CN109046282B - Adsorbent for removing endotoxin by hemoperfusion and water solution and preparation method thereof - Google Patents
Adsorbent for removing endotoxin by hemoperfusion and water solution and preparation method thereof Download PDFInfo
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- CN109046282B CN109046282B CN201811014894.4A CN201811014894A CN109046282B CN 109046282 B CN109046282 B CN 109046282B CN 201811014894 A CN201811014894 A CN 201811014894A CN 109046282 B CN109046282 B CN 109046282B
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- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
- B01J20/26—Synthetic macromolecular compounds
- B01J20/265—Synthetic macromolecular compounds modified or post-treated polymers
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61M—DEVICES FOR INTRODUCING MEDIA INTO, OR ONTO, THE BODY; DEVICES FOR TRANSDUCING BODY MEDIA OR FOR TAKING MEDIA FROM THE BODY; DEVICES FOR PRODUCING OR ENDING SLEEP OR STUPOR
- A61M1/00—Suction or pumping devices for medical purposes; Devices for carrying-off, for treatment of, or for carrying-over, body-liquids; Drainage systems
- A61M1/36—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits
Abstract
The invention relates to an adsorbent for blood perfusion and endotoxin removal by aqueous solution and a preparation method thereof, wherein the adsorbent is mainly formed by connecting non-woven fabrics and polyethyleneimine by a covalent bond or a covalent group through a grafting chain; the preparation method mainly comprises the steps of grafting and activating fibers on the surface of the non-woven fabric and immobilizing the polyethyleneimine ligand. The preparation method is simple, the environmental pollution is less, and the obtained adsorbent has good endotoxin adsorption performance, blood compatibility and safety.
Description
Technical Field
The invention relates to the field of blood purification and water treatment, in particular to an adsorbent for removing endotoxin by blood perfusion and an aqueous solution and a preparation method thereof.
Background
Endotoxemia in humans is caused by a variety of conditions, including: under severe trauma, infection and other stress conditions; the reticuloendothelial system of the whole body has dysfunction, the immune function is reduced, and the endotoxin absorbed by the intestinal tract is excessive and exceeds the clearing capability of the organism; gastrointestinal mucosa ischemia, necrosis, barrier damage, release of large amount of endotoxin into blood; endotoxin absorbed by intestinal tract directly enters systemic circulation from collateral circulation due to liver dysfunction; infection of certain tissues and organs causes exogenous endotoxins to enter the blood.
Endotoxemia usually causes fatal infectious shock, multiple organ failure, disseminated intravascular coagulation and the like, and the mortality rate is extremely high. The endotoxin adsorption column can be applied to the fields of a plurality of diseases, including sepsis, severe traumatic infection, severe hepatitis, hepatic encephalopathy, severe acute pancreatitis complicated infection, lung infection and the like, and the number of patients suffering from the diseases is huge.
The endotoxin adsorbent is usually obtained by activating and loading a ligand by using macroporous resin, activated carbon, fiber or the like as a carrier. Currently, endotoxin adsorbents are mainly distributed and comprise deoxycholic acid, polymyxin B, arginine, recombinant proteins, Polyethyleneimine (PEI) and the like.
Macroporous adsorption resin used as endotoxin adsorbent carrier usually uses highly toxic and carcinogenic chloromethyl ether and nitrobenzene in the preparation process, the residue of the reagents has great safety hazard to the health of patients, and the pressure to the production environment is also great. After being coated, the activated carbon still drops a large amount of impurities when being subjected to blood perfusion, enters human blood and still has potential safety hazards to patients.
In hemodialysis, the endotoxin contained in the dialysis water produced from the water production equipment usually exceeds the specification of relevant standards, a series of dialysis complications can occur when the excessive endotoxin enters the human blood through the dialysis membrane, the living quality and the dialysis effect of a patient are seriously affected, and therefore, an endotoxin adsorption column is also needed to be adopted for adsorbing and filtering the excessive endotoxin in the dialysis water.
