The role of UV-B radiation in aquatic and terrestrial ecosystems--an experimental and functional analysis of the evolution of UV-absorbing compounds

We analysed and compared the functioning of UV-B screening pigments in plants from marine, fresh water and terrestrial ecosystems, along the evolutionary line of cyanobacteria, unicellular algae, primitive multicellular algae, charophycean algae, lichens, mosses and higher plants, including amphibious macrophytes. Lichens were also included in the study. We were interested in the following key aspects: (a) does the water column function effectively as an 'external UV-B filter'?; (b) do aquatic plants need less 'internal UV-B screening' than terrestrial plants?; (c) what role does UV screening play in protecting the various plant groups from UV-B damage, such as the formation of thymine dimers?; and (d) since early land 'plants' (such as the predecessors of present-day cyanobacteria, lichens and mosses) experienced higher UV-B fluxes than higher plants, which evolved later, are primitive aquatic and land organisms (cyanobacteria, algae, lichens, mosses) better adapted to present-day levels of UV-B than higher plants? Furthermore, polychromatic action spectra for the induction of UV screening pigments of aquatic organisms have been determined. This is relevant for translating 'physical' radiation measurements of solar UV-B into 'biological' and 'ecological' effects. From the action spectra, radiation amplification factors (RAFs) have been calculated. These action spectra allow us to determine any mitigating or antagonistic effects in the ecosystems and therefore qualify the damage prediction for the ecosystems under study. We summarize and discuss the main results based on three years of research of four European research groups. The central theme of the work was the investigation of the effectiveness of the various screening compounds from the different species studied in order to gain some perspective of the evolutionary adaptations from lower to higher plant forms. The induction of mycosporine-like amino acids (MAAs) was studied in the marine dinoflagellate Gyrodinium dorsum, the green algal species Prasiola stipitata and in the cyanobacterium Anabaena sp. While visible (400-700 nm) and long wavelength UV-A (315-400 nm) showed only a slight effect, MAAs were effectively induced by UV-B (280-315 nm). The growth of the lower land organisms studied, i.e. the lichens Cladina portentosa, Cladina foliacaea and Cladonia arbuscula, and the club moss Lycopodiumannotinum, was not significantly reduced when grown under elevated UV-B radiation (simulating 15% ozone depletion). The growth in length of the moss Tortula ruralis was reduced under elevated UV-B. Of the aquatic plants investigated the charophytes Chara aspera showed decreased longitudinal growth under elevated UV-B. In the 'aquatic higher plants' studied, Ceratophyllum demersum, Batrachium trichophyllum and Potamogeton alpinus, there was no such depressed growth with enhanced UV-B. In Chara aspera, neither MAAs nor flavonoids could be detected. Of the terrestrial higher plants studied, Fagopyrum esculentum, Deschampsia antarctica, Vicia faba, Calamagrostis epigejos and Carex arenaria, the growth of the first species was depressed with enhanced UV-B, in the second species length growth was decreased, but the shoot number was increased, and in the latter two species of a dune grassland there was no reduced growth with enhanced UV-B. In the dune grassland species studied outdoors, at least five different flavonoids appeared in shoot tissue. Some of the flavonoids in the monocot species, which were identified and quantified with HPLC, included orientin, luteolin, tricin and apigenin. A greenhouse study with Vicia faba showed that two flavonoids (aglycones) respond particularly to enhanced UV-B. Of these, quercetin is UV-B inducible and mainly located in epidermal cells, while kaempferol occurs constitutively. In addition to its UV-screening function, quercetin may also act as an antioxidant. Polychromatic action spectra were determined for induction of the UV-absorbing pigments in three photosynthetic organisms, representing very different taxonomic groups and different habitats. In ultraviolet photobiology, action spectra mainly serve two purposes: (1) identification of the molecular species involved in light absorption; and (2) calculation of radiation amplification factors for assessing the effect of ozone depletion. Radiation amplification factors (RAFs) were calculated from the action spectra. In a somewhat simplified way, RAF can be defined as the percent increase of radiation damage for a 1% depletion of the ozone layer. Central European summer conditions were used in the calculations, but it has been shown that RAF values are not critically dependent on latitude or season. If only the ultraviolet spectral region is considered, the RAF values obtained are 0.7 for the green alga Prasiola stipitata, 0.4 for the dinoflagellate Gyrodinium dorsum, and 1.0 for the cyanobacterium Anabaena sp. In the case of P. stipitata, however, the effect of visible light (PAR, photosynthetically active radiation, 400-700 nm) is sufficient to lower the RAF to about 0.4, while the PAR effect for G. dorsum is negligible. RAFs for some damage processes, such as for DNA damage (RAF=2.1 if protective effects or photorepair are not considered [1]), are higher than those above. Our interpretation of this is that if the ozone layer is depleted, increased damaging radiation could overrule increased synthesis of protective pigments. In addition to investigating the functional effectiveness of the different screening compounds, direct UV effects on a number of key processes were also studied in order to gain further insight into the ability of the organisms to withstand enhanced UV-B radiation. To this end, the temperature-dependent repair of cyclobutane dimers (CPD) and (6-4) photoproducts induced by enhanced UV-B was studied in Nicotiana tabacum, and the UV-B induction of CPD was studied in the lichen Cladonia arbuscula. Also, photosynthesis and motility were monitored and the response related to the potential function of the screening compounds of the specific organism.     

