A method for continuous production of medium and long chain triglycerides by enzymatic means

Abstract

The present application belongs to the technical field of oil refining and enzymatic modification, and particularly relates to a method for continuously preparing medium-long chain triglyceride by enzyme method. In the present application, medium-chain triglyceride (MCT) and long-chain triglyceride (LCT) are used as substrates, mixed, and then fed into a packed bed continuous flow reactor filled with micron-sized immobilized lipase particles to perform high-efficiency and controllable transesterification reaction, so as to realize continuous synthesis of MLCT. After the obtained product is separated and purified by low-pressure distillation combined with molecular distillation, flavor compounding and regulation are performed, and MLCT edible oil with both nutrition and flavor is obtained. The present application has the advantages of short reaction time, stable process, and suitability for large-scale continuous production, and provides an innovative solution for green manufacturing of high-quality flavor type medium-long chain triglyceride.

Description

Technical Field

[0001] This invention belongs to the field of oil and fat refining and enzymatic modification technology, specifically relating to a method for the continuous enzymatic preparation of medium- and long-chain triglycerides. Background Technology

[0002] Medium- and long-chain triglycerides (MLCTs) are a class of structural lipids with special functions. Their molecular structure contains both medium-chain fatty acids (C6-C12) and long-chain fatty acids (≥C14), giving them the dual advantages of rapid metabolism for energy supply and nutrient transport. They exhibit significant biological activity in improving obesity, enhancing immunity, and promoting the absorption of fat-soluble nutrients, and are widely used in functional foods, infant formula, and clinical nutritional preparations.

[0003] Enzymatic synthesis of MLCTs has become an ideal technological route in the field of green synthesis due to its advantages such as mild reaction conditions, high regioselectivity, and environmental friendliness. However, traditional batch stirred reactors generally suffer from problems such as low mass transfer efficiency and high mechanical shear force, which lead to easy catalyst breakage and detachment. Consequently, immobilized lipases are prone to breakage and detachment, have short service life, and low reusability, which seriously restricts the realization of continuous production.

[0004] To overcome the bottleneck of continuous production, related technologies have attempted to optimize processes using packed bed reactors. For example, patent CN105087694B discloses a scheme for preparing MLCT by filling a reactor with commercially available Novozymes Lipozyme TL IM immobilized lipase, and patent CN116555024B reports the application of packed bed reactors in continuous enzyme catalysis processes for non-natural amino acids. Although continuous flow enzyme reactors have shown significant technical advantages in terms of heat and mass transfer efficiency, precise process control and large-scale scale-up, existing packed bed technology still has core defects that are difficult to avoid in the MLCT enzymatic synthesis scenario: the above patents all rely on commercial immobilized enzymes, and their carriers are mostly conventional rigid resins or unmodified silica gels. These carriers lack directional design for MLCT synthesis and generally have an inherent contradiction between "mass transfer efficiency" and "enzyme immobilization stability": (1) The carrier particle size is too large (usually >20 μm) or the pore structure is unreasonable (wide pore size distribution and poor connectivity), which not only leads to high diffusion resistance of substrate (oil-based hydrophobic substrate) in the pores, but also easily causes enzyme molecule desorption and leakage; at the same time, after the rigid carrier is filled, a dense bed is formed, which easily produces channeling phenomenon, resulting in uneven flow of substrate solution and significant differences in local reaction efficiency, which further restricts the MLCT conversion rate. (2) The surface chemical properties of the carrier are simple and lack specific action sites, making it difficult to balance the high loading capacity of the enzyme with the stable anchoring effect. This leads to the easy loss of enzyme molecules under continuous flow scouring and shortens the catalytic life. In addition, conventional carriers cannot specifically enrich hydrophobic substrates, but instead easily adsorb hydrophilic impurities in the system (such as reaction by-product water and water-soluble impurities in the raw materials). These impurities occupy enzyme active sites and form competitive inhibition, significantly reducing catalytic efficiency. (3) Rigid carriers have insufficient compressive strength and bed stability. During long-term continuous operation, particle breakage and bed compaction are prone to occur, leading to a sudden increase in fluid resistance or even pipeline blockage. This seriously affects the long-term stable operation of the production system and cannot meet the needs of continuous industrial production of MLCT.

[0005] Furthermore, vegetable oils such as rapeseed oil, peanut oil, and sea buckthorn oil are highly favored by consumers due to their unique flavor characteristics and rich content of functionally active fatty acids, offering both sensory appeal and nutritional advantages. However, in the current production process of MLCT (Molecularly Extracted Chemicals), molecular distillation technology is commonly used for separation and purification. This process relies on high temperature and high vacuum environments, which, while effectively separating components, easily leads to the decomposition or loss of heat-sensitive and chemically unstable volatile flavor substances—such as key aroma components like esters and aldehydes—significantly weakening the original natural flavor of the final product. Although existing research has focused on the functional development and synthetic pathway optimization of MLCT, less attention has been paid to the critical issue of flavor preservation during processing. Therefore, compared to traditional products, MLCT that can fully preserve the unique flavor of vegetable oils will have greater market competitiveness and commercial value.

[0006] Therefore, there is an urgent need to develop a new process that can efficiently prepare MLCT while effectively maintaining the natural flavor of the raw materials, in order to meet consumers' demand for both "functionality and deliciousness" and promote the innovative development of high-end functional oil products. Summary of the Invention

[0007] In view of this, the present invention aims to solve the key problems existing in the preparation process of medium- and long-chain triglycerides (MLCTs), such as the difficulty of commercially available enzyme preparations in achieving both high loading capacity and pressure resistance, the lack of efficient and stable immobilized enzymes, the tendency of continuous flow enzyme packed bed reactors to experience increased pressure drop or even blockage during long-term operation, affecting system stability, and low product flavor retention. The invention proposes a method for the continuous enzymatic preparation of MLCTs, which is a method for the continuous production of MLCT edible oils that combine nutritional functions and natural flavor. This technical solution has advantages such as short reaction time, stable process, and suitability for large-scale continuous production, providing an innovative solution for the green manufacturing of high-quality flavorful MLCTs.

