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Camellia oleifera active glycoprotein with modified configuration, and preparation method and application of same

A glycoprotein and structural modification technology, which is applied in the preparation method of peptides, medical preparations containing active ingredients, and peptide/protein components, etc., can solve the structure-activity relationship between the structure and functional activity of Camellia oleifera glycoproteins. The refined quantitative utilization of proteins lacks effective and extensive data support, mechanism research gaps, etc., to achieve the effect of efficient optimization of structure-effectiveness modification methods

Active Publication Date: 2018-08-24
NANJING FORESTRY UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The inventor found in previous studies that Camellia oleifera also contains active glycoproteins. A series of cell experiments and animal experiments have shown that the camellia oleifera glycoprotein has good anti-tumor and antioxidant activities. The relationship has not yet been elucidated, especially the relationship between the glycosylation site in the glycoprotein and its activity function, the weight of sugar chains in the activity function of the glycoprotein, and the mechanism research at the cellular or molecular level is still blank. There is also a lack of effective and extensive data support

Method used

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  • Camellia oleifera active glycoprotein with modified configuration, and preparation method and application of same
  • Camellia oleifera active glycoprotein with modified configuration, and preparation method and application of same
  • Camellia oleifera active glycoprotein with modified configuration, and preparation method and application of same

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Effect test

Embodiment 1

[0023] Modification of camellia oleifera glycoprotein by trifluoromethanesulfonic acid (TFMS) method

[0024] Camellia oleifera is pulverized and degreased with petroleum ether, and the saponin is removed with 90% ethanol by volume fraction, and dried for later use. According to the liquid-to-solid ratio of 20mL / g, extract with water at 60°C for 3 hours, then filter, add absolute ethanol to the filtrate to make the final volume concentration of ethanol 70%, and freeze-dry the precipitate after centrifugation. The dried product was dissolved in 0.01mol / L Tris-HCl buffer (pH 8, containing 0.02mol / LNaCl) and separated and purified with DEAE Sepharose F.F. ion exchange column. Part of the 0.2mol / L NaCl eluate, after the eluate was dialyzed to remove salt, was separated and purified by Sephadex G-100 gel filtration column, the first peak was collected, and then passed through the AKTA protein separation and purification instrument Superdex TM The G-75 gel column was used for sepa...

Embodiment 2

[0027] Periodic acid (NaIO 4 ) to transform camellia oleifera glycoprotein

[0028] Camellia oleifera is pulverized and degreased with petroleum ether, and the saponin is removed with 90% ethanol by volume fraction, and dried for later use. According to the ratio of liquid to material of 20mL / g, extract with water at 70°C for 2.5h and then filter, add absolute ethanol to the filtrate to make the final volume concentration of ethanol 80%, centrifuge and freeze-dry the precipitate. The dry matter was dissolved in 0.02mol / L Tris-HCl buffer solution (pH 8, containing 0.02mol / L NaCl) and separated and purified with DEAE Sepharose F.F. ion exchange column, based on the results of in vitro anti-tumor and antioxidant activities. Collect the part of the 0.3mol / L NaCl eluate. After the eluate is dialyzed to desalt, it is separated and purified by Sephadex G-100 gel filtration column, the first peak is collected, and then passed through the AKTA protein separation and purification instr...

Embodiment 3

[0031] Enzymatic transformation of camellia oleifera glycoprotein with PNGase F

[0032] Camellia oleifera is pulverized and degreased with petroleum ether, and the saponin is removed with 90% ethanol by volume fraction, and dried for later use. According to the liquid-to-solid ratio of 20mL / g, extract with water at 80°C for 2 hours, then filter, add absolute ethanol to the filtrate to make the final volume concentration of ethanol 85%, and freeze-dry the precipitate after centrifugation. The dried product was dissolved in 0.03mol / L Tris-HCl buffer (pH 8, containing 0.02mol / LNaCl) and separated and purified with DEAE Sepharose F.F. ion exchange column. Part of the 0.4mol / L NaCl eluate, after the eluate was dialyzed to remove salt, was separated and purified by Sephadex G-100 gel filtration column, the first peak was collected, and then passed through the AKTA protein separation and purification instrument Superdex TM The G-75 gel column was used for separation and purificati...

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Abstract

A camellia oleifera active glycoprotein with modified configuration, and a preparation method and an application of same. The preparation method includes steps of: degreasing camellia oleifera powderand removing saponin, extracting the camellia oleifera powder with hot water and precipitating the extract with anhydrous ethanol, freeze-drying the precipitate, and performing separation and purification successively through a DEAE Sepharose F.F. ionic exchange column, a Sephadex G-100 gel filtering column, and a Superdex TM G-75 gel column of an AKTA protein separation and purification device, thus producing the camellia oleifera active glycoprotein, molecular weight being 25 kDa, finally performing configuration modification to the glycoprotein via a chemical process and / or an enzyme process. The method high-effectively solves preparation and configuration modification to the glycoprotein, wherein the modified glycoprotein is significantly changed in active functions, some gaining higher activity. The method supplies an effective technical approach for analysis of structure-function relationship of the glycoprotein.

Description

technical field [0001] The invention relates to the field of food biotechnology, in particular to a structure-effect modified active glycoprotein of camellia oleifera and its preparation method and application. Background technique [0002] Camellia oleifera (Camellia oleifera Abel.) is a plant of the genus Camellia in the family Theaceae. It is famous for producing camellia seed oil. It is an excellent woody oil plant unique to my country. A multi-purpose plant with multiple functions. Camellia oleifera recorded in traditional medicine has the effects of clearing away heat and detoxifying, promoting blood circulation and dissipating blood stasis, and relieving pain. Modern research reports have shown that camellia oleifera contains rich active ingredients such as saponin, polysaccharides, and flavonoids, which can regulate immunity, regulate blood lipids and blood sugar, and resist oxidation. However, the main way to develop and utilize Camellia oleifera is to extract oil f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/12C07K1/16C07K1/18C12P21/00A61K38/16A61K39/00A61P35/00
CPCA61K38/00C07K1/12C07K1/16C07K1/18C12P21/005
Inventor 李婷婷徐云巧吴彩娥范龚建
Owner NANJING FORESTRY UNIV
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