A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil and its application

Abstract

The present invention belongs to the technical field of enzymatic 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 catalyzing algal oil with lipase and its application. The method uses algal oil containing DHA, fully hydrolyzed sunflower oil, and medium and long-chain fatty acids as raw materials and is prepared by a multi-step enzymatic method. Through this method, the content of sn-2 DHA in the total DHA can be relatively stably increased, and the increase ratio can reach more than 60%.

Description

Technical Field

[0001] The present invention belongs to the technical field of enzymatic 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 catalyzing algal oil with lipase and its application. Background Art

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

[0003] sn-2DHA is a special form of DHA, which is located at the second acyl position in the triglyceride structure, so it is called DHA at the sn-2 position. This structure makes the bioavailability and biological activity of sn-2DHA in the human body higher than those of other forms of DHA, and it 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 loss during the absorption process in the human body, and maintain the original nutritional value. Existing methods usually use excessive free fatty acids for the transesterification reaction of DHA in algal oil. Since this reaction is a reversible reaction, the content of sn-2DHA is unstable and the increased proportion is not large. Moreover, if the transesterification reaction is excessive, DHA at the sn-1,3 positions will be removed in excess, reducing the total DHA content and degrading the product quality.

[0004] Algal oil is a natural plant source of DHA, and the relative content of its sn-2DHA is 35 - 40%, lower than the proportion of 50 - 60% in breast milk, which cannot meet the needs of infants. At the same time, increasing the content of sn-2DHA can also better meet the needs of adults, such as people in need of health care, patients with retinal diseases, and injured people with brain damage problems, etc. 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 catalyzing algal oil with lipase in view of the deficiencies of the existing technologies. This is a method for synthesizing triglycerides (TAG) with a high content of DHA at the sn-2 position by multi-step combined immobilized enzymes. In this method, through multi-step reactions, the content of sn-2DHA in the total DHA can be increased relatively stably, and the increased proportion can reach more than 60%. The obtained triglycerides with a high content of DHA at the sn-2 position can be preferably used in foods.

[0006] To achieve the above invention objectives, the technical solution of the present invention is as follows:

[0007] A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, which uses algal oil containing DHA, fully hydrolyzed sunflower oil, and medium- and long-chain fatty acids as raw materials and is prepared by a multi-step enzymatic method; specifically includes the following steps:

[0008] First step: Add immobilized lipase I to the algal oil containing DHA, then add medium- and long-chain fatty acids, and carry out an acidolysis reaction under vacuum conditions. Distill the reaction product by molecular distillation to remove the free fatty acids in the distillate, and collect the residue TAG.

[0009] Second step: Mix the residue TAG with medium- and long-chain fatty acids, then add immobilized lipase I, and carry out an acidolysis reaction under vacuum conditions. Then distill the reaction product by molecular distillation to remove the free fatty acids in the secondary distillate, and collect the secondary residue TAG.

[0010] Third step: Add immobilized lipase II to the secondary residue TAG for a hydrolysis reaction. After the reaction ends, centrifuge the mixture to separate the water layer, and the oil layer is the product. Add n-hexane and an ethanol solution to the product, ultrasonically mix evenly, centrifuge to separate the layers, and collect the lower ethanol aqueous phase; wash the ethanol aqueous phase with n-hexane, and remove ethanol and water under vacuum to obtain the product of the third step.

[0011] Fourth step: Add immobilized lipase I to the product of the third step, and add fully hydrolyzed sunflower oil. Carry out an acidolysis reaction under vacuum conditions. After the reaction ends, add an ethanol solution to the product, ultrasonically mix evenly, centrifuge to separate the layers, collect the lower ethanol aqueous phase, wash it with the ethanol solution again, and remove ethanol and water under vacuum to obtain the final product, that is, triglycerides with a high content of DHA at the sn-2 position.

[0012] Further, in the raw materials used in the method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, the mass percentage content of DHA in the algal oil is ≥40%; the mass percentage content of oleic acid OA in the fully hydrolyzed sunflower oil is 20-30%, and the mass percentage content of linoleic acid LA is 50-65%; the medium- and long-chain fatty acids are selected from at least one of n-caprylic acid, capric acid, lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, and linoleic acid.

[0013] Further, in the immobilized lipase used in the method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, the addition amount of the immobilized enzyme is the percentage content of the substrate mass.