[0008] It should be noted that this invention combines the continuous flow characteristics of packed beds with the specific requirements of MLCT enzymatic synthesis, developing a co-loading technology of micron-sized mesoporous silica spheres immobilized with enzyme carriers and hydrophobic cellulose, achieving multi-dimensional technological breakthroughs: the micron-sized mesoporous silica spheres are directionally designed with an average particle size of 2–20 μm and a pore size distribution concentrated in 8–15 nm (precisely matching the lipase molecule size and lipid substrate diffusion requirements). The silica sphere surface is rich in modifiable functional groups, which can achieve highly stable anchoring of lipases through hydrogen bonding, hydrophobic interactions, etc., significantly reducing enzyme molecule loss during continuous flow, significantly extending service life, and improving catalytic turnover per unit time. The hydrophobic cellulose and micron-sized mesoporous silica-based immobilized enzyme are used together as bed padding. The softness of cellulose buffers fluid impacts, avoiding the defects of a dense bed formed by a single rigid carrier. This creates flexible and highly interconnected pore channels, preventing channeling problems and ensuring uniform flow of the substrate solution within the bed. It also avoids pressure drop increases and pipe blockages caused by rigid particle breakage and compaction, significantly reducing fluid resistance during long-term continuous operation and ensuring stable system operation. Simultaneously, the hydrophobic cellulose can adsorb and enrich hydrophobic oily substrates through hydrophobic interactions, significantly increasing the local substrate concentration around the enzyme molecules, strengthening the contact between the enzyme active site and the substrate, and reducing ineffective loss of enzyme activity. Furthermore, its weak adsorption capacity for hydrophilic impurities effectively reduces the competitive inhibition of enzyme active sites by impurities, ultimately achieving a simultaneous improvement in MLCT synthesis efficiency, product conversion rate, and production stability.

[0009] Furthermore, this invention innovatively uses natural plant oils such as rapeseed oil, peanut oil, and sea buckthorn oil as raw materials. Through precise selection of specific fatty acid combinations, optimization of transesterification process parameters in a continuous flow packed bed enzyme reactor, and a flavor blending and control strategy combining vacuum distillation and molecular distillation, it has successfully achieved continuous enzymatic preparation of MLCT products that combine nutritional fortification and flavor harmony. The resulting product not only inherits the health benefits of MLCT but also retains the unique taste of plant-derived oils. This overcomes the limitations of existing research that focuses solely on the functional attributes of MLCT while neglecting sensory quality, significantly enhancing consumer acceptance and market competitiveness, and giving the product higher added value.

[0010] In summary, this invention, based on a high-performance micron-scale immobilized carrier and a flavor-oriented synthesis concept, constructs a highly efficient enzyme catalytic system suitable for continuous flow packed bed reactors, providing key technical support for the clean, energy-saving, and continuous production of high-quality flavor-oriented MLCTs.

[0011] To achieve the above objectives, the present invention adopts the following technical solution:

[0012] The technical objective of this invention is to provide a method for the continuous enzymatic preparation of medium- and long-chain triglycerides (MLCTs). The method involves mixing oils rich in long-chain fatty acid triglycerides and oils rich in medium-chain fatty acid triglycerides, and then feeding them into a continuous flow reactor filled with an immobilized lipase catalyst. The mixture undergoes an enzymatic transesterification reaction at a constant temperature to obtain an oil mixture rich in MLCTs. After distillation and recombining, an oil product rich in MLCTs with both nutritional value and flavor is obtained.

[0013] Specifically, the method for continuous enzymatic preparation of medium- and long-chain triglycerides includes the following steps:

[0014] (1) Preparation of micron-sized immobilized lipase: The immobilized lipase carrier is a hydrophobic micron-sized mesoporous silica sphere. It is prepared by acid catalysis and co-hydrolysis polycondensation. Acid catalysis and co-hydrolysis polycondensation of polyethyl silicate-40 are used to obtain a prepolymer. The prepolymer is used as the dispersed phase, water-alcohol is used as the continuous phase, and Triton-100 and sodium dodecyl sulfate are used as emulsifiers to construct an emulsion system. Under alkaline conditions, the prepolymer is further hydrolyzed and condensed, and the emulsion droplets are solidified into spheres to obtain micron-sized porous spherical silica. The micron-sized silica spheres are expanded, and the prepared microspheres are uniformly dispersed in a reactor containing an aqueous solution. Ammonia is added, the reactor is sealed, and the microspheres are subjected to alkaline heat treatment. After filtration, washing and drying with water and ethanol, micron-sized mesoporous silica spheres are obtained. The surface of the microspheres is hydrophobically modified. Finally, the microspheres are washed with ethanol, filtered, and dried to obtain hydrophobic hollow mesoporous silica spheres. The lipase is immobilized on the hydrophobic silica sphere carrier by physical adsorption to obtain immobilized lipase.

[0015] (2) Preparation of packed bed enzyme reactor: A certain amount of immobilized enzyme is mixed with hydrophobic cellulose and placed in a packed column to make a packed bed enzyme reactor. Then, it is placed in a constant temperature drying oven and the packed column is heated to the required reaction temperature.

[0016] (3) Continuous flow enzymatic preparation of medium- and long-chain triglycerides (MLCT): Long-chain triglycerides (LCT) and medium-chain triglycerides (MCT) are mixed and passed through a packed bed enzyme reactor. The transesterification reaction is completed under the catalysis of micron-sized immobilized lipase. The substrate molar ratio and flow rate are linearly controlled to obtain an oil mixture rich in MLCT.

[0017] (4) Separation and purification of medium- and long-chain triglycerides: Volatile aroma components are extracted from the oil mixture rich in MLCT by vacuum distillation to obtain natural flavor compounds; then the remaining oil is subjected to molecular distillation to remove free fatty acids to obtain high-purity MLCT oil; the obtained MLCT oil is compounded with the obtained natural flavor compounds to finally obtain a medium- and long-chain triglyceride edible oil product with both nutrition and flavor.

