Method for synthesizing sn-2-site triglyceride with high DHA content by catalyzing algae oil through lipase and application of sn-2-site triglyceride

Abstract

The invention belongs to the technical field of grease biological enzyme method synthesis, and particularly relates to a method for synthesizing sn-2 site triglyceride with high DHA content by catalyzing algae oil through lipase and application of the method. According to the method, DHA-containing algal oil, sunflower seed fully-hydrolyzed oil and medium-chain and long-chain fatty acid are used as raw materials, and the DHA-containing algal oil is prepared through a multi-step enzyme method. By means of the method, the content of sn-2DHA in the total DHA can be stably increased, and the increasing proportion can reach 60% or above.

Description

Technical Field

[0001] The present invention belongs to the technical field of bioenzymatic synthesis of oils and fats, and specifically relates to a method for synthesizing triglycerides with a high content of DHA at the sn-2 position by using lipase to catalyze algae oil and its application. Background Art

[0002] DHA (docosahexaenoic acid) is an important component of brain cell membranes. It participates in the formation and development of brain cells and plays an important role in the extension of nerve cell axons and the formation of new protrusions. It can maintain the normal physiological activities of nerve cells and participate in the brain's thinking and memory formation process. DHA is rich in the retina and can maintain the normal function of the retina and help prevent vision loss and eye diseases.

[0003] sn-2DHA is a special form of DHA, located at the second acyl position in the triglyceride structure, so it is called sn-2 DHA. This structure makes sn-2DHA more bioavailable and bioactive in the human body than other forms of DHA, and can be absorbed and utilized by the human body more efficiently. Due to its special structural position, sn-2DHA has higher stability, can reduce oxidative losses during human absorption, and maintain its original nutritional value. Existing methods usually use excess free fatty acids to transesterify DHA in algae oil. Since this reaction is a reversible reaction, the sn-2DHA content is unstable and the increase ratio is not large. If the transesterification reaction is excessive, the DHA at the sn-1,3 positions will be removed excessively, reducing the total DHA content and causing the product quality to decline.

[0004] Algae oil is a natural plant source of DHA, and its relative content of sn-2DHA is 35-40%, which is lower than the 50-60% in breast milk and cannot meet the needs of infants and young children. At the same time, increasing the content of sn-2DHA can also better meet the needs of adults, such as those who need health care, patients with retinal diseases, and injured people with brain damage problems. Therefore, it is necessary to study a method to increase the relative content of sn-2DHA. Summary of the invention

[0005] The purpose of the present invention is to provide a method for synthesizing triglycerides with a high content of DHA at the sn-2 position by using lipase to catalyze algae oil in view of the deficiencies of the current prior art. This is a method for synthesizing triglycerides with a high content of DHA at the sn-2 position (TAG) by combining a multi-step method with an immobilized enzyme. In this method, the content of sn-2DHA in the total DHA can be stably increased by more than 60% through multi-step reactions. The obtained triglycerides with a high content of DHA at the sn-2 position can be preferably used in food.

[0006] In order to achieve the above invention object, the technical solution of the present invention is as follows: A method for synthesizing triglycerides with a high content of DHA at the sn-2 position from algae oil using lipase catalysis, wherein the triglycerides are prepared by a multi-step enzymatic method using algae oil containing DHA, fully hydrolyzed sunflower seed oil and medium-chain fatty acids as raw materials; the method specifically comprises the following steps: Step 1: adding immobilized lipase I to algae oil containing DHA, and then adding medium-chain fatty acids, performing acid hydrolysis reaction under vacuum conditions, removing free fatty acids from the reaction product by molecular distillation, and collecting the residue TAG; Step 2: Mix the distillation residue TAG with medium-chain fatty acids, add immobilized lipase I, perform acid hydrolysis under vacuum conditions, remove free fatty acids in the secondary distillation product by molecular distillation, and collect the secondary distillation residue TAG; Step 3: adding immobilized lipase II to the secondary distillation residue TAG for hydrolysis reaction, after the reaction is completed, centrifuging the mixture to separate water to obtain an oil layer as the product, adding n-hexane and ethanol solution to the product, ultrasonically mixing, centrifuging and stratifying, collecting the lower ethanol-water phase; washing the ethanol-water phase with n-hexane, and removing ethanol and water in vacuo to obtain the third step product; Step 4: Add immobilized lipase I to the product of step 3, and add fully hydrolyzed sunflower seed oil, and carry out acid hydrolysis reaction under vacuum conditions. After the reaction, add ethanol solution to the product, mix evenly with ultrasound, centrifuge and separate, collect the lower ethanol-water phase, wash with ethanol solution, and remove ethanol and water in vacuo to obtain the final product, i.e., triglyceride with high content of DHA at the sn-2 position.

