Metabolism of Slices of the Tomato Stem

The rate of respiration of tomato stem slices varied considerably,the highest values (Qo₂, 2–3) being obtained for plantsin a good nutritional state, and the lowest (Qo₂, 1) in starvedplants. The respiratory quotient of 1.0 remained constant. Glucose fermentation was found to follow both glycolytic andalcohol fermentation pathways, the ratio of ethyl alcohol: lacticacid being 6.6:1. Fermentation seems to take place accordingto the Embden-Meyerhof scheme, as shown by the presence of someof these enzymes operative in this scheme and by inhibitionexperiments. In the presence of oxygen there was no formationof alcohol or lactic acid. Pyruvate added to tomato stem slices was metabolized by directoxidation to acetic acid and by dismutation to lactic and aceticacids and CO₂ The metabolism of acetic acid was demonstratedby its condensation with oxaloacetic acid to form citrate, thisbeing the second time that synthesis of citric acid by thismechanism has been found in plants. The presence of aconitase,of isocitric dehydrogenase, of succinic dehydrogenase, and ofmalic dehydrogenase, as well as the inhibition of respirationby malonic acid, favour the hypothesis that oxidation of carbohydratein tomato stem slices proceeds via the citric acid cycle. Thepossibility of an auxiliary route, the malic acid oxidationpathway, also was demonstrated. Tomato stem tissue anaerobicallysplit malic acid into glycolic acid. The further oxidation ofglycolic, glyoxylic, and formic acid was demonstrated. In experiments with C¹⁴-labelled acetate and butyrate a dilutionof the C¹⁴-labelled acids was found after incubation indicatingnew formation of these acids and of active participation offatty acid metabolism in the metabolic activities of the tissue. With the exception of alanine, added amino acids produced adefinite increase in O₂ uptake without extra formation of ammonia. Experimental demonstration of the possibility of electron transportfrom substrate to molecular oxygen in respiration via polyphenoloridase was provided by the attainment in a tomato tissue homogenateof a coupled oxidation-reduction between -ketoglutarate andcatechol with DPN and tyrosinase as the catalysts. The presenceof cytochrome oxidase was also demonstrated. Thus both systemspossibly may take part in the respiration of tomato stem slices.     

Organic Acid Metabolism of Sedum praealtum

1. The organic acids present are citric, isocitric, and l-malic,with a small residue of unidentified acids.
2. The diurnal variationin acidity is due chiefly to changes,in malic acid, with aparallel fluctuation shown by citric acid.Under these conditionsisocitric acid shows little change.
3. The importance of carbondioxide during acidification is confirmed,and it is shown thatat room temperatures or higher the CO₂produced in respirationis sufficient to produce maximum acidification.At lower temperaturesthe supply of CO₂ limits acid production.
4. In the absence ofoxygen no acidification occurs, but even smallquantities (approx.1 per cent.) are sufficient to cause someacid production.
5. Completebalance-sheets are presented for acids, carbohydrates,CO₂ andoxygen for leaves maintained in the dark at high andlow temperatures.As acids are produced there is a correspondingloss of carbohydrate(chiefly starch). A scheme of reactionsis suggested to explainthe experimental results.

     

Regulatory mechanisms of cellular respiration. III. Enzyme distribution in the cell. Its influence on the metabolism of pyruvic acid by bakers' yeast

The rate of the aerobic metabolism of pyruvic acid by bakers' yeast cells is determined mainly by the amount of undissociated acid present. As a consequence, the greatest rate of oxidation was observed at pH 2.8. Oxidation, at a slow rate, started at pH 1.08; at pH 9.4 there was no oxidation at all. The anaerobic metabolism, only a fraction of the aerobic, was observed only in acid solutions. There was none at pH values higher than 3. Pyruvic acid in the presence of oxygen was oxidized directly to acetic acid; in the absence of oxygen it was metabolized mainly by dismutation to lactic and acetic acids, and CO₂. Acetic acid formation was demonstrated on oxidation of pyruvic acid at pH 1.91, and on addition of fluoroacetic acid. Succinic acid formation was shown by addition of malonic acid. These metabolic pathways in a cell so rich in carboxylase may be explained by the arrangement of enzymes within the cell, so that carboxylase is at the center, while pyruvic acid oxidase is located at the periphery. Succinic and citric acids were oxidized only in acid solutions up to pH 4. Malic and α-ketoglutaric acids were not oxidized, undoubtedly because of lack of penetration.     

