Plant Physiol. (1989) 91, 357-3610032-0889/89/91/0357/05/$01 .00/0

Received for publication January 19, 1989and in revised form April 28, 1989

Enhanced Ethylene Emissions from Red and NorwaySpruce Exposed to Acidic Mists

Yi-Min Chen and Alan R. Wellburn*Division of Biological Sciences, Institute of Environmental and Biological Sciences, University of Lancaster,

Bailrigg, Lancaster, LA 1 4YQ, United Kingdom

ABSTRACT

Acidic cloudwater is believed to cause needle injury and todecrease winter hardiness in conifers. During simulations of theseadverse conditions, rates of ethylene emissions from and levelsof 1-aminocyclopropane-1-carboxylic acid (ACC) in both red andNorway spruce needles increased as a result of treatment withacidic mists but amounts of 1-malonyl(amino)cyclopropane-1-carboxylic acid remained unchanged. However, release of signif-icant quantities of ethylene by another mechanism independentof ACC was also detected from brown needles. Application ofexogenous plant growth regulators such as auxin, kinetin, ab-scisic acid and gibberellic acid (each 0.1 millimolar) had noobvious effects on the rates of basal or stress ethylene productionfrom Norway spruce needles. The kinetics of ethylene formationby acidic mist-stressed needles suggest that there is no activeinhibitive mechanism in spruce to prevent stress ethylene beingreleased once ACC has been formed.

Environmental stresses, such as wounding, chilling, airpollution, drought, infection by fungi or insect attack, are allknown to induce the formation of stress ethylene. This gasmay be derived from methionine, via S-adenosylmethionineand l-aminocyclopropane-l-carboxylic acid (ACC') as shownbelow. Synthesis of ACC is the key regulatory step and 1-malonyl(amino)cyclopropane-l-carboxylic acid (MACC) isformed as a side reaction (2, 3, 36). During the wilting ofwheat leaves, for example, there is a sharp rise followed by adecline in the levels of endogenous ACC and the rates ofproduction of ethylene, although the levels ofMACC rise andremain high throughout wilting (9). Similar changes in therates of ethylene emission and levels ofACC and MACC havebeen reported in S02-fumigated wheat seedlings (18). Therapid decline ofACC levels and consequential fall in ethyleneproduction in both cases can be attributed, in part, to theefficient conjugation of ACC to MACC. Thus, the malony-lation of ACC has been suggested (9, 36) to be a regulative

EFE

Methionine -- SAM -.ACC -T-*C2H4e CO2. HCN . H20

C-; ------ 02 MACC

'Abbreviations: ACC, 1-aminocyclopropane-l-carboxylic acid;MACC, l-malonyl(amino)cyclopropane- 1-carboxylic acid; EFE, eth-ylene-forming enzyme.

mechanism to dissipate excess ACC and thereby regulateethylene biosynthesis.

Regulation of ethylene emission may also take place byinteraction with other plant growth regulators. In wilted wheatleaves, levels of ABA rapidly increase and these then mayinhibit production of stress ethylene (33). Similarly, IAA andcytokinins (BA) are known to enhance ethylene formation(24, 34, 35, 37).

Ethylene-forming enzyme (EFE, see reaction schemeabove) may also play a part in the regulation of stress ethylene.The increase of ethylene and ACC levels during fumigationwith sulphur dioxide or wilting of wheat leaves are accom-panied by an increase ofEFE (18, 24). In some circumstances,such as fruit ripening, activation of EFE may be an inhibitingfactor which limits the rate of ethylene production (10).Over the past two decades, effects of air pollutants such as

