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CN-121975715-A - Nanometer vesicle derived from alpinia galanga as well as preparation method and application thereof

CN121975715ACN 121975715 ACN121975715 ACN 121975715ACN-121975715-A

Abstract

The invention relates to the technical field of biological medicine, in particular to a nano vesicle derived from alpinia galanga, and a preparation method and application thereof. The preparation method comprises the steps of crushing rhizome of galangal, mixing the rhizome of galangal with a buffer solution, soaking the rhizome of galangal in water bath, filtering to remove solids to obtain a crude extract, and sequentially carrying out four centrifugal treatments on the crude extract to obtain the nanometer vesicle from galangal. The invention provides a method for extracting nano vesicles from galangal, and discovers that the obtained vesicles can promote osteogenic differentiation of human bone marrow mesenchymal stem cells with high efficiency and low toxicity for the first time, the effect of the nano vesicles is obviously superior to that of galangin which is a known active ingredient, the cytotoxicity is obviously lower, and the nano vesicles provide a potential raw material and a method for developing novel and safe anti-osteoporosis drugs or inducers.

Inventors

  • WANG JIAFENG
  • XU BO
  • ZHOU YULAN
  • JIANG YOUPING
  • CHENG ZHIYONG
  • LIN BIYUN

Assignees

  • 广东医科大学附属医院

Dates

Publication Date
20260505
Application Date
20260206

Claims (9)

  1. 1. The preparation method of the nano vesicle from the alpinia galanga is characterized by comprising the following steps of: (1) Crushing rhizome of galangal, mixing with buffer solution, soaking in water bath, and filtering to remove solid to obtain crude extract; (2) Performing first low-speed centrifugation on the crude extract, taking supernatant, and filtering by using a filter with the aperture of 70 mu m; (3) Performing a second low-speed centrifugation on the filtrate obtained in the step (2), taking supernatant, and filtering by using a filter with the pore diameter of 40 mu m; (4) Performing medium-speed centrifugation on the filtrate obtained in the step (3) for the third time, taking supernatant, and filtering by using a filter with the pore diameter of 0.45 mu m; (5) And (3) performing fourth ultracentrifugation treatment on the filtrate obtained in the step (4), discarding supernatant, and collecting precipitate to obtain the galangal-derived nano vesicles.
  2. 2. The preparation method of claim 1, wherein the galangal in step (1) is mixed with a buffer solution according to a mass-volume ratio of 1:1-1.5 g/mL, and the buffer solution is PBS buffer solution.
  3. 3. The method according to claim 1, wherein the water bath in step (1) has a temperature of 55 to 65 ℃ and a time of 10 to 14 hours.
  4. 4. The method according to claim 1, wherein the first low-speed centrifugation is performed at a relative centrifugal force of 300 to 500 Xg for 20 to 30 minutes, and the second low-speed centrifugation is performed at a relative centrifugal force of 3,000 to 5,000 Xg for 25 to 35 minutes.
  5. 5. The method according to claim 1, wherein the third intermediate speed centrifugation is performed at a relative centrifugal force of 12,000 to 15,000 Xg for 25 to 35 minutes, and the fourth intermediate speed centrifugation is performed at a relative centrifugal force of 100,000 to 120,000 Xg for 60 to 80 minutes.
  6. 6. A nanovesicle derived from galangal prepared by the method of any one of claims 1-5.
  7. 7. The nanovesicle according to claim 6, wherein the nanovesicle has a phospholipid bilayer structure, and has an average particle diameter of 50 to 200 nm.
  8. 8. Use of the nanovesicle of claim 6 or 7 in the preparation of a formulation for promoting osteogenic differentiation of bone marrow mesenchymal stem cells.
  9. 9. Use of the nanovesicles according to claim 6 or 7 for the preparation of a medicament or biomaterial for the prevention or treatment of osteoporosis.

