Enzymatically prepared rice bran fatty alkanols, and methods of making and using the same

Abstract

This invention discloses an enzymatic method for preparing fatty alkanols from rice bran, along with its preparation method and applications, belonging to the field of deep oil processing technology. Using rice bran wax paste, a byproduct of rice bran oil processing, as raw material, the method utilizes the specific and efficient hydrolysis of rice bran wax, monoglycerides, and diglycerides by a glyceryl lipase, without affecting triglycerides. After the reaction, centrifugation separates the upper oil phase and the lower aqueous phase. The upper oil phase contains higher fatty alkanols, fatty acids, and unreacted triglycerides, while the lower aqueous phase contains glycerol, water, and the enzyme. Finally, molecular distillation is used to separate and purify the upper oil phase, yielding high-purity rice bran fatty alkanols. This invention features mild reaction conditions, high specificity, and a simple and easily purifiable product composition, enabling the efficient preparation of rice bran fatty alkanols. The product can be widely used in the food, pharmaceutical, daily chemical, and feed industries, demonstrating significant economic and social benefits.

Description

Technical Field

[0001] This invention belongs to the field of deep oil processing technology, specifically relating to an enzymatically prepared rice bran fatty alkyl alcohol, its preparation method, and its application. Background Technology

[0002] Rice bran wax paste is a byproduct of rice bran oil processing and is rich in the high-value active ingredient octacosanol. This component has outstanding physiological activity and broad application prospects, but its resource utilization rate is low, which restricts the added value of the rice bran oil processing industry. Existing methods for preparing octacosanol are difficult to achieve green, efficient, and low-cost industrial production, and cannot simultaneously meet product quality and environmental protection requirements, resulting in significant technological bottlenecks in the industry.

[0003] Currently, the preparation of octacosanol mainly relies on chemical methods. One method uses rice bran wax paste as raw material, employing reactions such as saponification, transesterification, and high-temperature pyrolysis, combined with extraction and distillation to achieve purification. The other method is chemical synthesis, using specific fatty acids as raw materials through esterification and reduction. Related patents all focus on optimizing chemical processes. Chemical extraction requires a high-temperature, strongly alkaline environment, which easily leads to the oxidative degradation of octacosanol, destroying its physiological activity, and also results in poor product color, strong odor, high energy consumption, and a large amount of alkaline waste liquid, leading to high environmental costs. Chemical synthesis methods have limited raw material sources, high costs, still rely on strong acid and reduction reactions, face significant environmental pressure, and are difficult to control in terms of purity. Overall, the existing processes are cumbersome, lack controllability, and do not conform to the trend of green manufacturing, urgently requiring breakthroughs in green preparation technologies. Summary of the Invention

[0004] To address the problems of difficult process control and poor quality in existing methods for preparing fatty alkyl alcohols from rice bran, this invention provides an enzymatic method for preparing fatty alkyl alcohols from rice bran, along with its application. This method features mild reaction conditions, rapid reaction rate, and easy product separation and purification, offering an efficient and green new pathway for the high-value utilization of rice bran wax paste, and possesses promising industrialization prospects.

[0005] To achieve the above objectives, the present invention adopts the following technical solution: The first aspect of this invention provides a method for preparing fatty alkanols from rice bran using an enzymatic method, comprising the following steps: Add metaglyceryl lipase and buffer solution to rice bran wax paste to carry out hydrolysis reaction to obtain reaction product; The reaction products were separated, and the upper oil phase was recovered. The upper oil phase was then separated and purified to obtain rice bran fatty alkyl alcohols.

[0006] Furthermore, the glycerol lipase is one or more of Lipase G "Amano" 50, Lipase SMG1, or Lipase AOL; Based on the quality of the rice bran wax paste, the amount of glyceryl lipase added is 100~300 U / g.

[0007] Furthermore, the rice bran wax paste contains 55% to 65% triglycerides and 35% to 45% rice bran wax.

