Chromatographic device and method for removing p-dihydroxybenzene in alpha-matrix maclura fruit glycoside

Abstract

The present application relates to a kind of chromatographic device and method for removing p-benzenediol in alpha-matilisin.The device includes vertically arranged DEAE-anion high molecular polymer packing chromatographic column, detector arranged above the chromatographic column and communicated with the chromatographic column, upper pipeline system arranged above the detector and communicated with the detector, lower pipeline system arranged below the chromatographic column and communicated with the chromatographic column, and control system adapted to the upper pipeline system and the lower pipeline system respectively, the control system includes controller and time relay.Compared with the prior art, the present application can complete the separation of p-benzenediol in alpha-matilisin by one-step operation, the separation process is simple, efficient, and the DEAE-anion high molecular polymer packing chromatographic column used as a weak anion exchange medium does not need to use acid and alkali for regeneration, and is friendly to the environment.The DEAE-anion high molecular polymer packing chromatographic column has long service life, high separation efficiency, can greatly reduce production cost, and is suitable for industrial production application.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of α-arbutin preparation, and relates to a chromatographic device and method for removing the impurity hydroquinone in α-arbutin synthesized by an enzyme catalysis method. BACKGROUND

[0002] Arbutin is a hydroquinone glycoside compound and is an inhibitor of tyrosinase, which can effectively inhibit the synthesis of dopamine and dopaquinone, thereby effectively inhibiting the generation of melanin and having a whitening effect. Natural extracts contain β-arbutin, but the whitening effect of α-arbutin is more than 10 times that of β-arbutin, and the effectiveness and safety are higher than those of β-arbutin. However, it is difficult to chemically synthesize α-arbutin, and it is easy to form a mixture of α-arbutin and β-arbutin, which is difficult to separate. Therefore, more and more researches tend to synthesize α-arbutin by a microbial method or an enzyme method.

[0003] The enzyme catalysis method for synthesizing α-arbutin uses maltose and hydroquinone as raw materials, and a small amount of hydroquinone is left in the product of α-arbutin obtained after the enzyme catalysis reaction is completed. Hydroquinone is a known skin sensitizer, and is prohibited to be used in skin care products. Therefore, if the hydroquinone in α-arbutin cannot be effectively removed, the product quality will be directly affected and even the product will be unqualified.

[0004] At present, the market has very high requirements for the residual amount of hydroquinone, which needs to be less than 10 ppm or not detected. However, in the separation and purification process of the current α-arbutin production process, the pigment molecules are first removed by a weak polar macroporous adsorption resin, and then adsorbed by a strong polar macroporous adsorption resin, so that α-arbutin and hydroquinone are separated according to the difference in polarity. Because the difference in polarity between α-arbutin and hydroquinone is not large, it is difficult to completely remove the hydroquinone, the efficiency is low, and it is difficult to be applied to industrial production.

[0005] Chinese Patent Publication No. CN103483401A discloses a method for separating hydroquinone from α-arbutin fermentation broth, which is extracted and back-extracted by tributyl phosphate for multiple times, and needs to introduce inorganic salts. Therefore, the method is complex in process, and the used tributyl phosphate is not friendly to the environment. SUMMARY

[0006] The purpose of the present application is to provide a chromatographic device and method for removing hydroquinone in α-arbutin, which has a high recovery rate of α-arbutin, completely removes hydroquinone, and has good color and no organic solvent residue.

[0007] The purpose of the present application can be achieved by the following technical solutions:

[0008] A kind of chromatographic device for removing pyrocatechol in alpha-arbutin, the device includes vertically arranged DEAE-anion high molecular polymer packing chromatographic column, detector being arranged above DEAE-anion high molecular polymer packing chromatographic column and being communicated with DEAE-anion high molecular polymer packing chromatographic column, upper pipeline system being arranged above detector and being communicated with detector, lower pipeline system being arranged below DEAE-anion high molecular polymer packing chromatographic column and being communicated with DEAE-anion high molecular polymer packing chromatographic column, and control system being adapted to upper pipeline system and lower pipeline system respectively, the control system includes controller and time relay.Controller is electrically connected with time relay, and plays the role of automatic control and delay control.

