Flavonols dihydroflavonols

The flavonoid group is very diverse and contains several compounds including flavanones, flavones, flavonols, dihydroflavonols, isoflavonoids, anthocyanins, flavan-3,4-diols, flavan-4-ols, and flavan-3-ols. Flavan-3-ols are the structural units of the polymeric compounds termed condensed tannins abundant in plants. 

The precursors of these reactions are, on one hand, proanthocyanidins and, on the other hand, any kind of flavonoid that can act as a nucleophile. The latter include flavonols, dihydroflavonols, flavanol monomers, proanthocyanidins, and anthocyanins under their hemiketal form (for anthocyanin reactivity, see Chapter 9A). 

The phenolics in the grape berry are monomeric and polymeric molecules and are located in the juice (hydroxycinnamoyl tartaric acid esters), the solid part of the pulp (proanthocyanidins, hydroxybenzoic acids with structures reported in Figure 2.1), seeds (flavan-3-ols, proanthocyanidins, gallic acid) and the skin (anthocyanins, flavan-3-ols, proanthocyanidins, flavonols, dihydroflavonols, hydroxycinnamoyl tartaric acid esters, hydroxybenzoic acids, hydroxystilbens). Their levels in the grape are mainly linked to the variety, but can also be influenced by environmental variables, cultural techniques and the ripening state of the grape. 

A second proof comes from wounding experiment of Pseudowintera colorata Raoul (Gould et al, 2002b). In contrast to green leaves, the red ones, enriched in vacuolar antho-cyanins, flavonols, dihydroflavonols, and HCAs, alleviate intensity and duration of the localized burst of H2O2 induced in palisade mesophyll cells by wounding. Vacuolar phenolics participate to scavenge ROS. 

In 1934 the transformation of 2 -hydroxychalcones to flavonols in the presence of hydrogen peroxide and sodium hydroxide was reported simultaneously by Algar and Flynn in Ireland and Oyamada in Japan However, many reports following the original disclosures showed that the Algar-Flynn-Oyamada reaction could lead to several products including aurones 4, dihydroflavonols 5, 2-benzyl-2-hydroxydihydrobenzofuran-3-ones 6, and 2-arylbenzofuran-3-carboxylic acids... 

Flavonols and flavones have a double bond between C2 and C3 in the flavonoid structure and an oxygen atom at the C4 position. Furthermore, flavonols also have a hydroxyl group at the C3 position. Dihydroflavonols have the same structure as flavonols without the double bond between C2 and C3. 

Food and plant phenolics are commonly detected using DAD detectors (Tan and others 2008). Photodiode array detection allows collection of the entire UV spectrum during the elution of a chromatographic peak, which makes it possible to identify a phenolic compound by its spectra. Simple phenols, phenolic acids, flavanones, benzophenones, isoflavones, and flavan-3-ols have maximum absorbance at 280 nm, hydroxycinnamic acids at 320 nm, flavonols, flavones, and dihydroflavonols at 365 nm, and anthocyanins at 520 nm (Ibern-G6mez and others 2002 Merken Hand Beecher 2000). Hydrolyzable tannins show a characteristic shoulder at 300 nm, suitable for identifying them (Arapitsas and others 2007). For stilbenes, maximum absorbance of trans-forms are at 306 nm and at 285 nm for cA-forms (Lamuela-Raventos and others 1995). 

Figure 5.4. Abbreviated scheme for biosynthesis of major flavonoid subclasses, showing the primary enzymes and substrates leading to different subclasses. Bold-faced, uppercase abbreviations refer to enzyme names, whereas substrate names are presented in lowercase letters. PAL, phenylalanine ammonia lyase C4H, cinnamate 4-hydroxylase 4CL, 4-coumarate CoA ligase CHS, chalcone synthase CHI, chalcone isomerase CHR, chalcone reductase IPS, isoflavone synthase F3H, flavonone 3-hydroxylase F3 H, flavonoid 3 -hydroxylase F3 5 H, flavonoid 3 5 -hydroxylase FNSI/II, flavone synthase DFR, dihydroflavonol 4-reductase FLS, flavonol synthase ANS, anthocyanidin synthase LAR, leucoanthocyanidin reductase ANR, anthocyanidin reductase UFGT, UDP-glucose flavonoid 3-O-glucosyltransferase. R3 = H or OH. R5 = H or OH. Glc = glucose. Please refer to text for more information.

Table 6.1 Abbreviations BAN, BANYULS bHLH, basic helix-loop-helix CHS, chalcone synthase CHI, chalcone isomerase DFR, dihydroflavonol reductase F3H, flavonol 3-hydroxylase F3 H, flavonoid 3 -hydroxylase FLS, flavonol synthase icx, increased chalcone synthase expression LDOX, leucoanthocyanidin dioxygenase LCR, leucoanthocyanidin reductase MATE, multidrug and toxic compound extrusion NR, not yet reported tt, transparent testa ttg, transparent testa glabrous the WD40 and WRKY transcription factors are named for conserved amino acid sequences within these proteins. PC = personal communication.

Fig. 6.2 The flavonoid core biosynthetic pathway. The P450s and 2-oxoglutarate-dependent dioxygenases (2-ODDs) are indicated by underlined and bolded titles, respectively. F3 H and F3 5 H are capable of using flavanones (2-3 = single bond, R1 = H), flavones (2-3 = double bond, R1 = H), dihydroflavonols (2-3 = single bond, R1 = OH), or flavonols (2-3 = double bond, R1 = OH) for a substrate. FNS activity has been indicated with two different enzymes, FNS-11 (P450) and the less-common FNS-I (2-ODD)...

