Camellia oleifera active glycoprotein with structure-effect modification and its preparation method and application

A technology for glycoprotein and structural transformation, which is applied in the preparation method of peptides, medical preparations containing active ingredients, peptide/protein components, etc. It can solve the structure-activity relationship between the structure and functional activity of camellia oleifera glycoproteins has not yet been clarified, and the structure-activity relationship of camellia oleifera sugar Protein fine quantification takes advantage of problems such as lack of effective and extensive data support, blank mechanism research, etc., to achieve the effect of efficient optimization of structure-efficiency modification methods

Active Publication Date: 2021-09-07
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 structure-effect modification and its preparation method and application
  • Camellia oleifera active glycoprotein with structure-effect modification and its preparation method and application
  • Camellia oleifera active glycoprotein with structure-effect modification and its preparation method and application

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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 20 mL / 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 SephadexG-100 gel filtration column, the first peak was collected, and then passed through the AKTA protein separation and purification instrument Superdex TMThe G-75 gel column was used for separa...

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 liquid-to-solid ratio of 20 mL / g, extract with water at 70°C for 2.5 hours, then filter, add absolute ethanol to the filtrate to make the final volume concentration of ethanol 80%, and freeze-dry the precipitate after centrifugation. The dried product was dissolved in 0.02mol / L Tris-HCl buffer (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 inst...

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 20 mL / g, add water to extract 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 SephadexG-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 purifica...

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Abstract

A structurally modified active glycoprotein from camellia oleifera and its preparation method and application, including degreasing and removing saponin from camellia oleifera powder, extracting with hot water for ethanol precipitation, freeze-drying the precipitate, and passing through DEAE Sepharose F.F. ion exchange column in sequence , Sephadex G‑100 gel filtration column, AKTA protein separation and purification instrument Superdex TM The G-75 gel column was used for separation and purification to obtain the active glycoprotein of camellia oleifera with a molecular weight of 25 kDa; then chemical and / or enzymatic methods were used to modify the structure of the glycoprotein. The method of the present invention can efficiently realize the preparation of glycoprotein and its structure-activity transformation. After the transformation, the activity and function of camellia oleifera glycoprotein have changed significantly, and some have been endowed with higher activity, which provides a basis for the analysis of glycoprotein structure-activity relationship. an effective technical approach.

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 Patents(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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