Measuring photosynthetic parameters in individual algal cells by Fast Repetition Rate fluorometry

A Single Cell Fast Repetition Rate (SCFRR) fluorometer was developed to measure the quantum yield of photochemistry, the functional absorption cross section of PS II and the kinetics of electron transport on the acceptor side of PS II in individual algal cells. These parameters are used to quantify the cell-specific photosynthetic performance in natural phytoplankton assembledges in aquatic ecosystems. The SCFRR technique measures chlorophyll fluorescence transients induced by a precisely controlled series of excitation flashlets that cumulatively saturate PS II within 120 s. To meet the requirement in the analysis for single algal cells, the measurements are conducted in micro volumes, such that the probability of probing more than one cell at a time is vanishingly low. We designed a novel, computer-controlled hydromechanical system to deliver a portion of the sample into the measuring chamber and, following measurement, remove it into one of six sorting containers. The fluorescence signal is induced by a series of high frequency flashlets obtained from high luminosity blue light-emitting diodes and is acquired by a novel red-sensitive PMT-based detection system exhibiting both high sensitivity and a very wide dynamic range. The wide dynamic range of the detector allows SCFRR measurements for a wide variety of cell sizes ranging from 1 to 100 m equivalent spherical diameter. The compact and light-weight design makes the SCFRR Fluorometer applicable for both laboratory and field studies.     

The contribution of phytoplankton productivity in turbid freshwaters to their trophic status

Certain factors influencing phytoplankton productivity are accentuated in turbid waters. They include mixing, spectral quality shifts, scattering, light fluctuations, and overall light attenuation. Measurements of productivity is influenced by the presence of inorganic turbidity. Together with the above factors high turbidity causes difficulties to assess and model phytoplankton productivity. Estimations of ^B, P_m ^B, I_k and _m only reflect on the physiological condition of the phytoplankton, which differs little between water types of temperate regions. Measurement of integral vertical productivity, efficiency and fractional absorption by the phytoplankton of light energy conversion, however, are greatly influenced by inorganic turbidity. Because of high ratios of mixing to euphotic depth, the critical mixing depth is one of the most important factors influencing overall productivity in turbid waters.     

Seasonal and spatial patterns in phytoplankton photosynthetic parameters in a tidal freshwater river

The photosynthetic response to irradiance wasquantified for phytoplankton from the tidalfreshwater Potomac River biweekly to monthly over aperiod of six years. Samples were collected from twoshallow embayments and portions of the deeper rivermainstem. Photosynthetic rate was measured in thelaboratory at in situ temperature over a range ofirradiance levels and photosynthetic parameters werecalculated using nonlinear regression.P^B _max,the maximum photosynthetic ratestandardized to chlorophyll a, increased withtemperature up to 25 °C with a Q₁₀ of 2.02. Above 25 °C, P^B _max was essentiallyconstant with temperature. Lesser correlationbetween P^B _max and ambient irradiance couldbe explained by the correlation of irradiance withtemperature. , the slope of the P–I curve atlow light, was correlated with both ambientirradiance and temperature. Highest valueswere found in late summer when high temperature andintermediate ambient irradiance were observed. Spring and early summer were characterized by low. Despite low light penetration, I_k and values were indicative of sun limitationpossibly due to intermittent high light levelsexperienced during mixing. I_k showed a clearseasonal trend directly related to days from summersolstice. Spatial patterns were minimal except thatI_k was consistently lower in one shallowembayment than in the other two areas. Seasonalpatterns in photosynthetic parameters correspondedroughly to changes from a spring diatom populationto summer cyanobacterial assemblage.     

Thermal stratification, nutrient dynamics, and phytoplankton productivity during the onset of spring phytoplankton growth in Lake Baikal, Russia