Effects of elevated ultraviolet radiation and endophytic fungi on plant growth and insect feeding in Lolium perenne, Festuca rubra, F. arundinacea and F. pratensis.

Plants of perennial ryegrass (Lolium perenne L.), red fescue (Festuca rubra L.), tall fescue (F. arundinacea Schreb.) and meadow fescue (F. pratensis Huds) were exposed at an outdoor facility located in Edinburgh, UK to modulated levels of UV-B radiation (280-315 nm) using banks of cellulose diacetate filtered UV-B fluorescent lamps that also produce UV-A radiation (315-400 nm). The plants were derived from a single clone of each species and were grown both with and without colonization by naturally-occurring fungal endophytes. The UV-B treatment was a 30% elevation above the ambient erythemally-weighted level of UV-B during July to October. Growth of treated plants was compared with plants grown under elevated UV-A radiation alone produced by banks of polyester filtered lamps and with plants grown at ambient levels of solar radiation under banks of unenergized lamps. At the end of the treatment period, sample leaves were collected for feeding trials with the desert locust Schistocerca gregaria (Forsk). The UV-B treatment produced no effects on the aboveground biomass of any of the four grasses. The UV-B treatment and the UV-A control exposure both increased plant height and the number of daughter plants formed by rhizome growth in F. rubra. There were significant effects of endophyte presence on the total fresh and dry weights of F. arundinacea and F. rubra, on fresh weight only in F. pratensis, and on the fresh and dry weights of inflorescence in F. arundinacea and L. perenne. There were no effects of UV treatments on the absolute amounts of leaf consumed or on the feeding preferences of locusts for leaves with or without endophyte in three species: F. rubra, F. arundinacea and L. perenne. In F. pratensis there was no effect of UV treatment on the weight of leaves consumed but a significant UV x endophyte interaction caused by a marked change in feeding preference between leaves with and without endophyte that differed between the UV-B treatment and UV-A control exposures. The alkaloid compounds known as lolines were analysed in leaves of F. pratensis and were only found in plants grown with endophyte. However, there was no significant relationship between total loline content and insect feeding preference. These effects illustrate the potential complexities of species interactions under increasing levels of UV-B. The experiment also demonstrates the importance of appropriate controls in UV lamp supplementation experiments for interpretation of both plant growth and insect feeding effects.     

Non-destructive analysis of pigments and other organic compounds in lichens using Fourier-transform Raman spectroscopy: a study of Antarctic epilithic lichens

Lichens in Antarctic habitats are subjected to environmental extremes, including UVB radiation, desiccation and low temperatures, as well as to rapid fluctuations in these. Lichens synthesise a variety of chemical compounds in response to their environmental conditions which contribute towards their colour, and which act as protectants against physiological stresses. The fluorescence generated by the lichens at 532 nm can be used in epifluorescence microscopy to identify their presence on substrata but this can severely affect the Raman spectra using visible excitation. The advantage of the near infrared excitation used in FT-Raman spectroscopy in minimising fluorescence emission facilitates the molecular characterisation of lichen encrustations without having to remove the thallus from its substrate or remove or otherwise damage any part of the thallus. Spectroscopic biomarkers are proposed which allow the lichens to be characterised by the identification of characteristic lichen substances; the use of these biomarkers for the preliminary taxonomic identification of Antarctic lichens is examined and some potential pitfalls are described.     