[0014] As a preferred embodiment of the present application, in the first step of the method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, the addition amount of algal oil is calculated based on the DHA content, and the addition amount of sunflower seed fully hydrolyzed oil is calculated based on the oleic acid and linoleic acid contents. The molar ratio of DHA, oleic acid, and linoleic acid is 1:2-10:3-15. Preferably, the molar ratio is DHA:OA:LA = 1:(4-8):(5-10), and more 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 1-24 h, preferably 6-20 h, and most preferably 6-12 h. The molar ratio of DHA and medium- and long-chain fatty acids is 1:2-10, preferably 1:2-6; more preferably, the molar ratio is 1:3-5

[0015] As a preferred embodiment of the present application, in the second step of the method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, the residual TAG is mixed with a medium- and long-chain fatty acid. The molar ratio of DHA and medium- and long-chain fatty acids in the residual TAG is 1:2-10, preferably 1:2-8, and most preferably 1:3-6. Then, 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 1-24 h, preferably 6-20 h, and most preferably 9-12 h.

[0016] As a preferred embodiment of the present application, in the third step of the method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, 3-15% of immobilized lipase II is added to the secondary residual TAG, preferably 3-12%, and most preferably 3-6%. The hydrolysis reaction temperature is 40-60°C, preferably 45-50°C; the reaction time is 1-48 h, 12-32 h, and most preferably 18-24 h. After the reaction, n-hexane with an m / v ratio of 1:1-10 and an 80-95% ethanol solution with an m / v ratio of 1:1-10 are added to the product; the lower ethanol-water phase is collected and washed 2-5 times with n-hexane with a v / v ratio of 1:1-10, and ethanol and water are removed under vacuum at 85°C.

[0017] As a preferred embodiment of the present application, in the fourth step of the method for synthesizing triglycerides with a high DHA content at the sn-2 position by catalyzing algal oil with lipase, 3-15% of immobilized lipase I, preferably 3-12%, most preferably 6-12%, is added to the product of the third step, and sunflower whole hydrolysis oil with an m / m ratio of 1:2-6 is added; the acidolysis reaction temperature is 45-60 °C, preferably 50-60 °C; the reaction time is 1-12 h, preferably 6-10 h, most preferably 3-6 h; after the reaction is completed, an ethanol solution with a concentration of 80-95% and an m / v ratio of 1:1-10 is added to the product; the lower ethanol-aqueous phase is collected and washed 2-5 times with an ethanol solution with a concentration of 80-95% and a v / v ratio of 1:1-10, and ethanol and water are removed under vacuum at 85 °C.

[0018] As a preferred embodiment of the present application, in the method for synthesizing triglycerides with a high DHA content at the sn-2 position by catalyzing algal oil with lipase, the above lipase is an sn-1,3-specific lipase, and the lipase addition amount is 3-15% of the total substrate amount.

[0019] As a preferred embodiment of the present application, in the vacuum reaction of the method for synthesizing triglycerides with a high DHA content at the sn-2 position by catalyzing algal oil with lipase, the vacuum pressure is 10-100 Pa, preferably 30-50 Pa.

[0020] As a preferred embodiment of the present application, in the method for synthesizing triglycerides with a high DHA content at the sn-2 position by catalyzing algal oil with lipase, the method for removing free fatty acids is molecular distillation, with an evaporation temperature of 160 °C - 200 °C, a rotary vane speed of 1000-1100 rpm, and a pressure of 2-10 Pa.

[0021] As a preferred embodiment of the present application, in the method for synthesizing triglycerides with a high DHA content at the sn-2 position by catalyzing algal oil with lipase, the method for removing free fatty acids with a high acid value is ethanol-water extraction, in which an ethanol solution with a concentration of 80-95% and an m / v ratio of 1:1-10 is added; the lower ethanol-aqueous phase is collected and washed 2-5 times with an ethanol solution with a concentration of 80-95% and a v / v ratio of 1:1-10, and residual ethanol and water are removed under vacuum at 85 °C.

