Combined Gas Exchange Measurements Research Articles

Mesophyll conductance exerts a significant limitation on photosynthesis during light induction.

Past studies have established mesophyll diffusion conductance to CO2 (gm ) as a variable and significant limitation to plant photosynthesis under steady-state conditions. However, the role of gm in influencing photosynthesis (A) during the transient period of light induction is largely unknown. We combined gas exchange measurements with laser-enabled carbon isotope discrimination measurements to assess gm during photosynthetic induction, using Arabidopsis as the measurement species. Our measurements revealed three key findings: (1) we found that the rate at which gm approached steady state during induction was not necessarily faster than the induction rate of the carboxylation process, contradictory to what has been suggested in previous studies; (2) gm displayed a strong and consistent coordination with A under both induction and steady-state settings, hinting that the mechanism driving gm -A coupling does not require physiological stability as a prerequisite; and (3) photosynthetic limitation analysis of our data revealed that when integrated over the entire induction period, the relative limitation of A imposed by gm can be as high as > 35%. The present study provides the first demonstration of the important role of gm in limiting CO2 assimilation during photosynthetic induction, thereby pointing to a need for more research attention to be devoted to gm in future induction studies.

Light Quality Affects Chloroplast Electron Transport Rates Estimated from Chl Fluorescence Measurements.

Chl fluorescence has been used widely to calculate photosynthetic electron transport rates. Portable photosynthesis instruments allow for combined measurements of gas exchange and Chl fluorescence. We analyzed the influence of spectral quality of actinic light on Chl fluorescence and the calculated electron transport rate, and compared this with photosynthetic rates measured by gas exchange in the absence of photorespiration. In blue actinic light, the electron transport rate calculated from Chl fluorescence overestimated the true rate by nearly a factor of two, whereas there was closer agreement under red light. This was consistent with the prediction made with a multilayer leaf model using profiles of light absorption and photosynthetic capacity. Caution is needed when interpreting combined measurements of Chl fluorescence and gas exchange, such as the calculation of CO2 partial pressure in leaf chloroplasts.

Effect of water and salt stress on energy partitioning of two grapevine rootstock genotypes: a quantitative assessment

The ability of plants to adapt and/or acclimate to different environments is related to the plasticity and resilience of photosynthesis. This study aims at assessing the effects of salinity, drought and recovery on photosynthesis being either direct (stomatal limitations) or secondary (oxidative stress). Combined measurements of gas exchange and chlorophyll fluorescence enabled to quantify the absorbed energy used for net photosynthesis and that dissipated by photoprotective mechanisms. Measurements were performed under controlled conditions on leaves of susceptible (101.14) and new putative tolerant selection (M4) grapevine rootstocks in control and progressive drought and salt loading conditions. Leaves of 101.14 showed higher photosynthetic performances than the M4 in well-watered conditions. However, M4 showed a greater ability to tolerate increasing salt concentration and drought and an almost complete recovery after re-watering. Quenching analysis revealed the photoprotective thermal dissipation (ΦNPQ) as the major responsible for the different stress responses among susceptible and tolerant rootstock studied.

(13) CO2 /(12) CO2 exchange fluxes in a clamp-on leaf cuvette: disentangling artefacts and flux components.

Leaks and isotopic disequilibria represent potential errors and artefacts during combined measurements of gas exchange and carbon isotope discrimination (Δ). This paper presents new protocols to quantify, minimize, and correct such phenomena. We performed experiments with gradients of CO2 concentration (up to ±250 μmol mol(-1) ) and δ(13) CCO2 (34‰), between a clamp-on leaf cuvette (LI-6400) and surrounding air, to assess (1) leak coefficients for CO2 , (12) CO2 , and (13) CO2 with the empty cuvette and with intact leaves of Holcus lanatus (C3 ) or Sorghum bicolor (C4 ) in the cuvette; and (2) isotopic disequilibria between net photosynthesis and dark respiration in light. Leak coefficients were virtually identical for (12) CO2 and (13) CO2 , but ∼8 times higher with leaves in the cuvette. Leaks generated errors on Δ up to 6‰ for H. lanatus and 2‰ for S. bicolor in full light; isotopic disequilibria produced similar variation of Δ. Leak errors in Δ in darkness were much larger due to small biological : leak flux ratios. Leak artefacts were fully corrected with leak coefficients determined on the same leaves as Δ measurements. Analysis of isotopic disequilibria enabled partitioning of net photosynthesis and dark respiration, and indicated inhibitions of dark respiration in full light (H. lanatus: 14%, S. bicolor: 58%).

