Study On Identification Technology Of Arisaematis Rhizoma And Typhonii Rhizoma Before And After Processing

Arisaematis Rhizoma is the dry tuber from Arisaema erubescens(Wall.)Schott、Arisaema heterophyllum B1 and Arisaema amurense Maxim.Arisaematis Rhizoma is bitter,pungent,warm and toxic,with the effect of dampness phlegm,removing wind and antispasm,and reducing swelling.Typhonii Rhizoma is dry tuber from araceae Typhonium giganteum Engl.Typhonii Rhizoma ispungent,warm and toxic,with the effect of dispelling wind and phlegm,arresting convulsion,diminishing stagnation by detoxification and alleviating pain.Both Arisaematis Rhizoma and Typhonii Rhizoma are toxic,showing strong irritant toxicity,which could be used clinically after processing to reduce toxicity.There is common processing method of Arisaematis Rhizoma and Typhonii Rhizoma in the 2020 edition of Chinese Pharmacopoeia,and this method belongs to the repeatedly processing method,and it is fussy and time-consuming.Therefore,there is no lack of phenomenon that it is disordered of raw drugs and processed drugs in the market,and it brought great hidden trouble for clinical medication safety of Arisaematis Rhizoma and Typhonii Rhizoma.Both Arisaematis Rhizoma and Typhonii Rhizoma the quality standards of Arisaematis Rhizoma and Typhonii Rhizoma are lack of toxicity control index in the 2020 edition of Chinese Pharmacopoeia.Previous studies of our project team show that the common toxic chemical components of Arisaematis Rhizoma and Typhonii Rhizoma are needle crystal complex composed of calcium oxalate needle crystal and lectin protein.Therefore,considering the current status of lack of toxicity control index for Arisaematis Rhizoma and Typhonii Rhizoma,based on the previous research results of our project team,The method for the content determination of calcium oxalate needle crystal and lectin protein of endogenous toxic substances of Arisaematis Rhizoma and Typhonii Rhizoma was established in this paper,and the corresponding limit standard was also established.The discriminant models of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing and Arisaematis Rhizoma and Typhonii Rhizoma mixed with Arisaematis Rhizoma Preparatum and Typhonii Rhizoma Preparatum was established respectively with pattern recognition of stoichiometry.The models was verified and it showed excellent performance and accuracy.It provide methods for recognizing Arisaematis Rhizoma and Typhonii Rhizoma before and after processing.In addition,The difference of chemical compositions of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing was analysed based on UPLC-TOF-MS.This paper illustrated the the difference of chemical compositions of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing and the differential components was determined.This paper provide scientific data and research basis for further study on the material basis of detoxification and synergism by processing.The detailed research contents are as follows:1.Study on the content changes of endogenous toxic components of Arisaematis Rhizoma and Typhonii Rhizoma before and after processingEndogenous toxic components of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing was determined:UPLC was applied to determinate the content of calcium oxalate crystal.Western-Blot was applied to detect the content of lectin protein.Results showed that the content of calcium oxalate crystal of all batches of Arisaematis Rhizoma is 0.63%～2.33%,and average content is 1.39%;the content of calcium oxalate crystal of all batches of Arisaematis Rhizoma Preparatum is 0.20%～0.95%,and average content is 0.50%;the decreasing amplitude of calcium oxalate crystal is 51.0～78.3%,and average decreasing amplitude is 64.1%;the content of calcium oxalate crystal of all batches of Typhonii Rhizoma is 0.49%～1.76%,and average content is 0.91%;the content of calcium oxalate crystal of all batches of Typhonii Rhizoma Preparatum is 0.00%～0.46%,and average content is 0.26%;the decreasing amplitude of calcium oxalate crystal is 51.1%～100%;and average decreasing amplitude is 70.6%.The results of content of lectin protein showed that there is no lectin protein was detected in all batches of Arisaematis Rhizoma Preparatum and Typhonii Rhizoma Preparatum.Based on the results of content of calcium oxalate crystal and lectin protein of all batches of Arisaematis Rhizoma and Typhonii Rhizoma,it is suggested that the lectin protein should not be detected in Arisaematis Rhizoma Preparatum,and the reduction of calcium oxalate should not be less than 50.0%;It is suggested that the lectin protein should not be detected