CN111116353A - Method for purifying EPA in microalgae oil - Google Patents
Method for purifying EPA in microalgae oil Download PDFInfo
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- CN111116353A CN111116353A CN202010000718.6A CN202010000718A CN111116353A CN 111116353 A CN111116353 A CN 111116353A CN 202010000718 A CN202010000718 A CN 202010000718A CN 111116353 A CN111116353 A CN 111116353A
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Abstract
The invention discloses a method for purifying EPA in microalgae oil. According to the technical scheme, the purity and the recovery rate of EPA in the microalgae oil are improved by performing a two-step urea inclusion reaction on the microalgae oil.
Description
Technical Field
The invention relates to the technical field of fatty acid purification, in particular to a method for purifying EPA in microalgae oil.
Background
Eicosapentaenoic acid (EPA, C20: 5n-3) (i.e. the eicosapentaenoic acid molecular formula contains 20 carbon atoms and 5 unsaturated bonds) has important physiological significance in nutrition and medicine, and not only can promote brain development, improve brain function and promote the health of circulatory system, but also has positive effects on preventing and treating diseases such as rheumatoid arthritis, hypertension, diabetes and the like. The consumer market for EPA is also expanding with the demand of human beings. At present, people mainly obtain EPA from deep sea fish oil, but the EPA has the problems of poor stability, fishy smell of products and the like. In addition, fish oil contains a large amount of other polyunsaturated fatty acids, which leads to a complex post-treatment process and higher cost. The unicellular marine microalgae are large in storage amount and various in variety, are easier to obtain than deep sea fish oil, have simpler fatty acid composition, high EPA content and rapid cell propagation and growth, and are considered to be the most potential EPA production raw material source.
The method for purifying polyunsaturated fatty acid in the prior art is more than ten kinds at home and abroad, and comprises a molecular vacuum distillation method and a supercritical fluid CO2Extraction, chromatographic separation, urea inclusion and the like. The urea inclusion method is favored by researchers at home and abroad by the advantages of low raw material cost, mature process, simple operation, suitability for large-scale production, no high-temperature operation, and contribution to keeping the molecular structure, the physicochemical property and the like of unsaturated fatty acid. However, with the intensive research findings: in the process for purifying EPA by a urea inclusion method, the recovery rate or purity of EPA is not high; even because of the complex composition of the raw materials, EPA cannot be separated from polyunsaturated fatty acids.
Disclosure of Invention
The invention mainly aims to provide a method for purifying EPA in microalgae oil so as to improve the purity and recovery rate of EPA in the microalgae oil.
In order to achieve the above object, the present invention provides a method for purifying EPA in microalgae oil, which is characterized in that the steps of the method for purifying EPA in microalgae oil comprise:
a. weighing urea and a polar organic solvent according to a mass ratio of 1: 1-1: 20, adding the weighed urea into the polar organic solvent, and uniformly mixing to obtain a uniformly mixed solution;
b. b, adding microalgae oil into the uniformly mixed solution obtained in the step a, uniformly mixing, cooling and crystallizing to obtain a crystal mixed solution; wherein the mass ratio of the total mass of saturated fatty acids and monounsaturated fatty acids in the microalgae oil to the mass of the urea in the step a is 1: 1-1: 10;
c. separating the crystal mixed liquid obtained in the step b to obtain a filtrate;
d. evaporating the filtrate obtained in the step c to obtain an oil phase substance; washing and drying to obtain a first part of product.
Optionally, separating the crystal mixed liquid obtained in the step b to obtain crystals;
adding the polar organic solvent into the crystal, stirring and washing to obtain a washing solution; separating the washing liquid to obtain secondary filtrate; evaporating the secondary filtrate to obtain a secondary oil phase; and (5) washing and drying. Obtaining a second part of product.
Optionally, the crystallization temperature in the step b is-20 ℃ to 20 ℃.
Optionally, the mass ratio of the total mass of saturated fatty acids and monounsaturated fatty acids in the microalgae oil in the step b to the mass of the urea in the step a is 1: 2-1: 8.
Optionally, the polar organic solvent is one or a combination of methanol, ethanol, propylene glycol and glycerol.
Optionally, the polar organic solvent is absolute ethanol.
Alternatively, the stirring speed for stirring the crystal is 100rpm or more and 600rpm or less.
Alternatively, the washing time for washing the crystals is greater than 0 hour and 2 hours or less.
