Method for extracting camellia oleosa seed oil through wet method assisted by enzyme method

Abstract

The invention provides a method for extracting oil-tea camellia seed oil by an enzymic method assisted wet method, which comprises the following steps: shelling oil-tea camellia seeds to obtain oil-tea camellia seed kernels; crushing and sieving the camellia seed kernels to obtain camellia seed kernel powder; adding 1%-3% of an oil extraction aid NaCl and 2%-4% of a compounded enzyme preparation into the camellia seed kernel powder by taking the weight of the camellia seed kernel powder as a reference, adding 10%-15% of a buffer solution, and stirring and rubbing for 40-60 minutes at the temperature of 50-60 DEG C until aggregated particles are clustered and are difficult to stir, so as to obtain camellia seed oil; the enzyme preparation is formed by compounding pectinase of an enzyme-linked enzyme preparation, cellulase of the enzyme-linked enzyme preparation and protease of the enzyme-linked enzyme preparation. The method has the beneficial effects that the oil extraction rate of the camellia oleosa seed oil can be obviously improved, the residual oil rate of the camellia oleosa seed cakes can be reduced and the nutritional ingredients of the camellia oleosa seed oil can be well reserved by compounding the three enzymes, namely the intimate cellulase, the intimate pectinase and the intimate protease to perform enzyme treatment and then performing wet-process oil extraction and selecting the oil extraction auxiliary agent NaCl, so that the oil extraction rate of the camellia oleosa seed oil can be obviously improved and the residual oil rate of the camellia oleosa seed cakes can be reduced.

Description

Technical Field

[0001] The invention belongs to the technical field of extracting camellia oil, and mainly relates to a method for extracting camellia oil by an enzyme-assisted wet method. Background Art

[0002] Camellia oleifera Abel. belongs to the genus Camellia of the Theaceae family. It is known as one of the four major woody oil crops in the world, along with olive, oil palm and coconut. The fatty acid composition of camellia oil is similar to that of olive oil. It is a green, ecological, highly nutritious and healthy high-grade edible oil, known as the "Oriental Olive Oil". The oil extracted from its seeds contains many unsaturated fatty acids (90%), mainly oleic acid (80%). In addition, camellia oil also contains many healthy ingredients and beneficial bioactive compounds, such as sterols, squalene and tocopherol. Wang et al. detected 9 squalenes and 13 sterols from three types of camellia oil, with a total content of 2860.18-7626.23 mg / kg, and found that camellia oil had the highest content of sterols and squalene, and its tea oil quality was better. Wang Yaping and others tested six kinds of oils and fats, among which the α-tocopherol content of camellia oil was 19.30 mg / 100 g, which is much higher than that of other oils and fats. It can be seen that among the six oils, camellia oil has higher physiological activity. In life, the application range of camellia oil and its cake is very wide. In particular, camellia oil is rich in oleic acid, triterpenes, polyphenols and other bioactive substances with health-promoting properties, and has broad application prospects in food, cosmetics, medicine and other fields. With the increase in people's health needs, the development and utilization of the nutritional and health value of camellia oil has gradually become a hot topic in current research.

[0003] The main extraction methods of camellia oil include pressing, water replacement, solvent extraction, and water enzyme method. The oil yield of the pressing method is low, the protein is denatured by high temperature and high shear, the availability is low, and the production process is energy-consuming and time-consuming. The solvent extraction method has a high oil extraction rate, a large processing volume, a low oil extraction cost, a high quality of the extracted meal, and a high resource utilization rate. However, during the extraction, once the solvent evaporates into the environment, it is easy to cause environmental pollution. The water replacement method is low-cost and simple in equipment, but an emulsion layer is easily formed between oil and water, which needs to be treated by demulsification technology, and the water and volatile content cannot meet the requirements of GB / T 11765-2018 "Camellia Oil". The wet oil extraction is based on the study of the traditional water replacement method to use the cohesive force between hydrophilic molecules to squeeze out oil and other oil extraction principles, and draws on and refers to the ALCON conditioning principle to develop a new wet extraction technology for camellia oil. Although wet oil extraction can extract camellia oil at low temperatures and retain most of the bioactive components in the oil and is environmentally friendly, the oil extraction effect of this extraction method is not ideal and is time-consuming and labor-intensive. The water enzyme method is based on mechanical crushing, using enzymes that can degrade plant oil cells or have degradative effects on complexes such as lipoproteins and lipopolysaccharides to act on oil, so that oil is released from the oil, and the oil and non-oil components are separated by using the difference in affinity of non-oil components (proteins and carbohydrates) to oil and water and the different specific gravity of oil and water. The addition of enzymes in the water enzyme method destroys the membrane structure of the plant oil cell wall and liposomes, thereby promoting the release of oil. The present invention studies the influence of enzyme types on the extraction efficiency of camellia oil in enzyme-assisted wet oil extraction through this feature of the enzyme method, in order to develop a new method for enzyme-assisted wet oil extraction. Summary of the invention

