Joint production process and application of monoglyceride and diglyceride

Abstract

The invention provides a joint production process and application of monoglyceride and diglyceride, and belongs to the technical field of organic synthesis. The joint production process comprises the following steps: (1) dissolving a reaction substrate in a certain volume of solvent, and uniformly mixing to obtain a mixed substrate; (2) transferring the mixed substrate in the step (1) into a packed bed enzyme reactor filled with immobilized lipase, and reacting under a vacuum condition to obtain a reaction product; (3) carrying out molecular distillation on the reaction product in the step (2), and separating to obtain monoglyceride and diglyceride; according to the invention, a mixed solution of tert-butyl alcohol and propylene glycol monomethyl ether is used as a solvent, a mixture of microbial lipase-candida rugosa and commercial immobilized lipase Lipozyme RM IM is used as a lipase system, and the ratio and reaction temperature are controlled, so that the mixed enzyme can show better catalytic efficiency and enzyme stability; and the monoglyceride and diglyceride with high yield and high purity are further obtained.

Description

Technical Field

[0001] The invention belongs to the technical field of organic synthesis, and in particular relates to a combined production process of monoglyceride and diglyceride and application thereof. Background Art

[0002] Oil is a mixture of triglycerides of fatty acids. It is one of the indispensable ingredients in food. It not only provides calories, but also provides essential fatty acids and various fat-soluble vitamins that cannot be synthesized by the human body and must be obtained from plant oils. However, studies have found that excessive intake of triglycerides can cause cardiovascular diseases, hypertension, obesity and other diseases in the human body. In the late 1980s, a diglycerol with similar physical and chemical properties to triglycerides was developed. Compared with triglycerides, it has two major nutritional functions: one is to inhibit the increase of serum triglycerides after meals, mainly because 1,3-diglycerol in diglycerol plays a role in reducing serum triglycerides after meals; the other is to inhibit the accumulation of fat in the body. Studies have found that compared with triglycerides, individuals who eat diglycerol have lower hunger scores, lower appetite, lower food intake, and lower desire to eat after eating. Therefore, replacing ordinary cooking oil with diglycerol will increase the oxidation rate of fat in the body and inhibit the appetite of individuals. Although monoglycerides are natural components of oils and fats, their content is relatively low, and they are also intermediate products of oil metabolism in the human body. Therefore, the functional characteristics, biochemical effects and mechanisms of monoglycerides have attracted much attention and become a hot topic in the oil and fat industry. Monoglycerides are an important polyol nonionic surfactant. As an important food emulsifier and additive, they have emulsifying, dispersing and wetting effects, and are widely used in grain and oil food, medicine, cosmetics, detergent industry and polymer synthesis. Therefore, the preparation and purification of monoglycerides are also one of the research hotspots.

[0003] Fatty acids and glycerol can be esterified under the action of a catalyst to obtain diglycerides, and monoglycerides and triglycerides can be obtained as byproducts. The preparation methods are divided into chemical methods and enzymatic methods. The chemical method has the disadvantages of high energy consumption, easy degradation of reaction products, and difficulty in separation and purification, and is not suitable for the production of glycerides. In recent years, people have begun to explore enzymatic preparation.

[0004] For example, Chinese patent CN103361387A discloses an enzymatic method for producing diglycerol for co-producing unsaturated monoglycerides, comprising the following steps: (1) adding anhydrous ethanol to natural oils and fats, and performing an ethanolysis reaction under the catalysis of immobilized lipase to make the fatty acid ethyl ester production rate 5-40% w / w, removing the remaining ethanol, and obtaining an alcoholysis product; (2) adding 5-15% w / w glycerol to the alcoholysis product of step (1), and then reacting the above mixed substrate in a packed bed enzyme reactor equipped with immobilized lipase under vacuum conditions; (3) the reaction product of step (2) is subjected to molecular distillation, and the high-temperature fraction obtained by separation is a product containing diglycerol, and a low-temperature fraction mainly composed of monoglycerides is obtained. The reaction system of the invention does not need to add water, thereby solving the problem of product acid value, so that monoglycerides can be co-produced; and glycerol does not need to be pre-adsorbed, simplifying the production operation.

