Blood perfusion adsorbents, methods of preparation and use thereof

Abstract

The application belongs to the technical field of biological medicine, and relates to a hemoperfusion adsorbent, a preparation method and application thereof. The hemoperfusion adsorbent is a microsphere structure, the carrier of the microsphere is a chitosan microsphere, the ligand is type I collagen, and the particle size of the microsphere is 250-1000 mu m. The hemoperfusion adsorbent provided in the application has the characteristics of large adsorption amount of bilirubin, high selective adsorption, small adsorption of albumin and the like based on the synergistic effect of collagen and chitosan, has excellent specific adsorption effect of bilirubin, is low in cost, and has good clinical application prospect.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a blood perfusion adsorbent, its preparation method, and its application. Background Technology

[0002] Bilirubin is one of the main products of hemoglobin metabolism and is a pathogenic toxin. It binds to albumin and is transported to the liver for excretion. However, in patients with liver dysfunction, due to abnormal liver function and obstructed metabolic pathways, bilirubin cannot be excreted in time, leading to hyperbilirubinemia. Excessive bilirubin accumulates in various tissues such as the brain, eventually causing brain damage or even death. Generally, hyperbilirubinemia is defined as total bilirubin >12 mg / dL, while the normal serum bilirubin level is 0.4-1.8 mg / dL. Therefore, it is crucial to remove excess bilirubin from the blood of patients with hyperbilirubinemia. To address this issue, various techniques such as hemodialysis, phototherapy, plasma exchange, and hemoperfusion have been used to remove excess bilirubin. Among these techniques, hemoperfusion has been proven to be one of the most effective methods for removing bilirubin (J. Mater. Chem. B, 2017, 5(29): 5763-5773), and its core functional unit is the adsorbent material. Currently, various bilirubin adsorbents, such as activated carbon, chitosan, or polystyrene, as well as novel materials like MoFs or PAFs, have been widely developed. However, these adsorbents exhibit limitations in bilirubin adsorption performance or blood compatibility, restricting their clinical application (ACS Applied Materials & Interfaces, 2020(12): 25546-25556). Therefore, developing novel bilirubin adsorbents remains an urgent priority. Designing bilirubin adsorbents with high adsorption performance, good blood compatibility, and low cost is not only a huge demand for hemoperfusion applications but also a significant challenge.

[0003] Chitosan is a natural high-molecular-weight polysaccharide produced by the deacetylation of chitin. Due to its good biocompatibility and blood compatibility, it is widely used in the field of adsorbents. However, chitosan itself has limited adsorption capacity for free bilirubin and bilirubin in albumin-rich solutions, and has a higher adsorption capacity for albumin (Colloids Surfaces B: Biointerfaces, 2013, 112(12): 103-107). Chitosan is usually enhanced to enhance its adsorption capacity for bilirubin by combining it with other inorganic substances or grafting ligands (J. Mater. Chem. B, 2022, (10): 8650-8663). However, these inorganic materials have high adsorption capacity for components such as albumin in blood, and their safety needs to be further improved and verified by long-term experiments (Topics in Current Chemistry, 2020, 378(1): 1-41). Bilirubin is negatively charged and strongly hydrophobic, and common ligands typically include polylysine, cyclodextrin, and quaternary ammonium groups (Artificial Organs, 1992, 16(6):568-576; Journal of Applied Polymer Science, 2013, 130(1):563-571). However, these ligands often have limited adsorption capacity for bilirubin; or they have high adsorption capacity for other components in the blood, such as albumin, resulting in poor blood compatibility. Summary of the Invention

[0004] The purpose of this invention is to address the shortcomings of existing technologies by providing a hemoperfusion adsorbent, its preparation method, and its application. The hemoperfusion adsorbent provided by this invention, based on the synergistic effect of collagen and chitosan, exhibits characteristics such as high bilirubin adsorption capacity and high selective adsorption, while showing low albumin adsorption, thus demonstrating excellent bilirubin-specific adsorption performance.

