A method for removing salt in the production of sodium mannose phosphate

Abstract

The application belongs to the field of biotechnology separation, and discloses a desalination method in the production process of sodium mannose phosphate, which comprises the following steps: centrifugal ultrafiltration is performed on the sodium mannose phosphate reaction solution obtained through enzyme catalysis reaction, so that the membrane-passing clear liquid is obtained; and the membrane-passing clear liquid is subjected to desalination through electrodialysis, so that the sodium mannose phosphate product is obtained. The method is suitable for the removal of phosphate, hypophosphite and other ions with small molecular weight, and the method is simple in operation, efficient in desalination and green in environmental protection.

Description

Technical Field

[0001] This invention belongs to the field of bioseparation technology, specifically relating to a desalination method in the production process of sodium mannose phosphate. Background Technology

[0002] Sodium mannose phosphate, as a cosmetic active ingredient, has a variety of effects such as anti-aging, anti-wrinkle, and whitening. In particular, it is closely related to cell autophagy and metabolism. Therefore, sodium mannose phosphate has received increasing attention and application in the cosmetic industry.

[0003] Sodium mannose phosphate is generally produced by chemical synthesis or bio-enzymatic catalysis. During the production process, the product system contains some unreacted hexoses, as well as large amounts of orthophosphates, metaphosphates, and polyphosphates. Due to their unique physicochemical properties, phosphates are the most significant destabilizing factor in the product, causing phenomena such as turbidity during concentration, precipitation at low temperatures, and sedimentation during long-term storage, which greatly affects production and application.

[0004] Currently, the main desalination methods used are precipitation and ion exchange. Precipitation uses metal ions such as calcium and magnesium ions that can form insoluble compounds with phosphate groups. Phosphate is removed by precipitation and filtration, and anion exchange columns remove introduced anions such as chloride. Because mannose phosphate itself also has phosphate groups, precipitation has poor selectivity, leading to partial precipitation and significant product loss. Ion exchange involves diluting the product and then using cation exchange columns and anion exchange columns to replace phosphate ions (metaphosphate ions) and sodium ions in the system. This method is limited by the capacity of the ion exchange resin, requiring pre-dilution and subsequent concentration after desalination, making the entire process complex. Ion exchange resins also require high selectivity for mannose phosphate and phosphates; furthermore, the large amounts of eluent and waste resin increase environmental pressure.

[0005] Therefore, to address the aforementioned issues, it is necessary to develop a simple, efficient, green, and low-cost desalination method suitable for removing various small-molecule ionic impurities such as phosphates and nonionic impurities such as mannose during the production of sodium mannose phosphate, so as to better serve the production and application of sodium mannose phosphate. Summary of the Invention

[0006] (I) Purpose of the Invention

[0007] The purpose of this invention is to provide a desalination method in the production process of sodium mannose phosphate. This method uses electrodialysis to effectively remove phosphates, hypophosphites and other small molecular weight ions doped into the products during the production process of sodium mannose phosphate. The method is simple to operate, highly efficient in desalination, environmentally friendly and low in desalination cost.

[0008] (II) Technical Solution

[0009] To address the above problems, this invention provides a desalination method during the production of sodium mannose phosphate, comprising:

[0010] The sodium mannose phosphate reaction solution obtained by the enzyme-catalyzed reaction was centrifuged and ultrafiltered to obtain a membrane-filtered clear solution.

[0011] The membrane-passed solution was desalted by electrodialysis to obtain sodium mannose phosphate product.

[0012] Furthermore, the ion exchange membrane for the electrodialysis desalination method employs a homogeneous cation membrane and an anion membrane, with the homogeneous cation membrane and the homogeneous anion membrane forming a membrane module in pairs.

[0013] Furthermore, both the cation homogeneous membrane and the anion homogeneous membrane are based on styrene-divinylbenzene.

[0014] Furthermore, the voltage of the membrane array is 0.2 to 0.8 V per array.

