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8 3/14 eFOOD-Lab international Table 1: Example of samples used for volatiles analysis with Tenax®, SPME, TwisterTM and MonoTrapTM Fig. 2: Results of a clustering analysis (lowest concentration-blue, highest concentration-red) of volatiles significantly varying between the treatments (t-test, p < 0,05, in min. 70 % of the samples) and PCA on the metabolites contained in plants at 1 and 2 years old Sampling technique Plant/Organ Reference TwisterTM Tomato fruits 2,3 TwisterTM Tomato leaves 4 Tenax®, TwisterTM Leaves of Camelia sinensis (Fig. 1B) 1 Tenax®, SPME Flowers of Osmanthus fragrans, enzymatic products of the reaction 5,6 Tenax®, SPME Leaves of Camelia sinensis 7 SPME Algae Enteromorpha compressa (L.) Nees 8 SPME Flowers of Camelia sinensis 9 SPME Green algae Ulva prolifera, Ulva linza, Monostroma nitidum 10 MonoTrapTM, Tenax®, SPME Flowers of Rosa chinensis ‚Mutabilis‘, transformation of β-Ionone (Figs. 1A & 1C) 11 provement of the identification of the compounds, the GC-ToF Example of application (Time of Flight) will always provide the best accuracy. Data analysis In a general way, the use of commercial database (NIST, Wiley) enables the identification of compounds. Additionally, calculations based on the Retention Index (Kovat´s Index) improves identification. Nevertheless, internal standards are commonly used to enable a precise quantification eg. 1-9. Non-targeted analyses are commonly carried out to compare samples of different treatments by modern coupling techniques combined with modern software solutions. These software packages combine peak identification, statistical analysis and identification of compounds. Data processing is achieved by following a similar approach as for metabolomics: the signals of the metabolites are extracted. Then, the compounds varying significantly between the treatments are retained and identification is achieved by comparisons with databases. Statistical analyses such as clustering analysis or PCA (Principal Component Analysis) enable to distinguish between sample groups (Fig. 2). Summary Tenax®, TwisterTM, SPME and MonoTrapTM can be used for direct-sampling to analyse volatile compounds without using any organic solvent. Modern analytical methods combined with software solutions offer various possibilities to analyse metabolites and to study the influence of different eco-physiological factors. References 1. Katsuno, T, Kasuga, H, Kusano, Y, Yaguchi, Y, Tomomura, M, Cui, J, Yang, Y, Baldermann, S, Nakamura, Y, Ohnishi, T, Mase, M, Watanabe, N, Characterisation of odorant compounds and their biochemical formation in green tea with a low temperature storage process. Food Chemistry, 148 (1) 388-395 (2014). 2. Krumbein, A, Schwarz D, Grafting: A possibility to enhance healthpromoting and flavour compounds intomato fruits of shaded plants? Scientia Horticulturae 149, 97–107 (2013). 3. Krumbein, A., Hilfert, L. & Krause, K. Stir bar sorptive extraction of tomato aroma volatiles. In: State-of-the-Art in Flavour Chemistry and Biology T Hofmann, W. Meyerhof, P. Schieberle (Eds.) 342-345 (2005). 4. Errard, A, Baldermann, S, Mewis, I, Kühne, S, Parolin, P, Ulrichs, C. Comparison between single and multiple pest infestation on plant biochemistry and physiology. Annual Conference of the Ecological Society of Germany, Switzerland and Austria (GfÖ), Potsdam, Germany p. 1999 (2013). 5. Baldermann, S, Kato, M, Kurosawa, M, Kurobayashi, Y, Fujita, A, Fleischmann, P, Watanabe, N. Functional characterization of a carotenoid cleavage dioxygenase 1 and its relation to the carotenoid accumulation and volatile emission during the floral development of Osmanthus fragrans Lour.. Journal of Experimental Botany, 61(11):2967-2977 (2010). 6. Baldermann, S, Kato, M, Fleischmann, P, Watanabe, N. Biosynthesis of α- and β-ionone, prominent scent compounds, in flowers of Osmanthus fragrans. Acta Biochimica Polonica, 59, 79–81 (2012). 7. Dong, F, Yang, Z, Baldermann, S, Sato, Y, Asai T, Watanabe, N. Herbivore-induced volatiles from tea (Camellia sinensis) plants and their involvement in intra-plant communication leading to changes in endogenous metabolites. Journal of Agricultural and Food Chemistry, 59, 13131–13135 (2011). 8. Baldermann, S, Fleischmann, P, Bolten, M, Watanabe, N, Winterhalter, P, Ito, Y. Centrifugal precipitation chromatography, a powerful technique for the isolation of active enzymes from tea leaves (Camellia sinensis). Journal of Chromatography A, 1216:4263–4267 (2009). 9. Dong, F, Yang, Z, Baldermann, S, Kajitani, Y, Ota, S, Kasuga, H, Imazeki, Y, Ohnishi, T, Watanabe, N. Characterization of L-phenylalanine metabolism to acetophenone and 1-phenylethanol in the flowers of Camellia sinensis using stable isotope labeling. Journal of Plant Physiology, 169, 217-225 (2012). 10. Yamamoto, M, Baldermann, S, Yoshikawa, K, Fujita, A, Mase, N, Watanabe N, Determination of volatile compounds in four commercial samples of Japanese green algae using solid phase microextraction gas chromatography mass spectrometry. The Scientific World Journal, Volume 2014, Article ID 289780, 8 (2014). 11. Baldermann, S, Hirata, H, Ueda, Y, Winterhalter, P, Fleischmann, P, Watanabe, N. Changes in pigmentation and scent release during the floral development in Rosa chinensis ‘Mutabilis’. In: Advances and Challenges in Flavour Chemistry and Biology, T Hofmann, W. Meyerhof, P. Schieberle (Eds.) 201-206 (2010). 12. Chen, XM, Kobayashi, H, Sakai, M, Hirata, H, Asai, T, Ohnishi, T, Baldermann, S, Watanabe N. Functional characterization of rose phenylacetaldehyde reductase (PAR), an enzyme involved in the biosynthesis of the scent compound 2-phenylethanol. Journal of Plant Physiology, 168:88-95 (2011). Innovative Processi ng Tech nologies


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