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Secondary metabolites of ache-inhibiting secondary metabolites of Portunus trituberculatus endogenous Cladomycetes pt-2 and preparation methods thereof

A kind of technology of secondary metabolite, budding branch mold, applied in biological field

Inactive Publication Date: 2016-01-06
QINGDAO AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are relatively few reports on endophytic fungi from marine sources against Alzheimer's disease, and endophytic fungi from Portunus trituberculatus have not been reported

Method used

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  • Secondary metabolites of ache-inhibiting secondary metabolites of Portunus trituberculatus endogenous Cladomycetes pt-2 and preparation methods thereof
  • Secondary metabolites of ache-inhibiting secondary metabolites of Portunus trituberculatus endogenous Cladomycetes pt-2 and preparation methods thereof
  • Secondary metabolites of ache-inhibiting secondary metabolites of Portunus trituberculatus endogenous Cladomycetes pt-2 and preparation methods thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] 1. Isolation of strains

[0046] 1.1. Medium

[0047] Medium for separation: Potato agar (PDA) medium, adding penicillin potassium and 150U / ml streptomycin sulfate at a final concentration of 200U / mL.

[0048] Liquid fermentation medium (improved Chase medium): 30g sucrose, 3gNaNO 3 , 0.5gKCL, 0.5gMgSO 4 ·7H 2 O, 0.01g FeSO 4 , 1gK 2 HPO 4 Add 1000mL seawater in turn, stir well to dissolve, then add 1g of yeast extract to slightly heat to dissolve, and sterilize at 121°C for 20min for later use.

[0049] 1.2. Strain isolation method

[0050] Remove the shell of the crab, take its gills and intestines and stomach, soak it in 75% alcohol for 5 minutes for surface disinfection, then cut it into pieces and add 3mL sterile water to grind it, spread it on the separation medium, place the plate in a constant temperature incubator, Inverted culture at 28°C. When mycelium grows on the plate, the endophytic fungus is isolated and purified by cutting the mycelium tip, and...

Embodiment 2

[0092] The preparation process of the secondary metabolites of AchE inhibited by endogenous Cladomycetes trituberculatus Pt-2 is as follows:

[0093] (1), with endophytic branch mold Pt-2 of portunus trituberculatus as bacterial classification, adopt the method for liquid fermentation, under the conditions of 10% inoculum size, 200mL / 500mL Erlenmeyer flask, 28 ℃ of conditions in the fermentation medium Fermented in medium for 9 days to obtain the fermented product; the formula of the fermentation medium is: 30g glucose, 3gNaNO 3 , 0.5gKCl, 0.5gMgSO 4 ·7H 2 O, 0.01g FeSO 4 , 1gK 2 HPO 4 , 1g yeast extract, 1000mL sea water;

[0094] (2), concentrating the fermentation broth in vacuum at 40°C to 1 / 3 of the original volume, then extracting with ethyl acetate, and concentrating the extract to dryness;

[0095] (3) The organic phase extract of the fermented product of Cladomycetes condensed to dryness was subjected to silica gel column chromatography and chloroform / methanol s...

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Abstract

The invention discloses an AchE-inhibiting secondary metabolite and a preparation method of endophytic Cladomycetes trituberculatus Pt-2, belonging to the field of biotechnology. The secondary metabolites are prepared by the following method: (1), using the endophytic Cladomycetes Pt-2 of Portunus trituberculatus as the bacterial species, carrying out liquid fermentation to obtain a fermented product; (2), making the fermented liquid Concentrate in vacuum at 40°C to 1 / 3 of the original volume, then extract with ethyl acetate, and concentrate the extract to dryness; (3), the organic phase extract of the fermented product of Cladomycetes Pt-2 concentrated to dryness , using silica gel column chromatography, chloroform / methanol system gradient elution, to obtain 6 polar segments, the most active segment was separated by C18 reverse-phase column and Sephadex gel chromatography, and the inhibition rate of AchE was 70-90 % secondary metabolites. The invention has the advantages of overcoming the disadvantages of high price, side effects and difficult chemical synthesis of current medicines, providing a new medicine and a preparation method, and providing a theoretical basis for the development of biological medicines for preventing senile dementia.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a secondary metabolite of AchE inhibited by Endophytic Cladomycetes Pt-2 in Portunus trituberculatus and a preparation method thereof. Background technique [0002] Alzheimer's disease (AD) is a common senile brain neurodegenerative disease with a relatively high incidence, and has become one of the diseases that seriously threaten the lives of the elderly in modern society. Its death rate ranks fourth after cardiovascular disease, cancer and stroke. The pathogenesis of AD is complex and is a disease of multiple etiologies. Typical pathological changes include β-amyloid protein (β-amyloidprotein, Aβ) deposition to form senile plaques (senile plaques, SP), neurofibrillary tangles (neurofibrillary tangles, NFT), basal Cholinergic dysfunction in the forebrain and widespread neuronal loss and altered synapse morphology in the cortex and hippocampus. In recent years,...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P1/02C12N1/14C12R1/645
Inventor 王凤舞朴美子
Owner QINGDAO AGRI UNIV