Compared with the prior art, the invention has the following beneficial effects:
the invention provides a method for purifying EPA in microalgae oil, which comprises the following steps: a. weighing urea and a polar organic solvent according to a mass ratio of 1: 1-1: 20, adding the weighed urea into the polar organic solvent, and uniformly mixing to obtain a uniformly mixed solution; b. b, adding microalgae oil into the uniformly mixed solution obtained in the step a, uniformly mixing, cooling and crystallizing to obtain a crystal mixed solution; wherein the mass ratio of the total mass of saturated fatty acids and monounsaturated fatty acids in the microalgae oil to the mass of the urea in the step a is 1: 1-1: 10; c. separating the crystal mixed liquid obtained in the step b to obtain a filtrate; d. evaporating the filtrate obtained in the step c to obtain an oil phase substance; washing and drying to obtain a first part of product. According to the technical scheme, the purity and the recovery rate of EPA in the microalgae oil are improved by performing a two-step urea inclusion reaction on the microalgae oil. And the obtained first part of purified product and the second part of product are separated and purified again, and can be directly used for producing medicines, health-care foods and the like.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The invention provides a method for purifying EPA in microalgae oil, which is suitable for efficiently extracting EPA in the microalgae oil.
A method of purifying EPA in microalgal oil, the steps of the method of purifying EPA in microalgal oil comprising:
a. weighing urea and a polar organic solvent according to a mass ratio of 1: 1-1: 20, adding the weighed urea into the polar organic solvent, and uniformly mixing to obtain a uniformly mixed solution;
b. b, adding microalgae oil into the uniformly mixed solution obtained in the step a, wherein the mass ratio of the total mass of saturated fatty acids and monounsaturated fatty acids in the microalgae oil to the mass of urea in the step a is 1: 1-1: 10, uniformly mixing, cooling and crystallizing to obtain a crystal mixed solution;
c. separating the crystal mixed liquid obtained in the step b to obtain a filtrate;
d. evaporating the filtrate obtained in the step c to obtain an oil phase substance; washing and drying to obtain a first part of product.
Optionally, the step of separating the crystal mixture obtained in step b to obtain a filtrate further includes:
separating the crystal mixed liquid obtained in the step b to obtain crystals;
adding the polar organic solvent into the crystal, stirring and washing to obtain a washing solution; separating the washing liquid to obtain secondary filtrate; evaporating the secondary filtrate to obtain a secondary oil phase; washing and drying to obtain a second part of product.
In this example, the purity and recovery rate of EPA in microalgal oil was improved by performing a "two-step" urea inclusion reaction on the microalgal oil. The two-step urea inclusion reaction comprises the following specific steps: in the first step, the filtrate after the urea inclusion reaction is purified to obtain a first part of products; in the second step, the second part of the product is obtained by washing and purifying the crystals after the urea inclusion reaction. And the obtained first part of purified product and the second part of product are separated and purified again, and can be directly used for producing medicines, health-care foods and the like. Specifically, according to the method for purifying EPA in microalgae oil provided by the embodiment of the invention, urea and a polar organic solvent are mixed, then microalgae oil is added and mixed uniformly to perform a urea inclusion reaction, and then the reacted liquid is naturally cooled to room temperature and then is kept stand at a low temperature for several hours to crystallize the reacted liquid. It should be further explained that the microalgae oil selected in the embodiment of the present invention is ethyl ester type microalgae oil. Specifically, the ethyl ester type microalgae oil is obtained by performing a series of operations such as drying, extraction, transesterification, molecular distillation and the like on microalgae, and related experiments can refer to the prior art and are not repeated herein. Optionally, the low-temperature standing time is 24-48 hours, so that the urea inclusion reaction can be completely carried out. It is further explained that saturated fatty acid, monounsaturated fatty acid and polyunsaturated fatty acid such as EPA are preferentially included in the precipitated urea crystal according to the molecular structure. In this embodiment, most of saturated fatty acids and monounsaturated fatty acids in the microalgae oil are removed through the urea inclusion reaction of the "two-step method", so as to improve the purity of EPA in the microalgae oil. And the EPA contained in the filtrate and the crystal obtained after the reaction are respectively purified, so that the recovery rate of the EPA of the whole microalgae oil is improved.
Specifically, in the step of washing and drying the oil-phase substance to obtain the purified product, the oil-phase substance is washed by a salt solution firstly during washing, so that the whole washing solution is an electrolyte, an emulsion formed by mixing water and the oil-phase substance can be better electrically separated, and impurities contained in the oil-phase substance can be more easily washed and removed; and then washing with water to further remove urea and salt solution in the oil phase, wherein the washing process is at least twice. Meanwhile, when the washed oil phase substance is dried, a drying agent is needed to be used for drying, and optionally, anhydrous sodium sulfate is selected for drying.
Optionally, the crystallization temperature in the step b is-20 ℃ to 20 ℃.
