Mesoporous ZnNA derived carbon material as well as preparation method and application thereof

Abstract

The invention provides a mesoporous ZnNA derived carbon material as well as a preparation method and application thereof. The mesoporous ZnNA derived carbon material provided by the invention can be used for technologies of lipase immobilization and grease high-value conversion, effectively solves the problems that the traditional MOFs material is unstable, is difficult to recover due to small particle size, is difficult to be used for macromolecular lipase immobilization due to small external specific surface area and the like, and shows excellent performance in lipase immobilization and grease high-value conversion; good development and application prospects are realized.

Description

Technical Field

[0001] The present invention relates to the field of biochemistry, and in particular, to a mesoporous ZnNA-derived carbon material, a preparation method thereof, and an application thereof. Background Art

[0002] As an emerging material, metal-organic framework materials (MOFs) have important development and application prospects for lipase immobilization and subsequent high-value conversion of oils and fats. However, there are three major challenges that need to be addressed urgently when MOFs are applied to lipase immobilization: the problem of carrier stability, the problem of pore size compatibility, and the problem of recycling.

[0003] MOFs are formed by relatively unstable coordination bonds and have stability problems in different environments. MOF-immobilized lipases are generally applied to non-aqueous or oil-water two-phase systems to catalyze oil and fat substrates, mainly involving various different reaction types such as hydrolysis, esterification, and transesterification. During the catalytic process, MOF-immobilized lipases will inevitably come into contact with various substrates such as water and fatty acids. In order to enhance the reusability of MOF-immobilized lipases, it is necessary to develop appropriate strategies to prepare MOF carriers that are stable in the application system. In addition, the problem caused by the negative impact on the immobilization effect and catalytic effect due to the mismatch in size between the pore size of MOFs and enzyme molecules and substrate molecules is called the pore size compatibility problem. Conventionally prepared MOFs are generally microporous structures (<2 nm), while the size of lipase molecules is generally between 5 and 10 nm, which results in the inability of lipase molecules to enter the interior of MOF pores during immobilization. Therefore, only surface immobilization or in-situ embedding methods can be used. However, both of these methods have certain problems: surface immobilization has low immobilization efficiency and poor protection ability, resulting in unsatisfactory immobilization effects such as enzyme loading and reusability. Although in-situ embedding has a better immobilization effect, the immobilization process requires biocompatible synthesis conditions, and the available MOF materials are very limited. In addition, the molecular size of the most common triglyceride substrate during the lipase catalytic process is close to 3.5 nm, and the immobilized lipase prepared by in-situ embedding using microporous MOFs can hardly catalyze it. Based on this, it is necessary to construct mesoporous (2 - 50 nm) or macroporous (>50 nm) MOFs for pore immobilization of lipase molecules, which can not only make full use of the protective effect of the MOF framework but also eliminate the limitation of substrate size, and is the key to solving the pore size compatibility problem of MOF-immobilized lipases. In addition, the crystal size of MOFs is usually in the micron to nanometer range, which makes its recycling after being used for lipase immobilization challenging. It often needs to be separated from the application system by high-speed centrifugation or filtration, and the operation process is complex and brings greater energy consumption and production costs. Summary of the Invention

[0004] The object of the present invention is to provide a mesoporous ZnNA-derived carbon material, a preparation method thereof and an application thereof.

[0005] The inventive concept of the present invention is as follows: Aiming at the three major challenges faced by MOFs-immobilized lipase in the actual application system, namely the carrier stability, pore size compatibility and difficult recycling, the present invention proposes to perform high-temperature pyrolysis on mesoporous ZnNA, and introduce magnetism during the pyrolysis process to achieve its stability, reusability and generate more mesopores to increase its external specific surface area. The prepared material can be effectively used for the immobilization of macromolecular lipase.

[0006] To achieve the object of the present invention, in the first aspect, the present invention provides a preparation method of a mesoporous ZnNA-derived carbon material, comprising the following steps: 1) Performing high-temperature pyrolysis on ZnNA under a nitrogen atmosphere to obtain a mesoporous ZnNA-derived carbon material; or, Introducing iron element during the synthesis of ZnNA, and then performing high-temperature pyrolysis on Fe-ZnNA under a nitrogen atmosphere to produce a magnetic mesoporous Fe-ZnNA-derived carbon material; 2) Further performing low-temperature oxidation on the mesoporous material obtained in step 1) under an air atmosphere to obtain the product.