Chinese patent publication No. CN104525151A discloses that polystyrene divinyl benzene resin and polymethyl methacrylate resin are used as carriers, and the residual double bonds of the carrier resin are subjected to epoxidation modification, and then ligands are immobilized. However, since the porous carrier has a small number of double bonds remaining on the surface thereof, the amount of ligand immobilized is small, and the adsorption performance for endotoxin is low. In addition, although this approach reduces the residue of carcinogenic substances, it still uses a number of highly toxic chemical agents. Japanese patent publication No. JPH04270965A discloses an endotoxin adsorbent which is a woven fiber of polystyrene having polymyxin B on the surface, which has considerable toxic effects, mainly nephrotoxicity, neurotoxicity and neuromuscular blockade, in addition to being expensive.
Disclosure of Invention
In order to overcome the disadvantages and shortcomings of the prior art, the present invention provides an adsorbent for removing endotoxin in hemoperfusion and aqueous solution, which has good blood compatibility and selective adsorption performance for endotoxin.
Another object of the present invention is to provide a method for preparing an adsorbent for endotoxin removal in hemoperfusion and aqueous solutions.
In order to achieve the main object of the present invention, the present invention provides an adsorbent for endotoxin removal by hemoperfusion and aqueous solution, which is formed by forming a graft chain having an epoxy group or a hydroxyl group on a nonwoven fabric and covalently bonding polyethyleneimine through the epoxy group or covalently bonding polyethyleneimine through the hydroxyl group.
According to the scheme, the adsorbent is characterized in that a grafting chain with an epoxy group or a hydroxyl group is grafted on the surface of the non-woven fabric, and then polyethyleneimine is immobilized through the grafting chain. Wherein, the grafted chain with epoxy group or hydroxyl group can be obtained by various reaction modes, and the preparation method is relatively simple. The epoxy group or the hydroxyl group has higher reaction activity, is easy to immobilize the polyethyleneimine ligand, and forms a stable covalent bond or covalent group after reaction to prevent the ligand from being separated. The polyethyleneimine has good selective adsorption performance on endotoxin by combining the endotoxin in blood of a patient or the endotoxin in an aqueous solution through electrostatic complexation, and can realize efficient removal of the endotoxin. The non-woven fabrics can graft a large amount of grafting chains, and the grafting chains stretch out from the surface of the non-woven fabrics and serve as middle arms for connecting the non-woven fabrics and polyethyleneimine ligands, so that the steric hindrance is reduced when the ligands are immobilized, more polyethyleneimine can be immobilized, and the adsorption performance of the adsorbent is improved. The grafted chain and the polyethyleneimine are combined by a covalent bond or a covalent group, the covalent bond combination means a covalent bond connection mode formed by the direct reaction of a group on the grafted chain and a group in the polyethyleneimine, and the covalent group combination means that the group on the grafted chain and the group in the polyethyleneimine are connected together through a cross-linking agent or other molecules.
The further technical proposal is that when the grafted chain has epoxy groups, the density of the epoxy groups on the non-woven fabric is 0.7mmol/g to 1.6 mmol/g; or when the grafted chain has hydroxyl, the density of the hydroxyl on the non-woven fabric is 0.64mmol/g to 2.0mmol/g, and the hydroxyl reacts with the polyethyleneimine by glutaraldehyde to form covalent groups.
According to the scheme, the grafted chain in the adsorbent provided by the invention is used for immobilizing the polyethyleneimine ligand through the epoxy group or the hydroxyl group on the grafted chain, and when the density of the epoxy group or the hydroxyl group is in the range, the grafted chain is beneficial to connecting more polyethyleneimine, and meanwhile, the phenomenon that the polyethyleneimine chain is overcrowded and the adsorption performance is reduced due to steric hindrance is avoided. The epoxy group density and the hydroxyl group density refer to the number of moles of the nonwoven fabric group having a graft chain per unit mass.
The further technical proposal is that the graft chain is formed by graft polymerization of Glycidyl Methacrylate (GMA) on non-woven fabrics; or the grafted chain is formed by graft polymerization of hydroxyethyl methacrylate (HEMA), hydroxyethyl acrylate (HEA) or a mixture of the two on the non-woven fabric.
According to the scheme, the grafted chain in the adsorbent can be formed by graft polymerization of a reactive monomer with double bonds and epoxy groups or hydroxyl groups on the non-woven fabric, wherein the double bonds participate in the graft polymerization, and the epoxy groups or the hydroxyl groups are remained in the grafted chain and react with polyethyleneimine. The grafting chain formed by the method has a large amount of epoxy groups or hydroxyl groups, and can be used for immobilizing a large amount of polyethyleneimine, so that the adsorption performance of the adsorbent is improved.