[0009] Further, the detector is UV detector.

[0010] Further, the upper pipeline system includes mixed liquid outlet pipe, and the lower pipeline system includes mixed liquid inlet pipe.Mixed liquid containing alpha-arbutin and pyrocatechol enters DEAE-anion high molecular polymer packing chromatographic column from the bottom of DEAE-anion high molecular polymer packing chromatographic column, and is discharged from the top of DEAE-anion high molecular polymer packing chromatographic column after removing pyrocatechol.

[0011] Further, the upper pipeline system further includes regeneration liquid inlet pipe, and the lower pipeline system further includes regeneration liquid outlet pipe.During regeneration, regeneration liquid enters DEAE-anion high molecular polymer packing chromatographic column from the top of DEAE-anion high molecular polymer packing chromatographic column, and is discharged from the bottom of DEAE-anion high molecular polymer packing chromatographic column after eluting pyrocatechol.

[0012] Further, the upper pipeline system further includes upper balance liquid pipeline, and the lower pipeline system further includes lower balance liquid pipeline.Upper balance liquid pipeline serves as balance liquid outlet pipe, and lower balance liquid pipeline serves as balance liquid inlet pipe during separating pyrocatechol.Upper balance liquid pipeline serves as balance liquid inlet pipe, and lower balance liquid pipeline serves as balance liquid outlet pipe during regeneration.

[0013] Further, electromagnetic valve is arranged on mixed liquid outlet pipe, upper balance liquid pipeline, regeneration liquid inlet pipe, mixed liquid inlet pipe, lower balance liquid pipeline and regeneration liquid outlet pipe, and the control system is electrically connected with electromagnetic valve.Control system controls the on-off of corresponding pipeline through electromagnetic valve on each pipeline.

[0014] Further, one-way valve is arranged on mixed liquid outlet pipe, regeneration liquid inlet pipe, mixed liquid inlet pipe and regeneration liquid outlet pipe.One-way valve is used to ensure the one-way flow of liquid in corresponding pipeline.

[0015] Further, the lower pipeline system is provided with a filler column between the filler column and the DEAE-anion high polymer filler chromatographic column, and the lower pipeline system, the filler column and the DEAE-anion high polymer filler chromatographic column are sequentially connected.

[0016] Further, the filler column is provided with a stainless steel capillary tube between the filler column and the DEAE-anion high polymer filler chromatographic column, and the filler column and the DEAE-anion high polymer filler chromatographic column are connected through the stainless steel capillary tube. The filler column and the chromatographic column are connected through the stainless steel capillary tube, which is not only more stable, but also improves the separation efficiency of the chromatographic column due to the small inner diameter of the stainless steel capillary tube, i.e. the small column volume of the DEAE-anion high polymer filler chromatographic column.

[0017] A method for removing p-benzenediol in alpha-arbutin based on the chromatographic device, characterized in that the method comprises the following steps: adjusting a mixed solution containing alpha-arbutin and p-benzenediol to be alkaline, so that the p-benzenediol reacts to form p-benzenediol salt, and then the mixed solution is introduced into the DEAE-anion high polymer filler chromatographic column through the lower pipeline system, the p-benzenediol salt in the mixed solution is adsorbed on the DEAE-anion high polymer filler chromatographic column, and the purified alpha-arbutin solution is discharged through the upper pipeline system.

[0018] Preferably, the mixed solution is adjusted to pH 9-10 by sodium hydroxide.

[0019] In application, the mixed solution containing alpha-arbutin and p-benzenediol is first added with sodium hydroxide to adjust the pH to 9-10, so that the p-benzenediol reacts to form a negatively charged and ionizable phenolate under weak alkaline conditions, and then enters the DEAE-anion high polymer filler chromatographic column, removes the p-benzenediol impurities through adsorption, and penetrates the alpha-arbutin with no charge and small adsorption force. The recovery rate of alpha-arbutin separated by the adsorption-penetration method is high, the removal of p-benzenediol is more thorough, and the prepared alpha-arbutin has good color and no organic solvent residue. Through the cooperation of the controller and the time relay, the functions of automatic liquid feeding, liquid switching, collection of purified alpha-arbutin solution, chromatographic column regeneration, and time delay operation can be realized.