[0005] To achieve the above-mentioned objectives, the technical solution adopted by the present invention is as follows:

[0006] The first aspect of this invention provides a blood perfusion adsorbent, which has a microsphere structure. The carrier of the microspheres is chitosan microspheres, the ligand is type I collagen, and the particle size of the microspheres is 250-1000 μm. It has the following chemical structure:

[0007]

[0008] in, X represents chitosan microspheres; X represents polyamines.

[0009] According to the above scheme, the blood perfusion adsorbent uses chitosan microspheres freeze-dried and pore-forming as a carrier.

[0010] According to the above scheme, the blood perfusion adsorbent is prepared by the following methods: chitosan microspheres are prepared by reverse suspension polymerization, and the chitosan microspheres are freeze-dried to form pores and then used as a carrier structure to react with collagen; or they are activated by epichlorohydrin and then reacted with collagen; or they are activated by epichlorohydrin and then reacted with collagen using polyamines as spacers.

[0011] According to the above scheme, the surface of the blood perfusion adsorbent microspheres has pores. Preferably, the porosity of the blood perfusion adsorbent surface is 35-50%; the collagen loading on the surface of the chitosan microspheres is 0.5-10 mg / g.

[0012] According to the above scheme, the polyamine is a diamine with a carbon chain length of 2-6, diethylenetriamine, or triethylenetetramine; the collagen loading on the surface of the chitosan microspheres obtained by reacting the chitosan microspheres with collagen grafting after activation by epichlorohydrin and using the polyamine as a spacer arm is 2-8 mg / g.

[0013] According to the above scheme, the molecular weight of the collagen is 20kDa-1000kDa.

[0014] According to the above scheme, the collagen-grafted modified chitosan microspheres are brownish-brown spheres with a bilirubin clearance rate of 75-95% and an albumin clearance rate of less than 3%, meeting the current clinical requirement of an albumin clearance rate of less than 15%.

[0015] A second aspect of this invention provides a method for preparing a blood perfusion adsorbent, comprising the following steps:

[0016] (1) Preparation of chitosan microspheres by reverse suspension polymerization;

[0017] (2) Pore formation was achieved by freeze-drying chitosan microspheres;

[0018] (3) The chitosan microspheres obtained in step (2) are modified with collagen to prepare collagen-grafted modified chitosan microspheres, which are blood perfusion adsorbents. The collagen modification method is one of the following three methods:

[0019] Method 1: React chitosan microspheres with collagen solution to perform surface grafting modification;

[0020] Method 2: Chitosan microspheres are activated with epichlorohydrin to obtain activated chitosan microsphere carriers; the activated chitosan microsphere carriers are then reacted directly with a collagen solution.

[0021] Method 3: Chitosan microspheres are activated with epichlorohydrin to obtain activated chitosan microsphere carriers; after adding polyamines and reacting, the microspheres are washed with water to remove excess polyamines and then transferred to an organic phase containing carbonyl diimidazole for reaction. After the reaction is completed, the microspheres are washed with the organic phase and then reacted with collagen solution.

[0022] (4) After the above reaction is completed, post-processing is performed to obtain collagen-grafted modified chitosan microspheres.

[0023] According to the above scheme, step (1) is as follows: dissolve chitosan powder in acetic acid to prepare a chitosan / acetic acid solution with a mass fraction of 0.5%-5%. Then pour the chitosan / acetic acid solution into the dispersed oil phase, wherein the volume ratio of chitosan / acetic acid solution to oil phase is 1:5-20. After dispersing into uniform small droplets by adjusting the rotation speed, add a crosslinking agent. After crosslinking for a period of time, collect the microspheres and then process them to obtain chitosan microspheres.

[0024] According to the above scheme, the rotational speed in step (1) is 200-500 rpm.

[0025] According to the above scheme, the crosslinking agent in step (1) is formaldehyde, glutaraldehyde, genipin or tripolyphosphate, etc.; after adding the crosslinking agent, the reaction temperature is 40-60℃ and the reaction time is 2-6h.

[0026] According to the above scheme, the post-processing of step (1) is to wash with hexane, ethanol and water in sequence.