The further technical proposal is that the non-woven fabric is polyester non-woven fabric, polyethylene non-woven fabric or polypropylene non-woven fabric.
According to the scheme, the non-woven fabric is preferably prepared from a polymer which has good blood compatibility, is non-toxic and is not easy to decompose.
The further technical proposal is that the aperture of the non-woven fabric is 1 to 300 mu m, the fiber diameter is 1 to 15 mu m, and the single piece thickness is 0.2 to 3.0 mm.
According to the scheme, the non-woven fabric with the aperture, the fiber diameter and the single thickness can graft more graft chains on the fiber surface of the non-woven fabric on the basis of ensuring the strength of the non-woven fabric carrier, so that the adsorption performance is improved.
In order to achieve another object of the present invention, the present invention provides a method for preparing an adsorbent for removing endotoxin by hemoperfusion and an aqueous solution, the method comprising the steps of:
the method comprises the following steps: carrying out grafting reaction on the non-woven fabric in a grafting solution containing a reactive monomer, an initiator and a solvent to form a grafting chain with a hydroxyl group or an epoxy group on the non-woven fabric;
step two: when the grafted chain has an epoxy group, reacting the non-woven fabric with the grafted chain obtained in the step one in a polyethyleneimine solution; when the grafted chain has hydroxyl, adding glutaraldehyde into the non-woven fabric with the grafted chain obtained in the step one in a polyethyleneimine solution for reaction.
According to the scheme, the preparation method for preparing the endotoxin adsorbent is simple, mainly comprises the steps of activating the non-woven fabric and the immobilized ligand, and the adsorbent with good adsorption performance on endotoxin is obtained by firstly carrying out graft modification activation on the surface of the non-woven fabric substrate fiber and then connecting the polyethyleneimine ligand through the graft chain. When the graft chain has an epoxy group, ethanolamine may be further added to react with an excess epoxy group after the graft chain reacts with polyethyleneimine.
The further technical proposal is that the non-woven fabric is polyester non-woven fabric, polyethylene non-woven fabric or polypropylene non-woven fabric; the aperture of the non-woven fabric is 1-300 mu m, the fiber diameter is 1-15 mu m, and the thickness of each single piece is 0.2-3.0 mm; the reactive monomer is glycidyl methacrylate, or the reactive monomer is hydroxyethyl methacrylate, hydroxyethyl acrylate or a mixture thereof; the initiator is a photosensitizer and is selected from at least one of benzoin ethyl ether, anthraquinone-2, 6-disulfonic acid sodium, azo dye, Benzophenone (BP) or xanthone; the solvent is n-Butanol (BA).
According to the scheme, the carrier can be connected with more graft chains by selecting the specific pore diameter, the fiber diameter, the single-piece thickness and the specific type of non-woven fabric, and the blood compatibility or safety is improved; by selecting reactive monomers with epoxy groups or hydroxyl groups for graft polymerization, the content of the reactive groups of the grafted chain can be increased, so that more ligands are immobilized; the environment-friendly and nontoxic initiator and solvent are adopted, so that the biocompatibility and safety of the adsorbent are improved, and the damage of the preparation process to the environment is reduced.
The further technical proposal is that in the step one, the non-woven fabric is wetted by the grafting solution before the grafting reaction; the grafting reaction is carried out under the conditions of ultraviolet radiation and ventilation; after the grafting reaction, the nonwoven fabric with the grafted chains was rinsed with absolute ethanol.
According to the scheme, the grafting solution is used for fully infiltrating the non-woven fabric between grafting reactions so as to ensure that the reactions are fully carried out. Grafting and polymerization reaction are carried out under ventilation condition by ultraviolet rays, thus being safe, environment-friendly and high in efficiency. And after the grafting reaction, redundant photosensitizer, monomer and copolymer are removed, so that the treatment is convenient.
The further technical scheme is that the polyethyleneimine solution is a polyethyleneimine aqueous solution, and the polyethyleneimine is straight-chain type or branched-chain type polyethyleneimine with the molecular weight of 600-70000.
