Method for separating and purifying linolenic acid by using composite membrane

Abstract

The invention discloses a method for separating and purifying linolenic acid by using a composite membrane, and relates to the technical field of substance separation.The method comprises the steps that pretreated grease and ethyl alcohol serve as raw materials, and ethyl linolenate is prepared through an immobilized lipase catalytic esterification reaction; intercepting lipase and macromolecular impurities by using an ultrafiltration membrane; separating an oil phase from a water phase by adopting a modified PVDF (Polyvinylidene Fluoride) microfiltration membrane; ethyl linolenate and raw materials are separated based on polarity difference through a PDMS / TiO2 composite membrane; and removing residual ethanol by molecular distillation to obtain a high-purity ethyl linolenate product. PVP is added into the modified membrane, so that the pore diameter and the porosity of the membrane are increased, and the flux of the membrane is favorably improved; a proper amount of SDS is added into a membrane casting solution, so that the hydrophilicity of the surface of the membrane is improved, and the separation effect of the membrane on a water phase and an oil phase is improved.

Description

Technical Field

[0001] The invention belongs to the technical field of material separation, and in particular relates to a method for separating and purifying linolenic acid by utilizing a composite membrane. Background Art

[0002] Ethyl linolenate is a derivative of linolenic acid, sharing the same physiological effects as linolenic acid. Linolenic acid, an essential ω-3 fatty acid, can form the physiologically active EPA and DHA in the body. Long-term consumption of products containing linolenic acid or its derivatives can suppress allergies and prevent cardiovascular and cerebrovascular diseases. Ethyl linolenate also has greater antioxidant stability than linolenic acid, making linolenic acid in its ethyl linolenate form easier to store and transport than pure linolenic acid. Furthermore, ethyl linolenate is an intermediate in the preparation of many pharmaceuticals and other organic chemicals, leading to a high market demand for higher-purity ethyl linolenate.

[0003] In the industrial production of ethyl linolenate, traditional methods such as distillation are inefficient and energy-intensive; extraction methods are prone to introducing impurities; and column chromatography methods are costly and difficult to scale up. Membrane separation technology, due to its advantages such as high efficiency, energy conservation, and environmental friendliness, has garnered widespread attention in the field of material separation. However, conventional single membrane materials suffer from low flux, poor selectivity, and contamination when used to separate ethyl linolenate, making them inadequate for the high-purity, large-scale separation and purification of ethyl linolenate required for industrial production. To address these challenges, the following solutions are proposed. Summary of the Invention

[0004] The purpose of the present invention is to provide a method for separating and purifying linolenic acid using a composite membrane.

[0005] To solve the above technical problems, the present invention is achieved through the following technical solutions: The present invention provides a method for separating and purifying linolenic acid using a composite membrane. The purification method comprises the following steps: Step S1, Preparation of Ethyl Linolenate: Pretreated oil and ethanol are added to a reaction vessel at a certain molar ratio and stirred thoroughly to evenly distribute the substrate; immobilized lipase is added to the reaction system at a certain enzyme amount and stirred continuously to ensure full contact between the enzyme and the substrate, thereby initiating the esterification reaction; Step S2, lipase separation: the mixture obtained in step S1 is passed through an ultrafiltration membrane of a certain molecular weight under pressure to intercept the lipase and some impurities; Step S3, two-phase separation: the filtrate of step S2 is passed through a modified PVDF membrane to separate the aqueous phase and the oil phase; Step S4, separation of ethyl linolenate: the separated oil phase is passed through a composite membrane prepared from PDMS under pressure to obtain ethyl linolenate of a certain purity; Step S5, molecular distillation: the residual ethanol is subjected to molecular distillation to obtain the final product ethyl linolenate.

[0006] Furthermore, in step S1, the pretreated oil refers to a plant oil rich in linolenic acid, such as linseed oil, perilla oil, or peony seed oil, as the starting material. The raw oil is filtered to remove solid impurities and then dried using vacuum distillation or molecular sieve adsorption to ensure that the moisture content in the raw oil is less than 0.5% (mass fraction) to prevent moisture from adversely affecting the subsequent enzyme-catalyzed reaction. The molar ratio of oil to ethanol is preferably between 3:1 and 6:1, and the co-solvent is 20% tert-butanol.