[0027] According to the above scheme, when freeze-drying the chitosan microspheres to create pores, the freezing temperature is -20 to 80°C, and the time is 12 to 48 hours.

[0028] According to the above scheme, the collagen is type I collagen with a molecular weight of 20kDa-1000kDa and a collagen solution mass fraction of 0.01-2%.

[0029] According to the above scheme, in step (3) of method one, the reaction time between chitosan microspheres and collagen solution is 12-36h.

[0030] According to the above scheme, the activated chitosan microsphere carrier described in method two or method three of step (3) reacts with the collagen solution for 12-36 hours.

[0031] According to the above scheme, the polyamine in step (3) of method three is a diamine with a carbon chain length of 2-6, diethylenetriamine, or triethylenetetramine. The reaction after adding the polyamine is: react at 40-60℃ for 2-6 hours.

[0032] According to the above scheme, the reaction after adding carbonyl diimidazole in step (3) is a reaction at room temperature for 6-24 hours.

[0033] According to the above scheme, the post-processing of step (4) is to wash with acetic acid aqueous solution and distilled water in sequence.

[0034] The third aspect of this invention provides the application of a hemoperfusion adsorbent as a filler for preparing a hemoperfusion device, specifically for the specific removal of bilirubin from patients with severe jaundice or liver failure.

[0035] The fourth aspect of the present invention relates to a blood perfusion device obtained by using a blood perfusion adsorbent as a filler in a blood perfusion device.

[0036] This invention, based on extensive research including theoretical simulations of collagen spatial conformation maintenance under different carriers and specific experimental explorations, initially selected carriers. Subsequently, collagen modification was performed on these carriers, and combined with the exploration of modification methods, ultimately providing the hemoperfusion adsorbent of this invention. Based on the synergistic effect of collagen and chitosan, it exhibits high adsorption capacity and selectivity for bilirubin, and low adsorption of albumin, demonstrating excellent bilirubin-specific adsorption effect. It is also low in cost. Using chitosan as a carrier, compared to pure collagen microspheres (which have a higher water content), it possesses better mechanical properties and high structural stability, avoiding problems such as structural collapse during use that prevent normal perfusion. It shows promising clinical application prospects.

[0037] This invention employs a reverse suspension polymerization method to prepare chitosan microspheres. The chitosan microspheres are then freeze-dried to create pores and used as a carrier structure for reaction with collagen; alternatively, they are activated with epichlorohydrin before reacting with collagen; or activated with epichlorohydrin and then reacted with collagen using a polyamine as a spacer arm, resulting in collagen-grafted modified chitosan microspheres. The blood perfusion adsorbent prepared by this method, based on the synergistic effect of collagen and chitosan, exhibits high adsorption capacity and selectivity for bilirubin, and low adsorption for albumin. This is significantly superior to chitosan microspheres, as well as collagen-modified materials based on other carriers such as polystyrene microspheres, and chitosan / collagen raw materials obtained by other methods such as direct mixing. Furthermore, it requires less collagen, greatly reducing costs (the collagen consumption of this method is approximately one-third of the cost of the direct mixing method and one-tenth of the cost of preparing collagen microspheres), making it suitable for mass production and with broad application prospects.

[0038] The beneficial effects of this invention are:

[0039] The hemoperfusion adsorbent provided by this invention is based on the synergistic effect of collagen and chitosan. It has the characteristics of large adsorption capacity and high selective adsorption of bilirubin, and low adsorption of albumin, and has excellent bilirubin-specific adsorption effect. It is low in cost, has good mechanical properties, and high structural stability. It will not have problems such as failure to perfusion due to structural collapse during use, and has good prospects for clinical application.