According to the scheme, the immobilization reaction is carried out in an aqueous system, and the method is safe and environment-friendly. The polyethyleneimine may have a different structure, or a different molecular weight, such as 600, 1800, 10000, 70000, or other suitable molecular weight, as desired.
The further technical scheme is that in the second step, when the grafted chain has an epoxy group, the reaction is carried out under the condition that the polyethyleneimine solution circularly flows and the room temperature is kept; after the reaction is finished, washing the product to be neutral by using distilled water, and drying at the temperature of 45 ℃; when the grafted chain has hydroxyl, the reaction is carried out under the condition of circulating flow of polyethyleneimine solution, firstly, glutaraldehyde solution is dropwise added, and then the reaction is continued at room temperature; after the reaction, the product was washed with distilled water to neutrality and then dried at 45 ℃.
According to the scheme, the immobilized reaction method is simple, the process is mild, and the operation is safe and environment-friendly.
Detailed Description
The following detailed description of the products, methods and benefits of the present invention will be presented in conjunction with the following examples to fully illustrate the features and benefits of the present invention. The types of raw materials, process conditions and ranges of important parameters that may be employed in the examples of the present invention are shown in table 1 below.
TABLE 1 raw material types, Process conditions or ranges
The specific steps of surface activation of the fibers of the substrate nonwoven fabric are described below by examples 1 to 8, and the specific steps of the immobilization reaction are described by examples 9 to 16.
Surface activation of substrate nonwoven fibers
Example 1
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of HEMA, BP and BA, wherein the volume fraction of HEMA is 10%, the volume fraction of BA is 90%, and the concentration of BP is 0.2g/100mL, and uniformly stirring and dissolving for later use.
2) Fully wetting the nonwoven with the grafting solution: a polybutylene terephthalate (PBT) disc having a pore size of 300 μm, a thickness of 2mm and a diameter of 10cm was immersed in an excess of the grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet radiation grafting reaction for 20 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 30W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:12, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the surface activated and containing a large number of hydroxyl groups is obtained, and the hydroxyl group density of the fiber surface of the non-woven fabric is 0.64 mmol/g.
Example 2
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of HEMA, BP and BA, wherein the volume fraction of HEMA is 25%, the volume fraction of BA is 75%, and the concentration of BP is 2g/100mL, and uniformly stirring and dissolving for later use.
2) Fully wetting the nonwoven with the grafting solution: a polyethylene terephthalate (PET) wafer having a pore size of 100 μm, a thickness of 1mm and a diameter of 10cm was immersed in the excess grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet radiation grafting reaction for 40 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 30W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:20, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the surface activated and containing a large number of hydroxyl groups is obtained, and the hydroxyl group density of the fiber surface of the non-woven fabric is 1.5 mmol/g.
Example 3
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of HEA, BP and BA, wherein the volume fraction of HEA is 10%, the volume fraction of BA is 90%, and the concentration of BP is 1.0g/100mL, and uniformly stirring and dissolving for later use.
2) Fully wetting the PBT non-woven fabric with a grafting solution: PBT disks with a pore size of 50 μm, a thickness of 0.3mm and a diameter of 10cm were immersed in an excess of the grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet radiation grafting reaction for 15 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 50W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:10, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the surface activated and containing a large number of hydroxyl groups is obtained, and the density of the hydroxyl groups on the fiber surface of the non-woven fabric is 0.8 mmol/g.
Example 4
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of HEA, BP and BA, wherein the volume fraction of HEA is 30%, the volume fraction of BA is 70%, and the concentration of BP is 2.0g/100mL, and uniformly stirring and dissolving for later use.
2) Fully wetting the nonwoven with the grafting solution: a PET wafer having a pore size of 50 μm, a thickness of 1.0mm and a diameter of 10cm was immersed in the excess grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet radiation grafting reaction for 15 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 70W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the surface activated and containing a large number of hydroxyl groups is obtained, and the hydroxyl group density of the fiber surface of the non-woven fabric is 2.0 mmol/g.
Example 5
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of GMA, BP and BA, wherein the volume fraction of GMA is 15%, the volume fraction of BA is 85%, and the concentration of BP is 1.0g/100mL, and stirring to dissolve uniformly for later use.