03, SO2, and heavy metals on the rates of ethylene productionhave been widely studied (1, 19, 23, 36). Many short-termexperiments have shown that after exposure ethylene produc-tion reaches a maximum in just a few hours and falls off tothe control levels within 1 d (23). Kimmerer and Kozlowski(13) observed that, in birch seedlings exposed to SO2, stressethylene formation continued until about 80% of the tissuesshowed injury while Fuhrer (8) reported that both ethyleneemissions and the levels ofACC and MACC rose in damagedfir needles.Exposure of conifers to acidic cloudwater and mists has

often been suspected of causing injury to needles and to affectthe hardening of needles during autumn. In a series of jointexperiments between the Institute of Terrestrial Ecology inScotland and the University of Lancaster, simulated acidicmistings have produced symptoms of injury and evidence ofreduced winter hardiness in red spruce (6). As part of thatstudy, we report data from red spruce trees which have beentreated with different acidic mistings in the autumn and earlywinter, during the period when chilling stress may also occur,and a similar treatment of Norway spruce in midwinter whentrees were fully hardened and most exposure to acidic-mistsoccurs in forests.

MATERIALS AND METHODS

Acidic Mist Treatments

Two-year-old red spruce (Picea rubens L.) seedlings wereexposed to a range of acidic mist treatments in open-topchambers, at the Institute of Terrestrial Ecology (ITE) at Bushin Scotland, which provide filtered air but exclude rainfall by

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means of a large translucent dish beneath the opening andabove the trees (6). Treatments with acidic mists were carriedout for 20 weeks from July to December 1987 by means of aspinning disc droplet generator, producing drops with a meansize of about 80 ,um and giving deposition rates of water inthe chambers of 3 mm h-1. Mist acidities ranged from pH 2.5to 5.0 and the application frequency was twice a week for 40min on each occasion. In a second experiment carried outbetween January and March 1988, 3-year-old Norway spruce(Picea albies L.) seedlings were exposed in the solardomefumigation systems at Lancaster (21) to similar acidic mistsonce a day for 10 weeks (approximately 80 mL tree-') gen-erated as previously described (31). The composition of theacidic mists applied to spruce included S02-, NO-, NH' andH+ from an equimolar mixture of HNO3 and (NH4)2SO4diluted to provide a pH 2.5 treatment equivalent to 21 kg Nha-' month-', a pH 3.0 treatment equal to 7 kg N ha-'month-' and a pH 5.0 control treatment of 0.07 kg N ha-'month-'.

Measurement of Ethylene and Ethane Emissions

Rates of release of hydrocarbons from shoots of red orNorway spruce were determined by the procedure describedby Wolfenden et al. (32).

Determination of ACC and MACC

Spruce needles were immediately frozen in liquid N2 andstored at -20°C. Batches of needles (0.2 g) were homogenizedwith 1.2 mL 2% (w/v) metaphosphate acid solution cooledby ice and, after centrifugation at 11,000g for 10 min, thesupernatants were directly used for determinations of ACCaccording to the method of Lizada and Yang (20). MACC insimilar extracts was measured by passing 0.25 mL of thesupernatant through an Amberlite resin (acid-form of IR- 120;2 mL bed volume) to remove the ACC and hydrolyzing theeffluents containing MACC with 2 M HCI for 3 h at 100°C.Following neutralization with NaOH, the acid hydrolysateswere then assayed for MACC like those ofACC beforehand.

Application of ACC, ABA, BA, IAA, and GA3

ACC and plant growth regulators (0.1 mm in 66 mm sodiumphosphate buffer) were incubated with detached needles for24 h in sealed glass bottles before ethylene determinations byGLC. The needles were recut below the surface of the incu-bation media beforehand to avoid problems with trapped airbubbles.

RESULTS

Ethylene, ACC, and MACC Changes

In the ITE experiment, red spruce needles exposed to pH3.0 and pH 2.5 mistings showed needle necrosis after treat-ment for 10 weeks. The overall extent of injury ranged from15% to 40% but the pH 5.0 control trees showed no damagewhatsoever. Significant increases in both the rates of ethyleneemission and the levels of ACC were also observed in themore acidic mist treatments (Fig. 1). At pH 2.5, rates of