[0020] Compared with the prior art, the present application has the following characteristics:

[0021] 1) The present application can separate the p-benzene diol in alpha-matilisin by one-step operation, and the separation process is simple and efficient. The DEAE-anion high polymer filler chromatographic column is used as a weak anion exchange medium, and does not need to use acid and alkali for regeneration, which is environment-friendly. The DEAE-anion high polymer filler chromatographic column has long service life and high separation efficiency, and can greatly reduce the production cost, and is suitable for industrialized production application.

[0022] 2) The separation of small molecule compounds is mainly based on differential separation. If the low-pressure chromatographic column is filled from the top end, the column wall penetration phenomenon will occur, which causes a part of the mixed liquid to move down along the column wall, so that the mixed liquid is not evenly distributed on the surface of the chromatographic medium, and the separation effect is poor. Therefore, the present application adopts the way of filling from the bottom end of the chromatographic column, which avoids the above phenomenon and improves the separation effect.

[0023] 3) When the chromatographic column is regenerated, the adsorption amount of the mixed liquid at the lower end of the chromatographic column is large, and the retention strength is relatively weak and easy to dissociate. Therefore, the present application adopts the way of introducing the regeneration liquid from the top end of the chromatographic column, which improves the regeneration effect. BRIEF DESCRIPTION OF DRAWINGS

[0024] Figure 1 Figure 1 is a structural schematic diagram of the chromatographic device in the present application;

[0025] Figure 2 Figure 2 is a high performance liquid chromatography (HPLC) analysis diagram of the alpha-matilisin liquid collected in the example;

[0026] Marking explanation in the figure:

[0027] 1-DEAE-anion high polymer filler chromatographic column, 2-detector, 3-controller, 4-time relay, 5-mixed liquid outlet pipe, 6-upper equilibrium liquid pipe, 7-regeneration liquid inlet pipe, 8-mixed liquid inlet pipe, 9-lower equilibrium liquid pipe, 10-regeneration liquid outlet pipe, 11-solenoid valve, 12-filler column, 13-stainless steel capillary. DETAILED DESCRIPTION

[0028] The present application will be described in detail below in combination with the drawings and specific examples. The present embodiment is implemented on the premise of the technical scheme of the present application, and gives detailed implementation mode and specific operation process, but the protection scope of the present application is not limited to the following examples.

[0029] Example:

[0030] As Figure 1The chromatographic device for removing pyrocatechol in alpha-matrix includes a DEAE-anion high polymer filler chromatographic column 1 arranged vertically, a detector 2 arranged above the DEAE-anion high polymer filler chromatographic column 1 and communicated with the DEAE-anion high polymer filler chromatographic column 1, an upper pipeline system arranged above the detector 2 and communicated with the detector 2, a lower pipeline system arranged below the DEAE-anion high polymer filler chromatographic column 1 and communicated with the DEAE-anion high polymer filler chromatographic column 1, and a control system matched with the upper pipeline system and the lower pipeline system respectively, wherein the control system includes a controller 3 and a time relay 4.

[0031] The detector 2 is a UV detector.

[0032] The upper pipeline system includes a mixed liquid outlet pipe 5, and the lower pipeline system includes a mixed liquid inlet pipe 8. The upper pipeline system further includes a regeneration liquid inlet pipe 7, and the lower pipeline system further includes a regeneration liquid outlet pipe 10. The upper pipeline system further includes an upper balance liquid pipe 6, and the lower pipeline system further includes a lower balance liquid pipe 9. Electromagnetic valves 11 are arranged on the mixed liquid outlet pipe 5, the upper balance liquid pipe 6, the regeneration liquid inlet pipe 7, the mixed liquid inlet pipe 8, the lower balance liquid pipe 9 and the regeneration liquid outlet pipe 10, and the control system is electrically connected with the electromagnetic valves 11. One-way valves are arranged on the mixed liquid outlet pipe 5, the regeneration liquid inlet pipe 7, the mixed liquid inlet pipe 8 and the regeneration liquid outlet pipe 10.

