The Manufacture And Component Analysis Of Albumin Microsphere By Sonication, A New Left-Heart Contrast Agent For Echocardiography

PURPOSE:To study the manufacture of 5% albumin microspheres by sonication,and the possible mechanism of microsphere forming. To analyze the component by many methods and study the methodology of the quality controls.MOTHEDS:1. To study the manufacture of human albumin microsphere and the possible mechanism of microspheres forming.(1) Selected the best manufacture craft by orthogonal experiment design. In asepsis surroundings,sonicated three concentration (1,2,5%) of albumin-NS (normal saline) solutions with three ultrasound frequency ( 8,9,10 grade) and three exposure times ( 50,60,70 second),then determined the concentration of microspheres by Coulter Multisizer. Dealed with the date (the value of the concentration and size) by statistics methods and took the best craft condition.(2) Determined the diameter of microspheres made under the best craft by Coulter Multisizer.(3) Observed the outward appearance of microspheres by the light microscope.(4) The transmission electron microscope.Chemilluminescence was used to measure oxygen free radicals induced by sonication on the albumin solution. Laser time-flight mass spectrum was used to analyze the changes of albumin molecule sonicated. The mechanism of microspheres forming wasdiscussed for the results.2. To analyze the component by many methods and study the methodologyof the quality controls.(1) Studied the method that was used to determine the concentration and diameter of microspheres.Including determining condition,method,the liner of method.and recovery rate,etc.(2) Studied the method that was applied to determine the content of air in microspheres.(3) Determined the protein content on wall of albumin microspheres by Gel column high-performance liquid chromaography,cellulose acetate membrane electrophoresis and semimcro Kjeldall method.(4) SDS-PAGE electrophoresis analyses and else routine quality control methods.Including pH value,determining albumin polymeride,sterility test,pyrogen test and determination of HCV\ HIV> HbsAg.etc.RESULT:1. The best manufacture craft condition was 5% albumin-NS solution,9-grade ultrasound frequency and 60 seconds' exposure time. Two microscopes viewed that microspheres were well- distributed,smooth,plump and regulation. The acoustic sonication only caused reversible changes in spatial structure of albumin molecule.2. The result of Coulter Multisizer viewed that the mean concentration was 3.5 + 1.3 x108/ml,and the mean diameter was 4.2+0.2 u m. The content of protein on the wall was 2.52% to 4.64%. The results of routine methods were the same to albumin's in the Chinese Rules of Biological Product.CONCLUSION:Succeeded developing the albumin microspheres filled with air by orthogonal experiment design,and it could improve effect of contrast in intravenous myocardial contrast echocardiogragy( IMCE). It's stable,safe,effective and quality-controlled,and can meet the needs of the clinical diagnosis.