Detection method of L-erythrothioneine

A technology for ergothioneine and a detection method, applied in the field of chemical analysis, can solve problems such as being difficult to achieve effective separation of a target peak and an impurity peak, affecting the accurate quantitative detection of ergothioneine, and many pretreatment procedures, and achieving low cost and improved use. Longevity and cost reduction effect

Active Publication Date: 2014-04-23
TIANJIN SINONOCY BIOLOGICAL TECH CO LTD
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Problems solved by technology

[0008] Ergothioneine was prepared by mushroom liquid submerged fermentation. Due to the complexity of the fermentation products, it was difficult to effectively separate the target peak from the impurity peak by using the HPLC detection conditions and methods reported at present; there were many pretreatment procedures for samples, which affected the ergothioneine. Accurate quantitative detection; more organic solvents are used and the cost is high

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  • Detection method of L-erythrothioneine
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  • Detection method of L-erythrothioneine

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Embodiment 1

[0040] The pretreatment method of the mycelium: the fermentation broth is first filtered by suction to collect the mycelium, and then add the same volume of pure water as the fermentation broth to suspend to make a mycelium suspension. The mycelium suspension was leached once by a 600rpm magnetic stirrer, the leaching temperature was 90°C, and the leaching time was 60min. After leaching, centrifuge for 10 minutes by ultrafiltration, the pore size of the ultrafiltration membrane is 3kDa, and the centrifugation condition is 12840×g. The permeate is the ergothioneine sample to be tested.

Embodiment 2

[0042] The pretreatment method of the mycelium: the fermentation broth is first filtered by suction to collect the mycelium, and then add the same volume of pure water as the fermentation broth to suspend to make a mycelium suspension. The mycelium suspension is directly extracted without stirring, the extraction temperature is 100° C., and the extraction time is 10 minutes. After leaching, centrifuge for 10 minutes by ultrafiltration, the pore size of the ultrafiltration membrane is 3kDa, and the centrifugation condition is 12840×g. The permeate is the ergothioneine sample to be tested.

Embodiment 3

[0044] The pretreatment method of the mycelium: the fermentation broth is first filtered by suction, the mycelium is collected, and one-third of the volume of the fermentation broth is added into pure water to suspend to make a mycelium suspension. The mycelium suspension was leached in a water bath, the leaching temperature was 50°C, the leaching time was 30min, and the stirring speed was 500rpm. After leaching, the solid-liquid separation is realized through suction filtration, and the obtained mycelium is subjected to suction filtration again, and the method is the same as above, and the leaching process is repeated three times in this way, and the total extraction time is 90min, and the three extractions are mixed uniformly to obtain A mixed extract with the same volume as the fermentation broth. After leaching and mixing, ultrafiltration and centrifugation for 10 minutes, the pore size of the ultrafiltration membrane is 3kDa, and the centrifugation condition is 12840×g. ...

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Abstract

The invention relates to an HPLC detection method of L-erythrothioneine in mycelium or fermentation broth. The method comprises a pretreating step of the mycelium or the fermentation broth and an HPLC detection step. The pretreating step of the mycelium includes performing solid-liquid separation of the fermentation broth, collecting the mycelium, adding pure water for dissolution, leaching to obtain leaching liquor, and performing ultrafiltration by an ultrafiltration membrane to obtain permeate of a sample containing the L-erythrothioneine. The pretreating step of the fermentation broth includes leaching the fermentation broth directly, and performing ultrafiltration by an ultrafiltration membrane to obtain permeate of a sample containing the L-erythrothioneine. According to the method, simple pretreatment is performed before detection by utilization of the HPLC, thus removing most impurities in the fermentation broth and the mycelium, prolonging the service lifetime of chromatographic columns, increasing the separation degree of the sample, making detection results more accurate and more stable and reducing the detection cost.

Description

technical field [0001] The invention relates to the field of chemical analysis, in particular to a method for detecting ergothioneine in fermentation broth and mycelium. Background technique [0002] Ergothioneine (L-Ergothioneine, EGT), chemical name 2-mercaptohistidine trimethyl salt, is a rare natural chiral amino acid, it is the only known natural 2-thio Imidazole amino acid (2-thio-imidazole), the chemical structure formula is as follows: [0003] [0004] Ergothioneine is an important natural active substance in the body, which has strong antioxidant activity: scavenging active oxides and active nitrogen compounds, activating antioxidant enzymes, inhibiting superoxide dismutase (Aruoma OI, Whiteman M, England TG, Halliwell B.Antioxidantaction of ergothioneine assessment of its ability to scavenge peroxynitrite.BBRC,1997,231:389~391.), inhibit the oxidation reaction of various heme proteins (Romaro R, et al., The reactivity of thiols and disulfides with different r...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
Inventor 姜文侠刘琦周涛
Owner TIANJIN SINONOCY BIOLOGICAL TECH CO LTD
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