| Tea(Camellia sinensis),belong to Camelliaceae,evergreen shrubs or small tree,tea leaves made of tea which soaked in water for drinking,is China's traditional drinks.Tea is rich in Tea polyphenols,Caffeine,L-Theanine,Polysaccharides and other bioactive substances,with lower blood pressure,blood sugar,anti-aging,digestion,relieve fatigue and other effects.In this paper,We based on the ultra-high performance liquid chromatography-mass spectrometry method to establish several major active ingredients in tea analysis and detection methods,while using chemical colorimetric method to rapid detection of tea polyphenols.Designed to provide analysis and testing technical support for the evaluation of tea quality and the development of Hanzhong tea industry.Specific research contents can be mainly divided into the following aspects.(1)On optimized sample preparation condition,a determination method was developed using UPLC-MS/MS to simultaneously determined eight kinds of effective components in tea(Catechin,Epicatechin,Epigallocatechin,Gallocatechingallate,Epicatechingallate,Epigallocatechingallate,L-Theanine and Caffeine).Using Waters ACQUITY BEH C18 column separation,series quadrupole mass spectrometry multi-reaction ion monitoring and determination,external standard method quantitative.The chromatographic system flow rate was 250 ?L/min,the column temperature was 30°C,the mobile phase was 0.1% formic acid and acetonitrile,gradient elution.The results showed that the linearity of the 8 kinds of active components was good in the linear range,the correlation coefficient was more than 0.991,and the RSD of the precision was less than 5%,which indicateed that the method had good precision;the stability RSD of stability test value was less than 5%,which indicating that the method has good stability.The RSD values of the repeat experiments were less than 3%,which indicates that the method was reproducible.The average recoveries(n=5)ranged from 94.7% to 104.7%,and the detection limit of the method was between 3.0 and 60.0 ng/mL,and the method had a limit of quantitation between 6.0 and 200.0 ng/mL.(2)On the basis of single factor experiment,the preparation method of Tea polyphenols was optimized by using the response surface.The optimum conditions were as follows: the ratio of liquid to material was 26:1,the extraction time was 24 min,the extraction temperature was 91?,the pH value was 7.4,and the maximum yield was 20.56%.The samples were prepared under optimized conditions and the yield of tea polyphenols detecting and calculating by ultra performance liquid chromatography and mass spectrometry was 20.75%.The results showed that the preparation process and conditions were feasible,combined with UPLC-MS/MS could be used to qualitatively and quantitatively analyze tea polyphenols in tea.(3)A determination method was developed using UPLC-MS/MS to simultaneously determining 4 kinds of theaflavin(TF,TF-3-G,TF-3'-G,TF-3-3'-G)in black tea.Sample was extracted with 70% methanol by water bath,using Waters ACQUITY UPLCTMBEH C18 chromatographic column and Mass spectrometry was conducted by using positive(ESI+)electrospray ionization and Multi-reactions monitoring(MRM)models,with external standard method.The flow rate of chromatographic system was 250 ?L/min and at 30?.The mobile phase of acetonitrile was 0.1% methanoic acid and acetonitrile,with gradient elution.The results showed that the 4 kinds of theaflavins ingredients in their respective inside had good linear range and the correlation coefficient was greater than 0.9991.The average recovery was between 91.95% and 105.37%(n=5),RSD between 2.37% and 4.03%;Method detection limit was between 13.2 and 19.1 ng/mL,method of quantitative limit between 39.7 and 52.6 ng/mL.The detection method could quickly detect the content of theaflavins in black tea,reasonably control the black tea production quality.(4)Using chemical colorimetric technology,explored the rapid detection method for polyphenols by 0.02 g/mL ferric chloride solution,0.002 g/mL potassium permanganate solution and ferrous tartrate.The results showed that the chromaticity curve of ferric chloride was in the range of 0.10 to 0.70 mg/mL,which was unconfirmed than 0.70 mg/mL.The chroma curve of potassium permanganate was 0 to 0.80 mg/mL,which was not distinguished greater than 0.80 mg/mL.The ferrous tartrate curve had a corresponding accuracy in the concentration range between 0 and 0.50 mg/mL,which was unconfirmed after the concentration more than 0.50 mg/mL.The samples were tested using the established chromaticity curve,the results showed that the content of tea polyphenols detected by ferric chloride was 1.5 to 2.0 mg/mL,potassium permanganate was 1.20 to 1.80 mg/mL,the ferrous tartrate measured was 1.5 to 2.0 mg/mL.Based on the results of the three methods,it could be assumed that the concentration of the sample was between 1.50 and 1.80 mg/mL,but the sensitivity of the detection technology remained to be further studied.The method for the analysis of tea polyphenols,tea pigments,L-theanine and caffeine,had the characteristics of fast analysis,high sensitivity and high accuracy,and provided a new technology for tea quality analysis and evaluation.The construction of rapid detection technology of polyphenols provided a way for rapid analysis of active ingredients in tea. |
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