A method for physically synergistically extracting high-purity nervonic acid from malanerds

Abstract

The application belongs to the technical field of natural product extraction and separation, and particularly relates to a method for physically and synergistically extracting high-purity nervonic acid from Malania oleifera. The method comprises the following steps: (1) raw material pretreatment and oil extraction, wherein steam explosion and pulse electric field treatment are used to destroy seed kernel cell walls and improve oil yield; (2) saponification and metal salt precipitation, wherein alkali liquor and a non-aqueous solvent are used to saponify oil, and glycerol and toxic protein impurities are effectively removed through precipitation separation; (3) acidification and washing, wherein metal soaps are converted into mixed fatty acids; (4) fractional precipitation, wherein a first organic solvent is used to separate saturated fatty acids and low-melting-point impurities through two-stage variable-temperature precipitation, and crude nervonic acid is obtained; and (5) recrystallization and drying, wherein the crude nervonic acid is further purified to obtain a finished product. The method has the advantages of simple process and low cost, and solves the problems of existing technologies, such as difficult trade-off between purity and yield, residual toxic proteins and the like. The obtained nervonic acid has a purity of greater than or equal to 98% and a yield of greater than or equal to 70%, and has extremely high industrial application value and safety.

Description

Technical Field

[0001] This invention belongs to the field of natural product extraction and separation technology, specifically relating to a method for the physical and synergistic extraction of high-purity nervonic acid from garlic fruit. Background Technology

[0002] Nervonic acid (cis-15-tetracosenoic acid) is a long-chain monounsaturated fatty acid that plays an important role in repairing brain nerve fibers and promoting nerve cell growth, and has broad application prospects in the fields of medicine, health products, and functional foods. The market demand for nervonic acid continues to grow, but efficient extraction technology of natural nervonic acid has become a key bottleneck for the industry's development.

[0003] The garlic tree (Malania oleifera) is a unique and rare relict tree species, and its nervonic acid content ranks among the highest of all known plants (reaching 40-50%), making it an ideal raw material for the preparation of natural nervonic acid and possessing extremely high development and utilization value. However, garlic tree resources are scarce, and how to efficiently utilize garlic tree raw materials to achieve high-purity and high-yield extraction of nervonic acid has become an urgent problem to be solved in the industry.

[0004] Currently, the main methods for extracting nervonic acid from garlic fruit include column chromatography, molecular distillation, urea inclusion method, solvent extraction, and recrystallization. However, these methods generally have many shortcomings and cannot meet the needs of industrial production and high-quality products. The specific defects are as follows: The difficulty in balancing purity and yield: Existing methods generally suffer from the inability to simultaneously achieve ideal levels of product purity and yield. For example, the nervonic acid extraction and separation method disclosed in patent application CN105503580A can achieve a nervonic acid purity of over 90%, but the yield is less than 30%, resulting in significant resource waste. The separation and preparation method of cis-15-tetracosenoic acid disclosed in patent authorization announcement CN1050117C can achieve a yield of around 60%, but the product purity is only around 80%, which is insufficient to meet the purity requirements of pharmaceuticals and high-end health products.

[0005] Complex processes and high production costs: Some extraction methods (such as multiple column chromatography and multi-stage molecular distillation) are cumbersome to operate, require high-precision equipment, and need to use large amounts of organic solvents. This not only increases production costs but also brings environmental pressure and prolongs the production cycle, making them unsuitable for large-scale industrial production. For example, the patent application with publication number CN116283551A uses multi-stage molecular distillation combined with multiple column chromatography purification, which can obtain nervonic acid with high purity, but the process steps are cumbersome, the equipment investment is large, the solvent consumption is high, and the economic efficiency is poor.

[0006] Impurities are difficult to remove efficiently: In addition to the target product nervonic acid, garlic fruit oil also contains long-chain fatty acid analogs such as oleic acid, erucic acid, and behenic acid, as well as naturally occurring toxic proteins. These impurities have similar physicochemical properties to nervonic acid, making efficient and thorough separation and removal difficult using existing methods. Residual toxic proteins can affect product safety and limit its application in the food and health product sectors; while residual long-chain fatty acid analogs can reduce product purity and affect product quality.

[0007] Low raw material utilization: In traditional extraction methods, the cell walls of garlic kernels are difficult to be fully destroyed, resulting in a low oil extraction rate. The potential extraction amount of nervonic acid is not fully utilized, further reducing the overall yield and wasting the rare resource of garlic.

