Method for preparing dha in phospholipid form and microencapsulation method

Abstract

A method for preparing phospholipid type DHA, comprising the following steps: (1) hydrolysis of phospholipid to obtain hydrolyzed phospholipid; (2) preparation of DHA ethyl ester: slowly adding NaOH-anhydrous ethanol mixed solution in which NaOH has been completely dissolved into DHA oil to react, removing ethanol and water in the reaction product to obtain fatty acid ethyl ester; molecular distillation of the fatty acid ethyl ester to obtain DHA ethyl ester; (3) immobilized lipase transesterification reaction: mixing DHA ethyl ester with hydrolyzed phospholipid, and then adding into an immobilized enzyme column filled with immobilized lipase to react; (4) adding the reaction product obtained in step (3) into a winterization tank to winterize, and filtering to obtain phospholipid type DHA in crystalline state; (5) molecular distillation of the phospholipid type DHA in crystalline state obtained in step (4) after warming, and the heavy phase is the phospholipid type DHA. The method for preparing phospholipid type DHA is beneficial to improving the content of DHA in the phospholipid type DHA product, and can increase the oxidation stability of the phospholipid type DHA through microcapsule embedding, and prolong the shelf life.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of oil modification, and particularly relates to a preparation method of phospholipid type DHA and a microcapsule embedding method of phospholipid type DHA. BACKGROUND

[0002] Phospholipid type DHA is referred to as DHA when the R1 or R2 fatty acyl group in the phospholipid component contains DHA. Studies have shown that phospholipid type DHA has a variety of biological activities on the basis of the respective functional characteristics of phospholipids and DHA. Phospholipid type DHA has the advantages and functions of enhancing the antioxidant property of DHA, facilitating the storage of DHA, facilitating the absorption of DHA, enhancing the permeability of cell membranes, enhancing the stress resistance of organisms, anti-inflammatory effect, etc. DHA and phospholipids are important components and nutrients of the brain and nervous system, and can promote the normal development of the brain and nervous system and guarantee the normal function of the nervous system. In the prevention and treatment of diseases such as coronary heart disease, inflammation, autoimmune disease, diabetes and Alzheimer's syndrome, as well as in promoting the physiological functions such as the brain and visual development of infants, there are obvious advantages, and phospholipid type DHA has important nutritional value and broad application prospects in the fields of food, nutrition, medicine and health care products.

[0003] At present, the methods for obtaining phospholipid type DHA mainly include natural extraction, chemical synthesis, microbial fermentation and enzyme method.

[0004] Natural phospholipid type DHA mainly comes from marine resources such as marine krill and deep-sea fish seeds, and is separated and produced by methods such as organic extraction, column chromatography, complex salt precipitation, supercritical extraction and membrane separation. The natural source phospholipid type DHA faces the problems of raw material exhaustion and being not conducive to sustainable development, the raw material source is easy to fluctuate and is easy to be contaminated by heavy metals, the phospholipid type DHA content is low, and the purification cost is high.

[0005] Phospholipid type DHA is prepared by chemical catalysts, the reaction is violent, there are many by-products, and the product has safety hazards, and a chemical catalyst system with mild conditions, low toxicity and good catalytic effect still needs to be further developed.

[0006] Enzymatic catalysis has the advantages of position specificity, can select a specific site for reaction, has few by-products, mild reaction conditions and environmental friendliness, and is the preferred method for artificial synthesis of phospholipid type DHA. However, the existing enzyme catalysis directly uses phospholipids and DHA oil to carry out ester exchange reaction, and has the problem of low insertion rate of DHA, and the obtained phospholipid type DHA product has low DHA content.

[0007] The DHA in the molecular structure of the phospholipid type DHA contains unsaturated double bonds, is sensitive to light and heat, is easily oxidized, is easily affected by external factors during processing, storage and transportation, is oxidized and deteriorated, produces bad smell, and causes product quality to decrease and shelf life to be shortened. Therefore, how to protect the phospholipid type DHA and make it not easy to deteriorate and prolong the shelf life is also a key problem to be solved in the production of the phospholipid type DHA. SUMMARY

[0008] The technical problem to be solved by the present application is to provide a preparation method and a microcapsule embedding method of the phospholipid type DHA, the preparation method is beneficial to improve the content of DHA in the phospholipid type DHA product, and the oxidation stability of the phospholipid type DHA can be increased and the shelf life thereof can be prolonged through the microcapsule embedding.

