A Simultaneous Analytical Method to Profile Non-Volatile Components with Low Polarity Elucidating Differences Between Tobacco Leaves Using Atmospheric Pressure Chemical Ionization Mass Spectrometry Detection
A comprehensive analytical method using liquid chromatography atmospheric pressure chemical ionization mass spectrometry detector (LC/APCI-MSD) was developed to determine key non-volatile components with low polarity elucidating holistic difference among tobacco leaves. Nonaqueous reversed-phase chromatography (NARPC) using organic solvent ensured simultaneous separation of various components with low polarity in tobacco resin. Application of full-scan mode to APCI-MSD hyphenated with NARPC enabled simultaneous detection of numerous intense product ions given by APCI interface. Parameters for data processing to filter, feature and align peaks were adjusted in order to strike a balance between comprehensiveness and reproducibility in analysis. 63 types of components such as solanesols, chlorophylls, phytosterols, triacylglycerols, solanachromene and others were determined on total ion chromatograms according to authentic components, wavelength spectrum and mass spectrum. The whole area of identified entities among the ones detected on total ion chromatogram reached to over 60% and major entities among those identified showed favorable linearity of determination coefficient of over 0.99. The developed method and data processing procedure were therefore considered feasible for subsequent multivariate analysis. Data matrix consisting of a number of entities was then subjected to principal component analysis (PCA) and hierarchical clustering analysis. Cultivars of tobacco leaves were distributed far from each cultivar on PCA score plot and each cluster seemed to be characterized by identified non-volatile components with low polarity. While fluecured Virginia (FCV) was loaded by solanachromene, phytosterol esters and triacylglycerols, free phytosterols and chlorophylls loaded Burley (BLY) and Oriental (ORI) respectively. Consequently the whole methodology consisting of comprehensive method and data processing procedure proved useful to determine key-components among cultivars of tobacco leaves, and was expected to additionally expand coverage that metabolomics study has ensured. [Beitr. Tabakforsch. Int. 27 (2016) 60-73]
1. Davis, D. and M.T. Nielsen: Tobacco: Production, Chemistry and Technology; edited by D. Davis and M.T. Nielsen, Blackwell Science, Oxford, UK,1999; ISBN-13: 978-0632047918.
3. Rodman, A. and T.A. Perfetti: The Chemical Components of Tobacco and Tobacco Smoke, Second Edition; CRC Press, Taylor and Francis Group, Boca Raton, FL, USA, 2013; ISBN: 9781466515482.
4. Leffingwell, J.C.: Basic Chemical Constituents of Tobacco Leaf and Differences among Tobacco Types; in: Tobacco: Production, Chemistry and Technology; edited by D. Davis and M.T. Nielsen, Blackwell Science, Oxford, United Kingdom, 1999, pp. 265-284; ISBN-13: 978-0632047918.
5. Stedman, R.L.: The Chemical Composition of Tobacco and Tobacco Smoke; Chem. Rev. 68 (1968) 153-207; DOI: 10.1021/cr60252a002.
6. Schmeltz, I. and D. Hoffmann: Nitrogen-Containing Compounds in Tobacco and Tobacco Smoke; Chem. Rev. 77 (1977) 295-311; DOI: 10.1021/cr60307a001.
7. Krishnan, P., N.J. Kruger, and R.G. Ratcliffe: Metabolite Fingerprinting and Profiling in Plants Using NMR; J. Exp. Bot. 56 (2005) 255-265; DOI:10.1093/jxb/ eri010.
8. Zhang, L., X. Wang, J. Guo, Q. Xia, G. Zhao, H. Zhou, and F. Xie: Metabolic Profiling of Chinese Tobacco Leaf of Different Geographical Origins by GC-MS; J. Agric. Food Chem. 61 (2013) 2597-2605; DOI: 10.1021/jf400428t.
