Laisk Method Research Articles

Estimating leaf day respiration from conventional gas exchange measurements.

Leaf day respiration (Rd ) strongly influences carbon-use efficiencies of whole plants and the global terrestrial biosphere. It has long been thought that Rd is slower than respiration in the dark at a given temperature, but measuring Rd by gas exchange remains a challenge because leaves in the light are also photosynthesizing. The Kok method and the Laisk method are widely used to estimate Rd . We highlight theoretical limitations of these popular methods, and recent progress toward their improvement by using additional information from chlorophyll fluorescence and by accounting for the photosynthetic reassimilation of respired CO2 . The latest evidence for daytime CO2 and energy release from the oxidative pentose phosphate pathway in chloroplasts appears to be important to understanding Rd .

Laisk measurements in the nonsteady state: Tests in plants exposed to warming and variable CO2 concentrations.

Light respiration (RL) is an important component of plant carbon balance and a key parameter in photosynthesis models. RL is often measured using the Laisk method, a gas exchange technique that is traditionally employed under steady-state conditions. However, a nonsteady-state dynamic assimilation technique (DAT) may allow for more rapid Laisk measurements. In 2 studies, we examined the efficacy of DAT for estimating RL and the parameter Ci* (the intercellular CO2 concentration where Rubisco's oxygenation velocity is twice its carboxylation velocity), which is also derived from the Laisk technique. In the first study, we compared DAT and steady-state RL and Ci* estimates in paper birch (Betula papyrifera) growing under control and elevated temperature and CO2 concentrations. In the second, we compared DAT-estimated RL and Ci* in hybrid poplar (Populus nigra L. × P. maximowiczii A. Henry "NM6") exposed to high or low CO2 concentration pre-treatments. The DAT and steady-state methods provided similar RL estimates in B. papyrifera, and we found little acclimation of RL to temperature or CO2; however, Ci* was higher when measured with DAT compared to steady-state methods. These Ci* differences were amplified by the high or low CO2 pre-treatments. We propose that changes in the export of glycine from photorespiration may explain these apparent differences in Ci*.

CO<sub>2</sub> Demand-Supply Coordination in Photosynthesis Reflecting the Plant-Environment Interaction: Extension and Parameterization of Demand Function and Supply Function

Photosynthesis consists of a biochemical process named demand and a CO2 diffusion process named supply function. The intersection (Ci, An) at equal to the demand function and the supply function reflects a steady state of the plant subjected to the environment. The intersections of these demand-supply functions under different photosynthetically active radiation (PAR) can be fitted to a regression line (names DSF) in which slope (ΔAn/ΔCi) can be defined as dsf. We found that DSF information was embedded in both Laisk method (CO2 response curve (A/Ci) measured at three sub-saturated PARs, and their intersections were used to estimate daytime respiration (Rd), and CO2 compensation point (Ci*) and light response curve measurements, which could be used to estimate dsf values. This study investigated the relationship between dsf and the parameters related to the biochemical process and the CO2 diffusion process of photosynthesis. The results showed that dsf was negatively correlated with gs, apparent carboxylation efficiency, and apparent quantum yield. This suggests that DSF may coordinate the influence of environmental conditions (light, CO2 and water) on photosynthesis in the biochemical and CO2 diffusion process. Moreover, dsf was independent of gas exchange measurement conditions and showed species specificity. In conclusion, we speculated that dsf seems to be a comprehensive parameter that might be related to the intrinsic adaptation mechanism of plants to environmental conditions. We proposed an auxiliary line perpendicular to the DSF and used it to improve the stability of Ci* and Rd estimated from the Laisk dataset.

Using a reaction-diffusion model to estimate day respiration and reassimilation of (photo)respired CO2 in leaves.