[0007] Furthermore, in step S1, the lipase is Novozym 435 lipase, and the enzymatic reaction is carried out by using a glycerol pre-adsorption method and resin enzyme immobilization. The enzymatic reaction specifically includes the following steps: Novozym 435 lipase was mixed with a 15% glycerol solution at a ratio of 5% to 20%, and adsorbed in a pH 7.0 phosphate buffer at 25°C with shaking for 6 hours. The lipase was washed with n-hexane and then dried in vacuo. The lipase was immobilized with a tertiary amino resin to obtain an immobilized lipase pre-adsorbed with glycerol. The immobilized lipase treated with pre-adsorption of glycerol is mixed with a mixture of oil and ethanol, wherein the addition amount of the immobilized lipase is 5% to 10% of the total weight of the vegetable oil, and the enzymatic esterification reaction is carried out under stirring conditions of 40°C to 55°C and 200r / min to 300r / min.

[0008] Furthermore, in step S1, the amount of immobilized lipase added is 3% to 8% of the total weight of the mixed raw materials, the reaction temperature is adjusted to 35°C to 55°C, the carbon dioxide pressure is adjusted to 10 MPa to 25 MPa, and the stirring speed is 150 r / min to 300 r / min, and the enzyme-catalyzed transesterification reaction is carried out for 4 to 8 hours.

[0009] Furthermore, in step S2, the pore size of the ultrafiltration membrane is 10 kDa-30 kDa, the applied pressure is generally 0.1 MPa-0.2 MPa, and the filtration operation temperature is preferably controlled at 25°C-40°C.

[0010] Furthermore, in step S3, the modified membrane is a polyvinylidene fluoride (PVDF) microfiltration membrane modified with polyvinyl pyrrolidone (PVP) porogen and surfactant sodium dodecyl sulfate (SDS), and the pore size of the membrane is 0.1 μm-10 μm.

[0011] Furthermore, in step S4, the composite membrane is a functional membrane prepared by composite of polydimethylsiloxane (PDMS) and modified nano-titanium dioxide (TiO2).

[0012] Furthermore, in step S5, the temperature of the molecular distillation evaporator is controlled between 120° C. and 150° C., and the specific temperature needs to be adjusted according to the actual separation effect.

[0013] The present invention has the following beneficial effects: 1. The content of PVP added to the modified membrane of the present invention affects the pore size and porosity of the membrane. As the PVP content increases, the pore size and porosity of the membrane increase, which is beneficial to improving the flux of the membrane. When an appropriate amount of SDS is added to the casting solution, the SDS molecules are enriched on the membrane surface during the membrane formation process, thereby improving the hydrophilicity of the membrane surface. The increase in hydrophilicity helps the water phase to wetting and penetrate the membrane surface, while reducing the adsorption of the oil phase on the membrane surface, thereby improving the separation effect of the membrane on the water phase and the oil phase.

[0014] 2. In the step of the glycerol pre-adsorption enzyme reaction of the present invention, glycerol pre-adsorption can form a hydration protective layer on the enzyme surface, reduce the direct impact of the substrate or solvent on the enzyme active center, maintain the natural conformation of the enzyme, and thus improve the stability of the enzyme; the presence of glycerol can regulate the micro-water environment around the enzyme. Enzymatic reactions usually require a small amount of water to maintain activity, but excess water may promote side reactions; the pre-adsorbed glycerol combines with water to form a stable "enzyme-glycerol-water" complex, providing the enzyme with appropriate water activity, which not only ensures enzyme activity but also inhibits hydrolysis side reactions. At the same time, the glycerol pre-adsorbed enzyme can be more effectively dispersed in the oil phase or the oil-water interface.

[0015] 3. The ultrafiltration membrane of the present invention can intercept lipase and separate it from low-molecular-weight substrates or products, thereby realizing the reuse of the enzyme and reducing costs. After being filtered through the ultrafiltration membrane, the immobilized lipase can be reused 5 to 10 times, and the enzyme activity retention rate can reach more than 80%. In addition, the membrane filtration process is gentle, avoiding mechanical damage to the enzyme structure caused by traditional methods such as centrifugation and precipitation, and reducing the risk of enzyme inactivation due to physical effects. The membrane reactor can realize the simultaneous enzymatic reaction and product separation: the substrate continuously flows into the reactor, the product flows out with the filtrate, and the enzyme is retained in the reactor to continue catalysis, thereby improving production efficiency.

