| Background: Xinhui chenpi is the the dry peel of Citrus reticulata ‘Chachi’,which is cultivated in xinhui district of Jiangmen city,Guangdong province.According to the different harvest stages,it can be divided into green peel,semi-red peel and red peel.Pharmacological studies have illustrated that xinhui chenpi exhibit the good effects of regulating qi-flowing for strengthening spleen,harmonize the stomach and check vomiting,removing dampness to reduce phlegm and so on,which is highly praised by many traditional famous doctors and consumed as genuine medicinal herb.And it is widely used in the fields of Chinese medicine treatment,food,cosmeceuticals and so on.According to statistical analysis,there are nearly 100,000 mu in xinhui chenpi planting and more than 5,000 tons xinhui chenpi produced per year.However,the production and planting of xinhui chenpi are decentralized,and the lack of systemical study of harvesting,processing and storage leads to the different quality,which is not conducive to the development of xinhui chenpi industrialization.Thus,in this paper,the dynamic changes in the chemical components during harvesting,processing and storage of xinhui chenpi were analyzed comprehensively,including flavonoids,volatile oil,alkaloids,phenolic acids,and limonins.It is benefit to solve the important industrialization issues during harvesting,processing,storage and to provide scientifical reference for its quality control.In addition,the inhibitory effect of polymethoxyflavones on nasopharyngeal carcinoma would be clarified through a series of pharmacological experiments,which provided scientific basis for the development and application of xinhui chenpi.Aims:(1)To evaluate the dynamic changes of chemical components in xinhui chenpi during harvesting,drying and two storage years.(2)To study the inhibitory effect of nobiletin and 3,5,6,7,8,3’,4’-heptamethoxyflavone(HMF)from xinhui chenpi on human nasopharyngeal carcinoma 5-8F cells.Methods:(1)Ultraviolet-visible spectrophotometry(UV)was used to determine the total contents of flavonoids,polymethoxyflavones,phenolic acids,and high performance liquid chromatography(HPLC)was applied for the simultaneous determination of synephrine,hesperidin,ferulic acid,limonin,nobiletin,HMF,tangeretin,5-hydroxy-6,7,8,3’,4’-pentamethoxyflavone(5-HPMF).(2)The volatile oil was extracted by steam distillation and futher identified by gas chromatographymass spectrometry(GC-MS).Then the main volatile components were screened for principal component analysis(PCA).(3)The cell counting kit 8(CCK8)assay,colony formation assay,flow cytometry assay,hoechst assay,scratch-wound assay and transwell assay were used to investigate the effects of nobiletin and HMF on the proliferation,apoptosis,migration and invasion of 5-8F cells.Results:(1)The total flavonoids,total polymethoxyflavones,total phenolic acids,synephrine,hesperidin,limonin,nobiletin,HMF,tangeretin,5-HPMF of xinhui chenpi all showed a decreasing trend with the delay of harvest time.There was no significant difference among the extraction rate of volatile oil from different harvest periods.Besides,the content of volatile alcohols decreased with the delay of harvest time,while the content of volatile aldehydes,phenols and ketones increased firstly and then decreased.It is effective to apply PCA to distinguish xinhui chenpi from different harvest periods.(2)The content of flavonoids,especially polymethoxyflavones,was decreased significantly in xinhui chenpi dried at 60℃,while the other components had no significant change under different drying conditions.There was no significant difference among the extraction rate of volatile oil under different drying conditions.In addition,the content of volatile alcohols,phenols,aldehydes,ketones and esters were decreased,while the content of volatile alkenes were increased at 60℃ drying.PCA was effectively applied for distinguishing xinhui chenpi from different drying conditions.(3)The synephrine,hesperidin,limonin,HMF,5-HPMF all showed a decreasing trend after two years of storage,while ferulic acid increased significantly after one year of storage.There was no significant difference in nobiletin and tangeretin within two years.As for the volatile oil,the green peel showed an obvious decreasing trend,while the semi-red peel and red peel showed no obvious change.The numbers of volatile components decreased significantly after storage for one year,but there was little change during storage for one to two years.Whta’s more,the green peel showed a decreased trend in alcohols during storage,while increased in phenols and esters.The semi-red peel and red peel showed a decreased trend in alcohols,phenols,esters and aldehydes during storage,whereas increased in alkenes.The result of PCA showed that there was some differences among the volatile components of xinhui chenpi during two years of storage.(4)The pharmacological results showed that the viability,migration and invasion rate of 5-8F cells decreased,while the apoptosis rate increased with the increase of nobiletin and HMF concentration.Conclusion:(1)The non-volatile components of xinhui chenpi showed a decreasing trend with with the delay of harvest time.And the volatile components changed in different harvest periods.(2)High temperature drying at 60℃ induced a significant transformation in the flavonoids(especially polymethoxyflavones)and volatile substances of xinhui chenpi.Thus,it is believed that the fresh peel should be naturally sun-dried or dried at low temperature(30℃ or 45℃)rather than at high temperature(60℃).(3)Most of the non-volatile components in xinhui chenpi showed a decreasing trend during two storage years except ferulic acid,nobiletin and tangeretin.And there was significant difference in volatile components within two years.Therefore,it is believed that the xinhui chenpi had better effect when stored for a longer time.(4)Nobiletin and HMF inhibited proliferation,migration,invasion and induced apoptosis of nasopharyngeal carcinoma 5-8F cells.And the inhibition effect of HMF was stronger than nobiletin. |
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