[0008] Furthermore, the buffer solution is a phosphate buffer solution with a pH of 5.5 to 7.5, an ionic strength of 20 mM, and an addition amount of phosphate buffer solution of 5% to 30% of the mass of rice bran wax paste.

[0009] Furthermore, the hydrolysis reaction is carried out at a temperature of 30-45°C and a pH of 5.5-7.5.

[0010] Furthermore, the separation is centrifugal separation, with a rotation speed of 3000~5000 rpm and a centrifugation time of 10~20 min.

[0011] Furthermore, the upper oil phase comprises rice bran fatty alkanols, higher fatty acids, and triglycerides; wherein the rice bran fatty alkanols include one or more of octadecanoic fatty alkanols, triadecanoic fatty alkanols, and thirty-two-carbon fatty alkanols.

[0012] Furthermore, the separation and purification process employs molecular distillation, with a feed flow rate of 1-2 L / h, a first-stage evaporation surface temperature of 140-145℃, a second-stage evaporation surface temperature of 155-160℃, a third-stage evaporation surface temperature of 175-180℃, and a fourth-stage evaporation surface temperature of 180-185℃.

[0013] A second aspect of the present invention provides a rice bran fatty alkyl alcohol, which is prepared by the above-described enzymatic method for preparing rice bran fatty alkyl alcohol.

[0014] The third aspect of this invention provides the application of the above-mentioned rice bran fatty alkanols in the food, pharmaceutical, daily chemical and feed industries.

[0015] Compared with the prior art, the beneficial effects of the present invention are as follows: This invention provides an enzymatic method for preparing fatty alkanols from rice bran. Using rice bran wax paste as raw material, a glyceryl ester lipase is selected as a catalyst, and a buffer solution is used for hydrolysis. The target product is obtained after simple phase separation and purification steps. This method utilizes the high efficiency and specificity of glyceryl ester lipase for rice bran wax, almost completely converting it into fatty alkanols and higher fatty acids. The reaction conditions are mild, avoiding the damage to heat-sensitive components such as fatty alkanols caused by the high temperature and strong alkaline environment of traditional chemical methods. It also significantly improves the raw material conversion rate and product purity. Furthermore, the reaction process is green and environmentally friendly, producing no harmful byproducts, making it easy to scale up for industrial production and possessing good industrial feasibility. Compared to products prepared by traditional chemical methods, the fatty alkanols obtained by this method are not destroyed by high temperature and strong alkaline conditions, retaining their natural activity and physicochemical properties. They exhibit stable structure and excellent performance, possessing both the inherent advantages of fatty alkanols and avoiding the performance loss caused by chemical modification, laying the foundation for their efficient application in multiple fields.

[0016] Furthermore, the glycerol lipase is Lipase G "Amano" 50, Lipase SMG1, or Lipase AOL, which ensures that the enzyme can specifically hydrolyze rice bran wax without acting on triglycerides, thereby efficiently and specifically converting rice bran wax in rice bran wax paste into higher fatty alkanols and higher fatty acids.

[0017] The rice bran fatty alkyl alcohols prepared by the provided enzymatic method can achieve a purity of over 96% with the specific catalytic action of glyceryl lipase and subsequent separation and purification processes. The products are characterized by their single composition and high purity, effectively removing impurities and harmful components from the raw materials and improving the quality and safety of the products.

[0018] The rice bran fatty alkyl alcohols prepared by this invention can be widely used in multiple fields such as food, medicine, daily chemicals and feed industry, with rich application scenarios and strong practicality; its high purity and high activity characteristics can be adapted to the use needs of various fields, and its application effect is better than that of rice bran fatty alkyl alcohols obtained by ordinary preparation methods. Attached Figure Description

[0019] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly described below: Figure 1 This is a schematic diagram illustrating the principle of the enzymatic preparation of fatty alkanols from rice bran according to the present invention. Detailed Implementation To make the technical problem to be solved, the technical solution, and the beneficial effects of the present invention clearer, the present invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the present invention and are not intended to limit the present invention.