[0018] According to the above scheme, the hydrophobic micron-sized mesoporous silica spheres have a diameter of 2-20 μm, a mesopore diameter of 8-15 nm, and a contact angle of 110°-130°.

[0019] According to the above scheme, the specific preparation process of hydrophobic micron-sized mesoporous silica spheres is as follows:

[0020] Polyethyl silicate-40 and anhydrous ethanol were placed in a three-necked flask and thoroughly mixed by mechanical stirring in a water bath. Hydrochloric acid was added dropwise and stirring continued. Then, the mixture was subjected to low-pressure rotary evaporation to remove ethanol, hydrochloric acid, and other substances, yielding a prepolymer. The prepolymer was dissolved in a continuous phase composed of water, isopropanol, Triton-100, and sodium dodecyl sulfate. The mixture was stirred mechanically for a period of time to form an O / W emulsion. Ammonia was added for secondary hydrolysis and condensation, and stirring continued. A certain amount of water was added, and the mixture was allowed to stand for a period of time. This process was repeated three times. The mixture was then filtered, washed, and dried sequentially with water and ethanol. An appropriate amount of the material was placed in a three-necked flask, a certain amount of water was added, and the mixture was ultrasonically treated. Ammonia was added, and the mixture was placed in an oil bath and mechanically stirred for a period of time to expand the pores. After filtration, washing, and drying, micron-sized mesoporous silica spheres (MMSS) were obtained. Alkyl trichlorosilane was added dropwise to a hexane solution containing MMSS for surface hydrophobic modification. Finally, the mixture was washed with ethanol, filtered, and dried to obtain hydrophobic hollow mesoporous silica spheres MMSS-Cn.

[0021] Furthermore, in the specific preparation process of the hydrophobic micron-sized silicon spheres, the volume ratio of polyethyl silicate-40 to anhydrous ethanol is 2:1-4:1, the amount of 0.1 mol / L hydrochloric acid added is 1-4 mL, the stirring time is 30-60 min, the low-pressure rotary evaporation temperature is 40-80 ℃, and the time is 10-40 min. The volume ratio of prepolymer, water, and isopropanol is 1:3:1, the amount of Triton-100 added is 0.5-1.5% of the prepolymer, the amount of sodium dodecyl sulfate added is 0.3-0.9% of the prepolymer, the amount of ammonia added is 3-6%, the secondary stirring time is 20-50 min, and the settling time is 12-36 h.

[0022] Furthermore, in the specific preparation process of hydrophobic hollow mesoporous silica spheres, the alkyl trichlorosilane is one or more of methyl trichlorosilane, vinyl trichlorosilane, butyl trichlorosilane, n-octyl trichlorosilane, and octadecyl trichlorosilane. The amount of alkyl trichlorosilane added is 30%-50% of the mass of MMSS, the reaction temperature is 20-30 °C, and the reaction time is 1-3 h.

[0023] According to the above scheme, the specific process of preparing immobilized lipase by adsorption is as follows: the free enzyme is dissolved in phosphate buffer to prepare an enzyme solution, and then the enzyme solution is mixed with a hydrophobic carrier at a certain solid-liquid ratio (the ratio between mass and enzyme solution volume) for adsorption and immobilization. After centrifugation, filtration and washing, the immobilized lipase is obtained by freeze drying.

[0024] Furthermore, in the specific process of preparing immobilized lipase by adsorption, the free enzyme is one or more of the following: *Candida foldii* lipase, *Thermophilus sparsely cottony* lipase, *Candida antarcticis* lipase, and *Candida lipolyticis* lipase; the pH of the enzyme solution is 7.0-10.0, and the concentration is 10-100 mg / mL; the ratio of MMSS-Cn mass to enzyme solution volume is 1:100-3.5:100 (m / v, g / mL); the immobilization time is 30-50 min, and the temperature is 25-40 °C; the pH of the phosphate buffer is 7.0-10.0. During enzyme immobilization, the phosphate buffer has a significant impact on the amount of enzyme immobilized, therefore the pH of the phosphate buffer is set to neutral to alkaline 7.0-10.0; the temperature has a relatively small impact, and room temperature is generally sufficient.

[0025] Specifically, the preparation steps of the micron-scale immobilized lipase include:

[0026] (1) Synthesis of prepolymer: In a 200 mL three-necked flask, polyethyl silicate-40 (65 g) was mixed with anhydrous ethanol (18 mL), and 0.1 mol / L hydrochloric acid (3 mL) was added dropwise under stirring at 35 °C and 700 rpm. After stirring for 30 minutes, the evaporation product was evaporated under reduced pressure at 80 °C for 20 minutes to obtain the prepolymer.

[0027] (2) Synthesis of silica spheres: Take the prepolymer (50 g) and dissolve it in a continuous phase containing water (150 mL), isopropanol (50 mL), Triton-100 (2.0 g) and sodium dodecyl sulfate (0.3 g); after homogenizing the mixture at 1000 rpm for 10 minutes, add ammonia solution (5 mL) to initiate secondary hydrolysis and continue stirring for 20 minutes; then dilute with 100 mL of water and let it stand for 24 hours to precipitate, and discard the supernatant; further purify by adding water (200 mL) and standing for 24 hours again, filter to collect the solid, wash with water and ethanol in sequence, and dry at 60 °C for 24 hours to obtain silica spheres (apparent diameter 2-20 μm).

[0028] (3) Pore expansion treatment: 10 g of silica spheres were dispersed in 100 mL of water and sonicated for 10 minutes; 1 mL of ammonia was added and stirred at 70 °C for 12 hours to expand the pores; the product was thoroughly washed with water / ethanol and dried at 60 °C to obtain micron-sized mesoporous silica spheres (mesopore diameter 8-15 nm).

[0029] (4) Hydrophobic modification: 1 g of micron-sized mesoporous silica spheres was dispersed in 10 mL of hexane containing 462 μL of octyltrichlorosilane and sonicated for 10 minutes. Then, the mixture was stirred at 220 rpm for 2 hours at 20 °C. After the reaction was completed, the mixture was filtered and washed with ethanol to obtain hydrophobic micron-sized mesoporous silica microspheres (contact angle 110°-130°).

