CN113578288B - Cell inflammatory factor adsorbent and preparation method thereof - Google Patents

Cell inflammatory factor adsorbent and preparation method thereof Download PDF

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CN113578288B
CN113578288B CN202110868949.3A CN202110868949A CN113578288B CN 113578288 B CN113578288 B CN 113578288B CN 202110868949 A CN202110868949 A CN 202110868949A CN 113578288 B CN113578288 B CN 113578288B
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inflammatory factor
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CN113578288A (en
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刘群奇
白珂
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Beijing Zhongke Taikang Technology Co ltd
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • 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
    • B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
    • B01J20/28014—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their form
    • B01J20/28016—Particle form
    • B01J20/28019—Spherical, ellipsoidal or cylindrical
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • 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
    • B01J20/261—Synthetic macromolecular compounds obtained by reactions only involving carbon to carbon unsaturated bonds
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • 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
    • B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
    • B01J20/28002—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their physical properties
    • B01J20/28011—Other properties, e.g. density, crush strength
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2220/00—Aspects relating to sorbent materials
    • B01J2220/40—Aspects relating to the composition of sorbent or filter aid materials
    • B01J2220/44—Materials comprising a mixture of organic materials
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
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    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

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Abstract

The invention discloses a cell inflammatory factor adsorbent and a preparation method thereof, wherein the framework is polystyrene chloride spheres, and glycine is coupled to a carrier by utilizing the activity of benzyl chloride on the framework. By utilizing the activity of benzyl chloride on the polystyrene chloride balls, the coupling efficiency is high, amino acids such as valine can be linked by further modifying the coupling amino acids, and the chain connecting efficiency is high.

Description

Cell inflammatory factor adsorbent and preparation method thereof
Technical Field
The invention relates to a high molecular biomedical material, in particular to a cell inflammatory factor adsorbent applied to blood perfusion and a preparation method thereof.
Background
With the economic development, the urban pace is continuously accelerated, the urban clusters and population density are continuously increased, the traffic convenience is continuously improved, the life and travel of people are facilitated, the risk of infectious disease transmission is increased, and the environmental pollution is increasingly serious due to the economic activity, so that new challenges are brought to public health and safety and health of people.
Recent studies of new cases of coronaries have shown that most elderly and people suffering from underlying diseases (e.g. hypertension, chronic obstructive pulmonary disease, diabetes, cardiovascular disease) rapidly develop acute respiratory distress syndrome, septic shock, and even death. Through laboratory observation, the inflammatory factors of a majority of severe patients, such as interleukin-6 (IL-6), tumor necrosis factor (TNF-alpha) and the like, are obviously increased.
The cell inflammatory factors are low molecular proteins or polypeptides which can transmit information among cells and have immunoregulation and effect functions, and can be classified into Interleukin (IL), interferon (IFN), tumor necrosis factor (TNF-alpha), growth factors and the like. Cytokines stimulate and regulate the intensity and duration of the immune system response to help the body resist infection, however, cytokines are a double-edged sword that when overactivated in the body can cause "cytokine storms" in the body. The systemic inflammatory response caused by various causes such as infection, medicine and the like can damage human bodies, and cause autoimmune diseases of the human bodies. Cytokine storms are closely related to influenza, sepsis/septic shock, acute respiratory distress syndrome, etc., and ultimately can lead to multiple organ failure.
The molecular weight of the cytokines is mostly between several and tens of kDa, of which interleukin-6 (IL-6) is represented, and the molecular weight is 26kDa. In inflammatory response, IL-6 is elevated earlier than other cytokines and is of a longer duration and thus useful in assisting in early diagnosis of acute infections. IL-6 levels rapidly rise after infection in humans, peaking at 2 hours, with elevated levels consistent with the severity of the infection. Thus, IL-6 may serve as the most typical marker of inflammation.