Serious complications in experiments in which UV doses are effected by using different lamp heights

Many experiments examining plant responses to enhanced ultraviolet-B radiation (280–315 nm) simply compare an enhanced UV-B treatment with ambient UV-B (or no UV-B radiation in most greenhouse and controlled-environment studies). Some more detailed experiments utilize multiple levels of UV-B radiation. A number of different techniques have been used to adjust the UV dose. One common technique is to place racks of fluorescent UV-emitting lamps at different heights above the plant canopy. However, the lamps and associated support structure cast shadows on the plant bed below. We calculated one example of the sequence of shade intervals for two common heights of lamp racks and show the patterns and duration of shade which the plants receive is distributed differently over the course of the day for different heights of the lamp racks. We also conducted a greenhouse experiment with plants (canola, sunflower and maize) grown under unenergized lamp racks suspended at the same two heights above the canopy. Growth characteristics differed in unpredictable ways between plants grown under the two heights of lamp racks. These differences could enhance or obscure potential UV-B effects. Also, differences in leaf mass per unit foliage area, which were observed in this experiment, could contribute to differences in plant UV-B sensitivity. We recommend the use of other techniques for achieving multiple doses of UV-B radiation. These range from simple and inexpensive approaches (e.g., wrapping individual fluorescent tubes in layers of a neutral-density filter such as cheese cloth) to more technical and expensive alternatives (e.g., electronically modulated lamp control systems). These choices should be determined according to the goals of the particular experiment.     

AMELIORATION OF UV-B DAMAGE UNDER HIGH IRRADIANCE. I: ROLE OF PHOTOSYNTHESIS

Sensitivity to ultraviolet-B radiation (UV-B,280–315 nm) is generally reduced when background irradiance is high. We tested the involvement of photosynthesis in the amelioration of UV-B damage by treating plants at high PAR (photosynthetically-active radiation, 400–700 nm; 1000 μmol m-² s-¹) with supplemental UV-B at double ambient levels of biologically-effective radiation (18 kJ m-²d-¹) and either “ambient” (450 μmol mol-¹) or short term elevated (750 μmol mol-¹) CO₂ levels. Responses to UV-B were assessed by photosynthetic gas exchange, leaf expansion and production of UV-absorbing compounds (presumptive flavonoids) in cultivars of cucumber (Cucumis sativus L.) previously demonstrated to be relatively sensitive (cv. Poinsett) and insensitive (cv. Ashley) to UV-B. Except for marginal leaf interveinal chlorosis observed in Poinsett, both cultivars responded similarly. UV-B had little direct effect on leaf photosynthesis, but it did cause reductions in leaf area and corresponding increases in leaf dry matter per area. Increased CO, stimulated plant growth, counteracting the effect of UV-B on leaf growth and indicating an important role for photosynthesis. In contrast, the accumulation of UV-absorbing flavonoid compounds was enhanced by UV-B exposure but was not affected by COz enrichment.     

Plant responses to current solar ultraviolet-B radiation and to supplemented solar ultraviolet-B radiation simulating ozone depletion: an experimental comparison

Field experiments assessing UV-B effects on plants have been conducted using two contrasting techniques: supplementation of solar UV-B with radiation from fluorescent UV lamps and the exclusion of solar UV-B with filters. We compared these two approaches by growing lettuce and oat simultaneously under three conditions: UV-B exclusion, near-ambient UV-B (control) and UV-B supplementation (simulating a 30% ozone depletion). This permitted computation of "solar UV-B" and "supplemental UV-B" effects. Microclimate and photosynthetically active radiation were the same under the two treatments and the control. Excluding UV-B changed total UV-B radiation more than did supplementing UV-B, but the UV-B supplementation contained more "biologically effective" shortwave radiation. For oat, solar UV-B had a greater effect than supplemental UV-B on main shoot leaf area and main shoot mass, but supplemental UV-B had a greater effect on leaf and tiller number and UV-B-absorbing compounds. For lettuce, growth and stomatal density generally responded similarly to both solar UV-B and supplemented UV-B radiation, but UV-absorbing compounds responded more to supplemental UV-B, as in oat. Because of the marked spectral differences between the techniques, experiments using UV-B exclusion are most suited to assessing effects of present-day UV-B radiation, whereas UV-B supplementation experiments are most appropriate for addressing the ozone depletion issue.     