[0005] For example, Chinese patent CN103074388A discloses a method for synthesizing monoglycerides and diglycerides by lipase catalysis under ultra-high pressure, comprising the following steps: (1) dissolving the reaction substrate in a certain volume of solvent, adding a certain amount of lipase, and mixing evenly; (2) placing the substrate in an ultra-high pressure device for enzyme catalysis reaction to obtain a fatty acid and glyceride mixture; (3) separating and purifying the mixture obtained in step (2) by molecular distillation technology to obtain monoglycerides and diglycerides. The method for synthesizing monoglycerides and diglycerides by lipase catalysis under ultra-high pressure of the invention has mild reaction conditions, high conversion rate, and low enzyme activity loss, which is conducive to shortening the reaction time and reducing production costs, and is suitable for large-scale promotion and application.

[0006] However, although the existing preparation method achieves the co-production of monoglyceride and diglyceride, the yield and purity of the monoglyceride and diglyceride finally obtained cannot better meet the demand. Therefore, it is necessary to develop a co-production process with relatively high yield and purity of monoglyceride and diglyceride. Summary of the invention

[0007] Based on the deficiencies in the prior art, the present invention aims to provide a combined production process and application of monoglycerides and diglycerides. The present invention improves the speed of the enzymatic hydrolysis reaction by selecting lipase and solvent so that the substrate has a higher solubility in the solvent.

[0008] In order to achieve the above object, the present invention adopts the following technical solutions: The present invention provides a combined production process of monoglyceride and diglyceride, comprising the following steps: (1) dissolving the reaction substrate in a solvent and mixing them evenly to obtain a mixed substrate; (2) transferring the mixed substrate of step (1) into a packed bed enzyme reactor containing immobilized lipase, reacting under vacuum conditions to obtain a reaction product; (3) The reaction product of step (2) is subjected to molecular distillation to separate monoglyceride and diglyceride.

[0009] The reaction substrate in the above step (1) is a mixture of oil and glycerol; Wherein, the oil is selected from vegetable oil or animal oil; The animal fat is selected from one or more of fish oil, lard, beef tallow and chicken fat; The vegetable oil is selected from one or more of soybean oil, rapeseed oil, corn oil, palm oil, peanut oil, sunflower oil, camellia oil, olive oil and rice bran oil; preferably olive oil and / or rice bran oil.

[0010] The molar ratio of the oil to glycerol is 1:2-6; preferably 1:3-5; more preferably 1:3-4.

[0011] The solvent in the above step (1) is a mixture of tert-butyl alcohol and propylene glycol methyl ether; Preferably, the volume ratio of tert-butyl alcohol to propylene glycol methyl ether is 60-80:20-40; More preferably, the volume ratio of tert-butyl alcohol to propylene glycol methyl ether is 70:30.

[0012] The purpose of adding an organic solvent as a reaction medium is to increase the solubility of the substrate and solve the problem that glycerol and oil are difficult to dissolve each other. Tert-butyl alcohol has been proven to be a suitable solvent medium, but the freezing point of tert-butyl alcohol is too high, and its application is limited. In the implementation process of the present invention, propylene glycol methyl ether is added to tert-butyl alcohol, and the volume ratio of tert-butyl alcohol and propylene glycol methyl ether is controlled to be 60-80:20-40, so that the freezing point of the mixed solvent is reduced, and no obvious effect is produced on the reaction.

[0013] The mass volume ratio of the reaction substrate to the solvent is 0.2-0.5:1 (g / ml); Preferably, the mass volume ratio of the reaction substrate to the solvent is 0.3-0.4:1 (g / ml); More preferably, the mass volume ratio of the reaction substrate to the solvent is 0.35:1 (g / ml).

[0014] The immobilized lipase in step (2) is selected from one or more of the esterases of Candida cylindrica, Candida rugosa and Chromobacterium marcescens; The immobilized lipase further comprises Lipozyme TL IM and / or Lipozyme RM IM; Preferably, the lipase is a mixture of Candida rugosa and Lipozyme RM IM; More preferably, the mass ratio of the Candida rugosa to Lipozyme RM IM is 3-5:1; preferably 4:1.

