2 J.-H.B.Hatier,K.S.Gould anthocyanins may colour the entire blade,or else be restricted to the margins,stripes patches,or seer om spots on the upper,lower or both lamina surfaces.Ir some lea interveinal lamina tissue,or the stipules,or domatia that are anthocyanic 0m6 leaves turn red shortly before they abscise,others are red only while they are growing,yet others remain red throughout their lives.In many species,anthocyanin are produced only when the plant is unhealthy or has been exposed to environmenta stress,but there are some that develop the red pigments even under optimal growth environments.Given this enormous variation in location,timing,and inducibility of anthocyanins in vegetative tissues,it is not surprising that a unified explanation for the presence of these pigments has thus far eluded scientific investigation.Muriel Wheldale's statement that a unified explanation would require “reduction to absurdity"remains as valid today as it was when she wrote it in 1916. Although the selective pressure that has driven the evolution of anthocyanins in such disparate vegetative structures remains far from obvious,plant physiologists have nevertheless made significant progress over the past decade in elaborating the consequences of cellular anthocyanins on plant function.Reflecting the resurgence of scientific interest in anthocyanin (and betalain)function in vegetative organs several reviews have been written on this topic in recent years(Chalker-Scott 1999. 2002:Hoch et al.2001;Gould et al.2002b:Lee and Gould 2002a,2002b:Steyn et al.2002:Close and Beadle 2003;Gould 2004;Gould and Lister 2005:Stintzing and Carle 2005;Manetas 2006).Those reviews provide a comprehensive summar rary knowledge.particularly in relation to leaf physiolo on which most research has been done It is not our intention to duplicate that information in this chapter,although for completeness we do briefly summarise the three leading hypoth s for anthocyanin function in leaves.Rather.with the use of selected to demonstrate versatility in anthocva en ways and to different degrees.even though the chemical nature and histologica of the e identical. y,in acknowledg the para ole of re ecies (ROS)n and Noctor 2005) we develop arg ovel anth ne of a mod ator of signal trans ion physio cascades responses to stress 1.2 Anthocyanins and Stress Responses Foliar ant L anin commonlycur vacuor oon in epidema mos all (Pos aL2000- lins 2001; Post and 1992:Kunz et 1994 Hooijmaijers Kunz and Beck 1995 and G ould 2007).Irrespective of their cellular location,however anthocyanin biosynthesis in many leaves is generally upregulated in response to one or more environmental stressors.These include:strong light,UV-B radiation. temperature extremes,drought,ozone,nitrogen and phosphorus deficiencies
2 J.-H. B. Hatier, K.S. Gould anthocyanins may colour the entire blade, or else be restricted to the margins, stripes, patches, or seemingly random spots on the upper, lower or both lamina surfaces. In some leaves, only the petiole and major veins are pigmented red, in others it is the interveinal lamina tissue, or the stipules, or domatia that are anthocyanic. Some leaves turn red shortly before they abscise, others are red only while they are growing, yet others remain red throughout their lives. In many species, anthocyanins are produced only when the plant is unhealthy or has been exposed to environmental stress, but there are some that develop the red pigments even under optimal growth environments. Given this enormous variation in location, timing, and inducibility of anthocyanins in vegetative tissues, it is not surprising that a unified explanation for the presence of these pigments has thus far eluded scientific investigation. Muriel Wheldale’s statement that a unified explanation would require “reduction to absurdity” remains as valid today as it was when she wrote it in 1916. Although the selective pressure that has driven the evolution of anthocyanins in such disparate vegetative structures remains far from obvious, plant physiologists have nevertheless made significant progress over the past decade in elaborating the consequences of cellular anthocyanins on plant function. Reflecting the resurgence of scientific interest in anthocyanin (and betalain) function in vegetative organs, several reviews have been written on this topic in recent years (Chalker-Scott 