A method for purifying lutein esters

Abstract

The application discloses a lutein ester purification method, and belongs to the technical field of lutein ester purification. The purification method comprises the following steps: S1, lutein ester is coarsely dissolved in an organic solvent to prepare a crude product solution; S2, the crude product solution is loaded into a chromatographic column containing a composite adsorbent, gradient elution is carried out, and an eluate is collected; and S3, the eluate is concentrated to obtain a concentrated slurry, an anti-solvent is added for crystallization, and then filtration, washing and drying are carried out. The composite adsorbent in step S2 is composed of silica gel and macroporous adsorption resin. The gradient elution in step S2 is carried out by sequentially using a first eluent and a second eluent. The first eluent is petroleum ether and ethyl acetate with a volume ratio of 10-20:1. The second eluent is petroleum ether and ethyl acetate with a volume ratio of 2-5:1. The purity of the lutein ester is finally increased to more than 99%, and the yield is as high as more than 85%.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of lutein ester purification, and particularly relates to a lutein ester purification method. BACKGROUND

[0002] Lutein ester is an important carotenoid fatty acid ester, mainly existing in marigold flowers. It can be hydrolyzed into lutein in the human body, which is the main pigment in the macular area of the retina, has important physiological functions such as antioxidant, blue light filtering, vision protection, and prevention of age-related macular degeneration, and is widely used in food, health products, drugs and feed additives.

[0003] At present, lutein ester crude product is mainly obtained from marigold flower particles through organic solvent extraction and saponification (or without saponification) in the industry. The lutein ester crude product has complex components, contains a large amount of impurities such as oil, wax, free fatty acids and other carotenoids, and the purity is usually only between 5%-20%, which cannot directly meet the needs of high-end applications. At present, purification of lutein ester crude product is the key to improve the product value.

[0004] Chinese Patent Publication No. CN101891664A discloses a method for purifying lutein ester. First, marigold oil paste is dissolved in ethyl acetate at 25-80°C, and stirred for 1-6 hours. The filtered liquid is added to anhydrous ethanol, and stirred at 20-70°C for 0.5-5.0 hours to form a uniform solution. Then, after cooling at -5-30°C for 5-20 hours, filtration is performed. The filter cake is washed with low-grade alcohol, and vacuum drying is performed to remove solvent residues, thereby obtaining the desired high-purity lutein ester. The content of the obtained lutein ester is 70%-80%, and the yield of total lutein ester is about 45-69%. Chinese Patent Publication No. CN106588732A discloses a method for purifying lutein ester from marigold oil paste, comprising the following steps: a. dissolving marigold oil paste in an ether solvent with a mass of 1-5 times that of the marigold oil paste, and stirring to form a uniform solution x; b. filtering the solution x at 20-70°C, obtaining a filtrate, then adding deionized water to the filtrate, stirring for 0.5-5 hours, and filtering again to obtain crystals y; c. washing the crystals y with deionized water until the deionized water flowing out after washing is colorless; d. drying the washed crystals in a vacuum for 1-3 days. The content of the lutein ester obtained by the method is 75%-85%, and the yield is 72%-80%.

[0005] At present, the yield and purity of lutein ester purified from lutein ester crude product from marigold flowers are not ideal, and there is a large space for improvement. Developing a lutein ester purification method that can achieve high purity and high yield at the same time has important industrial application value. SUMMARY

[0006] In order to solve the above technical problems, the application discloses a purification method of lutein ester, which realizes efficient purification of lutein ester crude product through coupling of specific adsorbent combination, gradient elution and crystallization process, and the purity of the final product can reach more than 99%, and the yield is more than 85%.

[0007] In order to achieve the above-mentioned purpose, the application adopts the following technical scheme.

[0008] The application provides a purification method of lutein ester, comprising the following steps:

[0009] S1, lutein ester is dissolved in an organic solvent to prepare a crude product solution;

[0010] S2, the crude product solution is loaded into a chromatographic column containing a composite adsorbent, gradient elution is performed, and eluate is collected;

[0011] S3, the eluate is concentrated to obtain concentrated slurry, an anti-solvent is added for crystallization, filtration, washing and drying are performed.

