Preparation and verification method of Chlorella protein molecule

A technology of protein molecules and chlorella, applied in material separation, analysis of materials, algae/moss peptides, etc., can solve problems that do not involve the identification of protein molecular chemical structures

Inactive Publication Date: 2012-09-19
TIANJIN AGRICULTURE COLLEGE +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0012] 2. The traditional method does not involve the identification of the chemical structure of the protein molecule after separation and purification. It is very important to clarify the specific chemical structure of the protein molecule for the next step of artificial synthesis and application of functional proteins

Method used

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Examples

Experimental program
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Embodiment Construction

[0023] Suspend the chlorella mud in 0.001mol / L phosphate buffer solution (PBS) with pH 7.0, freeze and thaw 3 times; ultrasonically break in ice bath for 9 minutes; Add 30% saturated ammonium sulfate to the supernatant; let stand at 4°C for 24 hours, then centrifuge at 12000r / min for 30min at 4°C; concentrate the supernatant to 200mL with an ultrafiltration cup, and precipitate Dissolve with a small amount of PBS, place in a dialysis membrane, stir and dialyze in 0.001mol / LPBS at pH 7.0 for 2 days, embed and concentrate in polyethylene glycol-6000 to 60mL;

[0024] Through silica gel column chromatography: the metabolites are dry mixed with silica gel and then roughly separated by silica gel column chromatography. The eluent is petroleum ether, petroleum ether with a volume ratio of 1:1: dichloromethane (V / V), volume Petroleum ether at a ratio of 1:2: dichloromethane (1:2, V / V), dichloromethane, dichloromethane at a volume ratio of 5:1: methanol (5:1, V / V), a volume ratio of 5...

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PUM

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Abstract

The invention relates to a preparation and verification method of a Chlorella protein molecule, which comprises the steps of: suspending Chlorella mud in pH 7.0 0.001mol / L phosphate buffer solution (PBS), and repeatedly freezing and thawing three times; ultrasonically crushing in an ice bath for 9min; centrifuging at the temperature of 4 DEG C and 12000r / min for 30min, collecting supernatant and adding ammonium sulfate with saturation degree of 30%; standing at the temperature of 4 DEG C for 24h, and centrifuging at the temperature of 4 DEG C and 12000r / min for 30min; and concentrating the supernatant with an ultrafiltration cup to 200mL, dissolving precipitate with little PBS, placing in a dialysis membrane, dialyzing in pH 7.0 0.001mol / L PBS under stirring for 2d, and embedding with polyethylene glycol-6000 to 60mL. By the adoption of the method to separate the purified protein molecule of Chlorella, the protein is released from the tissue or cell, the original native state is kept, and the activity is not lost, so that the specific chemical structure of the protein molecule can be accurately determined.

Description

Technical field: [0001] The invention belongs to the preparation and identification of biopharmaceuticals and health food. In particular, it relates to a method for preparing and identifying chlorella protein molecules. Background technique: [0002] Currently available protein molecule preparations are: [0003] (1) Technical process for separation and purification of protein molecules [0004] 1. Material pretreatment and cell disruption [0005] The main methods are: ①Mechanical crushing method: use the shearing effect of mechanical force to break up the cells. Commonly used equipment includes high-speed tissue masher, homogenizer, mortar, etc.; ② osmotic crushing method: the cells are swollen and broken under hypotonic conditions; ③ repeated freezing and thawing method: after the biological tissue is frozen, the intracellular fluid freezes Swell to burst the cells; ④ Ultrasonic method: use an ultrasonic oscillator to cause uneven tension on the cell membrane to break...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/405C07K1/36C07K1/34C07K1/30C07K1/16G01N30/02G01N30/72G01N24/08
Inventor 周文礼陈成勋孙金辉毛海涛王庆奎毕相东乔秀亭邢克智何家瑞韩鈺松
Owner TIANJIN AGRICULTURE COLLEGE
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