Mesophyll Resistances to SO(2) Fluxes into Leaves

Uptake of label from solutions containing ³⁵SO₂, H³⁵SO₃⁻ and ³⁵SO₃²⁻ into mesophyll protoplasts, vacuoles, and chloroplasts isolated from young barley leaves was measured at different pH values. Uptake was fast at low pH, when the concentration of SO₂ was high, and low at high pH, when the concentration of SO₂ was low. When the resistance (R) of plasmalemma, tonoplast, and chloroplast envelope to the penetration of SO₂ was calculated from rates of uptake of label, comparable values were obtained for the different biomembranes at low pH values. R was close to 8000 seconds per meter and permeability coefficients were close to 1.25 × 10⁻⁴ meters per second. Under these conditions R may describe resistance to SO₂ diffusion across a lipid bilayer. At higher pH values, R decreased. As R was calculated on the assumption that SO₂ is the only penetrating molecular species, the data suggest that carrier-mediated anion transport contributes to the uptake of sulfur at physiological pH values thereby decreasing apparent R_SO2. The contribution of anion transport appeared to be smaller for transfer across the plasmalemma than for transfer across the tonoplast. It was large for transfer across the chloroplast envelope. The phosphate translocator of the chloroplast envelope catalyzed uptake of SO₃²⁻ into chloroplasts at neutral pH. Uptake was decreased in the presence of high levels of phosphate or sulfate and by pyridoxal phosphate. SO₂ transfer into cells leads to the intracellular liberation of one or two protons, depending on pH and oxidizing conditions. When the divalent sulfite anion is exchanged across the chloroplast envelope, bisulfite formation results in proton uptake in the chloroplast stroma, whereas SO₂ uptake into chloroplasts lowers the stroma pH.     

Can plants exposed to SO2 excrete sulfuric acid through the roots?

Hydroponically grown pea plants (Pisum sativum L., cv. Kleine Rheinländerin) and barley seedlings (Hordeum vulgare L., cv. Gerbel) were fumigated for several days with 1 or 2 μl l^?1 SO₂. Both species accumulated sulfate during fumigation, although the nutrient medium lacked sulfate. In pea, SO₂-dependent sulfate accumulation in different plant parts accounted for 60 percent of the SO₂ sulfur which, as calculated from a determination of boundary and stomatal flux resistances had entered the leaves. Up to 55% of the air-borne sulfate was translocated from pea leaves to roots during the period of fumigation, but no or only little sulfate was excreted into the nutrient solution. In contrast, barley retained sulfate in the leaves, and sulfate translocation from shoot to the root system could not be observed. In both species, protons were excreted by the roots. In fumigated plants, proton loss was higher than in untreated controls in pea, but not in barley. In pea, SO₂-dependent proton loss into the medium accounted for up to 50% of the sulfuric acid formed from SO₂. Proton excretion was strongly dependent on potassium availability in the nutrient medium. Cation uptake by the plants during fumigation was sufficient to compensate for proton loss, suggesting proton/cation exchange at the interface between root and medium. We conclude that by oxidation to sulfuric acid, plants are capable of detoxifying SO₂ taken up by the leaves. Depending on plant species, either both protons and sulfate anions can be exported from the leaves, or the proton load on leaf cells can be relieved by proton/cation exchange at the plasmalemma. Finally, the problem of airborne plant acidification may be solved by proton/cation exchange at the level of roots. The burden of acidification is then shifted from the plant to the nutrient medium. Appreciable amounts of sulfate can be excreted neither by pea nor by barley plants.     

Long-term effects of SO2 on plants,SO2 metabolism and regulation of intracellular pH