[0020] In this invention, the term "and / or" describes the relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent: A existing alone, A and B existing simultaneously, or B existing alone. A and B can be singular or plural. The character " / " generally indicates that the preceding and following related objects have an "or" relationship.

[0021] In this invention, "at least one" means one or more, and "more than one" means two or more. "At least one of the following" or similar expressions refer to any combination of these items, including any combination of single or multiple items. For example, "at least one of a, b, or c", or "at least one of a, b, and c", can both represent: a, b, c, ab (i.e., a and b), ac, bc, or abc, where a, b, and c can be single or multiple.

[0022] It should be understood that in various embodiments of the present invention, the order of the above-mentioned processes does not imply the order of execution. Some or all steps may be executed in parallel or sequentially. The execution order of each process should be determined by its function and internal logic, and should not constitute any limitation on the implementation process of the embodiments of the present invention.

[0023] The terminology used in the embodiments of this invention is for the purpose of describing particular embodiments only and is not intended to limit the invention. The singular forms “a,” “the,” and “the” as used in the embodiments of this invention and the appended claims are also intended to include the plural forms unless the context clearly indicates otherwise.

[0024] The weights of the relevant components mentioned in the embodiments of this invention can refer not only to the specific content of each component, but also to the proportional relationship between the weights of the components. Therefore, any scaling up or down of the content of the relevant components according to the embodiments of this invention is within the scope disclosed in the embodiments of this invention. Specifically, the mass described in the embodiments of this invention can be a mass unit known in the chemical industry, such as μg, mg, g, or kg.

[0025] Rice bran oil is a high-nutritional-value edible oil extracted from rice bran, a byproduct of rice processing. It is characterized by its mild taste, rich lipid content, and high smoke point. During the storage and refining process of rice bran oil, the wax in the oil precipitates due to decreased solubility. The byproduct obtained after filtration, mainly composed of rice bran oil and rice bran wax, is commonly referred to as rice bran wax paste. Currently, the resource utilization rate of rice bran wax paste is low. Traditional processing methods often involve its low-cost use as feed filler or purification into industrial wax, resulting in significant resource waste. Furthermore, the high-value components contained in rice bran wax paste have not been fully extracted and utilized, becoming a key issue restricting the added value of the rice bran oil processing industry.

[0026] Studies have shown that rice bran wax paste is rich in various high-value active ingredients, among which higher fatty alkanols have the greatest development potential. These saturated fatty alcohols containing long-chain alkyl groups are widely found in natural waxes such as rice bran wax and beeswax. Due to the wide availability and low cost of rice bran wax paste, it has become an ideal raw material for extracting higher fatty alkanols. Among the various fatty alkanols in rice bran wax paste, octacosanol has the highest content and the most prominent physiological activity. Octacosanol has the effects of regulating energy metabolism, improving aerobic endurance, anti-fatigue, and enhancing physical fitness. It can also inhibit intestinal cholesterol absorption and promote its excretion, having a positive impact on cardiovascular health. It also has application value in food preservation and anti-aging cosmetics. Furthermore, octacosanol has been approved by the National Health Commission of China, possessing clear safety and legality for consumption, and has a broad market prospect.

