Effects of Glycollate on the Growth of a Planktonic Chlorella

The duration of the lag phase in cultures of a planktonic strainof Chlorella pyrenoidosa growing from small inocula under conditionsof light limitation was shortened by the addition of 1 mg 1^–1of glycollate to the medium. Of eight related compounds tested,none, with the possible exception of lactate, had a similareffect. Addition of 1 mg 1^–1 of glycollate also considerablyincreased the relative growth-rate of the alga at low (500 lux)but not at higher light intensities. This effect was not, however,specific to glycollate. Concentrations of glycollate (20 mg1^–1) higher than those normally produced in cultures byexcretion had an inhibitory effect on growth.     

The Growth of Chlorella pyrenoidosa on Glycollate

The utilization of glycollate as a substrate for photoheterotrophicgrowth by a strain of Chlorella pyrenoidosa has been demonstrated.Glycollate stimulated growth of this alga in basal medium overthe pH range 4.0 to 8.0 in the light, but did not support growthin the dark. Stimulation of growth in the light occurred overa wide range of glycollate concentrations and was optimal at30 mM. Enzyme and inhibitor experiments suggested that the synthesisof cell constituents during growth on glycollate is achievedby the same pathway which operates during the metabolism ofexogenous glycollate by autotrophically-grown cells. For algaeto metabolize and grow on exogenous glycollate the cells mustbe readily permeable to this compound. When the cells readilytake up exogenous glycollate, the level of activity of enzymesin the cell, in particular glycollate:DCPIP oxidoreductase,may regulate the over-all rate of glycollate metabolism.     

The Oxidation of 14C-labelled Glucose by Chlorella vulgaris: 1. The Time Course of 14CO2 Production

Glucose, either uniformly labelled with¹⁴C, or specificallylabelled in the I, 2, or 6 position, was added to C. vulgaris.Radio-active carbon dioxide was produced initially ten timesfaster from glucose-I-¹⁴C than from glucose-6-¹⁴C. This differencewas found with carbohydrate-starved cultures, exponentiallygrowing cultures, and cultures assimilating ammonia or nitraterapidly. A similar difference was also found with C. pyrenoidosaand Ankistrodesmus. 37 per cent. of the ¹⁴C added as glucose-¹-¹⁴Cto exponentially growing cells was recovered as carbon dioxidebut generally the recovery was less than this. Only 5 per cent.of ¹⁴C added as glucose-6-¹⁴C was recovered as carbon dioxide.The specific activity of the carbon dioxide produced was considerablylower than that of the carbon in the added glucose.     

Rates of Photosynthesis and Respiration in Relation to Stomatal Movements in Leaves Treated with {alpha}-Hydroxysulphonate and Glycollate

Leaves of tobacco and Xanthium pennsylvanicum were treated withsolutions of -hydroxysulphonate and glycollate in normal air.During the treatment with -hydroxysulphonate the intercellular-spacecarbon dioxide concentration of the leaves increased owing toa decrease in photosynthetic activity. It seems probable thatthese increases in internal carbon dioxide concentration werethe cause of stomatal closure, because when treated leaves wereflushed with carbon dioxide free air the stomata reopened. Experimentalresults also showed that the accumulation of glycollate, resultingfrom treatment with -hydroxysulphonate, could not be held responsiblefor stomatal closure because treatment with glycollate alonehad no influence on either the internal carbon dioxide concentrationor on the stomata of the leaves.     

Some Physiological Responses of Chlorella pyrenoidosa to 2,4-Dichlorophenoxyacetic Acid

An 18-h treatment of synchronously-grown Chlorella pyrenoidosawith 2,4-D did not significantly alter the size, dry weight,degree of synchrony, or pigment content of the cells, nor weredetectable quantities of ethylene produced. When Chlorella pyrenoidosawas treated with 5?10^–4 M 2,4-D, there was a statisticallysignificant stimulation of both net oxygen uptake and productionwhile 5?10 M 2,4-D inhibited both processes. When Chlorellapyrenoidosa was treated with 5?10^–4 M and 5?10^–3M 2,4-D, significantly greater amounts of glycollate were presentin the culture medium, even though an assay for glycollate dehydrogenaseshowed that the activity of this enzyme from 2,4-D-treated Chlorellapyrenoidosa was three times greater than in control cells. Looselybound 2,4-D was partitioned from a nonaqueously isolated chloroplastfraction, while other cell fractions failed to show detectablequantities of 2,4-D. It is postulated that in Chlorella pyrenoidosathe chloroplast is a target for 2,4-D action and that interferencein photorespiratory processes may underlie the observed responses.     