[0008] Therefore, there is an urgent need to develop a method that simplifies the process, controls costs, increases raw material utilization, effectively removes harmful impurities, and achieves high-purity and high-yield extraction of nervonic acid. This would address many shortcomings of existing technologies and promote the efficient development and utilization of garlic fruit resources and the large-scale development of the nervonic acid industry. Summary of the Invention

[0009] This invention aims to provide a method for the physical and synergistic extraction of high-purity nervonic acid from garlic fruit, addressing the problems of complex processes, difficulty in balancing product purity and yield, ineffective removal of harmful impurities (such as toxic proteins) and long-chain fatty acid analogs, and low raw material utilization in existing technologies. The method of this invention should be simple to operate, cost-controllable, suitable for industrial production, and able to obtain nervonic acid products with high purity (>98%), high content (>95%), and high yield (>70%), while simultaneously improving raw material utilization and product safety. Technical solution

[0010] This invention provides a method for the physical and synergistic extraction of high-purity nervonic acid from garlic fruit, characterized by the following key steps:

[0011] 1. Raw material pretreatment: The garlic kernels are dried and pulverized to 100-200 mesh to remove impurities, resulting in garlic kernel powder. This ensures that the raw materials are evenly dispersed and increases the contact area in the subsequent extraction process.

[0012] Optionally, the garlic kernel powder is subjected to steam explosion treatment under the following conditions: steam pressure 3-8 MPa, heating power 4-8 kW, and treatment time 1-5 min. Through the mechanical and thermal effects of steam explosion, the cell wall structure of the kernel is destroyed, making it easier for the oil to seep out and increasing the oil yield.

[0013] Optionally, the kernel powder after steam explosion can be treated with a pulsed electric field (PEF) to further disrupt the cell wall, increase the oil yield, and simultaneously achieve preliminary denaturation of some toxic proteins. The treatment conditions are: pulse voltage 30-45kV, pulse frequency 750-850Hz, pulse width 5-15μs, and treatment time 120-240s.

[0014] 2. Oil extraction: Pretreated seed kernel powder can be extracted using a non-polar solvent (such as petroleum ether or n-hexane) via Soxhlet extraction or reflux extraction at a temperature of 60-85℃ for 4-10 hours to ensure full extraction of the oil; or a green solvent (such as 95% ethanol) can be used for low-temperature extraction at 40-60℃ 2-3 times, with each extraction lasting 2-3 hours, to reduce the risk of oil oxidation and retain the activity of nervonic acid.

[0015] After extraction, the solvent is recovered by distillation or rotary evaporation (the solvent can be reused) to obtain garlic seed oil.

[0016] Take the garlic seed oil obtained in step one, add an alkali (selected from one or more of NaOH, KOH, and Ca(OH)2, with a weight of 0.1-0.6 times the weight of the oil) and a non-aqueous solvent (selected from one or more of 85-95% ethanol and methanol, with a volume (L) of 8-15 times the weight (kg) of the oil), and stir well.

[0017] The mixture is heated and refluxed for saponification for 30-90 minutes, allowing the fatty acids in the garlic seed oil to fully react with the alkali and produce fatty acid metal salts (soap).

[0018] After the saponification reaction is complete, the saponification solution is cooled to room temperature to -5°C and allowed to stand for 1-4 hours to allow the fatty acid metal salts (soap) to fully precipitate out, while impurities such as glycerol and toxic proteins dissolve in the solution.

[0019] Filtration removes solutions containing impurities such as glycerol and toxic proteins, while retaining the filter residue (i.e., fatty acid metal salts, or metal soaps for short), thus achieving preliminary separation of impurities.

[0020] Add the filter residue (metal soap) obtained in step two to acid water (selected from one or more of 0.5-2 mol / L HCl and H2SO4, with a volume (L) of 5-20 times the weight (kg) of the oil), stir evenly, and acidify at 50-95℃ for 15-75 minutes to convert the fatty acid metal salt into free fatty acid, thus obtaining a mixed fatty acid solution.

[0021] After acidification, the mixture is allowed to stand at room temperature to separate into layers. The upper water layer (containing residual acid, salts and other water-soluble impurities) is discarded, and the lower oil layer (i.e., mixed fatty acids) is retained.