[0022] As a preferred embodiment of the present application, in the method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase, the lipase used is immobilized lipase I, which is an sn-1,3 position-selective lipase that can be used in transesterification reactions, such as: Aspergillus niger lipase, Rhizomucor miehei lipase, Thermomyces lanuginosus lipase, Candida rugosa lipase, porcine pancreatic lipase, etc.; immobilized lipase II is an sn-1,3 position-selective lipase that can be used in hydrolysis reactions, such as: Aspergillus niger lipase, porcine pancreatic lipase, DF Amano 15, Amano 250G, Rhizopus oryzae lipase, Rhizomucor miehei lipase, Rhizopus niveus, etc.

[0023] The present application also protects triglycerides with a high content of DHA at the sn-2 position obtained by combining any of the above methods and method steps.

[0024] 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 in TAG.

[0025] The present application also protects the application of the triglycerides with a high content of DHA at the sn-2 position as described above in foods.

[0026] Furthermore, the foods include formula foods, nutritional products, health foods, etc.

[0027] Compared with the existing technology, the beneficial effects of the present invention are:

[0028] (1) Through multiple-step reactions, the content of sn-2 DHA in the total DHA can be stably increased, and the increase ratio can reach more than 60%.

[0029] (2) In the intermediate steps, more fatty acids can be selectively introduced or removed, making the types of the final product structural esters richer and more suitable for the health needs of the human body.

[0030] (3) Molecular distillation, n-hexane-ethanol-water extraction method and ethanol-water extraction method effectively avoid side reactions of acyl migration.

[0031] (4) Residual free fatty acids in the product can be effectively removed, making it easier for the acid value of the product to meet the standard.

[0032] (5) The use of immobilized enzymes increases the reuse value of the enzymes, greatly reduces the cost of commercial lipases, and the production process is more economical.

[0033] (6) The process does not use any toxic and harmful solvents, and the raw materials used are all food-grade. The preparation method is safe, efficient, accurate, and practical in industrial production. Description of the Drawings

[0034] Figure 1 The gas chromatogram (GC) of the triglyceride product obtained in Example 1. In the figure, starting from the right, the rightmost peak is the total DHA;

[0035] Figure 2 The gas chromatogram (GC) of the triglyceride product obtained in Example 1. In the figure, starting from the right, the rightmost peak is the DHA at the sn-2 position;

[0036] Figure 3 The gas chromatogram (GC) of the triglyceride product obtained in Example 2. In the figure, starting from the right, the rightmost peak is the total DHA;

[0037] Figure 4 The gas chromatogram (GC) of the triglyceride product obtained in Example 2. In the figure, starting from the right, the rightmost peak is the DHA at the sn-2 position;

[0038] Figure 5 The gas chromatogram (GC) of the triglyceride product obtained in Example 3. In the figure, starting from the right, the rightmost peak is the total DHA;

[0039] Figure 6 The gas chromatogram (GC) of the triglyceride product obtained in Example 3. In the figure, starting from the right, the rightmost peak is the DHA at the sn-2 position;

[0040] Figure 7 The gas chromatogram (GC) of the triglyceride product obtained by the traditional method in Comparative Example 1. In the figure, starting from the right, the rightmost peak is the total DHA;

[0041] Figure 8 The gas chromatogram (GC) of the triglyceride product obtained by the method of the present invention in Comparative Example 1. In the figure, starting from the right, the rightmost peak is the total DHA;

[0042] Figure 9 The gas chromatogram (GC) of the triglyceride product obtained by the traditional method in Comparative Example 1. In the figure, starting from the right, the rightmost peak is the DHA at the sn-2 position;

[0043] Figure 10 The gas chromatogram (GC) of the triglyceride product obtained by the method of the present invention in Comparative Example 1. In the figure, starting from the right, the rightmost peak is the DHA at the sn-2 position. Detailed implementation manners

[0044] All features disclosed in this specification, or all steps in the disclosed methods or processes, except for mutually exclusive features and / or steps, can be combined in any manner.

[0045] Any feature disclosed in this specification (including claims and abstract), unless specifically recited, can be replaced by other equivalent or similar-purpose alternative features. That is, unless specifically recited, each feature is merely an example in a series of equivalent or similar features.

[0046] The features and properties of the present invention will be further described in detail below in conjunction with embodiments.

[0047] The immobilized porcine pancreatic lipase used in the following examples was obtained by immobilizing commercially available porcine pancreatic lipase through the prior art; the immobilized Aspergillus niger lipase was obtained by immobilizing commercially available Aspergillus niger lipase through the prior art; other raw material reagents are all prior art or can be obtained through the prior art.

