Dark CO2-fixation and diurnal malic acid fluctuations in the submerged-aquatic Isoetes storkii

Summary In the leaves (but not corms) of the submerged aquatic Isoetes storkii malic acid concentration fluctuated from 22 eg g FW^-1 in the evening to 171 eg g FW^-1 in the morning. Associated with this was a change in titratable acidity of 152 eg g FW^-1 between morning and evening. ¹⁴C carbon was fixed in both the light and the dark, though the amount of carbon fixed in the light was more than that fixed in the dark. Autoradiographs show 88% of ¹⁴CO₂ fixed in the dark is recovered after 1 h, in malic acid and the remainder in one other unidentified product, whereas these two products contain less than 15% of the ¹⁴C fixed after 1 h exposure to ¹⁴CO₂ in the light. It is suggested that CAM metabolism in this aquatic species may be related to the low availability of CO₂ for photosynthesis during the day in its aquatic environment and that this metabolic pathway may prove common in the genus Isoetes.     

CO2-fixation Sites in Leaves of Maize and Oats

The primary site of photosynthetic carbon dioxide fixation wasdetermined in the leaves of two monocot species by means ofmicroautoradiographic techniques. Carbon fixation occurred primarilyin the vascular-bundle sheath cells in maize whereas fixationin oat occurred uniformly throughout the leaf. The evidencepresented suggests that the bundle sheath cells of maize fixcarbon via the C.4 dicarboxylic acid pathway.     

Control of PAR-saturated CO2 exchange rate in some C3 and CAM plants

We measured PAR-saturated CO₂ exchange rate (CER), and leaf N, P and chlorophyll (Chl) concentrations in 21 plant species, selected to encompass as broad a range in specific leaf area (SLA) as possible, and encompassing non-succulent C3 as well as succulent CAM plants. We worked with plants growing under uniform conditions in the facilities of a biological research station to ensure that any correlations found were due to inherent, genetically controlled, relationships between the measured parameters and not due to variations in resource availability in different habitats. We found CER to be strongly correlated to SLA, leaf N concentration and Chl concentration. CER increased much faster with increasing leaf N concentration (CER ≈ N^3.1) than with increasing SLA (CER ≈ SLA^1.2). CER also increased much faster with leaf N concentration than with increasing Chl concentration (CER ≈ Chl^1.3), indicating the photosynthetic N-use efficiency (NUE) to be higher for plants with high N concentration than for plants with low N concentration (NUE ≈ N^2.1). Analysis of covariance showed that these relationships exist even when comparing plants of widely different growth forms - succulent or non-succulent, and of different photosynthetic pathways, as the C3 and CAM plants compared here. Testing against scaling coefficients calculated using dimensional analysis, showed that the scaling of N, Chl and CER against SLA was not merely a result of diluting N and Chl with carbon in thicker leaves but that SLA, probably through influencing light absorptio and/or CO2 diffusion pathway, played an independent role in controlling CER. This revised version was published online in July 2006 with corrections to the Cover Date.     

Regulation of CO2-fixation in Chlorella by light of varied wavelengths and intensities

1) The wavelength effects on ¹⁴CO₂-fixation by Chlorella cellswere studied, using monochromatic light of different light intensities. 2) Blue light (453 mµ) stimulated the incorporation of¹⁴C into aspartate, glutamate and malate. Red light (679 mµ),on the other hand, stimulated its incorporation into P-esters,free sugars and insoluble material. 3) The blue light effect was observed in the presence of CMUat concentrations completely suppressing ordinary photosyntheticCO₂-fixation. 4) The blue light effect in the presence of CMU was inducedat very low intensities. At 453 mµ, 300 erg cm^–2sec^–1 was sufficient for complete saturation. 5) Time courses of ¹⁴C-incorporation into individual compoundswere investigated. Irrespective of the wavelength of the illuminatinglight, the first stable CO₂-fixation product formed under weaklight (400–500 erg cm^–2 sec^–1) was citrulline.At higher light intensities (4,000–7,000 erg cm^–2sec^–1), PGA was the first stable CO₂-fixation product.The incorporation of ¹⁴C into citrulline was not inhibited byCMU. 6) Experimental results indicate that both blue light-inducedincorporation of ¹⁴C into amino and organic acids and the incorporationof ¹⁴C into citrulline induced by low intensity light are operatedby a mechanism(s) independent of ordinary photosynthetic CO₂-fixation.Possible effects of light regulating the carbon metabolism inalgal cells are discussed. (Received July 24, 1969; )     

