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Method for preparing phycoerythrin through combination of centrifugal method and anion exchange chromatography media

A technology of phycoerythrin and exchange chromatography, applied in chemical instruments and methods, algae/bryopeptides, peptide sources, etc., can solve the problems of difficult large-scale production, unstable product quality, high production cost, etc., and achieve recovery rate High, simple operation method, low cost effect

Inactive Publication Date: 2016-11-16
YANTAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, people mainly use sulfuric acid to prepare phycoerythrin by combining fractional precipitation, hydroxyapatite column chromatography, and gel filtration chromatography. The existing preparation process has complex operations, high production costs, and unstable product quality. , and it is difficult to achieve large-scale industrial production

Method used

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  • Method for preparing phycoerythrin through combination of centrifugal method and anion exchange chromatography media
  • Method for preparing phycoerythrin through combination of centrifugal method and anion exchange chromatography media

Examples

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Embodiment 1

[0018] (1) Centrifuge to collect 5 g of freshly cultivated Porphyridium algae, mix the algae mud with phosphate buffer (pH 6.8, 0.02mol / L) in a ratio of 1:6 to obtain a cell suspension;

[0019] (2) Ultrasonicate the cell suspension at 0-4˚C, with an output power of 500W, crush for 4s, intermittently for 4s, and the crushing time is 6min; obtain the broken wall liquid; freeze and centrifuge the broken wall liquid, and collect Serum;

[0020] (3) At room temperature, the supernatant was ultrafiltered by a membrane bag with a molecular cut-off of 50kD, ultrafiltered 5 times, and the filtrate was collected to obtain a crude phycoerythrin extract;

[0021] (4) Add chromatographic medium Source15Q (0.2g) pretreated with phosphate buffer (pH 6.8, 0.02mol / L) to the obtained phycoerythrin crude extract, mix well, refrigerate and centrifuge, and take the precipitate; Add 2 times the volume of phosphate buffer containing 0.1mo / L NaCl to the precipitation, mix well, refrigerate and cent...

Embodiment 2

[0024] (1) Centrifuge to collect 50 g of freshly cultivated Porphyridium algae, mix the algae mud with phosphate buffer (pH 6.8, 0.01mol / L) in a ratio of 1:8 to obtain a cell suspension;

[0025] (2) Ultrasonicate the cell suspension at 0-4˚C, with an output power of 500W, crush for 3s, intermittently for 3s, and the crushing time is 8min, to obtain the wall-breaking liquid; centrifuge the wall-breaking liquid at low temperature, and collect the supernatant liquid;

[0026] (3) At room temperature, the supernatant was ultrafiltered by a membrane bag with a molecular cut-off of 60kD, ultrafiltered 4 times, and the filtrate was collected to obtain a crude phycoerythrin extract;

[0027] (4) Add chromatographic medium Source15Q (2g) pretreated with phosphate buffer (pH 6.8, 0.01mol / L) to the obtained phycoerythrin crude extract, mix well, refrigerate and centrifuge, and take the precipitate; Add 2 times the volume of phosphate buffer containing 0.1 mol / L NaCl (pH 6.8, 0.01 mol / L...

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Abstract

The invention provides a method for preparing phycoerythrin through combination of a centrifugal method and anion exchange chromatography media. The method comprises the following steps of using fresh porphyridium cruentum as a raw material, performing centrifugation, mixing porphyridium cruentum paste with a phosphate buffer solution according to a certain proportion, and then performing ultrasonic crushing treatment so as to obtain a wall-breaking solution; and then performing refrigerated centrifugation on the wall-breaking solution, taking a supernatant, performing ultrafiltration on the supernatant with a membrane bag of which the molecule interception is 50-60kD for 4-6 times, adding strong anion chromatography media pretreated by the phosphate buffer solution, performing refrigerated centrifugation twice, and taking a supernatant so as to obtain the phycoerythrin of which the purity A<545> / A<280> is greater than 4.0. The preparation method disclosed by the invention is simple and convenient in technology, low in cost, good in purification effects, high in protein recovery rate and easy to amplify, and has a good popularization value and good application prospects.

Description

technical field [0001] The invention belongs to the technical field of preparation of marine biological active substances, and in particular relates to a method for preparing high-purity phycoerythrin by centrifugation combined with an anion exchange chromatography medium. Background technique [0002] Phycobiliprotein (PBP) is a kind of light-harvesting pigment protein present in cyanobacteria, red algae, cryptophyta and a few dinoflagellates. According to the characteristics of ultraviolet absorption spectrum, phycobiliproteins are divided into three categories: phycoeyanin (PC), allophycoeyanin (APC) and phycoerythrin (PE). Phycoerythrin is mainly distributed in marine red algae. It is composed of apoprotein and chromophore. The apoprotein is a kind of oligomeric protein, and the basic units are α and β subunits. There are a small number of gamma subunits. Due to the existence of pigment molecules, phycoerythrin has strong light absorption at 480-570nm in the visible li...

Claims

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Application Information

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IPC IPC(8): C07K14/405C07K1/36C07K1/34C07K1/18
CPCC07K14/405
Inventor 唐志红吕家森赵吉路张超
Owner YANTAI UNIV
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