[0004] The purpose of the present invention is to overcome the shortcomings of the prior art and provide a method for extracting camellia oil by enzyme-assisted wet method.

[0005] The object of the present invention is achieved by the following technical solution: A method for extracting camellia oil by enzyme-assisted wet method, the method comprising the following steps:

[0006] Step 1: Clean, remove impurities from fresh camellia seeds, dry them to a moisture content of 4%-6%, and then shell them to obtain camellia seed kernels;

[0007] Step 2: Grind the camellia seed kernels and pass them through a 50-60 mesh sieve to obtain camellia seed kernel powder;

[0008] Step 3: Add 1%-3% NaCl and 2%-4% compound enzyme preparation to the camellia kernel powder based on the weight of the camellia kernel powder, and add 10%-15% buffer solution, stir and knead at a temperature of 50° C.-60° C. for 40 min-60 min until the aggregated particles are agglomerated and difficult to stir, centrifuge and pour out the oil phase to obtain camellia seed oil;

[0009] The enzyme preparation is compounded by pectinase from British enzyme preparation, cellulase from British enzyme preparation and protease from British enzyme preparation.

[0010] As a preferred technical solution, in the step three, based on the weight of camellia oleifera seed powder, 1% of an oil extraction aid and 3% of a compound enzyme preparation are added, and 12% of a buffer solution is added, and the mixture is stirred and kneaded at a temperature of 55°C for 50 minutes until the aggregated particles are clumped and difficult to stir, and the mixture is centrifuged and the oil phase is poured out to obtain camellia oleifera oil.

[0011] As a preferred technical solution, the enzyme preparation is compounded by pectinase of British enzyme preparation, cellulase of British enzyme preparation and protease of British enzyme preparation in a weight ratio of 1-2:1-2:1-2.

[0012] As a preferred technical solution, the buffer solution is prepared from pure water and 1 mol / L NaOH solution or HCl solution.

[0013] The beneficial effects of the present invention are:

[0014] 1. Adding 1%-3% NaCl as an oil extraction auxiliary agent can not only prevent the tea saponin in camellia oil from emulsifying when it meets water, but also improve the separation efficiency of camellia oil and medium water, thereby increasing the oil yield.

[0015] 2. The oil was extracted by wet method after enzyme treatment with a combination of AB Cellulase, AB Pectinase and AB Protease. The complex enzyme has different activities of multiple enzymes, and the activities have a synergistic effect, which is more conducive to the rupture of cell walls to release oil. It can significantly increase the oil extraction rate of camellia seed oil and reduce the residual oil rate of camellia seed cake. The residual oil rate of cake can be 9.98±0.04%.

[0016] 3. The content of polyphenols, squalene and tocopherol in the three-enzyme compound of AB-cellulase, pectinase and protease is the highest among all compounding schemes. This also proves that the three-enzyme compound of AB-cellulase, pectinase and protease has the best effect in extracting nutrients. The nutrients of camellia oil can be well preserved, and the content of polyphenols, squalene, total sterols and tocopherol in the oil reaches 111.42±4.86mg / kg, 92.75±0.33mg / kg, 3182.0±31.50mg / kg and 315.77±1.56mg / kg respectively. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art or ordinary technicians, other drawings can be obtained based on these drawings without paying creative work. Figure 1 This is a process flow chart of enzyme-assisted wet oil extraction according to the present invention. DETAILED DESCRIPTION

[0018] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the protection scope of the present invention.