[0030] (5) Lipase immobilization: Select one or more of the following: Candida lipolytica lipase, Candida pleuropsis lipase and Candida antarcticis lipase, prepare a lipase solution (10-100 mg / mL, pH 7.5-9.5, 50 ml), mix it with pre-prepared hydrophobic micron-sized mesoporous silica microspheres, the solid-liquid ratio of the carrier and enzyme solution is 1:100-3.5:100 (m / v, g / mL), the immobilization time is 30-90 min, centrifuge, filter, freeze dry to obtain micron-sized immobilized lipase preparation (immobilization load 125-179 mg / g), and store for later use.

[0031] According to the above scheme, the oil rich in long-chain fatty acid triglycerides is one or a mixture of several of the following in any proportion: rapeseed oil, peanut oil, sea buckthorn oil, flaxseed oil, perilla oil, olive oil, hemp seed oil, evening primrose oil, peony seed oil, camellia oil, soybean oil, sunflower seed oil, conjugated linoleic acid glycerides, algae oil, fish oil, garlic fruit oil, and *Xanthoceras sorbifolium* oil; the medium-chain fatty acid triglycerides are caprylic / capric triglycerides.

[0032] According to the above scheme, the mass ratio of medium-chain fatty acid triglycerides to long-chain triglycerides is 2:3-3:2 (m / m), the reaction temperature is 30-90 °C, and the flow rate of the transesterification reaction is 5-60 mL / h.

[0033] Considering the flexibility required for dynamic formulations in actual production, in some embodiments, the continuous production system may include multiple parallel packed-bed continuous flow enzyme reactors.

[0034] According to the above scheme, the conditions for vacuum distillation are: constant temperature water bath at 50 ℃, circulating water temperature at 50 ℃, system pressure less than 10 Pa, and collection bottle placed in an environment of -20 ℃ to collect natural flavor compounds.

[0035] According to the above scheme, the molecular distillation conditions are as follows: feed rate of 10–50 mL / min, vacuum degree of molecular distillation less than 4 Pa, temperature of 180–220 ℃, and scraping speed of 240–350 rpm. The heavy phase is collected as medium- and long-chain triglyceride products (acid value between 0.3–2.0 mg KOH / g, medium- and long-chain triglyceride content of 75%–85%; total triglyceride content of 95–97%, and diglyceride content of 2.0–3.3%); the light phase is the fatty acid byproduct obtained by hydrolysis during the reaction.

[0036] According to the above scheme, the compounding conditions are as follows: natural flavor compounds and MLCT oils are slowly stirred and mixed at a rate of 50-100 rpm for 1 hour at a low temperature of 4 ℃ to obtain the final product.

[0037] The MLCT-rich oil products prepared by the above method have an acid value between 0.3-2.0 mg KOH / g, an MLCT content of 75%-85%, a total triglyceride content of 95-97%, a diglyceride content of 2.0-3.3%, and a functional fatty acid content of 55.12-61.73% in MLCT. All indicators meet the requirements of edible oil standards.

[0038] Compared with the prior art, the beneficial effects of the present invention are:

[0039] This invention uses medium-chain triglycerides (MCT) and long-chain triglycerides (LCT) as substrates. After mixing, the mixture is fed into a packed bed continuous flow reactor containing micron-sized immobilized lipase particles for efficient and controllable transesterification, achieving continuous synthesis of MLCT. The resulting product is purified by low-pressure distillation combined with molecular distillation, and then flavor blending is performed to obtain MLCT edible oil that combines nutrition and flavor.

[0040] This invention features a targeted design of high-performance micron-sized immobilized lipases, balancing high loading capacity with stress resistance, significantly improving enzyme stability and reusability while reducing production costs. Based on a packed bed continuous flow reaction system, it enables continuous operation of the reaction process, resulting in high mass transfer efficiency and low mechanical shear, which helps extend enzyme life and increase production capacity. Furthermore, it introduces a "flavor blending and regulation strategy" to effectively retain the characteristic flavor components of the raw oils, endowing the final product with a natural flavor and enhancing its sensory quality and market appeal.

[0041] In summary, this invention has advantages such as short reaction time, stable process, and suitability for large-scale continuous production, providing an innovative solution for the green manufacturing of high-quality flavorful medium- and long-chain triglycerides. Attached Figure Description

[0042] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0043] Figure 1 This is a diagram of the packed bed enzyme reactor used in this invention.

[0044] Figure 2 The images shown are SEM and TEM images of the carrier material obtained in Example 1.

[0045] Figure 3 The images show gas chromatograms of the medium- and long-chain triglycerides synthesized from rapeseed oil and MCT via enzymatic transesterification in Example 2, and gas chromatograms of the physical mixture of rapeseed oil and MCT.

[0046] Figure 4 This is an electronic nose flavor distribution map of rapeseed oil MLCT in Example 2.

[0047] Figure 5 This shows the content of medium- and long-chain triglycerides synthesized from rapeseed oil and MCT enzymatic transesterification reactions 20 times in a continuous flow enzyme reactor in Example 2. Detailed Implementation

[0048] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0049] The term "embodiment" used herein, as an example, is not necessarily to be construed as superior to or better than other embodiments. Performance testing in the embodiments of this application, unless otherwise specified, employs conventional testing methods in the art. It should be understood that the terminology used in this application is merely for describing particular implementations and is not intended to limit the scope of this disclosure.

[0050] Unless otherwise stated, the technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; other experimental methods and technical means not specifically mentioned herein refer to experimental methods and technical means commonly used by one of ordinary skill in the art.

[0051] To better illustrate the content of this application, numerous specific details are provided in the following detailed embodiments. Those skilled in the art should understand that this application can be implemented even without certain specific details. In the embodiments, some methods, means, instruments, and devices well-known to those skilled in the art are not described in detail in order to highlight the main points of this application.

