PES membrane modification method, PES membrane and its application in endotoxin removal

Abstract

The present invention discloses a PES membrane modification method, a PES membrane, and its application in endotoxin removal. The modification method, based on membrane surface treatment and chemical modification, significantly increases the number of amino groups on the PES membrane surface through a Schiff base / Michael addition reaction, thereby increasing the positive surface charge of the PES membrane. The method offers advantages such as high efficiency and energy saving, ease of operation, easy process control, and ease of scale-up and integration. The modified PES membrane positively modifies the original PES membrane surface, significantly increasing its positive surface charge, enabling electrostatic adsorption to remove endotoxins present in intravenous medications. Furthermore, the modified membrane introduces a large number of amino or hydroxyl groups, which readily bind to lipopolysaccharide molecules, onto the surface. These groups can be used to immobilize endotoxins on the membrane surface through molecular forces for removal, thereby ensuring that the modified membrane can remove endotoxins over a wider pH range.

Description

Technical Field

[0001] The present invention relates to the technical field of membrane modification, in particular to a PES membrane modification method, a PES membrane and application thereof in removing endotoxins. Background Art

[0002] Endotoxins, also known as lipopolysaccharides (LPS) or pyrogens, are core components of the outer cell wall of most Gram-negative bacteria. They carry a negative charge due to the presence of two phosphorylated groups in their backbone. Endotoxins have potent biological effects even at very low concentrations in the human body. Reported potent biological reactions to endotoxins include hypotension, vomiting, intense inflammation, fever, respiratory distress, intravascular coagulation, sepsis, and even death. Endotoxins are the most common pyrogenic contaminant in biological products. Even trace amounts of endotoxin in a sample can cause high fever, diarrhea, vasodilation, and even fainting or death. In the United States, over 750,000 patients suffer from sepsis and septic shock annually, with a mortality rate of 25-80%. In two separate incidents in Brazil, 33 infants and 35 newborns died after intravenous medications were diluted with endotoxin-contaminated distilled water. Therefore, developing membranes for the robust removal of endotoxins from injectable solutions is crucial to minimizing side effects such as inflammatory responses in patients. Because endotoxins have an amphiphilic supramolecular structure, their biological activity can only be destroyed by heating at 160°C for 2 to 4 hours, or by boiling with a strong base, strong acid, or strong oxidant for 30 minutes. Currently, there is no universally applicable technology for endotoxin removal. Common methods include activated carbon adsorption, chemical degradation, phase separation, chromatography, and tangential flow membrane filtration. However, these methods suffer from poor biocompatibility, low precision, complex operation, poor economic efficiency, and the tendency to easily remove useful substances.

[0003] Polyethersulfone (PES) membranes are made of melt-bonded microfibers. They are a hydrophilic polymeric membrane used for deep filtration. They feature a three-dimensional structure and are characterized by stable physical and chemical properties and excellent drug compatibility. They offer a wide range of pore sizes, high porosity, a large dirt-holding capacity, backflushing and high-temperature sterilization capabilities, and excellent pressure resistance, enabling the removal of insoluble microparticles. Their excellent hydrophilicity and unique microstructure allow for rapid and complete wetting, enabling high and stable liquid flow rates while maintaining high particle interception efficiency. They are stable to common acids, bases, aliphatic hydrocarbons, oils, and alcohols, and are suitable for physical and chemical sterilization, such as high-pressure steam, gamma-ray, and ethylene oxide sterilization. They possess a high bubble point and excellent membrane integrity. Polyethersulfone membranes have very low protein adsorption, ensuring the stability of filtered drug solutions. They are suitable for use in disposable precision filtration infusion sets, extension tubing, and drug filter cartridges. Polyethersulfone is often used to make hemodialyzers, blood concentrators, artificial lungs, etc. due to its excellent mechanical properties, stable wetting strength and blood compatibility. Currently, most of the liquid drug filtration membranes produced abroad are made of polyethersulfone materials.