Fitting mitochondrial respiration rates under light by photosynthetic CO<inf>2</inf> response models

Aims The objective of this study was to compare the values of respiration under light derived by fitting a photosynthetic CO2 response model and measurements, in order to provide information for model optimization. Methods Using combined gas exchange measurements and a low O2 (2% O2) method, the responses of photosynthetic rate (Pn) to CO2 at different light intensities (2 000, 1 500, 1 000 and 500 μmol·m·s) in the flag leaves of wheat were measured. The measured data were fitted by a biochemical model, a rectangular hyperbola model and a modified rectangular hyperbola model of the photosynthetic response to intercellular CO2 concentration (A/Ci) and air CO2 concentration (A/Ca), aiming to approach the reasonability of the fitted results obtained from the models. Important finding The sequence of fitting effect of the three CO2 response models in descending order was as follows: modified rectangular hyperbola model > rectangular hyperbola model > biochemical model. Fitted values of A/Ca curve was more reasonable than A/Ci curve, because the photorespiration and mitochondrial respiration under light (Rd) estimated by the former better matched the measured values. However, there were significant differences in the whole between the fitted and measured values. The reason could be that the effect of CO2 concentrations on Rd and apparent photorespiration (Rpa) is neglected in all the current CO2 response models. Our results showed that CO2 concentration had a marked effect on Rpa and Rd. With increasing CO2 concentration, Rpa and Rd increased first, and then decreased sharply. Take 2 000 μmol·m·s for example, Rpa varied between 5.035 and 11.670 μmol CO2·m·s, and Rd varied between 0.491 and 2.987 μmol CO2·m·s. Regression analyses 康华靖等: 植物光合 CO2响应模型对光下(暗)呼吸速率拟合的探讨 1357 doi: 10.3724/SP.J.1258.2014.00130 indicated that Rpa and Rd were well related to CO2 concentrations at different light intensities.

Causes of decreasing mitochondrial respiration under light in four crops

Aims Despite the increasing attention given to the rate of mitochondrial respiration under light (Rd), considerable confusion persists over whether mitochondrial respiration in the dark (Rn) is inhibited by light and whether Rd is affected by light intensity. The objective of this study is to test the hypotheses: 1) Rn is not inhibited by light; 2) the rate of Rd changes with light intensity; and 3) the photosynthetic refixation of CO2 produced by Rn accounts for the apparent disparity between Rd and Rn. Methods In the present study, 0.02 mol·mol O2 (i.e. 2% O2) was used to saturate Rn and to inhibit photorespiration (Rp). By using combined gas exchange measurements and a low O2 (2% O2) method, the post-illumination CO2 release rate of Rn, photosynthetic rate (Pn) in response to photosynthetically active radiation (PAR) in 2% O2 at either 380 or 0 μmol·mol CO2, of C3 (Triticum aestivum and Glycine max) and C4 (Zea mays and Amaranthus hypochondriacus) plants, were measured. Important finding Rn was not inhibited by light. At 2% O2 and 0 μmol·mol CO2, the measured parameters could be used to accurately estimate Rd when CO2 concentration was set for 0 μmol·mol. Rd decreased with increasing light intensity. Although Rd was lower in the dark, this could be accounted for by photosynthetic re-fixing of respiratory CO2. For all plants tested, CO2 recovery rates increased with increasing light intensity (from 50 and 2 000 μmol·m·s).

