阅读说明：本技术 一种磷脂酰丝氨酸的制备方法 (Preparation method of phosphatidylserine ) 是由 程彦 邓颖妍 于 2020-04-21 设计创作，主要内容包括：本发明属于食品保健品领域,具体涉及一种磷脂酰丝氨酸的制备方法。本发明提供的磷脂酰丝氨酸的制备方法,包括如下步骤：S1、将大豆卵磷脂和溶剂加入均质机中混合,得到混合液I；S2、向步骤S1所得的混合液I中加入酶液混合均匀,制得混合液II；S3、向步骤S2所得的混合液II制得加入有机溶剂萃取3～5次,制得磷脂酰丝氨酸半成品；S4、将步骤S3所得的磷脂酰丝氨酸半成品顺次通过硅胶柱和阳离子交换树脂,制得磷脂酰丝氨酸。本发明提供的磷脂酰丝氨酸的制备方法简单,易于实现工业化,制得的磷脂酰丝氨酸纯度较高,同时安全性较高,对我国磷脂工业,保健食品以及药品工业都将具有重要意义。(The invention belongs to the field of food health products, and particularly relates to a preparation method of phosphatidylserine. The preparation method of phosphatidylserine provided by the invention comprises the following steps: s1, adding soybean lecithin and a solvent into a homogenizer for mixing to obtain a mixed solution I; s2, adding the enzyme solution into the mixed solution I obtained in the step S1, and uniformly mixing to obtain a mixed solution II; s3, adding an organic solvent into the mixed liquid II obtained in the step S2, and extracting for 3-5 times to obtain a semi-finished product of phosphatidylserine; s4, making the semi-finished product of phosphatidylserine obtained in the step S3 pass through a silica gel column and cation exchange resin in sequence to prepare the phosphatidylserine. The preparation method of the phosphatidylserine provided by the invention is simple, the industrialization is easy to realize, the purity of the prepared phosphatidylserine is high, and the safety is high, so that the preparation method has important significance for the phospholipid industry, the health food industry and the pharmaceutical industry in China.)

1. A method for preparing phosphatidylserine, which comprises the following steps:

s1, adding soybean lecithin and a solvent into a homogenizer for mixing to obtain a mixed solution I;

s2, adding the enzyme solution into the mixed solution I obtained in the step S1, and uniformly mixing to obtain a mixed solution II;

s3, adding an organic solvent into the mixed liquid II obtained in the step S2, and extracting for 3-5 times to obtain a semi-finished product of phosphatidylserine;

s4, making the semi-finished product of phosphatidylserine obtained in the step S3 pass through a silica gel column and cation exchange resin in sequence to prepare the phosphatidylserine.

2. The method for preparing phosphatidylserine according to claim 1, wherein the amount of the soybean lecithin in the step S1 is 60 to 80 parts by weight, and the amount of the solvent is 260 to 400 parts by weight; the dosage of the enzyme solution in the step S2 is 70-80 parts by weight; the amount of the organic solvent used in step S3 is 300 to 800 parts by weight.

3. The method for preparing phosphatidylserine of claim 1 or 2, wherein the solvent in step S1 is deionized water, limonene, and cyclohexane-dissolved methanol solubles at a weight ratio of 20: 1-3: 5 to 12.

4. The method of claim 3, wherein the solvent used in step S1 is deionized water, limonene, and cyclohexane-soluble methanol in a weight ratio of 20: 2: 9.

5. The method for producing phosphatidylserine according to claim 1 or 2, wherein the enzyme solution in step S2 is composed of L-serine and phospholipase D at a weight ratio of 13-16: 2-6, the concentration of the L-serine is 0.15-0.2 g/ml, and the enzyme activity of the phospholipase D is 3-5U/ml.

6. The method for producing phosphatidylserine of claim 5, wherein the enzyme solution in step S2 comprises L-serine and phospholipase D at a weight ratio of 14:5, wherein the concentration of L-serine is 0.18g/ml, and the enzyme activity of phospholipase D is 4U/ml.

7. The method for producing phosphatidylserine of claim 1 or 2, wherein the organic solvent in step S3 is ethanol, methanol, and n-hexane at a weight ratio of 2:7: 3.