[0007] Among them, the preparation method of ZnNA includes: mixing a nicotinic acid solution and a zinc acetate solution, placing at 25-35 °C for about 0.5-2 h (preferably placing at 30 °C for about 1 h), then filtering, collecting the precipitate, washing the precipitate with pure water 2-4 times (preferably 3 times), and drying overnight in an oven at 45-60 °C (preferably 50 °C) to obtain the metal-organic framework material ZnNA.

[0008] The preparation method of Fe-ZnNA includes: mixing a nicotinic acid solution and a zinc acetate solution, placing at 25-35 °C for about 0.5-2 h (preferably placing at 30 °C for about 1 h), and then adding a substance containing Fe 2+ (such as FeSO4), stirring for 20-50 min (preferably 30 min), filtering, and drying overnight in an oven at 45-60 °C (preferably 50 °C) to obtain the magnetic metal-organic framework material Fe-ZnNA.

[0009] Further, step 1) includes: putting ZnNA or Fe-ZnNA into a tube furnace, and then heating to 400-1000 °C at a rate of 4-10 °C / min (preferably 5 °C / min) under a nitrogen atmosphere, and maintaining for 1-3 h (preferably 2 h), and then annealing to room temperature to obtain a mesoporous ZnNA-derived carbon material or a magnetic mesoporous Fe-ZnNA-derived carbon material.

[0010] Further, the conditions for performing low-temperature oxidation in step 2) under an air atmosphere are: 200-350 °C, 1-3 hours.

[0011] Further, in the preparation method of ZnNA, 100 - 300 mL of 0.154 - 0.616 mol / L nicotinic acid solution and 100 - 300 mL of 0.154 - 0.616 mol / L zinc acetate solution are mixed.

[0012] Further, in the preparation method of Fe - ZnNA, 100 - 300 mL of 0.154 - 0.616 mol / L nicotinic acid solution and 100 - 300 mL of 0.154 - 0.616 mol / L zinc acetate solution are mixed, placed at 25 - 35 °C for about 0.5 - 3 h (preferably placed at 30 °C for about 1 h), and then 100 - 200 mL of 0.08 - 0.9 mol / L FeSO4 solution is added.

[0013] In the second aspect, the present invention provides a mesoporous ZnNA - derived carbon material prepared according to the described method. Its average pore diameter is 6 - 12 nm.

[0014] In the third aspect, the present invention provides any one of the following applications of the mesoporous ZnNA - derived carbon material: 1) For the field of enzyme immobilization; 2) For the high - value conversion of oils and fats.

[0015] In the fourth aspect, the present invention provides a method for the high - value conversion of oils and fats by immobilizing lipase using the mesoporous ZnNA - derived carbon material, and the method includes: 1) The mesoporous ZnNA - derived carbon material is infiltrated in absolute ethanol, then deionized water is added, shaken well, and free lipase is added, shaken and mixed evenly, placed in a shaker at 30 - 50 °C, 100 - 400 rpm (preferably 40 °C, 200 rpm) for immobilization for 1 - 4 h, the immobilized enzyme is recovered by centrifugation, washed with deionized water, and freeze - dried for 8 - 16 h (preferably 12 h) to obtain the immobilized lipase; 2) The immobilized lipase is added to a single - stage or multi - stage enzyme reactor for the enzymatic conversion reaction of oils and fats.

[0016] Further, the lipase includes but is not limited to lipase derived from Candida antarctica ( Candida antarctica ), Thermomyces lanuginosus ( Thermomyces lanuginosus ), Aspergillus niger ( Aspergillus niger ), Aspergillus oryzae ( Aspergillus oryzae ), Rhizomucor miehei ( Rhizomucor miehei ), Rhizopus oryzae ( Rhizopus oryzae ); The oils and fats include but are not limited to biodiesel, structured phospholipids, and human milk substitute fats.

[0017] With the above technical solutions, the present invention has at least the following advantages and beneficial effects: The mesoporous ZnNA-derived carbon material provided by the present invention can be used in the technologies of lipase immobilization and high-value conversion of oils and fats, effectively solving the problems of instability of traditional MOFs materials, small particle size and difficult recovery, and small external specific surface area that is difficult to be used for the immobilization of macromolecular lipases. It exhibits excellent performance in lipase immobilization and high-value conversion of oils and fats, and has good development and application prospects. Description of the Drawings

[0018] Figure 1 Scanning electron microscope images of ZnNA synthesized at different precursor molar ratios (zinc acetate: nicotinic acid) in the preferred embodiment of the present invention; (a) 2:1; (b) 1:1; (c) 1:0.75; (d) 0.75:1; (e) 0.75:0.75.

