A method of monitoring and collecting CO samples in the field has been developed which has been used to study both temporal and spatial variations in canopy CO isotopic signatures in two contrasting tropical forest formations in Trinidad. These have been related to vertical gradients in the carbon isotope ratio (δC) of organic material in conjunction with measurements of other environmental parameters. The δC of leaf material from two canopies showed a gradient with respect to height, more negative values being found low in the understorey. The deciduous secondary forest, (Simla) showed a difference of 4.6‰ and the semi-evergreen seasonal canopy (Aripo), 2.8‰. The range of δC values at Simla was 4‰ less negative than those at Aripo. In order to relate these measurements to the interaction between diffusion or carboxylation limitation, and source CO effects, variations in environmental parameters through the canopy have been compared with changes in CO partial pressure (P ) and isotopic composition δC throughout the day during the dry season. Values of P 20 m above the ground at Aripo varied from 380 vpm at dawn to 340 vpm at midday, at which time the partial pressure 15 cm above the ground was 375 vpm. The CO partial pressure did not stabilise during the course of the day, and there was good correlation (r =0.82) between δ and P , with more negative values of δ occuring in the understorey. Diuraal changes of 2‰ were evident at all canopy positions. In the more open canopy at Simla, these gradients were similar, but less marked. Leaf-air vapour pressure deficit (VPD) showed no relationship with height, possibly as a result of minimal water flux from both the soil and the canopy due to low soil water content; VPD was 1.5 kPa higher at midday than dawn. A 3° C temperature gradient between the understorey and upper canopy was observed at Aripo but not in the more open Simla canopy. CO partial pressure stabilised for only 4 h in the middle of the day, while other parameters showed no stable period. The proportion of floor respired CO reassimilated at Aripo has been calculated as 26%, 19%, and 8% for the periods 0600-1000, 1000-1400, and 1400-1800 hours. In order to quantify source CO effects, measurements of the environmental parameters and assimilation rate must be made at all canopy positions and throughout the day.

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SUMMARYA study was made of photosynthesis and expression of crassulacean acid metabolism (CAM) in naturally exposed and shaded populations of Clusia minor L. during the transition from wet to dry season in Trinidad (mid-February to mid- April, 1990). At the start of the dry season, plants from exposed and shaded habitats showed a capacity for CAM either through the fixation of external or internal (respiratory) CO.^. Exposed plants showed continuous uptake of CO^ over 24 h although dark fixation accounted for only a small proportion of CO^ fixed over the day. The expression of CAM was considerably enhanced as the dry season progressed with substantial increases in the overnight accumulation of titratable acidity, particularly in leaves of exposed plants. This was accompanied by a reduction in day-time photosynthesis and an increase m dark fixation, with shaded plants showing only night-time fixation of CO,^. The magnitude of CAM in C. minor was substantial with a maximum AH" of 1410 mol m"'' measured in leaves from exposed branches. Both malic and citric acids were accumulated overnight. The highest citric.malic acid ratios were found in young leaves from exposed plants with 250 mol m~' * malic and 125 mol m'ĉ itric acid accumulated near the time of maximum CAM activity. Photosynthetic efficiency, measured as light responses of O^ evolution, also varied on a daily basis dependent on the incident photosynthetic photon flux density (PPFD). Apparent quantum yield and photosynthetic capacity showed marked reductions depending on the degree of exposure, suggesting that photoinhibitory responses are important under natural conditions. An analysis of three members of the Clusiaceae endemic to Trmidad showed that each had the capacity to induce CAM activity, despite being found in a narrow range of habitats which have higher rainfall than those of C. minor. However, despite the variable expression of CAM activity, carbon isotope composition suggested that when integrated throughout the year, carbon accumulation is predominantly mediated via the C^ pathway in all the species studied.

