Effects Of Different Concentrations Of Polysorbate - 80 On Two - Step Gel Method For Detecting Impurities In Traditional Chinese Medicine Injection

Traditional Chinese Medicine (TMC) injection is an innovative dosage forms, has become a common means of clinical treatment. With its increasingly widespread application in clinical, the number of adverse reaction reported significantly increased, one of the most important reasons is the existence of impurity protein which contained in the injection.The most common protein check item which is included in quality control of TMC injection is the sulfosalicylic acid turbidity method. This method is easy and fast, but has low sensitivity and poor specificity, and will encounter false positive interference, such as Tween-80 and so on. Tween-80 is commonly used in the TMC injections as solubilizing excipients for pHarmaceutical productions, can effect the accuracy and sensitivity of the determination method for impurity protein. This research applied the two-dimensional gel chromatography method, which was combined the dextran gel chromatography method with TSK-HPLC method, as the sulfosalicylic acid turbidity method for reference experiment, to study the effect of different concentration of Tween-80 on the detection of impurity protein in traditional Chinese medicine injections by two-dimensional gel chromatography method. The main contents are as following:Firstly, through literature research, this reseach selected 20 kinds of TMC injections as the research object, which included single Chinese medicine composition TMC injections, single herb TMC injections and compound medicine TMC injections, added and nun-added Tween-80 TMC injections.Secondly, this research improved the above two-dimensional gel chromatography method, then investigated the work-standard, reproducibility, tolerance and so on, after detect impurity protein in all the subjects. The results show that compared with sulfosalicylic acid turbidity method, two-dimensional gel chromatography method has lower detection limit, higher sensitivity, better stability and the result is more reliable.Finally, in order to detect the influence factors of protein detection methods, this research compared two-dimensional gel chromatography method and sulfosalicylic acid turbidity method under different Tween-80 concentrations. The results show that (1) for sulfosalicylic acid turbidity method, when Tween-80 concentration is below 0.5%, the detection limit can be maintained; when Tween-80 concentration is between 0.5% and 2%, the detection limit will increasing along with the concentration, whereas the sensitivity is descending; when the Tween- 80 concentration is higher than 2%, the blank group was positive, the sulfosalicylic acid turbidity experiment are easy to occur false positive results at this point. (2) when Tween-80 concentration is below 6.0%, it had no significant effect to two-dimensional gel chromatography method; when the Tween-80 concentration is higher than 6%, although Tween-80 had no significant effect on the retention time, the peak area was significantly decreased, and the detection limits and the limits of quantification were significantly increased.In conclusion, compared with sulfosalicylic acid turbidity method, the two-dimensional gel chromatography method has advantages of good stability, high repeatability, reliable results, and not subject to personnel and equipment for the protein examination of traditional Chinese medicine injections. This study has important guiding significance for the quality control of traditional Chinese medicine injection, and also provides reference for the rational and safe use of Chinese traditional medicine injection.