The national emphasis laboratory attached to the university of Zhejiang medical college for diagnosis and treatment of infectious diseases in the first hospital indicates that shock and hypoxia caused by novel coronaviruses are caused by cytokine storm in Zhejiang experience in the published 2019 coronavirus disease (COVID-19) diagnosis and treatment, and immunological indexes related to cytokine storm, especially IL-6 and the like, possibly have potential clinical values for measuring the severity of illness and predicting death risk of patients with heavy COVID-19.
The tumor necrosis factor (TNF-alpha) has a molecular weight of 17 kDa-25 kDa, can participate in the immune regulation of organisms, is also a causative factor of certain infectious diseases, and has toxicity to various biological cells when accumulated in a large amount.
The blood purifying system comprising blood adsorption, perfusion technology, etc. can eliminate inflammation medium fast, block the inflammatory chain reaction, restore the balance of inflammatory factor and restore the immune homeostasis of organism.
Because the molecular weight of the cell inflammatory factor is larger, the adsorption effect of the broad-spectrum adsorbent is not ideal, and the selectivity is poor. The general method is to modify the framework of the adsorbent to achieve the effect of specific adsorption. Patent No. 201510343163.4 mentions that adsorbents prepared by epoxidizing a polystyrene backbone followed by attachment of a selective adsorption group can be used for the removal of cytokines. However, the coupling ligand is not efficient because the coupling ligand is subjected to epoxidation.
Disclosure of Invention
The invention aims to provide the cell inflammatory factor adsorbent which has novel and unique structure and convenient use and does not need to modify an adsorbent framework; the specific technical scheme is as follows:
a cell inflammatory factor adsorbent has polystyrene chloride sphere as skeleton, and is prepared by coupling glycine to carrier by using activity of benzyl chloride on skeleton.
Further, the particle diameter of the carrier is 0.4-1.2 mm, the pore diameter of the carrier is 2-100 nm, the specific surface area is not less than 500 square meters per gram, and the chlorine content is not less than 17%.
Further, glycine is coupled to the carrier, and the concentration of glycine solution is 5% -25% by weight.
Further, the reaction temperature of the coupling glycine is 30-60 ℃.
Further, the reaction time of coupling glycine is 8-48 h.
Further, the use of group protecting reagents in coupling glycine
Further, the radical protection agent is chloroformate and its derivatives.
Further, the chloroformate may be benzyl chloroformate or tert-butyl chloroformate.
Further, the chloroformate can be benzyl chloroformate or tert-butyl chloroformate, and the molar ratio of the chloroformate to glycine is 1:1-3:1.
Further, a deprotection reagent is used after coupling glycine.
Further, the deprotecting reagent is a mixture of diethyl ether, ethyl acetate and trifluoroacetic acid.
Further, the mass fraction of the trifluoroacetic acid solution is 0.1-5%, preferably 2%.
Further, the carrier was further coupled to valine.
Further, the concentration of the solution of the coupling valine is 1-8%, preferably 8% by weight.
Further, the carboxylic acid group on valine is activated when valine is coupled, and the pH value is controlled to be 4-6 during activation.
Further, the activating reagent for activating the carboxylic acid group on valine is N-hydroxysuccinimide and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride.
Further, an activating reagent for activating the carboxyl group on valine can be disclosed in the prior art.
Furthermore, the adsorbent has good blood compatibility and can perform whole blood perfusion adsorption.
The beneficial effects of the invention are as follows:
the activity of the benzyl chloride on the polystyrene chloride balls is utilized to directly couple Gan Ji acid, so that the coupling efficiency is high; can also be linked with amino acids such as valine, and has high linking efficiency.
The amide bond formed by the linking increases biocompatibility.
The cell inflammatory factor is protein or glycoprotein, and valine bonded on the adsorbent can form a synergistic effect of hydrogen bond and electrostatic action with the inflammatory factor, so that the selective removal capability of the inflammatory factor is improved.
Valine is essentially a protein with carboxyl and amino groups and is charged, and the valine bonded on the adsorbent can form a synergistic effect of hydrogen bond and electrostatic action with inflammatory factors, so that the selective removal capacity of the inflammatory factors is improved.
