阅读说明：本技术 一种海藻功能油及其应用 (Seaweed functional oil and application thereof ) 是由 张翼 杨文聪 张嫄嫄 千忠吉 李亚娟 刘亚月 杨静明 聂影影 宋采 梁金月 马小 于 2019-11-20 设计创作，主要内容包括：本发明公开了一种海藻功能油及其应用。本发明首先提供了一种海藻功能油,所述海藻功能油是海藻羊栖菜粗提物先由石油醚和乙酸乙酯的混合溶液洗脱,再由石油醚、氯仿和甲醇的混合溶液洗脱后,收集满足薄层层析比移值为0.1～0.4且乙酰胆碱酯酶生物自显影为白色斑点的活性组分,即得到所述海藻功能油。该海藻功能油具有显著的抑制乙酰胆碱酯酶的活性,清除DPPH自由基的活性,抑制细胞内炎症因子诱导的NO生成,清除细胞内的ROS水平的功能,且对正常细胞无任何毒副作用；另外,该制备方法产量高,材料来源天然,所得海藻功能油无毒性,不会引起不良反应,在制备治疗阿尔茨海默氏症的药物方面具有很好的应用前景。(The invention discloses seaweed functional oil and application thereof. The invention firstly provides seaweed functional oil, which is obtained by eluting a crude extract of sargassum fusiforme by a mixed solution of petroleum ether and ethyl acetate, eluting by a mixed solution of petroleum ether, chloroform and methanol, and collecting active components which meet the requirements that a thin-layer chromatography specific transfer value is 0.1-0.4 and acetylcholinesterase is subjected to biological self-development to form white spots. The seaweed functional oil has the functions of obviously inhibiting the activity of acetylcholinesterase, eliminating the activity of DPPH free radicals, inhibiting the generation of NO induced by inflammatory factors in cells and eliminating the level of ROS in the cells, and has NO toxic or side effect on normal cells; in addition, the preparation method has high yield and natural material sources, and the obtained seaweed functional oil has no toxicity, does not cause adverse reactions and has good application prospect in the aspect of preparing medicines for treating Alzheimer's disease.)

1. The seaweed functional oil is characterized in that a seaweed sargassum fusiforme crude extract is eluted by a mixed solution of petroleum ether and ethyl acetate, then eluted by a mixed solution of petroleum ether, chloroform and methanol, and then an active component which meets the requirements that a thin-layer chromatography specific transfer value is 0.1-0.4 and acetylcholinesterase is subjected to biological self-development to form white spots is collected, so that the seaweed functional oil is obtained.

2. The seaweed functional oil according to claim 1, characterized in that the preparation method comprises the following steps:

s1, drying and crushing sargassum fusiforme, adding a mixed solution of methanol and chloroform for extraction, and concentrating under reduced pressure to obtain a sargassum fusiforme crude extract;

s2, subjecting the crude extract of the seaweed sargassum fusiforme obtained in the step S1 to reduced pressure column chromatography, and performing column chromatography by using a solvent with a volume ratio of 8: 2 and 7: 3, eluting the mixed solution of petroleum ether and ethyl acetate to obtain a crude component 1 and a crude component 2;

s3, mixing the crude component 1 and the crude component 2 obtained in the step S2, and treating the mixture by a silica gel column at a volume ratio of 9: 1, eluting with a mixed solution of petroleum ether and ethyl acetate, and passing through a gel column by using a solvent with a volume ratio of 2: 1: eluting with mixed solution of petroleum ether, chloroform and methanol to obtain the seaweed functional oil.

3. The seaweed functional oil as claimed in claim 2, wherein the filler of the reduced pressure column in step S2 is silica gel of 100-200 mesh.

4. The seaweed functional oil as claimed in claim 2, wherein the silica gel column of step S3 is filled with 200-300 mesh silica gel.

5. The seaweed functional oil of claim 2, wherein the filler of the gel column of step S3 is sephadex.

6. Use of the seaweed functional oil of any one of claims 1 to 5 in the preparation of a medicament for treating alzheimer's disease.

7. Use of the seaweed functional oil of any one of claims 1 to 5 in the preparation of a product for preventing and/or assisting in the improvement of Alzheimer's disease.

