N-15 marked microcystis and production method thereof

A technology of microcystin and N-15, which is applied in the preparation methods of peptides, microorganism-based methods, biochemical equipment and methods, etc., can solve the problem of unclear synthetic routes, single nitrogen source of medium, and uncontrollable processes, etc. problem, to achieve the effect of simple and easy to operate production method, low cultivation cost and easy quantification

Active Publication Date: 2019-12-24
青岛普瑞邦生物工程有限公司
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0004] At present, the synthesis pathway of microcystin molecules in Microcystis is not clear, and the sources of C, H, and O in the molecules are complicated. If the isotopes of these three elements are used to label...

Method used

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  • N-15 marked microcystis and production method thereof
  • N-15 marked microcystis and production method thereof
  • N-15 marked microcystis and production method thereof

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Embodiment 3

[0078] The purification of embodiment 3 microcystins

[0079] (1) Preliminary purification: The microcystin crude extract after three transfers (the original MC-LR / RR concentrations were 1.89±0.16ppm and 1.36±0.04ppm respectively) was subjected to vacuum rotary evaporation at 40°C , Preliminary purification was carried out through an adsorption column to obtain an initial pure liquid. In this example, D101 macroporous resin was selected as a filler to perform preliminary purification of the crude extract. The steps are as follows:

[0080] A. The pretreatment of adsorption column: the 100mm*1000mm chromatographic column that swells (95% ethanol) overnight 1000g macroporous adsorption resin is loaded into the bottom with 6 layers of gauze;

[0081] B. Rinse the column twice with 1000ml 95% ethanol;

[0082] C. Use 1000ml ddH 2 O rinse column 4 times;

[0083] D. Loading of algae toxin crude extract;

[0084] E. Elution steps: successively use 1000ml ddH 2 O was eluted twi...

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Abstract

The invention discloses N-15 marked microcystis and a production method thereof. The production method comprises the following steps of microcystis aeruginosa cleaning: using the microcystis aeruginosa in a logarithmic phase as microcystis aeruginosa seeds, performing suspension cleaning on the microcystis aeruginosa seeds by deionized water, and collecting microcystis aeruginosa bodies after centrifugation, wherein the number of cleaning times is at least two; microcystis aeruginosa culture: inoculating the cleanly cleaned microcystis aeruginosa into a special culture medium with N-15, performing culture in light (the light intensity is about 2500 to 5000LUX, and the temperature is 25 to 28 DEG C), shaking up a culture bottle for 1 to 5 times every day, and performing culture for 10 to 30days to obtain culture liquid; and microcystis extraction: performing lysis treatment on microcystis aeruginosa cells in the culture liquid, performing centrifugation, and collecting liquid supernatant to obtain microcystis crude extraction liquid. The production method is simple; the operation is easy; the N-15 marking rate of the prepared microcystis is high; when the prepared microcystis is used as a standard product, a matrix effect is reduced; and the quantification is more accurate. Compared with the existing non-standard microcystis, better application prospects are realized.

Description

technical field [0001] The invention relates to the technical field of production methods of microcystins, in particular to an N-15 labeled microcystin and a production method thereof. Background technique [0002] Microcystis aeruginosa is the dominant species of cyanobacteria "freshwater bloom". It has strong adaptability and fast reproduction speed. The microcystis produced are mainly LR, RR, YR, etc., which are a kind of biologically active monocyclic heptapeptide, which can be enriched in aquatic zooplankton and fish , when the algae cells rupture and die, the toxin will be released directly into the water body. Humans will induce liver damage or even liver cancer and other diseases by drinking polluted water or eating aquatic animals. This requires fast, efficient and accurate extraction and detection of water and food. Microcystin content in medium to ensure a healthy diet. Therefore, it is urgent to produce high-purity, high-stability microcystin standard products,...

Claims

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Application Information

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IPC IPC(8): C07K14/405C07K1/22C07K1/16C12N1/12C12R1/89
CPCC07K14/405C12N1/12
Inventor 吴振兴高春蕾刘金丽刘红兵
Owner 青岛普瑞邦生物工程有限公司
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