Summary Methods using gas exchange measurements to estimate respiration in the light (day respiration Rd) make implicit assumptions about reassimilation of (photo)respired CO 2; however, this reassimilation depends on the positions of mitochondria.We used a reaction‐diffusion model without making these assumptions to analyse datasets on gas exchange, chlorophyll fluorescence and anatomy for tomato leaves. We investigated how Rd values obtained by the Kok and the Yin methods are affected by these assumptions and how those by the Laisk method are affected by the positions of mitochondria.The Kok method always underestimated Rd. Estimates of Rd by the Yin method and by the reaction‐diffusion model agreed only for nonphotorespiratory conditions. Both the Yin and Kok methods ignore reassimilation of (photo)respired CO 2, and thus underestimated Rd for photorespiratory conditions, but this was less so in the Yin than in the Kok method. Estimates by the Laisk method were affected by assumed positions of mitochondria. It did not work if mitochondria were in the cytosol between the plasmamembrane and the chloroplast envelope. However, mitochondria were found to be most likely between the tonoplast and chloroplasts.Our reaction‐diffusion model effectively estimates Rd, enlightens the dependence of Rd estimates on reassimilation and clarifies (dis)advantages of existing methods.

Acclimation of light and dark respiration to experimental and seasonal warming are mediated by changes in leaf nitrogen in Eucalyptus globulus.

Quantifying the adjustments of leaf respiration in response to seasonal temperature variation and climate warming is crucial because carbon loss from vegetation is a large but uncertain part of the global carbon cycle. We grew fast-growing Eucalyptus globulus Labill. trees exposed to +3 °C warming and elevated CO2 in 10-m tall whole-tree chambers and measured the temperature responses of leaf mitochondrial respiration, both in light (RLight) and in darkness (RDark), over a 20-40 °C temperature range and during two different seasons. RLight was assessed using the Laisk method. Respiration rates measured at a standard temperature (25 °C - R25) were higher in warm-grown trees and in the warm season, related to higher total leaf nitrogen (N) investment with higher temperatures (both experimental and seasonal), indicating that leaf N concentrations modulated the respiratory capacity to changes in temperature. Once differences in leaf N were accounted for, there were no differences in R25 but the Q10 (i.e., short-term temperature sensitivity) was higher in late summer compared with early spring. The variation in RLight between experimental treatments and seasons was positively correlated with carboxylation capacity and photorespiration. RLight was less responsive to short-term changes in temperature than RDark, as shown by a lower Q10 in RLight compared with RDark. The overall light inhibition of R was ∼40%. Our results highlight the dynamic nature of leaf respiration to temperature variation and that the responses of RLight do not simply mirror those of RDark. Therefore, it is important not to assume that RLight is the same as RDark in ecosystem models, as doing so may lead to large errors in predicting plant CO2 release and productivity.

Day and night respiration of three tree species in a temperate forest of northeastern China

Leaf day respiration is one of the most fundamental plant physiological processes and plays a vital role in the plant carbon cycle. However, day respiration is inherently complex and difficult to measure. In this study, the Kok method and the Laisk method were used to measure leaf day respiration for the saplings of one evergreen conifer species (Pinus koraiensis) and two deciduous broadleaved species (Tilia amurensis and Fraxinus mandshurica) in a temperate forest. Results show that discrepancy between the corrected day respiration values estimated by the Kok and Laisk methods was only 4% for the three tree species. On average, day respiration was 55.9% and 52.6% lower compared to night respiration for the three tree species, as measured by the Kok and Laisk method, respectively. Day respiration of the evergreen conifer species estimated by the Kok method was 31.7% lower while that estimated by the Laisk method was 36.8% lower than that of the deciduous broadleaved species. Night respiration of the evergreen conifer trees was 40.7% lower than those of the deciduous broadleaved trees. Day respiration rate is positively correlated with night respiration rate. Notably, day respiration rate decreased with increased photosynthetic photon flux density, and even a small amount of light significantly inhibited leaf day respiration in all three species.

Estimating daytime ecosystem respiration to improve estimates of gross primary production of a temperate forest.

Leaf respiration is an important component of carbon exchange in terrestrial ecosystems, and estimates of leaf respiration directly affect the accuracy of ecosystem carbon budgets. Leaf respiration is inhibited by light; therefore, gross primary production (GPP) will be overestimated if the reduction in leaf respiration by light is ignored. However, few studies have quantified GPP overestimation with respect to the degree of light inhibition in forest ecosystems. To determine the effect of light inhibition of leaf respiration on GPP estimation, we assessed the variation in leaf respiration of seedlings of the dominant tree species in an old mixed temperate forest with different photosynthetically active radiation levels using the Laisk method. Canopy respiration was estimated by combining the effect of light inhibition on leaf respiration of these species with within-canopy radiation. Leaf respiration decreased exponentially with an increase in light intensity. Canopy respiration and GPP were overestimated by approximately 20.4% and 4.6%, respectively, when leaf respiration reduction in light was ignored compared with the values obtained when light inhibition of leaf respiration was considered. This study indicates that accurate estimates of daytime ecosystem respiration are needed for the accurate evaluation of carbon budgets in temperate forests. In addition, this study provides a valuable approach to accurately estimate GPP by considering leaf respiration reduction in light in other ecosystems.

