Multiple controls for the variability of hydrogen isotopic compositions in higher plant n-alkanes from modern ecosystems

We performed a global scale analysis of available leaf wax n -alkane δ D data compiled from our new results, as well as from the literature and expressed as average values of D/H ratios from three common lipids of n -alkanes with odd carbon numbers ( n -C₂₇, n -C₂₉, and n -C₃₁) from living higher plants. Our results clearly indicate multiple controls of hydrogen isotope composition and its variability in plants leaf wax. (1) At the global scale, precipitation δ D values play a dominating factor that exercises the first order of control for hydrogen isotopic compositions in plant leaf wax. The hydrogen isotopic composition of plant leaf wax tracks the decreasing trend of precipitation δ D with increasing latitude. (2) Because of different water acquisition systems, plant life form influences the hydrogen isotopic composition of leaf wax n -alkanes with woody plants and grasses having different responses to the change of global precipitation δ D. (3) Physiological difference, due to different photosynthesis pathways or different water usage strategies, can leave an imprint on δ D patterns of plant leaf waxes, causing δ D variations among plants using the same source water. While these results better explain the variability of hydrogen isotope composition in leaf wax, they also have important implications for the interpretation of n -alkane δ D data from fossils and ancient sediments.     

Modelling environmental controls on ecosystem photosynthesis and the carbon isotope composition of ecosystem-respired CO2 in a coastal Douglas-fir forest

We developed and applied an ecosystem-scale model that calculated leaf CO₂ assimilation, stomatal conductance, chloroplast CO₂ concentration and the carbon isotope composition of carbohydrate formed during photosynthesis separately for sunlit and shaded leaves within multiple canopy layers. The ecosystem photosynthesis model was validated by comparison to leaf-level gas exchange measurements and estimates of ecosystem-scale photosynthesis from eddy covariance measurements made in a coastal Douglas-fir forest on Vancouver Island. A good agreement was also observed between modelled and measured δ ¹³C values of ecosystem-respired CO₂ ( δ _R). The modelled δ _R values showed strong responses to variation in photosynthetic photon flux density (PPFD), air temperature, vapour pressure deficit (VPD) and available soil moisture in a manner consistent with leaf-level studies of photosynthetic ¹³C discrimination. Sensitivity tests were conducted to evaluate the effect of (1) changes in the lag between the time of CO₂ fixation and the conversion of organic matter back to CO₂; (2) shifts in the proportion of autotrophic and heterotrophic respiration; (3) isotope fractionation during respiration; and (4) environmentally induced changes in mesophyll conductance, on modelled δ _R values. Our results indicated that δ _R is a good proxy for canopy-level C _c/ C _a and ¹³C discrimination during photosynthetic gas exchange, and therefore has several applications in ecosystem physiology.     

Water vapour uptake by diapausing eggs of a tropical walking stick

Abstract. This study is the first to demonstrate the capacity of an arthropod egg, that of a tropical walking stick Extatosoma tiaratum (Macleuy), to absorb water vapour from the air. This species diapauses both as an early embryo and then again as a pharate first instar larva, and both stages are capable of absorbing water vapour. Water vapour absorption occurs at lower humidities and at a lower rate for an egg in early embryonic diapause (c. a_v 0.30, 0.516 mg h^-1δ_v^-1) than in the diapausing pharate first instar (c. a_v 0.60, 0.725 mgh^-1δa_v^-1) at 25C. In addition to having the capacity to gain water at very low vapour activities, water is efficiently conserved as indicated by the low rate of water loss (0.015% h^-1 in the early embryo and 0.046% h^-1 in the pharate larva at 25C). Eggs that have been killed lose water when held at a hydrating vapour activity, thus implying that active uptake contributes to net absorption. Wax block experiments suggest that water is absorbed over the entire chorionic surface. Eggs of five other insect species that were examined [Lymantria dispar (L.), Bombyx mori (L.), Antheraea polyphemus (Cram.), Oncopeltus fasciatus (Dallas) and Diaferomera femorata (Say)] lacked the ability to absorb atmospheric water.     