[0015] Furthermore, in the electrodialysis desalination process, the membrane supernatant is used as the dilute solution in the electrodialysis chamber, deionized water is used as the concentrate solution in the electrodialysis chamber, and 0.3M PBS is used as the protective solution.

[0016] Furthermore, the concentration of sodium mannose phosphate in the membrane-passing solution is 50–120 g / L.

[0017] Furthermore, the dilute solution is collected as sodium mannose phosphate product, and the concentrated solution and protective solution are mixed and then calcium chloride is added for dephosphorization treatment.

[0018] Furthermore, in the electrodialysis desalination process, the flow rate of the supernatant is 1–3 L / min / m. 2 .

[0019] Furthermore, the desalination time of the electrodialysis method is 30–180 min.

[0020] (III) Beneficial Effects

[0021] The above-mentioned technical solution of the present invention has the following beneficial technical effects: The present invention provides a desalination method for the production process of sodium mannose phosphate, applicable to the desalination of sodium mannose phosphate liquid obtained by bio-enzyme catalysis. The method involves centrifuging and ultrafiltration of the sodium mannose phosphate reaction solution obtained by the catalytic reaction, followed by desalination using electrodialysis. This method is simple, requiring only one electrodialysis step. During the electrodialysis control process, by controlling key process parameters such as the sodium mannose phosphate content range, membrane voltage, and electrodialysis time, desalination is highly efficient, with M6P loss not exceeding 10%. In addition to various phosphates, it can also remove other small molecular weight ionic impurities. Compared to the above-mentioned precipitation and ion exchange methods, the process of the present invention only produces a small amount of phosphorus-containing wastewater, which can be easily removed using calcium chloride during operation. It is green and environmentally friendly, with low desalination costs. Detailed Implementation

[0022] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments. It should be understood that these descriptions are merely exemplary and not intended to limit the scope of the invention. Furthermore, descriptions of well-known structures and technologies are omitted in the following description to avoid unnecessarily obscuring the concepts of the invention.

[0023] I. Testing Instruments and Methods

[0024] The instruments used in this invention include:

[0025] Electrodialysis unit: EX-3BT or EX-4S;

[0026] Water: All water used in the embodiments shall meet the Class I water requirements in GB / T 6682.

[0027] In this invention, the term "water" refers to ultrapure water unless otherwise specified. The terms "sodium mannose phosphate," "mannose phosphate," "sodium mannose-6-phosphate," "sodium mannose-6-phosphate," "mannose-6-phosphate," "sodium mannose phosphate," "mannose-6-phosphate," and "M6P" have the same meaning and can be used interchangeably. The specific technical solution of this invention is as follows:

[0028] This invention provides a desalination method in the production process of sodium mannose phosphate, comprising the following steps:

[0029] S1, the sodium mannose phosphate reaction solution obtained from the catalytic reaction is centrifuged and ultrafiltered to obtain a membrane-passed clear solution. This invention is applicable to the treatment of sodium mannose phosphate prepared by enzyme catalytic reaction. The content of sodium mannose phosphate that can be treated by electrodialysis is in the range of 50-120 g / L, that is, the concentration of sodium mannose phosphate in the membrane-passed clear solution is 50-120 g / L. Generally, if the content of sodium mannose phosphate in the membrane-passed clear solution is less than 50 g / L, it indicates that the reaction rate of the enzyme catalytic reaction is very low, and the membrane-passed clear solution contains a large amount of phosphates such as sodium polyphosphate and sodium polyphosphate with large molecular weights. Their large molecular weight cannot pass through the homogeneous membrane, resulting in the inability of electrodialysis to remove salt. At the same time, when the concentration of sodium mannose phosphate exceeds 120 g / L, it is easy to precipitate due to solubility, which affects the efficiency of electrodialysis. Therefore, the content of sodium mannose phosphate in the membrane-passed clear solution obtained from the enzyme catalytic reaction is finally determined to be 50-120 g / L.