9. Zhao, Y., C. Zhao, X. Lu, H. Zhou, Y. Li, J. Zhou, Y. Chang, J. Zhang, L. Jin, F. Lin, and G. Xu: Investigation of the Relationship between the Metabolic Profile of Tobacco Leaves in Different Planting Regions and Climate Factors Using a Pseudotargeted Method Based on Gas Chromatography/Mass Spectrometry; J. Proteome Res. 12 (2013) 5072-5083; DOI: 10.1021/ pr400799a.
10. Zhang, J., Y. Zhang, Y. Du, S. Chen, and H. Tang: Dynamic Metabonomic Responses of Tobacco (Nicotiana tabacum) Plants to Salt Stress; J. Proteome Res. 10 (2011) 1904-1914; DOI: 10.1021/pr101140n.
11. Cho, K., Y. Kim, S.J. Wi, J.B. Seo, J. Kwon, J.H. Chun, K.Y. Park, and M.H. Nam: Nontargeted Metabolite Profiling in Compatible Pathogen-Inoculated Tobacco (Nicotiana tabacum L. cv. Wisconsin 38) Using UPLCQ- TOF/MS; J. Agric. Food Chem. 60 (2012) 11015-11028; DOI: 10.1021/jf303702j.
12. Rowland, R.L., P.H. Latimer, and J.A. Giles: Flue- Cured Tobacco. I. Isolation of Solanesol, an Unsaturated Alcohol; J. Am. Chem. Soc. 18 (1956) 4680-4683; DOI: 10.1021/ja01599a041.
13. Ishida, N.: A Novel Method for Analyzing Solanesyl Esters in Tobacco Leaves Using Atmospheric Pressure Chemical Ionization/Mass Spectrometer; J. Chromatogr. A 1217 (2010) 5794-5801; DOI: 10.1016/j. chroma.2010.07.037.
14. Ishida, N.: Expanded Separation Technique for Chlorophyll Metabolites in Oriental Tobacco Leaf Using Non- Aqueous Reversed Phase Chromatography; J. Chromatogr. A 1218 (2011) 5810-5818; DOI: 10.1016/j. chroma.2011.06.082.
15. Ishida, N.: A Comprehensive Study on Triacylglycerols in Tobacco Leaves Using Liquid Chromatography and Atmospheric-Pressure Chemical-Ionization Mass Spectrometry; Beitr. Tabakforsch. Int. 25 (2013) 627-637; DOI: 10.2478/cttr-2013-0939.
16. Rowland, R.L.: Flue-Cured Tobacco. III. Solanachromene and α-Tocopherol; J. Am. Chem. Soc. 80 (1958) 6130-6133; DOI: 10.1021/ja01555a057.
17. Ishida, N.: A Method for Simultaneous Analysis of Phytosterols and Phytosterol Esters in Tobacco Leaves Using Non-Aqueous Reversed Phase Chromatography and Atmospheric Pressure Chemical Ionization Mass Spectrometry Detector; J. Chromatogr. A 1340 (2014) 99-108; DOI: 10.1016/j.chroma.2014.03.021.
18.Want, E.J., A. Nordström, H. Morita, and G. Siuzdak: From Exogenous to Endogenous: The Inevitable Imprint of Mass Spectrometry in Metabolomics; J. Proteome Res. 6 (2007) 459-468; DOI: 10.1021/ pr060505+.
19. Lu, W., B.D. Bennett, and J.D. Rabinowitz: Analytical Strategies for LC-MS-Based Targeted Metabolomics; J. Chromatogr. B 871 (2008) 236-242; DOI: 10.1016/ j.jchromb.2008.04.031.
20. Kuehnbaum, N.L. and P. Britz-McKibbin: New Advances in Separation Science for Metabolomics: Resolving Chemical Diversity in a Post-Genomic Era; Chem. Rev. 113 (2013) 2437-2468; DOI: 10.1021/cr300484s.
21. Hurtado-Fernández, E., T. Pacchiarotta, E. Longueira- Suárez, O.A. Mayboroda, A. Fernández-Gutiérrez, and A. Carrasco-Pancorbo: Evaluation of Gas Chromatography-Atmospheric Pressure Chemical Ionization-Mass Spectrometry as an Alternative to Gas Chromatography-Electron Ionization-Mass Spectrometry: Avocado Fruit as Example; J. Chromatogr. A 1313 (2013) 228-244; DOI: 10.1016/j.chroma.2013.08.084.