Limitation factors for photosynthesis in ‘Bluecrop’ highbush blueberry (Vaccinium corymbosum) leaves in response to moderate water stress

The levels of stomatal, mesophyll and biochemical limitations in CO2 assimilation of ‘Bluecrop’ highbush blueberry leaves were compared at two different levels of leaf water potential. The leaf water potentials were −1.49 and −1.94 MPa in daily-irrigated (DI) and non-irrigated (NI) shrubs, respectively. The NI shrubs represented plants under moderate water stress. Mesophyll conductance (g m) and chloroplastic CO2 concentration (C c) were estimated by combined measurements of gas exchange and chlorophyll fluorescence under various intercellular CO2 concentrations (C i). Net CO2 assimilation rates (A n) as a function of C c were used for calculating maximum carboxylation efficiency (α cmax) at the real sites of CO2 assimilation. Maximum A n (A nmax) from the light response curves at 400 μmol mol−1 air of ambient CO2 concentration (C a) were lower in the leaves of NI shrubs than in those of DI ones. However, electron transport rates were higher in the leaves of NI shrubs than in those of DI ones. The decrease in CO2 assimilation following water stress may be caused by a decrease in g m rather than a decrease in stomatal conductance (g s) according to limitation analysis. Limitation rates by g s, calculated at 400 μmol mol−1 air of C a in A n-C i curves, were not significantly different between the leaves of DI and NI shrubs. However, limitation rates by g m from A n-C c curves were significantly higher in the leaves of NI shrubs than in those of DI ones. Maximum carboxylation efficiency (α cmax) values calculated from the A n-C c curve, contrary to those calculated from the A n-C i curve, were higher in the leaves of NI shrubs than in those of DI ones. Consequently, mesophyll limitation than stomatal and biochemical limitations mainly down-regulated the photosynthesis in the leaves of ‘Bluecrop’ blueberry shrubs during moderate water stress.

Anatomical basis of variation in mesophyll resistance in eastern Australian sclerophylls: news of a long and winding path.

In sclerophylls, photosynthesis is particularly strongly limited by mesophyll diffusion resistance from substomatal cavities to chloroplasts (r m), but the controls on diffusion limits by integral leaf variables such as leaf thickness, density, and dry mass per unit area and by the individual steps along the diffusion pathway are imperfectly understood. To gain insight into the determinants of r m in leaves with varying structure, the full CO2 physical diffusion pathway was analysed in 32 Australian species sampled from sites contrasting in soil nutrients and rainfall, and having leaf structures from mesophytic to strongly sclerophyllous. r m was estimated based on combined measurements of gas exchange and chlorophyll fluorescence. In addition, r m was modelled on the basis of detailed anatomical measurements to separate the importance of different serial resistances affecting CO2 diffusion into chloroplasts. The strongest sources of variation in r m were S c/S, the exposed surface area of chloroplasts per unit leaf area, and mesophyll cell wall thickness, t cw. The strong correlation of r m with t cw could not be explained by cell wall thickness alone, and most likely arose from a further effect of cell wall porosity. The CO2 drawdown from intercellular spaces to chloroplasts was positively correlated with t cw, suggesting enhanced diffusional limitations in leaves with thicker cell walls. Leaf thickness and density were poorly correlated with S c/S, indicating that widely varying combinations of leaf anatomical traits occur at given values of leaf integrated traits, and suggesting that detailed anatomical studies are needed to predict r m for any given species.

Using combined measurements of gas exchange and chlorophyll fluorescence to investigate the photosynthetic light responses of plant species adapted to different light regimes