[0009] A preparation method of the phospholipid type DHA, characterized by comprising the following steps:

[0010] (1) phospholipid hydrolysis

[0011] The phospholipid is subjected to a hydrolysis reaction to obtain hydrolyzed phospholipid;

[0012] (2) preparation of DHA ethyl ester

[0013] According to the mass ratio of 1:5-1:8 of the DHA oil and ethanol, the DHA oil is first added to a reaction tank and preheated to 50-70 DEG C, then the NaOH-anhydrous ethanol mixed solution in which NaOH is completely dissolved is slowly added to the DHA oil, and the reaction is carried out at 70-80 DEG C for 1-2 hours under a closed condition;

[0014] The ethanol and water in the reaction product are evaporated by a thin film evaporator at 65-75 DEG C to obtain fatty acid ethyl ester;

[0015] The obtained fatty acid ethyl ester is subjected to molecular distillation under the condition that the vacuum degree is less than 10 Pa and the temperature is 120-150 DEG C, and the separated heavy phase is subjected to molecular distillation for 2-3 times to obtain DHA ethyl ester;

[0016] (3) immobilized lipase transesterification reaction

[0017] The DHA ethyl ester obtained in step (2) and the hydrolyzed phospholipid obtained in step (1) are mixed in a molar ratio of 3-6:1, and then added to an immobilized enzyme column filled with immobilized lipase, the enzyme amount of the immobilized enzyme column is 5-20% of the total mass of the DHA ethyl ester and the hydrolyzed phospholipid, and the reaction is carried out for 8-16 hours;

[0018] (4) winterization

[0019] The reaction product obtained in step (3) is added to a winterization tank, the stirring speed is 30-50 rpm, the temperature is lowered by 1-3 °C per hour, and after the temperature is lowered to 4-8 °C, the temperature is kept for 12-24 hours, and then filtration is performed to obtain phospholipid DHA in a crystalline state;

[0020] (5) Molecular distillation

[0021] The phospholipid DHA obtained in step (4) is melted after being warmed, and then molecular distillation is performed, and the heavy phase is phospholipid DHA.

[0022] In step (1), the role of phospholipid hydrolysis is to remove part or all of the fatty acid acyl groups in positions 1 and 2 of the phospholipid.

[0023] Preferably, in step (1), the method for obtaining hydrolyzed phospholipids by hydrolyzing the phospholipids is as follows: 20-40% of purified water by mass of the phospholipids is added to the phospholipids, and the mixture is uniformly mixed to obtain a phospholipid-purified water mixture; then the phospholipid-purified water mixture is added to an immobilized enzyme column filled with immobilized enzymes, the amount of enzyme added to the immobilized enzyme column is 5-20% of the phospholipid-purified water mixture, and the reaction is carried out at 50-55 °C for 6-12 hours; after the reaction is completed, the water phase is extracted with 2-5 times the volume of n-hexane for 2-4 times, and the water phase is evaporated at 65-75 °C using a thin film evaporator to obtain hydrolyzed phospholipids. In step (1), n-hexane is used for extraction, mainly to extract the fatty acids generated by enzymatic hydrolysis.

[0024] More preferably, the immobilized enzyme used in step (1) is one or a combination of several of phospholipase A1, phospholipase A2, and lipase.

[0025] Preferably, in step (2), nitrogen gas is filled in the reaction tank during the entire reaction process (after the DHA oil is added to the reaction tank, the inside of the reaction tank is vacuumed and then filled with nitrogen gas), which can effectively avoid the oxidation of DHA.

[0026] Preferably, in step (2), the mass ratio of anhydrous ethanol to sodium hydroxide in the NaOH-anhydrous ethanol mixture is 40-60:1, and the NaOH is dissolved in the anhydrous ethanol.

[0027] Preferably, in step (2), the NaOH-anhydrous ethanol mixture is added completely within 60-120 minutes. Usually, the NaOH-anhydrous ethanol mixture is added at a uniform speed.

[0028] In step (2), the mass ratio of DHA oil to ethanol is 1:5-1:8, and the ethanol refers to the ethanol contained in the NaOH-anhydrous ethanol mixture.

[0029] In step (2), the heavy phase obtained by molecular distillation of the fatty acid ethyl ester is DHA ethyl ester; and the light phase mainly contains glycerol, ethyl palmitate, and ethyl oleate.

