CN117718016A - Blood perfusion adsorbent, preparation method and application thereof - Google Patents
Blood perfusion adsorbent, preparation method and application thereof Download PDFInfo
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Abstract
The invention belongs to the technical field of biological medicines, and relates to a blood perfusion adsorbent, a preparation method and application thereof. The blood perfusion adsorbent is of a microsphere structure, the carrier of the microsphere is chitosan microsphere, the ligand is I-type collagen, and the particle size of the microsphere is 250-1000 mu m. The blood perfusion adsorbent provided by the invention has the characteristics of large bilirubin adsorption amount, high selective adsorptivity, small albumin adsorption and the like based on the synergistic effect of collagen and chitosan, and has the advantages of excellent bilirubin specific adsorption effect, low cost and good clinical application prospect.
Description
Technical Field
The invention belongs to the technical field of biological medicines, and particularly relates to a blood perfusion adsorbent, a preparation method and application thereof.
Background
Bilirubin is one of the major products of hemoglobin metabolism and is a pathogenic toxin. It binds to albumin and is transported to the liver for final excretion. In the case of patients with liver dysfunction, the metabolic pathway is blocked, and bilirubin cannot be discharged out of the body in time, so that hyperbilirubinemia is caused. Excessive bilirubin can accumulate in various tissues such as the brain, eventually leading to brain damage and even death. Hyperbilirubinemia is generally defined as total bilirubin > 12mg/dL, whereas normal serum bilirubin levels are 0.4-1.8mg/dL, and therefore, it is critical to remove excess bilirubin from the blood of hyperbilirubinemia patients. To address this problem, various techniques such as hemodialysis, phototherapy, plasma exchange, and blood perfusion are used to remove excess bilirubin. Among these techniques, blood perfusion has proven to be one of the most effective methods for bilirubin removal (J. Mater. Chem. B,2017,5 (29): 5763-5773), the core functional unit of which is an adsorbent material. Currently, various bilirubin adsorbents such as activated carbon, chitosan or polystyrene and other high molecular materials and novel materials such as MoFs or PAF have been widely developed. However, these adsorbents exhibit limitations in terms of bilirubin adsorption performance or blood compatibility, limiting their clinical use (ACS Applied Materials & Interfaces,2020 (12): 25546-25556). Therefore, developing a new bilirubin adsorbent is still urgent, and designing a bilirubin adsorbent with high adsorption performance, good blood compatibility, and low cost is not only a great demand for blood perfusion application, but also a great challenge.
Chitosan is a natural high molecular polysaccharide that is produced by deacetylation of chitin. Because of its good biocompatibility and hemocompatibility, it is widely used in the adsorbent field. However, chitosan itself has limited adsorption capacity for free bilirubin and bilirubin in albumin-rich solutions and has a high adsorption capacity for albumin (Colloids Surfaces B: biointerfaces,2013,112 (12): 103-107). Chitosan generally enhances the adsorption capacity for bilirubin by complexing with other inorganic substances or grafted ligands (J. Mater. Chem. B,2022, (10): 8650-8663). However, these inorganic materials have high adsorptivity to components such as albumin in blood, and further improvement in safety and long-term experimental verification have been required (Topics in Current Chemistry,2020,378 (1): 1-41). Bilirubin has the characteristics of negative charge and strong hydrophobicity, and common ligands typically include polylysine, cyclodextrin, and quaternary amine groups, among others (Artificial Organs,1992,16 (6): 568-576;Journal of Applied Polymer Science,2013,130 (1): 563-571). However, these ligands tend to have limited adsorption properties for bilirubin; or adsorption of other components in the blood such as albumin, etc., resulting in poor blood compatibility.
Disclosure of Invention
The invention aims at overcoming the defects of the prior art and provides a blood perfusion adsorbent, a preparation method and application. The blood perfusion adsorbent provided by the invention has the characteristics of large bilirubin adsorption amount, high selective adsorptivity, small albumin adsorption and the like based on the synergistic effect of collagen and chitosan, and has an excellent bilirubin specific adsorption effect.