[0016] 4. The PDMS of the present invention is a polymer material with good flexibility and chemical stability. Its molecular chain has high flexibility and can form a relatively loose network structure. This structure gives the membrane certain air permeability and liquid permeability. At the same time, PDMS itself has certain hydrophobicity and has good affinity for non-polar substances; nano-TiO2 has a large specific surface area and strong adsorption performance; the modified TiO2 can better combine with PDMS, improving the stability and performance of the composite membrane; at the same time, the modified TiO2 can adjust the pore size and distribution of the membrane, and exhibit different adsorption characteristics for substances of different polarities; the polarity difference is utilized to better separate ethyl linolenate and linolenic acid.

[0017] Of course, any product implementing the present invention does not necessarily need to achieve all of the advantages described above at the same time. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for describing the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0019] Figure 1 The present invention is a schematic flow chart of a method for separating and purifying linolenic acid using a composite membrane. DETAILED DESCRIPTION

[0020] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. 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 making any creative efforts shall fall within the scope of protection of the present invention.

[0021] Example 1 See also Figure 1 As shown, the present invention is a method for separating and purifying linolenic acid using a composite membrane, comprising the following steps: Step S1, Preparation of Ethyl Linolenate: Linseed oil was used as the starting material and filtered using a plate and frame filter to remove solid impurities. The oil was then dried using vacuum distillation at 50 Pa and 60°C for 2 hours to reduce the moisture content of the oil to less than 0.5%. 1g of Novozym 435 lipase was added to 10mL of a 15% glycerol solution (prepared in pH 7.0 phosphate buffer) and shaken at 25°C for 6 hours. After filtration, the lipase was washed three times with n-hexane and dried under vacuum. The lipase was then immobilized using a tertiary amino resin. Linseed oil and anhydrous ethanol were added to the co-solvent tert-butanol (20% of the total volume) at a molar ratio of 6:1. The immobilized enzyme was then added at 10% by weight of the oil. The mixture was stirred at 40°C and 200 rpm for 8 hours.

[0022] Step S2, lipase separation: the reaction mixture is passed through an ultrafiltration membrane with a pore size of 25 kDa, while applying a pressure of 0.1 MPa and controlling the filtration operation temperature at 30° C. to separate the lipase from the liquid.

[0023] Step S3, two-phase separation: The filtrate from step S2 is passed through a polyvinylidene fluoride (PVDF) microfiltration membrane modified with a polyvinyl pyrrolidone (PVP) porogen and a surfactant, sodium dodecyl sulfate (SDS), with a pore size of 10 μm, while applying a pressure of 0.2 MPa to separate the oil phase and the aqueous phase; Step S4, separation of ethyl linolenate: The separated oil phase is passed through a functional membrane prepared by a composite of polydimethylsiloxane (PDMS) and modified nano-titanium dioxide (TiO2) at a pressure of 0.2 MPa. The polarity difference is utilized to separate linolenic acid and ethyl linolenate. Ethyl linolenate preferentially passes through the membrane and enters the collection device, while most of the linolenic acid is retained.

[0024] Step S5, molecular distillation: slowly add the pretreated ethyl linolenate mixture into the evaporator of the molecular distillation device. The temperature of the evaporator is controlled at 150° C. to evaporate the residual alcohol and finally obtain the finished product ethyl linolenate.

[0025] Example 2 A method for separating and purifying linolenic acid using a composite membrane comprises the following steps: Step S1, Preparation of Ethyl Linolenate: Perilla seed oil was used as the starting material. Solid impurities were removed from the crude oil using plate and frame filtration. The crude oil was then dried under reduced pressure distillation at 60 Pa and 65°C for 2 hours to reduce the moisture content to less than 0.5%. 1 g of Novozym 435 lipase was added to 15 mL of a 15% glycerol solution (prepared in pH 7.0 phosphate buffer) and shaken at 25°C for 6 hours. After filtration, the mixture was washed three times with n-hexane and dried under vacuum. The lipase was then immobilized using a tertiary amino resin. Perilla seed oil and anhydrous ethanol were added to the co-solvent tert-butanol (20% of the total volume) at a molar ratio of 5:1. The immobilized enzyme was then added at 10% by weight of the oil. The mixture was stirred at 45°C and 200 rpm for 8 hours.

[0026] Step S2, lipase separation: the reaction mixture is passed through an ultrafiltration membrane with a pore size of 15 kDa, while applying a pressure of 0.2 MPa and controlling the filtration operation temperature at 30° C. to separate the lipase from the liquid.

