| Objective:The original processing is an important point in the production of traditional chinese medicine(TCM)materials.The quality of TCM decoction pieces will be directly affected by the quality of TCM materials processing.Salviae Miltiorrhizae Radix et Rhizoma(SM)is the dry root and rhizome of Salvia miltiorrhiza Bge..Its production areas are widely distributed,and the quality of SM may be affected by different production areas and different drying methods.Through investigation,there are differences in the cutting forms of SM decoction pieces from different origins.SM decoction pieces can be divided into origin slices and moistening slices.Moistening slices are sliced after moistening referring to the method of pharmacopoeia by decoction tablet factories.Origin slices are fresh TCM harvested and dried to a certain extent while fresh cutting.The origin slice has been included in the Shanghai Preparation Standard fo TCM Decoction Pieces(2018 edition)and the Zhejiang Preparation Standard fo TCM Decoction Pieces(2015 edition).Therefore,the quality of SM may be affected by the differences of appearance and internal quality between origin slices and moistening slices.On this basis,this subject investigates the influence of various factors on the quality of SM in the processing of the production area,in order to elaborate the change rule of chemical composition of SM in the process of producing area processing,to explore the influence of the change of appearance properties on the chemical composition of SM and its mechanism,to further standardize and select the optimal processing methods,and to improve the phase and quality of SM.This research also provides a theoretical and scientific basis for the origin processing technology of SM.Methods:1.Part 1:Study on the quality evaluation of commercial SM pieces.Commercial SM decoction pieces from different sources(including origin slices and moistening slices)were collected,and the quality evaluation of SM pieces was studied.The characteristics identification,moisture content,water-soluble and alcohol-soluble extracts,and total tanshinone(the sum of tanshinoneⅠ,tanshinoneⅡA,and cryptotanshinone)and the content of salvianolic acid B were determined according to the methods recorded in appendix of the Chinese Pharmacopeia(2020 edition).2.Part 2:Characterization of chemical constituents of SM.A rapid qualitative analysis of chemical components in SM was carried out by ultra performance liquid chromatography-quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS~E)combined with UNIFI platform.According to the automatic matching results of UNIFI platform,the identification results were further confirmed by the theoretical accurate molecular mass,chromatographic retention time,characteristic ion fragments and other information,as well as related literature reports.3.Part 3:Study on SM materials from different sources.Based on the above identification results,the UPLC-Q-TOF-MS~Etechnology combined with metabonomics analysis method was established.Combined with the sample information of the first part of the commercially available SM decoction pieces,the reasons affecting the quality of SM decoction pieces were analyzed,including the influence of different producing areas and different drying methods in the processing process on the quality of SM.4.Part 4:Study on fresh-cut and moisturized-cut of SM.UPLC-Q-TOF-MS~Etechnology combined with metabonomics analysis methods are also used to analyze self-made fresh TCM materials,origin slices and moistening slices,and the total content of tanshinone(the sum of tanshinone I,tanshinone IIA,and cryptotanshinone)and the content of salvianolic acid B were determined by HPLC.The quality evaluation of 5batches of commercial SM decoction pieces(including origin slices and moistening slices)of the same batch was studied.The characteristics identification,moisture content,water-soluble and alcohol-soluble extracts,and total tanshinone(the sum of tanshinone I,tanshinone IIA,and cryptotanshinone)and the content of salvianolic acid B were determined according to the methods recorded in appendix of the Chinese Pharmacopeia(2020 edition).5.Part 5:Study on the cause of appearance change of section of SM,enzymatic browning.During the processing of origin slice,the appearance change of cross section is prone to occur,which is related to enzymatic browning.Therefore,UPLC-Q-TOF-MS~Etechnology combined with metabonomics analysis method was also used to analyze the effect of enzymatic browning and the different browning degree on the quality of SM.The enzymatic browning products were extracted by alkaline hydrolysis,and then the substructure of the browning products was detected.The identification of browning substructure was mainly based on the fragment ion information of MS~1and MS~2collected by mass spectrometry.In order to reduce the effect of enzymatic browning on the quality of SM,UPLC-Q-TOF-MS~Etechnology combined with metabonomics analysis method was also used to investigate two intervention methods.The effect of different moisture content on the fresh cut and steaming on the quality of SM were studied.Finally,the same technology was used to analyze the commercial pieces of SM to verify the results.Results:1.Part 1:The quality of 21 batches of commercial SM decoction pieces from different sources were investigated.The results showed that there were 5 batches of unqualified decoction pieces,which indicated that the quality of SM decoction pieces from different sources was different.By reviewing the sample information of decoction pieces,it is speculated that the source of the possible differences lies in the differences between TCM materials from different places and