Coupling short-term changes in ambient UV-B levels with induction of UV-screening compounds

A substantial number of studies have been conducted over the last several decades to assess the potential impacts of long-term increases in ultraviolet-B radiation (UV-B between 280 and 320 nm) that will result from continued depletion of stratospheric ozone. However, seasonal changes, tropospheric chemistry and cloudiness are the dominant factors controlling ambient UV-B levels on a short-term or daily basis. The effects of short-term changes in UV-B on plant growth, phytochemistry and physiological processes have received relatively little attention. The USDA UV-B Monitoring and Research Program provides an excellent network of stations that provide an opportunity to monitor long-term changes in solar UV-B radiation and evaluate the responses of plants to short-term variation in UV-B levels on a near-real-time basis. In this study barley (Hordeum vulgare L.) and soybean (Glycine max [L] Merr.) were used as model systems. Emerging seedlings of these species were grown under either near-ambient levels of UV-B or under reduced levels (ca 90% reduction) in the field. Periodic measurements of foliar UV-screening compounds were made on separate groups of seedlings planted at intervals over the growing season during contrasting periods of ambient levels of UV radiation. The levels of UV-screening compounds correlated with UV-B levels in both species and with UV-A in soybean but the sensitivity of the response differed between the two species and among the soybean cultivars. Response differences among species may be related to unique secondary chemistry of each species, so one response estimate or action spectrum may not be appropriate for all species.     

EFFECTIVENESS OF UV-B RADIATION ON THE GROWTH AND PHYSIOLOGY OF FIELD-GROWN SOYBEAN MODIFIED BY WATER STRESS

Abstract— Soybeans [ Glycine max (L) Merr. cv Essex] were grown in field plots during May-October 1985 under ambient and an enhanced level of ultraviolet-B (UV-B) radiation (supplemental daily dose: 5.1 effective kJ m^-2). They were either subjected to water stress or supplementally irrigated, resulting in a 2.0 MPa lower soil water potential in stressed plots. Increased levels of UV-B radiation reduced leaf area, total plant dry weight and net photosynthesis under well-watered conditions, but no significant UV-B effects were detected in plants concurrently subjected to water stress. The insensitivity of growth and net photosynthesis to UV-B radiation in water-stressed plants may be related to anatomical and biochemical changes induced by water stress. These include an increase in the concentration of UV absorbing compounds in leaf tissues and leaf thickening.     

Solar ultraviolet-B radiation increases phenolic content and ferric reducing antioxidant power in Avena sativa

We examined the influence of solar ultraviolet-B radiation (UV-B; 280-320 nm) on the maximum photochemical efficiency of photosystem II (F(v)/F(m)), bulk-soluble phenolic concentrations, ferric-reducing antioxidant power (FRAP) and growth of Avena sativa. Treatments involved placing filters on frames over potted plants that reduced levels of biologically effective UV-B by either 71% (reduced UV-B) or by 19% (near-ambient UV-B) over the 52 day experiment (04 July-25 August 2002). Plants growing under near-ambient UV-B had 38% less total biomass than those under reduced UV-B. The reduction in biomass was mainly the result of a 24% lower leaf elongation rate, resulting in shorter leaves and less total leaf area than plants under reduced UV-B. In addition, plants growing under near-ambient UV-B had up to 17% lower F(v)/F(m) values early in the experiment, and this effect declined with plant age. Concentrations of bulk-soluble phenolics and FRAP values were 17 and 24% higher under near-ambient UV-B than under reduced UV-B, respectively. There was a positive relationship between bulk-soluble phenolic concentrations and FRAP values. There were no UV-B effects on concentrations of carotenoids (carotenes + xanthophylls).     