[0040] The collagen-grafted modified chitosan microspheres prepared by the method of this invention, which are blood perfusion adsorbents, have the characteristics of large adsorption capacity and high selective adsorption of bilirubin and low adsorption of albumin based on the synergistic effect of collagen and chitosan. They are significantly superior to chitosan microspheres, as well as collagen-modified materials based on other carriers such as polystyrene microspheres. They are also superior to chitosan / gel raw materials obtained by other methods such as direct mixing. Furthermore, they require less collagen, which can greatly reduce costs. They are suitable for mass production and have broad application prospects. Attached Figure Description

[0041] Figure 1 SEM image of CS-Col-3 microspheres prepared in Example 3;

[0042] Figure 2 Adsorption kinetics of bilirubin by the adsorbent. Detailed Implementation

[0043] Example 1

[0044] Chitosan powder was dissolved in acetic acid to prepare a 0.5% chitosan-acetic acid solution. This solution was then poured into a well-dispersed oil phase at a volume ratio of 1:10. The mixture was dispersed into uniform droplets at 200 rpm, and then formaldehyde, a crosslinking agent, was added. Crosslinking was carried out at 40°C for 4 hours. Microspheres were collected and washed with hexane, ethanol, and water to obtain chitosan microspheres. The chitosan microspheres were frozen at -20°C for 12 hours. The adsorbent that formed ice crystals was transferred to a vacuum freeze dryer for freeze-drying to create pores. The chitosan microsphere carrier was directly reacted with a 0.01% (w / w) solution of type I collagen (molecular weight 20 kDa) for 24 hours. The microspheres were washed with 0.1M acetic acid aqueous solution and distilled water to obtain collagen-grafted modified chitosan microspheres, designated CS-Col-1.

[0045] Example 2

[0046] Chitosan powder was dissolved in acetic acid to prepare a 2% chitosan-acetic acid solution. This solution was then poured into a well-dispersed oil phase at a volume ratio of 1:10. The mixture was dispersed into uniform droplets at 300 rpm, and then genipin, a crosslinking agent, was added. Crosslinking was carried out at 40°C for 4 hours. Microspheres were collected and washed with hexane, ethanol, and water to obtain chitosan microspheres. The chitosan microspheres were frozen at -40°C for 24 hours, and the adsorbent that formed ice crystals was transferred to a vacuum freeze dryer for freeze-drying to create pores. The chitosan microspheres were activated with epichlorohydrin to obtain activated chitosan microsphere carriers. The activated chitosan microsphere carriers were reacted with a 0.1% (w / w) solution of type I collagen (molecular weight 100 kDa) for 24 hours. The microspheres were washed with 0.1M acetic acid aqueous solution and distilled water to obtain collagen-grafted modified chitosan microspheres, designated CS-Col-2.

[0047] Example 3

[0048] Based on the activation of chitosan with epichlorohydrin, introducing polyamines such as diamines with carbon chain lengths of 2-6 (ethylenediamine, propylenediamine, butylenediamine, pentanediamine, hexamethylenediamine), diethylenetriamine, and triethylenetetramine as spacers, and reacting them with collagen, can further enhance the adsorption rate of bilirubin and achieve better specific adsorption effects. Specific experiments using hexamethylenediamine as a spacer are as follows:

[0049] Chitosan powder was dissolved in acetic acid to prepare a 3% chitosan-acetic acid solution. This solution was then poured into a well-dispersed oil phase at a volume ratio of 1:5. The mixture was dispersed into uniform small droplets at 300 rpm, and then glutaraldehyde, a crosslinking agent, was added. Crosslinking was performed at 40°C for 4 hours. The microspheres were collected and washed with hexane, ethanol, and water to obtain chitosan microspheres. The chitosan microspheres were then frozen at -80°C for 36 hours. The adsorbent that formed ice crystals was transferred to a vacuum freeze dryer for freeze-drying to create pores. The chitosan microspheres were then activated with epichlorohydrin to obtain an activated chitosan microsphere carrier. Activated chitosan microspheres were added to hexamethylenediamine as spacers and reacted at 60°C for 3 h. The microspheres were then washed with distilled water and transferred to an organic phase containing carbonyl diimidazole, where they were reacted at room temperature for 12 h. After the reaction was complete, the microspheres were washed again with the organic phase and then reacted with a 1% (w / w) solution of type I collagen (molecular weight 300 kDa) for 24 h. The microspheres were then washed with 0.1 M acetic acid aqueous solution and distilled water to obtain collagen-grafted modified chitosan microspheres, designated CS-Col-3.

