Experimental Studies of the Factors Controlling Transpiration: I. APPARATUS AND EXPERIMENTAL TECHNIQUE

Apparatus and experimental techniques are discussed for usein the investigation of transpiration rate of leaves as controlledby stomatal aperture and leaf water content. The leaf chambers used and the methods adopted for the estimationof the water transpired are described. The designs of the porometer cups used for the different typesof leaves (Pelargonium and wheat) employed in the work are described.To obviate the difficulty that stomata within the cup behaveabnormally, the design employed makes possible a removal ofthe cup from the leaf except during the short periods requiredto estimate stomatal resistance to air flow at intervals duringthe course of an experiment. In these experiments the water content of the leaf is changedat will by interrupting the water-supply and re-establishingit to permit recovery from wilting. The methods used to carryout this cycle of operations are fully dealt with. Determinationsof the transpiration and absorption rates during the experimentand of the final leaf water content make it possible to followchanges in leaf water content throughout the experiment. An account is given of the methods used for varying the speedof flow, the humidity, and the CO₂ concentration of the airstreams.     

The uptake and release of water vapour by the foam seal of a diffusion porometer as a source of bias

Abstract. The apparent resistance to diffusion of water vapour presented by a set of holes in a calibration plate when measured by a commercial porometer depended on materials previously measured in the sensor head of the instrument. The data reported show the variation in apparent resistance of a set of holes with a specified value of 13.7 s cm⁻¹ after a leaf of small resistance, a set of holes with a specified resistance of 3.75 s cm⁻¹, or a hydrophobic membrane had been in the sensor head and the effect of covering the foam seal around the mouth of the cup with aluminium foil. It was concluded that the results were profoundly affected by the history of water vapour accumulation and release from the foam seal which could give rise to biased readings both during calibration and in measurements of leaf resistance and might also explain much of the scatter when calibration data obtained at different times were pooled for the calculation of a calibration equation. Reliable estimates of resistance over the effective range of the instrument cannot be expected unless this transfer of water vapour between the seal and the cup is prevented.     

The effect of temperature differences between porometer head and leaf surface on stomatal conductance measurements

It is generally known that instantaneous values of leaf conductance as measured with a dynamic porometer need to be corrected for the temperature difference, ΔT, between the porometer cup and the sampled leaf. Leaf conductances, obtained with a Delta-T AP4 dynamic porometer, with and without correction for ΔT are compared for a bush species (Guiera senegalensis) and two forb species (Jacquemontia tamnifolia and Mitracarpus scaber). With temperature differences predominantly varying within the ±2.5 °C recommended by the manufacturer, it appears that the differences between uncorrected and corrected conductances are very large, up to 100% on average, especially for the two forbs. Furthermore, it is shown that, using the Mitracarpus data, a relatively small error of ±0.5 °C in ΔT can cause a difference of 25–50% in the final conductance value, in particular for the high conductance range. An error of ±0.5 °C may easily occur: the accuracy of ΔT as measured by the thermistors in the porometer is 0.2 °C and the temperature variation within a leaf can be much larger. This result will have implications for upscaling of leaf conductances to canopy values or may explain why upscaled values appear not to correspond with down-scaled values, obtained from eddy correlation measurements and an inverted canopy transpiration model.     

Studies in Stomatal Behaviour: V. THE ROLE OF CARBON DIOXIDE IN THE LIGHT RESPONSE OF STOMATA