[0019] like Figure 1 As shown, the present invention provides a method for enzyme-assisted wet extraction of camellia oil, the method comprising the following steps:

[0020] Step 1: Shelling the camellia seeds to obtain camellia seed kernels;

[0021] Step 2: Grinding and sieving the camellia seed kernels to obtain camellia seed kernel powder;

[0022] Step 3: Add 1%-3% of an oil extraction aid and 2%-4% of a compound enzyme preparation to the camellia seed kernel powder, based on the weight of the camellia seed kernel powder, and add 10%-15% of a buffer solution, stir and knead at a temperature of 50° C.-60° C. for 40 min-60 min until the aggregated particles are agglomerated and difficult to stir, centrifuge and pour out the oil phase to obtain camellia seed oil;

[0023] The enzyme preparation is prepared by compounding pectinase, cellulase and protease of British enzyme preparation in a weight ratio of 1-2: 1-2: 1-2, and the optimal ratio is 1: 1: 1. The buffer solution is prepared by pure water and 1 mol / L NaOH solution or HCl solution, and the function of the buffer solution is to adjust the optimal pH of the enzyme.

[0024] Experimental proof:

[0025] In the experiment, the present invention selected 17 kinds of enzymes to treat wet oil extraction, sought a solution to improve the efficiency of wet oil extraction and improve the quality of camellia oil, compared and screened out the enzyme-assisted wet oil extraction scheme with the best oil extraction effect and extraction of bioactive components in order to improve the effect of wet oil extraction.

[0026] 1. Materials and methods

[0027] 1.1 Materials and reagents

[0028] 1.1.1 Camellia oleifera

[0029] The camellia seeds used in the experiment were mature seeds of ordinary camellia oil produced in Hangzhou in 2023. They were dried, shelled and refrigerated for later use. The moisture content of the camellia oil seed kernels was determined to be 5.48±0.20%, and the oil content was 38.55±0.56%.

[0030] 1.1.2 Reagents and standards

[0031] Oil extraction additives; sodium chloride, sodium hydroxide, ethanol, acetic acid, petroleum ether, phenolphthalein, folin phenol, potassium hydroxide, anhydrous sodium bisulfate, sodium thiosulfate, hydrochloric acid, chloroform, potassium iodide, soluble starch, anhydrous sodium carbonate, isooctane were all analytical grade; methanol, n-hexane, 1,4-dioxane were all chromatographic grade.

[0032] Standard substances such as gallic acid, squalene (≥99.0%), α-tocopherol (≥96%), brassicasterol (≥97%), campesterol (≥95%), lanosterol (≥98%), stigmasterol (≥98%), β-amyrin (≥98%), Delta 5-avenasterol, Delta 7-Avenasterol, β-sitosterol (≥98.0%), and cholestanol (≥95%) were all chromatographically pure.

[0033] 1.1.3 Enzymes

[0034] The 17 enzymes used in the experiment came from five companies, namely, AB Biopharm, Aquabio, DSM, Novozymes and Yuanye. Their main characteristics are listed in Table 1-1.

[0035] Table 1-1 Types of enzymes

[0036]

[0037]

[0038] 1.2 Instruments and Equipment

[0039] The instruments and manufacturers used in this experiment are: PTY-B520 electronic balance, Huazhi (Fujian) Electronic Technology Co., Ltd.; HWS-26 electric constant temperature water bath, Shanghai Yiheng Scientific Instrument Co., Ltd.; XL-10B new sealed pulverizer, Guangzhou Xulang Mechanical Equipment Co., Ltd.; HN-50S electric constant temperature incubator, Bangsi Instrument Technology (Shanghai) Co., Ltd.; BRUKER nuclear magnetic resonance analyzer, Bruker Technology Co., Ltd.; GC2010Plus gas chromatograph, Shimadzu Instrument Co., Ltd.; 7000D gas mass spectrometer, Agilent Technologies Co., Ltd.; UV-2550 ultraviolet visible spectrophotometer, Shimadzu Instrument Co., Ltd.; BUCHI Syncore multi-sample parallel evaporator, Swiss BUCHI; SPD-15C liquid chromatograph, Shimadzu Instrument Co., Ltd., etc.