Lake Baikal, Russian Siberia, was sampled in July 1990 during the period of spring mixing and initiation of thermal stratification. Vertical profiles of temperature, dissolved nutrients (nitrate and soluble reactive phosphorus), phytoplankton biomass, and primary productivity were determined in an eleven-station transect encompassing the entire 636 km length of the lake. Pronounced horizontal variability in hydrodynamic conditions was observed, with the southern region of the lake being strongly thermally stratified while the middle and north basins were largely isothermal through July. The extent of depletion of surface water nutrients, and the magnitude of phytoplankton biomass and productivity, were found to be strongly correlated with the degree of thermal stratification. Horizontal differences likely reflected the contribution of two important factors: variation in the timing of ice-out in different parts of the lake (driving large-scale patterns of thermal stratification and other limnological properties) and localized effects of river inflows that may contribute to the preliminary stabilization of the water column in the face of intense turbulent spring mixing (driving meso-scale patterns). Examination of the relationships between surface water inorganic N and P depletion suggested that during the spring and early summer, phytoplankton growth in unstratified portions of the lake was largely unconstrained by nutrient supplies. As summer progressed, the importance of co-limitation by both N and P became more apparent. Uptake and regeneration rates, measured directly using the stable isotope ¹⁵N, revealed that phytoplankton in stratified portions of the lake relied primarily on NH₄ as their N source. Rates of NH₄ regeneration were in approximate equilibrium with uptake; both processes were dominated by organisms <2 µm. This pattern is similar to that observed for oligotrophic marine systems. Our study underscores the importance of hydrodynamic conditions in influencing patterns of biological productivity and nutrient dynamics that occur in Lake Baikal during its brief growing season.     

Physiological state of phytoplankton communities in the Southwest Atlantic sector of the Southern Ocean, as measured by fast repetition rate fluorometry

The majority of the Southern Ocean is a high-nutrient low-chlorophyll (HNLC) ecosystem. Localized increases in chlorophyll concentration measured in the wake of bathymetric features near South Georgia demonstrate variations in the factors governing the HNLC condition. We explore the possibility that the contrast between these areas of high-chlorophyll and surrounding HNLC areas is associated with variations in phytoplankton photophysiology. Total dissolvable iron concentrations, phytoplankton photophysiology and community structure were investigated in late April 2003 on a transect along the North Scotia Ridge (53–54°S) between the Falkland Islands and South Georgia (58–33°W). Total dissolvable iron concentrations suggested a benthic source of iron near South Georgia. Bulk community measurements of dark-adapted photochemical quantum efficiency (F _v/F _m) exhibited a sharp increase to the east of 46°W coincident with a decrease in the functional absorption cross-section (σ_PSII). Phytoplankton populations east of 46°W thus displayed no physiological symptoms of iron or nitrate stress. Contrasting low F _v/F _m west of 46°W could not be explained by variations in the macronutrients nitrate and silicic acid and may be the result of taxon specific variability in photophysiology or iron stress. We hypothesize that increased F _v/F _m resulted from local relief from iron-stress near South Georgia, east of Aurora Bank, an area previously speculated to be a “pulse point” source of iron. Our measurements provide one of the first direct physiological confirmations that iron stress is alleviated in phytoplankton populations near South Georgia.     

Detection of photosynthetic energy storage in a photosystem I reaction center preparation by photoacoustic spectroscopy

Thermal emission and photochemical energy storage were examined in photosystem I reaction center/core antenna complexes (about 40 Chl a/P700) using photoacoustic spectroscopy. Satisfactory signals could only be obtained from samples bound to hydroxyapatite and all samples had a low signal-to-noise ratio compared to either PS I or PS II in thylakoid membranes. The energy storage signal was saturated at low intensity (half saturation at 1.5 W m^-2) and predicted a photochemical quantum yield of >90%. Exogenous donors and acceptors had no effect on the signal amplitudes indicating that energy storage is the result of charge separation between endogenous components. Fe(CN)₆ ^-3 oxidation of P700 and dithionite-induced reduction of acceptors F_A-F_B inhibited energy storage. These data are compatible with the hypothesis that energy storage in PS I arises from charge separation between P700 and Fe-S centers F_A-F_B that is stable on the time scale of the photoacoustic modulation. High intensity background light (160 W m^-2) caused an irreversible loss of energy storage and correlated with a decrease in oxidizable P700; both are probably the result of high light-induced photoinhibition. By analogy to the low fluorescence yield of PS I, the low signal-to-noise ratio in these preparations is attributed to the short lifetime of Chl singlet excited states in PS I-40 and its indirect effect on the yield of thermal emission.Abbreviations FFT fast Föurier transform - HA hydroxyapatite - I₅₀ half saturation intensity for energy storage - PA photoacoustic - PS photosystem - PS I-40 photosystem I reaction center/core antenna complex containing about 40 Chl a/P700 - 201-1 photoacoustic energy storage signal - S/N signal-to-noise     

Effects of bottle size in determinations of primary productivity by phytoplankton

Determinations of primary productivity were made by the radiocarbon method at regular intervals over a year in samples, taken from the eutrophic waters of the Menai Strait, North Wales, and contained in bottles of capacities from 135 ml up to 2290 ml. Fixation per unit volume in particulate matter retained by 0 · 45 m pore size membrane filters was independent of bottle size over most of the year. There was a consistent divergence, however, during the period of rapid phytoplankton growth in the spring, when small bottles gave up to three times higher values than large ones. Possible effects of bottle size on primary productivity determinations are discussed but the cause of this particular result is not apparent.     