[0015] In the implementation process of the present invention, microbial lipase-Candida rugosa and commercial immobilized lipase Lipozyme RM IM are mixed, and their ratio is controlled so that they can be more suitable for the solvent system under the condition of relatively small addition amount, and can show better catalytic efficiency and enzyme stability.

[0016] It is well known in the art that the hydrogen bonds formed between the water molecules and the functional groups of the enzyme molecules in the aqueous solution will destroy the hydrogen bonds in the enzyme molecules. The enzyme molecules are loosened and in an "on" state, so that the enzyme molecules are in a dynamic balance of "tight" and "on", showing a certain flexibility, thereby exerting a catalytic effect. On the contrary, in organic solvents, due to the extremely few water molecules, the hydrogen bonds formed by the enzyme molecules are extremely few, so the enzyme molecule hydrogen bonds play a leading role, making the enzyme molecular structure "rigid", and this rigidity limits the transformation of the enzyme molecular structure under the hydrophobic environment, and can maintain the same structure and conformation as in water, showing catalytic activity. Its catalytic activity is closely related to the water content in the organic solvent reaction system. Only when the optimal balance point is reached between the rigidity and thermodynamic stability of the enzyme molecular structure at the optimum water content, the enzyme will show maximum activity. When the water content is lower than the optimum water content, the enzyme conformation is too "rigid", and it is reduced or even loses activity; when the water content is too high, the flexibility of the enzyme structure is too large, causing changes in the enzyme structure and even inactivation.

[0017] During the implementation of the present invention, it was found that the enzyme activity was best when the water content in the reaction system was 0.5-4.0% of the mass of glycerol; preferably, the water content of the reaction system was 1.5-2.5% of the mass of glycerol; further preferably, the water content of the reaction system was 2.0% of the mass of glycerol.

[0018] The amount of the immobilized lipase added in the above step (2) is 1-5% of the mass of the oil, preferably 2-4%, and more preferably 2.5%.

[0019] The reaction temperature in the above step (2) is 50-60°C, preferably 55°C, and the reaction time is 2-4 hours, preferably 3 hours.

[0020] The molecular distillation described in step (3) is carried out in five stages: the first stage molecular distillation is used for degassing; the second stage molecular distillation is used for removing glycerol; the third stage molecular distillation is used for removing free fatty acids; the fourth stage molecular distillation is used to obtain monoglycerides; the fifth stage molecular distillation is used to remove triglycerides to obtain diglycerides; As some preferred implementation schemes, the temperature of the first-stage molecular distillation is 50-90°C, and the vacuum degree is 1000-2000Pa; the temperature of the second-stage molecular distillation is 100-150°C, and the vacuum degree is 50-100Pa; the temperature of the third-stage molecular distillation is 160-180°C, and the vacuum degree is 1-50Pa; the temperature of the fourth-stage molecular distillation is 180-220°C, and the vacuum degree is 1-50Pa; the temperature of the fifth-stage molecular distillation is 200-220°C, and the vacuum degree is 0.5-20Pa.

[0021] As a preferred embodiment, the temperature of the first-stage molecular distillation is 65°C and the vacuum degree is 1500Pa; the temperature of the second-stage molecular distillation is 110°C and the vacuum degree is 65Pa; the temperature of the third-stage molecular distillation is 165°C and the vacuum degree is 2.5Pa; the temperature of the fourth-stage molecular distillation is 180°C and the vacuum degree is 2.5Pa; the temperature of the fifth-stage molecular distillation is 210°C and the vacuum degree is 1.5Pa.

[0022] When the molecular distillation described in the present invention separates the same components, the distillation can be cyclically performed 2-3 times under the same conditions.

[0023] The reaction product of step (3) mainly includes triglycerides, diglycerides, monoglycerides, fatty acid ethyl esters, and glycerol. The present invention uses monoglycerides and diglycerides as target products, and the purpose of separation is to obtain relatively pure monoglycerides and diglycerides. The commonly used separation method is molecular distillation, which is characterized by being carried out under vacuum conditions, which can greatly reduce the temperature required for separation.

[0024] The present invention also provides a monoglyceride or diglyceride prepared by the combined production process.