[0022] Wash the oil layer with deionized water 2-7 times. After each wash, let it stand to separate into layers, discard the water layer, until the oil layer is neutral. Remove residual acid and water-soluble impurities to obtain a water-washed mixed fatty acid oil layer.

[0023] Add a first organic solvent (selected from one or more of anhydrous or high-concentration acetone, methanol, and ethanol, with a concentration ≥80%) to the mixed fatty acid oil layer obtained in step three. The volume (L) of the first organic solvent is 2-8 times the weight (kg) of the garlic fruit oil. Stir evenly to obtain a mixed solution.

[0024] First stage precipitation (low-temperature impurity removal): The mixture is allowed to stand at 5°C to room temperature (preferably 10-25°C) for 1 hour to overnight to allow precipitation. This stage mainly causes saturated fatty acids and some low-melting-point unsaturated fatty acids (impurities with significantly different physicochemical properties from nervonic acid) to precipitate. Filter, discard the filter residue, and retain the filtrate (containing nervonic acid and a small amount of impurities).

[0025] Second-stage precipitation (target product crystallization): Cool the filtrate from the previous step to -20°C to 5°C (preferably -15°C to 0°C), and let it stand for 1 hour to overnight. During this stage, the target product, nervonic acid, crystallizes out due to decreased solubility. Filter and collect the filter residue to obtain crude nervonic acid.

[0026] Add the crude nervonic acid obtained in step four to a second organic solvent (selected from one or more of anhydrous or high-concentration methanol, ethanol, and acetone, with a concentration ≥80%). The volume (L) of the second organic solvent is 2-8 times the initial weight (kg) of garlic fruit oil.

[0027] The mixture is heated to 30-60°C and stirred until the crude nervonic acid is completely dissolved. Then, it is recrystallized at -20°C to room temperature (preferably -15°C to 10°C) and left to stand for 1 hour to overnight to further purify the nervonic acid.

[0028] Filter the residue, collect the residue, place the residue in a vacuum dryer, and vacuum dry it at 40-60℃ for 4-6 hours to remove residual solvent, thus obtaining high-purity nervonic acid product.

[0029] If higher purity of nervonic acid is required (e.g., ≥99%), the recrystallized nervonic acid product can be purified by rapid dry column chromatography. The specific operation is as follows: the recrystallized product is mixed with diatomaceous earth powder, loaded onto a silica gel column (40-200 mesh silica gel for column chromatography), and gradient elution is performed using different proportions of petroleum ether-ethyl acetate, dichloromethane-ethyl acetate, or diethyl ether-ethyl acetate as the mobile phase. The target fraction with high nervonic acid content is collected by high performance liquid chromatography (HPLC). After concentrating the target fraction, it is crystallized again and dried to obtain ultra-high purity nervonic acid product.

[0030] In the solid-liquid separation stages of the entire method, such as precipitation, filtration, and washing, a dedicated precipitation separation device can be used to improve separation efficiency and accuracy. This device is characterized by: an extraction precipitation tank, a top cover, a bottom filter plate that can move vertically (for compression filtration to increase filtration speed), a side wall with multi-stage outflow components (containing multiple outflow holes of different heights), and a flow connection pipe that can be adjusted vertically and mates with the multi-stage outflow holes; the extraction precipitation tank is equipped with an observation window for easy observation of the stratification interface of the mixture.

[0031] When using this product, place the mixture to be separated into the extraction sedimentation tank, allow it to stand and separate into layers, observe the separation interface through the observation window, move the flow connection tube to the corresponding height, and accurately extract the lower sediment or upper clear liquid. Avoid disturbing the separation interface, thereby improving the efficiency and purity of solid-liquid separation and reducing the loss of the target product.

[0032] Saponification alkali dosage: 0.2-0.4 times the weight of garlic fruit oil. This range ensures that the saponification reaction proceeds fully, while avoiding excessive alkali that would lead to excessive consumption of acid in subsequent acidification steps.

[0033] Non-aqueous solvent: 85-95 v / v% ethanol, used at a rate of 10-15 times (L / kg). Ethanol is a green solvent with low toxicity and easy recovery. This amount can ensure the smooth progress of the saponification reaction while reducing solvent consumption.

[0034] Saponification time: 45-75 minutes. This range allows for the full saponification of fatty acids, avoiding incomplete saponification that could lead to incomplete precipitation of fatty acids after subsequent acidification, while also shortening the production cycle.