[0019] Figure 2 Flow chart of preparing ZnNA-derived carbon material by pyrolysis of ZnNA in the preferred embodiment of the present invention.

[0020] Figure 3 XRD curve of Fe-ZnNA in the preferred embodiment of the present invention.

[0021] Figure 4 N2 (77 K) adsorption-desorption isotherm of mesoporous CMFe-ZnNA (average pore diameter about 11 nm) in the preferred embodiment of the present invention.

[0022] Figure 5 Pore size distribution diagram of mesoporous CMFe-ZnNA (average pore diameter about 11 nm) in the preferred embodiment of the present invention.

[0023] Figure 6 When the ZnNA-derived carbon material and Fe-ZnNA-derived magnetic carbon material in the preferred embodiment of the present invention are used as carriers for lipase immobilization, the specific enzyme activity is significantly improved. Left: after low-temperature oxidation detoxification in air atmosphere (CMFe-ZnNA), right: before low-temperature oxidation detoxification in air atmosphere. Detailed Embodiments

[0024] The present invention aims to provide a mesoporous ZnNA-derived carbon material and a preparation method thereof, and use the mesoporous ZnNA-derived carbon material in the technologies of lipase immobilization and high-value conversion of oils and fats.

[0025] The present invention adopts the following technical solutions: First, ZnNA is pyrolyzed at high temperature under a nitrogen atmosphere to obtain a mesoporous ZnNA-derived carbon material. Iron species can also be introduced during the synthesis of ZnNA, and then Fe-ZnNA is pyrolyzed at high temperature under a nitrogen atmosphere to produce a magnetic mesoporous Fe-ZnNA-derived carbon material. The obtained mesoporous material is further oxidized at low temperature under an air atmosphere to obtain an excellent carrier with chemically intrinsic stability, easy recyclability under a magnetic field, and a significantly increased external specific surface area, which exhibits excellent performance in lipase immobilization and high-value conversion of oils and fats.

[0026] The preparation technology of the present invention is high-temperature pyrolysis under a nitrogen atmosphere in the first step and low-temperature oxidation under an air atmosphere in the second step.

[0027] Furthermore, the material for high-temperature pyrolysis under a nitrogen atmosphere is ZnNA or Fe-ZnNA.

[0028] Among them, Fe-ZnNA is prepared by adding an Fe-containing 2+ substance during the synthesis of ZnNA. The Fe-containing 2+ substance includes but is not limited to FeSO4.

[0029] The synthesis method of the ZnNA is as follows: Mix 100 - 300 mL of a 0.154 - 0.616 mol / L nicotinic acid solution and 100 - 300 mL of a 0.154 - 0.616 mol / L zinc acetate solution, place it at 25 - 35 °C for 0.5 - 3 h, then filter, collect the precipitate, wash the precipitate with pure water 2 - 4 times, and dry it overnight in an oven at 45 - 60 °C to obtain the metal-organic framework material ZnNA.

[0030] The synthesis method of the Fe-ZnNA is as follows: Mix 100 - 300 mL of a 0.154 - 0.616 mol / L nicotinic acid solution and 100 - 300 mL of a 0.154 - 0.616 mol / L zinc acetate solution, stir and mix well, place it at 25 - 35 °C for 0.5 - 3 hours, then add 100 - 200 mL of a 0.08 - 0.9 mol / L FeSO4 solution, stir for 20 - 50 min, filter, and dry it overnight in an oven at 45 - 60 °C to obtain Fe-ZnNA.

[0031] Furthermore, the conditions for obtaining the mesoporous derived carbon material by high-temperature pyrolysis of ZnNA or Fe-ZnNA under a nitrogen atmosphere are as follows: Put ZnNA into a tubular furnace, then heat it to 400 - 1000 °C at a rate of 4 - 10 °C / min and maintain it for 1 - 3 h, and then anneal to obtain a series of mesoporous ZnNA or Fe-ZnNA-derived carbon materials.