The application of δ¹⁸O and δD for understanding water pools and fluxes in a Typha marsh

The δ¹⁸O and δD composition of water pools (leaf, root, standing water and soil water) and fluxes [transpiration (T), evaporation (E)] were used to understand ecohydrological processes in a managed Typha latifolia L. freshwater marsh. We observed isotopic steady‐state T and deep rooting in Typha. The isotopic mass balance of marsh standing water showed that E accounted for 3% of the total water loss, T accounted for 17% and subsurface drainage (D) accounted for the majority (80%). There was a vertical gradient in water vapour content and isotopic composition within and above the canopy sufficient for constructing an isotopic mass balance of water vapour during some sampling periods. During these periods, the proportion of T in evapotranspiration (T/ET) was between 56 ± 17% and 96 ± 67%, and the estimated error was relatively high (>37%) because of non‐local, background sources in vapour. Independent estimates of T/ET using eddy covariance measurements yielded similar mean values during the Typha growing season. The various T/ET estimates agreed that T was the dominant source of marsh vapour loss in the growing season. The isotopic mass balance of water vapour yielded reasonable results, but the mass balance of standing water provided more definitive estimates of water losses.     

Stomatal responses of pearl millet (Pennisetum americanum [L.] Leeke) to leaf water status and environmental factors in the field

Abstract. Factors affecting stomatal conductance (g₁) of pearl millet ( Pennisetum americanum [L.] Leeke), cultivar BJ 104, were examined in the field in India during the dry season.
Diurnal changes in g₁ were evaluated for upper expanded leaves at flowering on two occasions using plants subjected to varying degrees of water stress. Except for the most severely stressed treatment, diurnal changes in g₁ closely matched changes in irradiance ( I ), the promotive effect of which largely overcame opposing influences on g₁ of increasing atmospheric vapour pressure deficit, and decreasing leaf water and turgor potentials (Ψ, Ψ_p).
Two main effects of water stress on g₁ were evident: (i) a decrease in the amplitude of the mid-day peak in g₁, and (ii) a decrease in the time over which high g₁ was maintained, resulting in early (mid-day) closure and hysteresis in the relationship between g₁ and I .
Leaf conductance was greatest for upper leaves and decreased down the canopy. At equivalent depths in the canopy g₁ was higher in flowering than in photoperiodically-retarded plants of the same age. The magnitude of water stress-induced stomatal closure increased down the plant, and was more marked in retarded than in flowering plants.
Within individual stress treatments Ψ of upper leaves decreased linearly as transpiration flux increased. It is concluded that stomatal behaviour of upper leaves of pearl millet at flowering largely operates to maximize assimilation rather than to minimize water loss.     

Tracing carbon and oxygen isotope signals from newly assimilated sugars in the leaves to the tree-ring archive

The analysis of δ ¹³C and δ ¹⁸O in tree-ring archives offers retrospective insights into environmental conditions and ecophysiological processes. While photosynthetic carbon isotope discrimination and evaporative oxygen isotope enrichment are well understood, we lack information on how the isotope signal is altered by downstream metabolic processes.
In Pinus sylvestris , we traced the isotopic signals from their origin in the leaf water ( δ ¹⁸O) or the newly assimilated carbon ( δ ¹³C), via phloem sugars to the tree-ring, over a time-scale that ranges from hours to a growing season.
Seasonally, variable ¹³C enrichment of sugars related to phloem loading and transport did lead to uncoupling between δ ¹³C in the tree-ring, and the c _i/ c _a ratio at the leaf level. In contrast, the oxygen isotope signal was transferred from the leaf water to the tree-ring with an expected enrichment of 27‰, with time-lags of approximately 2 weeks and with a 40% exchange between organic oxygen and xylem water oxygen during cellulose synthesis.
This integrated overview of the fate of carbon and oxygen isotope signals within the model tree species P. sylvestris provides a novel physiological basis for the interpretation of δ ¹³C and δ ¹⁸O in tree-ring ecology.     

Diurnal and seasonal variation in the carbon isotope composition of leaf dark-respired CO2 in velvet mesquite (Prosopis velutina)

