Optimization Of Hydrochloric Acid Hydrolysis Extracting Mollugin From Rubia Cordifolia By Response Surface Methodology

Objective:Madder,the dried root and rhizome of Rubia cordifolia L.from madder family,has the functions of cooling blood,removing blood stasis,hemostasis,menstruation,and is widely used in the treatment of epistaxis,hematemesis,traumatic injury,blood stasis amenorrhea and other symptoms.Rubimaillin and purpurin are the index components of the quality of madder in the 2015 edition of the Chinese Pharmacopoeia,and have the effects of anti-tumor,antitoxic hormone,hemostatic and coagulation,immune system,anti-inflammatory,antioxidation,free radical elimination and radiation resistance.It was found that in the process of determination of the content of alizara,the sample preparation was more tedious and was easy to produce errors in the process of continuous transfer.In this study,a certain concentration of hydrochloric acid methanol was used as the extraction solvent to hydrolyze at corresponding temperature.It was found that this method had good extraction effect on rubimaillin and purpurin in madder.Therefore,in this experiment,through response surface optimization experiment,we determined the best concentration of hydrochloric acid,heating temperature and heating hydrolysis time with the content of rubimaillin and purpurin.Methods:1.The exclusive TLC identification method of Madder was developed by investigating the preparation methods,developing the system and developing the chromogenic agent.With the content of rubimaillin and purpurin as the quality control index,an HPLC method was established and was fully validated on precision,stability,repeatability and recovery.The content limits of rubimaillin and purpurin are determined by the contents of the 3 batches of samples.2.Response surface methodology was used to optimize the extraction conditions of rubimaillin from madder:based on single factor test,taking rubimaillin content as response value,select the concentration of hydrochloric acid,heating temperature and time as factors,using Box-Behnken to optimize the process of response surface design.Results:Using methanol solution for 30 minutes as the ultrasonic method,The preparation method was using methanol as solution and ultrasonic for 30 min.Petroleum ether:acetone(4:1)was as an unfolding agent.Under fluorescent observation,TLC spots were clear,negative control was not interfered,and specificity was strong.Established the determination method of rubimaillin and purpurin in madder by HPLC method:Rubimaillin show a good linear relationship in the range of 13.78?172.23 g/ml,the regression equation is Y=10.674X+20.078(R2=0.9946),the average recovery rate was 98.9%,and the RSD was 0.74%(n=6).Purpurin also show a good linear relationship in the range of 6.73?84.10 g/ml,the regression equation is Y=36.315X-0.0166(R2=0.9997),the average recovery rate was 97.7%,and the RSD was 1.13%(n=6).The content limit of madder should be determined by 80%of the 3 batches of samples.The content of rubimaillin should not be less than 0.96%,and the content of purpurin should not be less than 0.17%.The method is simple,reliable and reproducible,and can be used for quality control of madder.To optimize the extraction process of rubimaillin in madder,response surface methodology was used to optimize the quality control,and provide quality testing method to study the optimum process of rubimaillin extraction by the response surface method.Response surface methodology was applied to optimize the extraction process of Madarin from madder.When the concentration of hydrochloric acid is 10.5%,the heating time is 41 min,and the heating temperature is 82 degrees,the yield of rubimaillin is 2.53%,which is close to the predicted value 2.51%.Conclusion:In this study,TLC was used for qualitative identification of madder,and HPLC was used for the content determination of madder.The linear regression curve,precision,repeatability,stability and recovery rate were also studied.It is concluded that all the samples in this study contain rubimaillin and purpurin,and the content is generally higher than that of Chinese Pharmacopoeia.In this study,response surface methodology was used to optimize the extraction method of madder.The content of rubimaillin and purpurin extracted by the method is greater than that of the Pharmacopoeia method.The optimization process is simple,stable and reproducible,and can be used for the extraction and extraction of Madarin from madder.