WEEKSFigure 1. Weekly changes in ethylene emissions from and levels ofACC and MACC in red spruce needles treated with acidic mists for20 weeks. Results to the right of the vertical dashed line wereobtained after misting had been stopped. Each value shown in themean of 10 determinations. The error bars above each represent thestandard errors of the means and the astersks show the relativesignificances from the controls as revealed by Student's t-tests where* = P<0.05and ** = P<0.01.

ethylene production were about 5 to 10 times those of the pH5.0 treatment but levels of MACC throughout were littlechanged. These results indicate that stress ethylene emissionsfrom red spruce needles are induced by acidic mist treatmentsand sustained for a long period by means of high levels ofACC.Acid misting of red spruce during this experiment was

stopped on December 18, 1987, but the same seedlings weresampled twice more, 8 and 19 weeks after this treatment.Since rates of ethylene production can be influenced byenvironmental factors, the ratio between control (pH 5.0) andstressed (pH 2.5) is a means of expressing comparativechanges. Before the termination of misting, rates of ethyleneemissions from and levels of ACC in pH 2.5-treated needleswere 10.2- and 4.5-fold, respectively, those of the pH 5.0treatment. Even 4 months after the misting stopped, both ofthem were still about 2.5-fold greater.

In order to confirm some of the observations in red spruce,Norway spruce seedlings were also subjected to acidic misttreatments. Trends of ethylene production and changes inlevels of ACC and MACC (Fig. 2) were very similar to thosealready found in red spruce (cf Figs. 1 and 2). Visible symp-toms, enhanced rates of ethylene production and levels ofACC rose in stressed needles between the 5th and 7th weeksoftreatment and were sustained to the 10th week. Once again,MACC levels changed little during the entire period.

Changes in Needles with Different Degrees of InjuryIn acidic mist-treated spruce seedlings, needles could be

visually sorted into three classes by color representing the

CHEN AND WELLBURN358

ETHYLENE FROM ACIDIC MIST-TREATED SPRUCE

1 3 6 7 10

WEEKS

Figure 2. Weekly changes in ethylene emissions from and levels ofACC and MACC in Norway spruce needles treated with acidic mistsfor 10 weeks. Each value shown is the mean of 6 determinations.The error bars above each represent the standard errors of themeans and the asterisks show the relative significances from thecontrols as revealed by Student's t-tests where * = P < 0.05 and **

= P<0.01.

Table I. Rates of Ethylene or Ethane Emissions, with Levels of ACC,Associated with Spruce Needles with Different Degrees of Injury

Species,Needle Color, Ethylene Ethane ACCand Misting

nL g-' dry wt d-' nmol g' dry wt

Norway spruceGreen (pH 5.0) 12.4 ± 0.8a ND 1.4 ± 0.2Green (pH 2.5) 62.3 ± 5.5 ND 5.7 + 1.2Green/brown (pH 2.5) 189 ± 34.8 17.5 ± 2.1 13.3 ± 2.8Brown (pH 2.5) 96.9 ± 19.8 13.4 ± 1.3 22.1 ±2.0

Red spruceGreen (pH 5.0) 2.5 ± 0.2 ND 1.1 ±0.2Green (pH 2.5) 18.7 ± 1.6 ND 3.8 ± 0.4Green/brown (pH 2.5) 25.7 ± 3.0 5.6 ± 0.9 5.9 ± 0.5Brown (pH 2.5) 10.0 ± 1.5 2.1 ± 0.5 32.9 ± 5.6

a Means (± SE) taken from six replications each. ND, not detected.

extent of different degrees of injury-some all green, some allbrown, and many intermediate green-brown forms. Table Ishows the rates of ethylene and ethane releases from, andlevels of ACC in, these various selected groups of red andNorway spruce needles compared with those ofpH 5.0-treatedall green needles. Although the rates of ethylene emissionsfrom and levels of ACC in acidic mist-exposed (pH 2.5) are

all higher than those of the pH 5.0-treated needles, when theability to convert exogenous ACC to ethylene was tested in

the two different groups of needles (Table II) green needlesshowed high rates ofethylene formation from ACC but brownneedles had totally lost their EFE activity. Ethane productionby all pH 2.5-treated brown needles was significant but ratherless was emitted by those from red spruce by comparison totheir Norway spruce counterparts (Table I).