[0052] Without conflict, the technical features disclosed in the embodiments of this application can be combined arbitrarily, and the resulting technical solution belongs to the content disclosed in the embodiments of this application.

[0053] This invention discloses a method for the continuous enzymatic preparation of medium- and long-chain triglycerides.

[0054] To better understand the present invention, the following embodiments are provided for further detailed description of the present invention, but they should not be construed as limiting the present invention. Any non-essential improvements and adjustments made by those skilled in the art based on the above-described invention are also considered to fall within the protection scope of the present invention.

[0055] Testing of MLCT products:

[0056] The contents of MLCT, LCT, and MCT in MLCT products were determined by gas chromatography. Instrumentation: A gas chromatograph (Shimadzu GC2030, Japan) equipped with a flame ionization detector was used to detect MLCT content. The chromatographic column was a DB-5HT (15 m × 0.32 mm, 0.10 μm). Method: The column was stored at 170 °C for 2 min, then ramped up to 380 °C at a rate of 5 °C / min and held for 6 min, followed by injection at a 50:1 split ratio. Helium was used as the carrier gas at a flow rate of 2 mL / min, and air was supplied at a flow rate of 200 mL / min. The injector and detector temperatures were maintained at 380 °C.

[0057] The composition of glycerides was determined by liquid chromatography. The detection conditions were as follows: a Venusil XBPSilica 5 μm 4.6 × 250 mm column was used; an ELSD detector was employed; the sample injection volume was 10 μL (30 mg / mL); the column temperature was 35 ℃; the mobile phase was hexane / isopropanol / formic acid (15:1:0.03, v:v:v); and the flow rate was 1 mL / min.

[0058] The structure of the glycerol esters in the product was determined using liquid chromatography-mass spectrometry (LC-MS). The LC and MS parameters are as follows:

[0059] Chromatographic system: UPLC LC-30A system equipped with a Phenomenex Kinete C18 column (100*2.1 mm, 2.6 μm). Method: Mobile phases were solvent A (water / methanol / acetonitrile 1:1:1, v / v) and solvent B (isopropanol / acetonitrile 5:1, v / v), with a flow rate of 400 μL / min. Gradient elution conditions: 0–0.5 min, 20% B phase; 0.5–1.5 min, 40% B phase; 1.5–3 min, 60% B phase; 3–13 min, 95% B phase, hold for 7 min, 20.1 min, 20% B phase; 20.1–25 min, 20% B phase. Column and sample room temperatures were 60 °C and 4 °C, respectively. The sample volume for each analysis was 3.0 μL. Each lipid component was qualitatively analyzed using standards. The sample concentration and peak area showed a linear relationship. The relative composition of each substance was expressed by the area normalization method (%).

[0060] Mass spectrometry system: AB Sciex Triple TOF 6600 for MLCT mass spectrometry analysis. Method: ESI positive ion mode, mass number acquired by mass spectrometry in the range of m / z 100-1200. Mass spectrometry conditions were as follows: curtain gas: 35,000 psi; ion source Gas1: 50,000; ion source Gas2: 50,000; ion spray voltage: 5500,000 V; temperature: 600 °C.

[0061] The aroma characteristics of the product were analyzed using headspace solid-phase microextraction gas chromatography-mass spectrometry. The gas chromatography and mass spectrometry parameters are as follows:

[0062] Gas chromatography was performed using an Agilent 7890A-5975C (Agilent Technologies) DB-WAX column (30 m × 250 μm × 0.25 μm). The GC injector was set to splitless mode at 230 °C, and the column temperature was held at 30 °C for 1 minute, then increased to 240 °C at a rate of 5 °C / min and held for 1 minute. Helium was used as the carrier gas at a flow rate of 1 mL / min, and the mass spectrometer detector was selected to operate in ionization mode (MS-EI) with an ionization energy of 70 eV. The ion source temperature was 230 °C.

[0063] Example 1:

[0064] A method for preparing medium- and long-chain triglycerides using a silicon-based carrier-supported packed-bed enzyme reactor includes the following steps:

[0065] (1) Preparation of hydrophobic micron-sized silica spheres:

[0066] 65 g of polyethyl silicate-40 and 18 mL of anhydrous ethanol were placed in a 200 mL three-necked flask and thoroughly mixed by mechanical stirring at 700 rpm in a 35 °C water bath. 3 mL of 0.1 mol / L hydrochloric acid was added dropwise, and stirring continued for 30 min. The mixture was then subjected to low-pressure rotary evaporation at 80 °C for 20 min to remove ethanol, hydrochloric acid, and other substances, yielding the prepolymer. 50 g of the prepolymer was dissolved in a continuous phase consisting of 150 mL of water, 50 mL of isopropanol, 2.0 g of Triton-100, and 0.3 g of sodium dodecyl sulfate. The mixture was stirred at 1000 rpm for 10 min to form an O / W emulsion. 5 mL of ammonia was added for secondary hydrolysis and condensation, and stirring continued for 20 min. 100 mL of water was added, and the mixture was allowed to stand for 24 h. The supernatant was then poured off, 200 mL of water was added, and the mixture was allowed to stand for another 24 h. The mixture was filtered and washed sequentially with water and ethanol, and then dried at 60 °C for 24 h to obtain silica spheres. Take 10 g of the product and place it in a 150 mL three-necked flask. Add 100 mL of water and sonicate for 10 min. Then add 1 mL of ammonia and place the flask in a 70 °C oil bath with mechanical stirring at 150 rpm for 12 h to expand the pores. After filtration and washing with water and ethanol and drying at 60 °C, micron-sized mesoporous silica spheres (MMSS) are obtained.

[0067] 1 g MMSS was dissolved in 10 mL n-hexane, 462 uL n-octylsilane was added and sonicated for 10 min, and then shaken in a shaking bed at 220 rpm at 20 ℃ for 2 h. The mixture was then filtered through ethanol and dried to obtain hydrophobic micro silica spheres (MMSS-C8).