We evaluated diurnal and seasonal patterns of carbon isotope composition of leaf dark-respired CO₂ ( δ ¹³C_l) in the C₃ perennial shrub velvet mesquite ( Prosopis velutina ) across flood plain and upland savanna ecosystems in the south-western USA. δ ¹³C_l of darkened leaves increased to maximum values late during daytime periods and declined gradually over night-time periods to minimum values at pre-dawn. The magnitude of the diurnal shift in δ ¹³C_l was strongly influenced by seasonal and habitat-related differences in soil water availability and leaf surface vapour pressure deficit. δ ¹³C_l and the cumulative flux-weighted δ ¹³C value of photosynthates were positively correlated, suggesting that progressive ¹³C enrichment of the CO₂ evolved by darkened leaves during the daytime mainly resulted from short-term changes in photosynthetic ¹³C discrimination and associated shifts in the δ ¹³C signature of primary respiratory substrates. The ¹³C enrichment of dark-respired CO₂ relative to photosynthates across habitats and seasons was 4 to 6‰ at the end of the daytime period (1800 h), but progressively declined to 0‰ by pre-dawn (0300 h). The origin of night-time and daytime variations in δ ¹³C_l is discussed in terms of the carbon source(s) feeding respiration and the drought-induced changes in carbon metabolism.     

Transport of gases into leaves

Abstract. Transport of gases between the intercellular spaces of plant leaves and the surrounding air is analysed in terms of multicomponent collision processes through an isothermal, porous septum. Interaction of diffusing species with each other and with the pore walls is described using a modified Stefan–Maxwell equation and an equation relating the pressure gradient to the sum of the diffusive fluxes, weighted by their appropriate Knudsen diffusivities. Viscous How arising from an excess pressure within the leaf is also considered.
Equations are derived which describe the flux densities of water vapour and CO₂ through the stomata. The analysis is general and is applicable to trace gases other than CO₂. A simple conductance is defined for water vapour to relate the flux and mol fraction difference across the stomata, viz. N_w =− g_w , δ x_w / x_a . A simple conductance cannot be defined for CO₂ because the flux of water vapour has a significant influence on the CO₂ gradient. The equation derived for the intercellular mol fraction of CO₂ is in terms of the fluxes of CO₂ and water vapour and represents a 'large-pore' ( d > μm) approximation which requires no information about stomalal geometry. Analogous equations are developed for transfer of gases through the leaf boundary layer. Sample calculations are presented to illustrate the effect of neglecting the interaction of water vapour and CO₂ on the calculated intercellular and surface concentrations of CO₂. Equations for computing water vapour and CO₂ flux densities from leaf chamber measurements are also presented.     

Physiological and isotopic (δ13C and δ18O) responses of three tropical tree species to water and nutrient availability

Water-use efficiency and stable isotope composition were studied in three tropical tree species. Seedlings of Tectona grandis , Swietenia macrophylla and Platymiscium pinnatum were grown at either high or low water supply, and with or without added fertilizer. These three species previously exhibited low, intermediate and high whole-plant water-use efficiency ( TE ) when grown at high water supply in unfertilized soil. Responses of TE to water and nutrient availability varied among species. The TE was calculated as experiment-long dry matter production divided by cumulative water use. Species-specific offsets were observed in relationships between TE and whole-plant ¹³C discrimination (Δ¹³C_p). These offsets could be attributed to a breakdown in the relationship between Δ¹³C_p and the ratio of intercellular to ambient CO₂ partial pressures ( c _i/ c _a) in P. pinnatum , and to variation among species in the leaf-to-air vapour pressure difference ( v ). Thus, a plot of v · TE against c _i/ c _a showed a general relationship among species. Relationships between δ ¹⁸O of stem dry matter and stomatal conductance ranged from strongly negative for S. macrophylla to no relationship for T. grandis . Results suggest inter-specific variation among tropical tree species in relationships between stable isotope ratios ( δ ¹³C and δ ¹⁸O) and the gas exchange processes thought to affect them.     

Photosynthesis is improved by exogenous glycinebetaine in salt-stressed maize plants

The effects of exogenous application of glycinebetaine (GB) (10 m M ) on growth, leaf water content, water use efficiency, photosynthetic gas exchange, and photosystem II photochemistry were investigated in maize plants subjected to salt stress (50 and 100 m M NaCl). Salt stress resulted in the decrease in growth and leaf relative water content as well as net photosynthesis and the apparent quantum yield of photosynthesis. Stomatal conductance, evaporation rate, and water use efficiency were decreased in salt-stressed plants. Salt stress also caused a decrease in the actual efficiency of PSII ( Φ _PSII), the efficiency of excitation energy capture by open PSII reaction centers ( F _v'/ F _m'), and the coefficients of photochemical quenching ( q _P) but caused an increase in non-photochemical quenching (NPQ). Salt stress showed no effects on the maximal efficiency of PSII photochemistry ( F _v/ F _m). On the other hand, in salt-stressed plants, GB application improved growth, leaf water content, net photosynthesis, and the apparent quantum yield of photosynthesis. GB application also increased stomatal conductance, leaf evaporation rate, and water use efficiency. In addition, GB application increased Φ _PSII, F _v'/ F _m', and q _P but decreased NPQ. However, GB application showed no effects on F _v/ F _m. These results suggest that photosynthesis was improved by GB application in salt-stressed plants and such an improvement was associated with an improvement in stomatal conductance and the actual PSII efficiency.     