Overcoming Incompatibility in Brassica campestris L. by Carbon Dioxide, and Dark Fixation of the Gas by Self- and Cross-pollinated Pistils

Supplementing pollen suspension cultures with CO₂ (3–5per cent) caused a marked increase in germination and tube growthin vitro in Brassica campestris L. cv. toria. A weakening ofself-incompatibility by increased CO₂ levels from 3–5per cent was observed. The percentage of pollen tubes whichpenetrated the cuticle layer of stigmatic papilla cells in self-pollinatedpistils was high when CO₂ level was 5 per cent. Phosphoenolpyruvate (PEP) carboxylase activity was greater in the pollengerminated in 4 per cent CO₂ as compared to air (0.03 per cent).A possible role of CO₂ for self-recognition and control of pollentube growth is proposed, proposed. Brassica campestris L., carbon dioxide, self-incompatibility, phosphoenol pyruvate carboxylase     

The Photometabolism of Acetate by Chlorella pyrenoidosa

Chlorella pyrenoidosa can utilize sodium acetate as a carbonsource for growth in the light. Growth proceeds under aerobicconditions both in the presence and in the absence of carbondioxide, but under anaerobic conditions only in its presence.The assimilation of acetate does not result from oxidation tocarbon dioxide followed by photosynthetic fixation because theproducts of ¹⁴C-acetate assimilation are different from theproducts of ¹⁴CO₂ fixation in the presence of unlabelled acetate. In aerobic conditions 10-⁶ M DCMU induces a pattern of acetateassimilation in the light similar to that in the dark. Thus,in the presence of DCMU in the light, less acetate carbon isincorporated into cells, particularly into lipids, polysaccharide,and protein, and more is released as carbon dioxide than inits absence. The effect of 4 x 10^-3 M MFA on acetate assimilationin the presence of 10^-6 M DCMU is the same in light and dark.Acetate assimilation is unaffected by desaspidine and sodiumbisulphite. The mean generation time of C. pyrenoidosa growing on acetatein the light under aerobic conditions is 20 hours. When 10^-5M DCMU is added the mean generation time is 60 hours, the sameas that for Chlorella growing on acetate in the dark. The activityof the enzymes of the glyoxylate cycle, isocitrate lyase (E.C.4.1.3.1.)and malate synthetase (E.C.4.1.3.2.) is repressed in the light,but activity of both enzymes increases markedly when DCMU isadded.     

The Effects of pH and Carbon Dioxide on Ion Uptake by Excised Barley Roots

The effects of carbon dioxide concentrations up to 8 per centin air on uptake of potassium and chloride at two pH levels,nominally pH 6 and 8, werestudied. In all experiments, enhanced uptake of potassium occurred atthe higher pH level with carbon dioxidefree air, but chlorideuptake was generally unaffected. At nominal pH 6, 1 per cent carbon dioxide reduced and 6 percent increased potassium uptake. There was no effect on chlorideuptake except with 1 per cent carbon dioxide where a markedenhancement was recorded. At nominal pH 8, l and 2 per cent carbon dioxide increased potassiumuptake whereas 6 and 8 per cent were inhibitory. Chloride uptakewas favoured by 1 and 8 per cent concentrations of the gas.     

Extracellular Release of Organic Matter from Two Freshwater Diatoms

The ¹⁴C-labelled extracellular materials released from culturesof Synedra acus and Stephanodiscus hantzschii were separatedby paper chromatography. The major components were identifiedand their rates of release measured. Stephanodiscus in axenicculture released the same extracellular materials under optimumlight conditions as did the natural population from which thediatom was isolated. The major extracellular product of Stephanodiscuswas glycollate; polysaccharide and amino acids were also released.The major extracellular product of Synedra was polysaccharide;amino acids were also released, but no glycollate. Under lightintensities so high as to inhibit photosynthesis the arraysof extracellular substances were unchanged, but some rates ofrelease were reduced. The array of products and the rates ofrelease from Stephanodiscus in darkness are quite differentfrom extracellular release during photosynthesis. Glycollaterelease from Stephanodiscus is greatest when the cultures areyoung and growing rapidly, older cultures release less.     