In this example, the crystallization temperature was set to-20 ℃ or higher in order to sufficiently react and crystallize the saturated fatty acids and monounsaturated fatty acids in the raw material with urea and increase the recovery rate of the first partial product. Meanwhile, in order to ensure that crystals in the mixed solution can be fully crystallized and the whole urea inclusion reaction can be completely reacted, the crystallization temperature is set to be less than or equal to 20 ℃ so as to improve the purity of the first part of products. It will be appreciated that at temperatures around 60 ℃ the urea inclusion reaction does not proceed sufficiently due to the increased evaporation of ethanol, resulting in a substantial reduction in the purity of the first fraction. It should be understood that in other embodiments, the crystallization temperature may also range between-30 ℃ and-20 ℃, or the crystallization temperature may range between 20 ℃ and 30 ℃.
Optionally, the mass ratio of the total mass of saturated fatty acids and monounsaturated fatty acids in the microalgae oil in the step b to the mass of the urea in the step a is 1: 2-1: 8.
In this example, to remove more saturated and monounsaturated fatty acids from the raw microalgal oil, the ratio of the total mass of urea to the total mass of saturated and monounsaturated fatty acids in the microalgal oil was set to 1:1 to increase the purity in the first portion of product. To reduce the reaction with unsaturated fatty acids (including EPA) due to excessive urea content, the ratio of the total mass of urea to the total mass of saturated and monounsaturated fatty acids in the microalgal oil was set to 8:1 to increase the recovery of the first fractionated product.
Optionally, the polar organic solvent in step a is one or a combination of methanol, ethanol, propylene glycol and glycerol. In this embodiment, in order to reduce the influence of the polar organic solvent on the urea inclusion reaction, the polar organic solvent is anhydrous ethanol.
Optionally, the step a of mixing uniformly specifically comprises: heating and stirring at the temperature of 40-80 ℃ and uniformly mixing. Namely, in this embodiment, urea and a polar organic solvent are weighed according to a mass ratio of 1: 1-1: 20, and the weighed urea is added into the polar organic solvent, and is heated, stirred and mixed uniformly at a temperature of 40-80 ℃ to obtain a mixed solution.
Alternatively, the stirring speed for stirring the crystal is 100rpm or more and 600rpm or less.
In this example, the crystal stirring speed in step e was set to 100rpm or more in order to sufficiently wash the crystals, thereby increasing the recovery rate of the second fraction product. In order to reduce the dissolution of monounsaturated fatty acids and saturated fatty acids crystallized from the crystals in the washing liquid, the crystal stirring rotation speed is set to be less than or equal to 600rpm to reduce impurities in the second product, improve the purity of the second product and save energy. It should be understood that in other embodiments, the stirring speed may also be in the range of 0 to 100rpm, or in the range of 600 to 2000 rpm.
Alternatively, the washing time for washing the crystals is greater than 0 hour and 2 hours or less.
In this embodiment, in order to reduce the dissolution of the crystals caused by over-washing the crystals, the time for washing the crystals is limited to less than 2 hours, so as to reduce the impurities in the second part of the product and improve the purity of the second part of the product.
Optionally, in an embodiment, the washing time for washing the crystal is selected to be any time node of 3min or more and 30min or less for washing the crystal.
To better illustrate the purification and recovery efficiency of the method for purifying EPA from microalgae oil of the present invention, the following examples of specific components were selected as references in the present invention.
Group one of the embodiments:
the experimental procedure of group one of this example was: weighing urea and absolute ethyl alcohol (absolute ethyl alcohol is selected as the polar organic solvent in the first group of the embodiment, and absolute ethyl alcohol is selected as the following experimental groups) according to the mass ratio of 1:4, adding the urea and the absolute ethyl alcohol into a container, converging and stirring the mixture at 60 ℃ for 1 hour, slowly adding microalgae oil according to the proportion shown in the table 1, keeping stirring the mixture at 480r/min for 10 to 40 minutes, taking out the container, naturally cooling the mixture to room temperature, and standing the mixture at the low temperature of minus 20 ℃ for 24 to 48 hours to obtain a crystal mixed solution (namely the urea inclusion product in the embodiment). Rapidly filtering the crystal mixed solution by using a sand core funnel, and filtering under the vacuum pressure of 0.1MPa to obtain crystals and filtrate; placing the filtrate on a rotary evaporator for evaporation to remove the absolute ethyl alcohol, wherein the pressure of the rotary evaporator is 0.1MPa, and the evaporation temperature is 78 ℃, so as to obtain an oil phase substance; then washing the oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the oil phase substance with water twice until the oil phase substance is neutral; and drying the washed oil phase substance by using sodium sulfate to obtain a first part of product. Washing the crystal obtained by suction filtration for the second time, adding absolute ethyl alcohol into the crystal according to the mass ratio of 1:1 (it should be understood that the mass ratio of the absolute ethyl alcohol used for washing to the crystal is not limited to 1:1, and the washing ratio is 1: 1-20: 1), stirring for 5min at the rotating speed of 180r/min, and then carrying out suction filtration rapidly by using a sand core funnel to obtain a secondary filtrate; putting the secondary filtrate into a rotary evaporator for evaporation to remove absolute ethyl alcohol, so as to obtain a secondary oil phase substance; then washing the secondary oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the secondary oil phase substance with water twice until the secondary oil phase substance is neutral; and drying the washed secondary oil phase substance by using sodium sulfate to obtain a second part of product. Specific results are shown in table 2.