[0050] The surface morphology of the gold-sprayed microspheres, numbered CS-Col-3, prepared in Example 3, was observed using a scanning electron microscope (SEM). Figure 1 As we can see, the collagen-grafted modified chitosan microspheres prepared are relatively regular spheres with a large porosity on the surface, and collagen can be observed on the surface of the microspheres. The collagen loading on the surface of CS-Col-3 microspheres is about 2.88 mg / g.

[0051] Application Examples

[0052] Bilirubin adsorption experiment

[0053] Experimental group 1

[0054] Take 30 mg each of chitosan microspheres (CS), chloromethylated polystyrene microspheres (PS), microspheres synthesized by grafting collagen onto chloromethylated polystyrene microspheres (PS-Col), and microspheres prepared in Example 3 above (CS-Col-3), add them to a certain amount of high bilirubin solution (200 mg / L), place them in a constant temperature shaker, and shake and adsorb at 37°C for 3 h. Compare the difference in bilirubin concentration before and after adsorption. The adsorption rate is calculated by the following formula (1):

[0055] BP=(C1-C2) / C1×100% (1)

[0056] In the formula, BP is the adsorption rate of bilirubin (%), and C1 and C2 are the concentrations of bilirubin before and after adsorption (mg / L), respectively.

[0057] The experimental results are shown in Table 1. After 3 hours of adsorption, the results show that the chitosan microspheres used in this invention as the collagen-grafted modified carrier exhibit excellent bilirubin-specific adsorption. Compared with the unmodified chitosan, the adsorption rate of bilirubin by the collagen-grafted modified chitosan microspheres is nearly four times higher. However, after collagen grafting onto chloromethylated polystyrene microspheres, the adsorption rate of bilirubin did not change significantly compared with that before grafting. This indicates that different carriers directly affect the adsorption effect of the collagen-modified material. This invention initially selected carriers through screening, including theoretical simulation and experimental exploration. Then, collagen modification was carried out based on these carriers, and combined with the exploration of modification methods, ultimately providing the collagen-grafted modified chitosan microspheres with excellent bilirubin-specific adsorption, i.e., a blood perfusion adsorbent.

[0058] Table 1. Adsorption rate of bilirubin

[0059]

[0060] CS and polystyrene microspheres were synthesized using the same reverse suspension polymerization method as in Example 3 above. Then, the polystyrene microspheres were chloromethylated (PS) and then PS-Col microspheres were synthesized using the same collagen grafting method as in Example 3.

[0061] Experimental group 2

[0062] Take 30 mg each of chitosan microspheres (CS), chitosan / collagen microspheres (CS / Col) synthesized by direct mixing method, and microspheres prepared in the above examples, add them to a certain amount of high bilirubin solution (200 mg / L), place them in a constant temperature shaker, and shake at 37°C for 3 h to adsorb. Compare the difference in bilirubin concentration before and after adsorption, and calculate the bilirubin adsorption rate.

[0063] The experimental results are shown in Table 2. After 3 hours of adsorption, the adsorption rate of bilirubin by chitosan microspheres (CS) was 25.9% (which is basically consistent with the adsorption rate of bilirubin by chitosan microspheres reported in the literature). The adsorption rate of bilirubin by chitosan / collagen microspheres synthesized by the direct mixing method was 45.0%. This indicates that the adsorption effect of collagen-grafted modified chitosan microspheres prepared by grafting collagen with different methods on bilirubin varies greatly. However, the adsorption rate of bilirubin by the microspheres in the embodiments of this invention is >75%, which shows that the collagen-grafted modified chitosan microspheres provided by this invention have excellent adsorption performance for bilirubin. Among them, CS-Col-3 microspheres have the highest adsorption rate of bilirubin, reaching 92.1%. This may be because the introduction of the spacer arm can minimize steric hindrance, increase the accessibility of the ligand collagen, and graft more collagen, thereby improving the adsorption performance of bilirubin based on the synergistic effect of chitosan-collagen. Meanwhile, CS-Col-3 microspheres of the same mass can achieve a bilirubin adsorption rate comparable to that of collagen microspheres (the adsorption rate of bilirubin for collagen microspheres (Col) is 92.9%), but this method consumes about one-tenth of the collagen required to prepare collagen microspheres, which greatly reduces costs.