Flavonol synthase (FLS E.C.l.14.11.23) catalyzes the committed step in the production of fiavonols by introduction of a double bond between C2 and C3 of the corresponding dihydroflavonols. Like E3H, ELS has been described as a 2-oxoglutatarate-dependent dioxygenase based on its cofactor requirements for 2-oxoglutarate, Fe, and ascorbate. FLS was initially identified in enzyme preparations from illuminated parsley cell suspension cultures [67]. Subsequently, FLS was characterized from the flower buds of Matthiola incana and carnation (Dianthus caryophyllus L.), and it was suggested that there was regulation between flavonol and anthocyanidin biosynthesis [83, 84]. 

Flavanone 3 -hydroxylase (F3 H ECl.14.13.21 CYP75B) activity was initially identified in microsomal preparations of golden weed (Haplopappus gracilis) [110]. E3 H from irradiated parsley cell cultures was later biochemically analyzed and characterized as a cytochrome P450 having an absolute requirement for NADPH and molecular oxygen as cofactors [111]. The enzyme has been shown to have activity with flavanones, flavones, dihydroflavonols, and flavonols, but does not appear to have activity with anthocyanidins [111]. The first cDNA clone for E3 H was isolated from Petunia [112]. It has been suggested that E3 H may serve as an anchor for the proposed flavonoid multi-enzyme complex on the cytosolic surface of the endoplasmic reticulum [44]. 

SpribiUe R, Forkmann G (1984) Conversion of dihydroflavonols to flavonols with enzyme extracts from flower buds of Matthiola incana. Z Naturforsch 39C 714-719... 

Davies, K.M. et al.. Enhancing anthocyanin production by altering competition for substrate between flavonol synthase and dihydroflavonol 4-reductase. Euphytica, 131, 259, 2003. 

Since the early contributions of Willstatter and Robinson, several alternative approaches following mainly two routes have been considered for synthesis of anthocyanins.One of the routes includes condensation reactions of 2-hydroxybenzaldehydes with acetophenones, while the other uses transformations of anthocyanidin-related compounds like flavonols, flavanones, and dihydroflavonols to yield flavylium salts. The urge for plausible sequences of biosynthetic significance has sometimes motivated this latter approach. In the period of this review, new synthetically approaches in the field have also predominantly been following the same general routes however, some new features have been shown in synthesis of pyranoanthocyanidins. 

Data on this type of flavonols are summarized in Table 12.4. In contrast to the corresponding flavones, the number and complexity of derivatives is smaller. This concerns particularly the formation of furano-, pyrano- and other cyclic flavonols. There is a remarkable number of 0-prenylated flavonols known to date, contrasting to only very few flavones exhibiting this substitution pattern (see Table 12.3). Similar trends have been earlier documented in the review of Barron and Ibrahim. The occurrence of a series of glycosides based on C-prenylated structures is considerable. This substitution trend concerns also some of the dihydroflavonols, thus indicating specific enzyme activities probably dependent on the presence of a 3-OH group. 

Table 5.38 b. Structures and 13C Chemical Shifts (<5C in ppm) of Selected Chalcones, Flavones, Flavonols, Isoflavones, Dihydro-flavones, Dihydroflavonols and Flavons (Aglycones). Spectra were Recorded in DMSO-d6, Except those of Flavone (CDC13) and 2 -Hydroxy-, and 2,2 -Dihydroxychalcone (D20/DMS0-d6 (2 7)) [989, 990]. 

Figure 1.36 Schematic diagram of the stilbene and flavonoid biosynthetic pathway. Enzyme abbreviations SS, stilbene synthase CHS, chalcone synthase CHR, chalcone reductase CHI, chalcone isomerase IFS, isoflavone synthase FNS, flavone synthase F3H, flavanone 3-hydroxylase FLS, flavonol synthase F3 H, flavonoid 3 -hydroxylase DFR, dihydroflavonol 4-reductase LAR, leucoanthocyanidin 4-reductase LDOX, leucocyanidin deoxygenase ANR, anthocyanidin reductase EU, extension units TU, terminal unit.

Flavonoids constitute a large class of polyphenols found in fruits and vegetables that share a common skeleton of phenylchromane. This basic structure allows a large number of substitution patterns leading to several subclasses of flavonoids, such as flavonols, flavones, flavanones, flavanols, anthocyanidins, isoflavones, dihydroflavonols, and chalcones. Among the diverse flavonoid subclasses, flavonols (especially quercetin) and flavanols (catechins) are the most abundant in our food. Flavonols are present in foods as diverse glycosides, whereas flavanols are usually found as aglycones. 

In plants accumulating anthocyanins, flavonols, and proanthocyanidins, naringenin is stereospecifically hydroxylated at position 3 of the C-ring (C3) by the 2-oxoglutarate-dependent dioxygenase flavanone 3-hydroxylase (F3H, EC 1.14.11.9) to yield the 3-hydroxy-trans-flavanone (syn. dihydroflavonol) dihy-drokaempferol [Springob et al., 2003] (Fig.21 2). Dihydroquercetin (3, 4, 5,5, 7-... 

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