Method for removing zearalenone toxin in natural vitamin E oil

Abstract

The invention discloses a method for removing zearalenone toxin in natural vitamin E oil, and belongs to the technical field of purification. The method comprises the following steps: carrying out chromatography treatment by adopting a silica gel column, sequentially eluting for three times, and using an eluent in each time which is 0.8-1.5 times of the volume of the column; the first eluent is a mobile phase A, the second eluent is a mobile phase A and a mobile phase B, the volume ratio of the mobile phase A to the mobile phase B is 8: 3-10: 1, and the third eluent is a mobile phase C; the mobile phase A is selected from normal hexane, petroleum ether or cyclohexane, the mobile phase B is the same as or different from the mobile phase C, the mobile phase B is selected from ethyl acetate, acetone, isobutyl acetate or ethanol, and the mobile phase C is selected from ethyl acetate, acetone, isobutyl acetate or ethanol; and collecting the components eluted for the second time and the third time, and concentrating under reduced pressure to remove the eluent to obtain the product. The content of Zen is removed, and natural vitamin E with the purity of 50% can be refined to 90% or above.

Description

Technical Field

[0001] The invention relates to the technical field of purification, and in particular to a method for removing zearalenone toxin from natural vitamin E oil. Background Art

[0002] Zearalenone (Zen) is a fungal toxin isolated from moldy corn. It is a secondary metabolite produced by various Fusarium fungi, and its skeleton structure is dihydroxybenzoic acid lactone. The structure is as follows: .

[0003] Studies have found that Zen is commonly found in moldy corn, soybeans, rice and other grains and their products. It is one of the mycotoxins that seriously endangers human and animal health. Its toxicity is mainly manifested in: 1. Damage to the blood system, immune system, reproductive system and liver and kidney organs of mammals. 2. Promote cancer in animal liver, ovary and other tissues. 3. Large intake can cause acute poisoning and even death in animals. my country stipulates that the Zen content in corn feed raw materials shall not exceed 500μg / kg.

[0004] Natural vitamin E oil, also known as tocopherol, is extracted from byproducts of the refining process of soybeans, corn, rapeseed and other oils. It is an essential vitamin for the human body and is widely used as a nutritional enhancer and antioxidant in the fields of food, medicine, cosmetics and feed. The structural formula of tocopherol is as follows: .

[0005] Soybeans, corn, and rapeseed, which are the raw materials for natural vitamin E oil, are prone to mildew during transportation and storage, which can lead to excessive levels of zearalenone toxin in these raw materials. Zen toxin is stable and difficult to destroy, and most of it will be transferred to soybean oil, corn oil, rapeseed oil and their subsequent by-product, natural vitamin E oil. Therefore, the EU is currently paying strict attention to the zearalenone toxin in natural vitamin E oil, and the requirement is that it should not exceed 200μg / kg.

[0006] At present, there are few patents and papers on the removal of Zen toxins from oils and fats. Chinese patent CN117305281A discloses a method of using a composite enzyme preparation to detoxify grain and feed. This patent mainly targets solid feed, and uses a composite enzyme preparation to fully mix with the feed, and then removes the clear liquid by centrifugation to obtain the detoxified grain and oil product, which is then dried. This method obviously has great limitations and requires large-scale mixing and separation equipment as well as a large amount of wastewater and waste liquid, which is costly and difficult to use on a large scale.

[0007] Chinese patent CN115873658A discloses a process for removing zearalenone toxin from corn germ in vegetable oil, comprising the following steps: cleaning and removing impurities from corn germ, and then crushing it with a crusher; adding activated carbon to the crushed corn germ, and then detoxifying it under the conditions of 150-200°C and a steam pressure of 0.8-2.0MPa; softening the detoxified corn germ; rolling the softened corn germ to obtain oil germ flakes; pressing the oil germ flakes obtained by rolling to obtain pressed crude oil. The present invention uses a physical method to remove zearalenone toxin from the raw material, significantly reducing the content of zearalenone in the vegetable oil, and does not destroy the structure of triglycerides, does not introduce exogenous impurities, retains inherent nutrients, and the vitamin E in the crude oil is ≥1300mg / kg, and the phytosterol is ≥12500mg / kg. The present invention mainly utilizes high-pressure steam to boil the oil to destroy Zen toxins. Many studies have shown that the Zen toxins with a boiling point of up to 600°C have a relatively stable structure, and the boiling destruction method cannot ensure that it can stably achieve the effect.

