Preparation of alpha-1 antitrypsin and comparative analysis on two bioactivity measurement methods

FU Yan-li; GUO Yu-ting; KONG Tao; MA Ya-ru; LIANG Ling-yu; CHEN Yong

doi:10.20251/j.cnki.1003-3734.2025.11.008

Abstract：Objective:alpha-1 antitrypsin(AAT)was isolated and purified from the precipitation of Cohn component IV,and two methods for determination of the bioactivity of AAT were compared.Methods:AAT was isolated from the Cohn component IV following a series of processes,including precipitation dissolution,press filtration and clarification filtration,SD virus inactivation,3-step chromatography,ultrafiltration concentration,sterile filtration,freeze-drying,and finally dry heat virus inactivation.The activity of AAT was measured by trypsin method and elastin method,respectively.The linear range,specificity,accuracy,repeatability,intermediate precision,linearity,durability and consistency of the two methods were compared and analyzed.Results:AAT was purified from FIV.The specificity and linearity of the two methods were good for the determination of AAT bioactivity.The range of trypsin assay is 0～16 μg·mL-1,and the range of elastin enzymatic assay is 0～2 μg·mL-1.The RSD of reproducibility and intermediate precision verification ae all below 10%,and the recoveries of high,medium and low concentrations are all between 80%and 120%.The durability of the two methods was better when incubation time was 20～40 min,and there was no significant difference in AAT bioactivity between the two methods(P=0.173 0,>0.05).Conclusion:In this study,AAT was successfully isolated from FIV.Both trypsin method and elastin method have good specificity,linearity,repeatability,intermediate precision,accuracy and durability in the determination of AAT bioactivity.The two methods were simple and reliable,and could be used for determining the bioactivity of AAT products.