22. Cho, K., Y. Kim, S.J. Wi, J.B. Seo, J. Kwon, J.H. Chung, K.Y. Park, and M.H. Nam: Metabolic Survey of Defense Responses to a Compatible Hemibiotroph, Phytophthora parasitica var. nicotianae, in Ethylene Signaling-Impaired Tobacco; J. Agric. Food Chem. 61 (2013) 8477-8489; DOI: 10.1021/jf401785w.
23. Monton, M.R.N. and T. Soga: Metabolome Analysis by Capillary Electrophoresis-Mass Spectrometry; J. Chromatogr. A 1168 (2007) 237-246; DOI: 10.1016/ j.chroma.2007.02.065.
24. Sandra, K., M. Moshir, F. D'hondt, K. Verleysen, K. Kasa, and P. Sandra: Highly Efficient Peptide Separations in Proteomics Part 1. Unidimensional High Performance Liquid Chromatography; J. Chromatogr. B 866 (2008) 48-63. DOI: 10.1016/j.jchromb.2007.10.034.
25. Dugo, P., M. Beccaria, N. Fawzy, P. Donato, F. Cacciola, and L. Mondello: Mass Spectrometric Elucidation of Triacylglycerol Content of Brevoortia tyrannus (Menhaden) Oil Using Non-Aqueous Reversed-Phase Liquid Chromatography Under Ultra High Pressure Conditions; J. Chromatogr. A 1259 (2012) 227-236; DOI: 10.1016/j.chroma.2012.03.067.
26. Bamba, T., J.W. Lee, A. Matsubara, and E. Fukusaki: Metabolic Profiling of Lipids by Supercritical Fluid Chromatography/Mass Spectrometry; J. Chromatogr. A 1250 (2012) 212-21; DOI: 10.1016/j.chroma.2012.05.068.
27. Vrhovsek, U., D. Masuero, M. Gasperotti, P. Franceschi, L. Caputi, R. Viola, and F. Mattivi: A Versatile Targeted Metabolomics Method for the Rapid Quantification of Multiple Classes of Phenolics in Fruits and Beverages; J. Agric. Food Chem. 60 (2012) 8831-8840; DOI: 10.1021/jf2051569.
28. Abu-Reidah, I.M., M.M. Contreras, D. Arráez-Román, A. Segura-Carretero, and A. Fernández-Gutiérrez: Reversed-Phase Ultra-High-Performance Liquid Chromatography Coupled to Electrospray Ionization- Quadrupole-Time-of-Flight Mass Spectrometry as a Powerful Tool for Metabolic Profiling of Vegetables: Lactuca sativa as an Example of its Application; J. Chromatogr. A 1313 (2013) 212-227; DOI: 10.1016/ j.chroma.2013.07.020.
29. Gan, H.H., C. Soukoulis, and I. Fisk: Atmospheric Pressure Chemical Ionisation Mass Spectrometry Analysis Linked With Chemometrics for Food Classification - A Case Study: Geographical Provenance and Cultivar Classification of Monovarietal Clarified Apple Juices; Food Chem. 146 (2014) 149-156; DOI: 10.1016/j.foodchem.2013.09.024.
30. Parris, N.A.: Non-Aqueous Reversed-Phase Liquid Chromatography: A Neglected Approach to the Analysis of Low Polarity Samples; J. Chromatogr. A 157 (1978) 161-170; DOI:10.1016/S0021-9673(00)92332-X.
31. Yang, S., M. Sadilek, and M.E. Lidstrom: Streamlined Pentafluorophenylpropyl Column Liquid Chromatography- Tandem Quadrupole Mass Spectrometry and Global 13C-labeled Internal Standards Improve Performance for Quantitative Metabolomics in Bacteria; J. Chromatogr. A 1217 (2010) 7401-741; DOI: 10.1016/ j.chroma.2010.09.055.