[0064] Table 2. Adsorption rate of bilirubin in experimental group 2

[0065]

[0066] Chitosan and collagen were directly mixed at a mass ratio of 5:1, and then chitosan / collagen microspheres (CS / Col) were synthesized by the same reverse suspension polymerization method as in Example 3 above.

[0067] albumin adsorption experiment

[0068] Chitosan microspheres (CS), chitosan / collagen microspheres (CS / Col) synthesized by direct mixing method, and microspheres prepared in the above examples were added to a certain amount of albumin solution and placed in a constant temperature shaker. The mixture was shaken and adsorbed at 37°C for 3 hours. The difference in albumin concentration before and after adsorption was compared. The adsorption rate of albumin was calculated by the following formula (2):

[0069] AP=(C3-C4) / C3×100% (2)

[0070] In the formula, AP is the adsorption rate of albumin (%), and C3 and C4 are the concentrations of albumin before and after adsorption (mg / mL), respectively.

[0071] The experimental results are shown in Table 3. After 3 hours of adsorption, the adsorption rate of albumin by chitosan microspheres was 7.8%, while that of chitosan / collagen microspheres synthesized by the direct mixing method was 4.5%. In contrast, the adsorption rate of albumin by the microspheres in the example was only 2.3%, a reduction of more than three times. This indicates that the collagen-grafted chitosan microspheres have extremely low albumin adsorption performance but exhibit specific adsorption of bilirubin. This may be because the collagen on the chitosan microspheres affects the properties of the chitosan surface, thus preventing further albumin adsorption on the surface of the chitosan microspheres. These results demonstrate that collagen-grafted chitosan microspheres not only improve the adsorption performance for bilirubin but also reduce the adsorption of albumin, reflecting the synergistic effect of collagen and chitosan.

[0072] Table 3. Adsorption rate of albumin

[0073]

[0074] Adsorption kinetics experiment

[0075] Take 30 mg each of chitosan microspheres (CS), chitosan / collagen microspheres (CS / Col) synthesized by direct mixing method, and microspheres (CS-Col-3) prepared in the above example, add them to a certain amount of high bilirubin solution (200 mg / L), place them in a constant temperature shaker, and shake and adsorb at 37°C. Take samples at different time intervals.

[0076] Experimental results are as follows Figure 2 As shown, compared with chitosan microspheres and chitosan / collagen microspheres synthesized by direct mixing, the CS-Col-3 microspheres prepared in the examples exhibited rapid adsorption of bilirubin within the first 30 minutes and reached equilibrium within 120 minutes, at which point the adsorption rate reached as high as 90.3%. This indicates that collagen-grafted modified chitosan microspheres can improve the adsorption efficiency of bilirubin and reduce the time to reach adsorption equilibrium. This rapid adsorption process is essentially consistent with that of pure collagen microspheres.

[0077] In summary, this invention, through extensive experimental research including theoretical simulation studies on collagen spatial conformation maintenance under different carriers and specific experimental explorations, initially selected carriers, then modified collagen based on these carriers, and explored modification methods, ultimately providing collagen-grafted modified chitosan microspheres with excellent bilirubin-specific adsorption, i.e., a blood perfusion adsorbent. Based on the synergistic effect of collagen and chitosan, it has the characteristics of large adsorption capacity and high selective adsorption of bilirubin, and low adsorption of albumin, exhibiting excellent bilirubin-specific adsorption effect; low cost; good mechanical properties; high structural stability, and will not cause problems such as failure to perfusion due to structural collapse during use, showing good prospects for clinical application.

Claims

1. A blood perfusion adsorbent, characterized in that: It has a microsphere structure, with chitosan microspheres as the carrier and type I collagen as the ligand. The microspheres have a particle size of 250-1000 μm and have the following chemical structure: in, X represents chitosan microspheres; X represents polyamines.