Capacity of enzymes of the euphorbiacea Aleurites montana involved in CO2-fixation, compared to plants having C3-, C4- and Crassulacean acid metabolism

Capacities of phosphoenolpyruvate carboxylase (PEP-Co), ribulose bisphosphate carboxylase (Rubisco), NADP+ malic enzyme (ME) and of malate dehydrogenase (MDH) were measured in the Euphorbiacea Aleurites montana, grown under 700 ppm CO2 for four weeks prior to enzyme extraction. For comparison Bryophyllum daigremontiana (CAM). Saccharum officinarum (C4) and Capsicum frutescens (C3) were treated in the same way. PEP-Co capacity of Aleurites was in the range of 12-, that of Capsicum approx. 26 nmol x min(-1) x mg protein(-1), without significant influence of the light period or CO2-treatment. In contrast, the activity of the enzyme from Saccharum was, depending on the duration of light, 160- respectively 96 times higher than that of the tung-oil tree. In Bryophyllum a rather low activity in the morning was increased during the day to approx. 230 nmol x min(-1) x mg protein(-1) in plants grown in the greenhouse and to approx. 115 nmol x min(-1) x mg protein(-1) in those from the growth chamber. Malate was hardly detectable in extracts of Aleurites, whereas it was high in Bryophyllum, depending on the light period. The ratio of average PEP-Co to Rub-Co capacity was high for the CAM-plant (20:1), somewhat lower for sugar cane (10:1), but almost at equality for Aleurites (0.9:1) and chilli (0.8:1). For the NADP+ malic enzyme, low capacity (20 to 28 nmol x min(-1) x mg protein(-1)) was found for Aleurites and for Capsicum, whereas it was 10 to 17 times higher in Saccharum. In Bryophyllum, the activity was up to 80 nmol x min(-1) x mg protein, dependent on light period. MDH capacity was extremely high in all plants investigated. Highest rates (10-20 micromol x min(-1) x mg protein(-1)), were obtained for Bryophyllum, followed by sugar cane and Capsicum with 5-8 micromol x min(-1) x mg protein(-1). Again, the lowest capacity was found in extracts of Aleurites with approx. 1.3 to 1.6 micromol x min(-1) x m protein(-1). Thus, in Aleurites montana no indication for C4- or Crassulacean acid metabolism was obtained. Therefore, the earlier observed very efficient uptake of CO2 cannot be explained by a high expression of the PEP-Co protein, known to occur in CAM- and C4-plants.     

Distribution of radioactivity in carbon atoms of malic acid formed during light-enhanced dark 14CO2-fixation in maize leaves

Most of the radioactivity incorporated into malic acid duringlight-enhanced dark ¹⁴CO₂-fixation was found in C-4, supportingour conclusion that phosphoenolpyruvic acid serves as a primaryacceptor of ¹⁴CO₂ to form the C₄ acid. ¹This work was reported at the 13th Annual Meeting of JapaneseSociety of Plant Physiologists, April, 1972. (Received January 6, 1973; )     

Effects of disalicylidenepropandiamine and near far-red light on 14CO2-fixation in Chlorella cells

1) With Chlorella ellipsoidea cells, in the presence of 5x10^–6M DSPD, or in its absence, the amounts of ¹⁴CO₂ incorporatedin P-esters, serine-plus-glycine and alanine were larger underred light than under blue light, whereas blue light specificallyincreased ¹⁴CO₂-incorporation in aspartate, glutamate, malateand fumarate (blue light effect). The amount of total ¹⁴C fixedunder blue or red light was greatly decreased by the additionof DSPD. When the concentration of DSPD was raised to 5x10^–4M, practically no radioactivity was found, under blue or redlight, in aspartate, glutamate and fumarate. Radioactivity inalanine was greatly increased. Effects of higher concentrationof DSPD are explained as due to the inhibition of PEP carboxylaseactivity in Chlorella cells. 2) The percentage incorporation of ¹⁴C into aspartate and theother compounds mentioned above, under near infra-red illuminationwas significantly smaller than that under blue light and wasalmost equal to that under red light. These results along withthe effect of 5x10^–6 M DSPD, exclude the possibility thatcyclic photophosphorylation is involved in the "blue light effect"mechanism. (Received December 12, 1969; )     