One broad-leaved pioneer tree, Alnus formosana, two broad-leaved understory shrubs, Ardisia crenata and Ardisia cornudentata, and four ferns with different light adaptation capabilities (ranked from high to low, Pyrrosia lingus, Asplenium antiquum, Diplazium donianum, Archangiopteris somai) were used to elucidate the light responses of photosynthetic rate and electron transport rate (ETR). Pot-grown materials received up to 3 levels of light intensity, i.e., 100%, 50% and 10% sunlight. Both gas exchange and chlorophyll (Chl) fluorescence were measured simultaneously by an equipment under constant temperature and 7 levels (0-2,000 μmol m-2 s-1) of photosynthetic photon flux density (PPFD). Plants adapted to-or acclimated to high light always had higher light-saturation point and maximal photosynthetic rate. Even materials had a broad range of photosynthetic capacity [maximal photosynthetic rate ranging from 2 to 23 μmol(CO2) m-2 s-1], the ratio of ETR to gross photosynthetic rate (P G) was close for A. formosana and the 4 fern species when measured under constant temperature, but the PPFD varied. In addition, P. lingus and A. formosana grown under 100% sunlight and measured at different seasonal temperatures (15, 20, 25, and 30°C) showed increased ETR/P G ratio with increasing temperature and could be fitted by first- and second-order equations, respectively. With this equation, estimated and measured P G were closely correlated (r 2 = 0.916 and r 2 = 0.964 for P. lingus and A. formosana, respectively, p<0.001). These equations contain only the 2 easily obtained dynamic indicators, ETR and leaf temperature. Therefore, for some species with near ETR/P G ratio in differential levels of PPFD, these equations could be used to simulate dynamic variation of leaf scale photosynthetic rate under different temperature and PPFD conditions.

Using a biochemical C4 photosynthesis model and combined gas exchange and chlorophyll fluorescence measurements to estimate bundle‐sheath conductance of maize leaves differing in age and nitrogen content

Bundle-sheath conductance (g(bs) ) affects CO(2) leakiness, and, therefore, the efficiency of the CO(2) -concentrating mechanism (CCM) in C(4) photosynthesis. Whether and how g(bs) varies with leaf age and nitrogen status is virtually unknown. We used a C(4) -photosynthesis model to estimate g(bs) , based on combined measurements of gas exchange and chlorophyll fluorescence on fully expanded leaves of three different ages of maize (Zea mays L.) plants grown under two contrasting nitrogen levels. Nitrogen was replenished weekly to maintain leaf nitrogen content (LNC) at a similar level across the three leaf ages. The estimated g(bs) values on leaf-area basis ranged from 1.4 to 10.3 mmol m(-2) s(-1) and were affected more by LNC than by leaf age, although g(bs) tended to decrease as leaves became older. When converted to resistance (r(bs) = 1/g(bs)), r(bs) decreased monotonically with LNC. The correlation was presumably associated with nitrogen effects on leaf anatomy such as on wall thickness of bundle-sheath cells. Despite higher g(bs), meaning less efficient CCM, the calculated loss due to photorespiration was still low for high-nitrogen leaves. Under the condition of ambient CO(2) and saturating irradiance, photorespiratory loss accounted for 3-5% of fixed carbon for the high-nitrogen, versus 1-2% for the low-nitrogen, leaves.

Using combined measurements of gas exchange and chlorophyll fluorescence to estimate parameters of a biochemical C3 photosynthesis model: a critical appraisal and a new integrated approach applied to leaves in a wheat (Triticum aestivum) canopy

We appraised the literature and described an approach to estimate the parameters of the Farquhar, von Caemmerer and Berry model using measured CO(2) assimilation rate (A) and photosystem II (PSII) electron transport efficiency (Phi(2)). The approach uses curve fitting to data of A and Phi(2) at various levels of incident irradiance (I(inc)), intercellular CO(2) (C(i)) and O(2). Estimated parameters include day respiration (R(d)), conversion efficiency of I(inc) into linear electron transport of PSII under limiting light [kappa(2(LL))], electron transport capacity (J(max)), curvature factor (theta) for the non-rectangular hyperbolic response of electron flux to I(inc), ribulose 1.5-bisphosphate carboxylase/oxygenase (Rubisco) CO(2)/O(2) specificity (S(c/o)), Rubisco carboxylation capacity (V(cmax)), rate of triose phosphate utilization (T(p)) and mesophyll conductance (g(m)). The method is used to analyse combined gas exchange and chlorophyll fluorescence measurements on leaves of various ages and positions in wheat plants grown at two nitrogen levels. Estimated S(c/o) (25 degrees C) was 3.13 mbar microbar(-1); R(d) was lower than respiration in the dark; J(max) was lower and theta was higher at 2% than at 21% O(2); kappa(2(LL)), V(cmax), J(max) and T(p) correlated to leaf nitrogen content; and g(m) decreased with increasing C(i) and with decreasing I(inc). Based on the parameter estimates, we surmised that there was some alternative electron transport.