8. The use according to claim 6, in the manufacture of a medicament for inhibiting acetylcholinesterase activity.

9. The use according to claim 6, wherein the use is for the preparation of a medicament having DPPH radical scavenging activity.

10. The use according to claim 6, for the manufacture of a medicament for inhibiting the production of NO and ROS levels in a cell.

Technical Field

The invention belongs to the technical field of essential oil extraction. More particularly, relates to seaweed functional oil and application thereof.

Background

Alzheimer's Disease (AD), commonly known as senile dementia, is one of the major diseases endangering human health in the 21 st century, is a neurodegenerative disease accompanied by cognitive, behavioral and functional disorders, and is frequently seen in elderly people over 65 years old. The existing research shows that AD is closely related to oxidative stress in the brain, neurodegeneration caused by neuroinflammation, cholinergic neuron injury and the like. According to 2016 world AD report, the number of people suffering from AD in the current year is up to 4700 thousands, and with the increasing aging progress of population, the number of AD patients in China increases at the fastest global speed, and related experts predict that 2200 thousands of people will suffer from AD in the world by 2025 years. Meanwhile, the cost of AD patients for medicine and health care reaches trillion annually, which has great influence on the physical and mental health of the patients and the national economic development.

Currently, drugs for treating AD include acetylcholinesterase inhibitors and vaccines such as tacrine, tolazamide, rivastigmine, galantamine, memantine hydrochloride, and the like. However, the drugs or vaccines are expensive, have great toxic and side effects, have limited effects on treating AD, can only improve AD symptoms but cannot prevent disease progression, and can cause a plurality of adverse reactions.

The marine organisms generate and accumulate a large amount of secondary metabolites with new structures and strong activity in the growth and metabolism process due to the special living environment of the marine organisms, and the marine organisms are important resources for researching and discovering novel marine medicines. Research shows (Zhang Gan et al, 2005, research on inhibition of acetylcholinesterase activity by seaweed components), some kinds of seaweed components have activity of inhibiting acetylcholinesterase; but the content difference of the effective components of different algae is obvious, the related research is less at present, and the extraction results of the related active components of different algae by different extraction processes show great difference. Therefore, the medicine for treating Alzheimer's disease, which has good activity, high yield, no toxicity and natural sources, is screened out, and has important significance for preventing and treating AD.

Disclosure of Invention

The invention aims to provide the seaweed functional oil with the functions of inhibiting acetylcholinesterase, resisting oxidation and inflammation and the preparation method thereof.

The invention aims to provide seaweed functional oil.

The invention also aims to provide the application of the seaweed functional oil in preparing the medicine for treating the Alzheimer's disease.

The invention further aims to provide application of the seaweed functional oil in preparing a product for preventing and/or assisting in improving Alzheimer's disease.

The above purpose of the invention is realized by the following technical scheme:

the invention firstly provides seaweed functional oil, which is obtained by eluting a crude extract of sargassum fusiforme by a mixed solution of petroleum ether and ethyl acetate, eluting by a mixed solution of petroleum ether, chloroform and methanol, and collecting active components which meet the requirements that a thin-layer chromatography specific transfer value is 0.1-0.4 and acetylcholinesterase is subjected to biological self-development to form white spots.

The main components of the seaweed functional oil are myristic acid, (Z) -9-hexadecenoic acid, palmitic acid, phytol, arachidonic acid and 11,14, 17-eicosatrienoic acid, and the structures of the seaweed functional oil are respectively shown as follows:

the inventor obtains the seaweed functional oil when the thin-layer chromatography-biological self-development method and the 96-pore plate method are utilized to carry out activity tracing, organic solvent extraction and column chromatography are utilized to purify the active components of the seaweed sargassum fusiforme; through analysis and experimental verification, the seaweed functional oil has the functions of obviously inhibiting the activity of acetylcholinesterase, eliminating the activity of DPPH free radicals, inhibiting the generation of NO induced by inflammatory factors in brain-derived BV-2 microglia, eliminating the ROS level in the brain-derived BV-2 microglia, and has NO toxic or side effect on normal cells; therefore, the seaweed functional oil has good application prospect in preparing medicines for treating Alzheimer's disease and products for preventing and/or assisting in improving Alzheimer's disease.

