| The study using the two chemical fingerprints characteristics of chemical fingerprints: "integrity" and "fuzziness" established the fingerprint of polyphenols and sugars in Luoyang areas,in order to make more objective,comprehensive and simple identification of Luoyang tobacco leaves,the experimental results are as follows :(1)Determine the HPLC method for determination of polyphenols in flue-cured tobacco of Luoyang by the optimization of extraction methods,solvent concentration,solvent consumption and extraction time.the extraction conditions of polyphenols in flue-cured tobacco were determined.: the extraction solution is 30 ml.80% methanol solution and ultrasonic 30 min,the final chromatographic conditions: Waters Sunfire C18 column(4.6×150mm,5μm),UV Detector,eluted with pure methanol(A)-0.01mol/L KH2PO4(p H=4.5)as the mobile phases in gradient mode(0-10 min,75%B-50%B;10-15 min,50%B-25%B;15-20 min,25%-25%B;20-25 min,25-75%B)at a flow rate of 1 ml/min,column temperature of 30℃,the wavelength of 340 nm.(2)The analysis of the polyphenols fingerprints of 36 samples was obtained that the relative retention time RSDs of common peaks had small differences,but the differences of the relative peak area RSDs of common peaks were large,the results showed that the main components of the 36 samples were basically the same,but the content of component was quite different.the similarity of HPLC fingerprints of 36 samples was showed that the similarity was 0.985-1.000,the similarity between 36 samples and control fingerprints(R1 and R2)showed that each batch of samples still had some differences,thought the similarity between 36 samples was high.(3)36 samples were divided into two categories by component analysis and clustering verification,which were Ι(1.2.3.4.5.13.14.16.17.18.34.35.36)from the parts of Π(6.7.8.9.10.11.12.15.19.20.21.22.23.24.25.26.27.28.29.30.31.32.33),the indicator of component analysis is 72% comparing with the classified results by the sense of evaluating experts,certification the results of cluster analysis are consistent with principal component analysis.By analysis the influence of the main component and the peak areas of two categories,rutin was eventually treated as the difference between two categories.(4)Determine the GC-MS method for determination of carbohydrates in flue-cured tobacco of Luoyang by the optimization of the flow rate,the heating process and the split ratio,the extraction conditions of Carbohydrates in flue-cured tobacco were determined : HP-5MS(60m×0.25 mm,df=0.25μm)quartz capillary column,programmed temperature: column temperature 150 ℃,4 ℃ /min rose to 230 ℃ maintained 3 minutes,25℃/min rose to 260℃ maintained 20 minutes,MS conditions: Inlet temperature is 280℃,ion source temperature is 230℃.(5)The analysis of the carbohydrates fingerprints of 36 samples was obtained that the main difference in two categories was still the contents of compound,The similarity of GC-MS fingerprints of 36 samples compared with control fingerprints was showed that the similarity was 0.973-0.996,the similarity between 36 samples and control fingerprints(R1 and R2)showed that each batch of samples still had some differences,thought the similarity between 36 samples was high.(6)36 samples were divided into two categories by component analysis and clustering verification,which were Ι(2.3.4.11.12.14.15.19.29)and Π(1.5.6.7.8.9.10.13.16.17.18.20.21.22.23.24.25.26.27.28.30.31.32.33.34.35.36)the indicator of component analysis is 60% comparing with the classified results by the sense of evaluating experts,the results were slightly worse than polyphenols.By analysis the influence of the main component and the peak areas of component of two categories,glucose was eventually treated as the difference between two categories.In this study,selecting 36 samples from 7 reigns of Luoyang for fingerprint analysis.Using HPLC-UV and GC-MS established the fingerprint of polyphenols and categories,classifying the production areas and typical material mining relatively comprehensive and accurate,comparing with traditional method,this method had the advantages of simple operation and objective comprehensiveness,which can provided scientific basis for origin discrimination and quality judgment of flue-cured tobacco. |
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