[0008] Chinese patent CN106281657A discloses a method for removing zearalenone toxins from corn germs. The method for removing zearalenone toxins from corn germs of the present invention is as follows: the corn germs are metered, cleaned, crushed, detoxified by high-temperature and high-pressure steam, softened, rolled, and pressed to obtain corn pressed crude oil. Similar to the previous patent, high-temperature and high-pressure steam is also used for detoxification, and its effect is mainly for oil materials such as corn. If it is natural vitamin E oil that already contains Zen toxins, the method of high-temperature and high-pressure steam will be difficult to implement. The reasons are as follows: First, it is difficult for high-temperature and high-pressure steam to fully mix with natural vitamin E oil in water and oil phases, and second, the air carried in the high-temperature and high-pressure steam will destroy the activity of natural vitamin E (natural vitamin E is easily oxidized and has poor heat resistance). Therefore, the high-temperature and high-pressure cooking method has obvious limitations for removing Zen toxins from natural vitamin E oil. Summary of the invention

[0009] The method of this patent follows the following principles: 1. It does not change the nutritional properties of natural vitamin E oil; 2. It can quickly and effectively remove or destroy zearalenone; 3. It does not produce toxic residues or carcinogenic / mutagenic residues in the processed natural vitamin E; 4. It is technically feasible and economically affordable, and can significantly affect the cost of the final product.

[0010] The invention provides a method for removing zearalenone toxin from natural vitamin E oil, the method comprising: using a silica gel column for chromatography treatment, eluting three times in sequence, and the amount of each eluent is 0.8-1.5 times the volume of the column. The first eluent is mobile phase A, the second eluent is mobile phase A and mobile phase B, and the volume ratio of mobile phase A to mobile phase B is 8:3-10:1, and the third eluent is mobile phase C. Wherein, mobile phase A is selected from one of n-hexane, petroleum ether and cyclohexane, and is preferably n-hexane or petroleum ether. Mobile phase B is the same as or different from mobile phase C. Mobile phase B is selected from one of ethyl acetate, acetone, isobutyl acetate and ethanol, and is preferably ethyl acetate. Mobile phase C is selected from one of ethyl acetate, acetone, isobutyl acetate and ethanol, and is preferably ethyl acetate or isobutyl acetate. The components of the second and third elutions are collected, and the eluent is removed by reduced pressure concentration to obtain the product.

[0011] Among them, natural vitamin E oil is a by-product of processing soybean oil, corn oil or rapeseed oil (such as produced during the deodorization process), in which the tocopherol content is greater than 40wt%.

[0012] The silica gel column is a low-pressure chromatography column with a height-to-diameter ratio of 5:1-10:1, and the particle size of the silica gel used is 100-500 meshes. Preferably, the particle size of the silica gel used is 100-200 meshes.

[0013] The natural vitamin E oil is diluted with mobile phase A to obtain a sample solution, and then loaded onto the column. The volume mass ratio of the mobile phase A used to dilute the natural vitamin E oil to the natural vitamin E oil is 0.4-1.0 ml / g, and the mass ratio of the natural vitamin E oil to the silica gel in the silica gel column is 1:10-2:5.

[0014] The flow rate of the eluent is 3-8 ml / min.

[0015] Furthermore, after elution is completed, mobile phase A is used for regeneration, and the amount of mobile phase A used for regeneration is 0.8-1.5 times the volume of the column. In this patent, the regeneration of the silica gel column is simple and the number of times is greater than 10 times, which can significantly reduce costs and is suitable for industrial production.

[0016] Preferably, during the second elution, the volume ratio of mobile phase A to mobile phase B is 8:2-9:1.

[0017] Specifically, the first eluent is n-hexane, the second eluent is n-hexane and ethyl acetate, and the third eluent is ethyl acetate. Or the first eluent is petroleum ether, the second eluent is petroleum ether and ethyl acetate, and the third eluent is ethyl acetate. Or the first eluent is cyclohexane, the second eluent is cyclohexane and acetone, and the third eluent is acetone. Or the first eluent is petroleum ether, the second eluent is petroleum ether and ethyl acetate, and the third eluent is isobutyl acetate. Or the first eluent is petroleum ether, the second eluent is petroleum ether and ethanol, and the third eluent is isobutyl acetate.

[0018] More preferably, the first eluent is petroleum ether, the second eluent is petroleum ether and ethyl acetate and the volume ratio of petroleum ether to ethyl acetate is 85:15, and the third eluent is isobutyl acetate.