[0030] In step (3), the inside of the immobilized enzyme column is filled with nitrogen gas during the whole enzymolysis process (after the inside of the immobilized enzyme column is vacuumed and then filled with nitrogen gas), which can effectively avoid the oxidation of DHA.

[0031] In step (3), the immobilized lipase used is one or a combination of several of phospholipase A1, phospholipase A2 and lipase.

[0032] In step (4), the filter is a plate-frame filter.

[0033] Through a large number of experiments, it is concluded that the product of the immobilized lipase transesterification reaction is in a liquid state at a temperature of 4-8℃, and the unreacted DHA ethyl ester and the generated fatty acid ethyl ester are in a liquid state, and the phospholipid type DHA is in a crystalline state, so the phospholipid type DHA can be separated by filtration. The filtrate can be subjected to the molecular distillation method of step (2) to obtain DHA ethyl ester, which can be reused.

[0034] In step (5), the main function of molecular distillation is to remove the remaining fatty acid ethyl ester and to remove the odor.

[0035] In step (5), the working conditions of molecular distillation are as follows: the vacuum degree is less than 10 Pa, the oil inlet amount is 10-20 L / h, and the temperature of the heat conducting oil is 220-260℃; and the distillation is performed for 2-3 times.

[0036] The application also provides a microencapsulation method of the phospholipid type DHA prepared by the above method, which is characterized by comprising the following steps:

[0037] (1') Preparation of a mixed liquid

[0038] The following materials are prepared by weight: 15-30% of glucose syrup, 20-50% of modified starch, 3-6% of β-cyclodextrin, 2-4% of carrageenan, 1-3% of glycerol monostearate, 1-3% of sodium ascorbate, and the rest is the phospholipid type DHA prepared by the above method for preparing the phospholipid type DHA;

[0039] Then, the prepared glucose syrup, modified starch, β-cyclodextrin, carrageenan, glycerol monostearate, sodium ascorbate and phospholipid type DHA are uniformly stirred to obtain a mixed liquid;

[0040] (2') Spray drying

[0041] The mixed liquid obtained in step (1') is subjected to homogenization treatment and then spray dried to obtain the required phospholipid type DHA microcapsule powder.

[0042] The modified starch is obtained by modifying starch and has the advantages of good film forming property, low viscosity, easy atomization, easy dehydration and drying, less wall sticking, etc., and can effectively protect the core material phospholipid type DHA.

[0043] Beta-cyclodextrin (referred to as beta-CD) is a group of cyclic dextrin produced by cyclodextrin glucanotransferase acting on starch, which is formed by glucose with 1,4-glycosidic bond to form a chair conformation of cyclic oligomer. In the structure, the glucose monomer forms a closed ring type molecule without reducing group, forms a cavity with hydrophobic center and hydrophilic surface, and can interact with organic molecules to form inclusion complex to achieve effective embedding. Beta-CD can form more effective protection for the core material, can well prevent air from entering, and is very suitable for embedding in functional oil. The oil and beta-CD are closely combined in the cavity, which can greatly slow down the reaction process under the action of light, heat and oxygen, thereby improving the stability of the oil.

[0044] Carrageenan has strong stability, high solubility in hot water, good fluidity, and good coagulation property, and has excellent oxygen barrier ability.

[0045] Preferably, in step (2'), the mixed liquid is subjected to homogenization treatment under a pressure of 20-40 MPa, and is homogenized 1-3 times, each time for 20-30 min.

[0046] Preferably, in step (2'), after homogenization treatment, the mixed liquid is sterilized at 85-90 DEG C for 20-30 min, and then is subjected to spray drying.

[0047] Preferably, in step (2'), pressure spray drying machine is used for spray drying, the spray pressure is 10-20 MPa, the feeding temperature is 45-65 DEG C, the feeding pump rotation speed is 10-30 rpm, the air inlet temperature is 150-180 DEG C, and the air outlet temperature is 70-85 DEG C.

[0048] The present application has the following technical advantages and effects:

[0049] 1. The present application first hydrolyzes phospholipid to obtain hydrolyzed phospholipid, removes part or all of the fatty acid acyl groups in positions 1 and 2 of the phospholipid, thereby improving the low insertion rate of DHA in the phospholipid esterification reaction, i.e. increasing the insertion rate of DHA in the preparation of DHA ethyl ester reaction in step (2).