The ratio of the total mass of saturated and monounsaturated fatty acids to the mass of urea in the microalgal oil is set forth in table 1 below:
TABLE 1
| Group of embodiments | Saturated fatty acids and monounsaturated fatty acid ethyl esters: urea |
| Example 1 | 1:1 |
| Example 2 | 1:3 |
| Example 3 | 1:6 |
| Example 4 | 1:8 |
| Example 5 | 1:10 |
It should be noted that the EPA mass content, EPA to total fatty acid mass ratio and EPA recovery rate in this example group were calculated as follows: (this is true for the calculation of the following experimental groups and is not described in too much detail.)
The calculation formula of EPA mass content is as follows (the EPA mass content in the sample is quantified by the peak area of chromatographic peak):
wherein the content of the first and second substances,
X2-the content of EPA in the sample is in grams per hundred grams (g/100 g);
A2-the peak area of the EPA ethyl ester in the sample determination liquid;
ms2-the mass of EPA ethyl ester standard contained in the preparation of the standard assay solution in milligrams (mg);
the conversion coefficient of FEE-FA-EPA ethyl ester converted into fatty acid is 0.9151;
As2-peak area of EPA ethyl ester in standard assay;
m-the weighed mass of the sample in milligrams (mg);
100-conversion of the content to a factor of the content per 100g of sample.
The calculation formula of the EPA (eicosapentaenoic acid) to total fatty acid mass ratio is as follows:
wherein the content of the first and second substances,
X1-EPA on total fatty acid mass ratio;
X2-the content of EPA in the sample is in grams per hundred grams (g/100 g);
∑XTFA-sum of all fatty acid contents.
The EPA recovery is calculated as follows:
X3-EPA recovery;
X2-the content of EPA in the sample is in grams per hundred grams (g/100 g);
W2-the product quality;
Xfirst stage-the reactant EPA mass content;
Wfirst stage-mass of reaction.
The data of the results of example set one are shown in table 2 below: (% represents a mass ratio)
TABLE 2
As shown in the experimental data of the above example group one, it can be seen that: under the condition that the mass ratio of urea to absolute ethyl alcohol, the crystallization temperature, the crystal washing stirring speed and the crystal washing time are fixed, the higher the mass ratio of urea to saturated fatty acid, monounsaturated fatty acid ethyl ester and urea is, the higher the EPA content in the obtained product is, and the higher the EPA content in the total fatty acid is; but the recovery of EPA in the first portion of the product decreases with increasing urea mass fraction and the recovery of EPA in the second portion of the product increases with increasing urea mass fraction. Wherein when the mass ratio of the saturated fatty acid, the monounsaturated fatty acid ethyl ester and the urea in the microalgae oil is 1:10, the optimal EPA purity and the higher EPA recovery rate are obtained. The EPA mass content in the first part of products reaches 61.51g/100g, the EPA mass ratio in the total fatty acid of the microalgae oil reaches 81.69%, and the recovery rate reaches 71.1%; in the second part of products, the weight content of EPA reaches 45.08g/100g, the weight percentage of EPA in the total fatty acid of the microalgae oil reaches 67.05%, and the recovery rate reaches 25.75%; the overall recovery of the EPA in the overall product reached 96.85%.