1999, 2002; Hoch et al. 2001; Gould et al. 2002b; Lee and Gould 2002a, 2002b; Steyn et al. 2002; Close and Beadle 2003; Gould 2004; Gould and Lister 2005; Stintzing and Carle 2005; Manetas 2006). Those reviews provide a comprehensive summary of contemporary knowledge, particularly in relation to leaf physiology, on which most research has been done. It is not our intention to duplicate that information in this chapter, although for completeness we do briefly summarise the three leading hypotheses for anthocyanin function in leaves. Rather, with the use of selected examples, we hope to demonstrate the extraordinary versatility in anthocyanin function. Thus, any two species might benefit from anthocyanins in very different ways and to different degrees, even though the chemical nature and histological location of the pigment are identical. Finally, in acknowledgement of the recent paradigm shift in relation to the role of reactive oxygen species (ROS) in plants (see Foyer and Noctor 2005), we develop an argument for a novel function of anthocyanins in leaves – that of a modulator of signal transduction cascades in physiological responses to stress. 1.2 Anthocyanins and Stress Responses Foliar anthocyanins most commonly occur as vacuolar solutions in epidermal and/or mesophyll cells, although in certain bryophytes these red pigments bind to the epidermal cell wall (Post 1990; Gould and Quinn 1999; Gould et al. 2000; Lee and Collins 2001; Post and Vesk 1992; Kunz et al. 1994; Kunz and Becker 1995; Hooijmaijers and Gould 2007). Irrespective of their cellular location, however, anthocyanin biosynthesis in many leaves is generally upregulated in response to one or more environmental stressors. These include: strong light, UV-B radiation, temperature extremes, drought, ozone, nitrogen and phosphorus deficiencies
Anthocyanin Function in Vegetative Organs 3 bacterial and fungal infections,wounding.herbivory,herbicides,and various pollutants(McClure 1975;Chalker-Scott 1999).Because of their association with such biotic and abiotic stressors,anthocyanins are usually considered to be a stress symptom and/or part of a mechanism to mitigate the effects of stress.Much of the physiological work undertaken in recent years has attempted to unravel the phytoprotective functions of anthocyanins that would enhance tolerance to these stress factors 1.3 Photoprotection photoprotective roles of foliar anthocvanin have probably received more attention ir It had long been su sted that ins might shield photosynthetic cells from adverse effects of strong ligh (see Wheldale 1916).vet the first experimental confirmation of this was not a until the 1990s following the advent of the field. portable pulse a (PAM)chlorophyll fluor neter which r red ve es (Gould et al 1995 nthesis is d ta in requir ents of the light r ions car sely affect compo (antenna action centres acces ory protein and electro ep me brane 003) he te en le to excess chlorophy a. orome al 198 1991) 0 6.1 hi for dar ratio of maxim um yiel (M s0n2000 s are typically arour 0.83 for pre-dawn,healthy they can under stres asurem values for red and green leaves before and after exposing them to photoinhibitory light fluxes provide a convenient method to compare their relative tolerances to light stress Anthocyanic leaves typically absorb more light in the green and yellow wavebands than do acyanic leaves (Neill and Gould 1999;Gitelson et al.2001).The fate of these absorbed quanta is unknown,but it is very clear that their energy is not transferred to the chloroplasts.Indeed,the chlorenchyma of red leaves may receive considerably less green light than do those of structurally comparable green leaves (Gould et al 2002c),and red leaves may develop the morphological and physiological attributes of shade leaves(Manetas et al.2003).This light-filtering effect of anthocvanins has been shown many times both to reduce the severity of photoinhibition and to expedite photosynthetic recovery in red as compared to green leaves (see reviews by Steyn et al.2002;Gould and Lister 2005).In point of fact. sufficient experimental evidence of a photoprotective function of anthocyanins has accrued to justify its elevation from hypothesis to theory