[0032] Furthermore, the conditions for low-temperature oxidation under an air atmosphere are: 200 - 350 °C, 1 - 3 hours.

[0033] The obtained mesoporous ZnNA or Fe-ZnNA-derived carbon materials can be used as carriers for lipase immobilization. The lipases include, but are not limited to, those derived from Candida antarctica ( Candida antarctica ), thermophilic fungi ( Thermomyces lanuginosus ), Aspergillus niger ( Aspergillus niger ), Aspergillus oryzae ( Aspergillus oryzae ), Rhizomucor miehei ( Rhizomucor miehei ), Rhizopus oryzae ( Rhizopus Oryzae ).

[0034] The immobilized lipases can efficiently catalyze the high-value conversion of oils and fats, including, but not limited to, fields such as functional oil modification, biodiesel, structured phospholipids, and human milk substitute fats.

[0035] The following examples are used to illustrate the present invention, but do not limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0036] Example 1 Preparation of Mesoporous ZnNA-Derived Carbon Material 1. Synthesis of ZnNA Dissolve 4.74 g of nicotinic acid and 7.04 g of Zn(AC)2 in 250 mL of water respectively. Then mix the solutions, stir evenly, place at 30 °C for about 1 h, then filter, wash 3 times with 40 mL of pure water, and dry overnight in an oven at 50 °C to obtain MOFs-ZnNA.

[0037] Figure 1 is the electron micrograph of ZnNA crystals synthesized at different molar concentration ratios of Zn(AC)2 to nicotinic acid (the saturated concentration of nicotinic acid solution at room temperature is 0.154 mol / L, and 0.154 mol / L is defined as 1). Rhombic crystal morphologies are obtained at different precursor ratios (2:1, 1:1, 1:0.75, 0.75:1, 0.75:0.75), with no obvious difference in morphology. The size of ZnNA crystals is about one hundred micrometers, and there is no obvious difference in crystal size.

[0038] 2. Synthesis of ZnNA-Derived Carbon Material Weigh a certain mass of ZnNA and place it in a tube furnace for high-temperature treatment in an N2 atmosphere. The specific conditions are as follows: heat to 600 - 1000 °C at a rate of 5 °C / min, maintain for 2 h, and then anneal to room temperature to obtain the ZnNA-derived carbon material CZnNA. The pyrolysis process is as Figure 2 shown. The relevant characteristics of the derived carbon materials obtained at different pyrolysis temperatures are shown in Table 1.

[0039] Table 1 Average pore diameter, BET specific surface area, and t-plot external specific surface area of materials at different pyrolysis temperatures

[0040] Note: In Table 1, CZnNA-600, CZnNA-700, CZnNA-800, CZnNA-900, and CZnNA-1000 are materials at pyrolysis temperatures of 600 °C, 700 °C, 800 °C, 900 °C, and 1000 °C respectively.

[0041] Example 2 Synthesis of Fe-ZnNA and Its Derived Magnetic Carbon Materials 1. Synthesis of Fe-ZnNA Weigh 4.736 g of nicotinic acid and 7.046 g of Zn(AC)2 and dissolve them separately in 250 mL of water. Then mix the solutions, stir properly, and let it stand at 30 °C for about 1 h. Subsequently, add 100 - 200 mL of FeSO4 solution with different concentrations, stir for 30 min, filter, and place it in an oven at 50 °C to dry overnight to obtain Fe-ZnNA. Figure 3 This is its XRD diffraction pattern. Compared with the original ZnNA, the positions of the diffraction peaks do not change significantly, indicating that the introduction of Fe element has no effect on the crystal structure of ZnNA itself.

[0042] 2. Synthesis of CFe-ZnNA Weigh a certain mass of Fe-ZnNA and put it into a tubular furnace for high-temperature pyrolysis as Figure 2 shown. Heat it up to 800 °C at a rate of 5 °C / min and maintain for 2 h, then anneal to room temperature to obtain a magnetic derived carbon material named CFe-ZnNA.

[0043] 3. Preparation of Fe-ZnNA Derived Magnetic Carbon Material by Air Oxidation Detoxification The aforementioned magnetic mesoporous carbon material CFe-ZnNA is subjected to low-temperature oxidation detoxification at 300 °C in an air atmosphere in a muffle furnace for 2 hours to prepare mesoporous CMFe-ZnNA (average pore diameter about 11 nm). The N2 (77 K) adsorption-desorption isotherm ( Figure 4 ), pore size distribution is as Figure 5 shown.