2) Fully wetting the nonwoven with the grafting solution: a polypropylene PP nonwoven wafer having a pore size of 150 μm, a thickness of 1.0mm and a diameter of 10cm was immersed in the excess grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the ultraviolet irradiation grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet irradiation grafting reaction for 15 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 30W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the activated surface containing a large amount of epoxy groups is obtained, and the density of the epoxy groups on the surface of the non-woven fabric fibers is 0.7 mmol/g.
Example 6
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of GMA, BP and BA, wherein the volume fraction of GMA is 30%, the volume fraction of BA is 70%, and the concentration of BP is 2.0g/100mL, and stirring to dissolve uniformly for later use.
2) Fully wetting the nonwoven with the grafting solution: a polyethylene PE nonwoven fabric wafer having a pore size of 150 μm, a thickness of 1.5mm and a diameter of 10cm was immersed in the excess grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the ultraviolet irradiation grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet irradiation grafting reaction for 15 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 50W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:20, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the activated surface containing a large amount of epoxy groups is obtained, and the density of the epoxy groups on the surface of the non-woven fabric fibers is 1.6 mmol/g.
Example 7
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of HEMA, BP and BA, wherein the volume fraction of HEMA is 20%, the volume fraction of BA is 80%, and the concentration of BP is 2.0g/100mL, and uniformly stirring and dissolving for later use.
2) Fully wetting the nonwoven with the grafting solution: a PBT nonwoven wafer having a pore size of 15 μm, a thickness of 1mm and a diameter of 10cm was immersed in an excess of the grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the ultraviolet irradiation grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet irradiation grafting reaction for 25 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 70W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the surface activated and containing a large number of hydroxyl groups is obtained, and the density of the hydroxyl groups on the fiber surface of the non-woven fabric is 0.96 mmol/g.
Example 8
1) Preparation of the grafting solution for grafting by ultraviolet irradiation: preparing a grafting solution of GMA, BP and BA, wherein the volume fraction of GMA is 20%, the volume fraction of BA is 80%, and the concentration of BP is 2.0g/100mL, and stirring to dissolve uniformly for later use.
2) Fully wetting the nonwoven with the grafting solution: a PE nonwoven wafer having a pore size of 15 μm, a thickness of 1mm and a diameter of 10cm was immersed in the excess grafting solution for 3 minutes.
3) Ultraviolet irradiation grafting modified non-woven fabric: and taking the non-woven fabric fully wetted by the ultraviolet irradiation grafting solution out of the grafting solution, draining for 3 minutes at room temperature, and carrying out ultraviolet irradiation grafting reaction for 25 minutes under an ultraviolet light source under the ventilation condition. The main wave peak of the ultraviolet light source is 365nm, the power is 30W/cm, and the vertical distance from the ultraviolet light source to the non-woven fabric wafer is 30 cm.
4) Cleaning and drying the non-woven fabric with the activated fiber surface: rinsing the non-woven fabric subjected to ultraviolet radiation grafting treatment in excessive absolute ethyl alcohol for 4 times, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is subjected to circular flow during rinsing, and each rinsing is carried out for 30 minutes. After rinsing, the mixture is drained until no ethanol drops drop, and then is dried for 1 hour at 40 ℃. The non-woven fabric with the activated surface containing a large amount of epoxy groups is obtained, and the density of the epoxy groups on the surface of the non-woven fabric fibers is 0.82 mmol/g.
The parameters for preparing the surface-activated nonwoven fabrics from examples 1 to 8 are summarized in table 2 below.
Table 2 parameters of examples 1 to 8
Endotoxin adsorbent prepared by immobilization reaction
Example 9
The PBT nonwoven fabric having a large number of hydroxyl groups (0.64mmol/g) after the surface activation of the fiber obtained in example 1 was added 140mL of a 5% by mass aqueous solution of linear PEI having a molecular weight of 600, and circulated for half an hour. 50mL of glutaraldehyde solution with the mass concentration of 2% is gradually added dropwise. The reaction was then continued at room temperature for 4 hours with circulation. The nonwoven fabric was taken out, washed with distilled water to neutrality, and dried at 45 ℃.