[0033] A filler column 12 is arranged between the lower pipeline system and the DEAE-anion high polymer filler chromatographic column 1, and the lower pipeline system, the filler column 12 and the DEAE-anion high polymer filler chromatographic column 1 are sequentially communicated. A stainless steel capillary tube 13 is arranged between the filler column 12 and the DEAE-anion high polymer filler chromatographic column 1 and connected therewith.

[0034] The method for removing pyrocatechol in alpha-matrix based on the above chromatographic device includes the following steps: adjusting a mixed liquid containing alpha-matrix and pyrocatechol to be alkaline, so that the pyrocatechol is reacted to form pyrocatechol salt, and then the mixed liquid is introduced into the DEAE-anion high polymer filler chromatographic column 1 through the lower pipeline system, the pyrocatechol salt in the mixed liquid is adsorbed on the DEAE-anion high polymer filler chromatographic column 1, and the purified alpha-matrix liquid is discharged through the upper pipeline system.

[0035] The separation principle is as follows:

[0036] The phenolic hydroxyl group of hydroquinone has weak acidity, and can lose one or two protons to generate the corresponding phenate anion. The pH of the mixture containing α-arbutin and hydroquinone is adjusted to weak alkaline, and the hydroquinone reacts to generate a negatively charged phenate, which is adsorbed on the DEAE-anion high polymer chromatographic column 1. The interaction between α-arbutin and the anion chromatographic column is small, and α-arbutin is basically not adsorbed and passes through the column body, so that the hydroquinone is separated and removed.

[0037] The separation process of a sample is as follows:

[0038] The sample solution is composed of α-arbutin: 113.5 g / L, hydroquinone: 3.85 g / L, and water as the solvent.

[0039] Chromatographic packing: DEAE-anion high polymer packing (produced by Shanghai Yuji Biotechnology Co., Ltd.), particle size: 50-80 μm.

[0040] Column size: diameter 40 mm x length 500 mm, column volume: 0.628 L.

[0041] UV detection: 280 nm, separation flow rate: 0.5 BV / h.

[0042] Separation mobile phase: equilibration liquid: pure water.

[0043] Regeneration liquid: 80% (volume fraction) ethanol solution.

[0044] Sample injection: 1 L volume at a flow rate of 0.5 BV / h, the control system is set to automatically open the electromagnetic valves 11 on the mixed liquid inlet pipe 8 and the upper equilibration liquid pipe 6. When the UV 280 nm baseline starts to rise, the control system automatically closes the electromagnetic valve 11 on the upper equilibration liquid pipe 6 and opens the electromagnetic valve 11 on the mixed liquid outlet pipe 5 to collect the purified α-arbutin liquid. After the preset volume of sample injection is completed, the control system automatically opens the electromagnetic valve 11 on the lower equilibration liquid pipe 9. When the UV absorption peak drops to the baseline, the electromagnetic valve 11 on the mixed liquid outlet pipe 5 is closed. After the collection of the α-arbutin liquid is completed, the control system automatically opens the regeneration chromatographic column mode, opens the electromagnetic valves 11 on the regeneration liquid inlet pipe 7 and the lower equilibration liquid pipe 9, and when the UV baseline rises, the electromagnetic valve 11 on the lower equilibration liquid pipe 9 is closed, and the electromagnetic valve 11 on the regeneration liquid outlet pipe 10 is opened to collect the regeneration liquid. The chromatographic column is washed with 5 times the column volume of the regeneration liquid, then the electromagnetic valve 11 on the upper equilibration liquid pipe 6 is opened, and the electromagnetic valve 11 on the regeneration liquid outlet pipe 10 is closed by the time relay 4 for a delay of 20 minutes, and the electromagnetic valve 11 on the lower equilibration liquid pipe 9 is opened. The column is washed with 5 times the column volume of the equilibration liquid, and the regeneration of the chromatographic column is completed, and the next separation can be performed.