The impact of SO₂ on the ionic balance of plants and its implications for intracellular pH regulation was studied to find explanations for long-term effects of SO₂. When sulphur, taken up as SO₂ by the shoots of plants, is not assimilated in organic compounds, but stored as sulphate, an equivalent amount of H⁺ is produced. These H⁺ ions are not buffered chemically, but removed by metabolic processes.On the basis of knowledge on metabolic buffering mechanisms a conceptual model is proposed for the removal of shoot-generated H⁺ by (i) OH^- ions, produced in the leaves when sulphate and nitrate are assimilated in organic compounds and/or by (ii) OH^- ions produced by decarboxylation of organic anions (a biochemical pH stat mechanism). The form in which nitrogen is supplied largely determines the potential of the plant to neutralize H⁺ in the leaves during SO₂ uptake by the proposed mechanisms.In field experiments with N₂ fixing Vicia faba L. crops, the increase of sulphate in the shoots of SO₂-exposed plants was equivalent in charge to the decrease of organic anion content, calculated as the difference between inorganic cation content (C) and inorganic anion content (A), indicating that H⁺ ions produced in the leaves following SO₂ uptake were partly removed by OH^- from sulphate reduction and partly by decarboxylation of organic anions.The appearance of chronic SO₂ injury (leaf damage) in the field experiment at the end of the growing period is discussed in relation to the impact of SO₂ on the processes involved in regulation of intracellular pH. It is proposed that the metabolic buffering capacity of leaf cells is related to the rates of sulphate and nitrate reduction and the import rate of organic anions, rather than to the organic anion content in the vacuoles of the leaf cells.     

Interspecific Variation in SO(2) Flux : Leaf Surface versus Internal Flux, and Components of Leaf Conductance

The objective of this study was to clarify the relationships among stomatal, residual, and epidermal conductances in determining the flux of SO₂ air pollution to leaves. Variations in leaf SO₂ and H₂O vapor fluxes were determined using four plant species: Pisum sativum L. (garden pea), Lycopersicon esculentum Mill. flacca (mutant of tomato), Geranium carolinianum L. (wild geranium), and Diplacus aurantiacus (Curtis) Jeps. (a native California shrub). Fluxes were measured using the mass-balance approach during exposure to 4.56 micromoles per cubic meter (0.11 microliters per liter) SO₂ for 2 hours in a controlled environmental chamber. Flux through adaxial and abaxial leaf surfaces with closed stomata ranged from 1.9 to 9.4 nanomoles per square meter per second for SO₂, and 0.3 to 1.3 millimoles per square meter per second for H₂O vapor. Flux of SO₂ into leaves through stomata ranged from ~0 to 8.5 (dark) and 3.8 to 16.0 (light) millimoles per square meter per second. Flux of H₂O vapor from leaves through stomata ranged from ~0 to 0.6 (dark) to 0.4 to 0.9 (light) millimole per square meter per second. Lycopersicon had internal flux rates for both SO₂ and H₂O vapor over twice as high as for the other species. Stomatal conductance based on H₂O vapor flux averaged from 0.07 to 0.13 mole per square meter per second among the four species. Internal conductance of SO₂ as calculated from SO₂ flux was from 0.04 mole per square meter per second lower to 0.06 mole per square meter per second higher than stomatal conductance. For Pisum, Geranium, and Diplacus stomatal conductance was the same or slightly higher than internal conductance, indicating that, in general, SO₂ flux could be predicted from stomatal conductance for H₂O vapor. However, for the Lycopersicon mutant, internal leaf conductance was much higher than stomatal conductance, indicating that factors inside leaves can play a significant role in determining SO₂ flux.     

The relationship between absorption of sulfur dioxide (SO2) and inhibition of photosynthesis in several plants

Photosynthetic rate, transpiration rate and SO₂ absorption rate were simultaneously measured under exposure to SO₂ (0.1–1.0 μl l ^?1) for 5 or 8 hr in six species belonging to C₄ or C₃ plants (Zea mays, Sorghum vulgare, Amaranthus tricolor, Oryza sativa, Avena sativa andHelianthus annuus). Distinct interspecific differences were found as to the extent of inhibition of photosynthetic rate. Calculation of diffusive resistance to H₂O(r) and SO₂(r′) showed that the ratio of r′/r was 1.9 irrespective of species and coincided well with the theoretical value based on molecular diffusion. Thus it was made clear that the absorption of SO₂ was dependent upon the gas exchange capacity of leaf blade. Using the ratio of r′/r the rate of SO₂ absorption could be calculated from transpiration rate and was compared with the inhibition rate of photosynthesis. In three C₄ species, the inhibition of photosynthesis increased linearly with the amount of SO₂ absorbed during a 5-hour period. The pattern of inhibition of photosynthesis inA. sativa andH. annuus among C₃ species was similar to that of C₄ species until the amount of SO₂ absorbed reached 60 mg-SO₂ m^?2 above which the inhibition abruptly increased. The inhibition of photosynthesis inO. sativa was exceptionally severe even with only a small amount of SO₂ absorbed.     