The coating treatment is not needed, and the risk that the coating layer falls into the human body is avoided.
Detailed Description
The present invention will be described more fully with reference to the following examples. This invention may be embodied in many different forms and should not be construed as limited to the exemplary embodiments set forth herein.
Example 1:
1-1 pretreatment
Taking a 20% glycine aqueous solution by weight, adjusting the pH value of the solution to 9 by using 0.5% NaOH, and adjusting the pH value of the solution to be alkaline by using other low-concentration strong alkali; 50ml of the prepared glycine solution is measured, 26.4g of benzyl chloroformate is added, and the mixture is uniformly mixed, and the mixture is stirred for 1 hour at the speed of 100rpm of a stirrer at 40 ℃ for later use. The chloroformate can be benzyl chloroformate or tertiary butyl chloroformate, and the molar ratio of the chloroformate to glycine is controlled to be 1:1-3:1.
1-2 coupling Glycine
10g (commercially available, the parameters are: chlorine content 20%, aperture 2-60 nm, specific surface area not less than 500 square meters/g) of polystyrene chloride balls are taken, 50ml of dichloroethane is added to swell for 2 hours, then 10ml of thionyl chloride and 20ml of the solution obtained in the step 1-1 are respectively added, stirring reaction is carried out for 48 hours at 40 ℃, and the rotation speed of a stirrer is 100rpm. Then 10ml of ethyl acetate and 30ml of 2% cold trifluoroacetic acid solution are added, the rotation speed of a stirrer is reduced to 50rpm for continuous reaction for 8 hours, then ethyl acetate is used for extraction, then alcohols such as methanol or ethanol are used for alcohol washing, and the microspheres are washed by purified water and dried for later use.
1-3 coupling valine
Adding 50ml of valine solution with weight of 8% into 5g of microsphere obtained in the step 1-2, dripping acetic acid with weight of 0.5% into the solution to adjust the pH value of the solution to 5.0, adding 0.4g of N-hydroxysuccinimide, adding 0.3g of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, vibrating the mixture at normal temperature for reaction for 6 hours, washing the mixture with ethanol and purified water, and drying the mixture at 80 ℃ to obtain the cell inflammatory factor adsorbent.
Example 2:
2-1 pretreatment
Taking a 20 wt% glycine aqueous solution, dripping a 0.5% NaOH solution to pH 10, weighing 30ml of the prepared glycine solution, adding 12.8g of tertiary butyl chloroformate, uniformly mixing, and stirring at the rotating speed of a stirrer of 100rpm for 1h at normal temperature for later use.
2-2 coupling Glycine
10g (commercially available, the parameters are: chlorine content 18%, pore diameter 5-50 nm, specific surface area not less than 500 square meters per gram) of polystyrene chloride balls are taken, 50ml of dichloroethane is added to swell for 2 hours, then 10ml of thionyl chloride and 20ml of solution obtained by 2-1 are respectively added, stirring reaction is carried out for 24 hours at 40 ℃, and the rotation speed of a stirrer is 100rpm. Then 10ml of diethyl ether and 30ml of 2% cold trifluoroacetic acid solution are added, the rotation speed of a stirrer is reduced to 50rpm for continuous reaction for 8 hours, ethyl acetate is used for extraction for 8 hours, then alcohol washing, water washing and microsphere drying are carried out for standby.
2-3 coupling valine
Taking 5g of microspheres obtained in 2-2, adding 50ml of a weight 5% valine solution, adjusting the pH to 4.0 by using a 0.5% acetic acid solution, adding 0.3g of N-hydroxysuccinimide, adding 0.3g of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, carrying out oscillation reaction for 8 hours at normal temperature, washing by using ethanol and purified water respectively, and drying to obtain the cell inflammatory factor adsorbent.