[0025] The present invention also provides an application of the combined production process in preparing monoglycerides or diglycerides and in preparing foods and medicines.

[0026] Compared with the prior art, the present invention has the following beneficial effects: (1) The present invention uses a mixed solution of tert-butyl alcohol and propylene glycol methyl ether as a solvent, and controls the volume ratio of the two to be 60-80:20-40. The mixed solvent can not only achieve the mutual solubility of glycerol and oil, but also reduce the melting point of tert-butyl alcohol, and at the same time will not have a significant effect on the reaction; (2) During the implementation process, it was unexpectedly discovered that a small amount of water in the reaction system can better promote enzyme activity. The enzyme activity is best when the water content in the system is 0.5-4.0% of the mass of glycerol. (3) In the implementation process of the present invention, a mixture of microbial lipase - Candida rugosa and commercial immobilized lipase Lipozyme RM IM is used. By controlling their ratio, the mixture can be more suitable for the solvent system under the condition of a relatively small addition amount, and can show better catalytic efficiency and enzyme stability.

[0027] (4) The preparation method of the present invention is simple and can simultaneously obtain monoglycerides and diglycerides with high yield and high purity. DETAILED DESCRIPTION

[0028] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0029] The experimental methods used in the following examples are all conventional methods unless otherwise specified; the water used is all deionized water; the materials, reagents, etc. used are all commercially available unless otherwise specified.

[0030] Example 1 A combined production process of monoglycerides and diglycerides The steps include: (1) dissolving olive oil and glycerol in a molar ratio of 1:1 in a mixed solution of tert-butyl alcohol and propylene glycol methyl ether in a volume ratio of 60:40, and mixing them evenly to obtain a mixed substrate; the mass ratio of olive oil and glycerol to the volume ratio of tert-butyl alcohol and propylene glycol methyl ether is 0.2:1 (g / ml); (2) The mixed substrate of step (1) is transferred into a packed bed enzyme reactor containing immobilized lipase, wherein the water content is 0.5% of the mass of glycerol, and the reaction is carried out under vacuum conditions at a temperature of 50° C. for 4 hours to obtain a reaction product; the immobilized fat is a mixture of Candida rugosa and Lipozyme RM IM in a mass ratio of 3:1; and the amount of immobilized lipase added is 1% of the mass of the fat; (3) The reaction product of step (2) is subjected to molecular distillation in five stages, wherein the temperature of the first stage molecular distillation is 50°C and the vacuum degree is 1000Pa for degassing; the temperature of the second stage molecular distillation is 100°C and the vacuum degree is 50Pa for removing glycerol; the temperature of the third stage molecular distillation is 160°C and the vacuum degree is 10Pa for removing free fatty acids; the temperature of the fourth stage molecular distillation is 180°C and the vacuum degree is 10Pa for obtaining monoglycerides; the temperature of the fifth stage molecular distillation is 200°C and the vacuum degree is 2Pa for removing triglycerides to obtain diglycerides.

[0031] Example 2 A combined production process of monoglycerides and diglycerides The steps include: (1) dissolving olive oil and glycerol in a molar ratio of 1:3 in a mixed solution of tert-butyl alcohol and propylene glycol methyl ether in a volume ratio of 80:20, and mixing them evenly to obtain a mixed substrate; the mass ratio of olive oil and glycerol to the volume ratio of tert-butyl alcohol and propylene glycol methyl ether is 0.5:1 (g / ml); (2) The mixed substrate of step (1) is transferred into a packed bed enzyme reactor containing immobilized lipase, the water content is 4.0% of the mass of glycerol, and the reaction is carried out under vacuum conditions at a temperature of 60° C. for 2 hours to obtain a reaction product; the immobilized fat is a mixture of Candida rugosa and Lipozyme RM IM in a mass ratio of 5:1; the amount of immobilized lipase added is 5% of the mass of the fat; (3) The reaction product of step (2) is subjected to molecular distillation in five stages, wherein the temperature of the first stage molecular distillation is 90°C and the vacuum degree is 2000Pa for degassing; the temperature of the second stage molecular distillation is 150°C and the vacuum degree is 100Pa for removing glycerol; the temperature of the third stage molecular distillation is 180°C and the vacuum degree is 50Pa for removing free fatty acids; the temperature of the fourth stage molecular distillation is 220°C and the vacuum degree is 50Pa for obtaining monoglycerides; the temperature of the fifth stage molecular distillation is 220°C and the vacuum degree is 20Pa for removing triglycerides to obtain diglycerides.

