Study On The Quality Equality Of Different Cultivars Of Citri Reticulatae Pericarpium

In recent years,with the continuous reduction of wild medicinal plant resources,the industries for the cultivation of tradition Chinese medicines have developed rapidly.However,different cultivars were generated due to the influence of artificial breeding,cultivation methods,ecological environment and other factors during the cultivation of medicinal plants.The use of different cultivars as the same medicine has become a common phenomenon in the use of traditional Chinese medicines,which was difficult to guarantee the clinical efficacy of some medicinal herbs.In order to prevent the phenomenon that the efficacy could not always be obtained although the medicines used were the same,the study on the quality equality of different cultivars and the scientific evaluation on the medicinal value of different cultivars have been an important issue to solve for current study on the quality of traditional Chinese medicines.Citri Reticulatae Pericarpium(CRP)was firstly contained in Shen Nong's Herbal Classic.It was a commonly used traditional Chinese herbal medicine with long history of medicinal use.In recent years,the area for the cultivation of traditional variety of CRP was shrunk obviously and the new variety of citrus was introduced continuously for it was an economic fruit plant.The peel of some citrus cultivars whose medicinal quality was not evaluated was added into the medicine as “Hybrid Citri Reticulatae Pericarpium”(HCRP),which made the CRP circulated in the market have hybrid origins.The identification and quality evaluation on medicinal herbs became more difficult and the clinical efficacy was hard to be guaranteed.In order to scientifically evaluate the quality of different cultivars,a scientific contrast study on the quality of CRP from the aspects of characteristics,chemical properties and efficacy was firstly conducted with CRP,the representative traditional Chinese medicine with different cultivars,as the pattern drug and Chuan Citri Reticulatae Pericarpium(CCRP)(peel of C.reticulate ‘Dahongpao'),Guangdong Citri Reticulatae Pericarpium(GCRP)(peel of C.reticulata ‘Chachi')and the HCRP(peel of C.reticulate ‘Ponkan',C.sinensis cv.Newhall and C.unshiu×C.sinensis cv.Skiranui tangerine)as the objects.The rapid non-destructive identification and quality evaluation model of CRP was established by means of modern analytical technique.The main contents of the study were as follows:1.Literature textual researchTextual study on traditional Chinese medicines through the ages was conducted.The evolution of the name and variety of CRP was combed systemically: CRP was firstly contained in Shen Nong's Herbal Classic with “Citrus Tangelo” as the entry and citrus peel as its name.The name of CRP was firstly seen in Herbal Food Medicine with the meaning of old citrus peel.In the herbal before Song Dynasty,“Citrus Tangelo” was taken as the entry of CRP,while the “Citrus Tangelo” in the herbal referred to the peel of citrus with different cultivars in today's citrus reticulate excluding the peel of C.gradis(L.)Osb..The ancients had different evaluations on the quality of CRP from different origins.2.The investigation on commercial medicinal herbsA survey on the commercial CRP was conducted in term of market circulation and origin cultivation conditions.The results showed that as commercial crops,a great variety of citrus plants were widely distributed in China.Since much attention was paid on the edible value of economic benefits,the traditional medicinal cultivars(C.reticulata‘Dahongpao')were gradually replaced by the new varieties such as Ponkan,Skiranui Tangerine and Newhall.As a result,a large amount of peels of other cultivars was circulated in the market as “HCRP”,which made the mixed or uneven quality of CRP be a grim fact.It has exerted great influence on the efficacy and safety of clinical application of traditional Chinese medicines.3.The study on quality equality 3.1 The study on characteristicsA study on the characteristics of different cultivars of CRP indicated that different cultivars of CRP had a certain difference in their characteristics.GCRP was relatively large in size with three pulps connected tidily,thick peel and large spot-like cupped oil chambers.CCRP was relatively small in size with irregular flaky or several pulps connected,thin peel and small oil chamber.The HCRP was variable in size,shape and thickness.3.2 The study on chemical propertiesThe main active ingredients of CRP were flavonoids,alkaloids and volatile oils.A comparative study on the ingredients of different cultivars of CRP was conducted systemically for the first time.3.2.1 The comparative study on the main chemical ingredients(1)Flavonoids: The contents of total flavonoids,narirutin,hesperidin,nobiletin and tangeretin in the samples of medicinal herbs were determined by ultraviolet spectrophotometry and high performance liquid chromatography.The result showed that if it was judged as per current indicator for content detection(the content of hesperidin),only CCRP,GCRP and ponkan samples of HCRP complied with pharmacopeia standards.Besides,the content of hesperidin,nobiletin and tangeretin in CCRP and GCRP was significantly different from those in HCRP.It was higher than those in HCRP.The content of total flavonoids in CCRP,GCRP and HCRP had significant difference.The content of narirutin inCCRP and GCRP had no significant difference with that in ponkan of HCRP.