It was found that stomata on illuminated leaves, both of Pelargoniumand wheat, opened much wider where the leaf surface was enclosedin a small volume of air, as in a normal porometer cup, thanelsewhere. This was shown for both species by the infiltrationmethod, and for Pelargonium by Lloyd's method and direct microscopicalobservation also. The effect was shown not to be due to pressure of the porometercup or glass plate on the leaf, or to temperature differences,nor directly to the lack of movement or high humidity of theenclosed air. A considerable body of data was collected which appeared tosupport the hypothesis that the wide opening was due to accumulationof some volatile substance produced by the leaf, but all theresults were also consistent with the view that it was causedby reduction in the carbon dioxide content of the enclosed airbelow the normal 0·03 per cent. owing to photosynthesis.Further crucial experiments with both the porometer and infiltrationmethods left virtually no doubt that the latter hypothesis wascorrect. This extreme sensitivity of stomata to carbon dioxide concentrationwithin the range 0·03 per cent. to zero is discussedin relation to their operation in nature, and a possible biologicaladvantage is suggested. The bearing of the effect upon porometer investigations is alsodiscussed and it is concluded that for all quantitative or semi-quantitativeexperimentation it is essential to use a cup detached betweenreadings, or at least swept with air such as surrounds the restof the leaf, and to have the upper leaf surface above the cuparea freely exposed or similarly swept. For qualitative investigationof the light response of stomata the traditional form of cupmay be used. The importance is stressed of allowing porometer readings toreach equilibrium under one set of conditions before changingto another, when investigating the ‘closing’ or‘opening’ effects of external factors. Several subsidiary effects, observed in the course of the investigation,are discussed; in particular an effect of humidity upon therate of response to other factors.     

The Measurement of Stomatal Responses to Stimuli in Leaves and Leaf Discs

A comparison has been made of stomatal responses in intact leaves,leaf discs supplied with water via their cut edges and leafdiscs floating on water. Xanthium pennsylvanicum leaf discswatered via their cut edges appeared to be more turgid thanintact leaves; this considerably slowed down the rate of stomatalopening but it slightly increased the final steady-state stomatalopening. When the water potential of such leaf discs was loweredby pre-treatment with mannitol solutions rates of stomatal openingincreased whereas maximum steady-state openings decreased. In tobacco leaf discs floating on water the stomata in contactwith water were wider open than those in contact with normalair and they did not respond to treatment with carbon dioxide-freeair. The rate of photosynthesis was severely reduced in tobaccoleaf discs floating with the lower epidermis on water, mostprobably owing to the slow rate of diffusion of carbon dioxidein water. By floating such discs on osmotica the degree of stomatalopening was increased, however, a response to treatment withcarbon dioxide-free air was still not measurable. It is postulatedthat, on account of the relative unavailability of carbon dioxidefrom the water, the carbon dioxide concentration in the substomatalcavities of the lower surface is abnormally low, irrespectiveof whether ordinary air or carbon dioxide-free air is availableto the upper surface. A comparison between porometer readings and measurements ofsiliconerubber impressions of stomatal pores taken from insidethe porometer cup confirmed that the silicone-rubber impressionmethod of assessing stomatal responses to stimuli has severelimitations, especially at small stomatal apertures.     

Transit Time Diffusion Porometer Calibration: An Analysis taking into Account Temperature Differences and Calibration Non-Linearity

A method is described that determines the mean calibration ofa transit time diffusion porometer from a limited amount ofcalibration data. The calibration includes calculation of thetemperature coefficients over the range encountered. The commonassumption of a linear relation between transit time and leafresistance was found to give large errors, particularly at lowresistances, and a method of fitting the relation found is described.The improvement in fit is shown to improve significantly theaccuracy of estimates of changes in leaf conductance. Key words: Porometer, Stomata     

A Porometer for Laboratory and Field Operation

A porometer is described which is easily and rapidly attachedto plant leaves and appears to be a versatile and sensitiveinstrument for laboratory or field measurement of leaf resistanceto air flow. In the laboratory it can be connected to a normalresistance porometer circuit, and in the field to a sphygrnomanometer.A theoretical relationship has been developed and tested whichenables the conversion of leaf resistance obtained with a sphygmomanometerto that obtained with a normal porometer circuit.     