[0004] Therefore, developing a membrane separation technology that can simply, quickly and accurately remove endotoxins present during intravenous infusion is crucial to improving the safety of infusion, reducing separation costs and increasing the market competitiveness of products. Summary of the Invention

[0005] The present invention provides a PES membrane modification method, a PES membrane, and its application in removing endotoxins. The PES membrane modification is used to remove endotoxins in intravenous infusion drugs, thereby solving at least one of the above-mentioned defects in the prior art.

[0006] In view of this, the solution of the present invention is:

[0007] The first aspect of the present invention provides a method for modifying a PES membrane, comprising the following steps:

[0008] S1. A condensing agent is added to a mixed system of a Tris buffer solution, a precursor compound and an amino compound, and a condensation reaction occurs under stirring to obtain a mixed solution; the precursor compound is a compound that can react with an amino group;

[0009] S2. Immerse the PES membrane in the mixed solution obtained in step S1, take it out after reaction, and wash it with water and ethanol in sequence to obtain a modified PES membrane.

[0010] Furthermore, the pH of the Tris buffer solution is 7-9.

[0011] Furthermore, the precursor compound is selected from one of dopamine, O-carboxymethyl chitosan, N-carboxymethyl chitosan, and N,O-carboxymethyl chitosan; and in the mixed system, the concentration of the precursor compound is 0.01-3 wt%.

[0012] Furthermore, the amino compound is selected from one of serine, polyethyleneimine, propylenediamine, ethylenediamine, butylenediamine, tris(2-aminoethyl)amine, and diethylenetriamine; and in the mixed system, the concentration of the amino compound is 0.1-4 wt%.

[0013] Furthermore, the condensing agent is selected from one of dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI); and the concentration of the condensing agent is 0-1 wt%.

[0014] Preferably, in step S1: the condensation reaction temperature is 0-8°C, the reaction time is 0.05-6h; and / or; the stirring speed is 0-1000rpm.

[0015] Furthermore, in step S2, the reaction temperature is 5-75° C., and the reaction time is 0.5-5 days.

[0016] Furthermore, in step S2, the washing times are 3-6 times.

[0017] Preferably, the PES membrane is one of the commercially available PES filter membranes, including brands such as Pall, Cobetter, Nova, Longjin, etc., with a pore size of 0.1-5 μm and a diameter of 3-20 cm.

[0018] The second aspect of the present invention provides a PES membrane obtained by the PES membrane modification method of the first aspect, wherein the surface of the PES membrane is modified with amino groups.

[0019] The third aspect of the present invention provides an application of the PES membrane described in the second aspect, wherein the application is to remove endotoxins from intravenous infusion drugs.

[0020] The fourth aspect of the present invention provides a method for removing endotoxins from intravenous infusion drugs, comprising the step of filtering the intravenous infusion drugs using the PES membrane described in the second aspect.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] 1. The present invention utilizes membrane surface treatment methods and chemical modification techniques to significantly increase the number of amino groups on the PES membrane surface through a Schiff base / Michael addition reaction, thereby increasing the positive surface charge of the PES membrane. This method offers advantages such as high efficiency and energy saving, ease of operation, easy process control, and ease of scale-up and integration. Through the targeted selection of chemical modification methods, the original PES membrane surface is electropositively modified, significantly increasing the positive surface charge, which can be used for electrostatic adsorption to remove endotoxins present in intravenous medications. Furthermore, a large number of amino or hydroxyl groups, which readily bind to lipopolysaccharide molecules, are introduced onto the surface, allowing the endotoxin to be fixed to the membrane surface through molecular forces for removal, thereby ensuring that the modified membrane can remove endotoxins over a wider pH range.

[0023] 2. The chemical raw materials used in the modification method provided by the present invention have good biocompatibility and can avoid other adverse effects on the infusion preparation.