[0019] Preferably, the method for removing zearalenone toxin from natural vitamin E oil disclosed in the present invention comprises: using a silica gel column for chromatography, the height-to-diameter ratio of the silica gel column is 10:1, and the particle size of the silica gel used is 100-200 mesh. Dilute the natural vitamin E oil with petroleum ether, the mass volume ratio of petroleum ether to natural vitamin E oil is 0.625 ml / g, and the mass ratio of natural vitamin E oil to silica gel in the silica gel column is 32:100. Eluted three times in sequence, the amount of eluent used each time is 1 times the volume of the column, and the flow rate of the eluent is 5 ml / min. The first eluent is petroleum ether, the second eluent is petroleum ether and ethyl acetate, and the volume ratio of petroleum ether to ethyl acetate is 85:15, and the third elution is isobutyl acetate. Collect the components of the second and third elutions. Concentrate and remove the eluent to obtain the product.

[0020] Preferably, the method for removing zearalenone toxin from natural vitamin E oil disclosed in the present invention comprises: using a silica gel column for chromatography, the height-to-diameter ratio of the silica gel column is 10:1, and the particle size of the silica gel used is 100-200 mesh. Dilute the natural vitamin E oil with n-hexane, the mass volume ratio of n-hexane to natural vitamin E oil is 0.67 ml / g, and the mass ratio of natural vitamin E oil to silica gel in the silica gel column is 3:10. Eluted three times in sequence, the amount of eluent used each time is 1 times the volume of the column, and the flow rate of the eluent is 5 ml / min. The first eluent is n-hexane, the second eluent is n-hexane and ethyl acetate, and the volume ratio of n-hexane to ethyl acetate is 80:20, and the third elution is ethyl acetate. Collect the components of the second and third elutions. Concentrate and remove the eluent to obtain the product.

[0021] The present invention utilizes the difference in adsorption and analysis of different polar components by silica gel chromatography column to purify natural vitamin E and remove zearalenone toxin at the same time. Column chromatography is one of the important means for purifying natural vitamin E. Commonly used fillers include strong alkaline ion resins, alumina, etc. Silica gel is a low-cost, reusable conventional filler. The silanol group in its structure has a good effect of adsorbing polar components. The structure of natural vitamin E is a phenolic hydroxyl group, and the structure of zearalenone toxin is 2 phenolic hydroxyl groups and 1 carbonyl group. The polarity difference between the two is significant. Through different eluents, natural vitamin E and zearalenone toxin can be well separated. This method does not bring about a significant increase in separation costs, and the separation process is under normal temperature and pressure conditions, which will not affect the original nutritional properties of natural vitamin E oil, nor will it bring about the residue of toxic substances, and is very suitable for large-scale purification and production of natural vitamin E oil. Using the method of this patent, the recovery rate of natural vitamin oil is greater than 92%, and reaches 97.99% under optimal conditions; the Zen removal rate is greater than 99%; under optimal conditions, it reaches 99.69%, which is less than 20ug / kg, far less than the national standard value. DETAILED DESCRIPTION

[0022] In order to make the objectives, technical solutions and advantages of the present invention more clear, the embodiments of the present invention are described in further detail below.

[0023] In the following embodiments, the Zen content in the natural vitamin E oil raw material used is 3000.5 μg / kg, which is 6.0 times that specified in the national standard GB13078-2017, and the tocopherol content is 50.2%; among them, the proportions of various tocopherols are: α-tocopherol 17.2%, γ-tocopherol 59.3%, and δ-tocopherol 23.5%.

[0024] Example 1 The chromatography column was filled with commercially available 100-200 mesh industrial grade silica gel, which was dry packed according to a height-to-diameter ratio of 10:1, and the silica gel was infiltrated with n-hexane. After the sedimentation stabilized, the amount of silica gel was measured to be 100 g, the column volume was 200 ml, and the column was set aside. The following Examples 2-4 were all operated using this chromatography column.

[0025] Example 2 Take 30.0g of natural vitamin E oil, dilute it with 20ml of n-hexane to reduce the viscosity of the raw material, and load it onto the column through a peristaltic pump with a feed rate of 5ml / min. After loading, elute with 200ml of n-hexane, 200ml of ethyl acetate / n-hexane (20 / 80; V / V), and 200ml of ethyl acetate. Collect the 200-400ml elution segment as the product segment, and recover the dry solvent under reduced pressure to obtain 15.8g of high-content natural vitamin E oil (tocopherol content is 93.1%, and the proportion of each tocopherol is: α-tocopherol 17.3%, γ-tocopherol 59.3%, δ-tocopherol 23.4%), Zen content is 18.8ug / kg, natural vitamin E oil recovery rate is 97.67%, and Zen removal rate is 99.67%.