Effect of Exogenous Plant Growth Regulators

When excised Norway spruce needles were incubated withIAA, kinetin (BA), GA3, and ABA (0.1 mm each) for 24 h, noobvious enhancements or inhibitions were observed duringany treatment (Table III).

DISCUSSION

Most studies of the effects of air pollution on stress ethyleneformation by plants have used non-woody plants and short-term fumigations. During these experiments, the kinetics ofethylene production have shown a similar pattern when eth-ylene emissions have reached a maximum within a few hoursand then fallen back to their original level a day later (25, 27-30).

In hardwoods, a second pattern of stress ethylene evolutionhas been demonstrated. Bucher (4) reported a cyclical patternof stress ethylene production in forest trees exposed to S02while Kimmerer and Kozlowski (13) observed that, in birchseedlings exposed to SO2, ethylene formation continued untilabout 80% of the tissues were injured. Ethylene productionthen rapidly declined. This decrease was also observed in 03-

treated trees and shrubs showing a high degree of necrosis(17).Our results from acidic misting show a different behaviour

Table II. Ability to Convert Exogenous ACC to Ethylene in NorwaySpruce Needles

Ethylene EmissionsNeedle Colorand Misting - ACC + ACC

(0.1 mm)nL g-1 dry wt d-'

Green (pH 5.0) 24.4 ± 6.1a 87.1 ± 14.7Green (pH 2.5) 71.5 ± 8.2 402.0 ± 15.0Brown (pH 2.5) 181.0 ± 8.3 161.8 ± 6.0

a Means (± SE) taken from three replicates each.

Table IlIl. Effects of Plant Growth Regulators on Ethylene Emissionsfrom Norway Spruce Needles

Hormone Green (pH 5.0) Green (pH 2.5)Treatment

(±)a +Hormone -Hormone +Hormone -Hormone

nL g' dry wt d'ABA 11.6+3.3b 9.9± 1.8 100.4± 19.0 94.1 ± 19.3BA 17.8 2.3 14.7 ± 2.5 86.9 ± 2.1 79.6 ± 6.3GA3 10.4 2.0 12.7 ± 0.2 92.7 ± 6.3 96.9 ± 7.9IAA 9.0 1.5 12.7 ± 0.2 104.2 ± 16.3 96.9 ± 7.9

a At 0.1 mm of each in 66 mm phosphate buffer (blanks-bufferonly). b Means (± SE) taken from three replicates each.

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from both of these. With such a treatment of spruce, stressethylene is continuously emitted even when the needles be-come completely brown. As the damage on the stressedneedles increases, levels of ACC and activities of EFE gradu-ally rise but the levels of MACC are unchanged. Finally, asthe needles become brown, they lose their capacity to convertACC to ethylene but the evolution is maintained by largeemissions of ethylene apparently produced independently ofACC.

Patterns of ethylene evolution may depend on the plantspecies, the nature of the stress, and the physiological basis ofactive regulation. In wheat seedlings exposed to S02 (0.7 and1.3 ppm) for 9 h, for example, ethylene emissions and ACClevels reach a peak between 6 and 9 h before declining to theoriginal levels at 40 h afterward (18). By contrast, MACClevels rise steadily; noticeably so after 3 h of exposure. Pre-treatment with exogenous 6-BA enhances the rates ofethylenerelease and the levels ofACC in fumigated leaves but has littleeffects on this overall pattern of response (18). This indicatesthat, in wheat leaves at least, MACC accumulation is animportant aspect of regulation which inhibits excess forma-tion ofACC and reduces the release of stress ethylene inducedby the SO2 treatment.Under various environmental conditions, reasons for the

changes in rates of ethylene formation may be different (23).Needles of fir trees growing in stands affected by forest decline,for example, show elevated levels of MACC as well as ACCand enhanced rates of ethylene release (8), but little is knownabout the kinetics of such changes. In our acidic mist-stressedspruce needles, despite high rates ofethylene release and levelsof ACC being maintained for long periods, no simultaneousaccumulations of MACC took place. This implies that regu-lation of stress ethylene formation by means of a bypass toMACC does not take place in spruce needles.