Description

Nanometer vesicle derived from alpinia galanga as well as preparation method and application thereof Technical Field The invention relates to the technical field of biological medicine, in particular to a nano vesicle derived from alpinia galanga, and a preparation method and application thereof. Background Osteoporosis is a systemic metabolic chronic disease characterized by a decrease in bone density and mass due to deterioration of the bone trabecular microstructure, a decrease in number, and an accompanying increase in the risk of fracture. According to world health organization data, more than 2 million people worldwide have suffered from this disease. With the continuous increase of the aging degree of the social population, the prevention and treatment of osteoporosis have become an important public health challenge. Although the existing clinical commonly used synthetic drugs can improve bone metabolism to a certain extent, side effects with different degrees such as gastrointestinal reactions, cardiovascular risks and the like generally exist, so that the safety of long-term application of the synthetic drugs is limited. Therefore, the exploration of safe and sustainable control strategies, especially the intervention by dietary or natural active ingredients, has important practical significance for the prevention and auxiliary treatment of osteoporosis. In recent years, nanovesicles have received widespread attention as an emerging class of bioactive carriers. The cell-derived nano vesicle with a phospholipid bilayer structure can carry bioactive substances such as protein, lipid, nucleic acid, small molecules and the like, participate in intercellular communication and signal transduction, and play a key role in regulating cell growth, differentiation, metabolism and the like. It is worth noting that not only animal cells and plant-derived nano vesicles are found to have various biological activities, but also have potential application values in the aspects of anti-inflammation, anti-cancer, tissue regeneration promotion and the like. This provides a new idea for developing natural, low-toxicity food-borne active delivery systems. Galangal is a common zingiberaceous plant in Guangdong, hainan and the like, and the rhizome of galangal is used as a medicine and food homologous material for a long time, and the traditional Chinese medicine considers that galangal has the effects of warming stomach, dispelling cold, promoting digestion, relieving pain and the like. Modern researches have further revealed that the extract of galangal (e.g. galangin) has anti-inflammatory and antioxidant properties, and can promote osteoblast activity and inhibit osteoclast formation, thereby reducing bone loss and reducing the risk of osteoporosis. However, recent researches have found that plant nanovesicles isolated from galangal exhibit a more remarkable effect in promoting bone differentiation than traditional extracts of galangin-not only can significantly up-regulate the expression of human bone marrow mesenchymal stem cell osteogenic differentiation marker proteins, promote bone formation, but also have lower cytotoxicity and better safety. The nano vesicle derived from galangal is expected to be a natural active ingredient with more potential and safer osteoporosis prevention and treatment, and deserves further intensive research and development. Disclosure of Invention The invention provides a method for extracting nano vesicles from fresh galangal rhizome and application thereof. The method obtains the nano vesicle through crushing, soaking, filtering and high-speed centrifugation. Experiments show that the nano vesicle can obviously up-regulate the expression of the osteogenic differentiation marker protein, has better osteogenic induction effect on human bone marrow mesenchymal stem cells than galangin, and has lower cytotoxicity. Therefore, the nano vesicle can be used as an osteogenic differentiation inducer and has good application prospect in the development of anti-osteoporosis medicines. In order to achieve the above object, the present invention provides the following technical solutions: the invention provides a preparation method of a nano vesicle from alpinia galanga, which comprises the following steps: (1) Crushing rhizome of galangal, mixing with buffer solution, soaking in water bath, and filtering to remove solid to obtain crude extract; (2) Performing first low-speed centrifugation on the crude extract, taking supernatant, and filtering by using a filter with the aperture of 70 mu m; (3) Performing a second low-speed centrifugation on the filtrate obtained in the step (2), taking supernatant, and filtering by using a filter with the pore diameter of 40 mu m; (4) Performing medium-speed centrifugation on the filtrate obtained in the step (3) for the third time, taking supernatant, and filtering by using a filter with the pore diameter of 0.45 mu m; (5) And (3) performing fourth ultracentrif