In order to achieve the above purpose, the invention adopts the following technical scheme:
the first aspect of the present invention provides a hemoperfusion adsorbent which is in a microsphere structure, wherein the carrier of the microsphere is chitosan microsphere, the ligand is type I collagen, the particle size of the microsphere is 250-1000 μm, and the microsphere has the following chemical structure:
wherein,represents chitosan microspheres; x represents a polyamine.
According to the scheme, the hemoperfusion adsorbent takes chitosan microspheres as a carrier after freeze drying and pore making.
According to the scheme, the hemoperfusion adsorbent is prepared by the following method: preparing chitosan microspheres by a reversed-phase suspension polymerization method, and performing freeze drying and pore-forming on the chitosan microspheres to serve as a carrier structure to react with collagen; or after being activated by epoxy chloropropane, the collagen reacts with the epoxy chloropropane; or by reacting polyamine as a spacer with collagen after activation by epichlorohydrin.
According to the scheme, the surface of the blood perfusion adsorbent microsphere is provided with gaps, and preferably, the surface porosity of the blood perfusion adsorbent is 35-50%; the collagen loading on the surface of the chitosan microsphere is 0.5-10mg/g.
According to the scheme, the polyamine is diamine substance with carbon chain length of 2-6, diethylenetriamine or triethylenetetramine; after the chitosan microsphere is activated by epoxy chloropropane, the collagen graft modified chitosan microsphere surface collagen loading capacity obtained by taking polyamine as a spacer arm and reacting with collagen is 2-8mg/g.
According to the scheme, the molecular weight of the collagen is 20kDa-1000kDa.
According to the scheme, the collagen grafting modified chitosan microsphere is a tan sphere, the clearance rate of bilirubin can reach 75-95%, the clearance rate of albumin is less than 3%, and the requirement of clinical albumin clearance rate at present is met, wherein the clearance rate of albumin is less than 15%.
The second aspect of the present invention provides a method for preparing a hemoperfusion adsorbent comprising the steps of:
(1) Preparing chitosan microspheres by an inverse suspension polymerization method;
(2) Freeze drying chitosan microsphere to prepare holes;
(3) Collagen modification is carried out on the chitosan microsphere obtained in the step (2), and the collagen grafting modified chitosan microsphere, namely the blood perfusion adsorbent, is prepared, wherein the collagen modification method is one of the following three methods:
the method comprises the following steps: reacting chitosan microspheres with a collagen solution, and carrying out surface grafting modification;
the second method is as follows: activating chitosan microspheres by epoxy chloropropane to obtain an activated chitosan microsphere carrier; directly reacting the activated chitosan microsphere carrier with a collagen solution;
and a third method: activating chitosan microspheres by epoxy chloropropane to obtain an activated chitosan microsphere carrier; adding polyamine for reaction, washing with water to remove excessive polyamine, transferring to an organic phase containing carbonyl diimidazole for reaction, washing microspheres with the organic phase after the reaction is finished, and then reacting with collagen solution;
(4) And (3) performing post-treatment after the reaction is finished to obtain the collagen grafting modified chitosan microsphere.
According to the scheme, the step (1) is as follows: dissolving chitosan powder into acetic acid to prepare a chitosan/acetic acid solution with the mass fraction of 0.5% -5%, and then pouring the chitosan/acetic acid solution into a dispersed oil phase, wherein the volume ratio of the chitosan acetic acid solution to the oil phase is 1:5-20, dispersing into uniform droplets by adjusting the rotating speed, adding a cross-linking agent, cross-linking for a period of time, collecting microspheres, and processing to obtain chitosan microspheres.
According to the scheme, the rotating speed of the step (1) is 200-500rpm.
According to the scheme, the cross-linking agent in the step (1) is formaldehyde, glutaraldehyde, genipin or tripolyphosphate and the like; the reaction temperature is 40-60 ℃ and the reaction time is 2-6h after the crosslinking agent is added.
According to the scheme, the post-treatment in the step (1) is to sequentially wash with normal hexane, ethanol and water.
According to the scheme, when the chitosan microsphere is frozen, dried and holed, the freezing temperature is between-20 ℃ and 80 ℃ and the freezing time is between 12 hours and 48 hours.
