| The citris peels, Pericarpium Citri Reticulatae (PCR) and Pericarpium Citri Reticulatae Viride (PCRV),harvested at different growth stage, are from the Citrus reticulata Blanco and its cultivars of fruit. They are both featured as "regulate qi medicine", but for the clinical application, PCR has a better effect on regulating qi and strengthening spleen whereas PCRV is strong in soothing liver and breaking qi. As a result from the same plant,PCR and PCRV have some similarities in the terms of active ingredients, efficacy and so on. However, due to the different harvest period, the accumulated changes of secondary metabolites make them different.Current research shows that most of plants secondary metabolites with physiological activity, the medicine efficacy and effect of traditional Chinese medicine (TCM),is important efficacy material base. And the synthesis and accumulation of secondary metabolites by their genetic factors restriction and the influence of environmental factors, therefore, are needs to determine origin and plants to study them. Now there is no system to research the composition,levels and relative proportion of different growth period orange peels secondary metabolites; and Now there is no system to studies on dynamic changes of main Active Ingredient in orange peels of Citrus reticulata blanco from the same plant at different growth periods. So the content of difference between PCR and PCRV in different growth periods,in order to interpret the key factors that affect both drug efficacy differences and provide a theoretical basis.This study takes PCR and PCRV as the research object, in order to reduce environmental factors and their own genetic factors on the synthesis and accumulation of secondary metabolites of uncertainty.This paper specially collected the same plant orange peel to study. At the same time, this paper engaged to growth periods and used main pharmacological compounds of flavonoids, phenolic acids, alkaloids and essential oil as indicators. Systematically study dynamic changes of effective components in the entire growth cycle. Focus on the following research: 1. Studies on dynamic changes of main active constituents in orange peels from Citrus reticulata Blanco at different growth periods1.1FlavonoidsFlavonoids in orange peels are mainly narirutin, hesperidin, nobiletin and tangeretin. By the HPLC technology, this study takes the lead in building the method for double wavelength and determining simultaneously the contents of narirutin, hesperidin, nobiletin and tangeretin. By the UV technology, this study building the methods for determination of the total flavonoids. The results show that total flavonoids decreased from16.55%to10.37%with the increase of fruit maturity; the narirutin decreased from0.85%to0.17%with the increase of fruit maturity; the hesperidin decreased from10.58%to5.25%with the increase of fruit maturity. Nobiletin and tangeretin were significantly degressive trend.1.2Total phenolic acidsReported in the literature, phenolic acids not only have the function of antioxidant, anti-free radicals, anti-cancer, anti-inflammatory and anti-platelet aggregation,but also have inhibition of LDL oxidation and prevention of cardiovasc-ular disease and so on. This study refer to be on the basis of Pharmacopoeia of the People’s Republic of China (2010edition) of the tannins content determination method under appendix X B, this is the first comparative analysis of the variation of total phenolic acids content in the orange peels of Citrus reticulata Blanco from the same plant at different growth period. The results show that total phenolic acids decreased from6.454%to3.801%with the increase of fruit maturity. The results provided experimental basis for the raw material selection on phenolic acids extration.1.3AlkaloidsSynephrine is one of the characteristic components in orange peels, with biological activity, can be used as orange peels quality control index. By the HPLC technology, this study takes building the method for determining simultaneously the contents of synephrine. The results show that synephrine decreased from0.95%to0.39%with the increase of fruit maturity.1.4Volatile oil In the orange peels essential oil for one of its main component, with biological activity, such as limonene, terpineol, pine oil with anticancer, antibacterial activity. By the HS-SPME/GC/MS technology, this study takes building the method for determining simultaneously the contents of essential oil. The results show that a total of36compounds in the essential oils of13batches orange peel were identified, which represented88.26%,84.1%,92.25%,93.74%,91.41%,88.59%,88.79%,91.04%,96.53%,85.68%,81.93%,86.93%, and85.49%of their total relative contents, respectively. The major contains compound for limonene, gamma-terpinene, linalool, alpha-farnesene, thymol, palmitic acid. The compounds that have the highest content is Limonene, accounting for14.15%-31.70%. Different growth period of orange peel essential oil contains the same composition, the composition has the certain differences in content, r-terpinene