Changes in photosynthetic capability and carbohydrate production in an epiphytic CAM orchid plantlet exposed to super-elevated CO2

The effects on growth in super-elevated (1%) CO₂ in terms of photosynthetic capability and carbohydrate production were studied in an epiphytic CAM (Crassulacean acid metabolism) orchid plantlet, Mokara Yellow (Arachnis hookeriana×Ascocenda Madame Kenny). The growth of the plantlets was greatly enhanced after growing for 3 months at 1% CO₂ compared with the control plantlets (0.035% CO₂). CO₂ enrichment produced more than a 2-fold increase in dry matter production. The enhanced root growth at 1% CO₂ led to a higher root:shoot ratio. Plantlets grown at super-elevated CO₂ had higher F_v/F_m values, a higher photochemical quenching (q_P) and a relatively lower non-photochemical quenching (q_N). CO₂ at 1% appeared to enhance the utilization of captured light energy in the orchid plantlets. CO₂ enrichment also increased contents of soluble sugars (glucose and sucrose) and starch in the orchid plantlets. The extra starch formed under 1% CO₂ did not cause a disruption of the chloroplasts. Chlorophyll content was higher and a clear granal stacking was evident in young leaves and roots of plantlets grown at 1% CO₂. An extensive thylakoid system was observed in the young leaf chloroplasts of the CO₂-enriched plantlets indicating an improved development of the photosynthetic apparatus when compared to that of the control plantlets. The increased photosynthetic capacity and enhanced growth of the epiphytic roots under CO₂ enrichment would facilitate the generation of more photoassimilates and acquisition of essential resources, thereby increasing the survival rate of orchid plantlets under stressful field conditions.     

Effects of elevated CO2 concentration on seed production in C3 annual plants

The response of seed production to CO(2) concentration ([CO(2)]) is known to vary considerably among C(3) annual species. Here we analyse the interspecific variation in CO(2) responses of seed production per plant with particular attention to nitrogen use. Provided that seed production is limited by nitrogen availability, an increase in seed mass per plant results from increase in seed nitrogen per plant and/or from decrease in seed nitrogen concentration ([N]). Meta-analysis reveals that the increase in seed mass per plant under elevated [CO(2)] is mainly due to increase in seed nitrogen per plant rather than seed [N] dilution. Nitrogen-fixing legumes enhanced nitrogen acquisition more than non-nitrogen-fixers, resulting in a large increase in seed mass per plant. In Poaceae, an increase in seed mass per plant was also caused by a decrease in seed [N]. Greater carbon allocation to albumen (endosperm and/or perisperm) than the embryo may account for [N] reduction in grass seeds. These differences in CO(2) response of seed production among functional groups may affect their fitness, leading to changes in species composition in the future high-[CO(2)] ecosystem.     

Enzymatic synthesis of formic acid from H2 and CO2 and production of hydrogen from formic acid

Whole cells of Alcaligenes eutrophus (as well as isolated P. oxalaticus formate dehydrogenase and A. eutrophus hydrogenase coupled via NAD(+) or methyl viologen) have been shown to produce H(2) from formic acid. Immobilization of the cells in kappacarrageenan gel greatly enhances their stability at room temperature. The rate of hydrogen production catalyzed by immobilized A. eutrophus has been studied as a function of the concentrations of the cells and formate and also pH. An inhibition by high concentrations of formate has been found. Immobilized cells were also capable of synthesizingformate from H(2) and bicarbonate. Yields of formate up to 30% have been obtained. The catalytic efficiency of immobilized A. eutrophus cells was compared with that of palladium adsorbed on activated carbon.     

A simple apparatus for the continuous monitoring of 14CO2 production from several small reaction mixtures

A simple apparatus is described which permits the continuous monitoring of ¹⁴CO₂ production from ten separate reaction mixtures simultaneously. The device is relatively simple and inexpensive to construct, makes use of small disposable incubation vials, and allows complete trapping of all ¹⁴CO₂ evolved in scintillation vials, where it can be easily counted. The use of this apparatus to determine the rates of metabolism by glomeruli of ¹⁴C-labeled substrates to ¹⁴CO₂ is described.     