[0068] (2) Preparation of immobilized lipase:

[0069] 3.5 g of free Candida lipase (CSL) was dissolved in phosphate buffer (50 mL, pH 9.0, 50 mM), stirred at 300 rpm for 40 min at 4 °C, and centrifuged to separate the precipitate, yielding the enzyme solution. Hydrophobic hollow mesoporous silica beads were added at a solid-liquid ratio of 10 mg / mL, and the mixture was incubated on a shaker at 32 °C for 40 min. The precipitate was then separated by centrifugation and freeze-dried to obtain the immobilized lipase CSL@MMSS-C8. The enzyme loading was 163 mg / g.

[0070] (3) Preparation of continuous flow packed bed enzyme reactor

[0071] A mixture of dried immobilized lipase (0.9 g) and hydrophobic cellulose (0.6 g) was packed into a high-purity polypropylene hollow column with a diameter of 8.9 mm and a length of 63 mm. The column ends were sealed with sieve plates, silica sand, and hydrophobic cellulose to prevent leakage of the immobilized lipase. Silicone tubing was connected to the inlet and outlet of the hollow column, and a syringe pump was connected to the other end of the inlet to deliver the liquid. All equipment was placed in a constant temperature chamber to maintain a constant reaction temperature during the reaction.

[0072] like Figure 1 The diagram shows the packed bed reactor apparatus used in this invention. Connected sequentially by pipelines, it includes: a pump, a packed bed continuous flow enzyme reactor, an oven, a vacuum distillation unit, and a molecular distillation unit. The packed bed continuous flow enzyme reactor can be arranged independently or in parallel.

[0073] Example 2

[0074] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven, which was heated to 50°C. 10g of caprylic / capric triglyceride and 15g of rapeseed oil were mixed as reactants. After heating to the required reaction temperature, the mixture was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 15mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the packed column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain period, the syringe pump and the constant temperature oven were turned off. 500g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50°C, with circulating water at 50°C and a system pressure less than 10 Pa. The collection bottle was placed in an environment of -20°C to collect the flavor distillate. Heavy phase molecular distillation was performed at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 180 °C, and a scraper rotation speed of 240 r / min, yielding 390 g of MLCT-rich oil. The flavor distillate was mixed with the refined MLCT product at low temperature (4 °C) with slow stirring (50-100 rpm) for 1 h to obtain the final product.

[0075] The product was found to have an acid value of 0.6 mg KOH / g, an MLCT content of 73.24%, and a characteristic functional fatty acid (C18:1, C18:2) content of 59.21%.

[0076] Table 1 shows the mass spectrometry data of the medium- and long-chain triglycerides synthesized by the transesterification reaction of rapeseed oil and caprylic / capric triglycerides in Example 2 (the components with a content >0.5% are listed, and the total is the total). The MLCT content is 73.24%, the diglyceride content is 3.23%, and the triglyceride content is 96.45%.

[0077] Figure 3 The figures show the gas chromatograms of the medium- and long-chain triglycerides synthesized from rapeseed oil and MCT via enzymatic transesterification in Example 2, and the gas chromatogram of the physical mixture of rapeseed oil and MCT. The yellow portion represents MCT, the blue portion represents MLCT, and the light blue portion represents LCT. As can be seen from the figures, no MLCT component was found after physical mixing, but the target product appeared after the enzymatic transesterification reaction.

[0078] Figure 4 The electronic nose flavor distribution map of rapeseed oil MLCT in Example 2, combined with mass spectrometry analysis, revealed 53 major volatile compounds in rapeseed oil MLCT, including 16 ketones, 5 aldehydes, 9 pyrazines, 3 nitriles, 9 esters, 5 acids, 3 phenols and 3 olefins, with a flavor retention rate of 85%.

[0079] Table 1

[0080]

[0081] Example 3

[0082] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven, which was heated to 40 °C. 10 g of caprylic / capric triglyceride and 15 g of peanut oil were mixed as reactants. After heating to the required reaction temperature, the mixture was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 20 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and a system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Heavy phase molecular distillation was performed at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 180 °C, and a scraper rotation speed of 260 r / min, yielding 370 g of oil rich in MLCT. The flavor distillate was mixed with the refined MLCT product at low temperature (4 °C) with slow stirring (50-100 rpm) for 1 h to obtain the final product.

[0083] The product was found to have an acid value of 0.7 mg KOH / g, a MLCT content of 75.13%, a triglyceride content of 95.52%, and a diglyceride content of 2.37%. Fifty-seven major volatile compounds were identified in the peanut oil MLCT, including three ketones, 20 aldehydes, 14 pyrazines, nine esters, three phenols, and eight other compounds, with a flavor retention rate of 86%.

[0084] Example 4

[0085] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven, which was heated to 60 °C. 10 g of caprylic / capric triglyceride and 15 g of sea buckthorn oil were mixed as reactants. After heating to the required reaction temperature, the mixture was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 30 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and a system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Molecular distillation was performed in the heavy phase at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 180 °C, and a scraper rotation speed of 260 r / min, yielding 380 g of MLCT-rich oil.

[0086] The product was determined to have an acid value of 0.6 mg KOH / g, a MLCT content of 71.53%, a triglyceride content of 95.69%, a diglyceride content of 2.97%, and a characteristic functional fatty acid (C18:1, C18:2) content of 59.93%. Forty-nine major volatile compounds were identified in the MLCT of sea buckthorn oil, including two ketones, thirteen aldehydes, and thirty-four esters.

[0087] Example 5

[0088] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven and heated to 60 °C. 10 g of caprylic / capric triglyceride and 15 g of algal oil were mixed as reactants. After heating to the required reaction temperature, the mixture was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 25 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After a certain reaction time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Molecular distillation was performed in the heavy phase at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 180 °C, and a scraper rotation speed of 260 r / min, yielding 375 g of MLCT-rich oil.

[0089] The product was found to have an acid value of 0.7 mg KOH / g, an MLCT content of 72.37%, a triglyceride content of 97.77%, a diglyceride content of 2.93%, and a characteristic functional fatty acid (C22:5, C22:6) content of 59.97%.