regularity of change in orange peel growth period. The method employed in the study is convenient and efficient, and the study can serve as a reference to researches on aromatic and pungent pharmaceuticals with volatile constituents.These results showed that the contents of total flavonoids, total phenolic acids, hesperidin, narirutin and synephrine were significantly degressive trend with the increase of fruit maturity. But the contents of nobiletin and tangeretin showed a downward trend overall. Volatile oil itself instability leads to difficulties in the analysis, but can evaluate its belong to which chemical type. The results provided an experimental basis for the differences efficacy of PCR and PCRV.2. Comparative the different parts of the four kinds of flavonoidsBy the HPLC technology, this study takes the lead in building the method for double wavelength and determining simultaneously the contents of narirutin hesperidin, nobiletin and tangeretin. Studies on accumulation of secondary metabolites of the orange leaves and orange peel, and then compare the four kinds of flavonoids in the process of dynamic change rule.2.1Comparison the4kinds of flavonoids from orange leaves in different growth periodsThe results show that the compounds that have the highest content is hesperidin, accounting for4.283%,at the orange leaves in August,and the contents of narirutin, hesperidin, nobiletin and tangeretin showed a downward trend overall.2.2Preliminary comparison the4kinds of flavonoids from orange peeld and orange leaves in different growth periodThe components four kinds of flavonoids in the orange leaves are less than the content of orange peel on July15. Also on December14collection of sample also have those rule. While the other three flavonoid(narirutin, nobiletin and tangeretin) content had significant difference. This research for the orange leaf provides theory basis for the clinical application of herbs.3. Research the HPLC fingerprint3.1A comparative study from the same plant in different growth period orange peel HPLC fingerprintAt present, the mainly content of hesperidin as indicators of the quality control of orange peel, and single hesperidin content cannot fully reflect the quality of the orange peel, HPLC fingerprint spectrum in reflect the differences between chemical composition of species type, and quality control may play an important role. By the HPLC technology, this study takes the lead in comparing the fingerprint of methanol extraction from orange peels of Citrus reticulata Blanco from the same plant at different growth period. Using the system,"Chinese medicine chromatographic fingerprint similarity evaluation system"(2004edition), which the state pharmacopoeia commission issued it, analysis the fingerprint. The results show that different growth period orange peels contains the main chemical composition similar types. In young fruit period synephrine relative peak area values greater than the fruit mature period of relative peak area; Nobiletin and tangeretin were significantly degressive trend; those results are consistent with the chemical composition content determination results.3.2Research different citrus varieties of HPLC fingerprintThe citris peels are from the Citrus reticulata Blanco and its cultivars of fruit, Including the C. Hort. et Tanaka.the C. chachiensis Hort,the C. suhoiensis Hort. ex Tanaka, etc. Its cultivars of fruit are records by the Pharmacopoeia of the People’s Republic of China (2010edition):the C. tangerina Hort,the C. erythrosa Tanaka,the C. unshiu Marcoritch,the C. subcompressa Tanaka.By the HPLC technology, this study takes the lead in comparing the fingerprint of methanol extraction from orange peels of Citrus reticulata Blanco from the same plant at different growth period. Using the system,"Chinese medicine chromatographic fingerprint similarity evaluation system"(2004edition), which the state pharmacopoeia commission issued it, analysis the fingerprint. Different varieties of citrus relative front area ratio has significant difference. Also found that the ponkan and the reference spectra is similarly, but the proportion of each component and the control medicinal material have differences. The Pharmacopoeia of the People’s Republic of China (2010edition) contains citrus varieties similarity is very close, but in proportion to the various components are different.And the SPSS18statistical software is used to cluster analysis the difference between the samples.4. A study on the Dynamic Change law of the Synephrine and the total Flavone in Citrus reticulate ’Chachi’ from Different Storage PeriodBy the HPLC technology, this study takes building the method for determining simultaneously the contents of synephrine.By the UV technology, this study building the methods for determination of the total flavonoids. The results show that with the storage the extension of time, The total flavone content have a lower trend, but change tiny.The content of synephrine was in lower trend,the synephrine decreased from0.273%to0.1097%. |
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