AMELIORATION OF UV-B DAMAGE UNDER HIGH IRRADIANCE. II: ROLE OF BLUE LIGHT PHOTORECEPTORS

Abstract Sensitivity of plants to UV-B radiation (280–315 nm) is often reduced at high background irradiance. Interpretation of plant responses to potential increases in solar UV-B requires improved understanding of interactions between UV-B and other environmental parameters. In this study, photosynthetically active radiation (PAR, 400–700 nm) was kept approximately constant (38 mol m-² per day) while the daily blue light fluence (BL, 400–500 nm) was varied between 0.23 and 2.68 mol m-². Two lines of cucumber (cvs Ashley and Poinsett) with differential sensitivity to UV-B were compared. At low BL, 3 days of UV-B treatment (21 kJ m-² biologically effective radiation per 10 h per day) caused severe inhibition of growth in a developing leaf in both cultivars. Growth effects were detectable sooner and were accompanied by chlorotic lesions in the sensitive cultivar (cv Poinsett). Supplemental BL progressively reduced symptoms, consistent with an important role for BL photoreceptor(s) in prevention or repair of UV-B damage. Ultraviolet-induced increases in UV-absorbing compounds on an area basis were significant within 24 h of the start of the treatment but were independent of BL fluence over the range tested, suggesting that bulk accumulation of screening pigments did not contribute to BL-dependent amelioration of UV damage. However, BL did stimulate net increases in extractable UV-absorbing compounds on a total leaf busis, while high-performance liquid chromatography analysis indicated that BL and UV-B acted synergistically to increase specific components. Thus, the data do not necessarily exclude UV-absorbing compounds from an important role in overall UV-B protection nor do they rule out some more specific function for these compounds (e.g. antioxidants). Finally, BL effects on UV-B alteration of leaf growth and accumulation of UV-absorbing compounds were not saturated under the conditions used here, suggesting that BL may contribute to interactions between UV-B and natural levels of background irradiance. Caution is urged in the interpretation of data on UV-B effects obtained under conditions of low BL irradiance.     

Effects of UV-B radiation on growth, photosynthesis, UV-B-absorbing compounds and NADP-malic enzyme in bean (Phaseolus vulgaris L.) grown under different nitrogen conditions.

The effects of UV-B radiation on growth, photosynthesis, UV-B-absorbing compounds and NADP-malic enzyme have been examined in different cultivars of Phaseolous vulgaris L. grown under 1 and 12 mM nitrogen. Low nitrogen nutrition reduces chlorophyll and soluble protein contents in the leaves and thus the photosynthesis rate and dry-matter accumulation. Chlorophyll, soluble protein and Rubisco contents and photosynthesis rate are not significantly altered by ambient levels of UV-B radiation (17 microW m-2, 290-320 nm, 4 h/day for one week). Comparative studies show that under high nitrogen, UV-B radiation slightly enhances leaf expansion and dry-matter accumulation in cultivar Pinto, but inhibits these parameters in Vilmorin. These results suggest that the UV-B effect on growth is mediated through leaf expansion, which is particularly sensitive to UV-B, and that Pinto is more tolerant than Vilmorin. The effect of UV-B radiation on UV-B-absorbing compounds and on NADP-malic enzyme (NADP-ME) activity is also examined. Both UV-B radiation and low-nitrogen nutrition enhance the content of UV-B-absorbing compounds, and among the three cultivars used, Pinto exhibits the highest increases and Arroz the lowest. The same trend is observed for the specific activity and content of NADP-ME. On a leaf-area basis, the amount of UV-B-absorbing compounds is highly correlated with the enzyme activity (r2 = 0.83), suggesting that NADP-ME plays a key role in biosynthesis of these compounds. Furthermore, the higher sensitivity of Vilmorin than Pinto to UV-B radiation appears to be related to the activity of NADP-ME and the capacity of the plants to accumulate UV-B-absorbing compounds.     