(2)Alkaloids: The content of synephrine in the samples of each medicinal herb was determined by high performance liquid chromatography.The result showed that the content of synephrine in CCRP and that in ponkan of HCRP as well as the content of synephrine in CCRP and GCRP and that in ponkan of HCRP had no obvious difference,while the content of synephrine in other cultivars of medicinal herbs differed a lot.(3)Volatile components: A comparative analysis on the volatile ingredients of different cultivars of medicinal herbs was conducted for the first time by static headspace GC-MS technique in combination with automated mass spectral deconvolution & identification system and retention index.The results showed that except for the specific ingredient(benzoic acid,2-(methylamino)-,methyl ester)in GCRP,different cultivars of CRP had basically the same volatile ingredients(such as limonene,r-terpinene and linalool),while the relative proportions of the volatile components were significantly different,so they belonged to different chemotypes.CCRP belonged to the type of limonene/linalool,GCRP belonged to the type of limonene/?-terpinene/benzoic acid,2-(methylamino)-,methyl ester,the ponkan samples in HCRP belonged to limonene/?-terpinene/linalool,and the Shiranui and Newhall samples in HCRP belonged to limonene.3.2.2 The comparative study on fingerprint spectrumHPLC fingerprint spectrum of CCRP and GCRP was established.Based on the comparison of their control footprint spectrums,their chemical components had certain difference.The comparison between the spectrum of different cultivars and the control footprint spectrum of CCRP showed that the similarity of control footprint spectrums of GCRP,HCRP originating from ponkan and CCRP was greater than 0.9,which indicated that the three items had small difference in chemical components.The similarity between the control footprint spectrum of HCRP originating from Shiranui and Newhall and that of CCRP was less than 0.8,which indicated that they had great difference in chemical components.The results were identical with the content determination results.3.3 The preliminary comparative study on the main efficacyA preliminary comparative study on the main efficacy of CCRP,GCRP and HCRP was conducted for the first time based on the phlegm eliminating and qi regulating effect of CRP.The study on its phlegm eliminating effect was carried out by phenol red method.The results showed that CCRP,GCRP and HCRP had certain effect on the promotion of respiratory secretion and the effects of them had no obvious difference.In order to explore the difference of different cultivars in the effect of qi regulating,isolated rabbit intestine was taken as the object of this study to compare the difference of their effects.The results showed that all of the CRP had inhibition effect on isolated rabbit myenteron,which was concordant with the use of CRP for spasmolysis in case of gastric qi stagnation.The effect of CCRP,GCRP and HCRP from Ponkan was relatively strong.Remarkable difference was found between the effect intensity of the three items and HCRP from Shiranui and Newhall.The results of the study on quality equality showed that different cultivars of CRP differed a lot in quality.The quality of HCRP from Shiranui and Newhall had remarkable difference with that of CCRP and GCRP,so they could not be taken for the medicinal use of CRP.There was no obvious difference between the quality of HCRP from Ponkan and that of CCRP and GCRP.4.The study on the identification of CRP and the model of quality evaluation 4.1 The study on the identification of Citri Reticulatae Pericarpium based on color objectification.In the observation of the characteristics of CRP,it was found that the color of different cultivars was different.In order to realize the objectiveness and quantification of the color of medicinal herbs,spectrocolorimeter was adopted for the first time to obtain the color information of CRP,and thus establishing mathematical discriminant model of CCRP and GCRP based on L * a * b * chromaticity space and bilateral 90% reference range according to the parameters of the color characteristics.The samples of HCRP was identified in accordance with the determined range of reference value.The results demonstrated that the HCRP from Ponkan or Shiranui neither belonged to CCRP nor GCRP.This operation provided a simple and feasible method for the identification of traditional Chinese medicines.4.2 The study on the identification of Citri Reticulatae Pericarpium and quality evaluation based on near infrared spectroscopyThe near infrared spectroscopic information contained the physical and chemical information of the samples for test.The identification of traditional Chinese medicines based on the general characteristics could be achieved by the combination of the near infrared spectroscopic information with the stoichiometric method.In this study,CRP was analyzed by the near infrared spectroscopy in combination with a variety of chemical analysis methods,so as to identify the medicinal herbs based on the overall information and establish the near infrared identification model of CCRP and GCRP.The accuracy of the model was predicted to be 91.67%.At the same time,the near infrared quantitative model for the content of hesperidin,nobiletin and tangeretin in the CRP was established for the first time.Which not only provided a demonstration for the rapid identification of traditional Chinese medicines,but also provided a new optional means for the evaluation on the quality of traditional Chinese medicines.