8. The method for preparing phosphatidylserine of claim 1, wherein the mixing temperature in the mixing process of step S2 is 25 to 30 ℃ and the mixing time is 28 to 42 hours.

9. The phosphatidylserine prepared by the method of any one of claims 1 to 8.

10. Use of phosphatidylserine prepared by the method of any one of claims 1 to 8 in the preparation of a nutraceutical, a health product, a food, a pharmaceutical, or a cosmetic.

Technical Field

The invention belongs to the field of food health products, and particularly relates to a preparation method of phosphatidylserine.

Background

Phosphatidylserine (PS) is a naturally occurring family of phospholipids, an important component of cell membrane phospholipids, and is the only phospholipid that can regulate the functional state of key proteins in cell membranes. Especially in the nervous system of human body, it is one of the important components of the cell membrane of brain, and plays an important role in regulating various functions of brain (especially the memory and emotional stability of brain), such as it can affect the fluidity and permeability of cell membrane, and activate the metabolism and synthesis of various enzymes.

Phosphatidylserine is widely present in animal and higher plant seeds and in microbial biofilms, and has important regulatory effects on many cellular metabolic processes. Animal experiments and clinical application show that phosphatidylserine has obvious curative effects on preventing and treating senile dementia, improving memory and cognitive ability, and relieving mental stress and depression. Meanwhile, phosphatidylcholine is a natural emulsifier with unique functions of emulsification, permeation, wetting, antioxidation and the like, and has wide application in the fields of medicines, health products, foods, daily chemicals, feeds, leather and the like.

Generally, animals are the main source of phosphatidylserine, and the safety of animal-derived phosphatidylserine products is questioned due to mad cow disease and the like. Attempts have been made to extract phosphatidylserine or phosphatidylcholine from natural plants, but these methods also suffer from the problems of complicated steps, low yield, and high product price. Chinese patent application CN107266492A discloses an extraction method of phosphatidylserine, wherein the extraction method comprises: 1) adding chloroform and methanol into medulla bovis Seu Bubali, mixing, and leaching to obtain mixture M1; 2) filtering the mixture M1 prepared in the step 1), taking a filtrate, adding a sodium chloride aqueous solution into the filtrate, standing, and removing a solution layer after layering to obtain a grease layer M2; 3) adding a washing solution into the grease layer M2 prepared in the step 2), mixing, and removing the solution to obtain a mixture M3; 4) separating the mixture M3 prepared in the step 3) by adopting thin layer chromatography, and collecting to obtain phosphatidylserine. The method can extract phosphatidylserine from medulla bovis Seu Bubali, and can effectively utilize medulla bovis Seu Bubali. However, the method for extracting phosphatidylserine from animal cells has potential safety hazards due to animal diseases such as mad cow disease.

In recent years, attention is paid to a method for synthesizing phosphatidylserine by a phospholipase D conversion method at home and abroad, the enzyme conversion method has the advantages of mild reaction conditions, easiness in reaction control, high efficiency, simplicity and the like, and due to rapid development of a green biomass solvent in recent years, the enzyme method has a tendency of replacing a traditional solvent and can be developed and utilized.

Therefore, the invention provides a method for preparing phosphatidylserine with relatively simple process and high purity.

Disclosure of Invention

In order to solve the problems in the prior art, the invention aims to provide a preparation method of phosphatidylserine. The phosphatidylserine prepared by the method has high purity and high safety, and the method has simple integral process, is easy to realize industrialization, and has important significance to phospholipid industry, health food and pharmaceutical industry in China.

The technical scheme of the invention is as follows:

a method for preparing phosphatidylserine, comprising:

s1, adding soybean lecithin and a solvent into a homogenizer for mixing to obtain a mixed solution I;

s2, adding the enzyme solution into the mixed solution I obtained in the step S1, and uniformly mixing to obtain a mixed solution II;

s3, adding an organic solvent into the mixed liquid II obtained in the step S2, and extracting for 3-5 times to obtain a semi-finished product of phosphatidylserine;

s4, making the semi-finished product of phosphatidylserine obtained in the step S3 pass through a silica gel column and cation exchange resin in sequence to prepare the phosphatidylserine.

