Quantitative Chromatographic Fingerprinting Of Green Tea

Tea as a traditional beverage of China,and it has a history of thousands of years.The green tea is the largest yield and the most popular in China.In recent years,more bioactiveties of green tea extract have been published,such as antioxidant,anti-tumor,anti-inflammatory,reducing the risk of cardiovascular disease.The main factors that affect the quality of green tea inculde the surface temperature,average precipitation,altitude and sunshine duration.However,the current national standard of green tea is longer,and the analysis method is backward and time-consuming.How to establish a method for the analysis of the chemical components in green tea,and how to evaluate the quality of green tea effectively,quickly and accurately is an urgent problem to be solved.In this study,the green tea from different habitats were studied by high performance liquid chromatography with high resolution mass spectrometry:1.A total of 72 batches of green tea extracts from Yunnan and Hunan were analyzed by high performance liquid chromatography with high resolution mass spectrometry.The first,In order to make components of green tea fully extracted,the different green tea extraction solvent(water,pure methanol,70%methanol)and different dosage of samples(0.1g,0.5g,1.0g)were optimized.The results showed that using 70%methanol and 0.5g green tea sample extraction effect was the best.Secondly,in order to obtain more abundant mass spectrum information,the the mass spectrum conditions include in-source collision induced voltage(in-source CID)and high energy collision induced voltage(HCD)were optimized.So,to avoid excessive interference fragment,the in-source CID is chosen as 0eV.At the same time,in order to achieve more information about MS/MS spectrum and the stepted voltage(10 eV,30 eV,50 eV)was applied.Finally,the mixed standard and quality control samples were investigated in precision and stability of the method to ensure the stability of the experimental instrument in the process of analysis.The results showed that during the whole experiment,experimental instrument is stable and reliable.2.Qualitative and quantitative analysis of chemical components in green tea.The MZmine was applied in extraction and analysis of mass spectrometry.In order to extract the data of mass spectrometry as more effective as possible,the parameter of MZmine include peak correction,peak detection,peak extraction,deconvolution,peak alignment were optimized.We found that noise level is the most critical parameter.The results showed that noise level in 5.00E8 can eliminate background interference and can retain effective mass information.On this basis,the mass spectrometry data matched by the standards and the MS or MS/MS of several mass databases and chemical databases,75 compounds were identified in green tea inculded catechins,flavonoids,amino acids and so on,and 33 compounds were confirmed by standards.Taking 75 compounds as the chemcial features group,Semi quantitative analysis using internal standard method,and quantitative chromatographic fingerprint of Yunnan and Hunan green tea was established.Research shows that although the chemical fingerprint of two origin green tea is so similar,however,the content of each material is obvious difference.3.Analysis of the difference between Yunnan and Hunan green tea.First of all,the cluster analysis and principal component analysis which were the unsupervised pattern recognition method were applied to classified Yunnan and Hunan green tea.The results of classification show that two kinds of classification methods are to effectively distinguish Yunnan from Hunan green tea.On this basis,Partial least squares discriminant analysis(PLS-DA)combined with three variable selection methods(VIP,coefficient ?,Lasso)was used to discriminate green tea in Yunnan and Hunan.The results showed that PLS-DA combined with VIP method has the best ability of distinguishing,and it can be well divided into Yunnan and Hunan green tea.Three chemical features which Epicatechin-epigallocatechin-3-O-gallate(4beta->8),gallocatechin gallate(GCG)and vitexin were selected by VIP and the the content of GCG in Yunnan and Hunan was the most obvious.