[0030] S2, the clarified solution is desalted using electrodialysis to obtain sodium mannose phosphate product. The electrodialysis parameters are as follows: the membrane voltage for desalination is 0.2–0.8 V / membrane. This voltage range significantly reduces sodium mannose phosphate loss and improves the desalination rate. Too low a voltage significantly reduces desalination efficiency, prolongs the process, and significantly increases the loss rate of sodium mannose phosphate. Furthermore, due to the homogeneous membrane material, its voltage tolerance is limited; voltages exceeding 0.8 V will cause significant damage to the homogeneous membrane. The desalination time is 30–180 min, preferably 30–60 min. Experiments show that as the electrodialysis time increases, the material conductivity gradually decreases, but the rate of decrease gradually decreases, and the decrease is not significant after 60 min. The content and yield of sodium mannose phosphate decrease slowly, and the loss increases significantly after 60 min. The phosphate content and desalination rate change significantly in the first 30 min, but the change is not significant after 30 min. Therefore, the preferred time is 30–60 min. The ion exchange membrane used in the electrodialysis desalination process employs both cation-based and anion-based homogeneous membranes. Both the cation-based and anion-based homogeneous membranes are based on styrene-divinylbenzene. During the electrodialysis desalination process, the membrane supernatant is used as the dilute solution, deionized water as the concentrate solution, and 0.3M PBS as a protective solution to prevent damage to the module from excessive voltage or current. Other PBS concentrations can also be used. During the electrodialysis desalination process, the flow rate of the membrane supernatant is 1–3 L / min / m. 2 . II. Specific Implementation Methods

[0032] Example 1

[0033] In the production of sodium mannose phosphate, the reaction solution after the enzyme-catalyzed reaction was centrifuged and ultrafiltered, and an appropriate amount of the supernatant was collected. The sodium mannose phosphate content was determined to be 82.38 g / L using a K-MANGL kit for ATP removal, and the phosphate content was determined to be 23.25 g / L using the molybdenum blue method. The conductivity was 24.16 mS / cm. 1000 mL of the supernatant was added to the dilute chamber of the electrodialysis machine; 1000 mL of deionized water was added to the concentrate chamber, and 1000 mL of 0.3 M PBS was added to the electrode water chamber as a protective solution. Electrodialysis parameters: Electrodialysis machine model, EX-3BT; Electrodialysis membranes: styrene-divinylphenyl cation and anion membranes (homogeneous membranes); Homogeneous membrane stacks: 10 groups; Voltage per group: 0.5 V; Material flow rate: 3 L / min. Electrodialysis was performed for 180 min, with samples taken every 30 min to measure conductivity, sodium mannose phosphate, and phosphate content. The results are shown in Table 1. It can be seen that as the electrodialysis time increased, the conductivity of the material gradually decreased, but the rate of decrease gradually slowed down, and the decrease became insignificant after 90 min. Simultaneously, the content and yield of sodium mannose phosphate also decreased slowly, with a significant increase in loss after 90 min. The phosphate content and desalination rate showed significant changes in the first 60 min, but the changes were not substantial after 60 min.

[0034] Table 1. Measurement results of Example 1

[0035]

[0036]

[0037] Example 2

[0038] Take 1000 mL of the same batch of supernatant as in Example 1 (first batch sample) and add it to the dilute chamber of the electrodialysis apparatus. Operate according to the procedure in Example 1, except that the membrane voltage for each group is set to 0.2 V. Samples are taken every 30 minutes to measure conductivity, sodium mannose phosphate, and phosphate content. The results of 60 minutes of electrodialysis are shown in Table 2. Comparing the results of Example 2 and Example 1, it can be seen that in Example 1, the sodium mannose phosphate yield was 92.20% and the desalination rate was 85.21% after 60 minutes, while in Example 2, the sodium mannose phosphate yield was 96.09% and the desalination rate was 76.11% after 60 minutes. Therefore, reducing the voltage can slightly reduce the loss of sodium mannose phosphate, but it is significantly detrimental to the desalination efficiency.