Roles of Organic Acids and Nitrate in the Long-Distance Transport of Cobalt in Xylem Saps of <Emphasis Type="Italic">Alyssum murale</Emphasis> and <Emphasis Type="Italic">Trifolium subterraneum</Emphasis>

Roles of organic acids and nitrate in the long-distance transport of cobalt (Co) in xylem saps of hyperaccumulator Alyssum murale and non-hyperaccumulator Trifolium subterraneum were studied under hydroponic conditions. Organic acids (oxalic, malic, malonic, citric, and fumaric) and nitrate in xylem sap samples were separated and determined simultaneously by reversed-phase high performance liquid chromatography after solid-phase extraction with nanosized hydroxyapatite. Results indicated that Co treatment significantly increased the concentrations of xylem oxalic and malic acids for the hyperaccumulator A. murale compared to the control but significantly decreased the concentrations of xylem nitrate and malonic acid; concentrations of citric acid in xylem sap samples did not show significant difference between the control and Co treatments. By analyzing the relationship between the concentrations of organic acids, nitrate, and concentrations of Co in xylem saps, it could be concluded that oxalic and malic acids in xylem saps seemed to participate in the long-distance Co translocation process, and citric acid did not relate to the xylem Co transport of A. murale and T. subterraneum. Our work might be very useful for understanding the mechanism of long-distance transport of heavy metals in hyperaccumulator.     

EFFECT OF MANGANESE IONS ON THE IAA-INDUCED ELONGATION OF AVENA COLEOPTILE SECTIONS INHIBITED BY SOME ORGANIC ACIDS

1.Organic acids, such as citric, -ketoglutaric, succinic, fumaricand L-malic acids, inhibit the IAA-induced growth of Avena coleoptilesections. But pyruvic acid has no effect on the growth. 2.High concentrations of MnCl₂ (for example 10^–3 m) alleviatethe inhibition due to L-malic, -ketoglutaric, succinic and fumaricacids, but not that due to D-malic, tartaric and malonic acids. 3.A mechanism of the alleviating effect of Mn⁺⁺ on the inhibitiondue to the organic acids is discussed with the reference tothe activating effect of Mn⁺⁺ on "malic" enzyme. ¹Contribution No. 6 from the Botanical Gardens. Faculty of Science,University of Tokyo, Koishikawa, Tokyo.     

Quinate:NAD oxidoreductase of germinating Phaseolus mungo seeds: Partial purification and some properties

Quinate:NAD oxidoreductase, which catalyzes the interconversionof quinic acid and 3-dehydroquinic acid, has been extractedfrom liquid N₂-frozen powders of 2-day-old etiolated seedlingsof Phaseolus mungo. The enzyme was partially purified by ammoniumsulfate fractionation and by DEAE-cellulose and gel filtrationcolumn chromatographies, and was separable from shikimate: NADPoxidoreductase and 3-dehydroquinate hydrolyase. The activityappeared to be maximal at pH 8.6–9.0. The apparent Kmvalues at pH 8.6 were 0.48 mM for quinic acid and 0.043 mM forNAD. The involvement of sulfhydryl group in the reaction wasdemonstrated by the potent inhibitory action of both heavy metalions and sulfhydryl inhibitors. The purified preparation ofthe enzyme was reasonably stable for storage in the presenceof dithiothreitol. The metal ions tested, except Hg²⁺ and Ag⁺,showed practically no inhibitory action on the enzyme activity.Aromatic amino acids and other aromatic and alicyclic compoundstested had little or no effect on the activity. ¹ Part 9 of "Alicyclic acid metabolism in plants". (Received January 20, 1977; )     