Carbon-isotope ratios (PCs) were measured for various biochemical fractions quantitatively extracted from naturally exposed and shaded leaves of the C3-Crassulacean acid metabolism (CAM) intermediate C h i a minor, sampled at dawn and dusk on days during the wet and dry seasons in Trinidad. As the activity of CAM increased in response to decreased availability of water and higher photon flux density, organic acids and soluble sugars were enriched in 13C by approximately 3.5 to 4%0 compared to plants sampled during the wet season. The induction of CAM was accompanied by a doubling in size of the reserve carbohydrate pools. Moreover, stoichiometric measurements indicated that degradation of both chloroplastic reserves and soluble sugars were necessary to supply phosphoenolpyruvate for the synthesis of organic acids at night.Results also suggest that two pools of soluble sugars exist in leaves of C. minor that perform CAM, one a vacuolar pool enriched in 13C and the second a transport pool depleted in 13C. Estimates of carbon-isotope discrimination expressed during CAM, derived from the trafficking among inorganic carbon, organic acids, and carbohydrate pools overnight, ranged from 0.9 to 3.1%0. The 613C of structural material did not change significantly between wet and dry seasons, indicating that most of the carbon used in growth was derived from C3 carboxylation.Clusia minor L. is a tropical dicotyledonous tree that shows extreme flexibility in regulating the relative amounts of C 0 2 fixed during the day and night in response to changes in the environment (Franco et al., 1990Borland et al., 1992Borland et al., , 1993Winter et al., 1992;Zotz and Winter, 1993). Under natural conditions, short-term changes in on-line, instantaneous A have illustrated how shifts in C3 and C4 carboxylation occur on a daily basis in C. minor, reflecting changes in environmental parameters (Borland et al., 1993). These variations in carboxylation pathway may in the first instance

On-line instantaneous carbon isotope discrimination was measured in conjunction with net uptake of CO in leaves of exposed and shaded plants of the C-CAM intermediate Clusia minor growing under natural conditions in Trinidad. At the end of the rainy season (late January-early February, 1992) C photosynthesis predominated although exposed leaves recaptured a small proportion of respiratory CO at night for the synthesis of malic acid. Citric acid was the major organic acid accumulated by exposed leaves at this time with a citric: malic acid ratio of 11:1. Values of instantaneous discrimination (Δ) in exposed leaves during the wet season rose from 17.1‰ shortly after dawn to 22.7‰ around mid-day just before stomata closed, suggesting that most CO was fixed by Rubisco at this time. During the late afternoon, instantaneous Δ declined from 22.2‰ to 17‰, probably reflecting the limited contribution from PEPc activity and an increase in diffusional resistance to CO in exposed leaves. Shaded leaves showed no CAM activity and CO uptake proceeded throughout the day in the wet season. The decrease in instantaneous Δ from 27‰ in the morning to 19.2‰ in the late afternoon was therefore entirely due to diffusional limitation. Leaves sampled in the dry season (mid-March, 1992) had by now induced full CAM activity with both malic and citric acids accumulated overnight and stomata closed for 4-5 h over the middle of the day. Values of instantaneous Δ measured over the first 3 h after dawn (6.4-9.1‰) indicated that C carboxylation dominated CO uptake for most of the morning when rates of photosynthesis were maximal, implying that under natural conditions, the down regulation of PEPc in phase II occurs much more slowly than laboratory-based studies have suggested. The contribution from C carboxylation to CO uptake during phase II was most marked in leaves which accumulated lower quantities of organic acids overnight. In exposed leaves, measurements of instantaneous Δ during the late afternoon illustrated the transition from C to C carboxylation with stomata remaining open during the transition from dusk into the dark period. Uptake of CO by shaded leaves during the late afternoon however appeared to be predominantly limited by decreased stomatal conductance. The short-term measurements of instantaneous Δ were subsequently integrated over 24 h in order to predict the leaf carbon isotope ratios (δ) and to compare this with the δ measured for leaf organic material. Whilst there was close agreement between predicted and measured δ for plants sampled in the wet season, during the dry season the predicted carbon isotope ratios were 5-9‰ higher than the measured isotope ratios. During the annual cycle of leaf growth most carbon was fixed via the C pathway although CAM clearly plays an important role in maintaining photochemical integrity in the dry season.