Artefactual responses of mesophyll conductance to CO2 and irradiance estimated with the variable J and online isotope discrimination methods

Studies with the variable J method have reported that mesophyll conductance (gm ) rapidly decreases with increasing intercellular CO2 partial pressures (Ci ) or decreasing irradiance. Similar responses have been suggested with the online isotope discrimination method, although with less consistency. Here we show that even when the true gm is constant, the variable J method can produce an artefactual dependence of gm on Ci or irradiance similar to those reported in previous studies for any of the following factors: day respiration and chloroplastic CO2 photocompensation point are estimated with Laisk method; Ci or electron transport rate is positively biased; net photosynthetic rate is negatively biased; insufficient NADPH is assumed while insufficient ATP limits RuBP regeneration. The isotopic method produces similar artefacts if fractionation of carboxylation or Ci is positively biased or Δ(13) negatively biased. A non-zero chloroplastic resistance to CO2 movement results in a qualitatively different dependence of gm on Ci or irradiance and this dependence is only sensitive at low Ci . We thus cannot rule out the possibility that previously reported dependence of gm on Ci or irradiance is a methodological artefact. Recommendations are made to take advantage of sensitivities of the variable J and isotopic methods for estimating gm .

Influence of temperature on measurements of the CO2 compensation point: differences between the Laisk and O2-exchange methods.

The CO2 compensation point in the absence of day respiration (Γ*) is a key parameter for modelling leaf CO2 exchange. Γ* links the kinetics of ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) with the stoichiometry of CO2 released per Rubisco oxygenation from photorespiration (α), two essential components of biochemical models of photosynthesis. There are two main gas-exchange methods for measuring Γ*: (i) the Laisk method, which requires estimates of mesophyll conductance to CO2 (g m) and (ii) measurements of O2 isotope exchange, which assume constant values of α and a fixed stoichiometry between O2 uptake and Rubisco oxygenation. In this study, the temperature response of Γ* measured using the Laisk and O2-exchange methods was compared under ambient (25 °C) and elevated (35 °C) temperatures to determine whether both methods yielded similar results. Previously published temperature responses of Γ* estimated with the Laisk and O2-exchange methods in Nicotiana tabacum demonstrated that the Laisk-derived model of Γ* was more sensitive to temperature compared with the O2-exchange model. Measurements in Arabidopsis thaliana indicated that the Laisk and O2-exchange methods produced similar Γ* at 25 °C; however, Γ* values from O2 exchange were lower at 35 °C compared with the Laisk method. Compared with a photorespiratory mutant (pmdh1pmdh2hpr) with increased α, wild-type (WT) plants had lower Laisk values of Γ* at 25 °C but were not significantly different at 35 °C. These differences between Laisk and O2 exchange values of Γ* at 35 °C could be explained by temperature sensitivity of α in WT and/or errors in the assumptions of O2 exchange. The differences between Γ* measured using the Laisk and O2-exchange method with temperature demonstrate that assumptions used to measure Γ*, and possibly the species-specific validity of these assumptions, need to be considered when modelling the temperature response of photosynthesis.