Effects of SO(2) on Stomatal Metabolism in Pisum sativum L

Pea (Pisum sativum L. cv `Little Marvel') plants were exposed to SO₂ for short term (3 hours) and long term (2 days) at 0.2 and at 0.5 microliter per liter (ppm) levels. The effect of this treatment on the activity of phosphoenolpyruvate carboxylase, NAD- and NADP-malate dehydrogenases, and alanine aminotransferase from epidermis and whole leaves was investigated. Short-term exposure to SO₂ at 0.2 or 0.5 ppm decreased the activity of the carboxylase and the dehydrogenases in the epidermis. In contrast, the activity of the same three enzymes increased in whole leaves with either short- or long-term exposure to SO₂. Alanine aminotransferase in epidermis or whole leaves was not much affected by short-term exposure, but the epidermal activity was decreased and whole leaf activity was increased with long-term exposure. SO₂ exposure which was initiated prior to illumination decreased the free thiol content of both epidermis and of whole leaf. Net photosynthesis was reversibly inhibited by long-term exposure to SO₂ at 0.5 ppm. No effect of 0.5 ppm SO₂ on stomatal conductance was detectable after 3 hours. Stomatal conductance appeared to decrease after longer exposure times (2 days) at 0.5 ppm.     

Joint Action of O(3) and SO(2) in Modifying Plant Gas Exchange

The joint action of O₃ and SO₂ stress on plants was investigated by determining the quantitative relationship between air pollutant fluxes and effects on stomatal conductance. Gas exchange measurements of O₃, SO₂, and H₂O vapor were made for Pisum sativum L. (garden pea). Plants were grown under controlled environments, and O₃, SO₂, and H₂O vapor fluxes were evaluated with a whole-plant gas exchange chamber using the mass-balance approach. Maximum O₃ and SO₂ fluxes per unit area (2 sided) into leaves averaged 8 nanomoles per square meter per second with exposure to either O₃ or SO₂ at 0.1 microliters per liter. Internal fluxes of either O₃ or SO₂ were reduced by up to 50% during exposure to combined versus individual pollutants; the greatest reduction occurred with simultaneous versus sequential combinations of the pollutants. Stomatal conductance to H₂O was substantially altered by the pollutant exposures, with O₃ molecules twice as effective as SO₂ molecules in inducing stomatal closure. Stomatal conductance was related to the integrated dose of pollutants. The regression equations relating integrated dose to stomatal conductance were similar with O₃ alone, O₃ plus added SO₂, and O₃ plus SO₂ simultaneously; i.e. a dose of 100 micromoles per square meter produced a 39 to 45% reduction in conductance over nonexposed plants. With SO₂ alone, or SO₂ plus added O₃, a dose of 100 micromoles per square meter produced a 20 to 25% reduction in conductance. When O₃ was present at the start of the exposure, then stomatal response resembled that for O₃ more than the response for SO₂. This study indicated that stomatal responses with combinations of O₃ and SO₂ are not dependent solely on the integrated dose of pollutants, but suggests that a metabolic synergistic effect exists.     

The role of stomata in sensitivity of Betula papyrifera seedlings to SO2 at different humidities

Summary Stomata of paper birch (Betula papyrifera Marsh.) seedlings were more open at high humidity than at low humidity and responded rapidly to changes in vapor pressure deficit. SO₂ at 0.2 or 0.8 l l^-1 caused partial stomatal closure. Seedlings fumigated with SO₂ at 0.2 or 0.5 l l^-1 for 30 h or 0.2 l l^-1 for 75 h took up more SO₂ at high than at low humidity. Differences in pollutant uptake could be explained by stomatal conductance with no need to invoke changes in mesophyll conductance. Betula seedlings were more sensitive to SO₂ when fumigated at high humidity, as manifested in more leaf necrosis, increased leaf abscission, and greater growth inhibition compared to seedlings fumigated at low humidity. Amount of injury to leaves increased with rate of SO₂ uptake, and inhibition of root growth increased with total SO₂ uptake.Abbreviations RH relative humidity - VPD vapor pressure deficit - RGR mean relative growth rate - PPFD photosynthetic photon flux density (400–700 nm) - LDC leaf diffusive conductance - water potential Research supported by the College of Agricultural and Life Sciences, University of Wisconsin-Madison     

