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.e., room air) fruit. However, exposure to MAs of ultralow (e.g., less than 0.25%) O 2 and/or high (e.g., greater than 20%) CO 2 are frequently associated with the development of off-flavors (i.e., acetaldehyde, ethyl acetate, ethanol volatiles) in

International Ltd., Maidstone, Kent, England) at room temperature. The resulting mixtures were pooled and evaporated to the aqueous phase, which was subsequently adjusted to pH 10 with 1 m KOH and partitioned against 100% ethyl acetate. After centrifuging the

, higher acids, higher limonoids and some flavonoids, and lower top-note esters (ethyl acetate and ethyl butanoate) than juice of fruit from healthy trees or asymptomatic fruit from infected trees ( B aldwin et al., 2010 ; B assanezi et al., 2009 ; D

were further identified. α-Pinene (>99% purity; Fluka Chemical Corp., Ronkonkoma, NY) was selected as the external standard and ethyl acetate was used as the solvent (initial concentration was 0.858 g·mL −1 ). Four dilutions (×100, ×500, ×1000, and

acetate (7:0.96:0.04:2, mobile phase B) was pumped for 1 min. Finally, acetonitrile:methanol:water:ethyl acetate (7:0.96:0.04:8, mobile phase C) was pumped until all the components were eluted. The doubly distilled water was filtered with a HPLC solvent

vigorously. C-KODA was extracted using ethyl acetate, and isolated with high-performance liquid chromatography (NANOSPACE SI-1, Shiseido, Tokyo, Japan) using a CAPCELL PAK C 18 column (Shiseido). The mobile phase was 50% CH 3 CN in 0.05% trifluoroacetic acid

extracted twice with 15 mL n-hexane:ethyl acetate (9:1). The organic phase was collected, evaporated by Rotavapor (IKA, Labortechnik, Staufen, Germany), and the dry residue was dissolved in 1 mL 98% (v/v) methanol. A sample volume of 20 mL was separated by

with internal standards of [ 2 H 2 ]-ABA and reduced to aqueous phase. The aqueous phase was extracted with ethyl acetate at pH 3, then with potassium phosphate buffer at pH 8.5, again into ethyl acetate at pH 3, and further purified on C 18 Sep

to 2.5 with 0.1 m phosphoric acid and extracted three times with 20 mL 100% v/v ethyl acetate to concentrate it to dryness and then dissolved in 1 mL of 25% acetonitrile containing 20 m m acetic acid. The solution was filtered and then subjected to

.5% TTC in 0.06 mol·L −1 phosphatic buffer, pH 7.4) for 3 h at 37 °C in the dark. Then 2 mL of 1 mol·L −1 H 2 SO 4 was added to the tube to stop the reaction. The TTC solution was drained. Roots were ground and extracted with 10 mL of ethyl acetate and