Example set two:
the experimental procedure for group two of this example was: weighing absolute ethyl alcohol and urea according to the mass ratio in the table 3, adding the absolute ethyl alcohol and the urea into a container, mixing and stirring the absolute ethyl alcohol and the urea at the temperature of 60 ℃ for 1 hour, then weighing the absolute ethyl alcohol and the urea according to the mass ratio of saturated fatty acid to monounsaturated fatty acid ethyl ester in the urea and the microalgae oil of 3.5:1, slowly adding the microalgae oil, keeping stirring at the rotating speed of 480r/min for 10-40 minutes, taking out the container, naturally cooling to the room temperature, and standing at the low temperature of 5 ℃ for 24-48 hours to obtain a crystal mixed liquid (namely, a urea inclusion product in the embodiment). Rapidly filtering the crystal mixed solution by using a sand core funnel, and filtering under the vacuum pressure of 0.1MPa to obtain crystals and filtrate; placing the filtrate on a rotary evaporator for evaporation to remove the absolute ethyl alcohol, wherein the pressure of the rotary evaporator is 0.1MPa, and the evaporation temperature is 78 ℃, so as to obtain an oil phase substance; then washing the oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the oil phase substance with water twice until the oil phase substance is neutral; and drying the washed oil phase substance by using sodium sulfate to obtain a first part of product. Washing the crystal obtained by suction filtration for the second time, adding absolute ethyl alcohol into the crystal for washing according to the mass ratio of 1:1 (it should be understood that the mass ratio of the absolute ethyl alcohol used for washing to the crystal is not limited to 1:1, and the washing ratio is 1.1: 1-20: 1), stirring for 5min at the rotating speed of 180r/min, and then carrying out suction filtration by a sand core funnel quickly to obtain a secondary filtrate; putting the secondary filtrate into a rotary evaporator for evaporation to remove absolute ethyl alcohol, so as to obtain a secondary oil phase substance; then washing the secondary oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the secondary oil phase substance with water twice until the secondary oil phase substance is neutral; and drying the washed secondary oil phase substance by using sodium sulfate to obtain a second part of product. Specific results are shown in table 4.
The mass ratios of the absolute ethanol and the urea are as follows in table 3:
TABLE 3
The data of the results for example set two are shown in table 4 below: (% represents a mass ratio)
TABLE 4
As shown in the experimental data of the second embodiment, it can be seen that: under the condition that the mass ratio of urea to saturated fatty acid and monounsaturated fatty acid ethyl ester, the crystallization temperature, the crystal washing stirring speed and the crystal washing time are fixed, the lower the mass ratio of urea to absolute ethyl alcohol is, the higher the EPA mass content in the obtained product is, the higher the EPA mass ratio in the total fatty acid is, but the EPA recovery rate is reduced along with the reduction of the urea mass ratio; after a certain optimal mass ratio of absolute ethyl alcohol to urea, as the mass ratio of urea is further reduced, the EPA content in the obtained product is lower and lower, the EPA content in the total fatty acid is lower and lower, and the EPA recovery rate is improved. Wherein, when the mass ratio of the absolute ethyl alcohol to the urea is 4:1, the optimal EPA purity and the higher EPA recovery rate are obtained. The EPA content in the first part of products reaches 50.26g/100g, the EPA content in the total fatty acids of the microalgae oil reaches 70.44%, and the recovery rate reaches 76.00%; in the second part of products, the weight content of EPA reaches 31.25g/100g, the weight percentage of EPA in the total fatty acid of the microalgae oil reaches 45.34%, and the recovery rate reaches 22.19%; the overall recovery of the EPA in the overall product reached 98.19%.
Example group three:
the experimental procedure for group three of this example was: weighing absolute ethyl alcohol and urea according to a mass ratio of 4:1, adding the absolute ethyl alcohol and the urea into a container, mixing and stirring the mixture at 60 ℃ for 1 hour, weighing microalgae oil according to a mass ratio of 4:1 of saturated fatty acid and monounsaturated fatty acid ethyl ester in the urea to the microalgae oil, slowly adding the microalgae oil into the mixed liquid, stirring the mixture at 480r/min for 10-40 minutes, taking out the container, naturally cooling the container to room temperature, and standing the mixture at a crystallization temperature in the table 5 for 24-48 hours to obtain a crystal mixed liquid (namely a urea inclusion product in the embodiment). Rapidly filtering the crystal mixed solution by using a sand core funnel, and filtering under the vacuum pressure of 0.1MPa to obtain crystals and filtrate; placing the filtrate on a rotary evaporator for evaporation to remove the absolute ethyl alcohol, wherein the pressure of the rotary evaporator is 0.1MPa, and the evaporation temperature is 78 ℃, so as to obtain an oil phase substance; then washing the oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the oil phase substance with water twice until the oil phase substance is neutral; and drying the washed oil phase substance by using sodium sulfate to obtain a first part of product. Washing the crystal obtained by suction filtration for the second time, adding absolute ethyl alcohol into the crystal for washing according to the mass ratio of 1:1 (it should be understood that the mass ratio of the absolute ethyl alcohol used for washing to the crystal is not limited to 1:1, and the washing ratio is 1.1: 1-20: 1), stirring for 5min at the rotating speed of 180r/min, and then carrying out suction filtration by a sand core funnel quickly to obtain a secondary filtrate; putting the secondary filtrate into a rotary evaporator for evaporation to remove absolute ethyl alcohol, so as to obtain a secondary oil phase substance; then washing the secondary oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the secondary oil phase substance with water twice until the secondary oil phase substance is neutral; and drying the washed secondary oil phase substance by using sodium sulfate to obtain a second part of product. Specific results are shown in table 6.