Anthocyanin Function in Vegetative Organs 3 bacterial and fungal infections, wounding, herbivory, herbicides, and various pollutants (McClure 1975; Chalker-Scott 1999). Because of their association with such biotic and abiotic stressors, anthocyanins are usually considered to be a stress symptom and/or part of a mechanism to mitigate the effects of stress. Much of the physiological work undertaken in recent years has attempted to unravel the phytoprotective functions of anthocyanins that would enhance tolerance to these stress factors. 1.3 Photoprotection Photoprotective roles of foliar anthocyanin have probably received more attention in recent years than any other functional hypothesis. It had long been suggested that anthocyanins might shield photosynthetic cells from adverse effects of strong light (see Wheldale 1916), yet the first experimental confirmation of this was not achieved until the 1990s, following the advent of the field-portable pulse amplitude modulated (PAM) chlorophyll fluorometer which permitted non-invasive comparisons of the quantum efficiencies of photosynthesis in red versus green leaves (Gould et al. 1995; Krol et al. 1995). Although photosynthesis is driven by light, quanta in excess of the requirements of the light reactions can adversely affect the photosynthetic system components (antenna pigments, reaction centres, accessory proteins, and electron transport carriers), and can lead to secondary destructive and repair processes in thylakoid membranes (Adir et al. 2003). Photoinhibition, the term given to the decline in quantum yield of photosynthesis attributable to excessive illumination, can be quantified directly using PAM chlorophyll fluorometers (Genty et al. 1989; Krause and Weis 1991). One of the most useful parameters for this is the ratio of variable to maximum chlorophyll fluorescence (Fv/Fm) for dark-adapted leaves, which correlates to the maximum quantum yield of photosystem II (Maxwell and Johnson 2000). Fv/Fm values are typically around 0.83 for pre-dawn, healthy plants, but they can be considerably lower in plants under stress. Measurements of Fv/Fm values for red and green leaves before and after exposing them to photoinhibitory light fluxes provide a convenient method to compare their relative tolerances to light stress. Anthocyanic leaves typically absorb more light in the green and yellow wavebands than do acyanic leaves (Neill and Gould 1999; Gitelson et al. 2001). The fate of these absorbed quanta is unknown, but it is very clear that their energy is not transferred to the chloroplasts. Indeed, the chlorenchyma of red leaves may receive considerably less green light than do those of structurally comparable green leaves (Gould et al. 2002c), and red leaves may develop the morphological and physiological attributes of shade leaves (Manetas et al. 2003). This light-filtering effect of anthocyanins has been shown many times both to reduce the severity of photoinhibition and to expedite photosynthetic recovery in red as compared to green leaves (see reviews by Steyn et al. 2002; Gould and Lister 2005). In point of fact, sufficient experimental evidence of a photoprotective function of anthocyanins has accrued to justify its elevation from hypothesis to theory
4 J.-H.B.Hatier,K.S.Gould yanins confer me foliage of de ous trees (Feild et overwintering fol evergreen plants(Hughes et al.200 05 leaves can also benefit significantly from these pigments(Cai et al.2005). Indee nascent chloroplasts in immature leaves are particularly vul erable to the effects of light stress(Pettigrew and Vaughn 1998.Choinski et al.2003).Strong support for a photoprotective role of anthocyanins in developing leaves was provided recently by Hughes et al.(2007).who followed the timing of anthocyanin production and degradation across three unrelated species: Acer rubrum,Cercis canadensis,and Liquidambar styraciflua.In all three species,anthocyanins were produced early in leaf development,and persisted until leaf tissues had fully differentiated The subsequent decline in anthocyanin levels occurred only after leaves had synthesised approximately 50%of the total chlorophylls and carotenoids,and had attained close to their maximum photosynthetic assimilation rates.The authors suggested that the strong coupling between the timing of anthocyanin reassimilation and those of leaf developmental processes indicates that anthocvanins serve to protect tissues until other photoprotective mechanisms mature In view of the immutable property of the coloured anthocyanins