[0040] 1.3 Experimental methods

[0041] 1.3.1 Preliminary screening of wet oil extraction enzyme treatment

[0042] After the oil-tea camellia seeds are shelled, crushed and sieved, a certain amount of oil-tea camellia kernel powder is put into a 55°C constant temperature mortar, and an appropriate amount of oil extraction aid and 3% enzyme are added, and a suitable buffer solution is added. The buffer solution is prepared with pure water and 1mol / L NaOH solution and HCl solution adjusted to the optimal pH of the enzyme. Stir and knead for about 50 minutes at an appropriate temperature until the aggregated particles are agglomerated and difficult to stir, centrifuge and pour out the oil phase, repeat 3 times, collect the oil phase, and refrigerate for use. The wet oil extraction without enzyme treatment is called the blank group to serve as a control.

[0043] 1.3.2 Enzyme compounding

[0044] According to the above experimental results, the cellulase of British enzyme preparation was selected as the main enzyme, and compounded with British pectinase and British protease. The enzyme addition amount was 1:1, and other conditions were consistent with the wet oil extraction conditions. British cellulase and British pectinase were selected to compound with British protease, blue neutral protease, blue acid protease, and blue alkaline protease. The enzyme addition amount was 1:1:1. Other reaction conditions were consistent with the wet oil extraction conditions. The compounding scheme is shown in Table 1-2.

[0045] Table 1-2 Compound combination of enzymes for enzymatic assisted wet oil extraction

[0046]

[0047] 1.3.3 Quality Analysis

[0048] The method for determining the moisture content of camellia oleifera kernels shall refer to the first method of GB 5009.3-2016; the oil content of camellia oleifera kernels and the residual oil rate of cake shall be detected by nuclear magnetic resonance analyzer; the peroxide value and acid value of camellia oleifera oil shall refer to the first method of GB 5009.227-2016 and the first method of GB 5009.229-2016 respectively; the fatty acid composition of camellia oleifera oil shall refer to GB 5009.168-2016; the determination of squalene shall adopt the method of GB / T 43732-2024; the determination of polyphenols shall refer to LS / T 6119-2017; the determination of total sterols shall refer to GB25223-2010; the determination of tocopherol shall refer to GB / T 5009.82-2003.

[0049] 1.3.4 Data processing and statistics

[0050] All experimental results need to be repeated three times. Excel is used for data statistics to calculate the mean and standard deviation of the data. LSD and Duncan in IBM SPSS Statistics 26 are used for difference significance analysis. P < 0.05 is a significant difference, and P > 0.05 is an insignificant difference. Then, Origin 2018 is used to draw the graph based on the significant difference. The letters a, b, c, d, e, etc. represent significant differences, and different letters have significant differences.

[0051] 2. Experimental results

[0052] 2.1 Initial screening of enzymes

[0053] The residual oil rate of wet-process camellia oleifera cake after treatment with 17 kinds of enzymes, as well as the acid value and peroxide value of the obtained camellia oleifera oil were measured respectively. The results are shown in Table 2-1.

[0054] Table 2-1 Residual oil rate, acid value and peroxide value of various enzyme treatments

[0055]

[0056]

[0057] Note: Blank refers to wet-extracted oil without enzyme treatment.

[0058] It can be found from Table 2-1 that the residual oil rate of wet oil extraction assisted by enzyme treatment is lower than that of the blank group. The residual oil rate of wet oil extraction after enzyme treatment is 7.19%-43.12% lower than that of the blank group. This may be due to the effect of enzymes to increase the oil yield of camellia seeds. The corresponding enzyme preparations are used to hydrolyze the cell wall network, destroy the oil body membrane and the complex formed by oil and other macromolecules, thereby promoting the dissolution of cell contents and the release of free oil, so as to achieve the purpose of extracting oil from plant cells. Among them, the cake residual oil rate of the experimental group in which the cellulase of British enzyme preparation was added during the wet oil extraction process was the lowest, reaching 13.68±0.07%, which was 43.12% lower than that of the blank group. This may be because the cell wall of the camellia seed shell is mainly composed of hemicellulose, cellulose, and lignin, and each part has a layered structure. Cellulase can decompose the cellulose in the cell wall, which is conducive to the better release of camellia oil. The second is the pectinase of British enzyme preparations, which is 36.34% lower than the blank group. This may be because pectin is mainly present in the cell wall and the inner layer of the cell. Pectinase can effectively degrade pectin macromolecules, the main component of the cell wall of tea seeds, rupture the cells, and release the oil. Based on the above table, AB Cellulase, AB Pectinase, AB Protease, Azure Bio Acid Protease, and DSM Proline Specific Endoprotease were selected for the next experiment.