[0032] Example 3 A combined production process of monoglycerides and diglycerides The steps include: (1) dissolving olive oil and glycerol in a molar ratio of 1:2 in a mixed solution of tert-butyl alcohol and propylene glycol methyl ether in a volume ratio of 70:30, and mixing them evenly to obtain a mixed substrate; the mass ratio of olive oil and glycerol to the volume ratio of tert-butyl alcohol and propylene glycol methyl ether is 0.35:1 (g / ml); (2) The mixed substrate of step (1) is transferred into a packed bed enzyme reactor containing immobilized lipase, the water content is 1.0% of the mass of glycerol, and the reaction is carried out under vacuum conditions at a temperature of 55° C. for 3 hours to obtain a reaction product; the immobilized fat is a mixture of Candida rugosa and Lipozyme RM IM in a mass ratio of 4:1; the amount of immobilized lipase added is 2% of the mass of the fat; (3) The reaction product of step (2) is subjected to molecular distillation in five stages, wherein the temperature of the first stage molecular distillation is 65°C and the vacuum degree is 1500Pa for degassing; the temperature of the second stage molecular distillation is 110°C and the vacuum degree is 65Pa for removing glycerol; the temperature of the third stage molecular distillation is 165°C and the vacuum degree is 2.5Pa for removing free fatty acids; the temperature of the fourth stage molecular distillation is 180°C and the vacuum degree is 2.5Pa for obtaining monoglycerides; the temperature of the fifth stage molecular distillation is 210°C and the vacuum degree is 1.5Pa for removing triglycerides to obtain diglycerides.

[0033] Comparative Example 1 The difference from Example 3 is that in step (2), only Lipozyme RM IM is used as the immobilized lipase, and the rest is the same as Example 3.

[0034] Comparative Example 2 The difference from Example 3 is that in step (2), the mass ratio of Candida rugosa to Lipozyme RM IM is 1:1, and the rest is the same as Example 3.

[0035] Comparative Example 3 The difference from Example 3 is that no water is added to the reaction system in step (2), and the rest is the same as Example 3.

[0036] Comparative Example 4 The difference from Example 3 is that the water content in the reaction system of step (2) is 5% of the mass of glycerol, and the rest is the same as Example 3.

[0037] Comparative Example 5 The difference from Example 3 is that in step (1), propylene glycol methyl ether is replaced by tert-amyl alcohol, and the rest is the same as Example 3.

[0038] Performance data: The content and purity of monoglyceride and diglyceride were tested by gas chromatography, and the specific results are shown in the national food safety standard GB1886.65. The test results are shown in Table 1 below.

[0039] Table 1

[0040] According to the test results in Table 1 above, the combined production process provided by the present invention can achieve the separation and purification of monoglycerides and diglycerides, and the obtained monoglycerides can meet the condition that the purity is greater than 90% and the yield is higher than 38%, especially the purity of monoglycerides in Example 3 can reach 96%, and the yield is 46.3%; the obtained diglycerides can meet the condition that the purity is greater than 85% and the yield is higher than 32%, especially the purity of diglycerides in Example 3 can reach 92.4%, and the yield is 34.4%.

[0041] In Comparative Example 1, only microbial lipase-Candida rugosa was used to catalyze the reaction system, which would affect the catalytic efficiency to a certain extent, and thus affect the production of monoglycerides and diglycerides. According to the test results in Table 1 above, it can be seen that when the purity is equivalent to that of the embodiment, the yield of monoglycerides and diglycerides is significantly reduced.

[0042] Comparative Example 2 Changing the mass ratio of the two enzymes, namely Candida rugosa and Lipozyme RM IM, will also affect the catalytic efficiency of the enzyme to a certain extent, and thus affect the production of monoglycerides and diglycerides. According to the test results in Table 1 above, it can be seen that when the purity is equivalent to that of the embodiment, the yield of monoglycerides and diglycerides is increased compared with the yield of Comparative Example 1, indicating that the mixed use of the two enzymes has a higher catalytic efficiency than that of a single enzyme, but the yield is significantly lower than that of the embodiment, indicating that the mass ratio of the two enzymes has a significant effect on the catalytic efficiency.