Determinants of isotopic coupling of CO2 and water vapour within a Quercus petraea forest canopy

Concentration and isotopic composition (δ¹³C and δ¹⁸O) of ambient CO₂ and water vapour were determined within a Quercus petraea canopy, Northumberland, UK. From continuous measurements made across a 36-h period from three heights within the forest canopy, we generated mixing lines (Keeling plots) for δ_a ¹³CO₂, δ_a C¹⁸O¹⁶O and δ_a H₂ ¹⁸O, to derive the isotopic composition of the signal being released from forest to atmosphere. These were compared directly with measurements of different respective pools within the forest system, i.e. δ¹³C of organic matter input for δ_a ¹³CO₂, δ¹⁸O of exchangeable water for δ_a C¹⁸O¹⁶O and transpired water vapour for δ_a H₂ ¹⁸O. [CO₂] and δ_a ¹³CO₂ showed strong coupling, where the released CO₂ was, on average, 4 per mil enriched compared to the organic matter of plant material in the system, suggesting either fractionation of organic material before eventual release as soil-respired CO₂, or temporal differences in ecosystem discrimination. δ_a C¹⁸O¹⁶O was less well coupled to [CO₂], probably due to the heterogeneity and transient nature of water pools (soil, leaf and moss) within the forest. Similarly, δ_a H₂ ¹⁸O was less coupled to [H₂O], again reflecting the transient nature of water transpired to the forest, seen as uncoupling during times of large changes in vapour pressure deficit. The δ¹⁸O of transpired water vapour, inferred from both mixing lines at the canopy scale and direct measurement at the leaf level, approximated that of source water, confirming that an isotopic steady state held for the forest integrated over the daily cycle. This demonstrates that isotopic coupling of CO₂ and water vapour within a forest canopy will depend on absolute differences in the isotopic composition of the respective pools involved in exchange and on the stability of each of these pools with time. Received: 21 March 1998 / Accepted: 10 December 1998     

Towards a predictive description of forest canopies from litter properties

1. Specific leaf area (SLA), carbon isotope ratio (δ¹³C) and leaf nitrogen content (N) were measured in litter in 1993 and in the litter layer and canopy in 1996 in a pedunculate Oak ( Quercus robur ) wood at Wytham, Oxford.
2. The frequency distribution of SLA of leaves falling through the late summer and autumn of 1993 changed with a greater contribution of leaves of high SLA earlier in the period. For the same SLA leaves falling earlier in the season had less negative δ¹³C and higher N content than leaves falling later.
3. The frequency distribution of SLA of litter leaves was similar in both years but litter leaves had lower N and less negative δ¹³C in 1996.
4. In the canopy, there was a highly significant negative relationship between height in the canopy and SLA in green and senescent leaves. The relationship between SLA and both δ¹³C ratio and leaf N was different between green, senescent and litter leaves.
5. The frequency distribution of foliage in the canopy estimated from the frequency of SLA in the litter and the vertical pattern of SLA in the canopy was similar to the distribution measured by the 'drop-line' method.     

Fluoride inhibits root water transport and affects leaf expansion and gas exchange in aspen (Populus tremuloides) seedlings

The effects of sodium fluoride (0.3, 5 and 10 m M NaF) on root hydraulic conductivity, and gas exchange processes were examined in aspen ( Populus tremuloides Michx.) seedlings grown in solution culture. A long-term exposure of roots to NaF significantly decreased root hydraulic conductivity ( L _p) and stomatal conductance ( g _s). Root absorbed NaF significantly affected electrolyte leakage in leaf tissues and substantially restricted leaf expansion. NaF did not significantly affect leaf chlorophyll contents but decreased net photosynthesis ( P _n). A short-term exposure of excised roots to 5 m M NaF and KF significantly decreased root water flow ( Q _v) with a concomitant decline in root respiration and reduced g _s when applied through intact roots or excised stems. The same molar concentration of NaCl also decreased Q _v and g _s in intact seedlings, but to a lesser extent than NaF or KF, and did not significantly affect root respiration. The results suggest that fluoride metabolically inhibited Q _v or L _p, probably by affecting water channel activity. We suggest that the metabolic inhibition of L _p by root-absorbed fluoride affected gas exchange and leaf expansion in aspen seedlings.     