The crystallization temperatures of the urea inclusion reactions are shown in table 5 below:
TABLE 5
The data of the results for example group three are shown in table 6 below: (% represents a mass ratio)
TABLE 6
As shown in the experimental data of the third embodiment, it can be seen that: under the condition that the mass ratio of urea, absolute ethyl alcohol, saturated fatty acid and monounsaturated fatty acid ethyl ester, the crystal washing stirring speed and the crystal washing time are fixed, the lower the EPA mass content in the obtained product is along with the increase of the crystallization temperature, the lower the EPA mass ratio of the EPA in the total fatty acid is, and the EPA recovery rate is increased along with the increase of the crystallization temperature. Wherein the optimum EPA purity and high EPA recovery are obtained at a crystallization temperature of-20 ℃. The EPA content in the first part of products reaches 55.99g/100g, the EPA content in the total fatty acids of the microalgae oil reaches 80.68%, and the recovery rate reaches 69.38%; in the second part of products, the weight content of EPA reaches 38.43g/100g, the weight percentage of EPA in the total fatty acid of the microalgae oil reaches 47.70%, and the recovery rate reaches 27.32%; the overall recovery of the EPA in the overall product reached 96.70%.
Example group four:
the experimental procedure for group four of this example was: weighing absolute ethyl alcohol and urea according to a mass ratio of 4:1, adding the absolute ethyl alcohol and the urea into a container, mixing and stirring the mixture for 1 hour at 60 ℃, weighing microalgae oil according to a mass ratio of 4:1 of saturated fatty acid and monounsaturated fatty acid ethyl ester in the urea and the microalgae oil, slowly adding the microalgae oil into the mixed liquid, keeping stirring for 10-40 minutes at a rotating speed of 480r/min, taking out the container, naturally cooling to room temperature, and standing at a low temperature of-20 ℃ for 24-48 hours to obtain a crystal mixed liquid (namely a urea inclusion product in the embodiment). Rapidly filtering the crystal mixed solution by using a sand core funnel, and filtering under the vacuum pressure of 0.1MPa to obtain crystals and filtrate; placing the filtrate on a rotary evaporator for evaporation to remove the absolute ethyl alcohol, wherein the pressure of the rotary evaporator is 0.1MPa, and the evaporation temperature is 78 ℃, so as to obtain an oil phase substance; then washing the oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the oil phase substance with water twice until the oil phase substance is neutral; and drying the washed oil phase substance by using sodium sulfate to obtain a first part of product. Washing the crystals obtained by suction filtration for the second time, adding absolute ethyl alcohol into the crystals for washing according to the mass ratio of 1:1 (it should be understood that the mass ratio of the absolute ethyl alcohol used for washing to the crystals is not limited to 1:1, and the mass ratio of the absolute ethyl alcohol to the crystals is 1.1: 1-20: 1, and the washing ratio is 1.1: 1-20: 1), stirring for 5min at the rotating speed shown in the table 7, and then carrying out suction filtration rapidly by using a sand core funnel to obtain a secondary filtrate; putting the secondary filtrate into a rotary evaporator for evaporation to remove absolute ethyl alcohol, so as to obtain a secondary oil phase substance; then washing the secondary oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the secondary oil phase substance with water twice until the secondary oil phase substance is neutral; and drying the washed secondary oil phase substance by using sodium sulfate to obtain a second part of product. Specific results are shown in table 8.
The rotation speed of the stirring during the washing of the crystals is shown in the following table 7:
TABLE 7
The data of the results for example group four are shown in table 8 below: (% represents a mass ratio)
TABLE 8
As shown in the experimental data of the above example group four: under the condition that the mass ratio of urea, absolute ethyl alcohol, saturated fatty acid and monounsaturated fatty acid ethyl ester, the crystallization temperature and the crystal washing time are fixed, when the washing process is not performed on the crystal, the EPA recovery rate of the second part of products is extremely low. After the stirring is started, the EPA content by mass in the second part of products obtained by the method is lower as the crystal washing stirring speed is increased, the EPA content by mass in the total fatty acid is lower, but the EPA recovery rate is increased as the crystallization temperature is increased. Wherein, when the crystal washing and stirring speed is 180rpm, the optimal EPA purity and higher EPA recovery rate are obtained. The EPA content in the first part of products reaches 46.14g/100g, the EPA content in the total fatty acids of the microalgae oil reaches 64.81%, and the recovery rate reaches 68.56%; in the second part of products, the weight content of EPA reaches 28.19g/100g, the weight percentage of EPA in the total fatty acid of the microalgae oil reaches 35.83%, and the recovery rate reaches 25.45%; the overall recovery of the EPA in the overall product reached 94.01%.