[0026] Example 3 The column of Example 2 was used, and the replacement was completed with 100 ml of petroleum ether. The second batch of column loading was started. 30.0 g of natural vitamin E oil was taken, diluted with 20 ml of petroleum ether, and loaded onto the column by a peristaltic pump at a feed rate of 5 ml / min. After the loading was completed, 200 ml of petroleum ether, 200 ml of ethyl acetate / petroleum ether (20 / 80; V / V), and 200 ml of ethyl acetate were used for elution. The 200-400 ml elution section was collected as the product section, and the dry solvent was recovered under reduced pressure to obtain 15.1 g of high-content natural vitamin E oil (tocopherol content was 92.6%, and the proportion of each tocopherol was: α-tocopherol 17.4%, γ-tocopherol 59.4%, δ-tocopherol 23.2%), the Zen content was 30.5 ug / kg, the natural vitamin E oil recovery rate was 92.85%, and the Zen removal rate was 99.48%.

[0027] Example 4 The column of Example 3 was used, and the replacement was completed with 100 ml of cyclohexane. The third batch of column loading was started. 35.5 g of natural vitamin E was taken, diluted with 25 ml of cyclohexane, and loaded onto the column by a peristaltic pump at a feed rate of 4.5 ml / min. After the loading was completed, 200 ml of cyclohexane, 200 ml of acetone / cyclohexane (10 / 90; V / V), and 200 ml of acetone were used for elution. The 200-400 ml elution section was collected as the product section, and the dry solvent was recovered under reduced pressure to obtain 17.6 g of high-content natural vitamin E oil (tocopherol content was 95.1%, and the proportions of each tocopherol were: α-tocopherol 17.2%, γ-tocopherol 59.2%, δ-tocopherol 23.6%), the Zen content was 26.4 ug / kg, the natural vitamin E oil recovery rate was 93.92%, and the Zen removal rate was 99.61%.

[0028] Example 5 Referring to Example 1, two silica gel chromatography columns with a diameter-to-height ratio of 6:1 were newly installed, and the silica gel was selected to be 200-300 mesh industrial-grade amorphous silica gel. The columns were dry-packed, and the silica gel was infiltrated with n-hexane. After the sedimentation was stable, the amount of silica gel was measured to be 100 g, and the column volume was 200 ml. The columns were used for standby use. The following Examples 6-8 were all operated using this chromatography column.

[0029] Example 6 Using the column of Example 5, 32.0 g of natural vitamin E oil was taken, diluted with 20 ml of petroleum ether, loaded onto the column by a peristaltic pump at a feed rate of 5 ml / min. After loading, eluted with 200 ml of petroleum ether, 200 ml of ethyl acetate / petroleum ether (15 / 85; V / V), and 200 ml of isobutyl acetate, and the 200-400 ml elution segments were collected as product segments. The dry solvent was recovered under reduced pressure to obtain 16.8 g of high-content natural vitamin E oil (tocopherol content of 93.7%, the proportion of each tocopherol was: α-tocopherol 17.2%, γ-tocopherol 59.3%, δ-tocopherol 23.5%), the Zen content was 18.0 ug / kg, the natural vitamin E oil recovery rate was 97.99%, and the Zen removal rate was 99.69%.

[0030] Example 7 Using the column of Example 5, 32.0 g of natural vitamin E oil was taken, diluted with 20 ml of petroleum ether, loaded onto the column by a peristaltic pump at a feed rate of 5 ml / min. After loading, eluted with 200 ml of petroleum ether, 200 ml of ethanol / petroleum ether (10 / 90; V / V), and 200 ml of isobutyl acetate, and the 200th to 400th ml elution segments were collected as product segments. The dry solvent was recovered under reduced pressure to obtain 16.5 g of high-content natural vitamin E oil (tocopherol content of 94.3%, the proportion of each tocopherol was: α-tocopherol 17.3%, γ-tocopherol 59.4%, δ-tocopherol 23.3%), the Zen content was 36.6 ug / kg, the natural vitamin E oil recovery rate was 96.86%, and the Zen removal rate was 99.37%.

[0031] Summarize From the results of Examples 2-7, it can be seen that the content of Zen in natural vitamin E oil can be effectively reduced by silica gel chromatography, and the removal rate can basically reach more than 99%. This process is simple and easy, and has obvious specificity, and basically does not change the proportion of each tocopherol. While removing the Zen content, the 50% pure natural vitamin E can also be refined to more than 90%, significantly improving the economic value and use value of natural vitamin E.