to absorb light that might otherwise strike chlor oplasts,it is perhaps surprising that the degree to which anthocyanins contribute to the photoprotection of leaves seems to vary substantially from species to species.In Galax urceolata,for example,Fv/Fm values for green leaves decreased 36%more than did those for red leaves following osure to photoinhibitory conditions(Hughes and Smith 2007).Differences of a 9renephoenhibonoeantoenHeacsamd,Sahn2omlPierotcoi stolonifera (Feild et al 2001)and between green flushing leaves of Litsed dilleniifolia and the red flushing lea es of litsea pierrei and Anthocephalus chinensis (Cai et al.2005).Hov er,much larger differe s (ca 75%)hav eported for the green adult and red ju ile s of Pas and in Pici mis the decline in Fy/fm for green leaves was almost double that of red leav (Manetas et al.2002).In c ph ng red lea of gre thos light flux and Ma tas ed-lea species p20 ed worse than green- leafe specie thes inters ecific diffe ence in pho top tec phys else are th hic ere tal s(200 sugge capacity of an as a functio The arguedt gree to pno only in the low of th a (Sun et a eaves hich are relatively thick and whos sophyll contain large of chlorophyl would benefit most from the abatement of green light D anthocyanins. of thinner leaves that contain an chlorophyll would be driven almost exclusively by red and blue light,and therefore their propensity for photoinhibition would not be affected greatly by the presence of
4 J.-H. B. Hatier, K.S. Gould Anthocyanins confer measurable photoprotection when present both in senescing foliage of deciduous trees (Feild et al. 2001; Hoch et al. 2003) and in the mature, overwintering foliage of evergreen plants (Hughes et al. 2005). Young, developing leaves can also benefit significantly from these pigments (Cai et al. 2005). Indeed, nascent chloroplasts in immature leaves are particularly vulnerable to the effects of light stress (Pettigrew and Vaughn 1998; Choinski et al. 2003). Strong support for a photoprotective role of anthocyanins in developing leaves was provided recently by Hughes et al. (2007), who followed the timing of anthocyanin production and degradation across three unrelated species: Acer rubrum, Cercis canadensis, and Liquidambar styraciflua. In all three species, anthocyanins were produced early in leaf development, and persisted until leaf tissues had fully differentiated. The subsequent decline in anthocyanin levels occurred only after leaves had synthesised approximately 50% of the total chlorophylls and carotenoids, and had attained close to their maximum photosynthetic assimilation rates. The authors suggested that the strong coupling between the timing of anthocyanin reassimilation and those of leaf developmental processes indicates that anthocyanins serve to protect tissues until other photoprotective mechanisms mature. In view of the immutable property of the coloured anthocyanins to absorb light that might otherwise strike chloroplasts, it is perhaps surprising that the degree to which anthocyanins contribute to the photoprotection of leaves seems to vary substantially from species to species. In Galax urceolata, for example, Fv/Fm values for green leaves decreased 36% more than did those for red leaves following exposure to photoinhibitory conditions (Hughes and Smith 2007). Differences of a similar magnitude were noted between yellow and red senescent leaves of Cornus stolonifera (Feild et al. 2001), and between green, flushing leaves of Litsea dilleniifolia and the red flushing leaves of Litsea pierrei and Anthocephalus chinensis (Cai et al. 2005). However, much larger differences (ca. 75%) have been reported for the green adult and red juvenile leaves of Rosa sp., and in Ricinus communis the decline in Fv/Fm for green leaves was almost double that of red leaves (Manetas et al. 2002). In contrast, photosynthetic efficiencies of young red leaves of Quercus coccifera were only marginally greater than those of young green leaves under photoinhibitory light flux (Karageorgou and Manetas 2006), and red-leafed species of Prunus actually performed worse than green-leafed species under saturating light (Kyparissis et al. 2007). The reasons for these large interspecific differences in photoprotection by anthocyanin are unknown. It is uncertain whether they reflect true physiological differences, or else are the result of