Further, the soybean lecithin in the step S1 is used in an amount of 60 to 80 parts by weight, and the solvent is used in an amount of 260 to 400 parts by weight; the dosage of the enzyme solution in the step S2 is 70-60 parts by weight; the amount of the organic solvent used in step S3 is 300 to 800 parts by weight.

Further, the soybean lecithin was used in an amount of 70 parts by weight and the solvent was used in an amount of 350 parts by weight in step S1; the dosage of the enzyme solution in the step S2 is 75 weight parts; the amount of the organic solvent used in the step S3 is 600 parts by weight.

Further, the solvent in the step S1 is deionized water, limonene and cyclohexane dissolved methanol solubles in a weight ratio of 20: 1-3: 5 to 12.

Further, the solvent in step S1 is deionized water, limonene and cyclohexane dissolved methanol solubles in a weight ratio of 20: 2: 9.

Further, the enzyme solution in the step S2 is composed of L-serine and phospholipase D according to a weight ratio of 13-16: 2-6, the concentration of the L-serine is 0.15-0.2 g/ml, and the enzyme activity of the phospholipase D is 3-5U/ml.

Furthermore, the enzyme solution in the step S2 is composed of L-serine and phospholipase D according to the weight ratio of 14:5, the concentration of the L-serine is 0.18g/ml, and the enzyme activity of the phospholipase D is 4U/ml.

Further, the organic solvent in the step S3 is ethanol, methanol and n-hexane in a weight ratio of 2:7: 3.

Further, the mixing temperature in the mixing process in the step S2 is 25-30 ℃, and the mixing time is 28-42 hours.

Further, the mixing temperature in the mixing process in the step S2 is 28 ℃, and the mixing time is 35 h.

In addition, the invention also provides the phosphatidylserine prepared by the preparation method of the phosphatidylserine.

Further, another object of the present invention is to provide phosphatidylserine prepared by the method for preparing phosphatidylserine, and applications of phosphatidylserine in preparing nutraceuticals, health products, foods, medicines or cosmetics.

According to the invention, by combining physical and chemical properties of substances with the existing separation method, soybean lecithin is selected as a raw material, and phosphatidylserine is obtained by methods such as organic solvent extraction, silica gel column purification and the like, so that the phosphatidylserine can be used as a new resource food additive, and the economic added value is high; the whole production process takes 20-30 hours, the utilization value of the soybean lecithin is improved, production equipment is saved, and the production cost and time are reduced and shortened.

According to the preparation method of phosphatidylserine, a certain amount of limonene is added into the solvent in the step S2, the limonene is a monoterpene compound and has a certain lemon fragrance, can be extracted from citrus peel, and has important application in the flavor and fragrance industry. The present inventors have surprisingly found that the content of phosphatidylserine can be effectively increased when a certain amount of limonene is added into the solvent in step S2, and it can be seen from the experimental data of test example one that the content of phosphatidylserine prepared is greatly reduced when limonene is not added into the solvent in step S2.

Compared with the prior art, the phosphatidylserine prepared by the method has the following beneficial effects:

(1) the preparation method of the phosphatidylserine selects the soybean lecithin as the raw material, has the characteristic of higher safety compared with the method for obtaining the phosphatidylserine from the raw materials such as bovine brain, egg yolk and the like, has the characteristics of low cost, wide sources and short time consumption compared with an enzyme conversion synthesis method, and reduces the production cost.

(2) The preparation method of phosphatidylserine provided by the invention has simple integral process and easy realization of industrialization, and the prepared phosphatidylserine has the content of more than 90 percent and higher purity and has important significance for phospholipid industry, health-care food and pharmaceutical industry in China.

Detailed Description

The present invention is further illustrated by the following description of specific embodiments, which are not intended to limit the invention, and various modifications and improvements can be made by those skilled in the art based on the basic idea of the invention, but the invention is within the protection scope of the invention.

Wherein, the reagents used in the invention are all common reagents and can be purchased from common reagent production and sale companies.