Sulfur-dioxide fluxes into different cellular compartments of leaves photosynthesizing in a polluted atmosphere

A computer model is used to analyze fluxes of SO₂ from polluted air into leaves and the intracellular distribution of sulfur species derived from SO₂. The analysis considers only effects of acidification and of anion accumulation. (i) The SO₂ flux into leaves is practically exclusively controlled by the boundary-layer resistance of leaves to gas diffusion and by stomatal opening. At constant stomatal opening, flux is proportional to the concentration of SO₂ in air. (ii) The sink capacity of cellular compartments for SO₂ depends on intracellular pH and the intracellular localization of reactions capable of oxidizing or reducing SO₂. In the mesophyll of illuminated leaves, the chloroplasts possess the highest trapping potential for SO₂. (iii) If intracellular ion transport were insignificant, and if bisulfite and sulfite could not be oxidized or reduced, leaves with opened stomata would rapidly be killed both by the accumulation of sulfites and by acidification of chloroplasts and cytosol even if SO₂ levels in air did not exceed concentrations thought to be permissible. Acidification and sulfite accumulation would remain confined largely to the chloroplasts and to the cytosol under these conditions. (iv) Transport of bisulfite and protons produced by hydration of SO₂ into the vacuole cannot solve the problem of cytoplasmic accumulation of bisulfite and sulfite and of cytoplasmic acidification, because SO₂ generated in the acidic vacuole from the bisulfite anion would diffuse back into the cytoplasm. (v) Oxidation to sulfate which is known to occur mainly in the chloroplasts can solve the problem of cytoplasmic sulfite and bisulfite accumulation, but aggravates the problem of chloroplastic and cytosolic acidification. (vi) A temporary solution to the problem of acidification requires the transfer of H⁺ and sulfate into the vacuole. This transport needs to be energized. The storage capacity of the vacuole for protons and sulfate defines the extent to which SO₂ can be detoxified by oxidation and removal of the resulting protons and sulfate anions from the cytoplasm. Calculations show that even at atmospheric levels of SO₂ thought to be tolerable, known vacuolar buffer capacities are insufficient to cope with proton production during oxidation of SO₂ to sulfate within a vegetation period. (vii) A permanent solution to the problem of acidification is the removal of protons. Protons are consumed during the reduction of sulfate to sulfide. Proteins and peptides contain sulfur at the level of sulfide. During photosynthesis in the presence of the permissible concentration of 0.05l·l^-1 SO₂, sulfur may be deposited in plants at a ratio not far from 1/500 in relation to carbon. The content of reduced sulfur to carbon is similar to that ratio only in fast-growing, protein-rich plants. Such plants may experience little difficulty in detoxifying SO₂. In contrast, many trees may contain reduced sulfur at a ratio as low as 1/10 000 in relation to carbon. Excess sulfur deposited in such trees during photosynthesis in polluted air gives rise to sulfate and protons. If detoxification of SO₂ by reduction is inadequate, and if the storage capacity of the vacuoles for protons and sulfate is exhausted, damage is unavoidable. Calculations indicate that trees with a low ratio of reduced S to C cannot tolerate long-term exposure to concentrations of SO₂ as low as 0.02 or 0.03 l·l^-1 which so far have been considered to be non-toxic to sensitive plant species.     

SO(2) Effect on Photosynthetic Activities of Intact Sugar Maple Leaves as Detected by Photoacoustic Spectroscopy

Short-term (4 hours) effect of different concentrations of SO₂ fumigation on in vivo photochemical activities of sugar maple (Acer saccharum Marsh.) leaves was investigated using photoacoustic spectroscopy. The relative quantum yield of O₂ evolution (ratio of O₂ signal to the photothermal signal) and photochemical energy storage are increased by 0.05 microliter per liter of SO₂. This increase is more pronounced in 5 to 7 year old saplings than in 3 month old seedlings. Both oxygen-relative quantum yield and energy storage of seedlings are inhibited by increased concentrations of SO₂ and the inhibition is concentration dependent. The inhibition is greater in seedlings than in saplings at 2 microliters per liter of SO₂, indicating the more susceptible nature of seedlings. The present study indicates a concentration dependent differential effect of SO₂ on photochemical activities of sugar maple leaves.     