Effects ofother plant hormones may be an additional aspectof such regulation. In plants fumigated with 03 and NO2,endogenous levels of ABA rise (14, 15) while applications ofexogenous ABA can efficiently inhibit ethylene emissionsfrom 03-stressed rice seedlings (1 1). In water-stressed wheatleaves, ABA plays an important role in inhibiting ethyleneproduction (33, 34). Similarly, pretreatment with cytokinin(BA) enhances SO2 or drought-induced ethylene emissionsfrom wheat plants (18, 35). However, in our acidic mist-stressed Norway spruce needles, all exogenously applied plantgrowth substances had little influence on ethylene production(Table III) under the experimental conditions we used. Eitherthey failed to reach their site of action or, if they did, theyhad no subsequent effect.

After the acidic mist treatment stopped, rates of ethyleneproduction from and ACC levels in red spruce needles de-clined slowly (Fig. 1). Even 16 weeks afterward, ethyleneemissions and ACC levels were still higher in those needleswhich had been subjected to acidic mists ofpH 2.5. This slowdecline of ethylene release may be entirely due to the slowdecrease ofACC synthesis after the withdrawal ofthe imposedacidity. It appears that there is no active inhibition of stressethylene production in spruce needles once ACC has beenformed in the needles. Though the data support the fact thatACC is the usual precursor of ethylene from the stressed green

needles, other experiments, using Norway spruce needles,have shown that aminoethoxyvinylglycine, a strong inhibitorof ACC synthase, cannot inhibit stress ethylene formationexperimentally induced by bisulfite or ferric sulfate (our un-published results). This implies that the regulation ofethyleneformation by spruce needles is different from that in non-woody plants such as wheat even though aminoethoxyvinyl-glycine penetrates needles as well as softer leaf tissues (datanot shown).Many treatments, such as high temperature, osmotic shock,

homogenization, and detergents, which destroy the mem-brane structure, result in a loss ofEFE activity in plant tissue(36). This may indicate that such activity is membrane-bound.Other reports have shown that ethylene formation declines inbadly stressed tissues and, at the same time, large amounts ofethane are produced (13, 25). Our results support these find-ings. As Tables I and II show, EFE activity is totally lost asACC accumulates in brown needles.

Release of ethane is often interpreted as indicative of (lipid)peroxidation taking place within necrotic tissues (7, 12, 25,26). The rates of ethane we measured from brown needles(Table I) were about one tenth to one fifth of the rates ofethylene emissions not derived from ACC. It is to be expectedthat such ACC-independent ethylene releases are derived fromperoxidative breakdown but the precise characteristics of thisprocess remains unclear. Evidence using model in vitro sys-tems suggests that both ethane and ethylene may be producedfrom the oxidation of linolenate (12, 16). Our recent studieshave revealed that acid-stressed brown spruce needles providea highly suitable, reproducible and convenient experimentalsystem to study the details of this ACC-independent ethyleneproduction and to test this and other possibilities.

Since the production of stress ethylene is often correlatedwith visible leaf injuries, it has often been suggested as anindicator of the extent of injury from a wide range of envi-ronmental stress. Bucher (4), however, suggested that therelease of stress-ethylene may serve only as an indicator ofstress in controlled experiments, but could not recommend itfor use under field conditions. The kinetics of ethylene pro-duction are different among plant species and under differentstress conditions. Only when rates of ethylene emissions arehigh and continuous, as we have found in red and Norwayspruce exposed to acidic mists, are they suitable for diagnosticpurposes in the field. Details examination of the mass of datafrom the CEC 1986 Forest Decline Survey acrossNW Europe(5) revealed that, of the 30 different variables measured onsamples of Norway spruce taken from 12 sites on a transectfrom N. Scotland to S. Germany, only 3 parameters werefound to be site-independent and suitable for use as a predic-tors of the probability of forest decline (22). One of these wasthe measurement of the rates of ethylene emission.