Evaluating a new method to estimate the rate of leaf respiration in the light by analysis of combined gas exchange and chlorophyll fluorescence measurements

Day respiration (Rd) is an important parameter in leaf ecophysiology. It is difficult to measure directly and is indirectly estimated from gas exchange (GE) measurements of the net photosynthetic rate (A), commonly using the Laisk method or the Kok method. Recently a new method was proposed to estimate Rd indirectly from combined GE and chlorophyll fluorescence (CF) measurements across a range of low irradiances. Here this method is tested for estimating Rd in five C3 and one C4 crop species. Values estimated by this new method agreed with those by the Laisk method for the C3 species. The Laisk method, however, is only valid for C3 species and requires measurements at very low CO2 levels. In contrast, the new method can be applied to both C3 and C4 plants and at any CO2 level. The Rd estimates by the new method were consistently somewhat higher than those by the Kok method, because using CF data corrects for errors due to any non-linearity between A and irradiance of the used data range. Like the Kok and Laisk methods, the new method is based on the assumption that Rd varies little with light intensity, which is still subject to debate. Theoretically, the new method, like the Kok method, works best for non-photorespiratory conditions. As CF information is required, data for the new method are usually collected using a small leaf chamber, whereas the Kok and Laisk methods use only GE data, allowing the use of a larger chamber to reduce the noise-to-signal ratio of GE measurements.

Temperature response of photosynthesis and internal conductance to CO2: results from two independent approaches

The internal conductance to CO(2) transfer from intercellular spaces to chloroplasts poses a major limitation to photosynthesis, but few studies have investigated its temperature response. The aim of this study was to determine the temperature response of photosynthesis and internal conductance between 10 degrees C and 35 degrees C in seedlings of a deciduous forest tree species, Quercus canariensis. Internal conductance was estimated via simultaneous measurements of gas exchange and chlorophyll fluorescence ("variable J method"). Two of the required parameters, the intercellular photocompensation point (C(i)*) and rate of mitochondrial respiration in the light (R(d)), were estimated by the Laisk method. These were used to calculate the chloroplastic photocompensation point (Gamma*) in a simultaneous equation with g(i). An independent estimate of internal conductance was obtained by a novel curve-fitting method based on the curvature of the initial Rubisco-limited portion of an A/C(i) curve. The temperature responses of the rate of Rubisco carboxylation (V(cmax)) and the RuBP limited rate of electron transport (J(max)) were determined from chloroplastic CO(2) concentrations. The rate of net photosynthesis peaked at 24 degrees C. C(i)* was similar to reports for other species with a C(i)* of 39 micromol mol(-1) at 25 degrees C and an activation energy of 34 kJ mol(-1). Gamma* was very similar to the published temperature response for Spinacia oleracea from 20 degrees C to 35 degrees C, but was slightly greater at 10 degrees C and 15 degrees C. J(max) peaked at 30 degrees C, whereas V(cmax) did not reach a maximum between 10 degrees C and 35 degrees C. Activation energies were 49 kJ mol(-1) for V(cmax) and 100 kJ mol(-1) for J(max). Both methods showed that internal conductance doubled from 10 degrees C to 20 degrees C, and then was nearly temperature-independent from 20 degrees C to 35 degrees C. Hence, the temperature response of internal conductance could not be fitted to an Arrhenius function. The best fit to estimated g(i) was obtained with a three-parameter log normal function (R(2)=0.98), with a maximum g(i) of 0.19 mol m(-2) s(-1) at 29 degrees C.

Leaf respiration of snow gum in the light and dark. Interactions between temperature and irradiance.

We investigated the effect of temperature and irradiance on leaf respiration (R, non-photorespiratory mitochondrial CO(2) release) of snow gum (Eucalyptus pauciflora Sieb. ex Spreng). Seedlings were hydroponically grown under constant 20 degrees C, controlled-environment conditions. Measurements of R (using the Laisk method) and photosynthesis (at 37 Pa CO(2)) were made at several irradiances (0-2,000 micromol photons m(-2) s(-1)) and temperatures (6 degrees C-30 degrees C). At 15 degrees C to 30 degrees C, substantial inhibition of R occurred at 12 micromol photons m(-2) s(-1), with maximum inhibition occurring at 100 to 200 micromol photons m(-2) s(-1). Higher irradiance had little additional effect on R at these moderate temperatures. The irradiance necessary to maximally inhibit R at 6 degrees C to 10 degrees C was lower than that at 15 degrees C to 30 degrees C. Moreover, although R was inhibited by low irradiance at 6 degrees C to 10 degrees C, it recovered with progressive increases in irradiance. The temperature sensitivity of R was greater in darkness than under bright light. At 30 degrees C and high irradiance, light-inhibited rates of R represented 2% of gross CO(2) uptake (v(c)), whereas photorespiratory CO(2) release was approximately 20% of v(c). If light had not inhibited leaf respiration at 30 degrees C and high irradiance, R would have represented 11% of v(c). Variations in light inhibition of R can therefore have a substantial impact on the proportion of photosynthesis that is respired. We conclude that the rate of R in the light is highly variable, being dependent on irradiance and temperature.

