Research On The Sterilization Process Of Chinese Crude Medicine Powder In Tianma Shouwu Tablets And The Improvement Of Quality Standards

Objective:To optimize the sterilization technology of raw medicinal powder in Tianma shouwu tablets,and to investigate the effects of different doses of Co60 irradiation sterilization on raw medicinal powder.Method:(1)Respectively treat Rhizoma gastrodiae,Rhizoma ligustici chuanxiong,and radix polygoni multiflori preparata raw medicinal powder by wet heat sterilization,microwave sterilization,ethanol gas sterilization,and co60 irradiation sterilization method.The results of appearance,sterilization effect,TLC identification and content determination of effective components were used as evaluation indexes,and the optimal sterilization method was selected.Among them,gastrodin,ferulic acid and 2,3,5,4'-tetrahydroxystilbene-2-O-B-D-glucoside were selected as the index effective components respectively for gastrodia elata,Ligusticum wallichii and Polygonum multiflorum Thunb.(2)The same batch of gastrodia tuber,Ligusticum wallichii and Polygonum multiflorum Thunb are randomly divided into three groups and sterilized by co60 irradiation respectively at the doses of 2k Gy,4k Gy and 6k Gy,the effect of dosage on the chemical constituents of raw powder was investigated by TLC identification,determination of effective constituents and fingerprint.The same batch of Polygonum multiflorum Thunb.was randomly divided into three groups and sterilized by co60 irradiation,the doses were 2k Gy,4k Gy and 6k Gy,respectively.The treated samples,sterilized Polygonum multiflorum Thunb.and raw Polygonum multiflorum Thunb.were added to human normal hepatocytes(L02)in logarithmic phase for 24 hours.CCK-8 detection method was used to evaluate the growth inhibition of each group of samples on human normal hepatocytes(L02).(3)To establish the fingerprint of Tianma shouwu tablets: kromasil C18(250×4.6mm,5?m)column,acetonitrile(a)-0.3% phosphoric acid solution(b)as mobile phase gradient elution,0~18min,5%~15%a,18~32min,15%~26%a,32~35min,26%~27%a,35~48min,27%~40%a,detection wavelength 230 nm,flow rate is 1.0m L/min,column temperature is 30?,analysis time is 48 min.The fingerprints of 10 batches of Tianma shouwu tablets were determined.the common mode of Tianma tablets was established by similarity analysis software,and the chromatographic peaks were identified.Results:(1)Gastrodia elata: suitable for the use of microwave sterilization,Co60 irradiation sterilization,of which the best parameters of microwave sterilization are as follows: high fire 90 s when single flavor sterilization,high fire 3min when mixed sterilization.Temperature and humidity affect the content of Gastrodin in Gastrodia elata Blume,which may be related to the activity of enzyme in Gastrodia elata.Ligusticum chuanxiong: the powder of Ligusticum chuanxiong is suitable for microwave sterilization or Co60 irradiation sterilization.The best parameter of microwave sterilization is "medium fire 150s",and the mixed sample can be microwave sterilization,and the best parameter is "high fire 2min".The instability of ferulic acid is related to moisture and temperature.The increase of water and temperature accelerates the dissociation of ferulic acid.Polygonum multiflorum:suitable for microwave sterilization,Co60 irradiation sterilization,microwave sterilization of the best parameters are as follows: single flavor sterilization is "high fire 3min",mixed sterilization is "high fire 2min".The content of 2,3,5,4'-four hydroxyl two styrene-2-O-beta-D-glucoside in Radix Polygoni Multiflori is related to temperature.(2)The irradiation dose of 6k Gy had no significant effect on the chemical constituents of gastrodia elata,Ligusticum wallichii and Polygonum multiflorum Thunb.After 24 h treatment of normal human hepatocytes(L02)with raw Polygonum multiflorum,the cell survival rate was about 14.4 %.The cell survival rates of unsterilized Polygonum multiflorum and sterilized Polygonum multiflorum by 2k Gy,4k Gy and 6k Gy Co60 irradiation were 98.9%,109.7%,105.4%and 106.2%,respectively,there was no significant effect on cell viability.(3)The HPLC fingerprint of Tianma shouwu tablets was established,and the similarity data of 10 batches of samples were obtained.There were 18 peaks,17 of which belonged to each medicinal material and 6 were identified.Conclusion:(1)Microwave sterilization and Co60 irradiation are suitable for the medicinal powder of Gastrodia elata.The powder of Rhizoma Chuanxiong is suitable for microwave sterilization and Co60 irradiation.The Radix Polygoni Multiflori is suitable for sterilization by microwave and Co60 irradiation.In consideration of the effect of sterilization,the influence on chemical composition,the operation and the cost,the sterilization process of the raw powder in the Tianma Shouwu tablet is: the mixed powder of the three is sterilizing by microwave sterilization.(2)The irradiation dose of 6k Gy had no significant effect on the chemical constituents of gastrodia elata,Ligusticum wallichii and Polygonum multiflorum Thunb The toxicity of Polygonum multiflorum Thunb to human normal hepatocytes was not increased by Co 60 irradiation with dose lower than 6k Gy.(3)The HPLC fingerprint method was simple,accurate,stable and reproducible,which provided an effective method for the quality control of Tianma shouwu tablets.