[0044] Example 3 Immobilization of Lipase Using ZnNA Derived Carbon Material and Fe-ZnNA Derived Magnetic Carbon Material as Carriers 50 mg of the carrier (ZnNA derived carbon material or Fe-ZnNA derived magnetic carbon material) is infiltrated in absolute ethanol, then deionized water is added and shaken well. Then free lipase is added, shaken and mixed evenly, placed at 40 °C, and shaken in a shaker at 200 rpm for 1 - 4 h. The immobilized enzyme is recovered by centrifugation, washed with deionized water, and freeze-dried for 12 h to obtain immobilized lipase, and the specific enzyme activity is significantly improved ( Figure 6 )

[0045] Example 4 High-value conversion of oil and fat catalyzed by immobilized lipase Application Example 1: 100 g of rapeseed oil and methanol with a molar ratio of alcohol to oil of 6:1 were placed in an enzyme reactor, and Fe-ZnNA-derived carbon material-immobilized lipase (derived from Aspergillus niger), methanol was added uniformly within 2 hours, and the reaction temperature was controlled at 40 °C. After reacting for 8 hours, the yield of biodiesel was 94%. The above immobilized lipase was separated by centrifugation and reused. Under the same reaction conditions, the yield of biodiesel for 10 batches of the same reaction was 85%.

[0046] Application Example 2: 10 g of soybean phospholipids, docosahexaenoic acid ethyl ester with a molar ratio of phospholipids of 25, n-hexane and acetone with a volume ratio of 1:1 (total volume of 20 mL), Fe-ZnNA-derived magnetic carbon material-immobilized lipase based on 500 standard enzyme activities per unit mass of phospholipids (derived from Candida antarctica ), and Fe-ZnNA-derived magnetic carbon material-immobilized lipase based on 600 standard enzyme activities per unit mass of phospholipids (derived from Aspergillus oryza ), were placed in a first-stage or multi-stage reactor suitable for immobilized enzyme catalysis for reaction. After reacting at 60 °C for 20 hours, DHA on the structured phospholipids accounted for 68% of all fatty acids. The immobilized lipase was separated by magnetic field centrifugation and reused. Under the same reaction conditions, for 10 batches of the same reaction, DHA on the structured phospholipids accounted for 65% of all fatty acids.

[0047] Application Example 3: 10 g of stearyl palmitate, ethyl oleate 10 times the amount of stearyl palmitate in terms of amount of substance, Fe-ZnNA-derived magnetic carbon material-immobilized lipase based on 1000 standard enzyme activities per unit mass of stearyl palmitate (derived from Rhizomucor miehei ), and Fe-ZnNA-derived magnetic carbon material-immobilized lipase based on 500 standard enzyme activities per unit mass of stearyl palmitate (derived from Thermomyces lanuginosu ), were placed in a reactor suitable for immobilized enzyme-catalyzed transesterification reaction. After reacting at 60 °C for 2 hours, the material was then crystallized at 0 - 10 °C for 4 h, and then the solid and liquid phases were separated by centrifugation. The liquid phase rich in 1,3-dioleoyl-2-palmitoyl glycerol (OPO) was separated out, and the solid phase was further used for the second-step enzyme-catalyzed transesterification reaction. Then, 10 g of the obtained solid-phase triglyceride was taken, and ethyl oleate 7 times the amount of substance of the solid-phase triglyceride was used. Fe-ZnNA-derived magnetic carbon material-immobilized lipase based on 1000 standard enzyme activities per unit mass of the obtained solid-phase triglyceride (derived from Candida antarctica), and placed in a reactor suitable for immobilized enzyme catalysis for transesterification reaction. The reaction was carried out at 50 °C for 4 hours. The total yield of OPO was 92%. The total proportion of oleic acid in the sn-1,3 positions of the product triglyceride was 96%, and the proportion of palmitic acid in sn-2 was 94%. The above immobilized lipase was recycled repeatedly by magnetic separation. The reaction conditions were the same as above. The same reaction was catalyzed for 10 batches. The total yield of OPO was 90%. The total proportion of oleic acid in the sn-1,3 positions of the product triglyceride was 94%, and the proportion of palmitic acid in sn-2 was 92%.

