ABSTRACT
Iso‘avones are the largest group of iso‘avonoids, a distinctive subclass of ‘avonoids (Figure 19.1). By 2007, ~1600 iso‘avonoids had been characterized and described (Tahara, 2007). Depending on their substitution patterns, they can be divided into three different groups: those with simple O-substitutions (e.g., hydroxy or methoxy groups), prenylated forms, and glucosides (Veitch, 2007). The main structural difference between iso‘avonoids and ‘avonoids is the position of the B-ring, which is attached at the 3-position instead of the 2-position. The 3-phenylchroman skeleton of these compounds is derived by rearrangement of 2-phenylchroman (the skeleton of ‘avonoids) by 1,2-aryl rearrangement (Dewick, 1993). This rearrangement is catalyzed by an enzyme commonly referred to as iso‘avone synthase (Tahara and Ibrahim, 1995), although this is a term of convenience describing a sequence of reactions involving 2-hydroxyiso‘avone synthase (2HIS, a microsomal cytochrome P450 of the CYP93C subfamily) and 2-hydroxyiso‘avone dehydrase (HID or 2HID) (Veitch, 2009). In the ‚rst 2HIS-catalyzed step, a hydrogen is abstracted from C-3 by heme-bound oxygen in an NADPH-dependent reaction, followed
19.1 Introduction .................................................................................................................................. 405 19.1.1 Estrogenic Activities of Iso‘avones ................................................................................ 408
19.2 Analytical Methods ...................................................................................................................... 409 19.2.1 Sampling .......................................................................................................................... 409 19.2.2 Sample Storage ................................................................................................................ 409 19.2.3 Analyte Isolation ............................................................................................................. 409
19.2.3.1 Solvent Extraction .............................................................................................410 19.2.3.2 Ultrasound-Assisted Methods ...........................................................................411 19.2.3.3 Microwave-Assisted Methods ...........................................................................415 19.2.3.4 Supercritical Fluid Extraction ...........................................................................415 19.2.3.5 Pressurized Liquid Extraction ..........................................................................415 19.2.3.6 Sample Stability during Extraction ................................................................. 420
19.2.4 Sample Hydrolysis and Deconjugation ............................................................................ 420 19.2.5 Postextraction Treatment ................................................................................................. 420
19.2.5.1 Solid-Phase Extraction ......................................................................................421 19.2.5.2 High-Speed Countercurrent Chromatography ................................................ 422
19.2.6 Analytical Techniques ..................................................................................................... 422 19.2.6.1 Chromatography .............................................................................................. 423 19.2.6.2 Capillary Electrophoresis ................................................................................ 424 19.2.6.3 Immunoassays .................................................................................................. 425
19.2.7 Mass Spectrometric Methods for Iso‘avone Analysis .................................................... 425 19.2.7.1 Hyphenated Techniques ................................................................................... 425 19.2.7.2 Nonhyphenated Techniques ............................................................................. 426
References .............................................................................................................................................. 427
by the migration of the aryl group from C-2 to C-3, and the rebinding of the hydroxyl radical to the carbon radical at C-2 (Hashim et al., 1990). This reaction is stereoselective, using only 2S-‘avanones as substrate (Dixon, 2004), and requires a 2′- or 4′-hydroxyl group (Bhandari et al., 1992). The dehydration step from the 2-hydroxyiso‘avone is less well understood, although Akashi et al. (2005) suggested a mechanism in which amino acid residues of a catalytic triad and an oxyanion hole eliminate water cooperatively from the 2-position.