Phytotoxicity of Air Pollutants: Evidence for the Photodetoxification of SO(2) but Not O(3)

Pisum sativum L. cv Alsweet (garden pea) and Lycopersicon esculentum flacca Mill. (tomato) were used to evaluate the phytotoxicity of SO₂ and O₃ in the light and dark. Plants were grown in controlled environment chambers and exposed to SO₂ or O₃ in the light or dark at the same environmental conditions at which they were grown. The pea plants were treated with fusicoccin to ensure open stomata in the dark; the stomata of the tomato mutant remained open in the dark. Both species exhibited 64% to 80% less foliar necrosis following exposure to SO₂ (0.5 to 1.0 microliter per liter for 2 hours) in the light than in the dark. The decrease in SO₂ injury for light versus dark exposed plants was greater in fully expanded than expanding leaves. Both species exhibited 30% greater foliar necrosis following exposure to O₃ (0.2 microliter per liter for 2 hours) in the light than dark. The increase in O₃ injury in the light versus dark was similar for leaves at all stages of expansion. Leaf conductance to water vapor was 7% to 11% and 23% higher in the light than dark for fusicoccin-treated peas and tomato plants, respectively, indicating greater foliar uptake of both pollutants in the light than dark. Thus, the decreased SO₂ toxicity in the light was not associated with pollutant uptake, but rather the metabolism of SO₂. In contrast, the increased toxicity of O₃ in the light was at least in part associated with increased uptake or could not be separated from it.     

Foliar Fatty Acids and Sterols of Soybean Field Fumigated with SO(2)

Sixty-day-old soybean plants were exposed in the field to 78.7 parts per one-hundred million of SO₂ in an open-air fumigation system for 20 days. Leaves from the top one-fourth and bottom one-fourth of the plants were analyzed for chlorophyll, free fatty acids, fatty acid esters, polar lipid fatty acids, and sterols. Fumigated plants had a lower chlorophyll, free fatty acid, and polar lipid content, but a higher fatty acid ester content. Of the individual fatty acids, linoleic and linolenic acid increased with SO₂ fumigation while palmitic acid decreased. SO₂ fumigations had only a minor effect on leaf sterols. In general, the lower, more mature leaves showed a greater response to SO₂ exposure.     

Responses of dry weight growth under SO2 stress in an SO2-tolerant plant,Polygonum cuspidatum

The effects of exposure to 0.5–0.7 ppm SO₂ for about one month on the dry weight growth and net photosynthesis ofPolygonum cuspidatum were investigate. Furthermore, the carbon and nitrogen concentrations in each plant organ were measured. The results obtained showed no significant decrease in the total dry weight of SO₂-treated plants in comparison with controls. On the other hand, the leaf area (LA) and/or leaf dry weight of SO₂-treated plants were increased, and the root dry weight (RW) was decreased, in comparison with controls. The leaf carbon assimilation rate (CAR) in SO₂-treated plants was slightly decreased in spite of a clear decrease in net photosynthesis, and the value of (SW+RW)/LA (SW stem dry weight) was decreased in comparison with controls, thus minimizing the reduction in CAR. Furthermore, the ratio of total leaf carbon absorption (leaf area x CAR) to total root nitrogen absorption (root dry weight x nitrogen assimilation rate) in SO₂-treated plants was similar to that in controls. From these results, it can be concluded that an increase in leaf area and/or leaf dry weight and a decrease in root dry weight inP. cuspidatum under SO₂ stress may be induced in order to compensate for the decrease in CAR and to maintain the ratio of total leaf carbon absorption to total root nitrogen absorption in the early stage of vegetative growth.     