The Carbohydrate Nutrition of Tomato Roots: V. The Promotion and Inhibition of Excised Root Growth by Various Sugars and Sugar Alcohols

Iron is only consistently present in an available form in White'sroot culture medium if the inorganic salts are autoclaved withthe sugar. The substitution of ferric ethylenediamine-tetra-acetatefor the inorganic ferric salt of White's medium ensures ironavailability when the carbon source of the medium is renderedsterile by ether treatment and subsequently added to the remainderof the constituents which have been sterilized by autoclaving. The biological activity of sugars, and particularly of dextroseand laevulose, is altered by autoclaving them in presence ofthe inorganic salt solution of White's medium. The only sugar which supports a considerable growth of excisedtomato roots is sucrose. The activity of this sugar is not affectedby alcohol-precipitation nor is it dependent upon the simultaneouspresence of traces of its constituent mono-saccharides. Dextrose or laevulose or a mixture of the two sugars supporta low but sustained level of excised-root growth. All othersugars and sugar alcohols tested were inactive as carbon sources. The addition of sucrose at low concentration (0–2 percent.) to a medium containing dextrose as the main carbon compounddoes not make possible a level of growth comparable with thatobtained with an adequate sucrose supply. It has not been possibleto enhance the growth-rate of excised roots supplied with dextroseby previous presentation of this sugar under conditions permittingactive growth. Using media containing 'etherized' sucrose anddextrose, no evidence was obtained of any competitive inhibitionof sucrose utilization by dextrose. Certain sugars when added to a medium, containing the optimumconcentration of sucrose₁, markedly inhibited excised root growth.Thus l-sorbose, l- and d-arabinose, and d-xylose caused notless than 80 per cent, inhibition at a concentration of 0-5per cent. d-mannose and d-galactose completely inhibited growthat o-1 per cent. The oligosaccharides, dextrose, laevulose,and the sugar alcohols tested had, by contrast, very low inhibitoryactivity.     

Uptake of glycollate by Skeletonema costatum (Grev.) Cleve in bacterized culture

Glycollic acid, supplied at a concentration of 1 mg l^?1, increased the relative growth rate of Skeletonema costatum (Grev.) Cleve growing in bacterized culture at limiting light intensities. There was little or no such effect at intensities approaching saturation. The presence in the medium of alumina, an adsorbent for glycollate, prolonged the lag phase, the cells remaining viable for up to 5 days. Uptake of glycollate was not appreciably affected by the bicarbonate concentration of the medium. After 3 h, 80–92% of the glycollate carbon assimilated was found in the alcohol and benzene insoluble fraction of the cells. This is in agreement with the supposition that glycollate carbon is as-similated directly by the diatom rather than after degradation by bacteria to carbon dioxide.     

The Effect of Hydroxymethanesulphonate on Photosynthesis in Chlorella pyrenoidosa

Photosynthesis under conditions known to favour glycollate excretionby algae did not result in glycollate excretion in a strainof Chlorella pyrenoidosa unless an inhibitor of glycollate oxidase,-hydroxypyridin-2yl-methane sulphonate (-HPMS), was present.This inhibitor increased the total amount of glycollate presentin the supernatant from the cells during photosynthetic carbondioxide fixation and gave accumulation of ¹⁴C in glycollateduring ¹⁴CO₂ fixation under conditions favouring glycollatesynthesis. At pH 8.3 -HPMS did not stimulate photosynthetic¹⁴CO₂ fixation in C. pyrenoidosa as occurs with some algae.Photoassimilation of acetate was inhibited by -HPMS, and thiswas shown to result from acetyl-CoA synthetase inhibition by-HPMS.     