Effects of cadmium on the behaviour of citric acid in isolated tomato xylem cell walls

Effects of cadmium on the sorption of citric acid In isolatedxylem cell walls were Investigated. 2.5 nM to 9.5 mM [1.5–¹⁴]crticacid solutions were perfused through columns of xylem cell wallmaterial, isolated from tomato plants (Lycoperslcon esculentumMill, cv. Tiny Tim). The anion exchange potential of the column was estimated byamino acid analysis as approximately 46 meq dm whereas the apparentanion exchange capacity (AEC) was estimated as 1.65±0.1810^–4(citric acId units). This low AEC was attributed toa ‘zipper’ effect, a mutual screening of fixed R^–and A⁺ charges. Pre-loading with ¹¹⁵Cd²⁺ did not affect citric acid sorption,indicating the absence of Cd-effects on the availability offixed A⁺ charges, and the absence of the formation of effectiveR^–-Cd²⁺ and Donnan tree space (DFS) (Cd(cit)H₂]⁺ complexes. Simultaneous application of both citric acid and ¹¹⁵Cd²⁺,⁴⁵Ca²⁺or ²⁸Mg²⁺ resufted in increased sorption of citric acid, probablydue to capacity improvement rather than changes in valence-dependentanion sorption; this may be due to the presence of bulk (M(cit)H₂]⁺,held in the column as [M(cit)H₂]⁺ after protonation in the DFS.Sorption of citric acid was greatest in the presence of Ca²⁺which was discussed in the light of the differences betweenCa, Cd and Mg in their characteristics as co-ordinative M-complexes of citric acid. The overall results indicate the potentialimportance of the presence of metal ions for the xylem transportbehaviour of organic acids in plants. Key words: Cadmium, citric acid, ion exchange, ligand exchange, tomato, xylem cell walls     

Influence of Drought on Mitochondrial Activity, Photosynthesis, Nocturnal Acid Accumulation and Water Relations in the CAM Plants Prenia sladeniana(ME-type) and Crassula lycopodioides(PEPCK-type)

The activity of photosynthesis and mitochondrial respiration,nocturnal organic acid accumulation and water relations wereinvestigated in Prenia sladeniana L. Bol. [malic enzyme (ME)-type]andCrassula lycopodioides Lam. [phosphoenolpyruvate carboxykinase(PEPCK)-type] to compare the physiological responses to waterdeficit in crassulacean acid metabolism (CAM) plants differingin their decarboxylating enzyme systems. Withholding water inhibiteddaytime gas exchange within 2 d while night time CO₂gain andmalic acid accumulation remained relatively unchanged in bothspecies. In P. sladeniana, maximum photochemical efficiency(F_v/F_m) and photosynthetic electron transport declined to nearlythe same degree as CO₂supply was restricted during drought.Despite limited CO₂availability, photosynthetic activity waslargely unaffected in C. lycopodioides, as were mitochondrialproperties. There is no indication of a drought-induced increasein the capability to totally oxidize malate, yielding 4 CO₂, in either species. Nevertheless, the enhanced ratio of malateto glycine oxidation may have increased the in vivo capabilityfor malate oxidation in P. sladeniana. Although pressure potentialwas maintained throughout the experiment in both species, activeosmotic adaptation occurred only inP. sladeniana. The observeddecrease in photosynthetic and mitochondrial activity may haveresulted from the large increase in osmotic concentration inthis species. Copyright 2000 Annals of Botany Company Chlorophyll fluorescence analysis, Crassula lycopodioides Lam., crassulacean acid metabolism, citric acid, gas exchange, malic acid, mitochondria, photosynthetic electron transport, Prenia sladeniana L. Bol., water relations     

