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Extraction and purification method of ergothioneine

A technology of ergothioneine and purification method, which is applied in the fields of biochemical products, food and medicine, and achieves the effects of few steps, simple operation and convenient operation

Active Publication Date: 2015-07-15
TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the prior art, there is no report of adopting medium pressure preparation chromatography to purify ergothioneine

Method used

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  • Extraction and purification method of ergothioneine
  • Extraction and purification method of ergothioneine
  • Extraction and purification method of ergothioneine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: Preparation of Pleurotus otreatus CGMCC No.6232 mycelium fermentation broth

[0042] Slant medium: PDA medium (Becton, Dickinson and Company).

[0043] Liquid seed medium: corn flour 30g / L, soybean meal 15g / L, α-amylase 54U / L, KH 2 PO 4 3g / L, MgSO 4 ·7H 2 O1.5g / L, the rest is water, sterilized at 121°C for 20 minutes, and the liquid volume in a 500mL Erlenmeyer flask is 150mL.

[0044] Liquid fermentation medium: glycerol 50g / L, casein peptone 35g / L, KH 2 PO 4 3g / L, MgSO 4 ·7H 2 O1.5g / L, the rest is water, sterilized at 121°C for 20 minutes, and the liquid volume in a 500mL Erlenmeyer flask is 150mL.

[0045] The bacterial lawn of strain CGMCC No.6232 from the slant medium was picked and inserted into the seed medium, and cultured on a shaker at 25° C. at 150 rpm for 4 days to obtain a seed solution. The seed liquid was inserted into the fermentation medium with an inoculation amount of 5% by volume, and cultured on a shaker at 25° C. at 150 rpm f...

Embodiment 2

[0046] Embodiment 2: Extraction and purification of ergothioneine

[0047] The mycelium fermented liquid of Pleurotus otreatus CGMCC No.6232 is centrifuged, and 50 g of mycelium is collected, and water of 1:40 is added according to wet mycelium weight: water (g: mL) to make mycelium aqueous suspension. The mycelia suspension was placed in a water bath at 100°C, and leached for 5 minutes without stirring, and the ergothioneine was leached into the aqueous solution. Collect mycelia and filtrate respectively after filtration to obtain 1700 mL of ergothioneine aqueous solution.

[0048] The above-mentioned aqueous solution of ergothioneine is ultrafiltered by a hollow fiber ultrafiltration membrane with a pore size of 4kDa to obtain 1000mL of permeate, and the purity of ergothioneine measured by high pressure liquid phase reaches 31.6%. The permeated liquid was rotary evaporated at 50° C. to a concentrated solution with an ergothioneine concentration of 1.72 g / L or spray-dried t...

Embodiment 3

[0052] Embodiment 3: Extraction and purification of ergothioneine

[0053] Pleurotus otreatus (Pleurotus otreatus) CGMCC No.6232 mycelia after submerged fermentation, the mycelium fermentation liquid is centrifuged, 600g mycelium is collected, and water of 1:0.5 is added according to wet mycelium weight:water (g:mL) , to make an aqueous suspension of mycelium. The mycelia aqueous suspension is placed in a 70°C water bath, stirred and leached at 600rpm for 60min, and the ergothioneine is leached into the aqueous solution. Mycelia and filtrate were collected respectively after filtration, and mycelium was repeatedly leached 5 times by the above-mentioned method, and the filtrate after merging and leaching was obtained to obtain 1100mL ergothioneine aqueous solution.

[0054] The above-mentioned aqueous solution of ergothioneine is ultrafiltered by a hollow fiber ultrafiltration membrane with a pore size of 6kDa to obtain 600mL of permeate, and the purity of ergothioneine measur...

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Abstract

A method for extracting and purifying L-ergothioneine comprises the following steps: (1) soaking and extracting a mushroom mycelium fermentation liquid by using hot water, so as to obtain a mushroom and L-ergothioneine water solution; (2) performing ultrafiltration of the mushroom and L-ergothioneine water solution by using an ultrafiltration membrane with a molecular weight cut-off of 1kDa to 30kDa, and collecting an ultrafiltration liquid; and (3) performing chromatographic purification of the ultrafiltration permeation liquid, the chromatographic purification conditions being: a chromatographic filler being an HILIC filler, a mobile phase forming acetonitrile-water with a volume ratio of 80: 20 to 88: 12, and a quantity of feed of L-ergothioneine being 0.01g to 0.323 per 100g.

Description

technical field [0001] The invention belongs to the technical fields of biochemical products, food and medicine, and relates to a method for separating and refining ergothioneine, in particular to a method for extracting and purifying ergothioneine from mycelia. Background technique [0002] Ergothioneine (L-Ergothioneine, EGT) is a rare natural chiral amino acid, which is an important physiologically active substance in organisms. It has the functions of scavenging free radicals, detoxification, maintaining DNA biosynthesis, normal cell growth, cellular immunity, Anti-radiation, whitening and anti-aging functions are identified as a unique, multi-functional cell physiological protection agent, especially in anti-oxidation and energy regulation, so ergothioneine is widely used in food, cosmetics and Biomedicine and other industries have broad application prospects. [0003] Marvin S.Hausman MD, CEO of the Global Nutrition Society, pointed out: Since ergothioneine can inhibi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D233/84C12P13/04C12R1/645
CPCC07D233/84
Inventor 姜文侠刘琦王洪宇李运华刘伟梅保良张维亚
Owner TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
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