ACKNOWLEDGMENTS

We are grateful for all those at the ITE in Scotland, particularlyDrs. Neil Cape and David Fowler, for all their help with these studies,and for financial support from the Commission of the EuropeanCommunity (CEC, DG XII), the U.K. Overseas DevelopmentScheme and the Department of the Environment, and the North-eastern Forest Experiment Station's Spruce-Fir Research Cooperative

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ETHYLENE FROM ACIDIC MIST-TREATED SPRUCE

Forest Response Program, which is part of the U.S. National AcidPrecipitation Assessment Program (NAPAP). This paper has not beensubject to EPA or Forest Service peer review and should not beconstrued to represent the policies of either Agency.

LITERATURE CITED

1. Abeles FB (1973) Ethylene in Plant Biology. New York, Aca-demic Press, p 302

2. Adams DO, Yang SF (1977) Methionine metabolism in appletissue; implications of S-adenosylmethionine as an intermedi-ate in the conversion of methionine to ethylene. Plant Physiol60: 892-896

3. Adams DO, Yang SF (1979) Ethylene biosynthesis: identificationof 1-amino cyclopropane- -carboxylic acid as an intermediatein the conversion of methionine to ethylene. Proc Natl AcadSci USA 76: 170-174

4. Bucher JB (1981) S02-Induced ethylene evolution of forest treefoliage and its potential use as an indicator. Eur J For Pathol11: 369-373

5. Cape JN, Paterson IS, Wellburn AR, Wolfenden J, MehlhornH, Freer-Smith PH, Fink S (1988) Early Diagnosis of ForestDecline: A Report. NERC Institute of Terrestrial Ecology,Merlewood, Grange-over-Sands, UK

6. Cape JN, Sheppard LJ, Leith ID, Murray MB, Deans JD,Fowler D (1988) The effect of acid mist on the frost hardinesson red spruce seedlings. Aspects Appl Ecol 17: 141-149

7. Dumelin EE, Tappel AL (1977) Hydrocarbon gases producedduring in vitro peroxidation of polyunsaturated fatty acids anddecomposition of performed hydroperoxides. Lipids 12: 894-900

8. Fuhrer J (1985) Ethylene production and premature senescenceof needles from fir trees (Abies alba). Eur J For Pathol 15:227-236

9. Hoffman NE, Liu Y, Yang SF (1983) Changes in l-(malonylam-ino)cyclopropane-l-carboxylic acid content in wilted wheatleaves in relation to their ethylene production rates and 1-aminocyclopropane- 1 -carboxylic acid content. Planta 157:518-523

10. Hoffman NE, Yang SF (1980) Changes of l-aminocyclopropane-1-carboxylic acid content in ripening fruits in relation to theirethylene production rate. J Am Soc Hortic Sci 105: 492-495

11. Jeong YH, Nakamura H, Ota Y (1981) Physiological studies onphotochemical oxidant injury to rice plants. II. Effect of ab-scisic acid on ozone injury and ethylene production in riceplants. Nippon Sakumotsu Gakkai Kiji 50: 560-565

12. John WW, Curtis RW (1977) Isolation and identification of theprecursor of ethane in Phaseolus vulgaris L. Plant Physiol 59:521-522

13. Kimmerer TW, Kozlowski TI (1982) Ethylene, ethane, acetal-dehyde, and ethanol production by plants under stress. PlantPhysiol 69: 840-848

14. Kondo N, Maruta I, Oikawa T, Sugahara K (1981) Changes intranspiration rate and increase in stomatal sensitivity of ab-scisic acid in plants after fumigation with ozone and nitrogendioxide. Kokuritsu Kogai Kenkyusho Kenkyo Hokoku 28:23-30

15. Kondo N, Sugahara K (1984) Effects of air pollutants on tran-spiration rate in relation to abscisic acid content. KokuritsuKogai Kenkyusho Kenkyo Hokoku 65: 1-8