The Isolation of l-Quinic Acid from the Apple Fruit

1. A method is described by means of which several grammes ofl-quinicacid were isolated from young fruits of the WorcesterPearmainapple stored for a number of days in nitrogen.
2. Theessential stages in the method are the removal of colouringmatter from extracts of the frozen ground pulp tissue of thefruit with charcoal, the removal of amino acids by absorptionon cation exchange resin, and the separation of the organicacid from sugars by adsorption of the former on anion exchangeresin. Finally, the quinic acid was separated from malic acidby fractional displacement from the anion exchange resin.
3. Thecharacterization of the isolated quinic acid, and of malicacidalso isolated from the fruits, is described.
4. Citric acid isshown to be formed by oxidation of quinic acidwith hot hydrogenperoxide.
5. The possible function of quinic acid in the applefruit andin plants generally is discussed.

     

Role of extracellular peroxidase in the superoxide production by wheat root cells

Summary Extracellular peroxidase has been shown to contribute to superoxide production in wounded wheat (Triticum aestivum L. cv. Ljuba) root cells. The superoxide-synthesizing system of root cells was considerably inhibited by KCN and NaN₃ and activated by MnCl₂ and H₂O₂. Treatment of roots with salicylic acid and a range of di- and tri-carbonic acids (malic, citric, malonic, fumaric, and succinic acids) stimulated superoxide production in both root cells and extracellular solution. The H₂O₂-stimulated superoxide production in the extracellular solution was much higher when roots were preincubated with salicylic or succinic acid. Exogenous acids enhanced peroxidase activity in the extracellular solution. Pretreatment of root cells with the detergents trypsin and sodium dodecyl sulfate had similar effects on the peroxidase activity. Significant inhibition of both superoxide production and peroxidase activity by diphenylene iodonium suggests that the specificity of the latter as an inhibitor of NADPH oxidase is doubtful. Results obtained indicate that extracellular peroxidase is involved in the superoxide production in wheat root cells. The mobile form of peroxidase can be readily secreted to the apoplastic solution and serve as an emergency enzyme involved in plant wound response.Abbreviations DPI diphenylene iodonium - ECS extracellular solution - ROS reactive oxygen species - SA salicylic acid     

Alicyclic acid metabolism in plants V. Biosynthesis of alicyclic acids from 14C-labeled glucose and erythrose in some plant tissues

Etiolated seedlings of Phaseolus mungo were fed with ¹⁴C-glucoseand the incorporation of ¹⁴C into shikimic and quinic acidswas determined. The incorporation of ¹⁴C into shikimic acidwas enhanced when non-labeled shikimic, quinic or 5-dehydroquinicacid was not significantly affected by these alicyclic acids.To examine whether the difference in biosynthetic patterns betweenshikimic and quinic acids is common in higher plants, flowersand leaves of several plants were fed with ¹⁴C-glucose or ¹⁴C-erythroseand the effciencies of these labeled sugars as precursors ofshikimic and quinic acids were compared. In seven of eight plantsamples, erythrose was superior to glucose as the precursorof shikimic acid, while there was no great difference in theefficiency of either sugar as the precursor of quinic acid.The possibility that the biosynthetic mechanism for quinic aciddiffers from that for shikimic acid is discussed. (Received September 12, 1973; )     

Diurnal Rhythm and Characteristics of Photosynthesis and Respiration in the Leaf and Root of a Phalaenopsis Plant

A hybrid Phalaenopsis plant was grown hydroponically on a nutrientsolution of Hyponex in the greenhouse. Typical daily patternsof CO₂ fixation, production of malic acid and citric acid andpH of the leaf and root were determined. The rate of primaryfixation of CO₂ in the leaf increased markedly in the evening,remained high until well into the afternoon, then decreasedsharply. The pattern of production of organic acids resultedfrom the fact that the rate of uptake of CO₂ gas was highestat night and fell during the day. A high correlation betweenpH value and level of malic acid in the leaves was observed,and an exponential relationship appeared to exist between theseparameters. These rhythmic sequences were not observed in theroot. The results suggest that metabolism in the leaf had characteristicsof CAM (Crassulacean Acid Metabolism), while the root did not. (Received May 6, 1988; Accepted October 25, 1988)     

