Application of natural acid glycolipid molecules in neutralizing clostridium perfringens Epsilon toxin

A Clostridium perfringens, acidic technology, applied in the field of biomedicine, to achieve good neutralization ability, good neutralization effect, and prolong survival rate

Active Publication Date: 2019-11-19
KUNMING INST OF ZOOLOGY CHINESE ACAD OF SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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  • Application of natural acid glycolipid molecules in neutralizing clostridium perfringens Epsilon toxin
  • Application of natural acid glycolipid molecules in neutralizing clostridium perfringens Epsilon toxin
  • Application of natural acid glycolipid molecules in neutralizing clostridium perfringens Epsilon toxin

Examples

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Embodiment 1

[0029] Example 1: Isolation, purification and identification of Clostridium perfringens (NCTC 8346) Epsilon toxin

[0030] 1. Culture of Clostridium perfringens and collection of supernatant

[0031] Clostridium perfringens (NCTC 8346) was purchased from the British Culture Collection of Microorganisms. First, a few colonies were taken with a sterilized inoculation loop and inoculated into 5ml of sterilized thioglycollate broth (1L formula needs to add Tryptone 15g, yeast extract 5g, L-cystine 0.5g, sodium thioglycolate 0.5g, sodium chloride 2.5g, glucose 5.5g, pH7.1), placed in a shaker at 37°C overnight; Inoculate 5ml of the initial culture into 100ml of TGY medium (containing 3% soybean trypsin broth, 2% glucose, 1% yeast extract, 0.1% sodium thioglycolate), and culture it anaerobically at 37°C for 16-18h , inoculate it into 3.5L of freshly prepared TGY medium, and continue culturing at 37°C for 12h. Centrifuge at 8000rpm / min for 20min to collect the culture supernatant. ...

Embodiment 2

[0041] Example 2: Screening of three types of acidic glycolipid molecules with neutralizing ability to Epsilon toxin toxicity and verification of neutralizing ability

[0042] 1. Enzyme cleavage and activation of Epsilon protoxin

[0043] The prerequisite for the Epsilon toxin to play a toxic role is that the Epsilon protoxin should be cleaved by trypsin to become an active Epsilon toxin. Therefore, before studying the neutralizing activity of acidic glycolipid molecules on the toxicity of Epsilon toxin, trypsin digestion of Epsilon protoxin was carried out first. Specifically, 1 μg of Epsilon protoxin corresponds to 12.5 μg of trypsin. After mixing, place it at 37°C for 1 hour , followed by adding a trypsin inhibitor to stop the activity of trypsin. Therefore, in subsequent activity experiments, the Epsilon toxin / trypsin / trypsin inhibitor mixture was used to represent the Epsilon toxin, and the trypsin / trypsin inhibitor mixture was used as a control.

[0044] 2. Screening o...

Embodiment 3

[0052] Embodiment 3: Ganglioside, sulfatide and sialic acid are to the protective effect of mice after Epsilon toxin infection

[0053] As mentioned above, gangliosides, sulfatides, and sialic acid can all effectively inhibit and neutralize the toxicity of Epsilon toxin in vitro experiments. It is speculated that these three types of acidic glycolipid molecules are expected to be good antidotes for bacterial lethal toxins . In order to verify this speculation, we intend to conduct research on the protective effect of these three types of acidic glycolipid molecules on animals infected with Epsilon toxin at the level of animals. The specific experimental design is as follows:

[0054]For the experimental animals, 48 ​​C57BL / 6J adult male mice with a body weight of 20±2g were randomly divided into 6 groups with 8 mice in each group. Water feeding; (2) Epsilon toxin treatment group, mice in this group are given Epsilon toxin enema stimulation every day according to the dose of 5...

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Abstract

The invention relates to an application of natural acid glycolipid molecules in neutralizing clostridium perfringens Epsilon toxin, in particular, relates to the application in preparation of relateddrugs or reagents for prevention or treatment of clostridium perfringens infection in animals, and belongs to the biomedical field. The invention discloses one or more of natural acid glycolipid molecules sialic acid, ganglioside and sulfatide, wherein the natural acid glycolipid molecules have a good neutralization effect of Epsilon toxin, and have a good therapeutic effect on either animal-lethal clostridium perfringens infection or invasion and infection of Epsilon toxin. Three kinds of natural acid molecules sialic acid, ganglioside and glucosinolate have the advantages of simple molecularstructure, economic price, easily availability on the market and the like. The natural acid glycolipid molecules are expected to be used as substitute antibiotics or antibodies for prevention and treatment of clostridium perfringens infection, and have good application prospects in animal husbandry and veterinary.

Description

Technical field: [0001] The present invention relates to the application of natural acidic glycolipid molecules in neutralizing Clostridium perfringens Epsilon toxin, especially its application in the preparation of drugs or reagents related to the prevention or treatment of Clostridium perfringens infection in animals, which belongs to biomedical field. Background technique: [0002] Bacterial infectious diseases have always been the number one enemy that seriously threatens human or other animal life. Clostridium perfringens (Clostridium perfringens) is the main pathogenic bacterium that causes fatal gas gangrene and enterotoxemia in humans and animals, so it has caused great harm to human health and animal husbandry. Especially in animal husbandry, the animal clostridial disease caused by Clostridium perfringens that produces Epsilon toxin is characterized by "acute onset, fast course of disease, and high mortality", which has brought great troubles to the majority of he...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/7032A61P31/04
CPCA61K31/7032A61P31/04Y02A50/30
Inventor 郭小龙辛文文张云王景林李文辉
Owner KUNMING INST OF ZOOLOGY CHINESE ACAD OF SCI
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