[0048] In this application, % represents mass percentage unless otherwise specified.

[0049] Example 1:

[0050] A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil is prepared by a multi-step combination method and carried out in a stainless steel jacketed stirred reactor under a solvent-free system. The specific steps are as follows:

[0051] First step: Add 8% (calculated based on the mass percentage of the substrate, the same below) of the immobilized Aspergillus niger lipase to the algal oil with a DHA content of 44.06%, then add OA, with an m / m of 1:4, and carry out an acidolysis reaction at 50 °C under vacuum for 9 h. Distill the reaction product by molecular distillation to remove the distillate FFA, and collect the residue TAG.

[0052] Second step: Mix the residue TAG with new OA, with an m / m of 1:4, then add 8% of the immobilized Aspergillus niger lipase, and carry out an acidolysis reaction at 50 °C under vacuum for 9 h. Distill the reaction product by molecular distillation to remove the secondary distillate FFA, and collect the secondary residue TAG.

[0053] Third step: Add 4% of the immobilized porcine pancreatic lipase to the secondary residue TAG, carry out a hydrolysis reaction at 50 °C for 12 h. After the reaction is completed, centrifuge the mixture to obtain the product. Add n-hexane with an m / v of 1:2 and 85% ethanol solution with an m / v of 1:2 to the product, ultrasonically mix evenly, centrifuge and separate layers, collect the lower ethanol-aqueous phase, wash it 3 times with n-hexane with a v / v of 1:2, and remove ethanol and water under vacuum at 85 °C to obtain the product of the third step.

[0054] 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. Conduct acidolysis reaction at 50 °C under vacuum for 9 h. After the reaction, add a 90% ethanol solution with a m / v ratio of 1:2 to the product, ultrasonically mix evenly, centrifuge for stratification, collect the lower ethanol-aqueous phase, wash it 3 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 GC analysis results of the product are shown in Figure 1 、 Figure 2 。

[0055] Example 2

[0056] A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, comprising the following steps:

[0057] Step 1: Add 6% immobilized Aspergillus niger lipase to algal oil with a DHA content of 71.73%, and then add LA with a m / m ratio of 1:2. Conduct acidolysis reaction at 45 °C under vacuum for 12 h. Remove the distilled FFA from the reaction product by molecular distillation, and collect the residue TAG.

[0058] Step 2: Mix the residue TAG with fresh LA with a m / m ratio of 1:2, and then add 6% immobilized Aspergillus niger lipase. Conduct acidolysis reaction at 45 °C under vacuum for 12 h. Remove the secondary distilled FFA from the reaction product by molecular distillation, and collect the secondary residue TAG.

[0059] Step 3: Add 3% immobilized porcine pancreatic lipase to the secondary residue TAG, conduct hydrolysis reaction at 45 °C for 12 h. After the reaction, centrifuge the mixture to obtain the product. Add a n-hexane with a m / v ratio of 1:2 and an 85% ethanol solution with a m / v ratio of 1:2 to the product, ultrasonically mix evenly, centrifuge for stratification, collect the lower ethanol-aqueous phase, wash it 3 times with a n-hexane with a v / v ratio of 1:2, and remove ethanol and water under vacuum at 85 °C to obtain the product of Step 3.

[0060] 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. Conduct acidolysis reaction at 45 °C under vacuum for 12 h. After the reaction, add a 90% ethanol solution with a m / v ratio of 1:2 to the product, ultrasonically mix evenly, centrifuge for stratification, collect the lower ethanol-aqueous phase, wash it 3 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 GC analysis results of the product are shown in Figure 3 Figure 4 。

[0061] Example 3

[0062] A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase. Under a solvent-free system, it includes the following steps:

[0063] Step 1: Add 12% immobilized Aspergillus niger lipase to algal oil with a DHA content of 56.67%, then add n-caprylic acid, with a mass ratio (m / m) of 1:5. Conduct acidolysis reaction at 55 °C for 6 h under vacuum conditions. Subject the reaction product to molecular distillation to remove the distilled FFA, and collect the residue TAG.