Solid state fermentation: acid protease production in controlled CO2 and O2 environments

The effect of the partial pressure of O(2) and CO(2) on the acid protease production in solid state fermentation by Aspergillus niger on wheat bran was studied. A fermentation system was used, which allowed on-line reactor measurements and continuous data acquisition of pH, temperature, gas flow, pressure drop and CO(2) production. Six paired combinations of CO(2) and O(2) concentrations were studied. The results showed a direct relationship between pressure drop, production of CO(2) and temperature increase. The pH evolution patterns were similar in all cases but different if the measurements were made on-line or on a liquid homogenate of the fermented substrate. Acid protease production was increased when the gas had 4% CO(2), (vol/vol), and it reached its highest level, a 43% increase over air, with a mixture of 4% CO(2) and 21% O(2). The protease production was strongly related to the mold metabolic activity as represented by the total CO(2) evolved.     

Environmental Regulation of CO2 Exchange Pattern in Facultative CAM Plants

Three facultative CAM plants, Sedum spectabile, S. aizoon and Mesembryanthemum cordifolium, could take up CO2 throughout the night and daytime, and no phase Ill was observed during cloudy weather. The CO2 exchange patterns during cloudy day differed obviously from that during sunny day. But in the obligate CAM plants, Kalanchoe daigremontiana, Orostachys fimbriatus and Bryophyllum pinnatum, there were phase Ⅲ during cloudy day. These results showed that the COs exchange patterns with uptake of CO2 throughout the night and daytime were universal in facultative CAM plants during cloudy day, but not in obligate CAM plants, of which the CO2 exchange patterns were very stable. In the three facultative CAM plants, the difference of exchange patterns between cloudy and sunny days depended mainly on temperature change. The effect of the temperature on CO2 exchange patterns was mediated by the decarboxylation rate. At high temperature, the decarboxylation rate could be enhanced. It was found that the accumulation of malic acid at night in the three obligate CAM plants was much more than that in the three facultative C AM plants. So during cloudy day, the decarboxytion rate in the three obligate CAM plants was also higher. This might be an important cause that obligate CAM plants need not to take up CO2 during the daytime.     

Canopy CO2 concentrations and Crassulacean acid metabolism in Hoya carnosa in a subtropical rain forest in Taiwan: consideration of CO2 availability and the evolution of CAM in epiphytes

The potential importance of CO₂ derived from host tree respiration at night as a substrate for night time CO₂ uptake during CAM was investigated in the subtropical and tropical epiphytic vine Hoya carnosa in a subtropical rainforest in north-eastern Taiwan. Individuals were examined within the canopies of host trees in open, exposed situations, as well as in dense forests. Although night time CO₂ concentrations were higher near the epiphytic vines at night, relative to those measured during the day, presumably the result of CO₂ added to the canopy air by the host tree, no evidence for substantial use of this CO₂ was found. In particular, stable carbon isotope ratios of H. carnosa were not substantially lower than those of many other CAM plants, as would be expected if host-respired CO₂ were an important source of CO₂ for these CAM epiphytes. Furthermore, laboratory measurements of diel CO₂ exchange revealed a substantial contribution of daytime CO₂ uptake in these vines, which should also result in lower carbon isotope values than those characteristic of a CAM plant lacking daytime CO₂ uptake. Overall, we found that host-respired CO₂ does not contribute substantially to the carbon budget of this epiphytic CAM plant. This finding does not support the hypothesis that CAM may have evolved in tropical epiphytes in response to diel changes in the CO₂ concentrations within the host tree canopy.     

In vivo demonstration of acid phosphatase activity in the rhizosphere of soil-grown plants

For in vivo demonstration of acid phosphatase activity in the rhizosphere of soil-grown plants filter papers were treated with a mixture of 1-naphthyl phosphate as substrate and the diazonium salt Fast Red TR as an indicator. After enzymatic hydrolysis, 1-naphthol forms a red complex with Fast Red TR. This method was applied to 8-day old maize plants and 3-year old Norway spruce plants growing in rhizoboxes in soil under non-sterile conditions. The treated filter paper is placed at the surface of roots and soil and acid phosphatase activity is visualized as a red-coloured root print on the filter paper. The method can be used as a qualitative analysis of acid phosphatase in the rhizosphere. It also allows a rough estimate of phosphatase activity in different root zones.     