Example group five:
the experimental procedure for group five of this example was: weighing absolute ethyl alcohol and urea according to a mass ratio of 4:1, adding the absolute ethyl alcohol and the urea into a container, mixing and stirring the mixture for 1 hour at 60 ℃, weighing microalgae oil according to a mass ratio of 4:1 of saturated fatty acid and monounsaturated fatty acid ethyl ester in the urea and the microalgae oil, slowly adding the microalgae oil into the mixed liquid, keeping stirring for 10-40 minutes at a rotating speed of 480r/min, taking out the container, naturally cooling to room temperature, and standing at a low temperature of-20 ℃ for 24-48 hours to obtain a crystal mixed liquid (namely a urea inclusion product in the embodiment). Rapidly filtering the crystal mixed solution by using a sand core funnel, and filtering under the vacuum pressure of 0.1MPa to obtain crystals and filtrate; placing the filtrate on a rotary evaporator for evaporation to remove the absolute ethyl alcohol, wherein the pressure of the rotary evaporator is 0.1MPa, and the evaporation temperature is 78 ℃, so as to obtain an oil phase substance; then washing the oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the oil phase substance with water twice until the oil phase substance is neutral; and drying the washed oil phase substance by using sodium sulfate to obtain a first part of product. Washing the crystals obtained by suction filtration for the second time, adding absolute ethyl alcohol into the crystals for washing according to the mass ratio of 1:1 (it should be understood that the mass ratio of absolute ethyl alcohol used for washing to the crystals is not limited to 1:1, and the mass ratio of absolute ethyl alcohol to the crystals is 1.1: 1-20: 1, and the washing ratio is 1.1: 1-20: 1), stirring at the rotating speed of 180rpm for the time length shown in table 9, and then carrying out suction filtration rapidly by using a sand core funnel to obtain secondary filtrate; putting the secondary filtrate into a rotary evaporator for evaporation to remove absolute ethyl alcohol, so as to obtain a secondary oil phase substance; then washing the secondary oil phase substance with 10% sodium chloride solution, separating the organic phase, and then continuing to wash the secondary oil phase substance with water twice until the secondary oil phase substance is neutral; and drying the washed secondary oil phase substance by using sodium sulfate to obtain a second part of product. Specific results are shown in table 10.
The time for washing the crystals during the washing of the crystals is shown in the following table 9:
TABLE 9
The data for the results of example group five are shown in table 10 below: (% represents a mass ratio)
Watch 10
As shown in the experimental data of the above example group five, it can be seen that: under the condition that the mass ratio of urea, absolute ethyl alcohol, saturated fatty acid and monounsaturated fatty acid ethyl ester, the crystallization temperature and the stirring speed for crystal washing are constant, the EPA recovery rate of the second part of products is extremely low when the washing process is not stirred. After the stirring is started, the EPA mass content in the second part product is lower and lower as the stirring time of crystal washing is prolonged, the EPA mass ratio in the total fatty acid is lower, but the EPA recovery rate is increased as the crystal washing time is prolonged. Wherein, when the crystal washing and stirring time is 3min, the optimal EPA purity and higher EPA recovery rate are obtained. The EPA content in the first part of products reaches 51.69g/100g, the EPA content in the total fatty acids of the microalgae oil reaches 72.03%, and the recovery rate reaches 66.15%; in the second part of products, the weight content of EPA reaches 29.36g/100g, the weight percentage of EPA in the total fatty acid of the microalgae oil reaches 40.33%, and the recovery rate reaches 28.10%; the overall recovery of the EPA in the overall product reached 94.25%.
In conclusion, the method for purifying EPA in microalgae oil adopted by the invention greatly improves the EPA mass content in the product and the EPA proportion in total fatty acid, ensures the EPA recovery rate, avoids the waste of raw materials and saves the cost. In addition, the method for purifying EPA adopted in the embodiment group is simple and easy to operate, and is beneficial to industrial production.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not intended to limit the scope of the present invention; all equivalent changes made within the scope of the claims of the present invention are covered by the claims of the present invention.
