Method for simultaneously extracting hypocrellin A and Elsinochrome A from solid fermented material of Shiraia bambusicola

A technology for sclerostin and rhodopsin A, applied in the field of bioengineering, can solve the problems of no clear extraction rate, difficult to obtain raw materials, complicated elution steps, etc., and achieves large-scale production, high environmental protection index, Simple process effect

Active Publication Date: 2018-10-12
NANJING NORMAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] For Hypocretin A crystals, Yin Zhiqi and others first used highly polar ethanol in 15kg of Zhuhuang decoction pieces, and then extracted with ethyl acetate to obtain 65mg Hypocretin A crystals. The purity is not disclosed, but the inventor of this patent The experiment found that only using ethanol reflux to extract the extract, the extraction rate is low, and the elution steps are complicated by using petroleum ether, acetone, ethyl acetate ratio silica gel column, and the extraction rate is also low; a kind of invention invented by Yan Jiading et al. The preparation method of high-purity hypocrellin A uses natural bamboo yellow Chinese herbal medicines and does not disclose the crystallization purity of hypocrellin A and only uses ethanol solvent. There is no clear extraction rate and no separation using the polarity of organic solvents.
For the first time, Zheng Lixiong and others used 4.4kg bamboo red fungus crude powder to obtain about 74mg of coelomycin A crystals with acetone, ethanol, cyclohexane, ethyl acetate organic solvents and silica gel chromatography column, the purity of which was not determined. It is low and uses more solvents and is more complicated. It is extracted from the coarse powder of bamboo red fungus medicinal materials, and the raw materials are not easy to obtain, and only one kind of substance crystallization is obtained.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Acquisition of bamboo yellow solid-state fermentation material:

[0029] 1. Source of the strain: bamboo yellow fungus comes from the China Forestry Microorganism Collection and Management Center, Shiraiasp.cfcc84681. The bacterial strain is preserved on a PDA slant, which is a common medium for fungi, and the bacterial strain is activated when used.

[0030] 2. Preparation of seed inoculum: adopt medium formula: glucose 1%, sodium nitrate 0.2%, potassium dihydrogen phosphate 0.05%, magnesium sulfate 0.03%. Configure 10 bottles, each bottle 100mL, sterilized. Insert one block of activated strain into each bottle, and shake the flask at 28°C for 3 days of fermentation.

[0031] 3. Preparation of solid fermentation medium: Weigh 2000g of rice, soak it in water overnight, drain it and put it into a sterilization bag for high temperature and high pressure sterilization. Preparation of solid fermentation feed medium: glucose 2%, sodium nitrate 0.4%, potassium dihydrogen p...

Embodiment 2

[0035] 1. Weigh and extract:

[0036] The solid-state fermentation material finally obtained in Example 1 was crushed, weighed 1500g, firstly added 2000mL of petroleum ether, soaked for 1.5h, suction filtered to obtain petroleum ether extract, and then rotary evaporated to obtain petroleum ether extract, repeated 5 times to obtain Petroleum ether combined extract 3.2g. Then add 2000mL ethyl acetate to the solid-state fermentation material after suction filtration, after soaking for 1.5h, suction filtration to obtain ethyl acetate extract, and then rotary evaporation to obtain ethyl acetate extract, repeat 5 times to obtain ethyl acetate combined extract Cream 5.2g. Finally, 2000 mL of ethanol was added to the solid-state fermentation material after the previous suction filtration, soaked for 1.5 hours, and the ethanol extract was obtained by suction filtration, and then rotary evaporation was obtained to obtain the ethanol extract. Repeat 5 times to obtain 2.3 g of ethanol co...

Embodiment 3

[0047] Concentrated crystals of Hypocretin A and Escariacin A:

[0048] The chromatographic solution collected in steps 2-4 in Example 2 was detected by HPLC: mobile phase: aqueous phase (acetonitrile: water)=70:30, flow rate 0.8ml / min, column temperature 30°C, detection wavelength 254nm wavelength detection, The loading volume was 10 μL, and the eluate was detected. The 100% pure standard substance of hypocrellin and escharomycin was used as a control to determine the contents of the two. Combine the same substances with a content of more than 65% in the chromatographic liquid detected by HPLC, and spin-dry using a rotary evaporator to obtain a chromatographic liquid extract. The obtained extract uses ethyl acetate to separate Hypocretin A and The scabactin extract was fully dissolved, left to evaporate at room temperature, and when the ethyl acetate was quickly evaporated to dryness, washed with petroleum ether for 5 times to obtain crystals.

[0049] 0.32 g of hypocrellin...

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PUM

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Abstract

The invention discloses a method for simultaneously extracting hypocrellin A and Elsinochrome A from a solid fermented material of Shiraia bambusicola. The method comprises the following steps: (1) leaching the solid fermented material of Shiraia bambusicola separately by using organic solvents with different polarities from weak to strong according to the polarity; (2) concentrating extract intoextract paste, and performing sequential elution in a chromatography column by using organic solutions with gradient ratios separately so as to elute the two substances of hypocrellin A and Elsinochrome A from the same column in batches; and (3) performing identification, concentration and crystallization on the obtained chromatographic liquid so as to obtain the hypocrellin A and Elsinochrome A.By leaching the solid fermented material of Shiraia bambusicola through an organic solvent gradient method, simultaneous acquirement of the hypocrellin A and Elsinochrome A is achieved in the chromatography process of one chromatography column, meanwhile, a high extraction rate and high purity are achieved, and the organic solvents can be recycled without waste discharge; the extraction method isbeneficial to industrial production of hypocrellin A and Elsinochrome A.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a method for efficiently extracting hypocretin A and coelomycin A in bamboo yellow fungus fermentation materials. Background of the invention [0002] Bamboo yellow is a traditional medicinal fungus in my country. It has the functions of relieving cough and pain, relaxing tendons and activating collaterals, dispelling wind and dampness, nourishing the middle and replenishing qi, promoting blood circulation, dissipating blood stasis and stimulating menstruation. The content of hypocrellin and hypocretin in the secondary metabolites of bamboo yellow is relatively high. Coelomycins include coelomycin A, coelomycin B, coelomycin C, and coelomycin D, all of which belong to perylenequinone compounds. As good photosensitizers, perylenequinones have different degrees of anticancer activity and antiviral activity (Hudson J B, Imperial V, Haugland R P, et al. Photochemi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07C46/10C07C50/38
CPCC07C46/10C07C2603/54C07C50/38
Inventor 陈双林李强闫淑珍
Owner NANJING NORMAL UNIVERSITY
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