紫色小白菜有机酸的提取优化及UPLC定量分析

为建立一种紫色小白菜中精准的有机酸提取及定性定量分析方法。本研究在单因素试验的基础上,采用Box-Behnken的中心组合试验设计原理,设计响应面试验优化紫色小白菜有机酸的提取工艺。结果表明,紫色小白菜有机酸提取的最优工艺参数为:乙醇浓度73%,料液比1∶21,超声时间11 min,超声温度70℃。通过超高效液相色谱法(ultra performance liquid chromatography,UPLC)对紫色小白菜叶片和叶柄部位有机酸进行定性定量分析,检测出苹果酸、柠檬酸、丙二酸、琥珀酸和酒石酸,其中苹果酸含量最高,分别为15.968、5.019 mg/g;其次为柠檬酸,分别为9.293、1.385 mg/g。定性定量结果表明,紫色小白菜叶片及叶柄部位含有丰富有机酸类物质,叶片部位苹果酸、柠檬酸、丙二酸、酒石酸含量均高于叶柄部位,而琥珀酸含量较低。     

Mapping QTLs controlling fruit quality in peach (Prunus persica (L.) Batsch)

 The organoleptic quality of fleshy fruits is in a large part defined by their composition of soluble sugars and organic acids. An F₂ population issuing from a cross between two peach varieties, ‘Ferjalou Jalousia’, a non-acid peach, and ‘Fantasia’, an acid nectarine, was analysed over 2 successive years for agronomic characters and for molecular-marker (isoenzymes, RFLPs, RAPDs, IMAs and AFLPs) segregations. Blooming and maturity dates, as well as productivity, were noted for each tree. Four fruits per tree were analysed at maturity for fresh weight, colour, pH, titratable acidity, soluble-solids content (SSC), acid (malic, citric and quinic acids) and sugar (sucrose, glucose, fructose, sorbitol) contents. QTLs were detected for all fruit components analysed, except for fruit colour. The QTLs for nearly all components were present on two linkage groups. For productivity, fresh weight, pH, quinic acid, sucrose and sorbitol content, all the detected QTLs displayed the same effect as the parental phenotypes. By contrast, for maturity date, titratable acidity, malic and citric acids and fructose, some QTLs displayed the same effect as the parental phenotypes while others displayed the opposite effect. The fraction of the total variation in each trait throughout the population explained by the QTLs was very high and reached more than 90% for some characters. For most of the characters analysed, epistasis was observed between QTLs. Received: 10 October 1997 / Accepted: 18 August 1998     

The Regulation of Citric Acid Accumulation and Carbon Recycling During CAM in Ananas comosus