[0064] Step 2: Mix the residue TAG with fresh n-caprylic acid, with a mass ratio (m / m) of 1:5. Then add 12% immobilized Aspergillus niger lipase. Conduct acidolysis reaction at 55 °C for 6 h under vacuum conditions. Subject the reaction product to molecular distillation to remove the secondary distilled FFA, and collect the secondary residue TAG.

[0065] Step 3: Add 6% immobilized porcine pancreatic lipase to the secondary residue TAG. Conduct hydrolysis reaction at 45 °C for 24 h. After the reaction, centrifuge the mixture to obtain the product. Add n-hexane with a mass / volume ratio (m / v) of 1:2 and 85% ethanol solution with a mass / volume ratio (m / v) of 1:2 to the product, ultrasonically mix evenly, centrifuge to separate layers, collect the lower ethanol-aqueous phase, wash it 3 times with n-hexane with a volume / volume ratio (v / v) of 1:2, and remove ethanol and water under vacuum at 85 °C to obtain the product of Step 3.

[0066] Step 4: Add 12% immobilized Aspergillus niger lipase to the product of Step 3, and add sunflower seed fully hydrolyzed oil with a mass ratio (m / m) of 1:5. Conduct acidolysis reaction at 55 °C for 6 h under vacuum conditions. After the reaction, add 90% ethanol solution with a mass / volume ratio (m / v) of 1:2 to the product, ultrasonically mix evenly, centrifuge to separate layers, collect the lower ethanol-aqueous phase, wash it 3 times with 90% ethanol solution with a volume / volume ratio (v / v) 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 GC analysis results of the product are shown in Figure 5 Figure 6 。

[0067] Comparative Example 1

[0068] The steps of the traditional method are as follows:

[0069] Step 1: Add 12% immobilized Aspergillus niger lipase to algal oil with a DHA content of 44.06%, then add palmitic acid, with a mass ratio (m / m) of 1:5. Conduct acidolysis reaction at 55 °C for 6 h under vacuum conditions.

[0070] Step 2: Subject the reaction product to molecular distillation to remove the distilled FFA, and collect the residue TAG, which is the product. The GC analysis results of the product are shown in Figure 7 、 Figure 9 。

[0071] The steps of the method of the present invention are as follows:

[0072] Step 1: Add 12% immobilized Aspergillus niger lipase to algal oil with a DHA content of 44.06%, then add palmitic acid, with a m / m ratio of 1:5. Conduct an acidolysis reaction at 55°C for 6 h under vacuum conditions. Subject the reaction product to molecular distillation to remove the distilled FFA, and collect the residue TAG.

[0073] Step 2: Mix the residue TAG with fresh LA, with a m / m ratio of 1:5, and then add 12% immobilized Aspergillus niger lipase. Conduct an acidolysis reaction at 55°C for 6 h under vacuum conditions. Subject the reaction product to molecular distillation to remove the secondary distilled FFA, and collect the secondary residue TAG.

[0074] Step 3: Add 6% immobilized porcine pancreatic lipase to the secondary residue TAG, and conduct a hydrolysis reaction at 45°C for 24 h. 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, ultrasonically mix evenly, centrifuge to separate layers, collect the lower ethanol-aqueous phase, wash it 3 times with n-hexane with a v / v ratio of 1:2, and remove ethanol and water under vacuum at 85°C to obtain the product of the third step.

[0075] Step 4: Add 12% immobilized Aspergillus niger lipase to the product of the third step, and add sunflower seed fully hydrolyzed oil with a m / m ratio of 1:5. Conduct an acidolysis reaction at 55°C for 6 h under vacuum conditions. After the reaction, add 90% ethanol solution with a m / v ratio of 1:2 to the product, ultrasonically mix evenly, centrifuge to separate layers, collect the lower ethanol-aqueous phase, wash it 3 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 the fourth step, i.e., the final product. The GC analysis results of the product are shown in Figure 8 Figure 10 .

[0076] The above-described embodiments merely represent the specific implementation manners of the present application. The description is relatively specific and detailed, but it should not be construed as a limitation on the protection scope of the present application. It should be noted that for those of ordinary skill in the art, without departing from the concept of the technical solution of the present application, several deformations and improvements can still be made, and these all fall within the protection scope of the present application.

[0077] This background art section is provided to generally present the context of the present invention. The work of the currently named inventors, to the extent described in this background art section, and aspects that are not prior art at the time of filing this application are neither expressly nor impliedly admitted to be prior art of the present invention.