[0039] Table 2 Measurement results of the examples

[0040]

[0041] Example 3

[0042] Take 1000 mL of the same batch of membrane-exposed solution as in Example 1, and operate according to the procedure in Example 1, except that the membrane voltage for each group is set to 0.8 V. Samples are taken every 30 minutes to test conductivity, sodium mannose phosphate, and phosphate content. The results of 60 minutes of electrodialysis are shown in Table 3. Comparison with the results of Examples 1 and 2 shows that, within the tolerance range of the electrodialysis membrane, increasing the membrane voltage can improve the desalination rate, but the yield of sodium mannose phosphate decreases.

[0043] Table 3. Measurement results of Example 3

[0044]

[0045] Example 4

[0046] A batch of sodium mannose phosphate reaction solution (different from that in Example 1, the second batch sample) was used. The sodium mannose phosphate content was determined to be 85.06 g / L using a K-MANGL kit to remove ATP, and the phosphate content was determined to be 23.25 g / L using the molybdenum blue method. The conductivity was 24.06 mS / cm. 1000 mL of the supernatant was added to the dilute chamber of the electrodialysis apparatus; 1000 mL of deionized water was added to the concentrate chamber, and 1000 mL of 0.3 M PBS was added to the electrode water chamber as a preservative solution. The electrodialysis parameters were the same as in Example 1. After 60 min of electrodialysis, samples were taken every 30 min to measure the conductivity, sodium mannose phosphate, and phosphate content. The results are shown in Table 4. The analytical results are also shown in Table 4.

[0047] Table 4. Measurement results of Example 4

[0048]

[0049] Example 5

[0050] The supernatant from the third batch of samples (different from those in Examples 1-3) was used. The sodium mannose phosphate reaction solution was tested using a K-MANGL kit to remove ATP, and the sodium mannose phosphate content was found to be 81.55 g / L. The phosphate content was found to be 23.25 g / L using the molybdenum blue method, and the conductivity was 24.24 mS / cm. 1000 mL of the supernatant was added to the dilute chamber of the electrodialysis apparatus; 1000 mL of deionized water was added to the concentrate chamber, and 1000 mL of 0.3 M PBS was added to the electrode water chamber as a preservative solution. The electrodialysis parameters were the same as in Example 1. After 60 min of electrodialysis, samples were taken every 30 min to measure the conductivity, sodium mannose phosphate, and phosphate content. The results are shown in Table 5. Comparing the results of Examples 1, 4, and 5, it can be seen that for different batches of membrane-filtered clarified liquid in the production process of sodium mannose phosphate, the changes in the content and yield of sodium mannose phosphate, and the content and yield of phosphate, are not significantly different after treatment with the desalination method of the present invention.

[0051] Table 5. Measurement results of Example 5

[0052]

[0053] Examples 6-8

[0054] Three 1000 mL portions of the membrane-passing supernatant from the same batch as in Example 1 (first batch sample) were added to the dilute chamber of the electrodialysis apparatus as dilute solution. Separately, 1000 mL of deionized water was added to the concentrate chamber as concentrate solution, and 1000 mL of 0.3 M PBS was added to the electrode water chamber as a protective solution. Parallel experiments were conducted under the electrodialysis conditions of Example 1. The experimental results after 60 min of electrodialysis and the comparison with Example 1 are shown in Table 6. The RSD results show that the electrodialysis desalination method described in this invention has good repeatability.

[0055] Table 6 shows the measurement results of Examples 6-8.