[0012] In some embodiments, the organic solvent in step S1 is petroleum ether and / or n-hexane.

[0013] In some embodiments, the concentration of the crude product solution in step S1 is 50-150 mg / mL; preferably, 115-135 mg / mL.

[0014] In some embodiments, the composite adsorbent in step S2 is composed of silica gel and macroporous adsorption resin, preferably, the mass ratio of the silica gel and the macroporous adsorption resin is 3-1:1; preferably, the macroporous adsorption resin is HPD-100 or AB-8 type resin.

[0015] In some embodiments, the height-diameter ratio of the chromatographic column in step S2 is 10-15:1; preferably, 11-13:1.

[0016] In some embodiments, the gradient elution in step S2 is elution with a first eluent and a second eluent in sequence, preferably, the first eluent is petroleum ether and ethyl acetate with a volume ratio of 10-20:1, further preferably, 12-18:1; the second eluent is petroleum ether and ethyl acetate with a volume ratio of 2-5:1, further preferably, 3-4:1. Further preferably, the gradient elution is: elution with the first eluent until the effluent is colorless, then elution with the second eluent, collection of orange-red section eluate, and elution until the effluent is colorless.

[0017] In some embodiments, the concentration in step S3 is concentrated to a solution in a viscous state, and a crystal film is formed on the liquid surface.

[0018] In some embodiments, the volume ratio of the anti-solvent and the concentrated slurry in step S3 is 3-5:1; preferably 4-5:1.

[0019] In some embodiments, the anti-solvent in step S3 is a lower alcohol, preferably a C1-C4 alcohol, further preferably methanol or ethanol.

[0020] In some embodiments, the crystallization in step S3 is standing at 0-4°C for 4-8h; preferably standing at 1-3°C for 5-7h.

[0021] In some embodiments, the washing in step S3 is washing with the anti-solvent at 0-4°C.

[0022] In some embodiments, the drying in step S3 is freeze-drying, vacuum drying or spray drying. Preferably, it is vacuum drying.

[0023] Compared with the prior art, the present application has the following advantages:

[0024] 1. Purity and yield are significantly improved: the present application uses a composite adsorbent composed of silica gel and macroporous adsorption resin, which ingeniously combines the high separation efficiency of silica gel and the large adsorption capacity and anti-poisoning ability of macroporous adsorption resin. By controlling the gradient elution parameters, efficient and clear separation of impurities and target product is achieved. Combined with the optimized crystallization process (anti-solvent type, ratio and crystallization temperature), the purity of lutein ester is finally increased to more than 99%, and the comprehensive yield is as high as more than 85%.

[0025] 2. Low cost and easy to scale up: the composite adsorbent and solvent used in the present application are conventional chemical products, which are cheap and widely available. The chromatographic column can be scaled up, and the whole process conditions are mild, which is very suitable for large-scale industrial production.

[0026] 3. Good product quality: since strong acid, strong base or high temperature treatment is avoided in the process, isomerization and degradation of lutein ester are effectively prevented, and the obtained product has bright color (deep orange red), low impurity residue and good stability.

[0027] 4. Green and environmentally friendly: the solvents used (petroleum ether, ethyl acetate, ethanol) have low toxicity, and can be recycled by distillation, which reduces the production cost and environmental burden. DETAILED DESCRIPTION

[0028] The following description of the embodiments is merely exemplary in nature and is in no way intended to limit the scope of the application, its application, or its core teachings. It should be noted that for a person skilled in the art, certain improvements and modifications can be made to the application without departing from the principles of the application, and these improvements and modifications also fall within the scope of protection of the claims of the application. The following description of the disclosed embodiments enables any person skilled in the art to implement or use the application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the application. Accordingly, the present application is not to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application, the preferred methods and materials are described herein.

[0029] Unless otherwise defined, all technical and scientific terms used in the application have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The crude lutein ester in the application is a crude lutein ester from marigold flowers, and the purity detected by HPLC is 16.5%. The silica gel is column chromatography silica gel (100-200 mesh).