Example 10
The PET nonwoven fabric having a large number of hydroxyl groups (1.5mmol/g) after surface activation of the fiber obtained in example 2 was taken, 340mL of an aqueous solution of linear PEI having a molecular weight of 1800 and a mass concentration of 5% was added, and the mixture was circulated and flowed for half an hour. 150mL of glutaraldehyde solution with the mass concentration of 2% is gradually added dropwise. The reaction was then continued at room temperature for 8 hours with circulation. The nonwoven fabric was taken out, washed with distilled water to neutrality, and dried at 45 ℃.
Example 11
The PBT nonwoven fabric having a large number of hydroxyl groups (0.8mmol/g) after the surface activation of the fiber obtained in example 3 was added with 200mL of a 5% by mass aqueous solution of linear PEI having a molecular weight of 4800, and circulated for half an hour. 80mL of glutaraldehyde solution with the mass concentration of 2% is gradually added dropwise. The reaction was then continued at room temperature for 6 hours with circulation. The nonwoven fabric was taken out, washed with distilled water to neutrality, and dried at 45 ℃.
Example 12
The PET nonwoven fabric having a large number of hydroxyl groups (2.0mmol/g) after the surface activation of the fiber obtained in example 4 was taken, 500mL of an aqueous solution of linear PEI having a molecular weight of 70000 and a mass concentration of 5% was added, and the mixture was circulated and flowed for half an hour. 180mL of glutaraldehyde solution with the mass concentration of 2% is gradually added dropwise. The reaction was then continued at room temperature for 12 hours with circulation. The nonwoven fabric was taken out, washed with distilled water to neutrality, and dried at 45 ℃.
Example 13
The PP nonwoven fabric having activated fiber surfaces and a large number of epoxy groups (0.7mmol/g) obtained in example 5 was taken, 180mL of an aqueous solution of branched PEI having a molecular weight of 600 and a mass concentration of 5% was added, and a circulating flow reaction was carried out at room temperature for 4 hours. The nonwoven fabric was taken out and put into another container, and then 20mL of 1.5mol/L ethanolamine was added thereto, followed by reaction at room temperature for 4 hours. Taking out the non-woven fabric, rinsing the non-woven fabric for 4 times by using absolute ethyl alcohol, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is circulated and flows during rinsing for 30 minutes each time. And rinsed with water for injection for 30 minutes and dried at 45 c.
Example 14
The PE nonwoven fabric having a large number of epoxy groups (1.6mmol/g) after the surface activation of the fiber obtained in example 6 was added with 400mL of an aqueous solution of branched PEI having a molecular weight of 1800 and a mass concentration of 5%, and subjected to a circulating flow reaction at room temperature for 8 hours. The nonwoven fabric was taken out and put into another container, and then 20mL of 1.5mol/L ethanolamine was added thereto, followed by reaction at room temperature for 4 hours. Taking out the non-woven fabric, rinsing the non-woven fabric for 4 times by using absolute ethyl alcohol, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is circulated and flows during rinsing for 30 minutes each time. And rinsed with water for injection for 30 minutes and dried at 45 c.
Example 15
The PBT nonwoven fabric having a large number of hydroxyl groups (0.96mmol/g) after the surface activation of the fiber obtained in example 7 was added with 200mL of an aqueous solution of branched PEI having a molecular weight of 4800 and a mass concentration of 5%, and circulated for half an hour. 80mL of glutaraldehyde solution with the mass concentration of 2% is gradually added dropwise. The reaction was then continued at room temperature for 12 hours with circulation. The nonwoven fabric was taken out, washed with distilled water to neutrality, and dried at 45 ℃.
Example 16
The PE nonwoven fabric having a large number of epoxy groups (0.82mmol/g) after the surface activation of the fiber obtained in example 8 was added with 200mL of an aqueous solution of branched PEI having a molecular weight of 10000 and having a mass concentration of 5%, and subjected to a circulating flow reaction at room temperature for 12 hours. The nonwoven fabric was taken out and put into another container, and then 20mL of 1.5mol/L ethanolamine was added thereto, followed by reaction at room temperature for 4 hours. Taking out the non-woven fabric, rinsing the non-woven fabric for 4 times by using absolute ethyl alcohol, wherein the volume ratio of the non-woven fabric to the absolute ethyl alcohol is 1:15, and the absolute ethyl alcohol is circulated and flows during rinsing for 30 minutes each time. And rinsed with water for injection for 30 minutes and dried at 45 c.