According to the scheme, the collagen is type I collagen, the molecular weight is 20kDa-1000kDa, and the mass fraction of the collagen solution is 0.01-2%.
According to the scheme, in the first method of the step (3), the reaction time of the chitosan microsphere and the collagen solution is 12-36 hours.
According to the scheme, the activated chitosan microsphere carrier in the second or third method of the step (3) is reacted with the collagen solution for 12-36 hours.
According to the scheme, the polyamine in the third method in the step (3) is diamine substance, diethylenetriamine or triethylenetetramine with carbon chain length of 2-6. The reaction after the addition of the polyamine is: reacting at 40-60 deg.c for 2-6 hr.
According to the scheme, the reaction after adding carbonyl diimidazole in the step (3) is carried out for 6-24 hours at room temperature.
According to the scheme, the post-treatment in the step (4) is to sequentially wash with acetic acid aqueous solution and distilled water.
The third aspect of the present invention provides the use of a hemodynamic adsorbent as a hemodynamic filler for preparing a hemodynamic filler, particularly for specifically removing bilirubin from patients suffering from severe jaundice or liver failure.
The hemoperfusion cartridge of the fourth aspect of the present invention is obtained by using the hemoperfusion adsorbent as a packing material for hemoperfusion cartridges.
The invention provides the blood perfusion adsorbent which is based on the synergistic effect of collagen and chitosan, has the characteristics of large bilirubin adsorption amount, high selective adsorptivity, less albumin adsorption and the like, and has excellent bilirubin specific adsorption effect; the cost is low; the chitosan is used as a carrier, and compared with pure collagen microspheres (higher in water content), the collagen microsphere has better mechanical property and high structural stability, can not cause the problems of incapability of normal perfusion use and the like due to structural collapse in the use process, and has good clinical application prospect.
The invention adopts a reverse phase suspension polymerization method to prepare chitosan microspheres, and then the chitosan microspheres are freeze-dried to prepare holes and then are used as a carrier structure to react with collagen; or after being activated by epoxy chloropropane, the collagen reacts with the epoxy chloropropane; or after the epoxy chloropropane is activated, polyamine is used as a spacer arm to react with collagen, so as to obtain the collagen grafting modified chitosan microsphere. The blood perfusion adsorbent prepared by the method is based on the synergistic effect of collagen and chitosan, has the characteristics of large bilirubin adsorption amount, high selective adsorptivity, small albumin adsorption and the like, is obviously superior to chitosan microspheres, collagen modified materials based on other carriers such as polystyrene microspheres and the like, chitosan/collagen materials obtained by other methods such as a direct mixing method and the like, has small use amount of collagen, can greatly reduce the cost (the consumed collagen by the method is about one third of the cost of the preparation by the direct mixing method and one tenth of the cost of the preparation of the collagen microspheres), is suitable for batch production, and has wide application prospect.
The invention has the beneficial effects that:
the blood perfusion adsorbent provided by the invention has the characteristics of large bilirubin adsorption amount, high selective adsorptivity, small albumin adsorption and the like based on the synergistic effect of collagen and chitosan, and has an excellent bilirubin specific adsorption effect; the cost is low; has better mechanical property and high structural stability, the problems of incapability of normal perfusion and the like caused by structural collapse in the use process can be avoided, and the clinical application prospect is good.
The collagen grafting modified chitosan microsphere prepared by the preparation method, namely the blood perfusion adsorbent, is based on the synergistic effect of collagen and chitosan, has the characteristics of large bilirubin adsorption amount, high selective adsorption, small albumin adsorption and the like, is obviously superior to chitosan microsphere, is also superior to collagen modified materials based on other carriers such as polystyrene microsphere and the like, is superior to chitosan/collagen materials obtained by other methods such as a direct mixing method and the like, has less use amount of collagen, can greatly reduce cost, is suitable for batch production, and has wide application prospect.
Drawings
FIG. 1 is an SEM image of CS-Col-3 microspheres prepared in example 3;
figure 2 adsorption kinetics of the adsorbent to bilirubin.