disparities in the experimental conditions under which measurements were taken. Karageorgou and Manetas (2006) suggested that the photoprotective capacity of foliar anthocyanins might vary simply as a function of leaf thickness. They argued that because green light contributes to photosynthesis only in the lowermost tissues of the leaf lamina (Sun et al. 1998; Nishio 2000), then those leaves which are relatively thick and whose mesophyll contain large amounts of chlorophyll would benefit most from the abatement of green light by anthocyanins. Photosynthesis of thinner leaves that contain low amounts of chlorophyll would be driven almost exclusively by red and blue light, and therefore their propensity for photoinhibition would not be affected greatly by the presence of
Anthocvanin Function in vegetative Organs anthocyanins.Accordingly,the immature red leaves of Ouercus coccifera,which are less than 200 um thick and hold 11 ug mchlorophyll,show little evidence of photoprotection by anthocvanin (karage ou and Manetas 2006).In contrast,the mature leaves of that spe which are twice as thick and hold four times as much chlorophyll,show a sizeable benefit from anthocyanins (Manetas et al.2003).The "leaf-thickness hypothesis"warrants further testing although the recorded benefits of anthocvanins to the photosynthesis of thin.immature leaves in certain other At least some of the variation am rts of photon folia anthocvanins is likely to be attributable e to difepo nces in the e Photoinhibition xperimental protocol nsified when, in additio oton f nla ypes such as and free ng and defi empera nitroge ienc ha been trong ligh po suc Evide r this was presente green genotypes or ma n(0(02) app rent on a com on o Light quality is also important,reductions in Fv/Fm have been found to be greate for green than red leaves when irradiated with white or green light,yet they are similar in magnitude under red light (Hughes et al.2005). Thus,the experimenta conditions under which leaves are tested for photoinhibition can have a significant bearing on measurements of chlorophyll fluorescence. The interpretation of chlorophyll fluorescence signals can itself be problematic for red leaves. In a recent report describing the common pitfalls of chlorophyll fluorescence analvsis.Logan et al.(2007)explained that anthocvanins mav absorb a proportion of the measuring light issued from the PAM fluorometer,and therefore reduce the intensity of the emitted chlorophyll fluorescence that is collected for detection.This can lead to low signal to noise ratios,and therefore compromise the accuracy of the data.Fluorescence output can be improved by increasing the intensity of the measuring light,yet this runs the risk of the measuring beam becoming actinic (ie.driving photosynthesis).which would artifactually reduc Fv/Fm values.Some machines perform better than others for measuring chlorophyll fluorescence in red leaves;Pfundel et al.(2007)showed that because anthocyanins attenuate about half of the incident radiation at 470 nm a fluorometer that issues pulses of blue measuring light can be inferior to one that emits red pulses.It is alsc noteworthy that the chlo phvll fluor alter as leaf age and car even vary from re gion to reg n across a leaf lamina(Sestak and Siffel 1997).Thu the con mparison of young (red)and old(g en)leaves or else red and green parts of the sa ces in chlor escence that are unrelated.or only partially related to the presence of antho There are in addition to anthocyanins which plants ate light other mechanisms by gical featu uch as ha ticle hat and n from the
Anthocyanin Function in Vegetative Organs 5 At least some of the variation among reports of photoprotection by foliar anthocyanins is likely to be attributable to differences in the experimental protocol. Photoinhibition is often intensified when, in addition to excess photon flux, plants experience other types of abiotic stressor (Long et al. 1994). By limiting the rates of CO2 fixation, environmental factors such as chilling and freezing temperatures, high temperatures, and nitrogen deficiency have been shown to exacerbate the photoinhibitory responses to strong light. It seems possible, therefore, that the photoprotective capacity of anthocyanins would assume greater importance in plants that face combinations of such stressors. Evidence for this was presented recently in a comparison