Preferably, the preparation method of the seaweed functional oil comprises the following steps:

s1, drying and crushing sargassum fusiforme, adding a mixed solution of methanol and chloroform for extraction, and concentrating under reduced pressure to obtain a sargassum fusiforme crude extract;

s2, subjecting the crude extract of the seaweed sargassum fusiforme obtained in the step S1 to reduced pressure column chromatography, and performing column chromatography by using a solvent with a volume ratio of 8: 2 and 7: 3, eluting the mixed solution of petroleum ether and ethyl acetate to obtain a crude component 1 and a crude component 2;

s3, mixing the crude component 1 and the crude component 2 obtained in the step S2, and treating the mixture by a silica gel column at a volume ratio of 9: 1, eluting with a mixed solution of petroleum ether and ethyl acetate, and passing through a gel column by using a solvent with a volume ratio of 2: 1: eluting with mixed solution of petroleum ether, chloroform and methanol to obtain the seaweed functional oil.

Preferably, the filler of the reduced pressure column in the step S2 is 100-200 mesh silica gel.

Preferably, the filler of the silica gel column in the step S3 is 200-300 mesh silica gel.

Preferably, the filler of the gel column of step S3 is sephadex.

Preferably, the volume ratio of methanol to chloroform in step S1 is 1: 1 to 3.

More preferably, the volume ratio of methanol to chloroform in step S1 is 1: 2.

in addition, the application of the seaweed functional oil in preparing the medicine for treating the Alzheimer's disease and the product for preventing and/or assisting in improving the Alzheimer's disease are both within the protection scope of the invention.

Preferably, the product is a medicament, food or health product.

Preferably, the use is in the manufacture of a medicament for inhibiting acetylcholinesterase activity, in the manufacture of a medicament for scavenging DPPH free radical activity, and in the manufacture of a medicament for inhibiting the production of NO and ROS levels in a cell.

The invention has the following beneficial effects:

the invention provides seaweed functional oil and application thereof. Experiments prove that the seaweed functional oil provided by the invention has obvious acetylcholinesterase activity inhibition, antioxidant activity and anti-inflammatory activity, specifically has the functions of inhibiting the activity of acetylcholinesterase, eliminating the activity of DPPH free radicals, inhibiting the generation of NO induced by inflammatory factors in cells and eliminating the ROS level in the cells, has NO toxic or side effect on normal cells, and can be used as a medicine which has high activity, high yield, NO toxicity and natural source and inhibits the activity of acetylcholinesterase;

in addition, the preparation method of the seaweed functional oil has high yield, the seaweed functional oil takes natural seaweed and sargassum fusiforme as raw materials, the material source is natural and wide, the obtained seaweed functional oil has no toxicity, adverse reaction can not be caused, and the method for preparing the seaweed functional oil is simple and convenient and has low cost; therefore, the seaweed functional oil provided by the invention has good application prospects in preparation of medicines for treating Alzheimer's disease and products for preventing and/or assisting in improving Alzheimer's disease.

Drawings

FIG. 1 is a graph showing the results of tests on the main components of the functional oil of seaweeds of the present invention; wherein, A is the gas chromatogram of the non-methyl esterification of the seaweed functional oil; b is a gas chromatogram of the methyl esterification of the seaweed functional oil.

FIG. 2 is a graph showing the results of the acetylcholinesterase inhibition test using the algal oil of the present invention.

FIG. 3 is a graph showing the results of the antioxidant property test of the functional oil of seaweed of the present invention.

FIG. 4 is a graph showing the results of the toxicity test of the algal functional oils of the present invention to cells.

FIG. 5 is a graph showing the results of the activity test of the algal functional oils of the present invention for inhibiting the production of NO in cells.

FIG. 6 is a graph showing the results of the inhibition of LPS-induced ROS in BV-2 cells by algal functional oil of the present invention.

FIG. 7 is a graph showing the results of the activity test of the algal functional oils of the present invention to inhibit intracellular ROS levels.

Detailed Description

The present invention is further illustrated by the following specific examples, which are not intended to limit the invention in any way. Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated.

Unless otherwise indicated, reagents and materials used in the following examples are commercially available.