Generation of micro-particles of proteins for aerosol delivery using high pressure modified carbon dioxide
Generation of micro-particles of proteins for aerosol delivery using high pressure modified carbon dioxide
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DOI：
10.1023/a:1007551006782
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发表时间：
2000-11-01
影响因子：
3.7
通讯作者：
Foster, NR
中科院分区：
文献类型：
--
作者：
Bustami, RT;Chan, HK;Foster, NR
目的。目的探讨利用气雾剂溶剂提取系统(ASES)从水基溶液中制备适合气雾剂输送的蛋白质微粒的可行性。利用经乙醇修饰的高压二氧化碳的ASES技术从水溶液中产生微小的蛋白质颗粒(溶菌酶、白蛋白、胰岛素和重组人脱氧核糖核酸酶(RhDNase))。考察了颗粒大小、形貌、粒度分布和粉末气溶胶性能。用分子排斥层析法和溶菌酶生物测定法测定蛋白质的分子聚集水平，评价蛋白质的生物化学完整性。蛋白质沉淀为大小为100~500 nm的球形颗粒。初生纳米颗粒在沉淀过程中凝聚成微米级颗粒。颗粒的中值尺寸是操作条件的函数。体外气溶胶性能测试表明，溶菌酶、白蛋白和胰岛素的细小颗粒质量百分比(<；5M)分别约为65%、40%和20%。溶菌酶的单体含量或生物活性的损失可以忽略不计。胰岛素有轻微的聚集，加工后单体的保留率为93%。白蛋白受到工艺的影响，只有50-75%的单体被保留，而原始材料中的白蛋白保留了86%。但在炮制过程中，rhDNase发生了明显的变性，单体含量显著降低。利用改进的ASES技术，从水溶液中成功地制备出具有良好吸入性能的溶菌酶、白蛋白和胰岛素的微米级颗粒。加工蛋白质的生物化学完整性是操作条件和单个蛋白质性质的函数。
Purpose. To investigate the feasibility of using the Aerosol Solvent Extraction System (ASES) to generate microparticles of proteins suitable for aerosol delivery from aqueous-based solutions.Methods. The ASES technique using high- pressure carbon dioxide modified with ethanol was utilised for the generation of microparticles of proteins (lysozyme, albumin, insulin and recombinant human deoxyribonuclease (rhDNase)) from aqueous solutions. Particle size, morphology, size distributions and powder aerosol performance were examined. The biochemical integrity of the processed proteins was assessed by testing the level of molecular aggregation using size exclusion chromatography and by bioassay technique for lysozyme.Results. Proteins were precipitated as spherical particles ranging in size from 100 to 500 nm. The primary nano-sized particles agglomerated to form micron-sized particles during the precipitation process. The median size of the particles was a function of the operating conditions. In-vitro aerosol performance tests showed that the percent fine particle mass (< 5m) was approximately 65%, 40% and 20% for lysozyme, albumin and insulin, respectively. Negligible loss in the monomer content or biological activity was observed for lysozyme. Insulin exhibited slight aggregation and 93% of the monomer was retained after processing. Albumin was affected by processing and only 50-75% of the monomer was retained compared with 86% in the original material. However, rhDNase was substantially denatured during processing as shown by the significantly reduced monomer content.Conclusions. Micron-sized particles of lysozyme, albumin and insulin with satisfactory inhalation performance were successfully generated from aqueous solutions using the modified ASES technique. The biochemical integrity of the processed proteins was a function of the operating conditions and the nature of the individual protein.