Improved rates of CO2-fixation by intact chloroplasts isolated in media with KCl as the osmoticum

Intact chloroplasts were isolated from spinach leaves using media with either 330 mM sorbitol or 200 mM KCl as the osmoticum. Chloroplasts isolated in KCl exhibited higher rates of CO₂-dependent oxygen evolution in nine out of ten experiments, the average increase being 43%. Chloroplasts isolated in KCl routinely achieved rates of CO₂-dependent oxygen evolution of 200–300 mol·mg chlorophyll^-1·hour^-1 at 20°C. Intact chloroplasts were also isolated in media with 200 mM NaCl or choline chloride but the rates of CO₂ fixation were not superior to those isolated in sorbitol media. The K⁺ content of chloroplasts isolated in KCl media was higher than for chloroplasts isolated in sorbitol. It is suggested that the use of KCl as an osmoticum prevents the loss of chloroplast K⁺ which can occur during isolation in sorbitol media. Chloroplasts isolated in KCl lost, on average, 36% of the initial CO₂ fixation activity after storage for four hours on ice, compared to 24% loss of activity for chloroplasts isolated in sorbitol. This increased loss of activity was not observed if KCl was used in the grinding medium and sorbitol or glycinebetaine in the resuspension media. For measurement of the maximum photosynthetic capacity in vitro, the use of KCl in the grinding medium may be better than sorbitol.Abbreviations BSA bovine serum albumin - Chl chlorophyll - Pi inorganic orthophosphate - EDTA ethlenediamine tetraacetic acid     

Dependence of CO2 gas exchange and acid metabolism of the alpine CAM plant Sempervivum montanum on temperature and light

Summary The influence of light intensity and temperature on the diurnal course and magnitude of CO₂ gas exchange and on acid metabolism was studied in the laboratory with rooted rosettes of Sempervivum montanum collected at 2,200 m above sea level in the Central Alps. Under a temperature regime having a cool dark period and warm light period, S. montanum exhibited the time course of CO₂ gas exchange typical of a CAM plant; the response was very distinct even when the plants were well-watered. At day temperatures of less than 10° C and at night temperatures greater than 35° C, S. montanum behaved like a C₃ plant. Characteristic for S. montanum are a broad temperature optimum and a wide range of temperatures in which CO₂ uptake in light is possible (-2° to 45° C). Dark fixation of CO₂ is evident between-2° and 35° C, an apparent uptake of external CO₂, on the other hand, only as high as 20° C. Light saturation of CO₂ uptake is reached at 60–80 W m^-2 while the rate of deacidification is nearly maximal at 40 W m^-2. These results show that, due to their specific metabolism, CAM plants can be favored not only in xeric habitats, but also in heat stressed mountain habitats where the daily variation in temperature may be extreme.Dedicated with appreciation to Dr. K.F. Springer     

The effect of CO2 availability on the growth, iron oxidation and CO2-fixation rates of pure cultures of Leptospirillum ferriphilum and Acidithiobacillus ferrooxidans

Understanding how bioleaching systems respond to the availability of CO(2) is essential to developing operating conditions that select for optimum microbial performance. Therefore, the effect of inlet gas and associated dissolved CO(2) concentration on the growth, iron oxidation and CO(2) -fixation rates of pure cultures of Acidithiobacillus ferrooxidans and Leptospirillum ferriphilum was investigated in a batch stirred tank system. The minimum inlet CO(2) concentrations required to promote the growth of At. ferrooxidans and L. ferriphilum were 25 and 70 ppm, respectively, and corresponded to dissolved CO(2) concentrations of 0.71 and 1.57 μM (at 30°C and 37°C, respectively). An actively growing culture of L. ferriphilum was able to maintain growth at inlet CO(2) concentrations less than 30 ppm (0.31-0.45 μM in solution). The highest total new cell production and maximum specific growth rates from the stationary phase inocula were observed with CO(2) inlet concentrations less than that of air. In contrast, the amount of CO(2) fixed per new cell produced increased with increasing inlet CO(2) concentrations above 100 ppm. Where inlet gas CO(2) concentrations were increased above that of air the additional CO(2) was consumed by the organisms but did not lead to increased cell production or significantly increase performance in terms of iron oxidation. It is proposed that At. ferrooxidans has two CO(2) uptake mechanisms, a high affinity system operating at low available CO(2) concentrations, which is subject to substrate inhibition and a low affinity system operating at higher available CO(2) concentrations. L. ferriphilum has a single uptake system characterised by a moderate CO(2) affinity. At. ferrooxidans performed better than L. ferriphilum at lower CO(2) availabilities, and was less affected by CO(2) starvation. Finally, the results demonstrate the limitations of using CO(2) uptake or ferrous iron oxidation data as indirect measures of cell growth and performance across varying physiological conditions.