Method for separating and purifying high-purity phycobiliprotein from spirulina

Abstract

A method for separating and purifying high-purity phycobiliprotein from spirulina is characterized in that the following steps are carried out at the temperature of 4 DEG C: taking out the spirulina, first breaking spirulina body by ultrasound with the ultrasound time of 10 minutes in a way of gap breaking, and then obtaining initial spirulina extraction solution by centrifugating at high speed for 30 minutes; next, adding ammonium sulfate into the extraction solution to reach 40% saturation, extracting supernatant after centrifugating at medium speed for 15 minutes, adding the ammonium sulfate continuously to reach 50% saturation, and extracting precipitation after centrifugating at medium speed for 15 minutes; then centrifugating the precipitation at high speed for 30 minutes after dissolving the precipitation by using phosphate buffer solution with extremely low concentration, filtering the obtained supernatant by using microporous filtering film with low pore size, desalinating the filtered solution, and adding the desalinated solution to hydroxyapatite column; finally eluting and collecting the phycocyanin from the hydroxyapatite column by using 0.040 M to 0.060 M phosphate salt buffer solution (pH7.0, containing 1 mmol / L EDTA); and eluting and collecting the changed phycocyanin from the hydroxyapatite column by using 0.10 M to 0.12 M phosphate salt buffer solution (pH7.0, containing 1 mmol / L EDTA).

Description

Technical field: [0001] The invention relates to a method for separating and purifying high-purity phycobiliprotein, in particular to a method for separating and purifying high-purity phycobiliprotein from spirulina. Background technique: [0002] Phycocyanin and variable phycocyanin are good pure natural pigments with bright colors, which can be used as food pigments and in the cosmetics industry without causing artificial harm. They are ideal and safe additives. At the same time, Phycocyanin and variable phycocyanin are also active substances, both of which have stable fluorescence and generate free radicals, and can be used as antibody fluorescent labeling substances to replace synthetic fluorescent substances. Labeled immune antibody reagents are used for clinical molecular diagnosis of medical diseases and life science molecular detection. Both phycocyanin and phycocyanin can be used as photosensitizers, and more and more people are optimistic about the prospect of pho...

AI Technical Summary

Problems solved by technology

At present, phycobiliproteins can be extracted from several kinds of seaweed, but the price of high-purity phycobiliproteins is very expensive, and the price is still rising rapidly

[0003] Regarding the extraction and purification of phycobiliproteins, although there are many studies, and most of them are extracted from spirulina, some processes are simple, and the purity of the protein obtained is extremely low, and some processes are too complicated and easy to denature the protein. , not suitable for a large amount of extraction, some involve ultrafiltration, which can easily cause solutes to adhere and deposit on the surface of the membrane, resulting in serious concentration polarization and blockage, and some in the extraction process, food stabilizers, sugar Classes, inorganic salts, pH regulators, etc., generally only pharmaceutical grade (A 615nm / A 280nm >2) phycocyanin, prepare the extract of algae with a hypotonic method, and the purity of phycocyanin reaches reagent grade (A 615nm / A 280nm >4)

Just because of these defects that the above-mentioned separation techniques exist, the purity of the phycobiliproteins obtained is low, and the yield is not high, so that the price of high-purity phycobiliproteins is high, and finally, greatly affects the production of high-purity phycobiliproteins. Wide application, therefore, reducing the cost of phycobiliprotein purification is the key