[0043] In Comparative Example 3, no water was added to the reaction system during the reaction, which made the enzyme conformation too "rigid", which would affect the catalytic activity of the enzyme to a certain extent, reduce the catalytic effect of the enzyme, and further reduce the yield of monoglycerides and diglycerides.

[0044] In Comparative Example 4, too much water was added to the reaction system, which made the enzyme structure in the reaction system too flexible, causing changes in the enzyme structure and even inactivation, thereby reducing the yields of monoglycerides and diglycerides.

[0045] In Comparative Example 5, the change in the type of solvent will affect the freezing point of the reaction system to a certain extent, thereby affecting the yield of monoglyceride and diglyceride, causing the yield to decrease to a certain extent.

[0046] In summary, it can be seen that by adopting the co-production process provided by the present invention, monoglycerides and diglycerides with high yield and high purity can be obtained simultaneously.

[0047] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A combined production process of monoglycerides and diglycerides, characterized in that: The steps include: (1) dissolving the reaction substrate in a solvent and mixing them evenly to obtain a mixed substrate; (2) transferring the mixed substrate of step (1) into a packed bed enzyme reactor containing immobilized lipase, reacting under vacuum conditions to obtain a reaction product; (3) The reaction product of step (2) is subjected to molecular distillation to separate monoglyceride and diglyceride; The solvent in step (1) is a mixture of tert-butyl alcohol and propylene glycol methyl ether; the volume ratio of the two is 60-80:20-40; The immobilized lipase in step (2) is a mixture of Candida rugosa and Lipozyme RM IM; the mass ratio is 3-5:

1.

2. The combined production process according to claim 1, characterized in that: The mass volume ratio of the reaction substrate to the solvent is 0.2-0.5 g:1 mL.

3. The combined production process according to claim 1, characterized in that: The reaction substrate in step (1) also contains water; the content of the water is 0.5-4.0% of the mass of the glycerol.

4. The combined production process according to claim 3, characterized in that: The water content is 2.0% of the glycerol mass.

5. The combined production process according to claim 1, characterized in that: The added amount of the immobilized lipase is 1-5% of the mass of the oil.

6. The combined production process according to claim 1, characterized in that: The reaction temperature in step (2) is 50-60°C and the reaction time is 2-4 hours.

7. The combined production process according to claim 1, characterized in that: The molecular distillation described in step (3) is carried out in five stages. The first stage molecular distillation is used for degassing, the distillation temperature is 50-90°C, and the vacuum degree is 1000-2000Pa; the second stage molecular distillation is used for removing glycerol, the distillation temperature is 100-150°C, and the vacuum degree is 50-100Pa; the third stage molecular distillation is used for removing free fatty acids, the distillation temperature is 160-180°C, and the vacuum degree is 1-50Pa; the fourth stage molecular distillation is used to obtain monoglyceride, the distillation temperature is 180-220°C, and the vacuum degree is 1-50Pa; the fifth stage molecular distillation is used to remove triglyceride, the distillation temperature is 200-220°C, and the vacuum degree is 0.5-20Pa to obtain diglyceride.

8. The combined production process according to claim 7, characterized in that: The temperature of the first-stage molecular distillation is 65°C, and the vacuum degree is 1500Pa; the temperature of the second-stage molecular distillation is 110°C, and the vacuum degree is 65Pa; the temperature of the third-stage molecular distillation is 165°C, and the vacuum degree is 2.5Pa; the temperature of the fourth-stage molecular distillation is 180°C, and the vacuum degree is 2.5Pa; the temperature of the fifth-stage molecular distillation is 210°C, and the vacuum degree is 1.5Pa.

9. A monoglyceride or diglyceride prepared by the combined production process according to any one of claims 1 to 8.

10. The diglyceride prepared by the combined production process according to any one of claims 1 to 8 or the use of the diglyceride in food and medicine.

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