16. Konze JR, Elstner EF (1978) Ethane and ethylene formation bymitochondria as indicators of aerobic lipid degradation in

response to wounding of plant tissue. Biochim Biophys Acta528: 213-221

17. Li L (1984) Exposure of trees to ozone and hydrofluoric acidand stress release of ethylene. Linye Kexue 20: 359-365

18. Li Z, Ji Y, Yu Z, Tang C, Yu S (1986) Effects of sulfur dioxideexposure on contents of 1-aminocyclopropane-l-carboxylicacid and l-(malonylamino)cyclopropane-l-carboxylic acid inwheat leaves in relation to ethylene production. Acta Phyto-physiol Sin 12: 315-323

19. Lieberman M (1979) Biosynthesis and action of ethylene. AnnuRev Plant Physiol 30: 533-591

20. Lizada MCC, Yang SF (1979) A simple and sensitive assay forI-aminocyclopropane-l-carboxylic acid. Anal Biochem 100:140-145

21. Lucas PW, Cottam DA, Mansfield TA (1987) A large scalefumigation system for investigating interaction between airpollution and cold stress on plants. Environ Pollut 43: 327-329

22. Mehlhorn H, Francis BJ, Wellburn AR (1989) Prediction of theprobability of forest decline damage to Norway spruce usingthree simple site-independent diagnostic parameters. New Phy-tol 110: 525-534

23. Meyer A, Muller P, Sembdner G (1987) Air pollution and planthormones. Biochem Physiol Pflanzen 182: 1-21

24. McKeon TA, Hoffman NE, Yang SF (1982) The effect of planthormone pretreatments on ethylene production and synthesisof 1 -aminocyclopropane- 1 -carboxylic acid in water-stressedwheat leaves. Planta 155: 437-443

25. Peiser GD, Yang SF (1979) Ethylene and ethane productionfrom sulfur dioxide-injured plants. Plant Physiol 63: 142-145

26. Riley CA, Cohen G, Lieberman M (1974) Ethane evolution: anew index of lipid peroxidation. Science 183: 208-210

27. Rodecap KD, Tingey DT, Tibbs JH (1981) Cadmium-inducedethylene production in bean plants. Z Pflanzenphysiol 105:65-74

28. Tingey DT (1980) Stress ethylene production-a measure ofplant response to stress. Hortic Sci 15: 630-633

29. Tingey DT, Pettit N, Bard J (1978) Effect of chlorine on stressethylene production. Environ Exp Bot 18: 61-66

30. Tingey DT, Standley C, Field FW (1976) Stress ethylene evolu-tion: a measure of ozone effects on plants. Atmos Environ 10:969-974

31. Wolfenden J, Wellburn AR (1986) Cellular readjustment ofbarley seedlings to simulated acid rain. New Phytol 104: 97-109

32. Wolfenden J, Robinson DC, Cape JN, Paterson IS, Francis BJ,Mehlhorn H, Wellburn AR (1988) Use of carotenoid ratios,ethylene emissions and buffer capacities for the early diagnosisof forest decline. New Phytol 109: 85-95

33. Wright STC (1977) The relationship between leafwater potential({ leaf) and the levels of abscidic acid and ethylene in excisedwheat leaves. Planta 134: 183-189

34. Wright STC (1979) The effect of 6-benzyladenine and leafageingtreatment on the levels of stress-induced ethylene emanatingfrom wilted wheat leaves. Planta 144: 179-188

35. Wright STC (1980) The effect of plant growth regulator treat-ments on the levels of ethylene emanating from excised turgidand wilted wheat leaves. Planta 148: 381-388

36. Yang SF, Hoffman NE (1984) Ethylene biosynthesis and itsregulation in higher plants. Ann Rev Plant Physiol 35: 155-189

37. Yu YB, Yang SF (1979) Auxin-induced ethylene production andits inhibition by aminoethoxyvinylglycine and cobalt ion. PlantPhysiol 66: 281-285

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