[0024] 3. The PES membrane obtained by the modification method of the present invention is highly efficient in removing endotoxins from intravenous drugs during filtration and has significant application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a scanning electron microscope image of the modified PES membrane of the present invention.

[0026] Figure 2 Schematic diagram of the surface molecular structure of the modified PES membrane of the present invention.

[0027] Figure 3 Surface Zeta potential diagram of the modified PES membrane in Examples 1-5 of the present invention. DETAILED DESCRIPTION

[0028] The technical solutions of the present invention will be clearly and completely described below in conjunction with preferred embodiments. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.

[0029] The following are preferred implementation examples of modified separation membranes. Except for the methods specifically defined herein, the methods involved in the process are those known to those skilled in the art, and the reagents used are all commercially available standard products.

[0030] In one embodiment, a method for modifying a PES membrane is provided, comprising:

[0031] S1. A condensing agent is added to a mixed system of a Tris buffer solution, a precursor compound and an amino compound, and a condensation reaction occurs under stirring to obtain a mixed solution; the precursor compound is a compound that can react with an amino group;

[0032] S2. Immerse the PES membrane in the mixed solution obtained in step S1, take it out after reaction, and wash it with water and ethanol in sequence to obtain a modified PES membrane.

[0033] As described above, in step S1, the precursor compound and the amino-containing compound undergo a Schiff base / Michael addition reaction, which greatly increases the amino groups on the surface of the PES membrane and improves the surface positive charge of the PES membrane.

[0034] In some embodiments, the precursor compound is selected from dopamine, O-carboxymethyl chitosan, N-carboxymethyl chitosan, and N,O-carboxymethyl chitosan; in the mixed system, the concentration of the precursor compound is 0.01-3 wt %. The amino-containing compound is selected from serine, polyethyleneimine, propylenediamine, ethylenediamine, butylenediamine, tris(2-aminoethyl)amine, and diethylenetriamine; in the mixed system, the concentration of the amino-containing compound is 0.01-4 wt %. The condensing agent is selected from dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI); in the mixed system, the concentration of the condensing agent is 0-1 wt %.

[0035] The precursor compounds are exemplified by O-carboxymethyl chitosan, N-carboxymethyl chitosan, and N,O-carboxymethyl chitosan, with a general formula of R1COOH. The amino compound has a general formula of NH2R2, representing serine, polyethyleneimine, propylenediamine, ethylenediamine, butylenediamine, tris(2-aminoethyl)amine, and diethylenetriamine. The reaction process is as follows:

[0036]

[0037] The precursor compound, taking dopamine as an example, undergoes a Schiff base / Michael addition reaction with an amino compound having the general formula NH2R2, where NH2R2 represents serine, polyethyleneimine, propylenediamine, ethylenediamine, butylenediamine, tris(2-aminoethyl)amine, or diethylenetriamine. The process is as follows:

[0038]

[0039] In the above embodiment, the scanning electron microscopy image of the modified PES membrane is as follows: Figure 1 As shown, the schematic diagram of the molecular structure of the modified PES membrane surface is as follows Figure 2 As shown, it can be seen that a large number of amino groups are introduced into the modified PES membrane.

[0040] In the above-mentioned embodiment, the modified PES membrane can be used to remove endotoxins from intravenous drugs. This can be achieved by first preparing the intravenous drug to a pH of 2-5, then using the modified PES membrane for filtration and adsorption. At this pH, the PES membrane surface carries a positive charge, capable of adsorbing and removing endotoxins. After adsorption, the pH of the intravenous drug is readjusted to the prescribed range without affecting the active pharmaceutical ingredient. Furthermore, endotoxins can be removed within the normal pH range of intravenous drugs. Because the modified membrane surface incorporates a large number of amino and hydroxyl groups that readily bind to lipopolysaccharide molecules, endotoxins can be fixed to the membrane surface through molecular forces for removal, ensuring that the modified membrane can remove endotoxins over a wider pH range.

[0041] Example 1

[0042] This example provides a specific preparation process for a separation membrane for efficient endotoxin removal.