in Typhonii Rhizoma Preparatum,and the decrease of calcium oxalate should not be less than 50.0%.The results showed that endogenous toxic components of Arisaematis Rhizoma and Typhonii Rhizoma was decreased significantly.2.Identification of Arisaematis Rhizoma、Typhonii Rhizoma and Arisaematis Rhizoma Preparatum、Typhonii Rhizoma Preparatum based on Near Infrared SpectroscopyNear Infrared Spectroscopy(NIRS)was used to analyze the samples of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing and Arisaematis Rhizoma and Typhonii Rhizoma mixed with Arisaematis Rhizoma Preparatum and Typhonii Rhizoma Preparatum respectively.The spectra were collected in the wave number range of 12000～4000 cm-1,and the discriminant model was established with pattern recognition of stoichiometry.The Discriminant Analysis model and Distance Match model of Arisaematis Rhizoma before and after processing showed that it could distinguish between Arisaematis Rhizoma、partial processed Arisaematis Rhizoma and Arisaematis Rhizoma Preparatum.The Discriminant Analysis model and Distance Match model of Arisaematis Rhizoma mixed with Arisaematis Rhizoma Preparatum and Arisaematis Rhizoma Preparatum showed that it could distinguish between a series of Arisaematis Rhizoma mixed with Arisaematis Rhizoma Preparatum and Arisaematis Rhizoma Preparatum.The Discriminant Analysis model and Distance Match model of Typhonii Rhizoma before and after processing showed that it could distinguish between Typhonii Rhizoma、partial processed Typhonii Rhizoma and Typhonii Rhizoma Preparatum.The Discriminant Analysis model and Distance Match model of Typhonii Rhizoma mixed with Typhonii Rhizoma Preparatum and Typhonii Rhizoma Preparatum showed that it could distinguish between a series of Typhonii Rhizoma mixed with Typhonii Rhizoma Preparatum and Typhonii Rhizoma Preparatum.3.Study on chemical compositions of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing based on UPLC-TOF-MSTo explore the difference of chemical composition of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing,we analysed the chemical composition of 15 batches of Arisaematis Rhizoma and 15 batches of Typhonii Rhizoma before and after processing by UPLCTOF-MS.The characteristic peaks were extracted by Marker View software,and then the difference of chemical composition were analyzed by SIMCA software.The differential components were analyzed according to the chemical composition database of Arisaematis Rhizoma and Typhonii Rhizoma.The results showed that there were significant difference in the chemical composition of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing.There were 121 significantly discrepant chemical compositions in Arisaematis Rhizoma before and after processing,and 66 chemical compositions decreased、10 chemical compositions increased、24 chemical compositions disappearing、21 chemical compositions producing.Eightyfive chemical compositions were identified,including 10 amino acids、18 alkaloids、18 peptides、6 organic acids、1 flavonoid、11 other chemical compositions、4 volatile oils and 17 gingerols,and the molecular formula of the other 36 chemical compositions were deduced.There were 80 significantly discrepant chemical compositions in Typhonii Rhizoma before and after processing,and 26 chemical compositions decreased、27 chemical compositions increased、15 chemical compositions disappearing、12 chemical compositions producing.Fifty-four chemical compositions were identified,including 4 amino acids、11 alkaloids、9 peptides、4 organic acids、1 flavonoids、14 other chemical compositions、3 volatile oils and 8 gingerols.The molecular formula of the other 26 compounds were deduced.This chapter showed that chemical compositions of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing have changed significantly.This paper adopts HPLC to determinate the content of calcium oxalate crystal of and WesternBlot to detect the content of lectin protein in Arisaematis Rhizoma and Typhonii Rhizoma respectively,and establish the determination limitation of toxic components based on the results of content of all batches of Arisaematis Rhizoma and Typhonii Rhizoma.This paper established method to distinguish between Arisaematis Rhizoma and Typhonii Rhizoma before and after processing and raw mixed with processed products.The difference of chemical compositions in Arisaematis Rhizoma and Typhonii Rhizoma before and after processing was analysed based on UPLC-TOF-MS.Method which we established in this paper is reasonable and it provide basis for quality identification of Arisaematis Rhizoma and Typhonii Rhizoma before and after processing.