[0059] The acid value and peroxide value of the wet-extracted oils treated with various enzymes in Table 2-1 were significantly different (P < 0.05), but met the standards for first-grade oil in GB / T 11765-2018.

[0060] 2.2 Effect of single enzyme on camellia oil

[0061] (1) Fatty acids

[0062] The fatty acid composition of camellia oil obtained after single enzyme-assisted wet oil extraction is listed in Table 2-2.

[0063] Table 2-2 Fatty acid composition of oils treated with different enzymes

[0064]

[0065] The fatty acid composition of camellia oil is mainly composed of palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C18:1), linoleic acid (C18:2), linolenic acid (C18:3), and cis-11-eicosenoic acid (C20:1). The fatty acid composition and proportion of different enzyme-assisted wet oil extraction are basically the same. This shows that the effects of enzyme treatment and enzyme type on fatty acid composition are negligible or have no effect. The content of saturated fatty acid SFA in camellia oil is between 9.81% and 9.97%. The content of monounsaturated fatty acid MUFA in camellia oil is between 82.09% and 82.30%, of which the content of oleic acid is between 81.71% and 81.97%. In addition to reducing atherosclerosis and regulating blood lipids, oleic acid is also beneficial in promoting wound healing and fighting cancer. The content of polyunsaturated fatty acid PUFA in the wet extraction of oil by enzyme treatment is 9.81-9.97%, with no significant difference (P>0.05). In addition, polyunsaturated fatty acids in camellia oil include linoleic acid and linolenic acid. Among them, linoleic acid has the effects of lowering serum cholesterol, inhibiting arterial thrombosis, preventing atherosclerosis and osteoporosis, and linolenic acid has important physiological functions such as enhancing intelligence, increasing immunity and enhancing vision.

[0066] (2) Antioxidants such as polyphenols, squalene, total sterols, tocopherol, etc.

[0067] The contents of antioxidant substances such as polyphenols, squalene, total sterols, tocopherol, etc. in the camellia oil obtained after single enzyme-assisted wet oil extraction are listed in Table 2-3.

[0068] Table 2-3 Antioxidant content in oil under single enzyme treatment

[0069]

[0070] As can be seen from Table 2-3, the polyphenol content of wet oil extraction treated with AB Pectinase and DSM Proline Specific Endoprotease increased by 16.46% and 9.92% respectively compared with the blank group. This may be because the enzyme acts on the camellia seed cells to destroy the cell structure, causing more polyphenols in the camellia seed cells to flow out with the camellia seed oil. The polyphenol content of the other three enzymes is lower than that of the blank group. This may be because the presence of enzymes can reduce the interaction between polysaccharides and phenols. This interaction will form a complex that can embed polyphenol compounds, which is considered to be one of the main reasons for the loss of phenolic substances during the crushing and fusion of fruit paste. However, the polyphenol content of wet oil extraction is much higher than that of hot pressing.

[0071] The content of squalene under single enzyme treatment is mostly higher than that of the blank group, 4.9%-24.38% higher than the blank group. This may be due to the relatively mild reaction conditions of the aqueous enzyme method, and the loss of squalene is small, so the squalene content is relatively high. The same trend is also seen in the data of total sterols and tocopherols. The total sterol and tocopherol content measured by wet oil extraction treated with most enzymes is much greater than that of the blank group and there is a significant difference (p < 0.05). The total sterol content of wet oil extraction treated with enzymes increased by 1.72%-9.67% compared with the blank group. This may be because the complex compounds in the oil can be gently decomposed through the action of enzymes, improving the extraction efficiency of the target components. In the oil extraction process, enzyme treatment can act on compounds such as sterols in the oil in a targeted manner, making sterol compounds easier to extract. The tocopherol content of wet oil extraction under enzyme treatment increased by 7.21%-24.61% compared with the blank group. On the one hand, it is possible that the enzymes degrade the cell walls, allowing more tocopherols and polyphenolic active substances to be released into the camellia oil. On the other hand, it is possible that the tocopherols complexed with polysaccharides in the oil are degraded into the camellia oil under the action of enzymes.