[0027] Despite the significant value of octacosanol, existing preparation methods still have many shortcomings, making it difficult to achieve efficient, green, and low-cost industrial production. CN103232320A discloses a chemical hydrolysis process for producing octacosanol, which involves saponifying, washing, dehydrating, drying, pulverizing, extracting, filtering, and desolventizing rice bran wax to obtain fatty alcohols, followed by vacuum distillation to obtain octacosanol. This method is cumbersome and relies heavily on the saponification reaction. CN112225638A discloses a process for extracting octacosanol from rice bran wax, which requires a transesterification reaction (adding methanol and sodium hydroxide), followed by molecular distillation and acetone slurry filtration, drying the filter cake to obtain octacosanol. This method suffers from solvent volatility, difficulty in recovery, residue problems, and environmental unfriendliness. CN102795960A discloses a method for large-scale preparation of high-purity octacosanol and triacontanol, using calcium oxide or calcium hydroxide to react with natural wax at a high temperature of 200-250℃ for 8-10 minutes. After further ethanol extraction and molecular distillation, octacosanol and triacontanol with a purity of over 90% are obtained. This method requires harsh reaction conditions, and high temperatures can destroy the heat-sensitive components of fatty alkanols in rice bran. The disadvantages of this type of chemical method are quite obvious: high temperatures and strongly alkaline environments easily lead to the oxidative degradation of octacosanol, destroying its natural structure and physiological activity, while also causing the product to darken in color and develop an off-odor, affecting the final quality. Furthermore, the reaction consumes a lot of energy and generates large amounts of alkaline wastewater or solid waste, resulting in high environmental treatment costs, which is inconsistent with the development trend of green manufacturing.

[0028] Other patents employ chemical synthesis routes to prepare octacosanol. For example, CN119569536A uses octacosanic acid as a raw material, first esterifying it with a monohydric alcohol under strong acid catalysis to form octacosanic acid ester, then reducing it with a reducing agent to obtain the crude product, and finally recrystallizing it with a low-polarity organic solvent to obtain the purified product. Although this method provides a novel synthetic route, the raw materials are specific fatty acids, which are limited in source and costly. It still relies on strong acid catalysis and chemical reduction reactions, posing challenges such as environmental pressure and difficulty in controlling product purity.

[0029] Currently, the preparation of fatty alkanols from rice bran mainly relies on the aforementioned chemical methods, which involve complex reaction steps, difficult process control, environmental unfriendliness, and poor product quality. Therefore, developing a green bio-enzymatic preparation technology for fatty alkanols from rice bran to achieve green, efficient, and high-quality preparation is of significant practical importance for overcoming industrial technology bottlenecks and promoting the upgrading and utilization of rice bran oil by-products.

[0030] Based on this, such as Figure 1 As shown, the enzymatic method for preparing fatty alkanols from rice bran provided by the present invention includes the following steps: Add metaglyceryl lipase and buffer solution to rice bran wax paste to carry out hydrolysis reaction to obtain reaction product; The reaction products were separated, and the upper oil phase was recovered. The upper oil phase was then separated and purified to obtain rice bran fatty alkyl alcohols.

[0031] This application utilizes a metaglycerol lipase to efficiently hydrolyze rice bran wax paste, directionally converting it into rice bran fatty alkanols and higher fatty acids. In the rice bran wax paste system, the metaglycerol lipase specifically hydrolyzes rice bran wax and metaglycerols (diglycerides and monoglycerides), generating rice bran fatty alkanols, higher fatty acids, and glycerol. It exhibits no catalytic effect on triglycerides, achieving targeted substrate hydrolysis. Furthermore, the required reaction conditions are mild, reducing the complexity of the preparation process. Compared to traditional lipases that can act on triglycerides, this application avoids the hydrolysis of triglycerides, not only improving the efficiency of rice bran wax paste hydrolysis but also reducing the difficulty of subsequent product purification; it also increases the yield and purity of rice bran fatty alkanols.

[0032] In some specific embodiments, the rice bran wax paste contains 55% to 65% triglycerides and 35% to 45% rice bran wax.

[0033] In some specific embodiments, the metaglycerol lipase is one or more of Lipase G "Amano" 50, Lipase SMG1, or Lipase AOL; based on the mass of the rice bran wax paste, the amount of metaglycerol lipase added is 100~300 U / g, and the total enzyme activity (U) = mass of rice bran wax paste (g) × amount of enzyme added (U / g).

[0034] In some specific embodiments, the buffer solution is a phosphate buffer with a pH of 5.5 to 7.5 and an ionic strength of 20 mM. The amount of phosphate buffer added is 5% to 30% of the mass of rice bran wax paste. These conditions can provide a suitable aqueous environment and a stable acid-base environment for the enzyme reaction.