Variability in the carbon isotope signature of Prochilodus lineatus (Prochilodontidae, Characiformes) a bottom-feeding fish of the Neotropical region

The spatial and seasonal variability of stable carbon isotopes in detritivorous Prochilodus lineatus , primary producers and particulate organic carbon (POC) were examined to promote a better understanding of the trophic dynamics of food webs in Neotropical ecosystems. Spatial variability in carbon isotope composition in the species showed a decreasing gradient in the Upper Paraná River system, from the Paraná River and Itaipu Reservoir subsystems to the Baía and Ivinheima subsystems. Fish δ¹³C followed the isotopic trend of POC along the Paraná River and Baía subsystem. Seasonal differences were not observed in fish. The analysis of percentile contribution of C₃ and C₄ sources in the species diet revealed the predominance of C₃ plants (phytoplanktonic and periphytic algae and riparian vegetation) in most of the analysed subsystems and seasons. Nevertheless, the considerable participation of riparian vegetation was verified, especially in the Ivinheima subsystem in the dry season. C₄ macrophytes presented a maximum contribution along the Upper Paraná River. These results demonstrate the existence of specific variability for the consumer and their food sources in different environments of the same ecosystem.     

Bulk leaf δ18O and δ13C reflect the intensity of intraspecific competition for water in a semi-arid tussock grassland

We investigated the extent to which plant water and nutrient status are affected by intraspecific competition intensity and microsite quality in a monodominant tussock grassland. Leaf gas exchange and stable isotope measurements were used to assess the water relations of Stipa tenacissima tussocks growing along a gradient of plant cover and soil depth in a semi-arid catchment of Southeast Spain. Stomatal conductance and photosynthetic rate decreased with increasing intensity of competition during the wet growing season, leading to foliar δ ¹⁸O and δ ¹³C enrichment. A high potential for runoff interception by upslope neighbours exerted strong detrimental effects on the water and phosphorus status of downslope S. tenacissima tussocks. Foliar δ ¹⁵N values became more enriched with increasing soil depth. Multiple stepwise regression showed that competition potential and/or rhizosphere soil depth accounted for large proportions of variance in foliar δ ¹³C, δ ¹⁸O and δ ¹⁵N among target tussocks (57, 37 and 64%, respectively). The results presented here highlight the key role that spatial redistribution of resources (water and nutrients) by runoff plays in semi-arid ecosystems. It is concluded that combined measurement of δ ¹³C, δ ¹⁸O and nutrient concentrations in bulk leaf tissue can provide insight into the intensity of competitive interactions occurring in natural plant communities.     

Water vapour fluxes and their impact under elevated CO2 in a C4-tallgrass prairie

We measured leaf-level stomatal conductance, xylem pressure potential, and stomate number and size as well as whole plant sap flow and canopy-level water vapour fluxes in a C4-tallgrass prairie in Kansas exposed to ambient and elevated CO₂. Stomatal conductance was reduced by as much as 50% under elevated CO₂ compared to ambient. In addition, there was a reduction in stomate number of the C4 grass, Andropogon gerardii Vitman, and the C3 dicot herb, Salvia pitcheri Torr., under elevated CO₂ compared to ambient. The result was an improved water status for plants exposed to elevated CO₂ which was reflected by a less negative xylem pressure potential compared to plants exposed to ambient CO₂. Sap flow rates were 20 to 30% lower for plants exposed to elevated CO₂ than for those exposed to ambient CO₂. At the canopy level, evapotranspiration was reduced by 22% under elevated CO₂. The reduced water use by the plant canopy under elevated CO₂ extended the photosynthetically-active period when water became limiting in the ecosystem. The result was an increased above- and belowground biomass production in years when water stress was frequent.     