The Diffusion Resistance and Water Status of Leaves of Beta vulgaris

Pot-grown sugar-beet plants were deprived of water in two experimentsuntil they wilted completely. Leaf resistance to water-vapourtransfer was measured directly with a diffusion porometer andcompared with values calculated from measurements of transpiration.There was good agreement between the two estimates. Leaf turgor-pressuredecreased only gradually due, apparently, to an increase intissue-solute concentration with increasing water deficit. Relationshipsbetween leaf water potential, soil water potential, and leafresistance were established and the effective control of transpirationby small changes in leaf resistance was clearly demonstrated.     

Design calibration and field use of a stomatal diffusion porometer

Modifications of the design and calibration procedure of a diffusion porometer permit determinations of stomatal resistance which agree well with results obtained by leaf energy balance. The energy balance and the diffusion porometer measurements indicate that the boundary layer resistances of leaves in the field are substantially less than those predicted from heat transport formulas based on wind flow and leaf size.     

Estimating stomatal conductance with thermal imagery

Most thermal methods for the study of drought responses in plant leaves are based on the calculation of 'stress indices'. This paper proposes and compares three main extensions of these for the direct estimation of absolute values of stomatal conductance to water vapour (gs) using infrared thermography (IRT). All methods use the measured leaf temperature and two environmental variables (air temperature and boundary layer resistance) as input. Additional variables required, depending on the method, are the temperatures of wet and dry reference surfaces, net radiation and relative humidity. The methods were compared using measured gs data from a vineyard in Southern Portugal. The errors in thermal estimates of conductance were of the same order as the measurement errors using a porometer. Observed variability was also compared with theoretical estimates of errors in estimated gs determined on the basis of the errors in the input variables (leaf temperature, boundary layer resistance, net radiation) and the partial derivatives of the energy balance equations used for the gs calculations. The full energy balance approach requires accurate estimates of net radiation absorbed, which may not be readily available in field conditions, so alternatives using reference surfaces are shown to have advantages. A new approach using a dry reference leaf is particularly robust and recommended for those studies where the specific advantages of thermal imagery, including its non-contact nature and its ability to sample large numbers of leaves, are most apparent. Although the results suggest that estimates of the absolute magnitude of gs are somewhat subjective, depending on the skill of the experimenter at selecting evenly exposed leaves, relative treatment differences in conductance are sensitively detected by different experimenters.     

Experimental Studies of the Factors Controlling Transpiration: II. THE RELATION BETWEEN TRANSPIRATION RATE AND LEAF WATER CONTENT

Data are presented which show, when stomatal control is eliminated,that wheat leaves may lose 5–6 per cent. and Pelargoniumleaves 10–12 per cent. of their water without any reductionin the transpiration rate. Experiments in which Pelargonium and wheat leaves, with stomatalcontrol present, were submitted to cycles of changing watercontent also failed to establish any direct relation betweentranspiration rate and leaf water content. It is concluded that leaf water content over the range of 70–100per cent. of that present in the turgid state has no significanteffect in determining the rate of water loss from leaves. A repetition of Knight's experiment showed that stomata openedin still air and closed in moving air. This was not recordedby Knight, who used a porometer cup permanently attached tothe leaf. It is concluded that the higher transpiration raterecorded by Knight after a period of still air was due to widerstomatal aperture and not to the higher leaf water content assuggested by him.     

Photosynthesis, Transpiration, Leaf Stomatal and Mesophyll Resistance Measurements by the Use of a Ventilated Diffusion Porometer

A ventilated diffusion porometer was modified and adapted for simultaneous measurements of leaf resistance and photosynthesis (using ¹⁴C). The system enables measurements to be made under field and laboratory conditions with different concentrations of CO₂ and vapor pressure gradients between the evaporating surfaces inside the leaf and the external atmosphere. The leaf is subjected to the porometer's atmosphere only for short periods (up to 30 seconds) and it is assumed that stomata are not affected. Establishing the linear regression of the effect of CO₂ concentration on net photosynthesis makes it possible to extrapolate for CO₂ compensation point, to calculate the overall resistance to CO₂ and the mesophyll resistance to CO₂.     