Characteristics of CO2 exchange between peach stems and the atmosphere.

Gas exchange by stems is dominated by respiratory CO2 emission, but photosynthetic CO2 uptake might also occur in stem bark. We show that light-dependent CO2 uptake was present and often exceeded CO2 release by respiration in illuminated current-year peach (Prunus persica L.) stems. Respiration of peach stems, as detected by 12CO2 release into air in which the natural concentration of 12CO2 was replaced with 13CO2, was lower in the light than in the dark, but this accounted for only a fraction of the observed total CO2 uptake by illuminated stems. Stem photosynthesis was saturated at low light and was negatively affected by elevated assay temperatures (30°C), especially when combined with light intensities above saturation. An inefficient mechanism of heat dissipation by transpiration in stomata-free stems might help explain this effect. Photosynthesis was rapidly stimulated and the electron transport rate was reduced when photorespiration was suppressed by exposure to low (2 kPa) oxygen. The time-course of these changes was closely associated with a transient burst of CO2 uptake concurrent with a reduced inhibition of fluorescence yield. Photosynthesis was also stimulated by exposure to elevated (twice ambient) CO2 concentration. These combined measurements of gas exchange and fluorescence suggested that (a) photorespiration may also be active in the bark of peach stems, (b) O2 and CO2 concentrations in the bark of peach stems may be similar to ambient concentrations, (c) a large amount of electron transport unrelated to photosynthesis and photorespiration may also be present in peach stems, and (d) stem photosynthesis may be enhanced under future atmospheric conditions.

Inhibition of photosynthesis by high temperature in oak (Quercus pubescens L.) leaves grown under natural conditions closely correlates with a reversible heat‐dependent reduction of the activation state of ribulose‐1,5‐bisphosphate carboxylase/oxygenase

ABSTRACTInhibition of the net photosynthetic CO2 assimilation rate (Pn) by high temperature was examined in oak (Quercus pubescens L.) leaves grown under natural conditions. Combined measurements of gas exchange and chlorophyll (Chl) a fluorescence were employed to differentiate between inhibition originating from heat effects on components of the thylakoid membranes and that resulting from effects on photosynthetic carbon metabolism. Regardless of whether temperature was increased rapidly or gradually, Pn decreased with increasing leaf temperature and was more than 90% reduced at 45 °C as compared to 25 °C. Inhibition of Pn by heat stress did not result from reduced stomatal conductance (gs), as heat‐induced reduction of gs was accompanied by an increase of the intercellular CO2 concentration (Ci). Chl a fluorescence measurements revealed that between 25 and 45 °C heat‐dependent alterations of thylakoid‐associated processes contributed only marginally, if at all, to the inhibition of Pn by heat stress, with photosystem II being remarkably well protected against thermal inactivation. The activation state of ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco) decreased from about 90% at 25 °C to less than 30% at 45 °C. Heat stress did not affect Rubisco per se, since full activity could be restored by incubation with CO2 and Mg2+. Western‐blot analysis of leaf extracts disclosed the presence of two Rubisco activase polypeptides, but heat stress did not alter the profile of the activase bands. Inhibition of Pn at high leaf temperature could be markedly reduced by artificially increasing Ci. A high Ci also stimulated photosynthetic electron transport and resulted in reduced non‐photochemical fluorescence quenching. Recovery experiments showed that heat‐dependent inhibition of Pn was largely, if not fully, reversible. The present results demonstrate that in Q. pubescens leaves the thylakoid membranes in general and photosynthetic electron transport in particular were well protected against heat‐induced perturbations and that inhibition of Pn by high temperature closely correlated with a reversible heat‐dependent reduction of the Rubisco activation state.

