Preparation method of cyclic dipeptide compound

A technology of compound and cyclic dipeptide, which is applied in the field of preparation of cyclic dipeptide, can solve the problem that the ability to synthesize active secondary metabolites is not fully exerted, and achieve the effect of simple composition, simple process, and low fermentation cost

Active Publication Date: 2016-03-30
杭州水芭莎生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

It is an "outstanding" marine fungus, but from the perspective of the diversity of secondary metabolites synthesized by molds, the ability to synthesize active secondary metabolites has not been fully "played"

Method used

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  • Preparation method of cyclic dipeptide compound
  • Preparation method of cyclic dipeptide compound
  • Preparation method of cyclic dipeptide compound

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Embodiment 1: the selection of culture medium

[0028] Penicillium citrinum MNP12010101 was cultured under stress in high-concentration metal ion medium, the diversity of metabolites was analyzed by HPLC fingerprint method, and the antitumor activity of total fermented extract was detected by tetramethylazozole method (MTT method), so as to screen A metal ion can be added to the culture medium to promote the production of more anti-tumor active substances by Penicillium citrinum. Experimental metal ions include Mg 2+ , Mn 2+ , Fe 3+ 、Ba 2+ 、Co 2+ and Cu 2+ Wait. The results showed that Mg 2+ , Mn 2+ , Fe 3+ 、Ba 2+ and Cu 2+ When added to the fermentation medium, the diversity of metabolites of Penicillium citrinum was basically unchanged, and the antitumor activity of the total fermented extract was not significantly improved. Only the addition of 1.0g / L cobalt chloride (CoCl 2 ), the diversity of metabolites of Penicillium citrinum increases, and the antitu...

Embodiment 2

[0040] Embodiment 2: the selection of cobalt ion concentration of fermentation medium

[0041] The concentration of cobalt chloride in the fermentation medium was changed, and Penicillium citrinum MNP12010101 was cultured under stress. The diversity of metabolites was analyzed by HPLC fingerprint method, and the antitumor activity of total fermented extract was detected by tetramethylazozole method (MTT method). The specific experimental steps and methods are as follows:

[0042] (1) According to the method of Example 1, the seed liquid of Penicillium citrinum MNP12010101 was prepared;

[0043] (2) With the inoculation amount of step (2) seed liquid with 10% volume concentration, inoculate in the fermentation medium (250mL Erlenmeyer flask) that contains different concentrations of cobalt chloride of 50mL, see table 1 for cobalt chloride concentration, at 28 ℃, 200r / min shaking conditions, cultured for 5 days (see Table 1 for the dry weight yield of bacteria under different c...

Embodiment 3

[0051] Embodiment 3: the preparation of fermented total extract

[0052] Utilize the fermentation culture medium that contains the cobalt chloride of 1.0g / L, to Penicillium citrinum MNP12010101 large-scale fermentation culture, prepare fermentation total extract, concrete steps are:

[0053] Penicillium citrinum MNP12010101 was inoculated on the slant medium, and cultured at 28°C for 60 hours to obtain the activated strain spores; the activated strain spores were picked and inoculated into 100mL seed medium (500mL Erlenmeyer flask), Cultivate at 28°C and 200r / min shaking conditions for 36h to obtain seed liquid; transplant the seed liquid into 400mL fermentation medium (packed in a 1L Erlenmeyer flask) with an inoculum of 10% volume concentration, and inoculate at 28°C and 200r / min After culturing for 7 days under shaking conditions (the dry weight of the bacteria is 12.2 g / L), and then standing at 28° C. for 5 days, a culture was obtained. Following the steps above, 80 L of ...

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Abstract

The invention provides a preparation method of a cyclic dipeptide compound. The method comprises the following steps: inoculating penicillium citrinum MNP12010101 into a fermentation medium containing cobalt ions; carrying out cultivation for 5-7d under a constant temperature vibration condition that the temperature is 25-30 DEG C and the speed is 200-250r/min; then standing for 4-5d at 25-30 DEG C; and separating and purifying the culture solution to obtain cyclic (gly-pro) dipeptide, cyclic (phe-pro) dipeptide, cyclic (isoleucine-pro) dipeptide and cyclic (phe-val) dipeptide compounds. According to the culture method provided by the invention, cyclic dipeptides which cannot be synthesized under a conventional culture condition are synthesized in penicillium citrinum MNP12010101 cells; the culture medium is simple in composition, the process is simple and the fermentation cost is low; and the four types of cyclic dipeptides synthesized in the penicillium citrinum MNP12010101 cells have antitumor activity.

Description

(1) Technical field [0001] The invention relates to a method for preparing a cyclic dipeptide by using a marine fungus, Penicillium citrinum (Penicillium citrinum) MNP12010101 of the class Deuteromycetes, the order Calicinomycetes. (2) Background technology [0002] Marine natural products have become one of the important sources of drug lead compounds. Since the 1960s, the search for natural active substances from marine sources has become a major research focus. In the initial research, people mainly focused on some lower marine organisms, such as molluscs, algae, and plankton. With the deepening of research, more and more natural products with peculiar structures and good activities have been isolated one after another. From 1988 to 1992, the new compounds produced by marine microorganisms almost rose from zero to 12.7%, while the new compounds produced by terrestrial fungi dropped from 74.7% to 50.5%. [0003] Conventional methods to cultivate marine microorganisms, i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P17/12C12R1/80
Inventor 梅建凤金航李靓王鸿应国清易喻陈建澍张彦璐
Owner 杭州水芭莎生物医药科技有限公司
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