2. The blood perfusion adsorbent according to claim 1, characterized in that: The method used to prepare chitosan microspheres is as follows: chitosan microspheres are prepared by reverse suspension polymerization. After freeze-drying and pore-forming, the chitosan microspheres are used as a carrier structure and activated by epichlorohydrin to obtain activated chitosan microsphere carriers. Polyamine is added to react, and the microspheres are washed with water to remove excess polyamine. The microspheres are then transferred to an organic phase containing carbonyl diimidazole for reaction. After the reaction is completed, the microspheres are washed with the organic phase and then reacted with collagen solution to obtain collagen-grafted modified chitosan microspheres.

3. The blood perfusion adsorbent according to claim 1, characterized in that: The surface porosity of the blood perfusion adsorbent is 35-50%; the collagen loading on the surface of the chitosan microspheres is 0.5-10 mg / g.

4. The blood perfusion adsorbent according to claim 1, characterized in that: The polyamine is a diamine with a carbon chain length of 2-6, diethylenetriamine, or triethylenetetramine; the collagen loading on the surface of the chitosan microspheres is 2-8 mg / g.

5. The blood perfusion adsorbent according to claim 1, characterized in that: The molecular weight of the type I collagen is 20 kDa-1000 kDa.

6. The blood perfusion adsorbent according to claim 1, characterized in that: Collagen-grafted modified chitosan microspheres have a bilirubin clearance rate of 75-95% and a albumin clearance rate of less than 3%.

7. The method for preparing the blood perfusion adsorbent according to claim 1, characterized in that: Includes the following steps: (1) Preparation of chitosan microspheres by reverse suspension polymerization; (2) Create pores by freeze-drying chitosan microspheres; (3) The chitosan microspheres obtained in step (2) are modified with collagen. The collagen modification method is as follows: the chitosan microspheres are activated with epichlorohydrin to obtain activated chitosan microsphere carriers; after adding polyamines, the microspheres are washed with water to remove excess polyamines, and then transferred to an organic phase containing carbonyl diimidazole for reaction. After the reaction is completed, the microspheres are washed with the organic phase and then reacted with collagen solution. (4) After the above reaction is completed, post-processing is performed to obtain collagen-grafted modified chitosan microspheres, i.e., blood perfusion adsorbent.

8. The preparation method according to claim 7, characterized in that: Step (1) is as follows: Chitosan powder is dissolved in acetic acid to prepare a chitosan / acetic acid solution with a mass fraction of 0.5%-5%. Then, the chitosan / acetic acid solution is poured into the dispersed oil phase, wherein the volume ratio of chitosan / acetic acid solution to oil phase is 1:5-20. After dispersing into uniform small droplets by adjusting the rotation speed, a crosslinking agent is added. After crosslinking for a period of time, the microspheres are collected and then processed to obtain chitosan microspheres.

9. The preparation method according to claim 8, characterized in that: The rotation speed in step (1) is 200-500 rpm; the crosslinking agent in step (1) is formaldehyde, glutaraldehyde, genipin or tripolyphosphate, and the reaction temperature after adding the crosslinking agent is 40-60℃, and the reaction time is 2-6 h.

10. The preparation method according to claim 7, characterized in that: The molecular weight of the type I collagen is 20 kDa-1000 kDa, and the mass fraction of the collagen solution is 0.01-2%. When freeze-drying the chitosan microspheres to create pores, the freezing temperature is -80~-20℃ and the time is 12-48 h; In step (3), the activated chitosan microsphere carrier reacts with collagen solution for 12-36 h; The polyamine in step (3) is a diamine with a carbon chain length of 2-6, diethylenetriamine or triethylenetetramine; the reaction after adding the polyamine is: react at 40-60℃ for 2-6 h; the reaction after adding carbonyl diimidazole is: react at room temperature for 6-24 h.

11. The application of the blood perfusion adsorbent according to claim 1 as a packing material for preparing an adsorption column in a blood perfusion device.

12. A hemoperfusion device obtained by using the hemoperfusion adsorbent of claim 1 as a packing material for a hemoperfusion device.

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