[0090] Example 6

[0091] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven and heated to 50 °C. 10 g of caprylic / capric triglyceride and 15 g of camellia oil were mixed as reactants. After heating to the required reaction temperature, the reactor was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 15 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the packed column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and a system pressure less than 10 Pa. The collection bottle was placed in an environment of -20 °C to collect the flavor distillate. Molecular distillation was performed in the heavy phase at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 180 °C, and a scraper rotation speed of 260 r / min, yielding 365 g of oil rich in MLCT.

[0092] The product was found to have an acid value of 0.6 mg KOH / g and an MLCT content of 72.63%.

[0093] Example 6

[0094] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven, which was heated to 50 °C. 10 g of caprylic / capric triglyceride and 15 g of olive oil were mixed as reactants. After heating to the required reaction temperature, the reactor was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 20 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and a system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Molecular distillation was performed in the heavy phase at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 180 °C, and a scraper rotation speed of 260 r / min, yielding 385 g of MLCT-rich oil.

[0095] The product was found to have an acid value of 0.6 mg KOH / g, an MLCT content of 74.51%, and a content of 55.12% of characteristic functional fatty acids (C18:1, C18:2, C18:3).

[0096] Example 7

[0097] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven and heated to 50 °C. 10 g of caprylic / capric triglyceride and 15 g of linseed oil triglyceride were mixed as reactants. After heating to the required reaction temperature, the reactor was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 25 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the packed column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain period, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Heavy phase molecular distillation was performed at a feed rate of 15 mL / min, a distillation pressure of 30 Pa, a heating temperature of 170 °C, and a scraper rotation speed of 240 r / min, yielding 375 g of MLCT-rich oil.

[0098] The product was found to have an acid value of 0.7 mg KOH / g, an MLCT content of 73.74%, and a content of 61.73% of characteristic functional fatty acids (C18:1, C18:2, C18:3).

[0099] Example 8

[0100] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven and heated to 50 °C. 10 g of caprylic / capric triglyceride and 15 g of sunflower seed oil were mixed as reactants. After heating to the required reaction temperature, the reactor was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 10 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Molecular distillation was performed in the heavy phase at a feed rate of 20 mL / min, a distillation pressure of 30 Pa, a heating temperature of 200 °C, and a scraper rotation speed of 270 r / min, yielding 385 g of MLCT-rich oil.

[0101] The product was found to have an acid value of 0.7 mg KOH / g and an MLCT content of 74.21%.

[0102] Example 9

[0103] Continuous flow enzymatic transesterification reaction: The continuous flow enzyme reactor apparatus from Example 1 was placed in a constant temperature drying oven, which was heated to 50 °C. 10 g of caprylic / capric triglyceride and 15 g of garlic fruit oil were mixed as reactants. After heating to the required reaction temperature, the reactor was placed in a constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 30 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the packed column for enzymatic transesterification. The liquid flowing out from the top of the packed column was collected as the crude transesterification MLCT product. After the reaction had proceeded for a certain time, the syringe pump and the constant temperature oven were turned off. 500 g of the crude product was accumulated and vacuum distilled under constant temperature water bath at 50 °C, with circulating water at 50 °C and system pressure less than 10 Pa. The collection bottle was placed at -20 °C to collect the flavor distillate. Molecular distillation was performed in the heavy phase at a feed rate of 10 mL / min, a distillation pressure of 30 Pa, a heating temperature of 190 °C, and a scraper rotation speed of 250 r / min, yielding 380 g of MLCT-rich oil.

[0104] The product was found to have an acid value of 0.6 mg KOH / g and an MLCT content of 76.37%.

[0105] Example 10

[0106] Long-term stability test: The operation method is as in Example 2. A continuous flow enzyme reactor was used to catalyze the transesterification reaction of rapeseed oil and MCT. The operation was continuous for 30 days, and the daily product output was measured. The results are as follows: Figure 5 As the reaction time increases, the MLCT content gradually decreases, stabilizing at around 58% after 30 days.

[0107] Comparative Example 1

[0108] The dried immobilized lipase (1.5 g) was packed into a high-purity polypropylene hollow column with a diameter of 8.9 mm and a length of 63 mm. The ends of the column were plugged with sieve plates, quartz sand, and hydrophobic cellulose to prevent leakage of the immobilized lipase. The inlet and outlet of the hollow column were connected to silicone tubing, and the other end of the inlet was connected to a syringe pump to deliver liquid. All equipment was placed in a constant temperature chamber to maintain a constant reaction temperature during the reaction.

[0109] Continuous flow enzyme transesterification reaction: The continuous flow enzyme reactor was placed in a constant temperature drying oven and heated to 50 °C. 10 g of caprylic / capric triglyceride and 15 g of rapeseed oil were mixed as reactants. After heating to the required reaction temperature, the reactor was placed in the constant temperature oven and connected to a packed column. The syringe pump connected to the packed column was turned on at a flow rate of 15 mL / h, allowing the reactants to continuously enter the enzyme-packed bed reactor from the bottom of the column for the enzyme transesterification reaction. After 2 hours of continuous operation, the fluid resistance increased sharply, the bed compacted, and the flow rate gradually decreased to 3 mL / h, making stable continuous operation impossible.

[0110] The crude product was found to contain 70.84% ​​MLCT.