A summary of the parameters of the PEI immobilization reactions of examples 9 to 16 is shown in table 3 below.
TABLE 3 parameters of examples 9 to 16
Performance testing of endotoxin adsorbents
Adsorption rate of endotoxin in plasma
It is currently generally accepted that endotoxin concentrations in normal human plasma are below 0.1EU/mL, that plasma endotoxin concentrations in patients with endotoxemia are typically distributed between 0.1 and 1.0EU/mL, and that very few patients have endotoxin concentrations distributed between 1.0 and 10 EU/mL. Blood plasma with endotoxin concentrations of 0.5EU/mL, 1.0EU/mL and 2.0EU/mL, respectively, was prepared and adsorbed with endotoxin adsorbents prepared in examples 9 to 16 at a volume ratio of 1:8 with an oscillation frequency of 100. + -. 3Hz, a temperature of 37. + -. 1 ℃ and an adsorption time of 2 hours. The endotoxin concentration of the diluted plasma was measured by the turbidity method, and the endotoxin adsorption rate of the endotoxin adsorbent was calculated, and the results are shown in Table 3.
TABLE 3 adsorption rate of endotoxin in plasma by endotoxin adsorbents prepared in examples
According to the endotoxin adsorption result, the adsorbent in the embodiment of the invention has better endotoxin adsorption rate, and can obviously reduce the concentration of endotoxin in blood plasma. For example, the endotoxin adsorption rate of example 12 can be even 70% or more.
Adsorption rate of proteins in plasma
Total protein loss experiments during plasma adsorption were performed according to the standard YY1290-2016 Disposable bilirubin plasma adsorber.
Measuring 1mL of adsorbent, placing into a conical flask, wherein the volume ratio of the adsorbent to the blood plasma is 1:8, placing into a constant-temperature water bath oscillator, controlling the temperature at 37 +/-1 ℃ and the oscillation frequency at 100 +/-3 Hz, taking out the conical flask after oscillation adsorption for 2 hours, standing at room temperature for 3 minutes, sampling and measuring the concentration, and the results are shown in Table 4.
Table 4 shows the adsorption rate of endotoxin adsorbents for total protein in plasma
| Examples | Rate of protein loss |
| Example 9 | 3.6％ |
| Example 10 | 4.2％ |
| Example 11 | 5.3％ |
| Example 12 | 5.6％ |
| Example 13 | 7.2％ |
| Example 14 | 4.1％ |
| Example 15 | 3.9％ |
| Example 16 | 6.4％ |
As can be seen from the results of total protein adsorption tests, the adsorbent in the examples of the present invention has a total protein loss of less than 8%, which is significantly less than 15% specified in YY1290-2016 Disposable bilirubin plasma adsorber, and the endotoxin adsorbent disclosed in the present invention has good blood compatibility in terms of total protein loss.
Therefore, the hydrophilic middle arm with a large number of hydroxyl groups or epoxy groups is grafted on the surface of the matrix non-woven fabric fiber by the ultraviolet irradiation grafting technology, the main ligand polyethyleneimine for adsorbing endotoxin is fixed on the hydroxyl groups or epoxy groups of the middle arm through chemical covalent bonds, and the endotoxin adsorbent for hemoperfusion and water solution adsorption is prepared. The preparation method is simple, avoids the use of toxic and carcinogenic chemical reagents in the operation process, improves the safety of the adsorbent and reduces the damage to the environment.
The embodiments of the present invention have been described in detail, but the embodiments are merely examples, and the present invention is not limited to the embodiments described above. Any equivalent modifications and substitutions to those skilled in the art are also within the scope of the present invention. Accordingly, equivalent changes and modifications made without departing from the spirit and scope of the present invention should be covered by the present invention.