[0035] In some specific embodiments, the hydrolysis reaction is carried out at a temperature of 30-45°C, a reaction time of 1-4 hours, and a stirring speed of 400-600 rpm. By employing a glyceryl ester lipase, the conditions required for the preparation of fatty alkanols from rice bran are reduced, allowing for the maintenance of high enzyme catalytic efficiency and reaction stability under mild process conditions, thus significantly improving overall efficiency.

[0036] In some specific embodiments, the separation is carried out by centrifugation, with a centrifugation speed of 3000~5000 rpm and a centrifugation time of 10~20 min.

[0037] In some specific embodiments, the upper oil phase comprises rice bran fatty alkanols, higher fatty acids, and triglycerides; wherein the rice bran fatty alkanols include one or more of octadecanoic fatty alkanols, triadecanoic fatty alkanols, and thirty-two-carbon fatty alkanols.

[0038] In some specific embodiments, the separation and purification are carried out by molecular distillation, with a feed flow rate of 1~2 L / h, a first-stage evaporation surface temperature of 140~145℃, a second-stage evaporation surface temperature of 155~160℃, a third-stage evaporation surface temperature of 175~180℃, and a fourth-stage evaporation surface temperature of 180~185℃.

[0039] To make the technical problem to be solved, the technical solution, and the beneficial effects of the present invention clearer, the present invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the present invention and are not intended to limit the present invention.

[0040] The following examples use instruments and equipment conventional in the art. Experimental methods in the following examples, unless otherwise specified, are generally performed under standard conditions or as recommended by the manufacturer. All raw materials used in the following examples are conventional commercially available products with specifications in the art, unless otherwise stated.

[0041] Example 1 220 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 33 g of 20 mM pH 6.5 phosphate buffer (15% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 40°C, and then Lipase G "Amano" 50 (200 U / g rice bran wax paste, corresponding to a total enzyme activity of 44000 U) was added. A specific hydrolysis reaction was then carried out at 400 rpm for 2 h. After the reaction, the product was centrifuged at 4000 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 20.51% higher fatty alkanols, 32.42% higher fatty acids, and 47.07% triglycerides. The upper oil phase was separated and purified by four-stage molecular distillation. The feed flow rate was 1.5 L / h, the first-stage evaporation surface temperature was 145℃, the second-stage evaporation surface temperature was 155℃, the third-stage evaporation surface temperature was 180℃, and the fourth-stage evaporation surface temperature was 185℃. After distillation, the first and second stage light phases were fatty acids, the third and fourth stage light phases were rice bran fatty alkanols, and the fourth stage heavy phase was triglycerides. Liquid chromatography analysis of the third and fourth stage light phases revealed that the rice bran fatty alkanol content in the distillation product was 98.2%.

[0042] Example 2 200 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 20 g of pH 7.0, 20 mM phosphate buffer (10% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 30°C, and then Lipase SMG1 (150 U / g rice bran wax paste, corresponding to a total enzyme activity of 30,000 U) was added. A specific hydrolysis reaction was then carried out at 500 rpm for 3 h. After the reaction, the product was centrifuged at 3500 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 20.21% higher fatty alkanols, 33.25% higher fatty acids, and 46.54% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 1.0 L / h, and the temperatures of the first-stage evaporation surface were 140℃, the second-stage evaporation surface 150℃, the third-stage evaporation surface 170℃, and the fourth-stage evaporation surface 180℃. After distillation, the third and fourth stage light phases were rice bran fatty alkanols. Liquid chromatography analysis of the third and fourth stage light phases revealed that the purity of the rice bran fatty alkanols in the distillation products was 97.1%.