[0035] Acid concentration: 0.5-1.5 mol / L, dosage: 8-12 times (L / kg), acidification temperature: 55-75℃. This parameter range can ensure that the fatty acid metal salt is fully converted into free fatty acid, while avoiding the destruction of the nervonic acid structure due to excessively high acid concentration or temperature.

[0036] The first solvent for fractional precipitation is 90-100 v / v% acetone or methanol, used in 3-6 times (L / kg). The precipitation temperature for the first stage is 10-25℃, and the precipitation temperature for the second stage is -15 to 0℃. These parameters can accurately separate saturated fatty acids and low-melting-point unsaturated fatty acids, thereby improving the purity of crude nervonic acid.

[0037] Recrystallization solvent: 80-100 v / v% methanol or ethanol, 3-6 times (L / kg), recrystallization temperature -15 to 10℃. These parameters can further remove residual impurities in crude nervonic acid and ensure the purity of the finished product.

[0038] Raw material pretreatment: Steam explosion pressure 5-6MPa, heating power 6-7kW; PEF treatment pulse voltage 35-40kV, pulse frequency 800-810Hz, pulse width 8-10μs, treatment time 180-200s. This parameter range can maximize the destruction of seed kernel cell walls, increase oil yield, and achieve preliminary denaturation of toxic proteins. Beneficial effects

[0039] Compared with the prior art, the present invention has the following significant advantages: 1. High purity and high yield are achieved synergistically: The "saponification-metal salt precipitation" step effectively removes water-soluble impurities such as glycerol and toxic proteins. Combined with the innovative "fractional precipitation" core technology, fatty acid analogs with different carbon chain lengths and melting points are precisely separated, achieving highly efficient separation of nervonic acid from impurities. In the final product, the nervonic acid content, calculated on a dried basis, is not less than 95% (preferably not less than 97%), and the purity, calculated by HPLC area normalization, is not less than 98% (preferably not less than 99%). Based on the total amount of nervonic acid in garlic fruit oil, the yield is not less than 70% (preferably not less than 75%). Example data show that the method of this invention can achieve excellent results with purity >98%, content >98%, and yield >75%, significantly superior to existing technologies.

[0040] 2. Simplified process and low production cost: The main steps of this invention are all conventional saponification, acidification, precipitation and crystallization operations, avoiding complex and expensive column chromatography (except for optional steps) or molecular distillation equipment. The operation is simple and easy to master. The solvent used can be recovered and reused through distillation, which greatly reduces solvent consumption and production costs, making it suitable for large-scale industrial production.

[0041] 3. High product safety: Through the "saponification-metal salt precipitation" step, naturally occurring toxic proteins in garlic fruit oil can be effectively removed, solving the problem that toxic proteins are difficult to completely remove in existing technologies, improving the safety of the product as a raw material for food and health products, and expanding the scope of application of the product.

[0042] 4. Flexible operation and strong controllability: By adjusting the temperature, solvent ratio and time of the fractionation precipitation, as well as the key parameters of saponification and acidification, the purity and yield of the product can be flexibly controlled to meet the purity requirements of nervonic acid products in different fields; the dedicated precipitation separation device makes the solid-liquid separation operation more precise and efficient, reduces the loss of target products, and further improves the controllability of the process.

[0043] 5. High raw material utilization rate: Combining pretreatment technologies such as steam explosion and pulsed electric field can significantly destroy the cell walls of garlic kernels, enabling the oil extraction rate to reach over 99%, fully leveraging the potential value of garlic raw materials, reducing the waste of rare resources, and improving the overall economic efficiency of the process. Specific Implementation

[0044] The present invention will be described in detail below with reference to specific embodiments, but the scope of protection of the present invention is not limited to these embodiments. Experimental methods not specifically described in the embodiments are generally performed under conventional conditions or as recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products. Example

[0045] This embodiment provides a method for physically and synergistically extracting high-purity nervonic acid from garlic fruit, comprising the following steps: 1. Raw material pretreatment and oil extraction: Garlic kernels were dried in a vacuum drying oven at 60℃ until constant weight, then pulverized and passed through a 200-mesh sieve to remove impurities, yielding garlic kernel powder. The kernel powder was freeze-dried and then subjected to steam explosion treatment at a steam pressure of 5 MPa, a heating power of 6 kW, and a treatment time of 3 min. The exploded kernel powder was then placed in a pulsed electric field treatment chamber with a fixed electrode spacing of 45 mm, a pulse voltage of 38 kV, a residence time of 190 s, a pulse frequency of 805 Hz, and a pulse width of 9 μs. After treatment, a 95 v / v% ethanol solution (solid-to-liquid ratio 1:10, g / mL) was added to the kernel powder, and extraction was performed twice at 50℃ for 2.5 h each time. The two extracts were combined, and the ethanol was recovered using a rotary evaporator to obtain garlic kernel oil.