[0111] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for the continuous enzymatic preparation of medium- and long-chain triglycerides (MLCT), characterized in that, Includes the following steps: (1) Preparation of micron-sized immobilized lipase: Micron-sized mesoporous silica spheres were prepared by acid catalysis and co-hydrolysis polycondensation. The obtained microspheres were subjected to alkali treatment to expand the pores, and the surface of the microspheres was hydrophobically modified with alkyl functional groups to obtain hydrophobic hollow mesoporous silica spheres MMSS-Cn. Lipase was immobilized in the pores of MMSS-Cn by adsorption to obtain micron-sized immobilized lipase. (2) Preparation of packed bed enzyme reactor: Micron-sized immobilized lipase is thoroughly mixed with hydrophobic cellulose, filled into the reactor cavity and gently compacted. After filling, the top is pressed with a 100-mesh gasket and defatted cotton to ensure that the enzyme bed is free of channeling and leakage. The reactor inlet and outlet are connected to the system with oil-resistant rubber tubes. The entire device is placed in a constant temperature drying oven to maintain a constant reaction temperature. The mass ratio of micron-scale immobilized lipase to hydrophobic cellulose is 3:1-2:1; (3) Continuous flow enzymatic preparation of medium and long chain triglycerides: Long chain triglyceride (LCT) and medium chain triglyceride (MCT) are mixed and transesterification is carried out in a packed bed enzyme reactor under the catalysis of micron-sized immobilized lipase. By linearly controlling the substrate molar ratio and flow rate, an oil mixture rich in MLCT is obtained. (4) Separation and purification of medium- and long-chain triglycerides: Volatile aroma components are extracted from the oil mixture rich in MLCT by vacuum distillation to obtain natural flavor compounds; then the remaining oil is subjected to molecular distillation to remove free fatty acids to obtain high-purity MLCT oil; the obtained MLCT oil is compounded with the obtained natural flavor compounds to finally obtain a medium- and long-chain triglyceride edible oil product with both nutrition and flavor.

2. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 1, characterized in that, The specific preparation process of micron-sized mesoporous silica spheres is as follows: Polyethyl silicate-40 and anhydrous ethanol are placed in a three-necked flask and thoroughly mixed by mechanical stirring in a water bath. Hydrochloric acid is added dropwise and stirring is continued. Then, the ethanol and hydrochloric acid are removed by low-pressure rotary evaporation to obtain the prepolymer. The prepolymer is dissolved in a continuous phase composed of water, isopropanol, Triton-100 and sodium dodecyl sulfate and mixed under mechanical stirring to form an O / W type emulsion. Ammonia was added for secondary hydrolysis and condensation, and stirring was continued. Water was added to mix and allowed to stand. This process was repeated three times. Then, the mixture was filtered, washed, and dried with water and ethanol in sequence. An appropriate amount of material was placed in a three-necked flask, water was added, and the mixture was sonicated. Ammonia was then added, and the mixture was placed in an oil bath and mechanically stirred to expand the pores. After filtration, washing, and drying, micron-sized mesoporous silica spheres MMSS were obtained. Alkyl trichlorosilane was added dropwise to a hexane solution containing MMSS for surface hydrophobic modification. Finally, the mixture was washed with ethanol, filtered, and dried to obtain hydrophobic hollow mesoporous silica spheres MMSS-Cn.

3. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 2, characterized in that, The volume ratio of polyethyl silicate-40 to anhydrous ethanol is 2:1-4:1; the amount of 0.1 mol / L hydrochloric acid added is 1-4 mL; the stirring time is 30-60 min; the low-pressure rotary evaporation temperature is 40-80 ℃; and the time is 10-40 min. The volume ratio of prepolymer, water, and isopropanol is 1:3:1; the amount of Triton-100 added is 0.5-1.5% of the prepolymer; the amount of sodium dodecyl sulfate added is 0.3-0.9% of the prepolymer; the amount of ammonia added is 3-6%; the secondary stirring time is 20-50 min; and the standing time is 12-36 h. The alkyltrichlorosilane is one or a combination of methyltrichlorosilane, vinyltrichlorosilane, butyltrichlorosilane, n-octyltrichlorosilane, and octadecyltrichlorosilane. The amount of alkyltrichlorosilane added is 30%-50% of the mass of MMSS, the reaction temperature is 20-30 °C, and the reaction time is 1-3 h.

4. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 1, characterized in that, The specific process for preparing immobilized lipase by adsorption is as follows: the free enzyme is dissolved in phosphate buffer to prepare an enzyme solution, and then the enzyme solution is mixed with micron-sized mesoporous silica balls for adsorption and immobilization. After centrifugation, filtration and washing, the immobilized lipase is obtained by freeze drying.

5. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 4, characterized in that, The free enzyme is one or more of the following: Candida lipolytica lipase, Candida thermophila lipolytica lipase, Candida antarctica lipase, and Candida lipolytica lipase; the enzyme solution has a pH of 7.0-10.0 and a concentration of 10-100 mg / mL; the ratio of MMSS-Cn mass to enzyme solution volume is 1g:100mL-3.5g:100mL; the immobilization time is 30-50 min, and the temperature is 25-40 °C; the phosphate buffer has a pH of 7.0-10.

0.

6. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 1, characterized in that, Long-chain triglycerides (LCTs) are one or more of the following in any proportion: rapeseed oil, peanut oil, sea buckthorn oil, flaxseed oil, perilla oil, olive oil, hemp seed oil, evening primrose oil, peony seed oil, camellia oil, soybean oil, sunflower seed oil, conjugated linoleic acid glycerides, algae oil, fish oil, garlic fruit oil, and *Sapindus mukorossi* fruit oil.

7. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 1, characterized in that, The packed bed enzyme reactor has the following specifications: φ8.9 mm × 63 mm, and the amount of micron-sized immobilized lipase added is 0.5-5 g.

8. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 1, characterized in that, The mass ratio of medium-chain triglycerides to long-chain triglycerides is 2:3 to 3:2, the reaction temperature is 30-90 °C, and the flow rate of the transesterification reaction is 5-60 mL / h.

9. The method for continuous enzymatic preparation of medium- and long-chain triglycerides according to claim 1, characterized in that, The vacuum distillation conditions are: constant temperature water bath at 50 ℃, circulating water temperature at 50 ℃, system pressure less than 10 Pa, and collection bottle placed in an environment of -20 ℃ to collect natural flavor compounds. The conditions for molecular distillation are as follows: feed rate of 10-50 mL / min, vacuum degree of molecular distillation of less than 4 Pa, temperature of 180-220 ℃, scraping speed of 240-350 rpm, and the heavy phase collected is the MLCT product, while the light phase is the fatty acid byproduct obtained by hydrolysis during the reaction. Natural flavor compounds and MLCT oils were slowly stirred at 50-100 rpm for 1 hour at a low temperature of 4 °C to obtain the final product.

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