[0027] Step S3, two-phase separation: The filtrate from step S2 is passed through a polyvinylidene fluoride (PVDF) microfiltration membrane modified with a polyvinyl pyrrolidone (PVP) porogen and a surfactant, sodium dodecyl sulfate (SDS), with a pore size of 20 μm, while applying a pressure of 0.2 MPa to separate the oil phase and the aqueous phase; Step S4, separation of ethyl linolenate: The separated oil phase is passed through a functional membrane prepared by a composite of polydimethylsiloxane (PDMS) and modified nano-titanium dioxide (TiO2) at a pressure of 0.1 MPa. The polarity difference is used to separate linolenic acid and ethyl linolenate. Ethyl linolenate preferentially passes through the membrane and enters the collection device, while most of the linolenic acid is retained.

[0028] Step S5, molecular distillation: slowly add the pretreated ethyl linolenate mixture into the evaporator of the molecular distillation device. The temperature of the evaporator is controlled at 150° C. to evaporate the residual alcohol and finally obtain the finished product ethyl linolenate.

[0029] Example 3 A method for separating and purifying linolenic acid using a composite membrane comprises the following steps: Step S1, Preparation of Ethyl Linolenate: Peony seed oil was used as the starting material. Solid impurities were removed from the crude oil using plate and frame filtration. The crude oil was then dried under reduced pressure distillation at 60 Pa and 60°C for 2 hours to reduce the moisture content to less than 0.5%. 1 g of Novozym 435 was added to 15 mL of a 15% glycerol solution (prepared in pH 7.0 phosphate buffer) and shaken at 25°C for 6 hours. The mixture was filtered, washed three times with n-hexane, and dried under vacuum. Lipase was then immobilized using a tertiary amino resin. Peony seed oil and anhydrous ethanol were added to the co-solvent tert-butyl alcohol (20% of the total volume) in a ratio of 4:1. The immobilized enzyme was then added at 10% by weight of the oil. The mixture was stirred at 55°C and 200 rpm for 8 hours.

[0030] Step S2, lipase separation: the reaction mixture is passed through an ultrafiltration membrane with a pore size of 30 kDa, while applying a pressure of 0.2 MPa and controlling the filtration operation temperature at 30° C. to separate the lipase from the liquid.

[0031] Step S3, two-phase separation: The filtrate from step S2 is passed through a polyvinylidene fluoride (PVDF) microfiltration membrane modified with a polyvinyl pyrrolidone (PVP) porogen and a surfactant, sodium dodecyl sulfate (SDS), with a pore size of 20 μm, while applying a pressure of 0.2 MPa to separate the oil phase and the aqueous phase; Step S4, separation of ethyl linolenate: The separated oil phase is passed through a functional membrane prepared by a composite of polydimethylsiloxane (PDMS) and modified nano-titanium dioxide (TiO2) at a pressure of 0.2 MPa. The polarity difference is utilized to separate linolenic acid and ethyl linolenate. Ethyl linolenate preferentially passes through the membrane and enters the collection device, while most of the linolenic acid is retained.

[0032] Step S5, molecular distillation: slowly add the pretreated ethyl linolenate mixture into the evaporator of the molecular distillation device. The temperature of the evaporator is controlled at 150° C. to evaporate the residual alcohol and finally obtain the finished product ethyl linolenate.

[0033] Comparative Example 1 On the basis of Example 1, the two-phase separation membrane in step S3 is replaced by a polyvinylidene fluoride (PVDF) microfiltration membrane, and the ethyl linolenate separation membrane is replaced by a polydimethylsiloxane (PDMS) filter membrane.

[0034] Comparative Example 2 On the basis of Example 2, the two-phase separation membrane in step S3 is replaced by a 0.1 μm aluminum oxide (Al2O3) base membrane, and the ethyl linolenate separation membrane is a functional membrane prepared by combining polydimethylsiloxane (PDMS) and modified nano-titanium dioxide (SiO2).

[0035] Comparative Example 3 On the basis of Example 3, the two-phase separation membrane in step S3 is a polytetrafluoroethylene (PTFE) membrane, and the ethyl linolenate separation membrane is a cellulose acetate membrane.