Claims

1. A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by lipase-catalyzed algal oil, characterized in that: It is prepared from algal oil containing DHA, fully hydrolyzed sunflower seed oil and medium and long chain fatty acids through multi-step enzymatic methods, such that the content of sn-2 DHA accounts for more than 60% of the total DHA content in TAG; specifically including the following steps: The first step: Add immobilized lipase I to the algal oil containing DHA, then add medium and long chain fatty acids, and conduct an acidolysis reaction under vacuum conditions. Remove the free fatty acids from the reaction product by molecular distillation, and collect the residue TAG; the acidolysis reaction temperature is 45-60 °C, and the reaction time is 1-24 h; The second step: Mix the residue TAG with medium and long chain fatty acids, then add immobilized lipase I, conduct an acidolysis reaction under vacuum conditions, and then remove the free fatty acids of the secondary distillate from the reaction product by molecular distillation, and collect the secondary residue TAG; the acidolysis reaction temperature is 45-60 °C, and the reaction time is 1-24 h; The third step: Add immobilized lipase II to the secondary residue TAG for hydrolysis reaction. After the reaction ends, centrifuge the mixture to separate water to obtain the oil layer as the product. Add n-hexane and ethanol solution to the product, ultrasonically mix evenly, centrifuge and layer, and collect the lower ethanol aqueous phase; wash the ethanol aqueous phase with n-hexane, and remove ethanol and water under vacuum to obtain the product of the third step; the hydrolysis reaction temperature is 45-60 °C, and the reaction time is 1-48 h; The fourth step: Add immobilized lipase I to the product of the third step, and add fully hydrolyzed sunflower seed oil, and conduct an acidolysis reaction under vacuum conditions. After the reaction ends, add ethanol solution to the product, ultrasonically mix evenly, centrifuge and layer, collect the lower ethanol aqueous phase, wash with ethanol solution again, and remove ethanol and water under vacuum to obtain the final product, that is, triglyceride with high content of DHA at the sn-2 position; the acidolysis reaction temperature is 45-60 °C, and the reaction time is 1-12 h; The immobilized lipase I is an sn-1,3 position selective lipase for transesterification reaction, selected from Aspergillus niger lipase; the immobilized lipase II is an sn-1,3 position selective lipase for hydrolysis reaction, selected from porcine pancreatic lipase.

2. The method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase as claimed in claim 1, wherein: The addition amounts of the immobilized lipase I and the immobilized lipase II are both 3%-15% of the total mass of the substrates.

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

4. A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase as described in claim 1, characterized in that: In the first step, the addition amount of the algal oil is based on the DHA content, and the addition amount of the fully hydrolyzed sunflower seed oil is based on the oleic acid and linoleic acid contents. The molar ratio of DHA, oleic acid and linoleic acid is 1:2-10:3-15; the molar ratio of DHA and medium and long chain fatty acids is 1:2-10.

5. A method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase as described in claim 1, characterized in that: In the second step, mix the residue TAG with a medium and long chain fatty acid. The molar ratio of DHA and medium and long chain fatty acids in the residue TAG is 1:2-10, and then add 3%-15% of immobilized lipase I.

6. The method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase as claimed in claim 1, wherein: In the third step, the immobilized lipase II is added to the secondary still residue TAG; after the reaction, n-hexane with an m / v ratio of 1:1 to 10 and an ethanol solution of 80% to 95% with an m / v ratio of 1:1 to 10 are added to the product; the lower ethanol-aqueous phase is collected and washed 2 to 5 times with n-hexane with a v / v ratio of 1:1 to 10, and ethanol and water are removed under vacuum at 85°C.

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

8. The method for synthesizing triglycerides with a high content of DHA at the sn-2 position by catalyzing algal oil with lipase according to claim 1, characterized in that: The vacuum pressure in the vacuum condition is 10 to 100 Pa.

9. The method for synthesizing triglycerides with a high DHA content at the sn-2 position by catalyzing algal oil with lipase as claimed in claim 1, wherein: For the molecular distillation method, the evaporation temperature is 160°C to 200°C, the rotary vane speed is 1000 to 1100 rpm, and the pressure is 2 to 10 Pa.

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