Effect of water availability on leaf water isotopic enrichment in beech seedlings shows limitations of current fractionation models

Current models of leaf water enrichment predict that the differences between isotopic enrichment of water at the site of evaporation (Δ_e) and mean lamina leaf water enrichment (Δ_L) depend on transpiration rates ( E ), modulated by the scaled effective length ( L ) of water isotope movement in the leaf. However, variations in leaf parameters in response to changing environmental conditions might cause changes in the water path and thus L . We measured the diel course of Δ_L for ¹⁸O and ²H in beech seedlings under well-watered and water-limited conditions. We applied evaporative enrichment models of increasing complexity to predict Δ_e and Δ_L, and estimated L from model fits. Water-limited plants showed moderate drought stress, with lower stomatal conductance, E and stem water potential than the control. Despite having double E , the divergence between Δ_e and Δ_L was lower in well-watered than in water-limited plants, and thus, L should have changed to counteract differences in E . Indeed, L was about threefold higher in water-limited plants, regardless of the models used. We conclude that L changes with plant water status far beyond the variations explained by water content and other measured variables, thus limiting the use of current evaporative models under changing environmental conditions.     

Does the 13C of foliage‐respired CO2 and biochemical pools reflect the 13C of recently assimilated carbon?

Isotopic labelling experiments were conducted to assess relationships among ¹³C of recently assimilated carbon ( δC _A), foliage respiration ( δC _F), soluble carbohydrate ( δC _SC), leaf waxes ( δC _LW) and bulk organic matter ( δC _OM). Slash pine, sweetgum and maize were grown under ¹³C depleted CO₂ to label biomass and then placed under ambient conditions to monitor the loss of label. In pine and sweetgum, δC _F of labelled plants (∼−44 and −35‰, respectively) rapidly approached control values but remained depleted by ∼4–6‰ after 3–4 months. For these tree species, no or minimal label was lost from δC _SC, δC _LW and δC _OM during the observation periods. δC _F and δC _SC of labelled maize plants rapidly changed and were indistinguishable from controls after 1 month, while δC _LW and δC _OM more slowly approached control values and remained depleted by 2–6‰. Changes in δC _F in slash pine and sweetgum fit a two-pool exponential model, with the fast turnover metabolic pool (∼3–4 d half-life) constituting only 1–2% of the total. In maize, change in δC _F fits a single pool model with a half-life of 6.4 d. The ¹³C of foliage respiration and biochemical pools reflect temporally integrated values of δC _A, with change in isotopic composition dampened by the size of metabolic carbon reserves and turnover rates.     

Night-time transpiration can decrease hydraulic redistribution

C₃ plants dominate many landscapes and are critically important for ecosystem water cycling. At night, plant water losses can include transpiration (E_night) from the canopy and hydraulic redistribution (HR) from roots. We tested whether E_night limits the magnitude of HR in a greenhouse study using Artemisia tridentata , Helianthus anomalus and Quercus laevis . Plants were grown with their roots split between two compartments. HR was initiated by briefly withholding all water, followed by watering only one rooting compartment. Under study conditions, all species showed substantial E_night and HR (highest minus lowest soil water potential [ Ψ _s] during a specified diel period). Suppressing E_night by canopy bagging increased HR during the nightly bagging period (HR_N) for A. tridentata and H. anomalus by 73 and 33% respectively, but did not affect HR_N by Q. laevis . Total daily HR (HR_T) was positively correlated with the Ψ _s gradient between the rooting compartments, which was correlated with light and/or atmospheric vapour pressure deficit (VPDa) the prior day. For A. tridentata , HR_T was negatively correlated with night-time VPDa. Ecological implications of the impact of E_night on HR may include decreased plant productivity during dry seasons, altered ecosystem water flux patterns and reduced nutrient cycling in drying soils.     

The influence of transpiration on the equilibration of leaf water with atmospheric water vapour

Abstract. The authors examine the isotopic composition of leaf water, at natural abundance levels, as influenced by transpiration rate. The isotopic composition of water of wheat leaves ( Triticum aestivum L. var. Aroona) was followed while their transpiration rate adjusted to 'steady-state' environmental conditions. Leaf diffusive resistance was modified by short-term salt treatment and by plant culture in either nutrient solution, free-draining sand, or vermiculite. Resultant changes in ¹⁸O and ²H in leaf water are described and fitted to the model of Leaney et al. (1985). The treatments with lower transpiration rates were found to have a greater fraction of their leaf water equilibrated with water vapour in the atmosphere. Comparable results were obtained with both ¹⁸O and ²H, with some differences being interpreted in terms of turbulence in the vapour diffusion path. The fraction of the leaf water equilibrated with the atmosphere varied between leaves of different ages. However, this may have been due to their different positions in the canopy.