[0036] Ethyl linolenate was prepared using the methods of the Examples and Comparative Examples, and then its purity was tested. The test results are shown in Table 1: project Ethyl linolenate content% Example 1 75.34 Comparative Example 1 50.18 Example 2 70.25 Comparative Example 2 40.51 Example 3 88.25 Comparative Example 3 68.23 From the analysis of Implementation Case 1 and Comparative Case 1, the functional composite membrane has better separation effect than the single composite membrane; Implementation Case 1 and Comparative Case 2 analyzed that adding PVP pore-forming agent can optimize the hydrophilic-hydrophobic balance of the membrane, enhance selectivity, and better separate the oil phase and the water phase; Implementation Case 3 and Comparative Case 3 analyzed that the composite membrane prepared by PDMS and TiO2 has a better promoting effect on the separation of ethyl linolenate.

[0037] Throughout this specification, references to terms such as "one embodiment," "example," or "specific example" indicate that the specific features, structures, materials, or characteristics described in conjunction with that embodiment or example are included in at least one embodiment or example of the present invention. In this specification, schematic representations of these terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.

[0038] The preferred embodiments of the present invention disclosed above are intended only to help illustrate the present invention. These preferred embodiments do not exhaustively describe all details, nor do they limit the present invention to the specific embodiments described. Obviously, many modifications and variations are possible based on the content of this specification. These embodiments are selected and described in detail in this specification to better explain the principles and practical applications of the present invention, thereby enabling those skilled in the art to better understand and utilize the present invention. The present invention is limited only by the claims and their full scope and equivalents.

Claims

1. A method for separating and purifying linolenic acid using a composite membrane, characterized in that: The method of purification comprises the following steps: Step S1, preparation of ethyl linolenate: pre-treated oil and ethanol are stirred and mixed in a certain amount, and immobilized lipase is added to initiate the esterification reaction; Step S2, lipase separation: the mixed solution is passed through an ultrafiltration membrane for molecular interception, and the lipase and substrate impurities are intercepted under pressure; Step S3, two-phase separation: the filtrate is passed through a polyvinylidene fluoride microfiltration membrane modified with a polyvinyl pyrrolidone porogen and a surfactant, sodium lauryl sulfate, to separate the oil phase and the aqueous phase; Step S4, separation of ethyl linolenate: the oil phase passes through a functional membrane prepared by a composite of polydimethylsiloxane and modified nano-titanium dioxide, and the ethyl linolenate is separated from the remaining linolenic acid raw material by utilizing the polarity and molecular weight differences of ethyl linolenate; Step S5, molecular distillation: molecular distilling the separated product to obtain an ethyl linolenate product with a purity of ≥85%.

2. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, wherein: In step S1, the grease is pretreated by plate and frame filtration.

3. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, wherein: In step S1, the oil refers to vegetable oil, and the vegetable oil is one or more oils with high linolenic acid content, and the molar ratio of the vegetable oil to ethanol is 3:1 to 6:

1.

4. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, wherein: In step S1, the enzymatic reaction uses a glycerol pre-adsorption method to treat the lipase, and a tertiary amino resin to immobilize the enzyme, which specifically includes the following steps: Novozym 435 lipase was mixed with a 15% glycerol solution at a ratio of 5% to 20%, and adsorbed in a pH 7.0 phosphate buffer at 25°C with shaking for 6 hours. The mixture was washed with n-hexane and then vacuum-dried to obtain a glycerol-pre-adsorbed lipase. The adsorbed lipase was then immobilized with a tertiary amino resin to obtain an immobilized lipase. The immobilized lipase treated with pre-adsorption of glycerol is mixed with a mixture of oil and ethanol, wherein the addition amount of the immobilized lipase is 5% to 10% of the total weight of the vegetable oil, and the enzymatic esterification reaction is carried out under stirring conditions of 40°C to 55°C and 200r / min to 300r / min.

5. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, wherein: In step S1, the amount of immobilized lipase added is 5% to 10% of the total weight of the vegetable oil; the enzymatic reaction temperature is 40° C. to 65° C., and the enzymatic reaction time is 24 hours to 48 hours.

6. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, characterized in that: In step S2, the pore size of the ultrafiltration membrane is 10 kDa-50 kDa.

7. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, characterized in that: In the step S3, the pore size of the modified polyvinylidene fluoride microfiltration membrane is 0.1 μm-10 μm, and the pressure applied during separation is 0.1 MPa-0.2 MPa.

8. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, characterized in that: In step S4, the operating pressure of the functional membrane is 0.2 MPa.

9. The method for separating and purifying linolenic acid using a composite membrane according to claim 1, wherein: In step S5, the temperature of the molecular distillation process is 120° C. to 150° C., and the vacuum degree is 0.1 Pa to 10 Pa.

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