In vivo photosynthetic electron transport does not limit photosynthetic capacity in phosphate-deficient sunflower and maize leaves

The effects of extreme phosphate (Pi) deficiency during growth on the contents of adenylates and pyridine nucleotides and the in vivo photochemical activity of photosystem II (PSII) were determined in leaves of Helianthus annuus and Zea mays grown under controlled environmental conditions. Phosphate deficiency decreased the amounts of ATP and ADP per unit leaf area and the adenylate energy charge of leaves. The amounts of oxidized pyridine nucleotides per unit leaf area decreased with Pi deficiency, but not those of reduced pyridine nucleotides. This resulted in an increase in the ratio of reduced to oxidized pyridine nucleotides in Pi-deficient leaves. Analysis of chlorophyll a fluorescence at room temperature showed that Pi deficiency decreased the efficiency of excitation capture by open PSII reaction centres (φ_e), the in vivo quantum yield of PSII photochemistry (φ_PSII) and the photochemical quenching co-efficient (q_P), and increased the non-photochemical quenching co-efficient (q_N) indicating possible photoinhibitory damage to PSII. Supplying Pi to Pi-deficient sunflower leaves reversed the long-term effects of Pi-deficiency on PSII photochemistry. Feeding Pi-sufficient sunflower leaves with mannose or FCCP rapidly produced effects on chlorophyll a fluorescence similar to long-term Pi-deficiency. Our results suggest a direct role of Pi and photophosphorylation on PSII photochemistry in both long-and short-term responses of photosynthetic machinery to Pi deficiency. The relationship between φ_PSII and the apparent quantum yield of CO₂ assimilation determined at varying light intensity and 21 kPa O₂ and 35 Pa CO₂ partial pressures in the ambient air was linear in Pi-sufficient and Pi-deficient leaves of sunflower and maize. Calculations show that there was relatively more PSII activity per mole of CO₂ assimilated by the Pi-deficient leaves. This indicates that in these leaves a greater proportion of photosynthetic electrons transported across PSII was used for processes other than CO₂ reduction. Therefore, we conclude that in vivo photosynthetic electron transport through PSII did not limit photosynthesis in Pi-deficient leaves of sunflower and maize and that the decreased CO₂ assimilation was a consequence of a smaller ATP content and lower energy charge which restricted production of ribulose, 1-5, bisphosphate, the acceptor for CO₂.     

Inorganic carbon uptake by phytoplankton in vineyard sound,Massachusetts. I. Measurements of the photosynthesis-irradiance response of winter and early-spring assemblages

The photosynthesis-irradiance dependence of natural phytoplankton assemblages from surface waters of Vineyard Sound, Massachusetts, was investigated over a several month period during late winter —early spring, 1982, when water temperatures were ? 0.5 to 8.5°C. Maximal photosynthetic rates not only were consistently observed between 7–15% I₀, but were substantially higher than previously reported rates for cold-water assemblages, averaging 10–20 μg C · μg Chl a^?1 · h^?1. At higher light intensities photo-inhibition was severe and developed within minutes to tens of minutes of the start of the experiment. Several lines of evidence, however, suggest that photoinhibition in situ may be much less that that measured in incubations of > 30 min duration; residence time of phytoplankton at surface light intensities is sufficiently short, and adaptation of photosynthetic capacity appears to have occurred to approximately the mean depth of the water column. These results further highlight the importance of establishing the time-dependent photosynthetic responses of phytoplankton and the relationship to the physical mixing regime in estimating primary productivity.     