Effects of SO(2) and O(3) on Allocation of C-Labeled Photosynthate in Phaseolus vulgaris

A series of laboratory exposures of two varieties of bush bean (Phaseolus vulgaris L., var 274 and var 290) was conducted to determine the sensitivity of [¹⁴C]photosynthate allocation patterns to alteration by SO₂ and O₃. Experiments with the pollution-resistant 274 variety demonstrated short-term changes in both ¹⁴C and biomass allocation to roots of ¹⁴CO₂-labeled plants but no significant effect on yield by up to 40 hours of exposure to SO₂ at 0.50 microliters per liter or 4 hours of O₃ at 0.40 microliters per liter. Subsequent experiments with the more sensitive 290 variety demonstrated significant alteration of photosynthesis, translocation, and partitioning of photosynthate between plant parts including developing pods. Significant increases in foliar retention of photosynthate (+40%) occurred after 8 hours of exposure to SO₂ at 0.75 microliters per liter (6.0 microliters per liter-hour) and 11 hours of exposure to O₃ at 0.30 microliters per liter-hour (3.3 microliters-hours). Time series sampling of labeled tissues after ¹⁴CO₂ uptake showed that the disruption of translocation patterns was persistent for at least 1 week after exposures ceased. Subsequent longer-term exposures at lower concentrations of both O₃ (0.0, 0.10, 0.15, and 0.20 microliters per liter) and SO₂ (0.0, 0.20, and 0.40 microliters per liter) demonstrated that O₃ more effectively altered allocation than SO₂, that primary leaves were generally more sensitive than trifoliates, and that responses of trifoliate leaves varied with plant growth stage. Altered rates of allocation of photosynthate by leaves were generally associated with alterations of similar magnitude and opposite direction in developing pods. Collectively, these experiments suggest that allocation patterns can provide sensitive indices of incipient growth responses of pollution-stressed vegetation.     

Subcellular distribution of 35S-sulfur in spinach leaves after application of 35SO 4 2- , 35SO 3 2- , and 35SO2

³⁵SO₂, ³⁵SO ₃ ^2- , and ³⁵SO ₄ ^2- , respectively, were applied to leaves of Spinacia oleracea L. for 60 min in the light. Thereafter, the specific activity was determined in the organelles separated by means of sucrose density gradient centrifugation. In mitochondria and peroxisomes, the specific activity was equally distributed in their protein moieties. After application of ³⁵SO₂ or ³⁵SO ₃ ^2- , the chloroplast lamellae are characterized by elevated specific activity, which is not found after application of ³⁵SO ₄ ^2- . Chloroplast stroma shows a low specific incorporation rate after application of either compound, which may be due to the low turnover rate of Fraction I protein.     

Partitioning of carbon an SO2-sulfur in a native grassland

Summary The seasonal assimilation and within-plant partitioning of ¹⁴CO₂-carbon and ³⁵SO₂-sulfur in field plots of mixed-grass prairie was investigated, as was the dry deposition of ³⁵SO₂ onto surfaces of dead leaves, litter, and soil, and possible effects of continuous low-level SO₂ fumigation on these processes. The proportion of total net-assimilated carbon found below-ground was 45% in May, 51% in July, and 17% in September. As the season progressed, greater proportions of assimilate were partitioned to 5–20 cm depths and less to the 0–5 cm depth. Rhizomes and crowns received greater proportions in late season. Significant fractions of total ³⁴SO₂-deposited sulfur were recovered on dead leaf surfaces as well as litter and soil, suggesting estimates of SO₂ removal based on stomatal resistance alone are inadequate. Only 4% to 7% of total deposited sulfur was translocated belowground, with most going to 0–5 cm roots. In July much greater proportions of the total translocated SO₂-sulfur were found in deeper depths than in September. On SO₂-fumigated plots roots had lower total sulfur concentrations than controls. Furthermore, while on control plots total sulfur in roots at 5–20 cm increased from May to July and decreased from July to September, on fumigated plots there was a decrease followed by an increase suggesting that SO₂ uptake by shoots interferes with the normal pattern of root sulfur uptake and redistribution within the plant. Continuous SO₂ fumigation also seemed to stimulate root growth in July, possibly through a stimulation of photosynthesis.     