Example 3:
3-1 pretreatment
Taking a 20 wt% glycine aqueous solution, dripping 0.5% NaOH to adjust the solution to be alkaline, measuring 20ml of the prepared glycine solution, adding 8.5g of tertiary butyl chloroformate, uniformly mixing, and stirring at the speed of 100rpm at the temperature of 40 ℃ for 1 hour for later use.
3-2 coupling Glycine
10g of polystyrene chloride balls (the commercial parameters are that the chlorine content is 18 percent, the aperture is 5-50 nm, the specific surface area is not less than 500 square meters per gram) are taken, 50ml of dichloroethane is added to swell for 2 hours, then 10ml of thionyl chloride and 20ml of solution obtained by 2-1 are respectively added, stirring reaction is carried out for 24 hours at 40 ℃, the rotating speed is 100rpm, then 10ml of ethyl acetate and 30ml of 2% cold trifluoroacetic acid solution are added, the rotating speed is reduced to 50rpm to continue the reaction for 8 hours, then ethyl acetate is used for extraction for 8 hours, then alcohol washing and water washing are carried out on the microspheres, and the microspheres are dried for standby.
3-3 coupling valine
Taking 5g of microspheres obtained in 3-2, adding 50ml of a weight 5% valine solution, adjusting the pH to 6 by using a 0.5% acetic acid solution, adding 0.3g of N-hydroxysuccinimide, adding 0.3g of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, performing oscillation reaction for 8 hours at normal temperature, then respectively washing by using ethanol and purified water, and drying to obtain the cell inflammatory factor adsorbent.
Example 4:
4-1 pretreatment
Taking 20 weight percent glycine aqueous solution, dripping 0.5 percent NaOH to adjust the solution to be alkaline, weighing 30ml of the prepared glycine solution, then adding 15.8g of benzyl chloroformate, uniformly mixing, and stirring at the rotating speed of a stirrer of 100rpm for 1 hour at normal temperature for later use.
4-2 coupling Glycine
10g of polystyrene chloride balls (the commercial parameters are that the chlorine content is 18 percent, the aperture is 5-50 nm, the specific surface area is not less than 500 square meters per gram) are taken, 50ml of dichloroethane is added to swell for 2 hours, then 10ml of thionyl chloride and 20ml of solution obtained by 2-1 are respectively added, stirring reaction is carried out for 24 hours at 40 ℃ at the speed of 100rpm, then 10ml of diethyl ether and 30ml of 2% cold trifluoroacetic acid solution are added, the speed of rotation is reduced to 50rpm for continuous reaction for 8 hours, then ethyl acetate is used for extraction for 8 hours, then alcohol washing and water washing are carried out on the microspheres, and drying is carried out for standby.
4-3 coupling valine
Taking the microsphere obtained by 4-2, regulating the pH to 4.0 by using 0.5% acetic acid solution, adding a wt5% valine solution, adding 0.4g of N-hydroxysuccinimide, adding 0.5g of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, performing oscillation reaction for 10 hours at normal temperature, then respectively washing by using ethanol and purified water, and drying to obtain the cell inflammatory factor adsorbent.
Comparative example 1:
20g of commercial chlorine balls (with the particle size of 0.3-1.1 mm) are taken, 60ml of dichloroethane and 6g of aluminum trichloride are added, mechanical stirring is started, the normal temperature expansion is carried out for 5h, the temperature is increased to 100 ℃ for reaction for 8h, the crude product is obtained, the dichloroethane is distilled off by reduced pressure distillation, alcohol washing, water washing and suction filtration are carried out, drying is carried out, and the deep-color microspheres with the particle size of 0.35-1.2 mm are obtained by screening.
Comparative example 2:
taking 10g of commercial chlorine balls (with the particle size of 0.3-1.1 mm), adding 40ml of dichloroethane and 5g of aluminum trichloride, starting mechanical stirring, expanding for 3 hours at normal temperature, increasing the temperature to 100 ℃ for reacting for 12 hours to obtain a crude product, distilling under reduced pressure to distill out dichloroethane, washing with alcohol, washing with water, filtering, drying, and screening to obtain the deep-color microspheres with the particle size of 0.355-1.2 mm.