Intra-annual variation of phytoplankton community responses to factorial N,P, and CO2 enrichment in a temperate mesotrophic lake

1. Across primary producer communities in different lakes, nitrogen (N) and phosphorus (P) can exhibit many different patterns of limitation. Here, we look at the intra-annual variability of these patterns in a single lake. Furthermore, we investigate whether a third resource, carbon dioxide (CO₂) can have significant effects on phytoplankton biomass and community composition.
2. We performed five in situ lacustrine mesocosm experiments at different times of the year. In each experiment, we had a factorial design with two levels of N, P and CO₂ enrichment (no enrichment or double lake concentrations for N and P and atmospheric (400 ppm) and c. 1,000 ppm for CO₂) resulting in a total of eight treatments. Mesocosms of c. 1,600 L were suspended in a temperate, mesotrophic lake (Lac Hertel, Canada). Each experiment lasted 2 weeks and chlorophyll a biomass, coarse chemotaxonomic community composition (measured using fluorometry), and several environmental variables were recorded at a minimum of four time points.
3. We found that the limiting, synergistic, and community composition effects of N and P varied between experiments. TN:TP ratios explained, in part, some of this variability, along with insolation and water temperature.
4. Despite relatively high levels of CO₂ in the control mesocosms, we found a constant synergistic effect of CO₂ with N. In combination with the synergistic effect of P with N found in some experiments, this provides support for CO₂ as one of the multiple limiting resources in nutrient-enriched systems. This finding could have implications for eutrophic lakes exposed to increasing concentrations of CO₂.
5. We also found that the effects of CO₂ on community composition varied intra-annually. Thus, we conclude that generalised predictions about the effect of CO₂ on community composition at a coarse chemotaxonomic scale are unlikely to hold, but predictions specific to season and system are likely to be reliable.

     

地毯草的光合特性研究

地毯草是一种典型的暖季型草坪草。用Li 6400便携式光合分析仪测定地毯草的光合特性。地毯草的光合日进程呈单峰型,无"午休"现象且受非气孔限制,最大净光合速率出现在14∶00,变化趋势与光合有效辐射(PAR)、气温(Ta)、蒸腾速率(Tr)等因子相同,与空气相对湿度(RH)、胞间CO2浓度(Ci)等相反;其光饱合点超过2000μmol.m 2s 1,最大净光合速率达22μmol.m 2s 1,暗呼吸速率为1.34μmol.m 2s 1,CO2补偿点为0.97μmol.mol 1,光补偿点为22.2μmol.m 2s 1,羧化效率为0.2539μmol.m 2s 1,表观光量子效率高达0.0592μmol.mol 1;表明地毯草是一种具有较强耐荫性的C4型阳性的暖季型草坪草,除了在园林中可以广泛用作草坪地被外还可以在一些较荫湿的特殊地带作为绿化和防护之用。     

Measuring photosynthetic parameters at a distance: laser induced fluorescence transient (LIFT) method for remote measurements of photosynthesis in terrestrial vegetation

We have developed a laser induced fluorescence transient (LIFT) technique and instrumentation to remotely measure photosynthetic properties in terrestrial vegetation at a distance of up to 50 m. The LIFT method uses a 665 nm laser to project a collimated, 100 mm diameter excitation beam onto leaves of the targeted plant. Fluorescence emission at 690 nm is collected by a 250 mm reflective telescope and processed in real time to calculate the efficiency of photosynthetic light utilization, quantum efficiency of PS II, and the kinetics of photosynthetic electron transport. Operating with peak excitation power of 125 W m⁻², and duty cycle of 10–50%, the instrument conforms to laser safety regulations. The LIFT instrument is controlled via an Internet connection, allowing it to operate from remote locations or platforms. Here we describe the theoretical basis of the LIFT methodology, and demonstrate its applications in remote measurements of photosynthetic properties in the canopy of cottonwood and oak trees, and in the rosette of Arabidopsis mutants.     