Role of white light in reversing UV-B-mediated effects in the N2-fixing cyanobacterium Anabaena BT2

The effects of various irradiances of artificial UV-B (280-315 nm) in the presence or absence of visible light (photosynthetically active radiation) on growth, survival, 14CO2 uptake and ribulose 1,5-bisphosphate carboxylase (RuBISCO) activity were studied in the N2-fixing cyanobacterium Anabaena BT2. We tested the hypothesis whether or not visible radiation offers any protection against UV-B-induced deleterious effects on growth and photosynthesis in Anabaena BT2. Attempts were also made to determine the irradiances of UV-B where inhibitory effects could be mitigated by simultaneous irradiation with visible light. Exposure of cultures to 0.2 W m(-2) or higher irradiance of UV-B caused inhibition of growth and survival and growth ceased above 1.0 W m(-2). 14CO uptake and RuBISCO activity were found to be more sensitive to UV-B and around 60% reduction in 14CO2 uptake and RuBISCO activity occurred after exposure of cultures to 0.4 W m(-2) for 1 h. However, growth, 14CO2 uptake and RuBISCO activity were nearly normal when UV-B (0.4 W m(-2)) and visible light (14.4 W m(-2)) were given simultaneously. Blue radiation (450 nm) was found to be the most effective in photoreactivation against UV-B, better than UV-A or any other light wavelength band. Our results demonstrate that the studied cyanobacterium possesses active photoreactivation mechanism(s) against UV-B-mediated damage which in turn probably allow survival under natural conditions in spite of being continuously exposed to the UV-B component present in the solar radiation. Continued growth of many algae and cyanobacteria in the presence of intense solar UV-B radiation under natural conditions seems to be due to the active role of photoreactivation.     

UV-B-INDUCED INCREASE IN SPECIFIC LEAF WEIGHT OF CUCUMBER AS A CONSEQUENCE OF INCREASED STARCH CONTENT

Abstract Specific leaf weight (SLW), the ratio of leaf dry matter to area, often increases in plants exposed to elevated UV-B radiation (280–315 nm). Increased SLW can result from greater leaf thickness or increased leaf density ( e.g . accumulation of high density substances in cells). The basis for large increases in SLW was examined in the first and third leaves of cucumber differing in developmental stage at the start of UV treatment. Leaf 1 was approximately 50% fully expanded, while leaf 3 had just unfolded. It is shown here that up to 80% of the UV-generated change in SLW in leaf 1 was caused by accumulation of nonstructural carbohydrates, especially starch (increasing from 13 to 23% of total dry weight). Leaf 3 contained a much smaller proportion of nonstructural carbohydrates (less than 8%) and the effect on SLW was correspondingly less. As shown in the previous paper, UV-B inhibition of growth in leaf 3 was reversed by supplemental blue light (BL) in a fluence-dependent manner between 0.23 to 2.68 mol m ² perday. Fluence-response curves revealed that supplemental BL reversed both the UV-induced accumulation of starch and increase in SLW in leaf 1 over the same range. The data are consistent with a back-up of photosynthate into leaf 1 as a result of UV-B inhibition of growth in leaf 3. The data also demonstrate that increases in SLW cannot be assumed to represent increases in leaf thickness.     

Effects of solar radiation on the Patagonian macroalga Enteromorpha linza (L.) J. Agardh-Chlorophyceae.

The photosynthetic performance of Enteromorpha linza (L.) J. Agardh-Chlorophyceae was determined with a portable PAM instrument in situ and under seminatural radiation conditions in Patagonia, Argentina. Solar radiation was measured in parallel with a three-channel radiometer, ELDONET (Real Time Computer, M?hrendorf, Germany), in three wavelength ranges, UV-B (280-315 nm), UV-A (315-400 nm), and PAR (400-700 nm). The effective photosynthetic quantum yield decreased after 15-min exposure to solar radiation when the thalli were kept in a fixed position but recovered in the subsequent shade conditions within several hours. A 30-min exposure of free floating thalli, however, caused less photoinhibition. The photosynthetic quantum yield of E. linza was also followed over whole days under clear sky, partly cloudy and rainy conditions in a large reservoir of water (free floating thalli) and in situ (thalli growing in rock pools). Most of the observed effect was due to visible radiation; however, the UV wavelength range, and especially UV-B, caused a significant reduction of the photosynthetic quantum yield. Fluence rate response curves indicated that the species is a typical shade plant which showed non-photochemical quenching at intermediate and higher irradiances. This is a surprising result since these algae are found in the upper eulittoral where they are exposed to high irradiances. Obviously they utilize light only during periods of low irradiances (morning, evening, high tide) while they shut down the electron transport chain during intensive exposure. Fast induction and relaxation kinetics have been measured in these algae for the first time and indicated a rapid adaptation of the photosynthetic capacity to the changing light conditions as well as a fast decrease of PS II fluorescence upon exposure to solar radiation. There was a strong bleaching of chlorophyll due to exposure to solar radiation but less drastic bleaching of carotenoids.     