Example 5: hemolysis test
Part 4 of the biological evaluation of medical devices according to GB/T16886.4-2003: the hemolysis rate of the adsorbent was evaluated by selecting an experiment for interaction with blood.
Test set, negative control and positive control, 5g of the adsorbent prepared above was added to the test tube, 10ml of 0.9% aqueous NaCl solution was added to the negative control tube, and 10ml of distilled water was added to the positive control. 10ml of 0.9% NaCl aqueous solution was added to each of the test tube, the negative control tube and the positive control tube. All test tubes are placed in a constant temperature water area at the temperature of (37+/-1) ℃ for 60rpm vibration leaching (24+/-2) h, then the test liquid is taken out, the diluted rabbit blood is added into each test tube according to the proportion of 0.2ml of diluted rabbit blood to 10ml of test liquid, then the test liquid is uniformly mixed, placed in the constant temperature water area at the temperature of (37+/-1) ℃ for 60min, then the liquid in the test tube is poured out for centrifugation for 5min, the supernatant is sucked, and the absorbance is measured at 545nm by an ultraviolet spectrophotometer. Three groups were averaged over 3 tubes. The calculated hemolysis rate of the adsorbent is shown in the following table.
TABLE 1 adsorbent hemolysis rate
Project Average value of absorbance Hemolysis rate (%)
Negative control 0.0197 /
Positive control 08914 /
Example 1 0.0275 1.0
Example 2 0.0280 0.9
Example 3 0.0270 0.8
Example 4 0.0280 0.9
Comparative example 1 0.0640 5.1
Comparative example 2 0.0750 6.3
Compared with the comparative example, the hemolysis rate of the examples is less than or equal to 1.0 percent, and meets the requirement of less than or equal to 5 percent.
Example 6: cell inflammatory factor adsorption experiment
The non-pyrogen test tube was taken, 0.5g of the adsorbents prepared in examples and comparative examples were added, and then 2ml of plasma containing 140.3pg/ml of interleukin-6 (IL-6) and 336.4pg/ml of tumor necrosis factor (TNF-. Alpha.) were added, respectively, and the mixture was placed in a constant temperature water area shaker at (37.+ -. 1), (100.+ -. 10) rpm for 2 hours, and then the IL-6 and TNF-. Alpha. Concentrations were measured, and the decrease rates of IL-6 and TNF-. Alpha. Before and after adsorption were calculated.
TABLE 2 clearance of adsorbents to cytokines
Adsorbent and process for producing the same IL-6 concentration before adsorption (pg/ml) IL-6 concentration after adsorption (pg/ml) IL-6 decrease Rate (%) TNF-alpha concentration before adsorption (pg/ml) TNF-alpha concentration after adsorption (pg/ml) TNF-alpha decrease rate (%)
Example 1 140.3 26.4 81.2 336.4 67.3 80.0
Example 2 140.3 27.2 80.6 336.4 69.0 79.5
Example 3 140.3 28.2 79.9 336.4 71.7 78.7
Example 4 140.3 23.1 83.5 336.4 70.3 79.1
Comparative example 1 140.3 98.1 30.1 336.4 237.8 29.3
Comparative example 2 140.3 90.2 35.7 336.4 239.2 28.9
It can be seen that the modified adsorbent with amino acid groups having similar structures to inflammatory factors has a higher adsorption rate to both inflammatory factors, which is almost 2.5 times that of the unmodified broad-spectrum adsorbent, showing good selective adsorption capacity to both inflammatory factors.
Example 7: sorbent clotting assay
Respectively taking 0.5g of each of the adsorbents prepared in the examples and the comparative examples, soaking 30ml of each of the adsorbents in 0.9% NaCl, then respectively adding 1.5ml of human blood (sodium citrate for anticoagulation), placing the adsorbents in a water area with the temperature of (37+/-1) DEG C and the temperature of (100+/-10) rpm for incubation for 1 hour under constant-temperature shaking, and then measuring prothrombin time. Blank samples were prepared in the same way.