Contrasting modes of regulation of PS II light utilization with changing irradiance in normal and psbS mutant leaves of Arabidopsis thaliana

Complementary techniques of chlorophyll a fluorescence, steady state CO₂ exchange, and O₂ release during a multiple turnover flash were applied to compare responses to irradiance for leaves of wild type and psbS mutants. The latter included variants in which the psbS gene was deleted (npq4-1) or possessed a single point mutation (npq4-9). Nonphotochemical quenching (NPQ) was reduced by up to 80 and 50%, respectively, in these lines at high irradiance. Analysis of changes in steady-state fluorescence yields and quantum yield of linear electron transport in the context of the reversible radical pair model of Photosystem II (PS II) indicated that NPQ occurs by nonradiative deactivation of chlorophyll singlet states in normal leaves. In contrast, application of the same criteria together with the observed irreversibility of NPQ and decline in density of functional PS II reaction centers following excessive illumination indicated a change in reaction center properties for the psbS deletion phenotype (Npq4-1^–). Specifically, PS II reaction centers in Npq4-1^– convert to a photochemically inactive, yet strongly quenching, form in intense light. The possibility of formation of a carotenoid or chlorophyll cation quencher in the reaction center is discussed. Results for the point mutant phenotype (Npq4-9^–) were intermediate to those of wild-type and Npq4-1^–. Furthermore, wild-type leaves exhibited a significant reversible increase in the PS II in vivo rate constant for photochemistry (k_P0) in saturating compared to limiting light. Changes in k_P0 could not be accounted for in terms of a classic phosphorylation-dependent (state transition) mechanism. Changes in k_P0 may arise from alternate pigment—protein conformations that alter the way excitons equilibrate among PS II chromophores. The lack of similar irradiance-dependent changes in k_P0 for the psbS mutants suggests a role for the PS II-S protein in the regulation of exciton distribution.This revised version was published online in October 2005 with corrections to the Cover Date.     

Chlorophyll fluorescence measured using the Fraunhofer line-depth principle and relationship to photosynthetic rate in the field

Abstract. A field study was conducted to determine the relationship of solar-excited chlorophyll a fluorescence to net CO₂ assimilation rate in attached leaves. The Fraunhofer line-depth principle was used to measure fluorescence at 656.3 nm wavelength while leaves remained exposed to full sunlight and normal atmospheric pressures of CO₂ and O₂. Fluorescence induction kinetics were observed when leaves were exposed to sunlight after 10 min in darkness. Subsequently, fluorescence varied inversely with assimilation rate. In the C₄ Zea mays , fluorescence decreased from 2.5 to 0.8 mW m^-2 nm^-1 as CO₂ assimilation rate increased from 1 to 8 μmol m^-2 s^-1 (r²= 0.520). In the C₃ Liquidambar styraciflua and Pinus taeda , fluorescence decreased from 6 to 2 mW m^-2 nm^-1 as assimilation rate increased from 2 to 5 or 0 to 2 μmol m^-2 s^-1 (r²= 0.44 and 0.45. respectively). The Fraunhofer line-depth principle enables the simultaneous measurement of solar-excited fluorescence and CO₂ assimilation rate in individual leaves, but also at larger scales. Thus, it may contribute significantly to field studies of the relationship of fluorescence to photosynthesis.     

Sequential assembly of photosynthetic units in Rhodobacter sphaeroides as revealed by fast repetition rate analysis of variable bacteriochlorophyll a fluorescence

The development of functional photosynthetic units in Rhodobacter sphaeroides was followed by near infra-red fast repetition rate (IRFRR) fluorescence measurements that were correlated to absorption spectroscopy, electron microscopy and pigment analyses. To induce the formation of intracytoplasmic membranes (ICM) (greening), cells grown aerobically both in batch culture and in a carbon-limited chemostat were transferred to semiaerobic conditions. In both aerobic cultures, a low level of photosynthetic complexes was observed, which were composed of the reaction center and the LH1 core antenna. Interestingly, in the batch cultures the reaction centers were essentially inactive in forward electron transfer and exhibited low photochemical yields F_V/F_M, whereas the chemostat culture displayed functional reaction centers with a rather rapid (1-2 ms) electron transfer turnover, as well as a high F_V/F_M of ∼0.8. In both cases, the transfer to semiaerobiosis resulted in rapid induction of bacteriochlorophyll a synthesis that was reflected by both an increase in the number of LH1-reaction center and peripheral LH2 antenna complexes. These studies establish that photosynthetic units are assembled in a sequential manner, where the appearance of the LH1-reaction center cores is followed by the activation of functional electron transfer, and finally by the accumulation of the LH2 complexes.     