[0050] 2. The esterification reaction of the present application uses an immobilized enzyme column, the immobilized lipase is easy to separate from the reaction product, can improve the stability of the enzyme, effectively reduce the loss of immobilized lipase, can be reused, realizes the minimization of the loss of immobilized lipase and the maximization of reuse of enzyme activity, can greatly improve the reaction efficiency, the use frequency of immobilized lipase can reach more than 30 times, greatly reduces the cost of using enzyme, and the utilization rate of fatty acid can reach more than 90%. The immobilized enzyme can also effectively improve its catalytic activity and stereoselectivity, thereby increasing the yield of the product and improving the quality of the product. At the same time, the phospholipid has poor flowability, and the immobilized enzyme column can be pressurized to increase the flow rate, which is beneficial to improve the reaction efficiency.

[0051] 3. The present application uses molecular distillation for separation, high vacuum, reduces the oxidation of DHA at high temperature. The present application uses molecular distillation technology to prepare phospholipid type DHA, the distillation temperature of molecular distillation is low, the working vacuum degree is high, and the heating time of the material is short. Not only the separation efficiency is high, the separation cost is low, but also it is beneficial to protect the phospholipid type DHA.

[0052] 4. Winterization and molecular distillation are both pure physical methods of separation, without chemical reagents, especially organic solvent pollution, green and environmental protection.

[0053] 5. The phospholipid type DHA is oily, which limits its application in solid beverages, liquid beverages and the like. The microencapsulated phospholipid type DHA is microencapsulated, which not only increases the oxidation stability of the phospholipid type DHA, but also expands the application range of the phospholipid type DHA.

[0054] The microencapsulation embedding technology can prolong and control the release of active substances in the microcapsule, improve the stability of the product and prolong the shelf life of the product, so that it is not affected by the environment. The insoluble oil is isolated to change it from liquid oil to powder, and the appearance of the product is also changed to make it easy to carry and transport. The phospholipid type DHA is coated by the microencapsulation embedding method of the present application, which not only improves the oxidation characteristics of DHA, improves the stability of the phospholipid type DHA, and prolongs the shelf life, but also can mask the fishy smell of the phospholipid type DHA to a certain extent, enhances the acceptability of the phospholipid type DHA, and expands the market application field of the phospholipid type DHA. DETAILED DESCRIPTION

[0055] Example 1

[0056] In this embodiment, the preparation method of the phospholipid type DHA includes the following steps:

[0057] (1) Phospholipid hydrolysis

[0058] The phospholipid is mixed with purified water in a mass ratio of 30% to obtain a phospholipid-purified water mixture; then the phospholipid-purified water mixture is added to an immobilized enzyme column filled with immobilized enzymes, the amount of enzyme added to the immobilized enzyme column is 15% of the mass of the phospholipid-purified water mixture, and the reaction is carried out at 55°C for 8 hours; after the reaction is completed, the water phase is extracted with 2 volumes of n-hexane 4 times, and the water phase is evaporated at 75°C using a thin film evaporator to obtain hydrolyzed phospholipids;

[0059] The immobilized enzyme used in step (1) is a combination of phospholipase A1 and lipase 435 (the mass ratio of phospholipase A1 to lipase 435 is 1:1);

[0060] (2) Preparation of DHA ethyl ester

[0061] According to the mass ratio of DHA oil to ethanol 1:7, first add DHA oil to the reaction tank and preheat to 70°C, then slowly add the NaOH-anhydrous ethanol mixture in which NaOH is completely dissolved to the DHA oil (in the NaOH-anhydrous ethanol mixture, the mass ratio of anhydrous ethanol to sodium hydroxide is 50:1, and NaOH is dissolved in anhydrous ethanol; the NaOH-anhydrous ethanol mixture is added at a constant speed within 80 minutes), and the reaction is carried out under closed conditions at 75°C for 2 hours; during the entire reaction process, the inside of the reaction tank is filled with nitrogen (after adding DHA oil to the reaction tank, the inside of the reaction tank is vacuumed and then filled with nitrogen);

[0062] Evaporate ethanol and water in the reaction product at 65°C using a thin film evaporator to obtain fatty acid ethyl ester;

[0063] Molecularly distill the obtained fatty acid ethyl ester under the conditions of a vacuum degree of 1 Pa and a temperature of 125°C, and molecularly distill the separated heavy phase 3 times to obtain DHA ethyl ester (the DHA content is 81.5% after detection);