of green- and red-leafed genotypes of maize; the beneficial effects of anthocyanins were apparent only after the plants had experienced a combination of strong light (2000 µmol m−2 s−1 ) and a 5ºC chilling treatment (Pietrini et al. 2002). Light quality is also important; reductions in Fv/Fm have been found to be greater for green than red leaves when irradiated with white or green light, yet they are similar in magnitude under red light (Hughes et al. 2005). Thus, the experimental conditions under which leaves are tested for photoinhibition can have a significant bearing on measurements of chlorophyll fluorescence. The interpretation of chlorophyll fluorescence signals can itself be problematic for red leaves. In a recent report describing the common pitfalls of chlorophyll fluorescence analysis, Logan et al. (2007) explained that anthocyanins may absorb a proportion of the measuring light issued from the PAM fluorometer, and therefore reduce the intensity of the emitted chlorophyll fluorescence that is collected for detection. This can lead to low signal to noise ratios, and therefore compromise the accuracy of the data. Fluorescence output can be improved by increasing the intensity of the measuring light, yet this runs the risk of the measuring beam becoming actinic (i.e. driving photosynthesis), which would artifactually reduce Fv/Fm values. Some machines perform better than others for measuring chlorophyll fluorescence in red leaves; Pfündel et al. (2007) showed that because anthocyanins attenuate about half of the incident radiation at 470 nm, a fluorometer that issues pulses of blue measuring light can be inferior to one that emits red pulses. It is also noteworthy that the chlorophyll fluorescence signals can alter as a leaf ages, and can even vary from region to region across a leaf lamina (Šesták and Šiffel 1997). Thus, the comparison of young (red) and old (green) leaves, or else red and green parts of the same leaf blade, may yield differences in chlorophyll fluorescence that are unrelated, or only partially related to the presence of anthocyanins. There are in addition to anthocyanins many other mechanisms by which plants can avoid or dissipate excess light energy. These include morphological features, such as hairs or a waxy cuticle that reflect and scatter incident radiation from the anthocyanins. Accordingly, the immature red leaves of Quercus coccifera, which are less than 200 µm thick and hold 11 µg m−2 chlorophyll, show little evidence of photoprotection by anthocyanin (Karageorgou and Manetas 2006). In contrast, the mature leaves of that species, which are twice as thick and hold four times as much chlorophyll, show a sizeable benefit from anthocyanins (Manetas et al. 2003). The “leaf-thickness hypothesis” warrants further testing, although the recorded benefits of anthocyanins to the photosynthesis of thin, immature leaves in certain other species (Hughes et al. 2007) would suggest that the hypothesis is not universally applicable
6 J.-H.B.Hatier,K.S.Gould lamina Surface and physiological proce an sses such as thermal dissipation by the triplet phyll Ive,and The degree to which each 0 these mechanisms is utilised apparently varies from species to species,as well as with the ity and duration of exposure to abiotic stress(Demmig-Adams and Adams III 2006).Accordingly,the requirement for supplementary photoprotection such as that provided by anthocyanins,would also vary.Consistent with this,the young leaves of Rosa sp.and Ricinus commmnis contain only low levels of xanthophyll pigments,yet they are resistant to photoinhibitory damage possibly because of their high anthocyanin concentrations (Manetas et al.2002). the combined effects of pubescence and anthocyanins in certain cultivars of grapevine (Vitis vinifera)apparently compensate for their reduced xanthophyll contents relative to levels in green,glabrous cultivars(Liakopoulos et al.2006).In their analysis of mutants of Arabidopsis thaliana,Havaux and Kloppstech (2001) concluded that the flavonoids might actually be more important than the xanthophylls in regard to long-term protection from photoinhibitory damage although the anthocyanins were less effective as photoprotectants in that system than were the flavonols and dihydroflavonols.Interspecific differences in requirements 6C62rlmcsaopoeeiePeSibhpiSmnotvTooiteepcv 1.4 Protection