[0030] Example 1

[0031] A method for purifying lutein ester, comprising the following steps:

[0032] S1, dissolve the crude lutein ester in petroleum ether to prepare a crude product solution of 50 mg / mL;

[0033] S2, load the crude product solution into a chromatographic column (height:diameter ratio 10:1) containing a composite adsorbent (mass ratio of silica gel and HPD-100 macroporous adsorbent resin 3:1), and then perform gradient elution, use the first eluent (volume ratio of petroleum ether to ethyl acetate 10:1) for elution until the effluent is colorless, then use the second eluent (volume ratio of petroleum ether to ethyl acetate 2:1) for elution, collect the orange-red section of the eluent until the effluent is colorless;

[0034] S3, concentrate the eluent to a solution that is viscous with a crystal film on the liquid surface, add 3 times the volume of ethanol to the concentrated slurry, and stand in a freezer at 0°C for 4 hours. Filter, wash the crystals twice with pre-cooled (4°C) anhydrous ethanol, and dry the filter cake in a vacuum drying oven (35°C, -0.095 MPa) for 6 hours to obtain orange-red crystals.

[0035] Example 2

[0036] A method for purifying lutein ester, comprising the following steps:

[0037] S1, the lutein ester crude product is dissolved in n-hexane to prepare a crude product solution of 150 mg / mL;

[0038] S2, the crude product solution is loaded into a chromatographic column (height-diameter ratio of 15:1) containing a composite adsorbent (mass ratio of silica gel and HPD-100 macroporous adsorbent resin of 1:1) and gradient elution is performed, first eluent (volume ratio of petroleum ether to ethyl acetate of 20:1) is used for elution until the effluent is colorless, then second eluent (volume ratio of petroleum ether to ethyl acetate of 5:1) is used for elution, and orange-red eluent is collected until the effluent is colorless;

[0039] S3, the eluent is concentrated to a viscous solution with a crystal film formed on the liquid surface, 5 times the volume of ethanol is added to the concentrated slurry, and the mixture is placed in a refrigerator at 4°C for 8 hours. The crystals are filtered and washed twice with pre-cooled (4°C) anhydrous ethanol, and the filter cake is placed in a vacuum drying oven (35°C, -0.095 MPa) for drying for 6 hours to obtain orange-red crystals.

[0040] Example 3

[0041] A lutein ester purification method, comprising the following steps:

[0042] S1, the lutein ester crude product is dissolved in petroleum ether to prepare a crude product solution of 125 mg / mL;

[0043] S2, the crude product solution is loaded into a chromatographic column (height-diameter ratio of 12:1) containing a composite adsorbent (mass ratio of silica gel and HPD-100 macroporous adsorbent resin of 2:1) and gradient elution is performed, first eluent (volume ratio of petroleum ether to ethyl acetate of 15:1) is used for elution until the effluent is colorless, then second eluent (volume ratio of petroleum ether to ethyl acetate of 4:1) is used for elution, and orange-red eluent is collected until the effluent is colorless;

[0044] S3, the eluent is concentrated to a viscous solution with a crystal film formed on the liquid surface, 4 times the volume of ethanol is added to the concentrated slurry, and the mixture is placed in a refrigerator at 2°C for 6 hours. The crystals are filtered and washed twice with pre-cooled (4°C) anhydrous ethanol, and the filter cake is placed in a vacuum drying oven (35°C, -0.095 MPa) for drying for 6 hours to obtain orange-red crystals.

[0045] Comparative Example 1

[0046] The difference between this comparative example and Example 3 is that the composite adsorbent in step S2 is silica gel, and the elution method is different.

[0047] Specifically, a lutein ester purification method, comprising the following steps:

[0048] S1, the lutein ester crude product is dissolved in petroleum ether to prepare a crude product solution of 125 mg / mL;

[0049] S2, the crude product solution is loaded into a chromatographic column (height-diameter ratio 12:1) containing a composite adsorbent (silica gel) and gradient elution is performed, an eluent (petroleum ether and ethyl acetate in a volume ratio of 4:1) is used for elution, and orange-red eluate is collected until the effluent is colorless;

[0050] S3, the eluate is concentrated to a viscous solution with a crystal film formed on the liquid surface, 4 volumes of ethanol are added to the concentrated slurry, and the mixture is placed in a refrigerator at 2°C for 6 hours. The mixture is filtered, the crystals are washed twice with pre-cooled (4°C) anhydrous ethanol, and the filter cake is placed in a vacuum drying oven (35°C, -0.095 MPa) for drying for 6 hours to obtain orange-red crystals.