[0072] 2.3 Enzyme compounding

[0073] The residual oil rate of tea oil cake obtained after complex enzyme-assisted wet oil extraction is listed in Table 2-4.

[0074] Table 2-4 Enzyme compounding

[0075]

[0076]

[0077] When a single enzyme is used for wet oil extraction, the oil extraction rate is not high. From Table 2-4, it can be concluded that the oil extraction rate of tea seeds has been significantly improved by treating with a composite enzyme of AB cellulase, pectinase and protease, reaching 79.61±0.04%. The residual oil rate of the enzyme with the best single enzyme treatment effect is 27.05% lower than that of the wet oil extraction cake that has not been treated with enzymes. The residual oil rate is 58.50% lower than that of the wet oil extraction cake that has not been treated with enzymes. This may be because the cell wall of plant cells is mainly composed of polysaccharides such as pectin and cellulose, hemicellulose, and lignin, in which some functional proteins are embedded, and oils are usually combined with other macromolecules (proteins, carbohydrates, etc.) to form complexes such as lipopolysaccharides and lipoproteins. The composite enzyme has different activities of multiple enzymes, and there is a synergistic effect between the activities, which is more conducive to the rupture of the cell wall to release oil, so the oil extraction effect is better than that of a single enzyme. It can be concluded that wet oil extraction after enzyme treatment with a combination of AB cellulase, AB pectinase and AB protease can significantly increase the oil extraction rate of camellia seed oil and reduce the residual oil rate of camellia seed cake.

[0078] The nutritional content of camellia oil obtained after complex enzyme-assisted wet oil extraction is shown in Table 2-5.

[0079] Table 2-5 Effects of enzyme complexes on the nutritional components of camellia oil

[0080]

[0081] Note: Cellulase, pectinase and protease are all from British United Enzyme Preparations, and acid protease, alkaline protease and neutral protease are from Blue Biotechnology.

[0082] As can be seen from Table 2-5, the polyphenol content in oil after oil extraction by cellulase and protease is 13.9% higher than that by cellulase and pectinase. This may be because polyphenols are mainly present in the vacuoles of plant cells, pectinase mainly acts on pectin macromolecules in plant cell walls and inner layers of cells, while protease mainly acts on lipoproteins in cells. The action of protease is more likely to affect the release of polyphenols than the action of pectinase. After oil extraction by cellulase, pectinase and protease, the content of polyphenols in camellia oil is 12.9% and 28.6% higher than that in camellia oil after cellulase, protease or pectinase are combined, respectively, indicating that pectinase and protease have a synergistic effect. This may be because pectinase acts on pectin macromolecules in plant cell walls and inner layers of cells to release them better, and protease acts on lipoproteins in cells to carry more polyphenols. Therefore, the two will play a synergistic role to increase the content of polyphenols. Comparison of polyphenol content in oils extracted by cellulase and pectinase, acidic, alkaline and neutral proteases showed that the polyphenol content in oil-tea camellia seed oil extracted by neutral protease was 111.58±5.08mg / kg, which was comparable to that of composite protease. However, the polyphenol content of acidic and alkaline proteases combined with cellulase and pectinase was 10.6% and 22.5% lower than that of the combined protease. The reason may be that neutral protease has strong adaptability and is active in the range of pH 5.5-8.5 and temperature 10℃-60℃, which is very suitable for enzymatic hydrolysis at room temperature and neutral environment. In addition, neutral protease is widely present in microorganisms, animal and plant tissues, so neutral protease is more adaptable than acidic protease and alkaline protease for oil-tea camellia seed. The combined extraction of oil by cellulase, pectinase and protease has a positive effect on retaining polyphenols in camellia oil. The effects of neutral protease and protease are equivalent. The three-enzyme complex of cellulase, pectinase, acid protease and alkaline protease has a negative effect on retaining polyphenols in camellia oil compared with the blank group.