[0064] (3) Immobilized lipase transesterification reaction

[0065] Mix the DHA ethyl ester obtained in step (2) with the hydrolyzed phospholipids obtained in step (1) at a molar ratio of 5:1, and then add them to an immobilized enzyme column filled with immobilized lipase, the amount of enzyme added to the immobilized enzyme column is 10% of the total mass of DHA ethyl ester and hydrolyzed phospholipids, and the reaction is carried out for 16 hours; during the entire enzymatic hydrolysis process, the inside of the immobilized enzyme column is filled with nitrogen (the inside of the immobilized enzyme column is vacuumed and then filled with nitrogen);

[0066] The immobilized lipase used in step (3) is a combination of phospholipase A1 and lipase 435 (the mass ratio of phospholipase A1 to lipase 435 is 1:1);

[0067] (4) Winterization

[0068] The reaction product obtained in step (3) is added to a winterization tank, the stirring speed is 30 rpm, the temperature is lowered by 2°C per hour, and after the temperature is lowered to 6°C, the temperature is kept for 12 hours, and filtration (using plate and frame filtration) is performed to obtain phospholipid type DHA in a crystalline state;

[0069] (5) Molecular distillation

[0070] The phospholipid type DHA obtained in step (4) is warmed and melted, and then subjected to molecular distillation, and the heavy phase is phospholipid type DHA (the DHA content is 38.5%, and the phospholipid content is 96.3%).

[0071] In step (5), the working conditions of the molecular distillation are as follows: the vacuum degree is less than 0.1 Pa, the oil inlet amount is 20 L / h, and the temperature of the heat conducting oil is 250°C; and the distillation is performed for 3 times.

[0072] In the embodiment, the microencapsulation method of the phospholipid type DHA prepared by the above method includes the following steps:

[0073] (1') Preparation of mixed liquid

[0074] The following materials are prepared by weight: 18% glucose syrup, 26% modified starch, 4% β-cyclodextrin, 2% carrageenan, 1% glycerol monostearate, 1% sodium ascorbate, and the balance is the phospholipid type DHA prepared by the above method for preparing phospholipid type DHA;

[0075] Then, the prepared glucose syrup, modified starch, β-cyclodextrin, carrageenan, glycerol monostearate, sodium ascorbate and phospholipid type DHA are stirred uniformly to obtain a mixed liquid;

[0076] (2') Spray drying

[0077] The mixed liquid obtained in step (1') is subjected to homogenization treatment and then spray dried to obtain the required phospholipid type DHA microcapsule powder.

[0078] In the above step (2'), the mixed liquid is subjected to homogenization treatment at a pressure of 40 MPa, and the homogenization is performed for 2 times, each time for 25 min; after the homogenization treatment, the mixed liquid is sterilized at 85°C for 30 min, and then subjected to spray drying (pressure spray drying machine is used for spray drying, the spray pressure is 20 MPa, the feed temperature is 60°C, the feed pump rotation speed is 30 rpm, the air inlet temperature is 170°C, and the air outlet temperature is 75°C).

[0079] Example 2

[0080] In the embodiment, the method for preparing phospholipid type DHA includes the following steps:

[0081] (1) Hydrolysis of phospholipids

[0082] The phospholipid is mixed with purified water in a mass ratio of 35% to obtain a phospholipid-purified water mixture; then the phospholipid-purified water mixture is added to an immobilized enzyme column filled with immobilized enzymes, the amount of enzyme added to the immobilized enzyme column is 10% of the mass of the phospholipid-purified water mixture, and the reaction is carried out at 55°C for 8 hours; after the reaction is completed, the water phase is extracted twice with 3 times the volume of n-hexane, and the water phase is evaporated at 70°C using a thin film evaporator to obtain hydrolyzed phospholipids;

[0083] The immobilized enzyme used in step (1) is a combination of phospholipase A1 and lipase 435 (the mass ratio of phospholipase A1 to lipase 435 is 1:2);

[0084] (2) Preparation of DHA ethyl ester

[0085] According to the mass ratio of DHA oil to ethanol of 1:6, first add DHA oil to the reaction tank and preheat to 70°C, then slowly add the NaOH-anhydrous ethanol mixture in which NaOH is completely dissolved to the DHA oil (in the NaOH-anhydrous ethanol mixture, the mass ratio of anhydrous ethanol to sodium hydroxide is 60:1, and NaOH is dissolved in anhydrous ethanol; the NaOH-anhydrous ethanol mixture is added at a constant speed within 100 minutes), and the reaction is carried out under closed conditions at 75°C for 2 hours; during the entire reaction process, the inside of the reaction tank is filled with nitrogen (after the DHA oil is added to the reaction tank, the inside of the reaction tank is vacuumed and then filled with nitrogen);