[0051] Comparative Example 2

[0052] The difference between this comparative example and Example 3 is that the composite adsorbent in step S2 is different.

[0053] Specifically, a lutein ester purification method comprises the following steps:

[0054] S1, the lutein ester crude product is dissolved in petroleum ether to prepare a crude product solution of 125 mg / mL;

[0055] S2, the crude product solution is loaded into a chromatographic column (height-diameter ratio 12:1) containing a composite adsorbent (HPD-100 macroporous adsorption resin) and gradient elution is performed, a first eluent (petroleum ether and ethyl acetate in a volume ratio of 15:1) is used for elution until the effluent is colorless, a second eluent (petroleum ether and ethyl acetate in a volume ratio of 4:1) is used for elution, and eluate is collected until the effluent is colorless.

[0056] S3, the eluate is concentrated to a viscous solution with a crystal film formed on the liquid surface, 4 volumes of ethanol are added to the concentrated slurry, and the mixture is placed in a refrigerator at 2°C for 6 hours. The mixture is filtered, the crystals are washed twice with pre-cooled (4°C) anhydrous ethanol, and the filter cake is placed in a vacuum drying oven (35°C, -0.095 MPa) for drying for 6 hours to obtain orange-red crystals.

[0057] Comparative Example 3

[0058] The difference between this comparative example and Example 3 is that the mass ratio of silica gel and HPD-100 macroporous adsorption resin is different, and the first eluent and the second eluent are different.

[0059] A lutein ester purification method comprises the following steps:

[0060] S1, the lutein ester crude product is dissolved in petroleum ether to prepare a crude product solution of 125 mg / mL;

[0061] S2, the crude solution is loaded into a chromatographic column (height-diameter ratio 12:1) containing composite adsorbent (mass ratio of silica gel and HPD-100 macroporous adsorbent resin is 5:1) and gradient elution is performed, elution is performed using a first eluent (volume ratio of petroleum ether to ethyl acetate is 18:1) until the effluent is colorless, and then a second eluent (volume ratio of petroleum ether to ethyl acetate is 6:1) is used for elution, and orange-red eluent is collected until the effluent is colorless;

[0062] S3, the eluent is concentrated to a viscous solution with crystal film formed on the liquid surface, 4 times the volume of ethanol is added to the concentrated slurry, and the mixture is placed in a refrigerator at 2℃ for 6 hours. Filtration is performed, the crystals are washed twice with pre-cooled (4℃) anhydrous ethanol, and the filter cake is placed in a vacuum drying oven (35℃, -0.095 MPa) for drying for 6 hours to obtain orange-red crystals.

[0063] Comparative Example 4

[0064] The difference between this comparative example and Example 3 is that the macroporous adsorbent resin is different and the crystallization method is different.

[0065] A method for purifying a lutein ester, comprising the following steps:

[0066] S1, the lutein ester crude product is dissolved in petroleum ether to prepare a crude solution of 125 mg / mL;

[0067] S2, the crude solution is loaded into a chromatographic column (height-diameter ratio 12:1) containing composite adsorbent (mass ratio of silica gel and D-101 macroporous adsorbent resin is 2:1) and gradient elution is performed, elution is performed using a first eluent (volume ratio of petroleum ether to ethyl acetate is 15:1) until the effluent is colorless, and then a second eluent (volume ratio of petroleum ether to ethyl acetate is 4:1) is used for elution, and orange-red eluent is collected until the effluent is colorless;

[0068] S3, the eluent is concentrated to a viscous solution with crystal film formed on the liquid surface, 4 times the volume of ethanol is added to the concentrated slurry, and the mixture is placed in a refrigerator at 6℃ for 12 hours. Filtration is performed, the crystals are washed twice with pre-cooled (4℃) anhydrous ethanol, and the filter cake is placed in a vacuum drying oven (35℃, -0.095 MPa) for drying for 6 hours to obtain orange-red crystals.

