Method for extracting and purifying microcystin

A technology of microcystin and purification method, which is applied in the field of extraction and purification of microcystin, can solve the problems of low sample loading, high cost, and low price, and achieve simple processing, low requirements, and simple operation Effect

Inactive Publication Date: 2012-12-12
INST OF AQUATIC LIFE ACAD SINICA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the methods for extracting and purifying microcystins from Microcystis mainly include: separation by thin-layer chromatography (Chinese patent, a method for extracting and purifying microcystins, publication number CN1401660A, application date: 2002.9 .24), the method uses acetic acid solution and methanol to soak Microcystis, and extracts and purifies Microcystin through steps such as impurity removal, crude toxin preparation, toxin separation, and thin-layer chromatography technology. This method has thin-layer The analysis plate can only be used once, the cost is high and the sample volume is low; Wei Tao et al. Using cyanobacteria as raw materials, a method for the extraction, purification and preparation of microcystins was established with methanol solution extraction, solid phase extraction and semi-preparative chromatographic separation as the main steps
Although the price of this method is relatively low and the purity of the prepared toxin is high, the toxin is not fully eluted when passing through the column, resulting in a large loss of toxin

Method used

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  • Method for extracting and purifying microcystin
  • Method for extracting and purifying microcystin
  • Method for extracting and purifying microcystin

Examples

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Effect test

Embodiment 1

[0032] A method for extracting and purifying microcystins, the operation steps are as follows:

[0033] 1. Weigh 3.000g of algae powder, put it into a 250ml Erlenmeyer flask, add 90ml of 60v / v% methanol to dissolve,

[0034] Shake at 200rpm at 8°C for 1h, then sonicate at 20°C for 20min, transfer to four 40ml plastic centrifuge tubes, centrifuge at 12000rpm at 20°C for 20min;

[0035] 2. Pour out the supernatant and combine it into a 200ml beaker, soak and wash the 4 parts of the precipitate obtained in step 1 with 50ml of 60v / v% methanol, then transfer all the 50ml cleaning solution into the original 250ml Erlenmeyer flask, shake at 200rpm at 8°C Shake for 1 hour, then ultrasonically induce solubilization at 20°C for 20 minutes, transfer to two 40ml plastic centrifuge tubes, centrifuge at 12,000 rpm for 20 minutes at 20°C, pour out the supernatant and combine it into the above-mentioned 200ml beaker;

[0036] 3. Adjust the above 6 combined supernatants to pH 3.7 with 1...

Embodiment 2

[0068] A method for extracting and purifying microcystins, the operation steps are as follows:

[0069] 1. Weigh 3.00g of algae powder, put it into a 250ml Erlenmeyer flask, add 100ml of 60v / v% methanol to dissolve, oscillate on a shaker at 200rpm at 15°C for 1h, and then ultrasonically induce dissolution at 25°C for 20min, then transfer to four 40ml plastic bottles Centrifuge tube, centrifuge at 12000rpm at 20°C for 20min;

[0070] 2. Pour out the supernatant and combine it into a 200ml beaker. The four precipitates obtained in step 1 are soaked and washed in 50ml of 60v / v% methanol, and then all the 50ml of the cleaning solution is transferred to the original 250ml Erlenmeyer flask, and shaker at 200rpm at 15°C Shake for 1 hour, and then ultrasonically induce solubilization at 25°C for 20 minutes, then transfer to two 40ml plastic centrifuge tubes, centrifuge at 12,000 rpm for 20 minutes at 20°C, pour out the supernatant and combine it into the above-mentioned 200ml beaker; ...

Embodiment 3

[0086] A method for extracting and purifying microcystins, the operation steps are as follows:

[0087] 1. Weigh 3.00g of algal powder, put it into a 250ml Erlenmeyer flask, add 120ml of 60v / v% methanol to dissolve, oscillate on a shaker at 200rpm at 20°C for 1h, and then ultrasonically induce dissolution at 30°C for 20min, then transfer to four 40ml plastic bottles Centrifuge tube, centrifuge at 12000rpm at 20°C for 20min;

[0088] 2. Pour out the supernatant and combine it into a 200ml beaker. The 4 parts of the precipitate obtained in step 1 are soaked and washed with 50ml of 60v / v% methanol, and then all the 50ml of the cleaning solution is transferred to the original 250ml Erlenmeyer flask and shaken at 20°C at 200rpm 1h, after ultrasonic solubilization treatment at 30°C for 20min, transfer to two 40ml plastic centrifuge tubes, centrifuge at 12000rpm at 20°C for 20min, pour out the supernatant and combine it into the above-mentioned 200ml beaker;

[0089] 3. Adjust the a...

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Abstract

The invention discloses a method for extracting and purifying microcystin, in particular a method for extracting high-purity microcystin MC-RR and MC-LR from a raw material of dried powder of bloom cyanobacteria harvested in the field. The method comprises the following steps of: A, crudely extracting the microcystin MC-RR and MC-LR by using methanol; B, making the crude extracting solution pass through an ODS column to finely extract the microcystin MC-RR and MC-LR; C, separating the microcystin MC-RR from MC-LR by using a semi-preparative chromatograph; and D, saving the microcystin MC-RR and MC-LR. The method for extracting and purifying the microcystin has the advantages of wide sources of raw materials, low cost, simple operating process, purity of over 97 percent of the extracted microcystin, and capacity of meeting the requirement on toxin purity for scientific research.

Description

technical field [0001] The method of the invention relates to the technical field of biochemical separation, in particular to a method for extracting and purifying microcystins. The method of the invention is suitable for extracting and purifying two kinds of microcystins MC-RR and MC-LR from cyanobacteria in the bloom period in the field. Background technique [0002] In the past ten years, with the intensification of water pollution and eutrophication, the phenomenon of cyanobacterial blooms in freshwater lakes in my country has become more and more serious, and the relationship between the accompanying toxins and human health has received more and more attention. Among them, microcystin (hereinafter referred to as MC) is the most widely distributed and most harmful type of hepatotoxin among the various cyanotoxins that have been found. Body liver damage. The chemical structure of this type of toxin is relatively stable, and it can bring potential harm to the human body ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/405C07K1/20C07K1/16C07K1/14
Inventor 谢平王文静国晓春梁高道吴来燕张伟戴明张大文
Owner INST OF AQUATIC LIFE ACAD SINICA
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