the different drying methods in the processing of TCM materials or decoction pieces,resulting in the uneven quality of decoction pieces.2.Part 2:101 chemical components of SM were identified by UPLC-Q-TOF-MS~Etechnology combined with UNIFI platform,including 56 phenolic acids and 45tanshinones.3.Part 3:There are significant differences in chemical composition of SM materials from Zhongjiang in Sichuan,Linyi in Shandong,and Shangluo in Shaanxi.The relative content of total phenolic acids and total tanshinone in SM from Linyi in Shandong were the highest,while the relative contents of phenolic acids in SM from Shangluo in Shaanxi were the lowest,and the relative contents of total tanshinones in SM from Sichuan were the lowest.Different drying methods(freeze-drying,drying in the shade and drying at 40℃and 60℃)had no significant effect on the chemical components of SM.4.Part 4:The chemical components of self-made fresh TCM materials,origin slices and moistening slices were significantly different in negative ion mode.The differences were derived from phenolic acids.8 compounds with VIP>3 were identified,including6 known components and 2 unknown components m/z 671(M-671)and m/z 669(M-669),and the structure of unknown components was predicted.The relative content of total phenolic acid in fresh TCM material is the highest,and the relative content of total phenolic acid in the origin slice is similar to that of the moistening slice.The content of active ingredients in self-made fresh TCM materials,origin slices and moistening slices were determined by HPLC.The results showed that there was no significant difference between the origin slices and moistening slices.However,the fresh TCM materials were compared with the origin slices and moistening slices,respectively.There are significant differences in the content of total tanshinone and salvianolic acid B.The results were consistent with the previous results,which showed that there was a great loss of effective components in the processing of decoction pieces.The quality of the same batch of commercially available origin slices and moistening slices showed that there was no significant difference in the appearance characteristics,moisture content,water-soluble and alcohol-soluble extracts,and total tanshinone(the sum of tanshinoneⅠ,tanshinoneⅡA,and cryptotanshinone)and the content of salvianolic acid B between the origin slices and moistening slices.5.Part 5:There are obvious differences in chemical composition between fresh TCM materials and enzymatic browning samples in negative ion mode,and the differences are derived from phenolic acids.8 compounds with VIP>3 were identified,including 7known components and 1 unknown component M-669,which are basically consistent with those of fresh TCM materials,origin slices and moistening slices.It is also observed that the relative content of M-669 in enzymatic browning samples is relatively high,so M-669 is considered to be a marker of enzymatic browning.The results showed that the relative content of total phenolic acids was negatively correlated with the degree of browning.12 substructures of browning polymers were identified,including 8phenolic acids and 4 organic acids.The sustructures of 6 unknown components were predicted.The results showed that the browning polymers were mainly composed of bound polyphenols and had unique coumarin skeleton.The relative content of total phenolic acid increased and the marker M-669 decreased when the moisture content of fresh TCM materials increased from 52%to 24%.After steaming,the relative content of M-669 in SM can be effectively reduced and the relative content of total phenolic acid can be retained.By investigating 4 batches of the same batch of origin slice and steamed slice,he results showed that the steamed slice M-669 was significantly lower than the origin slice,and the relative content of total phenolic acid was significantly higher than that of the origin slice,which was basically consistent with the previous results.Conclusion:There are significant differences in chemical composition of SM materials from Zhongjiang in Sichuan,Linyi in Shandong,and Shangluo in Shaanxi.Different drying methods,including freeze-drying,shade drying,drying at 40℃and 60℃,have no significant effect on the chemical components of SM.There is no significant difference in the quality between the original slices and the moistened slices,but the original slices are prone to changes in cross-sectional properties and enzymatic browning during the cutting process.It is necessary to strictly control the moisture content of fresh TCM materials when they are cut fresh.The moisture content of fresh TCM materials should be kept at about 24%,and the decoction pieces should be processed by steaming,which can effectively retain the content of salvianolic acids and make the section of the decoction pieces more beautiful.Based on the above experimental results,it can be used to guide the production practice of the processing of SM.According to the production purpose,choose the appropriate methods of processing the decoction pieces of SM.If you focus on the production of salvianolic acids,you can use steaming to process the decoction pieces.It can effectively retain the content of phenolic acids;if you focus on the production of tanshinone ingredients,you can use the method of fresh-cut,which can simplify the processing technology,save time and effort,improve the processing efficiency of decoction pieces.If the TCM materials are cut fresh at the place of production,the manufacturer should move the GMP workshop forward,so that the production area is cut fresh before it is truly feasible. |
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