[0007] Furthermore, in the raw materials used in the method of synthesizing triglycerides with a high content of DHA in the sn-2 position by lipase-catalyzed algae oil, the mass percentage of DHA in the algae oil is ≥40%; the mass percentage of oleic acid OA in the fully hydrolyzed sunflower seed oil is 20-30%, and the mass percentage of linoleic acid LA is 50-65%; and the medium-chain fatty acids are selected from at least one of caprylic acid, capric acid, lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, and linoleic acid.

[0008] Furthermore, in the immobilized lipase used in the method of using lipase to catalyze algae oil to synthesize triglycerides with a high content of DHA at the sn-2 position, the amount of immobilized enzyme added is a percentage of the mass of the substrate.

[0009] As a better embodiment of the present application, in the first step of the method of synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algae oil, the amount of algae oil added is calculated based on the DHA content, the amount of sunflower seed fully hydrolyzed oil added is calculated based on the oleic acid and linoleic acid content, the molar ratio of DHA, oleic acid and linoleic acid is 1:2~10:3~15, preferably the molar ratio of DHA:OA:LA is 1:(4~8):(5~10), and further preferably the molar ratio is 1:(4~6):(5~8); 3~15% of immobilized lipase I is added, preferably 3~12%, and most preferably 6~12%, the acidolysis reaction temperature is 45~60°C, preferably 50~60°C; the reaction time is 1~24h, preferably 6~20h, and most preferably 6~12h. The molar ratio of DHA to medium-chain fatty acids is 1:2-10, preferably 1:2-6; more preferably 1:3-5 As a better embodiment of the present application, in the second step of the method of synthesizing triglycerides with a high content of DHA in the sn-2 position from algae oil using lipase, the distillation residue TAG is mixed with a medium-chain fatty acid, and the molar ratio of DHA to the medium-chain fatty acid in the distillation residue TAG is 1:2~10, preferably 1:2~8, and most preferably 1:3~6, and then 3~15% of immobilized lipase I is added, preferably 3~12%, and most preferably 6~12%, and the acidolysis reaction temperature is 45~60°C, preferably 50~60°C; the reaction time is 1~24h, preferably 6~20h, and most preferably 9~12h.

[0010] As a better embodiment of the present application, in the third step of the method of synthesizing triglycerides with a high content of DHA in the sn-2 position from algae oil using lipase, 3-15% of immobilized lipase II is added to the secondary distillation residue TAG, preferably 3-12%, most preferably 3-6%, the hydrolysis reaction temperature is 40-60°C, preferably 45-50°C; the reaction time is 1-48h, 12-32h, most preferably 18-24h; after the reaction is completed, n-hexane with a m / v of 1:1-10 and 80-95% ethanol solution with a m / v of 1:1-10 are added to the product; the lower ethanol aqueous phase is collected, washed 2-5 times with n-hexane with a v / v of 1:1-10, and ethanol and water are removed under vacuum at 85°C.