[0083] From Table 2-5, it can be found that the content of polyphenols, squalene and tocopherol in the three-enzyme combination of British United Cellulase, Pectinase and Protease is the highest among all the combination schemes. This can also prove that the three-enzyme combination of British United Cellulase, Pectinase and Protease has the best effect in the extraction of nutrients. The content of total sterols is the best for British United Cellulase, Pectinase and Acid Protease, reaching 3291.2 mg / kg. The total sterol content of British United Cellulase, Pectinase and Alkaline Protease increased by 3.71%. Therefore, according to the data in Table 2-5, it can be concluded that the three-enzyme combination of British United Cellulase, Pectinase and Protease is the best in extracting nutrients.

[0084] A variety of oil extraction aids and dosages were selected for experiments. After experimental confirmation, using 1%-3% NaCl as an oil extraction aid in combination with a complex enzyme can not only prevent the emulsification of tea saponins in camellia oil when it comes into contact with water, but also improve the separation efficiency of camellia seed oil and medium water, thereby increasing the oil yield.

[0085] The invention adopts 17 kinds of enzyme treatment to assist wet oil extraction of camellia oil, and screens out 5 kinds of enzymes with better effects: AB pectinase, AB cellulase, AB protease, blue acid protease, and DSM proline specific endoprotease. And the best compounding scheme is obtained through compound enzyme process optimization experiment: the addition combination of AB pectinase, AB cellulase, and AB protease is 1%, 1%, and 1%, the addition amount of buffer solution is 12%, and the oil extraction auxiliary agent is 1%. The cake residual oil rate can be obtained as 9.98±0.04%, and the nutritional components of camellia oil can be well retained, and the polyphenol, squalene, total sterol and tocopherol contents in the oil reach 111.42±4.86mg / kg, 92.75±0.33mg / kg, 3182.0±31.50mg / kg and 315.77±1.56mg / kg respectively. The overall quality of camellia oil extracted by enzyme-assisted wet oil extraction meets the quality requirements of premium camellia oil among special and premium camellia oils T / LYCY 001—2020.

[0086] The above is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions that can be easily thought of by a person skilled in the art within the scope disclosed by the present invention should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention shall be based on the protection scope of the claims.

Claims

1. A method for extracting camellia oil by enzyme-assisted wet extraction, characterized in that: The method comprises the following steps: Step 1: Clean, remove impurities from fresh camellia seeds, dry them to a moisture content of 4%-6%, and then shell them to obtain camellia seed kernels; Step 2: Grind the camellia seed kernels and pass them through a 50-60 mesh sieve to obtain camellia seed kernel powder; Step 3: Add 1%-3% of NaCl, an oil extraction aid, and 2%-4% of a compound enzyme preparation to the camellia kernel powder, based on the weight of the camellia kernel powder, and add 10%-15% of a buffer solution, stir and knead at a temperature of 50° C.-60° C. for 40 min-60 min until the aggregated particles are agglomerated and difficult to stir, centrifuge and pour out the oil phase to obtain camellia seed oil; The enzyme preparation is compounded by pectinase from British enzyme preparation, cellulase from British enzyme preparation and protease from British enzyme preparation.

2. The method for enzyme-assisted wet extraction of camellia oil according to claim 1, characterized in that: In the step three, based on the weight of camellia oleifera seed powder, 1% of an oil extraction aid and 3% of a compound enzyme preparation are added, and 12% of a buffer solution is added, and the mixture is stirred and kneaded at a temperature of 55° C. for 50 minutes until the aggregated particles are clumped and difficult to stir, and the mixture is centrifuged and the oil phase is poured out to obtain camellia oleifera oil.

3. The method for enzyme-assisted wet extraction of camellia oil according to claim 1, characterized in that: The enzyme preparation is prepared by compounding pectinase of British enzyme preparation, cellulase of British enzyme preparation and protease of British enzyme preparation in a weight ratio of 1-2:1-2:1-2.

4. The method for enzyme-assisted wet extraction of camellia oil according to claim 1, characterized in that: The buffer solution is prepared from pure water and 1 mol / L NaOH solution or HCl solution.

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