[0043] A tris (hydroxymethyl)aminomethane (Tris) buffer solution at pH 9 was prepared, to which O-carboxymethyl chitosan (3% wt) and tris(2-aminoethyl)amine (4% wt) were added. A condensing agent, DCC (1% wt), was added to the resulting mixture, stirred at 1000 rpm, and reacted at 8°C for 6 hours. A PES membrane (Pall, pore size: 0.2 μm, diameter: 5 cm) was immersed in the mixture and reacted at 75°C for 5 days. The membrane was then removed and washed three times with water and three times with ethanol to obtain the surface-modified PES membrane.

[0044] Example 2

[0045] A tris (hydroxymethyl)aminomethane (Tris) buffer solution was prepared at pH 8, to which dopamine (2% wt) and polyethyleneimine (4% wt) were added. The resulting mixture was then added with EDCI (0.001% wt), stirred at 10 rpm, and reacted at 8°C for 0.5 hours. A PES membrane (brand: Cobetter, pore size: 5 μm, diameter: 20 cm) was immersed in the mixture and reacted at 35°C for 4 days. The membrane was then removed and washed six times with water and then ethanol, yielding a surface-modified PES membrane.

[0046] Example 3

[0047] A tris (hydroxymethyl)aminomethane (Tris) buffer solution was prepared at pH 7, to which N-carboxymethyl chitosan (0.01 wt%) and diethylenetriamine (0.01 w % t) were added. A condensing agent, DIC (0 wt%), was added to the resulting mixture, stirred at 0 rpm, and reacted at 5°C for 5 hours. A PES membrane (brand: Xinxing, pore size: 0.1 μm, diameter: 15 cm) was immersed in the mixture and reacted at 5°C for one day. The membrane was then removed and washed six times with water and then ethanol, yielding a surface-modified PES membrane.

[0048] Example 4

[0049] A tris (hydroxymethyl)aminomethane (Tris) buffer solution (pH 7.5) was prepared, to which N,O-carboxymethyl chitosan (2% wt) and diaminobutyl amine (4% wt) were added. The resulting mixture was then added with EDCI (1% wt), stirred at 800 rpm, and reacted at 0°C for 1 hour. A PES membrane (brand: Longjin, pore size: 5 μm, diameter: 3 cm) was immersed in the mixture and reacted at 25°C for 3 days. The membrane was then removed and washed five times with water and five times with ethanol to obtain the surface-modified PES membrane.

[0050] Example 5

[0051] A Tris buffer solution (pH 8.8) was prepared, to which dopamine (1% wt) and serine (2% wt) were added. A condensing agent, DCC (0.01% wt), was added to the resulting mixture, stirred at 500 rpm, and reacted at 4°C for 3 hours. A PES membrane (brand: Cobetter, pore size: 3 μm, diameter: 8 cm) was immersed in the mixture and reacted at 45°C for 5 days. The membrane was then removed and washed four times with water and four times with ethanol to obtain a surface-modified PES membrane.

[0052] Experimental Example 1: Determination of zeta potential on the surface of modified PES membrane

[0053] The surface zeta potential of the modified PES membranes obtained in Examples 1-5 was measured. Figure 3 As shown, it can be found that the surface potential of the membranes prepared in Examples 1-5 is positive in the pH range of 2-5.

[0054] Experimental Example 2: Verification of the ability of modified PES membrane to remove endotoxins

[0055] 1) Two saline solutions containing 0.50 EU / mL endotoxin were prepared and adjusted to pH 3 and 5, respectively. The modified PES membranes obtained in Examples 1-5 were used to remove endotoxins from the saline solutions. Pumping / gravity delivery was used at a delivery rate of 80 ml / h. The endotoxin removal performance was calculated based on the reduction in endotoxin concentration in the saline solution. The results are shown in Table 1. The separation flux was found to be improved compared to that reported in J. Mater. Chem. B, 2022, 10, 2471-2480.