Claims (6)
1. An adsorbent for removing endotoxin by hemoperfusion and an aqueous solution, which is characterized in that:
the adsorbent is formed by forming a grafting chain with an epoxy group on a non-woven fabric and then connecting the epoxy group and polyethyleneimine in a covalent bond mode; the grafted chain is formed by ultraviolet irradiation graft polymerization of glycidyl methacrylate on the non-woven fabric, and the density of epoxy groups on the non-woven fabric is 0.7mmol/g to 1.6 mmol/g;
or the adsorbent is formed by forming a grafting chain with hydroxyl on non-woven fabric and then connecting the hydroxyl and polyethyleneimine in a covalent group mode; the grafting chain is formed by ultraviolet irradiation graft polymerization of hydroxyethyl methacrylate, hydroxyethyl acrylate or a mixture of the hydroxyethyl methacrylate and the hydroxyethyl acrylate on the non-woven fabric, and the density of hydroxyl on the non-woven fabric is 0.64mmol/g to 2.0 mmol/g; the hydroxyl groups are reacted with the polyethyleneimine by glutaraldehyde to form the covalent groups;
the non-woven fabric is polyester non-woven fabric, polyethylene non-woven fabric or polypropylene non-woven fabric; the aperture of the non-woven fabric is 1-300 mu m, the fiber diameter is 1-15 mu m, and the single-piece thickness is 0.2-3.0 mm.
2. A preparation method of an adsorbent for removing endotoxin by hemoperfusion and aqueous solution is characterized by comprising the following steps:
the method comprises the following steps: carrying out ultraviolet irradiation grafting reaction on the non-woven fabric in a grafting solution containing a reactive monomer, an initiator and a solvent to form a grafted chain with an epoxy group on the non-woven fabric; the reactive monomer is glycidyl methacrylate; the density of epoxy groups on the non-woven fabric is 0.7mmol/g to 1.6 mmol/g;
step two: reacting the non-woven fabric with the grafted chain obtained in the step one in a polyethyleneimine solution;
alternatively, the method comprises the steps of:
the method comprises the following steps: carrying out ultraviolet irradiation grafting reaction on the non-woven fabric in a grafting solution containing a reactive monomer, an initiator and a solvent to form a grafting chain with hydroxyl on the non-woven fabric; the reactive monomer is hydroxyethyl methacrylate, hydroxyethyl acrylate or a mixture thereof; the density of hydroxyl groups on the non-woven fabric is 0.64mmol/g to 2.0 mmol/g;
step two: adding glutaraldehyde into the non-woven fabric with the grafted chain obtained in the step one in a polyethyleneimine solution for reaction;
in any of the above methods, the nonwoven fabric is a polyester nonwoven fabric, a polyethylene nonwoven fabric, or a polypropylene nonwoven fabric; the aperture of the non-woven fabric is 1-300 mu m, the fiber diameter is 1-15 mu m, and the single-piece thickness is 0.2-3.0 mm.
3. The method for producing an adsorbent for endotoxin removal by hemoperfusion and aqueous solution according to claim 2, wherein:
the initiator is a photosensitizer and is selected from at least one of benzoin ethyl ether, anthraquinone-2, 6-disulfonic acid sodium, azo dye, benzophenone or xanthone;
the solvent is n-butanol.
4. The method for producing an adsorbent for endotoxin removal by hemoperfusion and aqueous solution according to claim 2 or 3, wherein:
in the first step, before the grafting reaction, the non-woven fabric is wetted by the grafting solution, and then the non-woven fabric is taken out and drained;
the grafting reaction is carried out under the condition of ventilation; after the grafting reaction, the nonwoven fabric with the grafted chains was rinsed with absolute ethanol.
5. The method for producing an adsorbent for endotoxin removal by hemoperfusion and aqueous solution according to claim 2, wherein:
the polyethyleneimine solution is a polyethyleneimine aqueous solution, the polyethyleneimine is straight-chain or branched polyethyleneimine, and the molecular weight of the polyethyleneimine is 600-70000.
6. The method for producing an adsorbent for endotoxin removal by hemoperfusion and aqueous solution according to claim 2 or 5, wherein:
in the second step, when the grafted chain has an epoxy group, the reaction is carried out under the condition that the polyethyleneimine solution circularly flows and the room temperature is kept; after the reaction is finished, washing the product to be neutral by using distilled water, and then drying;
when the grafted chain has hydroxyl, the reaction is carried out under the circulating flow of a polyethyleneimine solution, a glutaraldehyde solution is firstly dropwise added, and then the reaction is continued at room temperature; after the reaction is finished, washing the product to be neutral by using distilled water, and then drying.
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