[0011] As a better embodiment of the present application, in the fourth step of the method for synthesizing triglycerides with a high content of DHA in the sn-2 position from algae oil using lipase to catalyze the synthesis of triglycerides, 3-15% of immobilized lipase I, preferably 3-12%, most preferably 6-12%, is added to the product of the third step, and fully hydrolyzed sunflower seed oil with an m / m ratio of 1:2-6 is added; the acid hydrolysis reaction temperature is 45-60°C, preferably 50-60°C; the reaction time is 1-12h, preferably 6-10h, most preferably 3-6h; after the reaction is completed, an 80-95% ethanol solution with an m / v ratio of 1:1-10 is added to the product; the lower ethanol aqueous phase is collected, washed 2-5 times with an 80-95% ethanol solution with a v / v ratio of 1:1-10, and the ethanol and water are removed under vacuum at 85°C.

[0012] As a better embodiment of the present application, in the method of using lipase to catalyze algal oil to synthesize triglycerides with a high content of DHA at the sn-2 position, the lipase is a sn-1,3 position specific lipase, and the amount of lipase added is 3-15% of the total amount of substrate.

[0013] As a better embodiment of the present application, in the vacuum reaction in the method of synthesizing triglycerides with a high content of DHA at the sn-2 position from algae oil using lipase, the vacuum pressure is 10~100Pa, preferably 30~50Pa.

[0014] As a better embodiment of the present application, in the method of synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algae oil, the method of removing free fatty acids is molecular distillation, and its evaporation temperature is 160°C~200°C, the rotor speed is 1000~1100rpm, and the pressure is 2~10Pa.

[0015] As a better embodiment of the present application, in the method of synthesizing triglycerides with a high content of DHA in the sn-2 position by lipase-catalyzed algae oil, the method for removing high-acid free fatty acids is an ethanol-water extraction method, which adds an 80-95% ethanol solution with a m / v of 1:1-10; collects the lower ethanol-water phase, washes it 2-5 times with an 80-95% ethanol solution with a v / v of 1:1-10, and removes residual ethanol and water under vacuum at 85°C.

[0016] As a better embodiment of the present application, in the method of synthesizing triglycerides with a high content of DHA at the sn-2 position from algae oil using lipase-catalyzed algae oil, the lipase used is an immobilized lipase I, which is a sn-1,3 selective lipase that can be used for ester exchange reaction, such as: Aspergillus niger lipase, Rhizomucor miehei lipase, Thermomyces lanuginosus lipase, Candida albicans lipase, porcine pancreatic lipase, etc.; immobilized lipase II is a sn-1,3 selective lipase that can be used for hydrolysis reaction, such as: Aspergillus niger lipase, porcine pancreatic enzyme lipase, DF Amano 15, Amano 250G, Rhizopus oryzae lipase, Rhizomucor miehei lipase, Rhizopus nivalis, etc.

[0017] The present application also protects triglycerides with a high content of DHA at the sn-2 position obtained by using any of the above methods and method step combinations.

[0018] Preferably, in the obtained triglycerides with a high content of DHA at the sn-2 position, the content of sn-2 DHA accounts for more than 60% of the total DHA content of TAG.

[0019] The present application also protects the use of the above-mentioned sn-2-position high-DHA triglyceride in food.

[0020] Furthermore, the food includes formula food, nutritional supplement or health food, etc.

[0021] Compared with the prior art, the present invention has the following beneficial effects: (1) Through multi-step reactions, the content of sn-2DHA in total DHA can be stably increased, with the increase ratio reaching more than 60%.

[0022] (2) The intermediate steps can more selectively introduce or remove a variety of fatty acids, making the final product structure esters richer in variety and more suitable for human health needs.

[0023] (3) Molecular distillation, n-hexane-ethanol-water extraction and ethanol-water extraction effectively avoid the acyl migration side reaction.

[0024] (4) Effectively remove residual free fatty acids in the product, making it easier for the product acid value to meet the standard.

[0025] (5) The use of immobilized enzymes increases the recycling value of the enzymes, greatly reduces the cost of commercial lipases, and makes the production process more economical.