[0069] The purity and yield results of the purification method are shown in Table 1.

[0070] Table 1

[0071]

[0072] The results show that the purity of the lutein ester obtained by the purification methods of Examples 1-3 is more than 99% and the yield is more than 85%, achieving high purity and high yield.

[0073] From the comparison of Comparative Example 4, using the conventional adsorbent and crystallization method, the purity of lutein ester after purification reached more than 95%, but the yield was less than 70%, which could not take into account both high purity and high yield. From the comparison of Comparative Examples 1-3, using the conventional adsorbent and elution method, the yield of lutein ester after purification reached more than 70%, but the purity was low and could not break through 95%. From the comparison of Example 3 and Comparative Examples 1-4, the present application combines the composite adsorbent, elution method and crystallization step, and has achieved unexpected technical effects in improving the purity and yield of lutein ester.

[0074] The above is a further description of the present application in combination with specific examples, but these examples are only exemplary and do not constitute any limitation on the scope of the present application. Those skilled in the art should understand that the details and forms of the technical solutions of the present application can be modified or replaced without departing from the spirit and scope of the present application, and such modifications and replacements all fall within the protection scope of the present application.

Claims

1. A method for purifying lutein esters, characterized in that, Includes the following steps: S1, lutein esters are crudely dissolved in an organic solvent to prepare a crude solution; S2. Load the crude product solution onto a chromatography column containing a composite adsorbent, elute with gradient, and collect the eluent. S3. The eluent is concentrated to obtain a concentrated slurry. An antisolvent is added and the mixture is allowed to stand at 0-4℃ for 4-8 hours to crystallize. The slurry is then filtered, washed, and dried. The composite adsorbent described in step S2 is composed of silica gel and HPD-100 macroporous adsorption resin in a mass ratio of 3-1:

1. The gradient elution in step S2 involves eluting with a first eluent and a second eluent sequentially. The first eluent is petroleum ether and ethyl acetate in a volume ratio of 10-20:1; the second eluent is petroleum ether and ethyl acetate in a volume ratio of 2-5:

1. The crude lutein ester mentioned in step S1 is crude lutein ester derived from marigold flowers; The gradient elution described in step S2 involves eluting with the first eluent until the effluent is colorless, then eluting with the second eluent, collecting the orange-red eluent, until the effluent is colorless. The silica gel mentioned in step S2 is 100-200 mesh column chromatography silica gel.

2. The purification method according to claim 1, characterized in that, The organic solvent mentioned in step S1 is petroleum ether and / or n-hexane.

3. The purification method according to claim 1, characterized in that, The concentration of the crude product solution in step S1 is 50-150 mg / mL.

4. The purification method according to claim 3, characterized in that, The concentration of the crude product solution in step S1 is 115-135 mg / mL.

5. The purification method according to claim 1, characterized in that, The height-to-diameter ratio of the chromatography column in step S2 is 10-15:

1.

6. The purification method according to claim 5, characterized in that, The first eluent is petroleum ether and ethyl acetate in a volume ratio of 12-18:1; the second eluent is petroleum ether and ethyl acetate in a volume ratio of 3-4:

1.

7. The purification method according to claim 1, characterized in that, The concentration mentioned in step S3 is to concentrate the solution until it becomes viscous and a crystal film forms on the surface of the liquid.

8. The purification method according to claim 1, characterized in that, The volume ratio of the antisolvent to the concentrated slurry in step S3 is 3-5:

1.

9. The purification method according to claim 8, characterized in that, The antisolvent mentioned in step S3 is a C1-C4 alcohol.

10. The purification method according to claim 9, characterized in that, The antisolvent mentioned in step S3 is methanol or ethanol.

11. The purification method according to claim 1, characterized in that, The crystallization described in step S3 involves standing at 1-3°C for 5-7 hours.

12. The purification method according to claim 9, characterized in that, The washing described in step S3 is washing with an anti-solvent at 0-4°C.

Images