Changes in timing of seasonal peak photosynthetic activity in northern ecosystems

Seasonality in photosynthetic activity is a critical component of seasonal carbon, water, and energy cycles in the Earth system. This characteristic is a consequence of plant's adaptive evolutionary processes to a given set of environmental conditions. Changing climate in northern lands (>30°N) alters the state of climatic constraints on plant growth, and therefore, changes in the seasonality and carbon accumulation are anticipated. However, how photosynthetic seasonality evolved to its current state, and what role climatic constraints and their variability played in this process and ultimately in carbon cycle is still poorly understood due to its complexity. Here, we take the “laws of minimum” as a basis and introduce a new framework where the timing (day of year) of peak photosynthetic activity (DOY_Pmax) acts as a proxy for plant's adaptive state to climatic constraints on its growth. Our analyses confirm that spatial variations in DOY_Pmax reflect spatial gradients in climatic constraints as well as seasonal maximum and total productivity. We find a widespread warming‐induced advance in DOY_Pmax (?1.66 ± 0.30 days/decade, p < 0.001) across northern lands, indicating a spatiotemporal dynamism of climatic constraints to plant growth. We show that the observed changes in DOY_Pmax are associated with an increase in total gross primary productivity through enhanced carbon assimilation early in the growing season, which leads to an earlier phase shift in land‐atmosphere carbon fluxes and an increase in their amplitude. Such changes are expected to continue in the future based on our analysis of earth system model projections. Our study provides a simplified, yet realistic framework based on first principles for the complex mechanisms by which various climatic factors constrain plant growth in northern ecosystems.     

Development of leaf photosynthetic parameters in Betula pendula Roth leaves: correlations with photosystem I density

The global modelling of photosynthesis is based on exact knowledge of the leaf photosynthetic machinery. The capacities of partial reactions of leaf photosynthesis develop at different rates, but it is not clear how the development of photoreactions and the Calvin cycle are co-ordinated. We investigated the development of foliar photosynthesis in the temperate deciduous tree Betula pendula Roth. using a unique integrated optical/gas exchange methodology that allows simultaneous estimation of photosystem I and II (PS I and PS II) densities per leaf area, interphotosystem electron transport activities, and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) kinetic properties. We combined these measurements with in vitro determinations of Rubisco, soluble protein and chlorophyll contents. We observed a strong increase in leaf photosynthetic capacity in developing leaves per leaf area, as well as per dry mass, that was paralleled by accumulation of leaf Rubisco. Enhanced mesophyll conductance was the outcome of increased carboxylation capacity and increased CO(2) diffusion conductance. However, Rubisco was only partly activated in the leaves, according to in vivo measurements of Rubisco kinetics. The amount of active Rubisco increased in proportion with development of PS I, probably through a direct link between Rubisco activase and PS I electron transport. Since the kinetics for post-illumination P700 re-reduction did not change, the synthesis of cytochrome b(6)f complex was also proportional to PS I. The synthesis of PS II began later and continued for several days after reaching the full PS I activity, but leaf chlorophyll was shared equally between the photosystems. Due to this, the antenna of PS II was very large and not optimally organized, leading to greater losses of excitation and lower quantum yields in young leaves. We conclude that co-ordinated development of leaf photosynthesis is regulated at the level of PS I with subordinated changes in PS II content and Rubisco activation.     

Photosynthesis and photoinhibition in leaves of chlorophyll b-less barley in relation to absorbed light

The response of photosynthesis to absorbed light by intact leaves of wild-type ( Hordeum vulgare L. cv. Gunilla) and chlorophyll b -less barley ( H. vulgare L. cv. Dornaria, chlorina-f2²⁸⁰⁰) was measured in a light integrating sphere. Up to the section where the light response curve bends most sharply the responses of the b -less and wild-type barley were similar but not identical. Average quantum yield and convexity for the mutant light response curves were 0.89 and 0.90, respectively, times those of the wild-type barley. The maximum quantum yield for PSII photochemistry was also 10% lower as indicated by fluorescence induction kinetics (F_v/F_m). Just above the region where the light curve bends most sharply, photosynthesis decreased with time in the mutant but not in the wild-type barley. This decrease was associated with a decrease in F_v/F_m indicating photoinhibition of PSII. This photoinhibition occurred in the same region of the light response curve where zeaxanthin formation occurs. Zeaxanthin formation occurred in both the chlorophyll b -less and wild-type leaves. However, the epoxidation state was lower in the mutant than in the wild-type barley. The results indicate that chlorophyll b -less mutants will have reduced photosynthetic production as a result of an increased sensitivity to photoinhibition and possibly a lowered quantum yield and convexity in the absence of photoinhibition.