrile) ; ...

Embodiment 3

[0022] Example 3 Degradation of Citrinin in Monascus Red Pigment by Manganese Peroxidase

[0023] Dissolve citrinin in methanol to prepare a 5g / L stock solution, and use the following reaction system: 250 μL malonate buffer (0.2 M, pH 5.0), 25 μL citrinin solution (400 mg / L), 250 μL μL manganese sulfate (40 mM, apple juice as solvent), 125 μL manganese peroxidase (10000 U / L) prepared in Example 1, 100 μL hydrogen peroxide (10 mM), 250 μL 8 mg / mL Monascus red pigment. The system without adding manganese peroxidase was used as a control, and the reaction system was repeated three times. The reaction was carried out at 30°C. After 96 h, three times the volume of methanol was added to terminate the reaction. The degradation rate of citrinin was analyzed by high performance liquid chromatography (HPLC). The liquid chromatography is Shimadzu Nexera UHPLC high-performance liquid chromatography analysis system, the chromatographic separation column is Zorbax SB-C18 (4.6×250 mm, 5 μm)...

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Abstract

The invention relates to the field of biotechnology, and relates to the application and method of manganese peroxidase for degrading citrinin. The present invention provides the application of manganese peroxidase with the amino acid sequence shown in SEQ ID NO:1 for degrading citrinin. The manganese peroxidase derived from Moniliophthora roreri efficiently degrades the mycotoxin citrinin. The method has low cost and wide application range, and can be widely used in the field of food toxin degrading enzymes.

Description

technical field [0001] The invention relates to the field of biotechnology, and relates to the application and method of manganese peroxidase for degrading citrinin. Background technique [0002] Mycotoxins are secondary metabolites produced by fungi, which mainly contaminate stored grain, oil, food and feed, and seriously endanger the health of humans and animals. Citrinin is a nephrotoxic mycotoxin that has significant nephrotoxicity to mammalian kidneys. [0003] Monascus red pigment is a secondary metabolite produced by Monascus during the growth and metabolism process. As an edible natural pigment, compared with chemically synthesized pigments, Monascus red pigment has natural sources, reliable safety performance, no toxic side effects, Nutritious, bright colors and other advantages. However, during the secondary metabolism of Monascus, citrinin is always produced along with the production of Monascus red pigment. [0004] Traditional methods of controlling mycotoxin...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/08A23L5/20
CPCC12N9/0065A23L5/25C12Y111/01013
Inventor 苏小运王帅张伟姚斌王晓璐秦星徐欣欣王苑张红莲罗会颖
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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