TABLE 3 sorbent clotting assay
Project Prothrombin time(s) Positive control Negative ofControl(s)
Example 1 13.5 NC 13.0
Example 2 13.3 NC 13.1
Example 3 14.0 NC 13.9
Example 4 13.9 NC 13.2
Note that: NC indicates that coagulation has not occurred
The prothrombin time of the adsorbent is within the normal range (10-14 s) of the human prothrombin blood coagulation function.
The invention provides a novel method for preparing selectively adsorbed cell inflammatory factors, which does not need to modify an adsorbent framework, namely directly couples amino acid by utilizing the activity of benzyl chloride on polystyrene chloride spheres, and has high coupling efficiency.
By further modifying the coupling amino acid, amino acid such as valine can be connected, and the chain connection efficiency is high.
The amide bond formed by the reaction increases the biocompatibility of the material.
The cell inflammatory factor is protein or glycoprotein, and can form a synergistic effect of hydrogen bond and electrostatic action with the inflammatory factor by utilizing amino acid groups bonded on the adsorbent, so that the selective clearance capability of the inflammatory factor is improved.
Valine is an essential amino acid for human body, and can maintain the growth and development of the organism, and avoid neurological disorder, dyskinesia, anemia, etc. caused by deficiency.
No surface physical modification is needed, and the introduction of redundant chemical reagents is avoided.
The above examples are for illustration of the invention only and, in addition, there are many different embodiments which will be apparent to those skilled in the art after having the insight into the present invention and are not explicitly recited herein.

Claims (5)

1. A cell inflammatory factor adsorbent is characterized in that polystyrene chloride balls are used as carriers, and the surfaces of the carriers are coupled with glycerol
Amino acid, wherein valine is further coupled to the surface of the carrier; the pore diameter of the carrier is 2-100 nm, the specific surface area is not less than 500 square meters per gram, and the chlorine content is not less than 17%;
the preparation method of the cell inflammatory factor adsorbent comprises the following steps:
1) Preparing an alkaline glycine aqueous solution and adding a radical protection reagent;
2) Amination reaction is carried out on the carrier, and an amino connecting arm is introduced;
3) Mixing a carrier added with an amino connecting arm with the alkaline glycine aqueous solution, and adjusting the pH to 4-6 to enable glycine to be obtained
Coupling to a carrier, wherein the reaction time is 8-48 h;
4) Adding a deprotection reagent, wherein the reaction time is 8 hours;
5) Extracting with ethyl acetate for 8h, washing with alcohol, washing with water, and drying;
6) Adding the microspheres into valine solution, adding an activating agent for activation, and reacting for 6 hours;
7) And then washing the microspheres with alcohol and water to prepare the cell inflammatory factor adsorbent.
2. The method for preparing the cytokine adsorbent of claim 1, comprising the steps of:
1) Preparing an alkaline glycine aqueous solution and adding a radical protection reagent;
2) Amination reaction is carried out on the carrier, and an amino connecting arm is introduced;
3) Mixing a carrier added with an amino connecting arm with the alkaline glycine aqueous solution, and adjusting the pH to 4-6 to enable glycine to be obtained
Coupling to a carrier, wherein the reaction time is 8-48 h;
4) Adding a deprotection reagent, wherein the reaction time is 8 hours;
5) Extracting with ethyl acetate for 8h, washing with alcohol, washing with water, and drying;
6) Adding the microspheres into valine solution, adding an activating agent for activation, and reacting for 6 hours;
7) And then washing the microspheres with alcohol and water to prepare the cell inflammatory factor adsorbent.
3. The method of claim 2, wherein the group-protecting agent is chloroformate or its derivative.
4. The method of claim 2, wherein the deprotecting reagent is a mixture of diethyl ether, ethyl acetate and trifluoroacetic acid.
5. The method of preparing an adsorbent for cytokines according to claim 2, wherein the activating reagent is N-hydroxysuccinimide and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride.
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