The carbon balance of shade-grown Ananas comosus was investigatedwith regard to nitrogen supply and responses to high PAR. Netdark CO₂ uptake was reduced from 61.2 to 38.5 mmol CO₂ m^–2in N limited (–N) plants grown under low PAR (60 µmolm^–2 s^–1) and apparent photon yield declined from0.066 to 0.034 (mol 0².mol^–1 photon), although photosyntheticcapacities (measured under 5% CO₂) were similar. Following transferfor 7 d to high PAR (600. µmol m^–2 s^–1), netCO² uptake at night increased by 14% in +N plants, and daytimephotosynthetic capacity was higher, with a maximum value of7.8 µmol m^–2 s^–1. The magnitude of dark CO₂ fixation during CAM was measured asdawn—dusk variations in leaf-sap titratable acidity (H+)and as the proportion of malic and citric acids. The contributionfrom re-fixation of respiratory CO₂ recycling (measured as thedifference between net CO₂ uptake and malic acid accumulation)varied with growth conditions, although it was generally lower(30%) than reported for other bromeliads. Assuming a stoichiometryof 2H+: malate and 3H+: citrate, there was a good agreementbetween titratable protons and enzymatically determined organicacids. The accumulation of citric acid was related to nitrogensupply and PAR regime, increasing from 7.0 mol m–3 (+Nplants) to 18 mol m^–3 (–N plants) when plants weretransferred to high PAR; malate: citrate ratios decreased from13.1 to 2.5 under these conditions. Under the low PAR regime, leaf-sap osmotic pressure increasedat night in proportion to malic acid accumulation. However,following the transfer to high PAR for 7 d, there was a muchgreater depletion of soluble sugars at night which correspondedto a decrease in leaf-sap osmotic pressure. Although a rolefor citric acid in CAM has not been properly defined, it appearsthat the accepted stoichiometry for CAM in terms of gas exchange,titratable acidity, malic acid and osmotic pressure may nothold for plants which accumulate citric acid. Key words: Ananas comosus, CAM, citric acid accumulation, carbon recycling     

Effect of acetaldehyde on fatty acid oxidation and ketogenesis by hepatic mitochondria.

Acetaldehyde inhibited the oxidation of fatty acids by rat liver mitochondria as assayed by oxygen consumption and CO₂ production. ADP-stimulated oxygen uptake was more sensitive to inhibition by acetaldehyde than was uncoupler-stimulated oxygen uptake, suggesting an effect of acetaldehyde on the electron transport-phosphorylation system. This conclusion is supported by the decrease in the respiratory control ratio, associated with fatty acid oxidation. Acetaldehyde depressed ketone body production as well as the content of acetyl CoA during palmitoyl-1-carnitine oxidation. Acetaldehyde was considerably more inhibitory toward fatty acid oxidation than was acetate. Therefore, the inhibition by acetaldehyde is not mediated by acetate, the direct product of acetaldehyde oxidation by the mitochondria. Oxygen uptake was depressed by acetaldehyde to a slightly, but consistently, greater extent in the absence of fluorocitrate, than in its presence. This suggests inhibition of oxygen consumption from β-oxidation to acetyl CoA and that which arises from citric acid cycle activity. The inhibition of fatty acid oxidation is not due to any effect on the activation or translocation of fatty acids into the mitochondria.The depression of the end products of fatty acid oxidation (CO₂, ketones, acetyl CoA) as well as the greater sensitivity of palmitate oxidation compared to acetate oxidation, suggests inhibition by acetaldehyde of β-oxidation, citric acid cycle activity, and the respiratory-phosphorylation chain. Neither the activities of palmitoyl CoA synthetase nor carnitine palmitoyltransferase appear to be rate limiting for fatty acid oxidation.     

Changes in bulk protein of tobacco leaves on aerobic autolysis: Hydrogenation studies and identification of bound quinic acid

During aerobic autolysis and in commercial curing, the bulk proteins of tobacco leaves become coupled with quinic acid, presumably in consequence of coupling of chlorogenic acid congeners with lysine ε-NH₂ groups. Quinic acid derivatives, prepared from acid hydrolysates of such altered proteins, were identified by GC-MS. Such proteins were also hydrogenated over Rh/Al₂O₃ with a view to stabilizing the hypothetical linkages. Difficulties in removing contaminant Al had to be overcome. Evidence was then obtained (by GLC of derivatives) for several components, in acid hydrolysates of hydrogenated altered proteins, which were neither normal hydrogenation products of the common amino acids nor derivatives of quinic acid. Details of the chromatograms and mass spectra of quinic acid derivatives are provided in a supplementary publications.     