A Krypton Diffusion Porometer for the Direct Field Measurement of Stomatal Resistance

A stomatal diffusion porometer is described which measures directlythe diffusion of radioactive krypton through amphistomatousleaves. The porometer is relatively small and portable and iseasily used under field conditions. It consists of a miniaturediffusion chamber above an acrylic plastic reservoir which contains1200 cm³ of air enriched with ⁸⁵Kr. Geiger tubes in the diffusionchamber and in the reservoir monitor the relative concentrationsof 85Kr. Krypton is allowed to diffuse from the larger reservoirthrough the leaf into the diffusion chamber and the time forits concentration in relation to that in the reservoir to changebetween two fixed values is recorded. When this time lapse wascalibrated against known resistances a linear relationship,independent of temperature was found. Sources of error are analysedand some experiments are described in which the porometer wasused to measure diurnal changes in stomatal resistance. Resistancesof potted sunflower plants (Helianthus annuus L.) grown in agreenhouse were measured with both the krypton diffusion porometerand a condensation-type porometer and the results were usedto calculate both cuticular and stomatal resistances. Demonstrationof field measurements with the porometer include data from eucalyptustrees {Eucalyptus camaldulensis Dehn) and from an unirrigatedcotton crop {Gossypium hirsutum var. SJ 2) growing under semi-aridconditions. Stomatal conductance of the cotton crop during theopening phase was linearly related to solar radiation.     

Changes in Leaf Viscous Flow Resistance Following Excision, Measured with a New Porometer

The viscous flow rate of air through leaves was measured witha sensitive, rapidly responding, bulk flow porometer during,and immediately following, leaf excision. In all cases excisionwas followed immediately by a rapid transient fall in viscousflow resistance. Following this initial response, the courseof subsequent changes in viscous flow resistance varied considerably,and it was concluded that this variation reflected differencesin the initial leaf water status.     

Dark opening of stomata in successional trees

Summary Dark opening of stomata was found in shade intolerant tree species in three different sets of experiments. In the field, leaves of eight successional species were darkened in light-proof bags and leaf resistance measured with a diffusion porometer. White ash, quaking aspen and American beech were sampled more intensively using darkened cuvettes and a dew point hygrometer. In the lab, white ash seedlings were kept in constant darkness and temperature, and their leaves also monitored with a cuvette and hygrometer. The stomata of all sampled shade intolerant species showed a dark opening response, often to leaf resistance levels typical of illuminated leaves. Shade tolerant species did not. Evidence suggests that this response is endogenous and could contribute to the morning opening of stomata of species under moderate water stress growing in well illuminated habitats.     

冬小麦近轴和远轴叶面气孔对土壤水分胁迫反应的敏感性

当根层土壤水分含量不足,作物体内出现水分胁迫时,小麦叶片两面气孔的反应有明显差异。远轴叶面气孔对水分胁迫的反应比近轴叶面气孔敏感。当出现水分胁迫时,远轴叶面气孔首先收缩,且收缩的程度比近轴叶面气孔大。远轴与近轴叶面气孔阻力的比值(r_(ab)/r_(ab))与根层平均土壤水势(Ψ_s)有关,当Ψ_s大于-50 kPa时,r_(ab)/r_(ad)基本稳定在1.5左右,而当Ψ_s小于-50 kPa时,r_(ab )/r_(ab)随Ψ_s降低而明显增加。     

DIFFUSION RESISTANCE IN LEAVES AS RELATED TO THEIR STOMATAL ANATOMY AND MICRO-STRUCTURE

Transpiration from a plant leaf depends upon the water vapor pressure gradient between the substomatal cavity and the free air beyond the leaf. Transpiration also depends inversely on the resistance of the diffusion pathway through the substomatal cavity, stomate, and surface boundary layer. The value of the diffusion resistance is derived mathematically for Zebrina pendula, Medicago sativa, and Pinus resinosa. The vapor pressure gradient depends on the leaf temperature and therefore is related to the energy budget of the leaf. The exact solution of the diffusion equation is described and limiting examples discussed. The so-called “diameter law” is a special case which is distinctly limited in its application.     