[0026] (6) The process does not use any toxic or harmful solvents, and all raw materials used are food grade. The preparation method is safe, efficient, precise, and practical in industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1The gas chromatogram (GC) of triglyceride of the product obtained in Example 1, in which, starting from the right, the first peak on the right is total DHA; Figure 2 The gas chromatogram (GC) of triglyceride obtained in Example 1 shows that, starting from the right, the first peak on the right is sn-2 DHA; Figure 3 The gas chromatogram (GC) of triglycerides of the product obtained in Example 2, in which, starting from the right, the first peak on the right is total DHA; Figure 4 The gas chromatogram (GC) of triglyceride obtained in Example 2 shows that, starting from the right, the first peak on the right is sn-2 DHA; Figure 5 The gas chromatogram (GC) of triglycerides of the product obtained in Example 3, in which, starting from the right, the first peak on the right is total DHA; Figure 6 The gas chromatogram (GC) of triglyceride of the product obtained in Example 3, in which, starting from the right, the first peak on the right is sn-2 DHA; Figure 7 The gas chromatogram (GC) of triglyceride obtained by the conventional method in Comparative Example 1 is shown in FIG. 1 . In the figure, the first peak from the right is total DHA; Figure 8 The gas chromatogram (GC) of triglyceride obtained by the method of the present invention in Comparative Example 1 is shown in FIG. 1 . In the figure, the first peak from the right is total DHA; Fig. 9 The gas chromatogram (GC) of triglyceride obtained by the conventional method in Comparative Example 1 is shown in FIG. 1 . In the figure, the first peak from the right is sn-2 DHA; Fig.10 This is a gas chromatogram (GC) of triglyceride product obtained by the method of the present invention in Comparative Example 1. In the figure, the first peak from the right is sn-2 DHA. DETAILED DESCRIPTION

[0028] All features disclosed in this specification, or steps in all methods or processes disclosed, except mutually exclusive features and / or steps, can be combined in any manner.

[0029] Any feature disclosed in this specification (including claims and abstract), unless otherwise stated, may be replaced by other alternative features that are equivalent or have similar purposes. That is, unless otherwise stated, each feature is only an example of a series of equivalent or similar features.

[0030] The features and performance of the present invention are further described in detail below in conjunction with the embodiments.

[0031] The immobilized porcine pancreatic lipase used in the following examples is obtained by immobilizing commercial porcine pancreatic lipase using existing technology; the immobilized Aspergillus niger lipase is obtained by immobilizing commercial Aspergillus niger lipase using existing technology; other raw materials and reagents are all existing technology or can be obtained through existing technology.

[0032] In this application, % means mass percentage unless otherwise specified.

[0033] Embodiment 1: A method for synthesizing sn-2 high-content DHA triglycerides from algae oil using lipase catalysis, using a multi-step combined preparation method, in a solvent-free system, in a stainless steel jacketed stirred reactor. Specifically comprising the following steps: Step 1: 8% (based on the mass percentage of the substrate, the same below) of immobilized Aspergillus niger lipase was added to algae oil with a DHA content of 44.06%, and then OA was added at an m / m ratio of 1:4. Acid hydrolysis was carried out at 50°C under vacuum conditions for 9 hours. The reaction product was subjected to molecular distillation to remove the distillate FFA, and the distillate residue TAG was collected; Step 2: Mix the distillation residue TAG with new OA at an m / m ratio of 1:4, add 8% immobilized Aspergillus niger lipase, and perform acid hydrolysis at 50°C for 9 hours under vacuum conditions. Remove the secondary distillation FFA from the reaction product by molecular distillation, and collect the secondary distillation residue TAG; Step 3: Add 4% immobilized porcine pancreatic lipase to the secondary distillation residue TAG, and carry out hydrolysis reaction at 50°C for 12 hours. After the reaction, centrifuge the mixture to obtain the product. Add n-hexane with a m / v ratio of 1:2 and 85% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate the layers, collect the lower ethanol-water phase, wash it three times with n-hexane with a v / v ratio of 1:2, and remove the ethanol and water under vacuum at 85°C to obtain the third step product.