Claims (8)
1. A method for purifying EPA in microalgae oil, wherein the method for purifying EPA in microalgae oil comprises the following steps:
a. weighing urea and a polar organic solvent according to a mass ratio of 1: 1-1: 20, adding the weighed urea into the polar organic solvent, and uniformly mixing to obtain a uniformly mixed solution;
b. b, adding microalgae oil into the uniformly mixed solution obtained in the step a, uniformly mixing, cooling and crystallizing to obtain a crystal mixed solution; wherein the mass ratio of the total mass of saturated fatty acids and monounsaturated fatty acids in the microalgae oil to the mass of the urea in the step a is 1: 1-1: 10;
c. separating the crystal mixed liquid obtained in the step b to obtain a filtrate;
d. evaporating the filtrate obtained in the step c to obtain an oil phase substance; washing and drying to obtain a first part of product.
2. The method for purifying EPA in microalgae oil of claim 1, wherein the step of separating the crystal mixture obtained in step b to obtain the filtrate further comprises:
separating the crystal mixed liquid obtained in the step b to obtain crystals;
adding the polar organic solvent into the crystal, stirring and washing to obtain a washing solution; separating the washing liquid to obtain secondary filtrate; evaporating the secondary filtrate to obtain a secondary oil phase; washing and drying to obtain a second part of product.
3. The method for purifying EPA in microalgae oil as claimed in claim 2, wherein the crystallization temperature in step b is-20 deg.C.
4. The method for purifying EPA in a microalgae oil of claim 3 wherein the ratio of the total mass of saturated and monounsaturated fatty acids in the microalgae oil in step b to the mass of the urea in step a is 1:2 to 1: 8.
5. The method for purifying EPA in microalgae oil of claim 2 wherein the polar organic solvent is one or a combination of methanol, ethanol, propylene glycol and glycerol.
6. The method for purifying EPA in microalgae oil of claim 5 wherein the polar organic solvent is absolute ethanol.
7. The method for purifying EPA in microalgae oil as claimed in claim 2, wherein the stirring speed for stirring said crystal is greater than or equal to 100rpm and less than or equal to 600 rpm.
8. The method for purifying EPA from microalgae oil as in claim 7, wherein the washing time for washing said crystal is greater than 0 hour and less than or equal to 2 hours.
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4792418A (en) * | 1985-08-14 | 1988-12-20 | Century Laboratories, Inc. | Method of extraction and purification of polyunsaturated fatty acids from natural sources |
| US5679809A (en) * | 1994-05-09 | 1997-10-21 | Nestec S.A. | Concentrate of polyunsaturated fatty acid ethyl esters and preparation thereof |
| CN103396310A (en) * | 2013-07-25 | 2013-11-20 | 浙江大学 | Method for separating and purifying eicosapentaenoic acid ester and docosahexenoic acid ester from micro-algal oil or fish oil |
| CN104529739A (en) * | 2014-12-25 | 2015-04-22 | 北京阳光基业药业有限公司 | Purification method of unsaturated fatty acid |
| CN105779140A (en) * | 2014-12-23 | 2016-07-20 | 浙江医药股份有限公司新昌制药厂 | Preparation method of ethyl ester type fish oil with high EPA content |
| CN106117050A (en) * | 2016-06-24 | 2016-11-16 | 四川欣美加生物医药有限公司 | Improve the method for the purity of EPA-E in fish oil |
-
2020
- 2020-01-02 CN CN202010000718.6A patent/CN111116353A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4792418A (en) * | 1985-08-14 | 1988-12-20 | Century Laboratories, Inc. | Method of extraction and purification of polyunsaturated fatty acids from natural sources |
| US5679809A (en) * | 1994-05-09 | 1997-10-21 | Nestec S.A. | Concentrate of polyunsaturated fatty acid ethyl esters and preparation thereof |
| CN103396310A (en) * | 2013-07-25 | 2013-11-20 | 浙江大学 | Method for separating and purifying eicosapentaenoic acid ester and docosahexenoic acid ester from micro-algal oil or fish oil |
| CN105779140A (en) * | 2014-12-23 | 2016-07-20 | 浙江医药股份有限公司新昌制药厂 | Preparation method of ethyl ester type fish oil with high EPA content |
| CN104529739A (en) * | 2014-12-25 | 2015-04-22 | 北京阳光基业药业有限公司 | Purification method of unsaturated fatty acid |
| CN106117050A (en) * | 2016-06-24 | 2016-11-16 | 四川欣美加生物医药有限公司 | Improve the method for the purity of EPA-E in fish oil |
Non-Patent Citations (1)
| Title |
|---|
| 王玉仙: ""眼点微拟球藻二十碳五烯酸高效积累及其提取纯化研究"", 《中国优秀博硕士学位论文全文数据库（硕士）基础科学辑》 * |
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