Effects of drought on photosynthesis in grapevines under field conditions: an evaluation of stomatal and mesophyll limitations.

The effect of diffusional and photochemical limitations to photosynthesis was assessed in field-grown water-stressed grapevines (Vitis vinifera L.) by combined measurements of gas exchange and chlorophyll fluorescence. Drought was slowly induced, and the progressive decline of photosynthesis was examined in different grapevine cultivars along a continuous gradient of maximum mid-morning values of stomatal conductance (g), which were used as an integrative indicator of the water-stress conditions endured by the leaves. Initial decreases of g were accompanied by decreases of substomatal CO2 concentration (Ci), the estimated chloroplastic CO2 concentration (Cc) and net photosynthesis (AN), while electron transport rate (ETR) remained unaffected. With increasing drought, g, AN, Ci and Cc further decreased, accompanied by slight decreases of ETR and of the estimated mesophyll conductance (gmes). Severe drought led to strong reductions of both g and gmes, as well as of ETR. The apparent carboxylation efficiency and the compensation point for CO2 remained unchanged under severe drought when analysed on a Cc, rather than a Ci, basis, suggesting that previously reported metabolic impairment was probably due to decreased gmes.

Seasonal patterns and control of gas exchange in local populations of the Mediterranean evergreen shrub Pistacia lentiscus L.

Abstract We examined temporal and spatial variations in net photosynthesis, stomatal conductance, intrinsic water-use efficiency, sub-stomatal CO 2 concentration, apparent carboxylation efficiency and chlorophyll fluorescence in the Mediterranean shrub Pistacia lentiscus . The study was done at the extremes of a precipitation and temperature gradient on the coast and in the mountains of Mallorca, Spain, with gas exchange measurements at different times of the year, and combined measurements of gas exchange and chlorophyll fluorescence in a controlled experiment. The objectives were to relate annual variation in photosynthetic functions to environmentally induced constraints and to quantify to which extent local differences in climate can affect photosynthesis in shrub populations. In the mountain population, net photosynthesis peaked in spring and autumn, when water was abundant and temperature was moderately high. It was reduced in winter paralleling reduced carboxylation efficiency. Photosynthesis was at the annual minimum in summer at both sites due to drought-induced stomata closure combined with impaired function of the Calvin cycle. The coastal population maintained high photosynthesis in mid winter but had a pronounced decline in spring, and the summer decline lasted longer than in the mountains. Integrated over the seasons, net photosynthesis was about 25 % lower in the coastal than in the mountain population, in spite of maintained high mid winter photosynthesis. Hence, the reduction at the coast was mainly due to early onset of drought in spring and a long period of summer drought, showing that local climatic differences can cause pronounced spatial differences in plant carbon balance. As a consequence, similar differences probably also occur as a function of year-to-year variability of precipitation patterns and temperatures.

The influence of leaf thickness on the CO2 transfer conductance and leaf stable carbon isotope ratio for some evergreen tree species in Japanese warm‐temperate forests

1. The influence of leaf thickness on internal conductance for CO2 transfer from substomatal cavity to chloroplast stroma (gi) and carbon isotope ratio (δ13C) of leaf dry matter was investigated for some evergreen tree species from Japanese temperate forests. gi was estimated based on the combined measurements of gas exchange and concurrent carbon isotope discrimination. 2. Leaves with thicker mesophyll tended to have larger leaf dry mass per area (LMA), larger surface area of mesophyll cells exposed to intercellular air spaces per unit leaf area (Smes) and smaller volume ratio of intercellular spaces to the whole mesophyll (mesophyll porosity). 3. g i of these leaves was correlated positively to Smes but negatively to mesophyll porosity. The variation in gi among these species would be therefore primarily determined by variation of the conductance in liquid phase rather than that in gas phase. 4.δ13C was positively correlated to mesophyll thickness and leaf nitrogen content on an area basis. However, gi values did not correlate to δ13C. These results suggest that difference in δ13C among the species was not caused by the variation in gi, but mainly by the difference in long‐term photosynthetic capacity. 5. Comparison of our results with those of previous studies showed that the correlation between leaf thickness and gi differed depending on leaf functional types (evergreen, deciduous or annual). Differences in leaf properties among these functional types were discussed.

