Preparation And Application Of Endotoxin Adsorbent Based On ?-polylysine As Ligands

Sepsis can be caused by infection in any part,clinically common in pneumonia,peritoneal,urinary tract infection,meningitis,abscess and so on.Its pathogenic microorganisms include bacteria,fungi,viruses and parasites,and sepsis often occurs in patients with serious diseases,such as severe burns,multiple injuries,and post-surgical patients.It is currently widely believed that sepsis is a systemic inflammatory response syndrome caused by endotoxin entering the blood.Endotoxin is a component of the outer wall of Gramnegative bacteria.In the process of expression of contaminated surgical instruments,irrigation systems,biological products such as vaccines,antibodies,etc.,a large amount of endotoxin is produced,so endotoxin is almost ubiquitous.In addition,the use of antibiotics during the treatment of sepsis will once again bring a large amount of endotoxin release to aggravate the patient's condition,so the removal of endotoxin is very important.Most of the existing methods for removing endotoxin have limited removal ability and non-specificity,and excessive non-specific adsorption makes it impossible to remove endotoxin in biological products and clinical blood.Therefore,there is an urgent need for an endotoxinbinding material which has strong adsorption capacity for endotoxin and good blood compatibility.However,sputum-polylysine is produced by Streptomyces albus,and has spectral antibacterial properties and has been approved as a safe food preservative,which is also non-toxic and harmless to the human body from a safety point of view.In addition,ruthenium-polylysine has 25-30 lysines,which can pass the electrostatic interaction and hydrophobic action of cations,and have good adsorption to endotoxin.In this thesis,an endotoxin adsorbent was synthesized by using sepharoseCL 4FF as a carrier and ?-polylysine(?-PLL)as a ligand.The kinetics and thermodynamics experiments on the adsorption of endotoxin solution and plasma solution were carried out.The adsorption of FITC-LPS indicates that the adsorbent has good adsorption capacity for endotoxin,and the adsorbent is optimized to achieve better specific adsorption of endotoxin and non-special other components in plasma.Heterostatic adsorption.The specific research content is:(1)Synthesis of agarose-?-polylysine adsorbent: the hydroxyl group of epoxy activated agarose gel,coupled with agarose and ?-polylysine(?-PLL)to synthesize agarose-?-polylysine adsorbent,then activate its carboxyl group by EDC/NHS,couple some hydrophobic and aminocontaining chemicals,and synthesize agarose-?-polylysine-R adsorbent,in anticipation of reaching the inner Better adsorption of toxins.Adsorption experiments on endotoxin showed that the adsorption effect of agarose-?-polylysine was better than other adsorbents.(2)Establishment of endotoxin detection method: The endotoxin content was determined by dynamic chromogenic method(0.005-10 EU),which is superior to the previous chromogenic substrate method(0.01-0.1,0.1-1 EU)and different The blood of the anticoagulant was tested for the recovery rate,and the heparin sodium anticoagulated blood was determined to meet the requirements of the pharmacopoeia for the recovery of endotoxin detection.The subsequent experimental plasma was anticoagulated with heparin sodium.(3)Agarose-polymyxin B(PMB)endotoxin adsorbent was synthesized by epoxy activated agarose and coupled with polymyxin B(PMB)to control the amino acid density to be approximately the same,through low concentration endotoxin Adsorption of plasma solution showed that agarose-?-polylysine adsorbed endotoxin better than agarose-polymyxin B(PMB).(4)Optimize the above agarose-?-polylysine to optimize the density of epoxy groups,the density of amino groups,the kinetics and thermodynamics experiments on the adsorption of endotoxin aqueous solution and plasma solution,and the adsorption experiments on FITC-LPS.It was verified that the adsorbent had good adsorption capacity for endotoxin.When the plasma concentration of endotoxin was 10 EU/mL(1 ng/mL),the adsorption rate of the adsorbent was 90%,and the adsorption capacity was 46.53 EU/mL..(5)The hemolysis rate of the adsorbent is 3.75% <5%,which satisfies the pharmacopoeia regulations.In addition,the relative changes of plasma total protein and albumin before and after adsorption of different amino density adsorbents are below 10%,that is,plasma protein can recover more than 90%,and the adsorption effect on endotoxin is good,the results are in clinical Within the allowable range,this adsorbent has good blood compatibility and has potential clinical application prospects.