Experimental Studies of the Factors Controlling Transpiration: III. THE INTERRELATIONS BETWEEN TRANSPIRATION RATE, STOMATAL MOVEMENT, AND LEAF-WATER CONTENT

Transpiration rates of single leaves of Pelargonium and wheatwere measured under constant conditions of light, temperature,and air flow. Concurrently, stomatal movement was followed withthe resistance porometer during cycles of changing water contentof the leaf and changes induced by light and darkness. Stomatalmovement was found to exert a large controlling influence onthe transpiration rate, whereas water content had an extremelysmall or negligible effect. An approximately inverse linearrelation between transpiration rate and logarithm of resistanceto viscous flow through the leaf is believed to be the resultantof an inverse curvilinear relationship between the diffusiveconductance of the stomata and log. leaf resistance and thedecreasing difference of vapour pressure arising from the highertranspiration rates with increasing stomatal conductances. Nevertheless,the relation demonstrates that the transpiration rate is influencedby the degree of stomatal opening throughout its entire range. There was some evidence of lower transpiration rates duringand after recovery from wilting than before wilting. This isattributed to a decrease in a cell-wall conductance, the evaporatingsurface being located within the cell wall. During wilting partiallyirreversible contraction of the cell wall occurs. There wasalso evidence of slow changes in cell volume at full turgidityattributable to plastic flow. These occurred when the leaf wastransferred from environments of a high to low potential forevaporation. Extensive movement of the stomata followed changes in leaf water,passive opening resulting from decrease and closure from increaseof leaf water. It is suggested that the direction and extentof stomatal changes induced by water deficits is a consequenceof the rate of change of leaf water content and not of the absolutevalues. The stomata also showed an enhanced tendency to closein dry moving air following a period of wilting even after theleaf had regained turgidity.     

Environmental and genetic components of stomatal behavior in two genotypes of upland cotton

Two parental lines of Upland cotton (Gossypium hirsutum L.) plus their reciprocal F₁, F₂, and backcross populations were field-tested for environmental and genetic components of leaf diffusive resistance. Leaf resistance was measured with a diffusion porometer three times each day during 6 days in August. A large, consistent environmental component was present during the morning, afternoon, and evening, but the genetic component appeared mainly during the afternoon. Leaf water potential indicated that afternoon was the period of highest water stress. Genetic analyses revealed that leaf resistance was associated with both additive and dominance genetic variances, with an estimated narrow sense heritability of 25%. High leaf resistance was completely dominant to low resistance.     

After-effect of Water Stress on Stomatal Opening Potential: I. TECHNIQUES AND MAGNITUDES

The after-effect of a period of water stress upon the light-openingability of stomata was studied in tobacco (Nicotiana tabaccum)and broad bean (Vicia faba). Stomatal opening was always measuredafter floating leaf discs on water under favourable conditionsof light and temperature to obtain maximal opening at full leafturgor. Thus possible interference due to persistence of leaf-waterdeficits in the post-stress leaves was eliminated. Stomatalaperture was accurately estimated using a resistance porometer;values so obtained were substantiated by the direct measurementof aperture. Two to 4 days of wilting of tobacco, producing leaf-water deficitsof 30 to 40 per cent, had a marked after-effect on tobacco stomata.The ability to open in light was depressed and complete recoveryfrom this depression required 2 to 5 days after rewatering.In some cases over-recovery was observed; this was probablyrelated to a physiologically younger condition of leaves ofstressed plants following turgor recovery. In beans similarstresses caused after-effects smaller in magnitude but qualitativelysimilar to those in tobacco. In both tobacco and beans the magnitudeof the after-effect was approximately proportional to the leaf-waterdeficit attained immediately prior to rewatering.