[0046] 2. Saponification - Metal Salt Precipitation: Take 1 kg of garlic seed oil, add 0.3 kg of NaOH, then add 12 L of 90 v / v% ethanol, stir evenly, and heat under reflux for 60 min for saponification; cool the saponification solution to 5°C, let it stand for 2 h for precipitation, filter it using a special precipitation separation device to remove the solution containing impurities such as glycerol and toxic proteins, and obtain the filter residue.

[0047] 3. Acidification and washing: Add 10L of 1mol / L hydrochloric acid to the filter residue, stir evenly, and acidify at 70℃ for 45min; after acidification, let it stand at room temperature to separate into layers, discard the water layer, and wash the oil layer with deionized water 5 times until the oil layer is neutral, to obtain the water-washed mixed fatty acid oil layer.

[0048] 4. Graded sedimentation: Add 6L of 90v / v% acetone to the above oil layer, stir evenly, and let it stand at 20℃ for 4h (first stage precipitation). Filter using a special precipitation separation device and discard the filter residue. Cool the filtrate to -10℃ and let it stand for 6h (second stage precipitation). Filter and collect the filter residue to obtain crude nervonic acid.

[0049] 5. Recrystallization and drying: Add crude nervonic acid to 5L of 90v / v% ethanol, heat to 45℃ and stir until completely dissolved, then recrystallize at -5℃ for 4h, filter and collect the filter residue; place the filter residue in a vacuum dryer and dry at 50℃ and -0.09MPa for 4h to obtain the finished nervonic acid product.

[0050] Results: The obtained nervonic acid product had a content of 98.9% on a dried basis, a purity of 99.7% by HPLC area normalization method, and a yield of 76.5% calculated based on the total amount of nervonic acid in garlic seed oil. The test showed that the residual amount of toxic protein was below the detection limit, and the product safety met the standards for food and health products. Example

[0051] The difference between this embodiment and Embodiment 1 is that the steam explosion and pulsed electric field pretreatment steps in S1 are omitted, and the pulverized kernel powder is directly extracted with 95v / v% ethanol; at the same time, in the S4 fractionation precipitation step, only one-step low-temperature precipitation (-10℃, overnight standing) is used, without the first-stage room-temperature precipitation. The remaining steps are the same as in Embodiment 1.

[0052] Results: The obtained nervonic acid product had a content of 88.3% on a dried basis, a purity of 91.6% by HPLC area normalization method, and a yield of 68.2%. The residual amount of toxic protein was 0.03 mg / kg, which did not meet the food-grade safety standard. Compared with Example 1, the product purity, content, and yield were all significantly reduced, and the safety was insufficient, indicating that the raw material pretreatment and fractionation precipitation steps are crucial to improving product quality and safety. Example

[0053] The difference between this embodiment and Embodiment 1 is that in S2 saponification-metal salt precipitation, NaOH is replaced with an equal amount (0.3 kg) of KOH, and 90 v / v% ethanol is replaced with 90 v / v% methanol, while other conditions are the same as in Embodiment 1.

[0054] Results: The obtained nervonic acid product had a content of 97.9% on a dried basis, a purity of 98.8% by HPLC area normalization method, and a yield of 74.1%; the residual amount of toxic protein was below the detection limit. The results indicate that high-purity and high-yield nervonic acid products can also be obtained using KOH and methanol, but the effect is slightly inferior to the preferred scheme in Example 1 (NaOH + 90 v / v% ethanol). Example

[0055] The difference between this embodiment and Example 1 is that, after the S5 drying step, a column chromatography deep purification step is added: the dried nervonic acid product is mixed with diatomaceous earth powder, loaded onto a 120-mesh silica gel column, and gradient elution is performed using petroleum ether-ethyl acetate (volume ratio 1:1) as the mobile phase. The target fraction with a nervonic acid content ≥99.5% is collected by HPLC monitoring. After concentrating the target fraction, 4L of 95v / v% ethanol is added, and the mixture is recrystallized at -8℃ for 3h. After filtration, the mixture is vacuum dried at 50℃ to obtain an ultra-high purity nervonic acid product.