[0048] Although the present invention has been described in detail with general descriptions and specific embodiments above, based on the present invention, some modifications or improvements can be made, which are obvious to those skilled in the art. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection required by the present invention.

Claims

1. A preparation method of a mesoporous ZnNA-derived carbon material, characterized in that, It includes the following steps: 1) High-temperature pyrolysis of ZnNA in a nitrogen atmosphere to obtain mesoporous ZnNA-derived carbon materials; Or, Introduce iron element during the synthesis of ZnNA, and then perform high-temperature pyrolysis on Fe-ZnNA in a nitrogen atmosphere to produce magnetic mesoporous Fe-ZnNA-derived carbon materials; 2) Further perform low-temperature oxidation on the mesoporous material obtained in step 1) in an air atmosphere to obtain the product; Among them, the preparation method of ZnNA includes: mixing nicotinic acid solution and zinc acetate solution, placing at 25 - 35 °C for 0.5 - 2 h, then filtering, collecting the precipitate, washing the precipitate with pure water 2 - 4 times, placing in an oven at 45 - 60 °C and drying overnight to obtain the metal-organic framework material ZnNA; The preparation method of Fe-ZnNA includes: mixing a nicotinic acid solution and a zinc acetate solution, placing them at 25-35 °C for 0.5-2 h, then adding a substance containing Fe 2+ , stirring for 20-50 min, filtering, and drying overnight in an oven at 45-60 °C to obtain a magnetic metal-organic framework material Fe-ZnNA.

2. The method according to claim 1, characterized in that Step 1) includes: putting ZnNA or Fe-ZnNA into a tube furnace, then heating to 400 - 1000 °C at a rate of 4 - 10 °C / min in a nitrogen atmosphere, and maintaining for 1 - 3 h, then annealing to room temperature to obtain mesoporous ZnNA-derived carbon materials or magnetic mesoporous Fe-ZnNA-derived carbon materials.

3. The method according to claim 1, characterized in that The conditions for low-temperature oxidation in step 2) in an air atmosphere are: 200 - 350 °C, 1 - 3 h.

4. The method according to claim 1, characterized in that, In the preparation method of ZnNA, 100 - 300 mL of 0.154 - 0.616 mol / L nicotinic acid solution and 100 - 300 mL of 0.154 - 0.616 mol / L zinc acetate solution are mixed.

5. The method according to claim 1, characterized in that, In the preparation method of Fe-ZnNA, 100 - 300 mL of 0.154 - 0.616 mol / L nicotinic acid solution and 100 - 300 mL of 0.154 - 0.616 mol / L zinc acetate solution are mixed, placed at 25 - 35 °C for 0.5 - 3 h, and then 100 - 200 mL of 0.08 - 0.9 mol / L FeSO4 solution is added.

6. Mesoporous ZnNA-derived carbon materials prepared by the method according to any one of claims 1 - 5.

7. The mesoporous ZnNA-derived carbon material according to claim 6, wherein The average pore diameter is 6 - 12 nm.

8. Any one of the following applications of the mesoporous ZnNA-derived carbon materials described in claim 6 or 7: 1) For the field of enzyme immobilization; 2) For the high-value conversion of oils and fats.

9. A method for high-value conversion of oil and fat by immobilizing lipase with the mesoporous ZnNA-derived carbon material according to claim 6 or 7, characterized in that, The method includes: 1) Immerse the mesoporous ZnNA-derived carbon materials in absolute ethanol, then add deionized water, shake well, add free lipase, shake and mix evenly, place at 30 - 50 °C, and perform immobilization in a shaker at 100 - 400 rpm for 1 - 4 h, centrifuge to recover the immobilized enzyme, wash with deionized water, and then freeze-dry for 8 - 16 h to obtain immobilized lipase; 2) Add the immobilized lipase to a single-stage or multi-stage enzyme reactor to carry out the enzymatic conversion reaction of oils and fats.

10. The method according to claim 9, characterized in that The lipase includes lipases derived from Candida antarctica ( Candida antarctica ), thermophilic fungi ( Thermomyces lanuginosus ), Aspergillus niger ( Aspergillus niger ), Aspergillus oryzae ( Aspergillus oryzae ), Rhizomucor miehei ( Rhizomucor miehei ), Rhizopus oryzae ( Rhizopus oryzae ); The oils and fats include biodiesel, structured phospholipids, and human milk substitute fats.

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