[0086] Evaporate the ethanol and water in the reaction product at 65°C using a thin film evaporator to obtain fatty acid ethyl ester;

[0087] Molecularly distill the obtained fatty acid ethyl ester under the conditions of a vacuum degree of 1 Pa and a temperature of 120°C, and molecularly distill the separated heavy phase for 3 times to obtain DHA ethyl ester (the DHA content is 80.5% after detection);

[0088] (3) Immobilized lipase transesterification reaction

[0089] Mix the DHA ethyl ester obtained in step (2) with the hydrolyzed phospholipids obtained in step (1) at a molar ratio of 4:1, and then add them to an immobilized enzyme column filled with immobilized lipase, the amount of enzyme added to the immobilized enzyme column is 10% of the total mass of DHA ethyl ester and hydrolyzed phospholipids, and the reaction is carried out for 16 hours; during the entire enzymatic hydrolysis process, the inside of the immobilized enzyme column is filled with nitrogen (the inside of the immobilized enzyme column is vacuumed and then filled with nitrogen);

[0090] The immobilized lipase used in step (3) is a combination of phospholipase A1 and lipase 435 (the mass ratio of phospholipase A1 to lipase 435 is 1:2);

[0091] (4) Winterization

[0092] The reaction product obtained in step (3) is added to a winterization tank, the stirring speed is 30 rpm, the temperature is lowered by 2℃ per hour, and after the temperature is lowered to 4℃, the temperature is kept for 12 hours, and filtration (using plate and frame filtration) is performed to obtain phospholipid type DHA in a crystalline state;

[0093] (5) Molecular distillation

[0094] After the crystalline state phospholipid type DHA obtained in step (4) is warmed and melted, molecular distillation is performed, and the heavy phase is phospholipid type DHA (the DHA content is 37.6%, and the phospholipid content is 96.6%).

[0095] In step (5), the working conditions of molecular distillation are as follows: the vacuum degree is 0.1 Pa, the oil inlet amount is 10 L / h, and the temperature of the heat conducting oil is 260℃; and the distillation is performed twice.

[0096] In the embodiment, the microencapsulation method of the phospholipid type DHA prepared by the above method includes the following steps:

[0097] (1') Preparation of mixed liquid

[0098] The following materials are prepared by weight: 16% of glucose syrup, 28% of modified starch, 3% of β-cyclodextrin, 2% of carrageenan, 2% of glycerol monostearate, 1% of sodium ascorbate, and the balance is the phospholipid type DHA prepared by the above method for preparing phospholipid type DHA;

[0099] Then, the prepared glucose syrup, modified starch, β-cyclodextrin, carrageenan, glycerol monostearate, sodium ascorbate and phospholipid type DHA are stirred uniformly to obtain a mixed liquid;

[0100] (2') Spray drying

[0101] After the mixed liquid obtained in step (1') is subjected to homogenization treatment, spray drying is performed to obtain the required phospholipid type DHA microcapsule powder.

[0102] In the above step (2'), the mixed liquid is subjected to homogenization treatment under a pressure of 40 MPa, and the homogenization is performed for 3 times, each time for 20 min; after the homogenization treatment, the mixed liquid is sterilized at 85℃ for 30 min, and then spray drying is performed (pressure spray drying machine is used for spray drying, the spray pressure is 20 MPa, the feed temperature is 65℃, the feed pump rotation speed is 30 rpm, the air inlet temperature is 160℃, and the air outlet temperature is 75℃).

[0103] Phospholipid content detection: GB / T 5537-2008 Grain and oil inspection Determination of phospholipid content.

[0104] DHA content detection: The DHA content is determined according to the method specified in GB 5009.168-2016 National food safety standard Determination of fatty acids in food.