[0056]

[0057]

[0058] Examples 9-12

[0059] Take the same batch of membrane-passing solution (original membrane-passing solution) as in Example 1: 1) Concentrate the original membrane-passing solution to remove 1 / 3 of the water volume, and use it as Example 9 for later use; 2) Take an appropriate amount of the above concentrated solution (Example 9) and mix it evenly with the original membrane-passing solution at a volume ratio of 1:1, and use it as Example 10 for later use; 3) Take an appropriate amount of the above concentrated solution (Example 9) and mix it evenly with deionized water at a volume ratio of 1:1, and use it as Example 11 for later use; 4) Mix the original membrane-passing solution and deionized water evenly at a volume ratio of 1:1, and use it as Example 12 for later use. Take 1000 mL of each of the above four materials, measure their conductivity, sodium mannose phosphate, and phosphate content, and add them to the dilute solution tank of the electrodialysis instrument as dilute solution; take another 1000 mL of deionized water as concentrated solution and add it to the concentrated solution tank, and add 1000 mL of 0.3M PBS as protective solution to the electrode water tank. The electrodialysis parameters are the same as in Example 1. After 60 minutes of electrodialysis, the conductivity, sodium mannose phosphate, and phosphate content were measured. The test results for each batch and the comparison with Example 1 are shown in Table 7.

[0060] Table 7. Measurement results of Examples 9-12

[0061]

[0062] Example 13

[0063] Take 20.0 L of the same batch of membrane-passing supernatant as in Example 1 and add it to the dilute chamber of the electrodialysis apparatus; separately, take 20.0 L of deionized water and add it to the concentrate chamber; and add 20.0 L of 0.3M PBS as the protective solution to the electrode water chamber. Electrodialysis parameters: electrodialysis apparatus model, EX-4S; homogeneous membrane stack number of groups: 50 groups; set the voltage of each group of membranes to 0.8V, and the material flow rate to 9 L / min. After 60 min of electrodialysis, samples were taken every 30 min to detect conductivity, sodium mannose phosphate, and phosphate content. The results are shown in Table 8. It can be seen that the desalination process described in this invention can be used for industrial-scale production, and the desalination rate and sodium mannose phosphate yield after scale-up are not significantly different from those in the small-scale test.

[0064] Table 8. Measurement results of Example 13

[0065]

[0066] Comparative Example 1

[0067] 1000 mL of the same batch of membrane-passing solution as described in Example 1 was added to the dilute chamber of the electrodialysis instrument. 1000 mL of deionized water was added to the concentrate chamber, and 1000 mL of 0.3M PBS was added to the electrode water chamber as a protective solution. Electrodialysis parameters: Electrodialysis instrument model, EX-3BT; alloy membrane, 10 membrane groups; voltage per group of membranes set to 0.5V, material flow rate 3L / min. Electrodialysis was performed for 60 min, and samples were taken every 30 min to measure conductivity, sodium mannose phosphate, and phosphate content. The results are shown in Table 9. It can be seen that the homogeneous membrane based on styrene-divinylbenzene has a permeable molecular weight of 300 Da, while the alloy membrane has a permeable molecular weight of 400 Da. The alloy membrane has a higher permeable molecular weight than the homogeneous membrane, resulting in higher ion permeation efficiency in the material. Therefore, after electrodialysis using an alloy membrane, the desalination rate was significantly improved compared to the results of Example 1; however, correspondingly, since the molecular weight of mannose phosphate ions is 281, they can easily pass through the alloy membrane, so the yield of sodium mannose phosphate also decreased significantly.

[0068] Table 9 shows the results of the determination in Comparative Example 1.

