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Preparation method of monascus metabolite

A technology of metabolites and Monascus bacteria, applied in the biological field, can solve the problems of no specific research on separation methods and biological activities, and slow progress, and achieve the effects of short retention time, easy operation, and improved extraction rate

Active Publication Date: 2015-09-02
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

In the 1930s, since the spread of red yeast rice to Europe and the United States, all countries in the world began to study the classification and identification of red yeast rice, the optimization of the extraction process of red yeast pigment and its application in production, and the fermentation components of red yeast rice. Some studies have been carried out on the separation and purification, structural characterization and metabolic mechanism of the
[0003] Monascus metabolites Monascopyridine A (MCA) and Monascopyridine B (MCB) were developed by German scientists Wild et al. (Wild D, et al. New Monascus metabolites with a pyridine structure in red fermented rice. J Agric Food Chem, 2003, 51, 5493–5496) were obtained by liquid fermentation of red yeast rice or red yeast rice flour as the fermentation medium for the first time, and its structure was characterized. However, no specific research was done on the separation methods and biological activities of MCA and MCB

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1. Comparison of MCA and MCB yields of Monascus aurantiacus AS3.4384 and its pksCT gene-deleted strain PHDS26.

[0022] 1.1 According to the preparation method of the above-mentioned Monascus metabolites MCA and MCB, respectively prepare the mycelia of Monascus aurantium AS3.4384 and its pksCT gene deletion strain PHDS26, and then use the HPLC method to analyze the contents of MCA and MCB in different mycelia samples. Determination.

[0023] 1.2 Chromatographic conditions.

[0024] The chromatographic column Hypersil ODS-2 (5μm, 250×4.6mm) was used as the detection chromatographic column, acetonitrile-water (60:40, v / v) was used as the mobile phase, the column temperature was 25°C, and the flow rate was 0.8mL / min. The detection wavelength is 300nm.

[0025] 1.3 Analysis of results.

[0026] For Monascus aurantula AS3.4384, MCA and MCB were not detected in the mycelia and fermentation broth; MCA and MCB were not detected in the fermentation broth of the gene ...

Embodiment 2

[0027] Example 2. Optimization of conditions for ultrasonic-assisted extraction of MCA and MCB.

[0028] 1.1 A single factor experiment was carried out on the solvent concentration, ultrasonic frequency, extraction time, extraction temperature, solid-liquid ratio, etc., which affect the ultrasonic extraction effect, to determine the range of each influencing factor.

[0029] 1.1.1 The influence of solvents.

[0030] Take 7 parts of 0.50g dry mycelium powder, add methanol, ethyl acetate, ethanol, acetonitrile, acetone, petroleum ether, chloroform 5mL each to seven 10mL volumetric flasks, oscillate evenly, 50℃, ultrasonic extraction for 30min, put on The supernatant was centrifuged at 8000r / min for 20min, filtered through a 0.22μm microporous membrane, and detected according to the HPLC method in Example 1 to study the influence of different extraction solvents on the content of MCA and MCB.

[0031] 1.1.2 The influence of methanol concentration.

[0032] Take 10 parts of 0.50g...

Embodiment 3

[0045] Example 3. Bioactivity assays of MCA and MCB.

[0046] 3.1 Cell Culture

[0047] Resuscitate the cells stored in the liquid nitrogen tank and inoculate them in DMEM medium containing 0.1g / L streptomycin, 13.4g / L DMEM, 100000U / L penicillin, and 15% calf serum (NBCS) and 3% (v / v) glutamine. at 5% CO 2 Incubate at 37°C for 48-72 hours in an incubator.

[0048] When the cells grow to 80% confluence, the old medium is sucked off, and an appropriate amount of trypsin digestion solution is added. When the cells become well-defined and slightly retracted and rounded, the digestive fluid is aspirated, and serum is added to terminate the digestion. Then add an appropriate amount of complete DMEM culture medium, pipette into a single cell suspension, and adjust the cell concentration to 1×10 4 cells / mL, inoculated into a new culture bottle, incubator at 37°C, 5% CO 2 to cultivate.

[0049] 3.2 MTT method to determine the inhibitory effect of MCA and MCB on the proliferatio...

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PUM

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Abstract

The invention discloses a preparation method of a monascus metabolite. Mycelium is obtained through liquid state fermentation of monascus PHDS26, and then extraction, separation and purification are performed on the mycelium to obtain Monascopyridine A and Monascopyridine B. With the adoption of the preparation method, the yields of MCA and MCB can reach 2.07 and 1.96 g / kg respectively, and the extraction rate is increased by about 50% compared with that of the prior art. For the HPLC analytical method adopted by the invention, gradient elution is not required, phosphoric acid is not required to be added during the sample extraction process, the operation is simpler and more convenient, and the retention time of the MCA and MCB is short.

Description

technical field [0001] The invention belongs to the field of biotechnology. Involves the preparation of natural active products. Background technique [0002] Red yeast rice is a traditional fermented product in my country. It has always been considered to have dual functions of medicine and food in China. It is not only a precious scientific and cultural heritage of the motherland, but also has great significance in the history of microorganisms in the world. In the 1930s, since the spread of red yeast rice to Europe and the United States, all countries in the world began to study the classification and identification of red yeast rice, the optimization of the extraction process of red yeast pigment and its application in production, and the fermentation components of red yeast rice. Some studies have been carried out on the separation and purification, structural characterization and metabolic mechanism of phospholipids, but the progress is slow. In recent decades, with ...

Claims

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Application Information

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IPC IPC(8): C12P17/18C07D491/048C12R1/645
Inventor 黄志兵许杨苏保伟李燕萍涂追
Owner NANCHANG UNIV
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