[0056] Results: The obtained ultra-high purity nervonic acid product had a content of 99.3% on a dried basis, a purity of 99.9% by HPLC area normalization method, and a yield of 73.8%, which can meet the extremely high requirements of the pharmaceutical field for the purity of nervonic acid. Attached Figure Description

[0057] Figure 1 : Process flow diagram of this invention.

[0058] Garlic kernel → Drying and pulverizing → Optional (steam explosion + PEF pretreatment) → Solvent extraction → Solvent recovery → Garlic kernel oil → Saponification - metal salt precipitation → Filtration → Filter residue → Acidification → Layered washing → Mixed fatty acid oil layer → Graded precipitation (first stage impurity removal + second stage crystallization) → Filtration → Crude nervonic acid → Recrystallization → Filtration → Drying → High-purity nervonic acid product; Optional steps: Deep purification by column chromatography after recrystallization → Recrystallization and drying → Ultra-high purity nervonic acid product; Solid-liquid separation throughout the process can be achieved using a dedicated precipitation separation device.

Claims

1. A method for physically and synergistically extracting high-purity nervonic acid from garlic fruit, characterized in that, Includes the following steps: Saponification – metal salt precipitation, acidification, water washing, fractional precipitation and recrystallization.

2. The method according to claim 1, characterized in that, The graded precipitation includes a first-stage precipitation and a second-stage precipitation; the first-stage precipitation is carried out at 5°C to room temperature, mainly to remove saturated fatty acid impurities; the second-stage precipitation is carried out at -20°C to 5°C, causing nervonic acid crystals to precipitate.

3. The method according to claim 1, characterized in that, In the saponification-metal salt precipitation step, the alkali used is one or more of NaOH, KOH or Ca(OH)2, and the amount used is 0.2-0.4 times the weight of garlic fruit oil.

4. The method according to claim 1, characterized in that, In the saponification-metal salt precipitation step, the non-aqueous solvent used is 85-95% (v / v) ethanol or methanol, and the amount used is 8-15 times (L / kg, based on the weight of garlic fruit oil), and the saponification reaction time is 45-75 minutes.

5. The method according to claim 1, characterized in that, Before extracting the oil, the method further includes a step of steam explosion and / or pulsed electric field pretreatment of the garlic kernels; the conditions for steam explosion are: steam pressure 3-8MPa, heating power 4-8kW, and treatment time 1-5min; the conditions for pulsed electric field treatment are: pulse voltage 30-45kV, pulse frequency 750-850Hz, pulse width 5-15μs, and treatment time 120-240s.

6. The method according to claim 1, characterized in that, In the acidification step, the acid used is 0.5-1.5 mol / L HCl or H2SO4, and the amount used is 8-12 times (L / kg, based on the weight of garlic fruit oil). The acidification temperature is 55-75℃, and the acidification time is 15-75 minutes.

7. The method according to claim 1, characterized in that, In the fractionation precipitation step, the first organic solvent is 90-100 v / v% acetone or methanol, and the amount used is 3-6 times (L / kg, based on the weight of garlic fruit oil); in the recrystallization step, the second organic solvent is 80-100 v / v% methanol or ethanol, and the amount used is 3-6 times (L / kg, based on the weight of garlic fruit oil).

8. The method according to claim 1, characterized in that, The method uses a dedicated sedimentation separation device for solid-liquid separation. The device includes an extraction sedimentation tank, a top cover, a bottom filter plate that can move up and down, a side wall with multi-stage outflow components, and a flow connection pipe that can be adjusted up and down. The extraction sedimentation tank is equipped with an observation window.

9. The method according to claim 1, characterized in that, It also includes a column chromatography deep purification step: the recrystallized product is mixed with diatomaceous earth powder, loaded onto a silica gel column, and eluted with a gradient of petroleum ether-ethyl acetate, dichloromethane-ethyl acetate or diethyl ether-ethyl acetate as the mobile phase. The target fraction is collected, concentrated, recrystallized, and dried.

10. The method according to any one of claims 1-9, characterized in that, The obtained nervonic acid product has a content of ≥95% on a dried basis, a purity of ≥98% by HPLC area normalization method, and a yield of ≥70% based on the total amount of nervonic acid in garlic fruit oil.

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