In situ estimation of net CO2 assimilation, photosynthetic electron flow and photorespiration in Turkey oak (Q. cerris L.) leaves: diurnal cycles under different levels of water supply

ABSTRACTDiurnal time courses of net CO2 assimilation rates, stomatal conductance and light‐driven electron fluxes were measured in situ on attached leaves of 30‐year‐old Turkey oak trees (Quercus cerris L.) under natural summer conditions in central Italy. Combined measurements of gas exchange and chlorophyll a fluorescence under low O2 concentrations allowed the demonstration of a linear relationship between the photochemical efficiency of PSII (fluorescence measurements) and the apparent quantum yield of gross photosynthesis (gas exchange). This relationship was used under normal O2 to compute total light‐driven electron fluxes, and to partition them into fractions used for RuBP carboxylation or RuBP oxygenation. This procedure also yielded an indirect estimate of the rate of photorespiration in vivo. The time courses of light‐driven electron flow, net CO2 assimilation and photorespiration paralleled that of photosynthetic photon flux density, with important afternoon deviations as soon as a severe drought stress occurred, whereas photochemical efficiency and maximal fluorescence underwent large but reversible diurnal decreases. The latter observation indicated the occurrence of a large non‐photochemical energy dissipation at PSII. We estimated that less than 60% of the total photosynthetic electron flow was used for carbon assimilation at midday, while about 40% was devoted to photorespiration. The rate of carbon loss by photorespiration (R1) reached mean levels of 56% of net assimilation rates. The potential application of this technique to analysis of the relative contributions of thermal de‐excitation at PSII and photorespiratory carbon recycling in the protection of photosynthesis against stress effects is discussed.

The Relationship Between CO2 Transfer Conductance and Leaf Anatomy in Transgenic Tobacco With a Reduced Content of Rubisco

The CO2 transfer conductance in leaves quantifies the ease with which CO2 can diffuse from sub-stomatal cavities to sites of carboxylation within the chloroplast. The aim of this work was to test the hypothesis that the CO2 transfer conductance is proportional to the surface area of chloroplasts exposed to intercellular airspaces. We compared two genotypes, wild-type and transgenic tobacco, that had been transformed with an antisense gene directed at the mRNA of the Rubisco small subunit. Transgenic tobacco had lower rates of CO2 assimilation than wild-type but similar chlorophyll contents. Leaf anatomy was altered by growing plants in two different environments: a high daily irradiance in a growth cabinet (12 h photoperiod of 1 mmol quanta m-2 s-1) and a sunlit glasshouse. The growth cabinet gave at least twice the daily irradiance compared to the glasshouse. The CO2 transfer conductance was calculated from combined measurements of gas exchange and carbon isotope discrimination measured in 2% oxygen. Following gas exchange measurement, leaves were sampled for biochemical and anatomical measure- ment. In transgenic tobacco plants, Rubisco content was 35% of that found in the wild-type tobacco, the CO2 assimilation rate was 50% of the wild-type rate and the chlorophyll content was unaltered. While leaf mass per unit leaf area of transgenic tobacco was 82% of that of the wild-type, differences in leaf thickness and surface area of mesophyll cells exposed to intercellular airspace per unit leaf area (Smes) were small (92 and 87% of wild-type, respectively). Leaves grown in the growth cabinet under high daily irradiance were thicker (63%), had a greater Smes (41%) due to the development of thicker palisade tissue, had higher photosynthetic capacity (27%) and contained more chlorophyll (58%) and Rubisco (77%), than leaves from plants grown in the glasshouse. Irrespective of genotype or growth environment, CO2 transfer conductance varied in proportion to surface area of chloroplasts exposed to intercellular airspaces. While the method for calculating CO2 transfer conductance could not distinguish between limitations due to the gas or liquid phases, there was no reduction in CO2 transfer conductance associated with more closely packed cells, thicker leaves, nor with increasing chloroplast thickness in tobacco.