[0105] Test Example

[0106] I. SUMMARY

[0107] DHA (docosahexaenoic acid) is a polyunsaturated fatty acid, which is easily oxidized and deteriorated. Once oxidized and deteriorated, it will produce rancid smell, affecting the product quality and flavor. DHA microcapsule powder is obtained by embedding DHA oil through microcapsule technology, so as to isolate air and prolong the oxidation time of DHA oil. However, once the embedded DHA oil is oxidized and deteriorated, it will also produce rancid smell. Therefore, the anti-oxidation performance of DHA microcapsule powder can reflect the embedding index and stability of the microcapsule powder.

[0108] In the oil and fat industry, the Schaat oven method is usually used for anti-oxidation performance test to accelerate the destructive test of oil and fat, study the stability of oil and fat in application, and thus reflect the shelf life of oil and fat. According to the research on the anti-oxidation performance of ascorbyl palmitate in different oil products in China Oil and Fat, 2000, Vol. 25, No. 3 (empirical formula).

[0109] Temperature and shelf life coefficient relationship

[0110] Temperature °C 62 52 42 32 22 12 Shelf life factor 1 2 4 8 16 32

[0111] Using the Schaat oven method for anti-oxidation test, one day of experiment in a 62°C constant temperature oven is equivalent to one month of storage at 15°C. According to the edible oil anti-oxidation performance test method in the oil and fat industry, the accelerated destructive anti-oxidation stability test of DHA microcapsule powder is carried out to evaluate the product quality and shelf life.

[0112] Odor has always been the most intuitive way of identifying food quality, and is an important aspect of the food quality index system. Peroxide value can directly reflect the embedding effect of microcapsule powder, and is an important index of the stability of microcapsule powder.

[0113] II. SAMPLES

[0114] Phospholipid type DHA microcapsule powder of Example 1; phospholipid type DHA microcapsule powder of Example 2.

[0115] III. TEST METHOD

[0116] The phospholipid type DHA microcapsule powder prepared in Examples 1 and 2 was respectively packaged in aluminum foil bags (the phospholipid type DHA microcapsule powder prepared in Examples 1 and 2 was respectively packaged in 6 bags), vacuum sealed, and placed in an oven at 62°C. Every certain time (for example, 4 days), one sample was taken out, placed in a clean odorless container, smelled, then gargled with warm water, tasted, compared with the control sample not placed in the oven, tested for odor change, identified whether rancid odor was produced, and detected the content and peroxide value.

[0117] Detection method:

[0118] DHA content: determined according to the method specified in GB 5009.168-2016 Food Safety National Standard Determination of Fatty Acids in Food.

[0119] Peroxide value: determined according to the method specified in GB 5009.227-2016 Food Safety National Standard Determination of Peroxide Value in Food.

[0120] Four, test results

[0121] 1. Odor judgment standard: no obvious fishy odor, 0; slight fishy odor, 1; obvious fishy odor, 2; and heavy fishy odor, 3. The comprehensive evaluation score of 5 evaluators was taken as the odor judgment standard of the sample.

[0122] 2. Results: see the following table

[0123]

[0124] Five, result analysis

[0125] The phospholipid type DHA microcapsule powder of Examples 1 and 2 had no obvious change in odor at 62°C for 24 days. The DHA content of the phospholipid type DHA microcapsule powder had no obvious change. Although the peroxide value had a trend of increasing during storage, the peroxide value was still controlled within 5 meq / kg at the 24th day.

[0126] Six, conclusion

[0127] The phospholipid type DHA microcapsule powder of Examples 1 and 2 had no obvious change in odor and DHA content at 62°C for 24 days, and the peroxide value was still controlled within the standard requirement range. According to the experience formula calculation, one day of storage at 62°C is equivalent to one month of storage at 15°C. It can be inferred that the DHA microcapsule powder obtained according to the process of the application has a shelf life of at least 24 months.