[0045] The high performance liquid chromatography (HPLC) analysis diagram is as follows: Figure 1The volume of the collected α-melitoside liquid was 2.3 L, containing 109 g of α-melitoside, with a recovery rate of 96%, a purity of 99.7%, and no impurity hydroquinone detected.

[0046] The above description of the embodiments is to enable those skilled in the art to understand and use the invention. It is obvious to those skilled in the art that various modifications can be made to the embodiments and the general principles described herein can be applied to other embodiments without departing from the scope of the invention. Therefore, the invention is not limited to the above embodiments, and any improvements and modifications made by those skilled in the art based on the disclosure of the invention without departing from the scope of the invention should be within the scope of the protection of the invention.

Claims

1. A method for removing p-benzoquinone in alpha-matrix, based on a chromatographic device, characterized in that, the chromatographic device comprises a DEAE-anion high molecular polymer packing column (1) arranged vertically at the bottom end of the device, a detector (2) arranged above the DEAE-anion high molecular polymer packing column (1) and connected with the DEAE-anion high molecular polymer packing column (1), an upper pipeline system arranged above the detector (2) and connected with the detector (2), a lower pipeline system arranged below the DEAE-anion high molecular polymer packing column (1) and connected with the DEAE-anion high molecular polymer packing column (1), and a control system matched with the upper pipeline system and the lower pipeline system, respectively, the control system comprising a controller (3) and a time relay (4); the upper pipeline system comprises a mixed liquid outlet pipe (5) and a regeneration liquid inlet pipe (7), and the lower pipeline system comprises a mixed liquid inlet pipe (8) and a regeneration liquid outlet pipe (10); the method is as follows: adjusting a mixed liquid containing alpha-matrix and p-benzoquinone to be alkaline, so that the p-benzoquinone is reacted to form p-benzoquinone salt, and then the mixed liquid is introduced into the DEAE-anion high molecular polymer packing column (1) through the lower pipeline system, the p-benzoquinone salt in the mixed liquid is adsorbed on the DEAE-anion high molecular polymer packing column (1), and the purified alpha-matrix liquid is discharged through the upper pipeline system.

2. The method of removing p-benzoquinone from a- arbutin according to claim 1, characterized in that, the detector (2) is a UV detector.

3. The method of removing p-benzoquinone from a- arbutin according to claim 1, characterized in that, the upper pipeline system further comprises an upper balance liquid pipeline (6), and the lower pipeline system further comprises a lower balance liquid pipeline (9).

4. The method of removing p-benzoquinone from a- arbutin according to claim 3, characterized in that, electromagnetic valves (11) are arranged on the mixed liquid outlet pipe (5), the upper balance liquid pipeline (6), the regeneration liquid inlet pipe (7), the mixed liquid inlet pipe (8), the lower balance liquid pipeline (9), and the regeneration liquid outlet pipe (10), and the control system is electrically connected with the electromagnetic valves (11).

5. The method of removing p-benzoquinone from a- arbutin according to claim 3, wherein, one-way valves are arranged on the mixed liquid outlet pipe (5), the regeneration liquid inlet pipe (7), the mixed liquid inlet pipe (8), and the regeneration liquid outlet pipe (10).

6. The method of removing p-benzoquinone from a- arbutin according to claim 1, wherein, a packing column (12) is arranged between the lower pipeline system and the DEAE-anion high molecular polymer packing column (1), and the lower pipeline system, the packing column (12), and the DEAE-anion high molecular polymer packing column (1) are connected in sequence.

7. The method of removing p-benzoquinone from a- arbutin according to claim 6, characterized in that, a stainless steel capillary tube (13) is arranged between the packing column (12) and the DEAE-anion high molecular polymer packing column (1), and the packing column (12) and the DEAE-anion high molecular polymer packing column (1) are connected through the stainless steel capillary tube (13).

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