Role of superoxide dismutase in defense against SO2 toxicity and an increase in superoxide dismutase activity with SO2 fumigation

The role of superoxide dismutase (SOD) in defense against SO₂toxicity was investigated using leaves of poplar and spinach.Young poplar leaves having five times the SOD of the old leaveswere more resistant to the toxicity of SO₂. Spraying spinachleaves with diethyldithiocarbamate caused a marked loss of SODactivity which resulted in a decrease in their resistance tothe toxic effects of SO₂. The SOD activity in poplar leaveswas increased by fumigation with 0.1 ppm SO₂, and this was moreevident in young leaves than in old ones. The increased SODactivity was inhibited by cyanide. The poplar leaves havinghigh SOD activity induced with SO₂ fumigation were more resistantto 2.0 ppm SO₂ than the control leaves. These findings suggestthat SO₂ toxicity is in part due to the superoxide radical andthat SOD participates in the defense mechanism against SO₂ toxicity. (Received February 12, 1980; )     

Stomatal SO2 uptake and sulfate accumulation in needles of Norway spruce stands (Picea abies) in Central Europe

Monthly uptake rates and the annual deposition of gaseous SO₂ via the stomata of six Norway spruce canopies (Picea abies (L.) Karst.) in Germany (Königstein im Taunus, Witzenhausen, Grebenau, Frankenberg, Spessart, Fürth im Odenwald) were calculated (i) from statistical response functions of stomatal aperture depending on meteorological data, and (ii) from the synchronously measured SO₂ immission at these stands. The stomatal response functions had been derived on the basis of thorough stomatal water conductance measurements in the field. Calculations of the SO₂ conductance of spruce twigs and SO₂ uptake rates via stomata need continuously measured complete data sets of the (i) light intensity, (ii) air temperature, (iii) air humidity and (iv) SO₂ concentration in spruce forests from all the year. These data were recorded half hourly in different German spruce forests. The apparent needle water vapour pressure difference and transpiration rates were calculated from meteorological data. Additional use of canopy through flow data in dry years allowed the estimation of the mean stomatal conductance for H₂O and SO₂ of whole spruce canopies. The annual SO₂ uptake of a mean unit needle surface in spruce forests was 32% of the SO₂ uptake rate of exposed needles at the top of spruce crowns. There is significant SO₂ uptake all the year. The mean SO₂ dose at all sites and years received through the stomata was (0.25±0.07) mol SO₂ m^-2 (total needle surface) (nPa Pa^-1)^-1 (annual mean of SO₂ immission; 1 nPa (SO₂) Pa^-1 (air) = 1 ppb) day^-1 (vegetation period per year). Comparison of calculated SO₂ uptake rates into needles with measured SO₄ ^2- accumulation rates in needles from the mentioned sites and additionally from Würzburg, Schneeberg (Fichtelgebirge) and from three sites in the eastern Erzgebirge (Höckendorf, Kahleberg, Oberbärenburg) revealed that oxidative SO₂ detoxification (SO₄ ^2- formation) dominates only at sites with high SO₂ immission and short vegetation periods. Under these conditions 70 to 90% of the annual stomatal SO₂ uptake is detoxified via SO₄ ^2- accumulation in needles. Cations are needed for neutralization of accumulating SO₄ ^2- which are inavailable to support growth. Thus, SO₂ induces a dominant and competitive additional nutrient cation demand, cation deficiency symptoms and enhanced needle loss (spruce decline symptoms) mainly at sites, where the ratio R=(SO₂ immission): (length of the vegetation period) is higher than R=0.07 nPa Pa^-1 day^-1. Correlation analysis of the relative needle loss versus the SO₂-dependent SO₄ ^2- formation rate revealed a significant increase of needle loss at the 98% level (Student). At sites with small SO₂ immission and long vegetation periods (R<0.07 nPa Pa^-1 day^-1) reductive SO₂ detoxification via growth (and/or phloem export of SO₄ ^2-) is not kinetically overburdened. Under these conditions only 30% of the annual SO₂ uptake is detoxified via SO₄ ^2- formation and spruce decline is small or absent. On the basis of the critical value R0.07 nPa Pa^-1 day^-1 recommended SO₂ immission limits can be deduced on a mere ecophysiological basis. These deduced values are close to the proposed SO₂ immission limits of the IUFRO, WHO and the UNECE.     