[0056] Table 1:

[0057]

[0058]

[0059] As can be seen from Table 1, compared to the unmodified blank PES membrane, the endotoxin removal rates of Examples 1-5 after modification were significantly improved. This is due to the positive electrophoresis of the modified PES membrane surface at pH 4, resulting in a strong endotoxin adsorption capacity. Furthermore, the relatively strong removal capacity of Examples 2 and 4 may be related to the amount of amino groups introduced and their spatial structure.

[0060] 2) Three finished intravenous injection solutions with pH values ​​of 7, 8, and 9, respectively, were used to remove endotoxins using the modified PES membranes described in Examples 1-5. The process employed a pump / gravity delivery method at a delivery rate of 80 ml / h. The endotoxin removal performance was calculated as the relative percentage reduction of endotoxins in the intravenous injection solutions. The results are shown in Table 2.

[0061] Table 2:

[0062]

[0063] It is not difficult to see from Table 2 that under higher pH conditions, the separation flux is also improved compared to the prior art J.Mater.Chem.B, 2022, 10, 2471-2480. After Examples 1-5 were modified respectively, the endotoxin removal rate was still significantly improved compared to the unmodified PES membrane. The possible reason is that a large number of hydroxyl groups and amino groups are introduced on the surface of the modified PES membrane, which can form hydrogen bonds with endotoxin polysaccharides to improve the removal rate. Examples 2 and 4 have relatively strong removal capabilities, which may be related to the amount of amino groups introduced and the spatial structure.

[0064] Experimental Example 3: Surface stability test of modified PES membrane

[0065] The components of the physiological saline and intravenous injection solutions after endotoxin removal in Examples 1-5 in Experimental Example 2 were analyzed to detect the residual amounts of amino groups and secondary amines in the free components. The results are shown in Table 3.

[0066] Table 3:

[0067]

[0068]

[0069] As shown in Table 3, the modified PES membrane has good surface stability and can be used to filter intravenous infusion drugs to remove endotoxins without causing the release of surface modifiers and thus contaminating the filtered intravenous injection solution.

[0070] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A method for removing endotoxins from intravenous injections, comprising the step of filtering an intravenous infusion drug using a modified PES membrane, wherein the pH of the drug is 2-5; The preparation method of the modified PES membrane comprises the following steps: S1. A condensing agent is added to a mixed system of a Tris buffer solution, a precursor compound and an amino compound, and a condensation reaction occurs under stirring to obtain a mixed solution; the precursor compound is a compound that can react with an amino group; S2. The PES membrane was immersed in the mixed solution obtained in step S1 and removed after the reaction, and washed with water and ethanol to obtain a modified PES membrane; The precursor compound is selected from one of O-carboxymethyl chitosan, N-carboxymethyl chitosan, and N,O-carboxymethyl chitosan; the amino-containing compound is selected from one of serine, polyethyleneimine, propylenediamine, ethylenediamine, butylenediamine, tris(2-aminoethyl)amine, and diethylenetriamine.

2. The method according to claim 1, characterized in that The pH of the Tris buffer solution is 7-9.

3. The method according to claim 1, characterized in that In the mixed system, the concentration of the precursor compound is 0.01-3 wt %.

4. The method according to claim 1, wherein In the mixed system, the concentration of the amino compound is 0.01-4 wt %.

5. The method according to claim 1, wherein The condensing agent is selected from one of dicyclohexylcarbodiimide, diisopropylcarbodiimide and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide; and the concentration of the condensing agent is 0-1 wt %.

6. The method according to claim 5, characterized in that In step S1: the condensation reaction temperature is 0-8°C, the reaction time is 0.05-6h; and / or; the stirring speed is 0-1000rpm.

7. The method according to claim 1, characterized in that In step S2, the reaction temperature is 5-75° C., and the reaction time is 0.5-5 days.

8. The method according to claim 1, characterized in that In step S2, the washing times are 3-6 times.

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