[0034] Step 4: Add 8% immobilized Aspergillus niger lipase to the product of step 3, and add sunflower seed fully hydrolyzed oil with a m / m ratio of 1:4, and perform acid hydrolysis at 50°C under vacuum for 9 hours. After the reaction, add 90% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate, collect the lower ethanol-water phase, wash it three times with a 90% ethanol solution with a v / v ratio of 1:2, and remove ethanol and water under vacuum at 85°C to obtain the product of step 4, i.e., the final product. The product was analyzed by GC. Figure 1 , Figure 2 .

[0035] Example 2 A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by using lipase to catalyze algae oil comprises the following steps: Step 1: Add 6% immobilized Aspergillus niger lipase to algae oil with a DHA content of 71.73%, then add LA at a m / m ratio of 1:2, perform acid hydrolysis at 45°C for 12 hours under vacuum conditions, remove the FFA from the reaction product by molecular distillation, and collect the residue TAG; Step 2: Mix the distillation residue TAG with new LA at an m / m ratio of 1:2, add 6% immobilized Aspergillus niger lipase, and perform acid hydrolysis at 45°C for 12 hours under vacuum conditions. Remove the secondary distillation FFA from the reaction product by molecular distillation, and collect the secondary distillation residue TAG; Step 3: Add 3% immobilized porcine pancreatic lipase to the secondary distillation residue TAG, and carry out hydrolysis reaction at 45°C for 12 hours. After the reaction, centrifuge the mixture to obtain the product. Add n-hexane with a m / v ratio of 1:2 and 85% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate the layers, collect the lower ethanol-water phase, wash it three times with n-hexane with a v / v ratio of 1:2, and remove the ethanol and water under vacuum at 85°C to obtain the third step product.

[0036] Step 4: Add 6% immobilized Aspergillus niger lipase to the product of step 3, and add sunflower seed fully hydrolyzed oil with a m / m ratio of 1:2, and perform acid hydrolysis at 45°C under vacuum for 12 hours. After the reaction, add 90% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate, collect the lower ethanol-water phase, wash it three times with 90% ethanol solution with a v / v ratio of 1:2, and remove ethanol and water under vacuum at 85°C to obtain the product of step 4, i.e., the final product. The product was analyzed by GC. Figure 3 Figure 4 .

[0037] Example 3 A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by using lipase to catalyze algae oil in a solvent-free system comprises the following steps: Step 1: Add 12% immobilized Aspergillus niger lipase to algae oil with a DHA content of 56.67%, then add n-octanoic acid at an m / m ratio of 1:5, perform acid hydrolysis at 55°C for 6 hours under vacuum conditions, remove the FFA from the reaction product by molecular distillation, and collect the residue TAG; Step 2: Mix the distillation residue TAG with new n-octanoic acid at an m / m ratio of 1:5, add 12% immobilized Aspergillus niger lipase, and perform acid hydrolysis at 55°C for 6 hours under vacuum conditions, remove the secondary distillation FFA from the reaction product by molecular distillation, and collect the secondary distillation residue TAG; Step 3: Add 6% immobilized porcine pancreatic lipase to the secondary distillation residue TAG, and carry out hydrolysis reaction at 45°C for 24 hours. After the reaction, centrifuge the mixture to obtain the product. Add n-hexane with a m / v ratio of 1:2 and 85% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate the layers, collect the lower ethanol-water phase, wash it three times with n-hexane with a v / v ratio of 1:2, and remove the ethanol and water under vacuum at 85°C to obtain the third step product.

[0038] Step 4: Add 12% immobilized Aspergillus niger lipase to the product of step 3, and add sunflower seed fully hydrolyzed oil with a m / m ratio of 1:5, and perform acid hydrolysis at 55°C for 6 hours under vacuum conditions. After the reaction, add 90% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate, collect the lower ethanol-water phase, wash it three times with a 90% ethanol solution with a v / v ratio of 1:2, and remove ethanol and water under vacuum at 85°C to obtain the product of step 4, i.e., the final product. The product was analyzed by GC. Figure 5 Figure 6 .