Claims

1. A method for preparing phospholipid-type DHA, characterized in that... Includes the following steps: (1) Phospholipid hydrolysis Phospholipids undergo hydrolysis to obtain hydrolyzed phospholipids; The method for hydrolyzing phospholipids to obtain hydrolyzed phospholipids is as follows: Add 20-40% purified water (by weight of phospholipids) to the phospholipids and mix thoroughly to obtain a phospholipid-purified water mixture. Then, add the phospholipid-purified water mixture to an immobilized enzyme column filled with immobilized enzymes. The amount of enzyme added to the immobilized enzyme column is 5-20% of the phospholipid-purified water mixture. React at 50-55℃ for 6-12 hours. After the reaction, extract 2-4 times with 2-5 times the volume of n-hexane. Evaporate the water from the aqueous phase using a thin-film evaporator at 65-75℃ to obtain the hydrolyzed phospholipids. The immobilized enzyme used in step (1) is a combination of phospholipase A1 and lipase 435; (2) Preparation of DHA ethyl ester According to the mass ratio of DHA oil to ethanol of 1:5-1:8, first add DHA oil to the reaction vessel and preheat it to 50-70℃. Then slowly add the NaOH-anhydrous ethanol mixture with completely dissolved NaOH to the DHA oil and react for 1-2 hours under sealed conditions at 70-80℃. Ethanol and water in the reaction products were evaporated at 65-75℃ using a thin-film evaporator to obtain fatty acid ethyl esters. The obtained fatty acid ethyl esters were molecularly distilled under a vacuum of less than 10 Pa and a temperature of 120-150 °C, and the separated heavy phase was molecularly distilled 2-3 times to obtain DHA ethyl esters. (3) Immobilized lipase transesterification reaction The DHA ethyl ester obtained in step (2) and the hydrolyzed phospholipid obtained in step (1) are mixed in a molar ratio of 3-6:1 and then added to an immobilized enzyme column filled with immobilized lipase. The amount of enzyme added to the immobilized enzyme column is 5-20% of the total mass of DHA ethyl ester and hydrolyzed phospholipid, and the reaction is carried out for 8-16 hours. The immobilized enzyme used in step (3) is a combination of phospholipase A1 and lipase 435; (4) Winterization Add the reaction product obtained in step (3) into a winterization tank, stir at 30-50 rpm, reduce the temperature by 1-3°C per hour, keep the temperature at 4-8°C for 12-24 hours, and filter to obtain crystalline phospholipid DHA. (5) Molecular distillation After the crystalline phospholipid DHA obtained in step (4) is melted at room temperature, molecular distillation is performed, and the heavy phase is the phospholipid DHA.

2. The method for preparing phospholipid-type DHA according to claim 1, characterized in that: In step (2), the reaction vessel is filled with nitrogen gas throughout the entire reaction process; In step (2), the mass ratio of anhydrous ethanol to sodium hydroxide in the NaOH-anhydrous ethanol mixture is 40-60:1, wherein NaOH is dissolved in anhydrous ethanol; In step (2), the NaOH-anhydrous ethanol mixture is added within 60-120 minutes.

3. The method for preparing phospholipid-type DHA according to claim 1, characterized in that: In step (3), nitrogen gas is filled inside the immobilized enzyme column throughout the entire enzymatic digestion process.

4. The method for preparing phospholipid-type DHA according to claim 1, characterized in that: In step (4), plate and frame filtration is used.

5. The method for preparing phospholipid-type DHA according to claim 1, characterized in that: In step (5), the working conditions for molecular distillation are: vacuum degree less than 10 Pa, oil feed rate of 10-20 L / h, heat transfer oil temperature of 220-260 ℃; distillation 2-3 times.

6. A method for microencapsulating phospholipid-type DHA obtained by the method according to any one of claims 1-5, characterized in that... Includes the following steps: (1') Preparation of the mixed liquid The following materials are prepared by weight: 15-30% glucose syrup, 20-50% modified starch, 3-6% β-cyclodextrin, 2-4% carrageenan, 1-3% mono- and di-stearylglycerol, 1-3% sodium ascorbate, and the balance is phospholipid-type DHA prepared by the method of claim 1. Then, the prepared glucose syrup, modified starch, β-cyclodextrin, carrageenan, mono- and di-stearyl glycerides, sodium ascorbate and phospholipid DHA are stirred evenly to obtain a mixed solution; (2') Spray drying After homogenizing the mixture obtained in step (1'), spray dry it to obtain the desired phospholipid-type DHA microcapsule powder.

7. The method for microencapsulating phospholipid-type DHA obtained according to claim 6, characterized in that: In step (2'), the mixed liquid is homogenized under a pressure of 20-40 MPa, and homogenized 1-3 times, each time for 20-30 min; In step (2'), after homogenization, the mixture is sterilized at 85-90℃ for 20-30 minutes, and then spray-dried. In step (2'), a pressure spray dryer is used for spray drying. The spray pressure is 10-20 MPa, the feed temperature is 45-65℃, the feed pump speed is 10-30 rpm, the inlet air temperature is 150-180℃, and the outlet air temperature is 70-85℃.

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