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ICOS 2013 Proceedings Identification and Characterization of Potent Odorants in Chinese Green Tea (Longjing, Maofeng, Biluochun) Infusions Ryoko Baba and Kenji Kumazawa Ogawa & Co., Ltd., 15-7 Chidori Urayasushi, Chiba 279-0032, Japan Correspondent email: [email protected] Summary The volatile fractions of three famous Chinese green tea cultivar (Longjing, Maofeng, Biluochun) infusions were isolated by a combination of the adsorptive column method and the SAFE techniques. The aroma extract dilution analysis (AEDA) applied to the volatile fractions revealed 58 odor-active peaks with FD factors between 41 and 47. Forty-six of the odorants, which included three newly identified odorants in Chinese green tea, were identified or tentatively identified by GC-MS and GC-O. Among the perceived odorants, seven odorants showed high FD factors in all the cultivars. On the other hand, thirteen odorants were characteristic of each cultivar. In addition, the formation mechanism of (E)-isoeugenol was also investigated, and it was assumed that (E)-isoeugenol is formed from a nonvolatile precursor such as a glycoside via coniferyl alcohol. These findings suggested that the formation of (E)-isoeugenol has a close connection with the manufacturing process of the tea leaves. Introduction Chinese green tea, known for its characteristic pan-fired and floral aromas, is the most popular tea in China. There are numerous Chinese green tea cultivars classified by the subtle differences in the producing area and/or the manufacturing process. Furthermore, most of Chinese green tea products are said to be affected by the endogenous enzymatic reaction during the manufacturing process. The potent odorants of Chinese green tea have been reported by Kumazawa and Masuda (2002). However, this investigation was only intended for a single cultivar, despite there being many. Therefore, the potent odorants common to many Chinese green tea cultivars and characteristic of each cultivar are still unclear. The objective of the present investigation was to elucidate the potent odorants common to the three famous cultivars (Longjing, Maofeng, Biluochun) and characteristic of each cultivar. In addition, the formation mechanism of (E)-isoeugenol was also investigated. Materials and methods Materials: Chinese green tea products Longjing, Maofeng, and Biluochun were produced in the Zhejiang, Anhui, and Jiangsu province (China) in 2012. Sen-cha was produced in the Shizuoka prefecture (Japan) in 2008. These products were high grade based on the market price. Isolation of the Volatiles: Distilled water (90 ºC, 1500 g) was added to 75 g of Chinese green tea, and the leaves were filtered after standing for 5 min. The filtrate (1000 g) was passed through a column packed with 20 mL of a porous polymer adsorbed resin (SP700). The adsorbed compounds were eluted with methylene chloride (100 mL) and the eluate was distilled under reduced pressure (< 4.0 × 10-3 Pa, 40 ºC) using the SAFE technique. The distillate concentrated to ~100 µL was used as the sample for the AEDA and the GC-MS analysis. Model experiments for (E)-isoeugenol formation: The Sen-cha infusion (200 g) was passed through a column packed with 5 mL of SP700. The glycosidic compounds were eluted with methanol (50 mL). The lyophilized glycosidic fraction was solved in the buffer, and then treated with pectriase (pH 4.5, 40 ºC, 3 h.). The released volatile compounds were analyzed by the GC-MS. Coniferyl alcohol mixed with cellulose powder was heated (150 ºC, 30 min.), and then analyzed by the GC-MS using the Solvent Phase Microextraction (SPME) technique. 1 Results and discussion The AEDA applied to the volatile fractions of the Chinese green tea cultivar (spring and summer crops of each sample) infusions revealed 58 odor-active peaks with FD factors between 41 and 47. Forty-six of the odorants, which included three newly identified odorants in Chinese green tea, were identified or tentatively identified by GC-MS and GC-O. Among the perceived odorants, seven odorants (sweet, floral, burnt) showed high FD factors in all the cultivars, and therefore, it was suggested that those odorants are essential for the Chinese green tea flavor (Figure 1). On the other hand, it was expected that the Figure 1. The potent odorants common to all the cultivars amounts of the 13 odorants Table 1. The potent odorants characteristic of each cultivar influence the characteristics of each cultivar (Table 1). Coniferyl alcohol, which is known as an important intermediate of (E)-isoeugenol in plants (Koeduka, et al. (2006), Sakai (1994)) was released from the glycosidic fraction by pectriase treatment. Therefore, coniferyl alcohol is indicated to exist as a glycoside like coniferin in the tea leaves. In addition, (E)-isoeugenol was generated from coniferyl alcohol by heating. These results indicate that (E)-isoeugenol is formed from such a nonvolatile precursor such as a glycoside via coniferyl alcohol during the withering and pan-firing process of the tea leaves (Figure 2). Therefore, it was suggested that the formation of (E)-isoeugenol has a close connection with the manufacturing Figure 2. Hypothetical generation mechanism of (E)-isoeugenol process of the tea leaves. in the tea leaves during the manufacturing process of Chinese green tea products References Kumazawa, K., Masuda, H. (2002) Identification of potent odorants in different green tea varieties using flavor dilution technique. J. Agric. Food Chem. 50:5660-5663 Koeduka, T., Fridman, E., Gang, D., Vassao, D., Jackson, B., Kish, C., Orlova, I., Spassova, S., Lewis, N., Noel, J., Baiga, T., Dudareva, N., Pichersky, E. (2006) Eugenol and isoeugenol, characteristic aromatic constituents of spices, are biosynthesized via reduction of a coniferyl alcohol ester. PNAS 103: 10128-10133 Sakai, K. (1994) Organosolv delignification. JAPAN TAPPI JOURNAL 48:1003-1012 2 .
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