[0039] Comparative Example 1 The traditional method steps are as follows: Step 1: Add 12% of immobilized Aspergillus niger lipase to algae oil with a DHA content of 44.06%, then add palmitic acid with an m / m ratio of 1:5, and perform acid hydrolysis at 55°C under vacuum for 6 hours.

[0040] Step 2: The reaction product is subjected to molecular distillation to remove the evaporation FFA, and the evaporation residue TAG is collected, which is the product. The product is analyzed by GC. Figure 7 , Fig. 9 .

[0041] The method of the present invention comprises the following steps: Step 1: Add 12% immobilized Aspergillus niger lipase to algae oil with a DHA content of 44.06%, then add palmitic acid at an m / m ratio of 1:5, and perform acid hydrolysis at 55°C for 6 hours under vacuum conditions. Remove the FFA from the reaction product by molecular distillation, and collect the residue TAG; Step 2: Mix the distillation residue TAG with new LA at an m / m ratio of 1:5, add 12% immobilized Aspergillus niger lipase, and perform acid hydrolysis at 55°C for 6 hours under vacuum conditions. Remove the secondary distillation FFA from the reaction product by molecular distillation, and collect the secondary distillation residue TAG; Step 3: Add 6% immobilized porcine pancreatic lipase to the secondary distillation residue TAG, and carry out hydrolysis reaction at 45°C for 24 hours. After the reaction, centrifuge the mixture to obtain the product. Add n-hexane with a m / v ratio of 1:2 and 85% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate the layers, collect the lower ethanol-water phase, wash it three times with n-hexane with a v / v ratio of 1:2, and remove the ethanol and water under vacuum at 85°C to obtain the third step product.

[0042] Step 4: Add 12% immobilized Aspergillus niger lipase to the product of step 3, and add sunflower seed fully hydrolyzed oil with a m / m ratio of 1:5, and perform acid hydrolysis at 55°C for 6 hours under vacuum conditions. After the reaction, add 90% ethanol solution with a m / v ratio of 1:2 to the product, mix evenly with ultrasound, centrifuge and separate, collect the lower ethanol-water phase, wash it three times with a 90% ethanol solution with a v / v ratio of 1:2, and remove ethanol and water under vacuum at 85°C to obtain the product of step 4, i.e., the final product. The product was analyzed by GC. Figure 8 Fig.10 .

[0043] The above-mentioned embodiments only express the specific implementation methods of the present application, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the protection scope of the present application. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the technical solution concept of the present application, and these all belong to the protection scope of the present application.

[0044] This background section is provided to generally present the context of the invention, and the work of the presently named inventors, the work to the extent described in this background section, and aspects of this section that did not constitute prior art at the time of application are neither explicitly nor implicitly admitted to be prior art to the present invention.

Claims

1. A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by using lipase to catalyze algae oil, characterized in that: The method is prepared by a multi-step enzymatic method using algae oil containing DHA, fully hydrolyzed sunflower seed oil and medium-chain fatty acids as raw materials; specifically comprising the following steps: Step 1: adding immobilized lipase I to algae oil containing DHA, and then adding medium-chain fatty acids, performing acid hydrolysis reaction under vacuum conditions, removing free fatty acids from the reaction product by molecular distillation, and collecting the distillation residue TAG; Step 2: Mix the distillation residue TAG with medium-chain fatty acids, add immobilized lipase I, perform acid hydrolysis under vacuum conditions, remove free fatty acids in the secondary distillation product by molecular distillation, and collect the secondary distillation residue TAG; Step 3: adding immobilized lipase II to the secondary distillation residue TAG for hydrolysis reaction, after the reaction is completed, centrifuging the mixture to separate water to obtain an oil layer as the product, adding n-hexane and ethanol solution to the product, ultrasonically mixing, centrifuging and stratifying, collecting the lower ethanol-water phase; washing the ethanol-water phase with n-hexane, and removing ethanol and water in vacuo to obtain the third step product; Step 4: adding immobilized lipase I to the product of step 3, and adding fully hydrolyzed sunflower seed oil, and performing acid hydrolysis reaction under vacuum conditions. After the reaction, adding ethanol solution to the product, ultrasonically mixing evenly, centrifuging and stratifying, collecting the lower ethanol-water phase, and then washing with ethanol solution, and removing ethanol and water in vacuum to obtain the final product, i.e., triglyceride with high content of DHA at the sn-2 position; The immobilized lipase I is a sn-1,3 selective lipase for transesterification reaction, selected from Aspergillus niger lipase, Rhizomucor miehei lipase, Thermomyces lanuginosus lipase, Candida plicata lipase, and porcine pancreatic lipase; the immobilized lipase II is a sn-1,3 selective lipase for hydrolysis reaction, selected from Aspergillus niger lipase, porcine pancreatic lipase, Rhizomucor miehei lipase TL, Amano 250G, DF Amano 15, and Candida plicata lipase.

2. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: The addition amount of the immobilized lipase I and the immobilized lipase II is 3-15% of the total mass of the substrate.

3. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: Among the raw materials used, the mass percentage of DHA in algae oil is ≥40%; the mass percentage of oleic acid OA in the fully hydrolyzed sunflower seed oil is 20-30%, and the mass percentage of linoleic acid LA is 50-65%; the medium-chain fatty acids are selected from at least one of caprylic acid, capric acid, lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, and linoleic acid.

4. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: In the first step, the amount of algae oil added is calculated based on the DHA content, and the amount of sunflower seed fully hydrolyzed oil added is calculated based on the oleic acid and linoleic acid content. The molar ratio of DHA, oleic acid and linoleic acid is 1:2~10:3~15; the molar ratio of DHA and medium-chain fatty acids is 1:2~10; the acidolysis reaction temperature is 45~60℃, and the reaction time is 1~24h.

5. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: In the second step, the distillation residue TAG is mixed with a medium-chain fatty acid, the molar ratio of DHA to the medium-chain fatty acid in the distillation residue TAG is 1:2-10, and then 3-15% of immobilized lipase I is added, the acid hydrolysis reaction temperature is 45-60°C, and the reaction time is 1-24h.

6. The method of synthesizing triglycerides with a high content of DHA at the sn-2 position from algae oil using lipase as claimed in claim 1, characterized in that: In the third step, immobilized lipase II is added to the secondary distillation residue TAG, the hydrolysis reaction temperature is 45-60°C, and the reaction time is 1-48h; after the reaction is completed, n-hexane with a m / v of 1:1-10 and 80-95% ethanol solution with a m / v of 1:1-10 are added to the product; the lower ethanol-water phase is collected, washed 2-5 times with n-hexane with a v / v of 1:1-10, and ethanol and water are removed under vacuum at 85°C.

7. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: In the fourth step, immobilized lipase I is added to the product of the third step, and fully hydrolyzed sunflower seed oil with an m / m ratio of 1:2-6 is added; the acid hydrolysis reaction temperature is 45-60°C, and the reaction time is 1-12 hours; after the reaction, an 80-95% ethanol solution with an m / v ratio of 1:1-10 is added to the product; the lower ethanol-water phase is collected, washed 2-5 times with an 80-95% ethanol solution with a v / v ratio of 1:1-10, and the ethanol and water are removed under vacuum at 85°C.

8. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: The vacuum pressure in vacuum conditions is 10~100Pa.

9. The method of synthesizing triglycerides with high content of DHA at the sn-2 position by using lipase to catalyze algae oil as claimed in claim 1, characterized in that: The evaporation temperature of the molecular distillation method is 160℃~200℃, the rotor speed is 1000~1100rpm, and the pressure is 2~10Pa.

10. The sn-2-position high-DHA triglyceride obtained by the method according to any one of claims 1 to 9, characterized in that: The sn-2DHA content accounts for more than 60% of the total DHA content of TAG.

11. Use of the sn-2-position high-DHA triglyceride as claimed in claim 10 in food.

12. The use of the sn-2 high-DHA triglyceride in food according to claim 11, characterized in that: The food includes formula food, nutritional supplement or health food.

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