[0032] Comparative Example 1 Using the column of Example 1, 30.0g of natural vitamin E oil was taken, diluted with 20ml of n-hexane to reduce the viscosity of the raw material, and loaded onto the column by a peristaltic pump at a feed rate of 5ml / min. After loading, 600ml of ethyl acetate / n-hexane (20 / 80; V / V) was used for elution, and the 200th-400th ml elution section was collected as the product section. The dry solvent was recovered under reduced pressure to obtain 15.3g of high-content natural vitamin E oil (tocopherol content was 92.7%, and the proportion of each tocopherol was: α-tocopherol 16.9%, γ-tocopherol 59.4%, δ-tocopherol 23.7%), Zen content was 495.9ug / kg, natural vitamin E oil recovery rate was 94.18%, and Zen removal rate was 83.47%. It can be seen from Comparative Example 1 that adjusting the eluent not only affects the purity (smaller impact), but also changes the proportion of each tocopherol (smaller impact); it mainly affects the Zen removal rate, and Zen may be greater than the national standard value.

[0033] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for removing zearalenone toxin from natural vitamin E oil, characterized in that: The method comprises: using a silica gel column for chromatography treatment, eluting three times in sequence, the amount of eluent used each time being 0.8-1.5 times the volume of the column; the first eluent is mobile phase A, the second eluent is mobile phase A and mobile phase B, and the volume ratio of mobile phase A to mobile phase B is 8:3-10:1, and the third eluent is mobile phase C; the mobile phase A is selected from one of n-hexane, petroleum ether and cyclohexane, the mobile phase B is the same as or different from the mobile phase C, the mobile phase B is selected from one of ethyl acetate, acetone, isobutyl acetate and ethanol, and the mobile phase C is selected from one of ethyl acetate, acetone, isobutyl acetate and ethanol; collecting the components of the second and third elutions, and concentrating under reduced pressure to remove the eluent to obtain the product.

2. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: Natural vitamin E oil is a byproduct of soybean oil, corn oil or rapeseed oil processing, in which the tocopherol content is greater than 40wt%.

3. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: The silica gel column is a low-pressure chromatography column with a height-to-diameter ratio of 5:1-10:1, and the particle size of the silica gel used is 100-500 meshes.

4. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: The natural vitamin E oil is diluted with mobile phase A, the volume mass ratio of the mobile phase A to the natural vitamin E oil is 0.4-1.0 ml / g, and the mass ratio of the natural vitamin E oil to the silica gel in the silica gel column is 1:10-2:

5.

5. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: The flow rate of the eluent is 3-8 ml / min.

6. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: After elution is completed, mobile phase A is used for regeneration, and the amount of mobile phase A used for regeneration is 0.8-1.5 times the column volume.

7. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: During the second elution, the volume ratio of mobile phase A to mobile phase B is 8:2-9:

1.

8. The method for removing zearalenone toxin from natural vitamin E oil according to claim 7, characterized in that: The first eluent was n-hexane, the second eluent was n-hexane and ethyl acetate, and the third eluent was ethyl acetate; Alternatively, the first eluent is petroleum ether, the second eluent is petroleum ether and ethyl acetate, and the third eluent is ethyl acetate; Alternatively, the first eluent is cyclohexane, the second eluent is cyclohexane and acetone, and the third eluent is acetone; Alternatively, the first eluent is petroleum ether, the second eluent is petroleum ether and ethyl acetate, and the third eluent is isobutyl acetate; Alternatively, the first eluent is petroleum ether, the second eluent is petroleum ether and ethanol, and the third eluent is isobutyl acetate.

9. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: The first eluent was petroleum ether, the second eluent was petroleum ether and ethyl acetate with a volume ratio of petroleum ether to ethyl acetate of 85:15, and the third eluent was isobutyl acetate.

10. The method for removing zearalenone toxin from natural vitamin E oil according to claim 1, characterized in that: The method comprises: using a silica gel column for chromatography treatment, wherein the height-to-diameter ratio of the silica gel column is 10:1, and the particle size of the silica gel used is 100-200 meshes; The natural vitamin E oil was diluted with petroleum ether, the mass volume ratio of petroleum ether to the natural vitamin E oil was 0.625 ml / g, and the mass ratio of the natural vitamin E oil to the silica gel in the silica gel column was 32:100; Elution was performed three times in sequence, the amount of the eluent used each time was 1 times the column volume, and the flow rate of the eluent was 5 ml / min; the first eluent was petroleum ether, the second eluent was petroleum ether and ethyl acetate, and the volume ratio of petroleum ether to ethyl acetate was 85:15, and the third eluent was isobutyl acetate; the components of the second and third elutions were collected; The product was obtained by concentrating under reduced pressure to remove the eluent.