Detailed Description
Example 1
Dissolving chitosan powder in acetic acid to prepare 0.5% chitosan acetic acid solution, and pouring the chitosan acetic acid solution into a dispersed oil phase, wherein the volume ratio of the chitosan acetic acid solution to the oil phase is 1:10, dispersing into uniform small droplets at 200rpm, adding a cross-linking agent formaldehyde, and cross-linking for 4 hours at 40 ℃. Collecting the microspheres, and cleaning with n-hexane, ethanol, water and the like to obtain the chitosan microspheres. And (3) putting the chitosan microspheres into a refrigerator at the temperature of minus 20 ℃ to freeze for 12 hours, and transferring the adsorbent with the formed ice crystals into a vacuum freeze dryer to freeze dry and pore-form. The chitosan microsphere carrier was directly reacted with a 0.01% by mass type I collagen (molecular weight 20 kDa) solution for 24h. Washing with 0.1M acetic acid aqueous solution and distilled water to obtain collagen grafting modified chitosan microsphere with the number CS-Col-1.
Example 2
Dissolving chitosan powder in acetic acid to prepare 2% chitosan acetic acid solution, and pouring the chitosan acetic acid solution into a dispersed oil phase, wherein the volume ratio of the chitosan acetic acid solution to the oil phase is 1:10, dispersing into uniform small liquid drops at 300rpm, adding a cross-linking agent genipin, and cross-linking for 4 hours at 40 ℃. Collecting the microspheres, and cleaning with n-hexane, ethanol, water and the like to obtain the chitosan microspheres. And (3) putting the chitosan microspheres into a refrigerator at the temperature of minus 40 ℃ to freeze for 24 hours, and transferring the adsorbent forming the ice crystals into a vacuum freeze dryer to freeze dry and pore-form. And (3) activating the chitosan microsphere by epoxy chloropropane to obtain an activated chitosan microsphere carrier. The activated chitosan microsphere carrier was reacted with a 0.1% by mass type I collagen (molecular weight 100 kDa) solution for 24 hours. Washing with 0.1M acetic acid aqueous solution and distilled water to obtain collagen grafting modified chitosan microsphere with the number CS-Col-2.
Example 3
On the basis of chitosan epichlorohydrin activation, polyamine such as diamine substances (ethylenediamine, propylenediamine, butylenediamine, pentylene diamine, hexamethylenediamine) with carbon chain length of 2-6 are introduced to react with collagen as a spacer, so that the adsorption rate of bilirubin can be further improved, a better specific adsorption effect is achieved, and the specific experiment using hexamethylenediamine as the spacer is as follows:
dissolving chitosan powder in acetic acid to prepare a 3% chitosan acetic acid solution, and then pouring the chitosan acetic acid solution into a well-dispersed oil phase, wherein the volume ratio of the chitosan acetic acid solution to the oil phase is 1:5, dispersing into uniform small droplets at 300rpm, adding a crosslinking agent glutaraldehyde, and crosslinking for 4 hours at 40 ℃. Collecting the microspheres, and cleaning with n-hexane, ethanol, water and the like to obtain the chitosan microspheres. And (3) putting the chitosan microspheres into a refrigerator at the temperature of minus 80 ℃ to freeze for 36 hours, and transferring the adsorbent with the formed ice crystals into a vacuum freeze dryer to freeze dry and pore-form. And (3) activating the chitosan microsphere by epoxy chloropropane to obtain an activated chitosan microsphere carrier. Adding hexamethylenediamine into the activated chitosan microsphere carrier as a spacer arm, reacting at 60 ℃ for 3 hours, washing the microsphere with distilled water, transferring to an organic phase containing carbonyldiimidazole, reacting at room temperature for 12 hours, washing the microsphere with an organic phase after the reaction is finished, and then reacting with a 1% type I collagen (molecular weight of 300 kDa) solution for 24 hours. Washing with 0.1M acetic acid aqueous solution and distilled water to obtain collagen grafting modified chitosan microsphere with the number CS-Col-3.
The surface morphology of the microspheres after metal spraying was observed by a Scanning Electron Microscope (SEM) using the numbered CS-Col-3 microspheres prepared in example 3. As can be seen from FIG. 1, the prepared collagen grafting modified chitosan microsphere is in a more regular sphere shape, the surface has larger porosity, the existence of collagen can be observed on the surface of the microsphere, and the collagen loading capacity of the CS-Col-3 microsphere surface is about 2.88mg/g.