INFLUENCE OF PHOTON FLUX DENSITY IN THE 400–700 nm WAVEBAND ON INHIBITION OF PHOTOSYNTHESIS BY UV-B (280–320 nm) IRRADIATION IN SOYBEAN LEAVES: SEPARATION OF INDIRECT AND IMMEDIATE EFFECTS

Abstract— Visible radiation can substantially influence the degree to which plant photosynthesis is inhibited by UV-B radiation. This study was designed to separate the immediate effects of visible radiation on UV-B photosynthetic inhibition from the indirect influence of visible irradiation on morphological and physiological properties of leaves during leaf development. Soybean plants were pretreated in growth chambers with either high or low visible irradiance (750 and 70 μmol m^-2s^-1 quantum flux in the 400–700 nm waveband, respectively) during the development of leaves used subsequently for UV irradiation. Test leaves still attached to the plant were exposed to 5 h of polychromatic UV-B irradiation and the photosynthetic capacity (net CO₂ exchange) was determined before and after the UV irradiation. During the UV irradiation, plants from both pretreatment groups received either high or low visible flux. Development of leaves in the high visible flux pretreatment conditions resulted in thicker leaves, higher chlorophyll a/b ratios, more UV-absorbing pigments, and reduced sensitivity to the UV-B irradiation. However, higher visible flux during the UV-B irradiation resulted in greater depression of photosynthesis by the UV-B irradiation. The relative magnitude of photosynthetic depression under these treatment combinations was the same when photosynthesis was measured under either light-limited or light-saturated conditions.     

In situ imaging of usnic acid in selected Cladonia spp. by vibrational spectroscopy

In this study, we demonstrate the first in situ detection of usnic acid (UA) in selected species of the lichen Cladonia, using FPA-FTIR imaging and Raman microscopy. Fruticose lichens present a variety of defensive mechanisms, one of which is the production of UA. This polyketide secondary metabolite, produced by certain lichenized fungi, has a protective function for the lichen that includes a strong absorption in the ultraviolet range. Upon confirming the distinct spectral signature of UA in lichen tissue, we mapped its distribution in Cladonia arbuscula, Cladonia uncialis and Cladonia sulphurina tissues. Spectroscopic images were obtained from cryosectioned lichen fragments embedded in media and from hand-sectioned fragments that were media-free. UA was present in the pycnidia, and younger walls of C. arbuscula and C. uncialis, the spore-producing region of a C. uncialis apothecium, and in both the younger and older soredia of C. sulphurina. The localization of UA in lichens is an important precursor to future work that includes the identification of the gene cluster responsible for its biosynthesis. Our results show that FTIR and Raman imaging can be an effective way to study the distribution of natural products in lichens with micron-scale precision.     

Influence of PAR and UV-A in determining plant sensitivity and photomorphogenic responses to UV-B radiation

The role of photosynthetically active radiation (400-700 nm) (PAR) in modifying plant sensitivity and photomorphogenic responses to ultraviolet-B (280-320 nm) (UV-B) radiation has been examined by a number of investigators, but few studies have been conducted on ultraviolet-A (320-400 nm) (UV-A), UV-B and PAR interactions. High ratios of PAR-UV-B and UV-A-UV-B have been found to be important in ameliorating UV-B damage in both terrestrial and aquatic plants. Growth chamber and greenhouse studies conducted at low PAR, low UV-A and high UV-B often show exaggerated UV-B damage. Spectral balance of PAR, UV-A and UV-B has also been shown to be important in determining plant sensitivity in field studies. In general, one observes a reduction in total biomass and plant height with decreasing PAR and increasing UV-B. The protective effects of high PAR against elevated UV-B may also be indirect, by increasing leaf thickness and the concentration of flavonoids and other phenolic compounds known to be important in UV screening. The quality of PAR is also important, with blue light, together with UV-A radiation, playing a key role in photorepair of DNA lesions. Further studies are needed to determine the interactions of UV-A, UV-B and PAR.     