[0043] Example 3 240 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 48 g of pH 6.0, 20 mM phosphate buffer (20% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 37°C, and then Lipase G "Amano" 50 (200 U / g rice bran wax paste, corresponding to a total enzyme activity of 48000 U) was added. A specific hydrolysis reaction was then carried out at 450 rpm for 2.5 h. After the reaction, the product was centrifuged at 4000 rpm for 10 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 22.87% higher fatty alkanols, 31.05% higher fatty acids, and 46.08% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 1.5 L / h, and the temperatures of the first-stage evaporation surface were 145℃, the second-stage evaporation surface 155℃, the third-stage evaporation surface 175℃, and the fourth-stage evaporation surface 182℃. After distillation, the third and fourth stage light phases were rice bran fatty alkanols. Liquid chromatography analysis showed that the purity of the rice bran fatty alkanols in the distillation product was 97.6%.

[0044] Example 4 180 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 9 g of 20 mM pH 5.5 phosphate buffer (5% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 40°C, and then Lipase AOL (300 U / g rice bran wax paste, corresponding to a total enzyme activity of 54000 U) was added. A specific hydrolysis reaction was then carried out at 500 rpm for 2 h. After the reaction, the product was centrifuged at 4500 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 26.43% higher fatty alkanols, 29.18% higher fatty acids, and 44.39% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 1.2 L / h, and the temperatures of the first-stage evaporation surface were 145℃, the second-stage evaporation surface 155℃, the third-stage evaporation surface 180℃, and the fourth-stage evaporation surface 185℃. After distillation, the third and fourth stage light phases were rice bran fatty alkanols. Liquid chromatography analysis showed that the purity of the rice bran fatty alkanols in the distillation product was 98.0%.

[0045] Example 5 260 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 78 g of pH 7.5, 20 mM phosphate buffer (30% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 45°C, and then Lipase SMG1 (100 U / g rice bran wax paste, corresponding to a total enzyme activity of 26000 U) was added. A specific hydrolysis reaction was then carried out at 600 rpm for 1 h. After the reaction, the product was centrifuged at 3000 rpm for 20 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 20.07% higher fatty alkanols, 33.21% higher fatty acids, and 46.72% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 2.0 L / h, and the temperatures of the first-stage evaporation surface were 145℃, the second-stage evaporation surface 160℃, the third-stage evaporation surface 175℃, and the fourth-stage evaporation surface 180℃. After distillation, only the third and fourth stage light phases were collected as the target product. Liquid chromatography analysis revealed that the purity of fatty alkanols in the distillate was 96.4%.

[0046] Example 6 220 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 33 g of pH 6.0, 20 mM phosphate buffer (15% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 35°C, and then Lipase G "Amano" 50 (180 U / g rice bran wax paste, corresponding to a total enzyme activity of 39600 U) was added. A specific hydrolysis reaction was then carried out at 400 rpm for 4 h. After the reaction, the product was centrifuged at 4000 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 24.96% higher fatty alkanols, 30.22% higher fatty acids, and 44.82% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 1.3 L / h, and the temperatures of the first-stage evaporation surface were 140℃, the second-stage evaporation surface 150℃, the third-stage evaporation surface 170℃, and the fourth-stage evaporation surface 180℃. After distillation, the third and fourth stage light phases were rice bran fatty alkanols. Liquid chromatography analysis showed that the purity of the rice bran fatty alkanols in the distillation product was 97.4%.

[0047] Example 7 200 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 40 g of pH 6.5, 20 mM phosphate buffer (20% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 45°C, and then Lipase AOL (220 U / g rice bran wax paste, corresponding to a total enzyme activity of 44000 U) was added. A specific hydrolysis reaction was then carried out at 500 rpm for 2 h. After the reaction, the product was centrifuged at 4500 rpm for 10 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 23.54% higher fatty alkanols, 30.87% higher fatty acids, and 45.59% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 1.6 L / h, and the temperatures of the first-stage evaporation surface were 145℃, the second-stage evaporation surface 155℃, the third-stage evaporation surface 178℃, and the fourth-stage evaporation surface 185℃. After distillation, the third and fourth stage light phases were rice bran fatty alkanols. Liquid chromatography analysis showed that the purity of the rice bran fatty alkanols in the distillation product was 97.3%.