Physiological effects of long term exposure to low concentrations of SO2 and NH3 on poplar leaves

Shoots of poplar (Populus euramericana L. cv. Flevo) were exposed to filtered air, SO₂, NH₃ or a mixture of SO₂ and NH₃ for 7 weeks in fumigation chambers. After this exposure gas exchange measurements were carried out using a leaf chamber. As compared to leaves exposed to filtered air, leaves pretreated with 112 μg m^?3 SO₂ showed a small reduction in maximum CO₂ assimilation rate (P_max) and stomatal conductance (g_s). They also showed a slightly higher quantum yield and dark respiration. In addition, the fluorescence measurements indicated that the Calvin cycle of the leaves pretreated with 112 μg m^?3 SO₂ was more rapidly activated after transition from dark to light. An exposure to 64 μg m^?3 NH₃ had a positive effect on P_max, stomatal conductance and NH₃ uptake of the leaves. This positive effect was counteracted by an SO₂ concentration of 45 μg m^?3. The exposure treatments appeared to have no effect on the relationship between net CO₂-assimilation and g_s. Also, no injury of the leaf cuticle or of epidermal cells was observed. Resistance analysis showed that NH₃ transfer into the leaf can be estimated from data on the boundary layer and stomatal resistance for H₂O transfer and NH₃ concentration at the leaf surface, irrespective of whether the leaves are exposed for a short or long time to NH₃ or to a mixture of NH₃ and SO₂. In contrast SO₂ uptake into the leaves was only partly correlated to the stomatal resistance. The results suggest a large additional uptake of this gas by the leaves. The possibility of a difference in path length between SO₂ and H₂O molecules is proposed.     

Detoxification of SO2 in conifers differing in SO2-tolerance

Contents of organic sulfur, sulfate and the inorganic cations K⁺, Ca²⁺, Mg²⁺, Mn²⁺ and Na⁺ were compared in needles of three conifer species differing in tolerance to chronic SO₂ immissions. Sulfate and organic sulfur compounds were also measured in bark and wood. Field material was collected from Norway Spruce (Picea abies (L.) Karst.), Colorado Spruce (Picea pungens Engelm.) and Scots Pine (Pinus sylvestris L.) at sites where the SO₂ concentration in air was high, and at another site where it was low. In general, sulfate contents were higher but cation contents lower at the sites where SO₂ concentrations were high than where they were low. Up to 114mmol · (kg DW)^–1 sulfate was measured in fouryear-old needles of Norway Spruce from the Erzgebirge (annual mean of SO₂ in air 32 nl · 1^–1). Sulfate accumulation in this SO₂-sensitive conifer increased with SO₂ concentration in ambient air and with needle age, indicating that the main part of the sulfate resulted from the oxidative detoxification of SO₂. Loss of inorganic cations from ageing needles was reduced, or cation levels even increased, with increasing needle age, while sulfate accumulated. Apparently, cations served as counter-ions for sulfate, which is sequestered in the vacuoles. Individual trees differed in regard to the nature of cations which accumulated with sulfate. Calcium, potassium and magnesium were the dominating cations. Sodium levels were very low. Needles of the SO₂-tolerant conifers Colorado Spruce and Scots Pine growing next to Norway Spruce in the Erzgebirge did not accumulate, or accumulated less, sulfate with increasing needle age as compared to needles of Norway Spruce. However, somewhat more sulfate was found in the bark of the SO₂-tolerant species than in the bark of Norway Spruce. Scots Pine contained distinctly more sulfate in the wood than the other conifers. Since accumulation of organic sulfur compounds could not be observed with increasing needle age, or in bark and wood, reduction does not appear to play a major role in the detoxification of SO₂ by the investigated species. Physiological mechanisms permitting Colorado Spruce and Scots Pine to avoid the sulfate accumulation in the needles and the accompanying sequestration of cations that are observed in neighbouring Norway Spruce are discussed on the basis of the obtained data.Abbreviations S_org organic sulfur compounds Died June 10, 1991, aged 29, in a traffic accident. He initiated this work.This work was supported by the Sonderforschungsbereich 251 of the University of Würzburg and by the Projektgruppe Bayern zur Erforschung der Wirkung von Umweltschadstoffen (PBWU). The authors with to thank Prof. Dr. W Kaiser and Prof. Dr. W. Urbach (both Julius-von-Sachs-Institut, University of Würzburg, Germany) for HPLC-analysis and ICP-analysis.