Application examples
Bilirubin adsorption experiments
Experiment group 1
Taking 30mg of chitosan microsphere (CS), chloromethylated polystyrene microsphere (PS), microsphere (PS-Col) synthesized by grafting collagen with chloromethylated polystyrene microsphere as a carrier and microsphere (CS-Col-3) prepared in the above example 3, adding into a certain amount of high bilirubin solution (200 mg/L), placing into a constant temperature shaking table, oscillating and adsorbing for 3h at 37 ℃, comparing bilirubin concentration difference before and after adsorption, and calculating the adsorption rate by the following formula (1):
BP＝(C 1 -C 2 )/C 1 ×100％ (1)
wherein BP is the adsorption rate (%) of bilirubin, C 1 And C 2 The concentration (mg/L) of bilirubin before and after adsorption, respectively.
The experimental results are shown in table 1, and after 3 hours of adsorption, the results show that the collagen grafting modified chitosan microsphere obtained by using the chitosan microsphere as a collagen grafting modified carrier has excellent bilirubin specific adsorption, and compared with the chitosan before collagen grafting modification, the adsorption rate of the collagen grafting modified chitosan microsphere on bilirubin is improved by about 4 times. And compared with the prior art, the adsorption rate of bilirubin is not changed greatly after collagen is grafted and synthesized by taking chloromethylated polystyrene microspheres as carriers. This indicates that the different carriers directly affect the adsorption effect of the collagen-modified material. The invention provides the collagen grafting modified chitosan microsphere, namely the blood perfusion adsorbent, with excellent bilirubin specific adsorption through preliminary selection of carriers by screening, theoretical simulation, experimental fumbling research and the like, collagen modification based on the carriers and exploration of a modification method.
TABLE 1 adsorption Rate of bilirubin
CS and polystyrene microspheres were synthesized by the same inverse suspension polymerization method as in example 3, and then PS-Col microspheres were synthesized by the same graft collagen method as in example 3 after chloromethylation (PS) of the polystyrene microspheres.
Experiment group 2
Taking chitosan microsphere (CS), directly mixing to synthesize chitosan/collagen microsphere (CS/Col) and 30mg of microsphere prepared in the above example, adding into a certain amount of high bilirubin solution (200 mg/L), placing into a constant temperature shaking table, oscillating and adsorbing at 37deg.C for 3 hr, comparing bilirubin concentration difference before and after adsorption, and calculating bilirubin adsorption rate.
The experimental results are shown in table 2, after 3h of adsorption, the adsorption rate of the chitosan microsphere (CS) to bilirubin is 25.9% (basically the adsorption rate of the chitosan microsphere to bilirubin in the report of the literature is the same), and the adsorption rate of the chitosan/collagen microsphere synthesized by the direct mixing method to bilirubin is 45.0%, which shows that the adsorption effects of the collagen grafting modified chitosan microsphere prepared by grafting collagen by different methods to bilirubin are far different, and the adsorption rate of the collagen grafting modified chitosan microsphere provided by the embodiment of the invention to bilirubin is more than 75%, which shows that the collagen grafting modified chitosan microsphere provided by the invention has excellent adsorption performance to bilirubin. Wherein, the absorption rate of CS-Col-3 microsphere to bilirubin is highest and reaches 92.1%, which is probably because the introduction of the spacer arm can reduce steric hindrance to the greatest extent, increase the accessibility of ligand collagen, and graft more collagen, thereby improving the absorption performance to bilirubin based on the synergistic effect of chitosan-collagen. Meanwhile, the CS-Col-3 microsphere with the same quality can achieve bilirubin adsorption rate (the bilirubin adsorption rate of the collagen microsphere (Col) is 92.9%) which is equivalent to that of the collagen microsphere, but the collagen consumed by the method is about one tenth of the cost for preparing the collagen microsphere, so that the cost is greatly reduced.
Table 2 adsorption rate of bilirubin from experimental group 2
Chitosan and collagen were mixed according to 5:1 (mass ratio) and then synthesizing chitosan/collagen microspheres (CS/Col) by the same inverse suspension polymerization method as in the above-mentioned example 3.