[0048] Example 8 240 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 36 g of pH 7.0, 20 mM phosphate buffer (15% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 40°C, and then Lipase G "Amano" 50 (200 U / g rice bran wax paste, corresponding to a total enzyme activity of 48000 U) was added. A specific hydrolysis reaction was then carried out at 500 rpm for 2 h. After the reaction, the product was centrifuged at 5000 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 21.84% higher fatty alkanols, 31.26% higher fatty acids, and 46.90% triglycerides. The upper oil phase was separated and purified using four-stage molecular distillation. The feed flow rate was 1.8 L / h, and the temperatures of the first-stage evaporation surface were 145℃, the second-stage evaporation surface 155℃, the third-stage evaporation surface 180℃, and the fourth-stage evaporation surface 185℃. After distillation, the third and fourth stage light phases were rice bran fatty alkanols. Liquid chromatography analysis showed that the purity of the rice bran fatty alkanols in the distillation product was 97.9%.

[0049] Comparative Example 1 220 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 33 g of 20 mM phosphate buffer (pH 6.5, 15% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 40°C, and then 100 L of Lipozyme TL (200 U / g rice bran wax paste, corresponding to a total enzyme activity of 44000 U) was added. A specific hydrolysis reaction was carried out at 400 rpm for 2 h. After the reaction, the product was centrifuged at 4000 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 12.38% higher fatty alkanols, 27.64% triglycerides, 3.87% diglycerides, and 56.11% fatty acids. The upper oil phase was separated and purified by four-stage molecular distillation. The feed flow rate was 1.5 L / h, the first-stage evaporation surface temperature was 145℃, the second-stage evaporation surface temperature was 155℃, the third-stage evaporation surface temperature was 180℃, and the fourth-stage evaporation surface temperature was 185℃. After distillation, the first and second stage light phases consisted of fatty acids and a small amount of diglycerides, the third and fourth stage light phases consisted of rice bran fatty alkanols, and the fourth stage heavy phase consisted of triglycerides. Liquid chromatography analysis of the third and fourth stage light phases revealed that the rice bran fatty alkanol content in the distillation product was 91.2%.

[0050] Comparative Example 2 220 g of rice bran wax paste was added to a 1000 mL Erlenmeyer flask, followed by 33 g of 20 mM phosphate buffer (pH 6.5, 15% of the rice bran wax paste mass). After thorough mixing, the mixture was heated to 40°C, and then 20000 L of Palatase (200 U / g rice bran wax paste, corresponding to a total enzyme activity of 44000 U) was added. A specific hydrolysis reaction was carried out at 400 rpm for 2 h. After the reaction, the product was centrifuged at 4000 rpm for 15 min. Liquid chromatography analysis of the upper oil phase revealed that the product contained 9.08% higher fatty alkanols, 5.63% diglycerides, 31.47% triglycerides, and 53.82% fatty acids. The upper oil phase was separated and purified by four-stage molecular distillation. The feed flow rate was 1.5 L / h, the first-stage evaporation surface temperature was 145℃, the second-stage evaporation surface temperature was 155℃, the third-stage evaporation surface temperature was 180℃, and the fourth-stage evaporation surface temperature was 185℃. After distillation, the first and second stage light phases consisted of fatty acids and a small amount of diglycerides, the third and fourth stage light phases consisted of rice bran fatty alkanols, and the fourth stage heavy phase consisted of triglycerides. Liquid chromatography analysis of the third and fourth stage light phases revealed that the rice bran fatty alkanol content in the distillation product was 88.6%.