Against Ultraviolet Radiation In addition to their capacity to protect plant tissues from excess visible radiation anthocvanins have also been implicated in the protection from ultraviolet (Uv radiation.UV radiation is often classified as UV-A (320-390 nm)UV-B (280-320 nm)and IV.C(s280 nm)Strat ospheric ozone(O:)absorbs most of the UV-C and part of the uv-B UV-A radiation ever is not filtered by stratospheric o:with an absorption maxin um (A. at 260 nm.DNA is particularly vulnerable to the effects of highl tie uv r nd Ror 11999 To fortify the hepedmuiro ves agai nst the harmful effects of UV radiation,plants have devel s to din penetration ng the synthesis of UV-abs and Hunt 2005).The biosyr ing phenolic ted in re (Taka hi et al 1991:Mende al 1999:Singh et al.1999),altho ha 6 ed(Jordan et al.1994 1995:Solove and M 2003 anl UV 982:G ar et al their vegetative orga buffer 1991 Lietal1993 da excess er Koost 99 ng th pro anthocyanin ide ards 96 noted tha e photos capac es of green-le varieties of Coleus were lower varieties
6 J.-H. B. Hatier, K.S. Gould lamina surface, and physiological processes such as thermal dissipation by the xanthophyll cycle pigments and the triplet chlorophyll valve, and the transfer of excess electrons to alternative sinks (Niyogi 2000). The degree to which each of these mechanisms is utilised apparently varies from species to species, as well as with the intensity and duration of exposure to abiotic stress (Demmig-Adams and Adams III 2006). Accordingly, the requirement for supplementary photoprotection, such as that provided by anthocyanins, would also vary. Consistent with this, the young leaves of Rosa sp. and Ricinus communis contain only low levels of xanthophyll pigments, yet they are resistant to photoinhibitory damage possibly because of their high anthocyanin concentrations (Manetas et al. 2002). Similarly, the combined effects of pubescence and anthocyanins in certain cultivars of grapevine (Vitis vinifera) apparently compensate for their reduced xanthophyll contents relative to levels in green, glabrous cultivars (Liakopoulos et al. 2006). In their analysis of mutants of Arabidopsis thaliana, Havaux and Kloppstech (2001) concluded that the flavonoids might actually be more important than the xanthophylls in regard to long-term protection from photoinhibitory damage, although the anthocyanins were less effective as photoprotectants in that system than were the flavonols and dihydroflavonols. Interspecific differences in requirements for supplementary photoprotection probably best explain why reports of the capacity of foliar anthocyanins to protect leaves from photoinhibiton vary so greatly. 1.4 Protection Against Ultraviolet Radiation In addition to their capacity to protect plant tissues from excess visible radiation, anthocyanins have also been implicated in the protection from ultraviolet (UV) radiation. UV radiation is often classified as UV-A (320–390 nm), UV-B (280–320 nm) and UV-C (<280 nm). Stratospheric ozone (O3) absorbs most of the UV-C and part of the UV-B. UV-A radiation, however, is not filtered by stratospheric O3. With an absorption maximum (Aλmax) at 260 nm, DNA is particularly vulnerable to the adverse effects of highly energetic UV rays (Hoque and Remus 1999). To fortify themselves against the harmful effects of UV radiation, plants have developed multifarious mechanisms to diminish UV penetration into plant tissues, including the synthesis of UV-absorbing phenolic compounds (Ryan and Hunt 2005). The biosynthesis of anthocyanins and other flavonoids is known to be activated in many plant species by UV exposure (Takahashi et al. 1991; Mendez et al. 1999; Singh et al. 1999), although exceptions have been noted (Jordan et al. 1994; Buchholz et al. 1995; Solovchenko and Merzlyak 2003). Most anthocyanins, especially those that are acylated, can absorb biologically-active UV radiation (Markham 1982; Giusti et al. 1999), and it has been suggested that their function in vegetative organs may be to buffer tissues against UV damage by attenuating the excess energy (Takahashi et al. 1991; Li et al. 1993; Koostra 1994). Support for a protective role of anthocyanins was provided by Burger and Edwards (1996), who noted that following exposure to UV-B or UV-C radiation, the photosynthetic capacities of green-leafed varieties of Coleus were lower than those of red-leafed varieties