Albumin adsorption experiments
Taking chitosan microspheres (CS), directly mixing to synthesize chitosan/collagen microspheres (CS/Col) and the microspheres prepared in the embodiment, adding the chitosan/collagen microspheres (CS/Col) and the microspheres into a certain amount of albumin solution, placing the albumin solution into a constant temperature shaking table, carrying out shaking adsorption at 37 ℃ for 3 hours, comparing the concentration difference of albumin before and after adsorption, and calculating the adsorption rate of albumin by the following formula (2):
AP＝(C 3 -C 4 )/C 3 ×100％ (2)
wherein AP is an adsorption rate (%) of albumin, C 3 And C 4 The concentration of albumin before and after adsorption (mg/mL) was determined.
The experimental results are shown in table 3, the adsorption rate of the chitosan microsphere to albumin is 7.8%, the adsorption rate of the chitosan/collagen microsphere synthesized by the direct mixing method is 4.5%, and the adsorption rate of the microsphere of the example to albumin is 2.3%, and the adsorption rate is reduced by more than 3 times, which shows that the collagen grafting modified chitosan microsphere has extremely low adsorption performance to albumin and has specific adsorption to bilirubin. This is probably because collagen on chitosan microspheres affects the properties of the chitosan surface, and the effect prevents further adsorption of albumin on the chitosan microsphere surface. These results demonstrate that the collagen graft modified chitosan microsphere not only can improve the adsorption performance of bilirubin, but also can reduce the adsorption of albumin, thus reflecting the synergistic effect of collagen and chitosan.
TABLE 3 adsorption Rate of albumin
Adsorption kinetics experiments
Taking 30mg of chitosan microsphere (CS), directly mixing to synthesize chitosan/collagen microsphere (CS/Col) and microsphere (CS-Col-3) prepared in the above example, adding into a certain amount of high bilirubin solution (200 mg/L), placing into a constant temperature shaking table, oscillating and adsorbing at 37deg.C, and sampling at different intervals.
As shown in FIG. 2, the CS-Col-3 microsphere prepared in the example rapidly adsorbed bilirubin within the first 30 minutes and reached equilibrium within 120 minutes, as compared with the chitosan microsphere and the chitosan/collagen microsphere synthesized by the direct mixing method, at which time the adsorption rate was as high as 90.3%. The collagen grafting modified chitosan microsphere can improve the adsorption efficiency of bilirubin, reduce the time for reaching equilibrium, and the rapid adsorption process can be basically consistent with that of pure collagen microsphere.
In conclusion, the invention provides the collagen grafting modified chitosan microsphere, namely the blood perfusion adsorbent, which has the characteristics of large bilirubin adsorption amount, high selective adsorptivity, less albumin adsorption and the like based on the synergistic effect of collagen and chitosan, and has excellent bilirubin specific adsorption effect, through a great number of experimental groping researches including the preliminary selection of carriers such as the theoretical simulation research of the collagen space conformation retention under different carriers, the specific experimental groping research and the like, then carrying out collagen modification based on the carriers, and combining with the exploration of a modification method; the cost is low; has better mechanical property and high structural stability, the problems of incapability of normal perfusion and the like caused by structural collapse in the use process can be avoided, and the clinical application prospect is good.
Claims (12)
1. The blood perfusion adsorbent is characterized in that: the microsphere has a microsphere structure, the carrier of the microsphere is chitosan microsphere, the ligand is collagen I, the particle size of the microsphere is 250-1000 mu m, and the microsphere has the following chemical structure:
wherein,represents chitosan microspheres; x represents a polyamine.
2. The hemodynamic sorbent of claim 1, wherein: the preparation method comprises the following steps: preparing chitosan microspheres by a reversed-phase suspension polymerization method, and performing freeze drying and pore-forming on the chitosan microspheres to serve as a carrier structure to react with collagen; or after being activated by epoxy chloropropane, the collagen reacts with the epoxy chloropropane; or the collagen grafting modified chitosan microsphere is obtained by taking polyamine as a spacer and reacting with collagen after the epoxy chloropropane is activated.