[0051] As shown in Table 1, when the reaction conditions and purification processes of Example 1, Comparative Examples 1 and 2 are completely identical, changing only the type of lipase results in significant differences in the synthesis efficiency and purification effect of rice bran fatty alkanols. When using a glyceryl ester lipase, the content of higher fatty alkanols in the upper oil phase remains stable at approximately 18%–21%, and after four-stage molecular distillation purification, the purity of the rice bran fatty alkanols can be maintained above 96%. In contrast, if triglyceride lipase is used as a catalyst, the content of higher fatty alkanols in the upper oil phase is only about 9%–12%, only 40%–50% of that in the examples, and the alkanol yield decreases significantly; even using the same molecular distillation process, the purity of the purified rice bran fatty alkanols still drops to 88%–92%. These results indicate that triglyceride lipase has a limited ability to release fatty alkanols from wax esters, while glyceryl ester lipase is the key to achieving the expected technical effect.

[0052] Table 1 compares the yield and purity of enzyme types and rice bran fatty alkanols in each example and comparative example.

[0053] The above examples demonstrate that the method of the present invention can stably prepare high-purity rice bran fatty alkanols under different process parameters, proving the reliability and superiority of the present invention.

[0054] The endpoints and any values ​​of the ranges disclosed in this invention are not limited to the precise ranges or values; these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein. In the following, various technical solutions can, in principle, be combined with each other to obtain new technical solutions, which should also be considered as specifically disclosed herein.

[0055] The above embodiments are only used to illustrate the technical solutions of the present invention and not to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art can still make modifications or equivalent substitutions to the specific implementation of the present invention. Any modifications or equivalent substitutions that do not depart from the spirit and scope of the present invention are within the protection scope of the claims of the present invention pending approval.

Claims

1. A process for the enzymatic preparation of a rice bran fatty alkanol, characterized in that, The method comprises the following steps: adding partial glyceride lipase and buffer to the rice bran wax paste to perform a hydrolysis reaction to obtain a reaction product; separating the reaction product, recovering the upper oil phase, and purifying the upper oil phase to obtain rice bran fatty alkanol.

2. The process for the enzymatic production of rice bran fatty alkanols according to claim 1, characterized in that, The partial glyceride lipase is one or more of Lipase G "Amano" 50, Lipase SMG1, and Lipase AOL. The amount of the partial glyceride lipase added is 100-300 U / g based on the mass of the rice bran wax paste.

3. The method of enzymatic production of rice bran fatty alkyl alcohol according to claim 1, characterized in that, The content of triglyceride in the rice bran wax paste is 55%-65%, and the content of rice bran wax is 35%-45%.

4. The process for enzymatic production of rice bran fatty alkanols according to claim 1, characterized in that, The buffer is a phosphate buffer, the pH of the phosphate buffer is 5.5-7.5, the ionic strength of the phosphate buffer is 20 mM, and the amount of the phosphate buffer added is 5%-30% of the mass of the rice bran wax paste.

5. The process for enzymatic production of rice bran fatty alkanols according to claim 1, wherein, The temperature of the hydrolysis reaction is 30-45℃.

6. The process for enzymatic preparation of rice bran fatty alkanols according to claim 1, wherein, The separation is centrifugal separation, the rotation speed is 3000-5000 rpm, and the centrifugal time is 10-20 min.

7. The process for enzymatic production of rice bran fatty alkanols according to claim 1, wherein, The upper oil phase comprises rice bran fatty alkanol, higher fatty acid, and triglyceride; the rice bran fatty alkanol comprises one or more of octacosanol, triacontanol, and dotriacontanol.

8. The process for enzymatic preparation of rice bran fatty alkanols according to claim 1, wherein, The separation and purification are performed by molecular distillation, the feed flow rate is 1-2 L / h, the temperature of the first evaporation surface is 140-145℃, the temperature of the second evaporation surface is 155-160℃, the temperature of the third evaporation surface is 175-180℃, and the temperature of the fourth evaporation surface is 180-185℃.

9. A rice bran fatty alkyl alcohol, characterized by, The method is prepared according to any one of claims 1-8.

10. The rice bran fatty alkanol of claim 9 is applied in the fields of food, medicine, daily chemical, and feed industry.

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