Quinate:NAP(P)+-oxidoreductase from Larix sibirica: purification,characterization and function

Quinate:NAP(P)⁺-oxidoreductase (QORase, EC 1.1.1.24), which catalyzes the interconversion of quinic and 3-dehydroquinic acids, was purified from the needles and developing xylem cells of Larix sibirica. The enzymes from these two tissues were partially characterized and compared. QORase from needles had optimum pH at 9.0 and apparent K_m values of 1.84 mM for quinic acid and 0.19 mM for NADP⁺. The enzyme was activated by phosphoenolpyruvate. Gallic and protocatechuic acids were formed in a reaction mixture of purified enzyme from needles as final products of quinic acid transformation. QORase from developing xylem cells showed pH optimum at 10.0 and had apparent Km values of 0.70 mM for quinic acid and 0.05 mM for NADP⁺. The enzyme was not affected by PEP. The divalent cations Co²⁺ and Mn²⁺ at least doubled activity of QORase from both sources but Mg²⁺ affected the enzyme from needles only. The spatial organization and regulation of quinic acid metabolism in the autotrophic and heterotrophic cells of conifers and the role of QORase in this process are discussed.     

The enzymic conversion of hydroxycinnamic acids to p-coumarylquinic and chlorogenic acids in tomato fruits

Two enzymes thought to be involved in the biosynthesis of chlorogenic acid have been separated and purified by ion exchange chromatography and their properties studied. These two enzymes, p-coumarate CoA ligase and hydroxycinnamyl CoA: quinate hydroxycinnamyl transferase, acting together catalyse the conversion of p-coumaric acid to 5′-p-coumarylquinic acid and of caffeic acid to chlorogenic acid. The ligase has a higher affinity for p-coumaric than for caffeic acid and will in addition activate a number of other cinnamic acids such as ferulic, isoferulic and m-coumaric acids but not cinnamic acid. The transferase shows higher activity and affinity with p-coumaryl CoA than caffeyl CoA. It also acts with ferulyl CoA but only very slowly. The enzyme shows high specificity for quinic acid; shikimic acid is esterified at only 2% of the rate with quinic acid and glucose is not a substrate. The transferase activity is reversible and both chlorogenic acid and 5′-p-coumarylquinic acids are cleaved in the presence of CoA to form quinic acid and the corresponding hydroxycinnamyl CoA thioester.     

The Effect of Temperature and Light on Gas Exchange and Acid Accumulation in the C3-CAM Plant Clusia minor L

Gas exchange and organic acid accumulation of the C₃-CAM intermediateClusia minor L. were investigated in response to various day/nighttemperatures and two light regimes (low and high PAR). For bothlight levels equal day/night temperatures between 20°C and30°C caused a typical C₃ gas exchange pattern with all CO₂uptake occurring during daylight hours. A day/ night temperatureof 15°C caused a negative CO₂ balance over a 24 h periodfor low-PAR-grown plants while high-PAR-grown plants showeda CAM gas exchange pattern with most CO₂ uptake taking placeduring the dark period. However, there was always a considerablenight-time accumulation of malic acid which increased when thenight-time temperature was lowered and had its maximum (54 mmolm^–2) at day/night temperature of 30/15°C. A significantamount of malic acid accumulation (23 mmol m^–2) in low-PAR-grownplants was observed only at 30/15°C. Recycling of respiratoryCO₂ in terms of malic acid accumulation reached between 2·0and 21·5 mmol m_–2 for high-PAR-grown plants whilethere was no significant recycling for low-PAR-grown plants.Both low and high-PAR-grown plants showed considerable night-timeaccumulation of citric acid. Indeed under several temperatureregimes low-PAR-grown plants showed day/night changes in citricacid levels whereas malic acid levels remained approximatelyconstant or slightly decreased. It is hypothesized that lowand high-PAR-grown plants have different requirements for citrate.In high-PAR-grown plants, the breakdown of citrate preventsphotoinhibition by increasing internal CO₂ levels, whereas inlow-PAR-grown plants the night-time accumulation of citric acidmay function as an energy and carbon saving mechanism. Key words: C. minor, C₃, CAM, citric acid, light intensity