In vitro AND in vivo ULTRAVIOLET-INDUCED ALTERATIONS OF OXY- AND DEOXYHEMOGLOBIN

Ultraviolet (UV) radiation was found to convert oxyhemoglobin and deoxyhemoglobin stoichiometrically into methemoglobin and a met-like product, respectively. The peak conversion efficiency for oxyhemoglobin occurred at 285 nm and decreased by a factor of 100 by 315 nm. The peak conversion efficiency for deoxyhemoglobin occurred at 290 nm and decreased by a factor of 30 by 320 nm. The transformation of oxyhemoglobin to methemoglobin was also documented in intact erythrocytes using UV-B radiation. Finally, similar transformations were found to occur in human skin with UV-B exposure but not on all volunteers tested. These results imply that methemoglobin will be formed in vivo on solar exposure and provide evidence that UV-B photons reach the blood vessels.     

Ultraviolet radiation and the snow alga Chlamydomonas nivalis (Bauer) Wille

Aplanospores of Chlamydomonas nivalis are frequently found in high-altitude, persistent snowfields where they are photosynthetically active despite cold temperatures and high levels of visible and ultraviolet (UV) radiation. The goals of this work were to characterize the UV environment of the cells in the snow and to investigate the existence and localization of screening compounds that might prevent UV damage. UV irradiance decreased precipitously in snow, with UV radiation of wavelengths 280-315 nm and UV radiation of wavelengths 315-400 nm dropping to 50% of incident levels in the top 1 and 2 cm, respectively. Isolated cell walls exhibited UV absorbance, possibly by sporopollenin, but this absorbance was weak in images of broken or plasmolyzed cells observed through a UV microscope. The cells also contained UV-absorbing cytoplasmic compounds, with the extrachloroplastic carotenoid astaxanthin providing most of the screening. Additional screening compound(s) soluble in aqueous methanol with an absorption maximum at 335 nm played a minor role. Thus, cells are protected against potentially high levels of UV radiation by the snow itself when they live several centimeters beneath the surface, and they rely on cellular screening compounds, chiefly astaxanthin, when located near the surface where UV fluxes are high.     

Removing UV-A and UV-C radiation from UV-B fluorescent lamp emissions. Differences in the inhibition of photosynthesis in the marine alga Dunaliella tertiolecta using chromate versus cellulose acetate-polyester filters

Ultraviolet-B (UV-B; 280-320 nm)-emitting lamps unavoidably emit ultraviolet-A (UV-A; 320-400 nm) and ultraviolet-C (UV-C; <280 nm) radiation. Short-wavelength-blocking filters are generally used to limit the wave bands of UV under investigation. The widespread use of such filters means that all exposures to UV-B radiation will have a significant UV-A component. Therefore, the physiological effects unique to UV-B exposure are difficult to clearly isolate. This study presents a method to remove the UV-A and UV-C "contamination" using a liquid potassium chromate (K(2)CrO(4)) filter, thus allowing more direct assessment of the effects of UV-B exposure. Cultures of the green marine alga Dunaliella tertiolecta were grown in the absence of UV radiation. Sunlamps supplied the UV radiation for a 24 h exposure (solar radiation was not used in this study). The UV radiation was filtered either by the standard method (i.e. cellulose acetate (CA) with polyester = Mylar controls) or by a liquid filter of potassium chromate. Photosynthetic responses were compared. Major decreases in the ratio of variable to maximal fluorescence in dark-adapted cells and photosynthetic capacity were observed in CA-filtered cultures, whereas no change was observed in cells exposed to the same UV-B flux with the UV-A removed by K(2)CrO(4). The use of a CA filter with a Mylar control does not link results unequivocally to UV-B radiation. Such results should be interpreted with caution.