[0069]

[0070] Comparative Example 2

[0071] Based on the phosphate content (23.25 g / L) of the membrane-passing solution described in Example 1, 23.25 g each of sodium hexametaphosphate (a), sodium trimetaphosphate (b), sodium metaphosphate (c), and trisodium phosphate (d), and 23.25 g of a mixed metaphosphate (e) with a mass ratio of sodium hexametaphosphate, sodium trimetaphosphate, and sodium metaphosphate of 1:1:1 were weighed and placed in 1000 mL of deionized water and stirred until completely dissolved to obtain five different phosphate solutions. These phosphate solutions were added to the dilute chamber of the electrodialysis apparatus as dilute solutions, and 1000 mL of deionized water was added to the concentrate chamber as concentrate. 1000 mL of 0.3 M PBS was added to the electrode water chamber as a protective solution. Electrodialysis was performed with the same parameters as in Example 1. After 60 min of electrodialysis, the conductivity and phosphate content were measured and are shown in Table 10. It is evident that, due to the molecular weight distribution of homogeneous electrodialysis membranes, larger molecular weight phosphates are less likely to permeate, and this also affects the permeation rate of smaller molecular weight phosphates. Therefore, when the sodium mannose phosphate content in the membrane supernatant is too low, a higher residual amount of large molecular weight phosphates will occur, which will affect the desalination rate of the electrodialysis process.

[0072] Table 10 shows the results of the comparative example 2.

[0073]

[0074] This invention provides a desalination method for the preparation and production of sodium mannose phosphate. This method is simple, efficient, and environmentally friendly, better serving the production and application of sodium mannose phosphate. By controlling key process parameters such as membrane stack voltage, electrodialysis time, and ion exchange membrane in the electrodialysis process, the yield and desalination rate of sodium mannose phosphate are significantly improved, with the yield reaching over 90%. It also achieves highly efficient desalination, with M6P loss not exceeding 10%. In addition to various phosphates, it can remove other small molecular weight ionic impurities. Furthermore, it is environmentally friendly. Compared to the two existing desalination methods, the electrodialysis process of this invention produces only a small amount of phosphorus-containing wastewater, which can be easily removed with calcium chloride. This method is environmentally friendly and has low desalination costs.

[0075] It should be understood that the specific embodiments described above are merely illustrative or explanatory of the principles of the invention and do not constitute a limitation thereof. Therefore, any modifications, equivalent substitutions, improvements, etc., made without departing from the spirit and scope of the invention should be included within the protection scope of the invention. Furthermore, the appended claims are intended to cover all variations and modifications falling within the scope and boundaries of the appended claims, or equivalent forms of such scope and boundaries.

[0076] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A desalination method in the production process of sodium mannose phosphate, characterized in that, include: The sodium mannose phosphate reaction solution obtained by the enzyme-catalyzed reaction was centrifuged and ultrafiltered to obtain a membrane-filtered clear solution. The membrane-passed solution was desalted by electrodialysis to obtain sodium mannose phosphate product.

2. The desalination method in the production process of sodium mannose phosphate according to claim 1, characterized in that, The ion exchange membranes used in the electrodialysis desalination method employ both cation homogeneous membranes and anion homogeneous membranes, with the cation homogeneous membranes and anion homogeneous membranes forming a membrane module in pairs.

3. The desalination method in the production process of sodium mannose phosphate according to claim 2, characterized in that, Both the cationic homogeneous membrane and the anionic homogeneous membrane are based on styrene-divinylbenzene.

4. The desalination method in the production process of sodium mannose phosphate according to claim 2, characterized in that, The voltage of the membrane array is 0.2 to 0.8 V per array.

5. The desalination method in the production process of sodium mannose phosphate according to claim 1, characterized in that, In the electrodialysis desalination process, the membrane-passed clear solution is used as the dilute solution in the electrodialysis chamber, and deionized water is used as the concentrated solution in the electrodialysis chamber.

6. The desalination method in the production process of sodium mannose phosphate according to claim 1, characterized in that, The concentration of sodium mannose phosphate in the membrane-passing solution is 50–120 g / L.

7. The desalination method in the production process of sodium mannose phosphate according to claim 1, characterized in that, In the electrodialysis desalination process, the flow rate of the supernatant is 1–3 L / min / m. 2 .

8. The desalination method in the production process of sodium mannose phosphate according to claim 1, characterized in that, The desalination time for the electrodialysis method is 30–180 min.

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