Leaf Respiration in Light and Darkness (A Comparison of Slow- and Fast-Growing Poa Species).

We investigated whether leaf dark respiration (nonphotorespiratory mitochondrial CO2 release) is inhibited by light in several Poa species, and whether differences in light inhibition between the species are related to differences in the rate of leaf net photosynthesis. Four lowland (Poa annua L., Poa compressa L., Poa pratensis L., and Poa trivialis L.), one subalpine (Poa alpina L.), and two alpine (Poa costiniana Vick. and Poa fawcettiae Vick.) Poa species differing in whole plant relative growth rates were grown under identical controlled conditions. Nonphotorespiratory mitochondrial CO2 release in the light (Rd) was estimated according to the Laisk method. Photosynthesis was measured at ambient CO2 partial pressure (35 Pa) and 500 [mu]mol photons m-2 s-1. The rate of photosynthesis per unit leaf mass was positively correlated with the relative growth rate, with the slow-growing alpine Poa species exhibiting the lowest photosynthetic rates. Rates of both Rd and respiration in darkness were also substantially lower in the alpine species. Nonphotorespiratory CO2 release in darkness was higher than Rd in all species. However, despite some variation between the species in the level of light inhibition of respiration, no relationship was observed between the level of inhibition and the rate of photosynthesis. Similarly, the level of inhibition was not correlated with the relative growth rate. Our results support the suggestion that rates of leaf respiration in the light are closely associated with rates in darkness.

Dark Leaf Respiration in Light and Darkness of an Evergreen and a Deciduous Plant Species.

Dark respiration in light as well as in dark was estimated for attached leaves of an evergreen (Heteromeles arbutifolia Ait.) and a deciduous (Lepechinia fragans Greene) shrub species using an open gas-exchange system. Dark respiration in light was estimated by the Laisk method. Respiration rates in the dark were always higher than in the light, indicating that light inhibited respiration in both species. The rates of respiration in the dark were higher in the leaves of the deciduous species than in the evergreen species. However, there were no significant differences in respiration rates in light between the species. Thus, the degree of inhibition of respiration by light was greater in the deciduous species (62%) than in the evergreen species (51%). Respiration in both the light and darkness decreased with increasing leaf age. However, because respiration in the light decreased faster with leaf age than respiration in darkness, the degree of inhibition of respiration by light increased with leaf age (from 36% in the youngest leaves to 81% in the mature leaves). This suggests that the rate of dark respiration in the light is related to the rate of biosynthetic processes. Dark respiration in the light decreased with increasing light intensity. Respiration both in the light and in the dark was dependent on leaf temperature. We concluded that respiration in light and respiration in darkness are tightly coupled, with variation in respiration in darkness accounting for more than 60% of the variation in respiration in light. Care must be taken when the relation between respiration in light and respiration in darkness is studied, because the relation varies with species, leaf age, and light intensity.

Comparison of Methods to Estimate Dark Respiration in the Light in Leaves of Two Woody Species.

Dark respiration in the light was estimated in leaves of two woody species (Heteromeles arbutifolia Ait. and Lepechinia fragans Greene) using two different approaches based on gas-exchange techniques: the Kok method and the Laisk method. In all cases, dark respiration in the light was lower (P < 0.05) than respiration in darkness, indicating that dark respiration was inhibited in the light. Rates of dark respiration in the light estimated by the Laisk method were 52% higher (P < 0.05) than those estimated by the Kok method. Differences between the methods could be explained by the low ambient CO2 concentrations required by the Laisk approach. The mean value of the inhibition of respiration by light for the two species, corrected for the ambient CO2 concentration effect, was 55%. Despite the differences in leaf characteristics between the species, values of the CO2 photocompensation point, at which the rate of photosynthetic CO2 uptake equaled that of photorespiratory CO2 evolution, were very constant, suggesting an excellent consistency in the results obtained with the Laisk approach.