3. The hemodynamic sorbent of claim 1, wherein: the surface porosity of the blood perfusion adsorbent is 35-50%; the collagen loading on the surface of the chitosan microsphere is 0.5-10mg/g.
4. The hemodynamic sorbent of claim 1, wherein: the polyamine is diamine substance with carbon chain length of 2-6, diethylenetriamine or triethylenetetramine; after the chitosan microsphere is activated by epoxy chloropropane, the collagen graft modified chitosan microsphere surface collagen loading capacity obtained by taking polyamine as a spacer arm and reacting with collagen is 2-8mg/g.
5. The hemodynamic sorbent of claim 1, wherein: the molecular weight of the type I collagen is 20kDa-1000kDa.
6. The hemodynamic sorbent of claim 1, wherein: the clearance rate of the collagen grafting modified chitosan microsphere to bilirubin is 75-95%, and the clearance rate of the collagen grafting modified chitosan microsphere to albumin is less than 3%.
7. The method for preparing the hemoperfusion adsorbent of claim 1, wherein: the method comprises the following steps:
(1) Preparing chitosan microspheres by an inverse suspension polymerization method;
(2) Freeze drying chitosan microsphere to prepare holes;
(3) Collagen modification is carried out on the chitosan microsphere obtained in the step (2), and the collagen grafting modified chitosan microsphere, namely the blood perfusion adsorbent, is prepared, wherein the collagen modification method is one of the following three methods:
the method comprises the following steps: reacting chitosan microspheres with a collagen solution, and carrying out surface grafting modification;
the second method is as follows: activating chitosan microspheres by epoxy chloropropane to obtain an activated chitosan microsphere carrier; directly reacting the activated chitosan microsphere carrier with a collagen solution;
and a third method: activating chitosan microspheres by epoxy chloropropane to obtain an activated chitosan microsphere carrier; adding polyamine for reaction, washing with water to remove excessive polyamine, transferring to an organic phase containing carbonyl diimidazole for reaction, washing microspheres with the organic phase after the reaction is finished, and then reacting with collagen solution;
(4) And after the reaction is finished, carrying out post-treatment to obtain the collagen grafting modified chitosan microsphere, namely the blood perfusion adsorbent.
8. The method of manufacturing according to claim 7, wherein: the step (1) is as follows: dissolving chitosan powder into acetic acid to prepare a chitosan/acetic acid solution with the mass fraction of 0.5% -5%, and then pouring the chitosan/acetic acid solution into a dispersed oil phase, wherein the volume ratio of the chitosan acetic acid solution to the oil phase is 1:5-20, dispersing into uniform droplets by adjusting the rotating speed, adding a cross-linking agent, cross-linking for a period of time, collecting microspheres, and processing to obtain chitosan microspheres.
9. The method of manufacturing according to claim 8, wherein:
the rotating speed of the step (1) is 200-500rpm; the cross-linking agent in the step (1) is formaldehyde, glutaraldehyde, genipin or tripolyphosphate, and the reaction temperature is 40-60 ℃ and the reaction time is 2-6h after the cross-linking agent is added.
10. The method of manufacturing according to claim 7, wherein:
the molecular weight of the type I collagen is 20kDa-1000kDa, and the mass fraction of the collagen solution is 0.01-2%;
when the chitosan microsphere is frozen and dried to prepare holes, the freezing temperature is between-20 ℃ and 80 ℃ and the freezing time is between 12 and 48 hours;
the chitosan microsphere carrier activated in the first method or the second method or the third method of the step (3) reacts with the collagen solution for 12-36 hours;
the polyamine in the third method in the step (3) is diamine substance, diethylenetriamine or triethylenetetramine with carbon chain length of 2-6; the reaction after the addition of the polyamine is: reacting for 2-6h at 40-60 ℃; the reaction after adding carbonyl diimidazole is carried out for 6-24 hours at room temperature.
11. Use of the hemodynamic adsorbent of claim 1 as a hemodynamic filler for preparing an adsorption column in a hemodynamic filler.
12. A hemodiafiltration device obtained by using the hemodiafiltration adsorbent of claim 1 as a hemodiafiltration device filler.
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