The Role of Glycollic Acid in the Photoassimilation of Acetate by Chlorella pyrenoidosa

The kinetics of ³H-acetate assimilation by Chlorella pyrenoidosain the light were examined. The primary products of assimilationwere glycollate and succinate. After 10 sec glycollate contained45 per cent and succinate 25 per cent of the tritium incorporatedby the cells. The percentage of the total tritium in glycollateand succinate fell with time while that in citrate increased.Initially the specific activities (µc of ³H per µmoleof acid) of succinate and glycollate were greater than citrate.When ³H-¹⁴C-2-acetate was added to the cells, total dpm for³H and ¹⁴C in glycollate rapidly reached a steady state andgave a ³H/¹⁴C ratio of 10, compared with a ³H/¹⁴C ratio of 4in the acetate. This ³H/¹⁴C ratio in glycollate is found because³H is derived from ³H-¹⁴C-2-acetate and because the ¹⁴C is dilutedwith cold carbon from elsewhere. The addition of ¹⁴CO₂ at thesame time as ³H-¹⁴C acetate decreased the ³H/¹⁴C ratio in glycollatebut incorporation of ¹⁴C from ¹⁴CO₂ into glycollate was slowerthan incorporation from ¹⁴C-2-acetate. Although ¹⁴C from acetaterapidly appeared in glycollate, ¹⁴C-labelled glyoxylate wasnot detected. The ³H/¹⁴C ratio observed in glycollate rulesout formation of glycollate from acetate via glycoaldehyde.The available evidence did not support glycollate formationvia the Calvin cycle. ¹⁴C from ¹⁴C-Z-acetate appeared in glycollatebefore it did in phosphoglyceric acid. Total dpm for ³H, ¹⁴C,and ³H/^l4C ratio in Calvin cycle intermediates were not in equilibriumwith glycollic acid.     

Effects of Nitrogen Nutrition on Leaf Expansion and Photosynthesis of Trifolium subterraneum L. 1. Comparison between Different Levels of Nitrogen Supply

Growth analysis showed that reductions in the relative growth-rateof subterranean clover plants (cv. Mt. Barker), even those dueto moderate nitrogen deficiencies, were reflected in reductionsof the leaf-area ratio and particularly of the net assimilationrate. A decline in nitrogen supply in the culture solutions was foundto depress net rates of carbon dioxide uptake per unit leafarea and leaf expansion per plant to about the same extent,even at moderate levels of nitrogen stress. Four days aftertransfer of plants grown with adequate nitrogen to solutionswithout nitrogen, leaf area and net carbon dioxide uptake haddeclined to 84 per cent and 89 per cent of the values for thecontrol plants. After a further 4 days these values had decreasedto 71 per cent and 52 per cent respectively. When net carbon dioxide uptake was expressed per unit weightof chlorophyll, the effect of changes in nitrogen supply onnet photosynthesis largely disappeared, indicating a close relationshipwith the chlorophyll content of the leaves. However, anotherand perhaps more direct effect of nitrogen on photosynthesiswas suggested by the fact that, during the early stages of recoveryfrom a severe nitrogen stress, photosynthesis began to increasebefore the chlorophyll content of the leaves.     

The Effect of Artificial Wind on the Growth-rate of Plants in Water Culture: With Plate and three Figures in the Text

Young plants of Brassica napus (rape), Hordeum vulgare, andPisum sativum growing in water culture have been exposed tofour continuous wind speeds for from 4 to 5 days. The plantswere exposed to natural daylight, humidity, and temperaturein a wind tunnel in which only air movement was controlled.The wind speeds found among the plants in the four sectionsof this tunnel were approximately 0·3, 0·7, 1·7,and 4·0 m./sec. The results showed no significant changein relative growth-rate or net assinilation rate with wind speed. Previously published results, obtained with plants in soil orsand culture, have differed from those in the present experimentsin showing a fall in the amount of growth as wind speed increased.It is suggested that the reduction in growth found in theseprevious experiments was caused by partial drying out of theplants, whilst in the present experiments water-supply to theroots was abundant and this effect was considerably reduced.     

Growth of Sclerotiumrolfsiiat Different Concentration of Oxygen and Carbon Dioxide

Sclerotium rolfsii was grown in various atmospheres, the compositionsof which were controlled by the diffusion column technique.Growth-rate of mycelium was constant within a range of oxygenconcentrations from 3 to 21 per cent, but dirminished steadilyas carbon dioxide concentration increased from about 0.03 percent. The rate of sclerotial germination was reduced by oxygenconcentrations less than 6 per cent and by carbon dioxide concentrationsgreater than about 10 per cent. No sclerotia were formed ifthe concentration of oxygen fell below 15 per cent or if thatof carbon dioxide exceeded 4 per cent. The effects of inverserationof gases, from 0 per cent oxygen: 20 per cent carbon dioxideto 21 per cent oxygen: 0 per cent carbon dioxide, were in eachcase similar to the effect of the corresponding carbon dioxideconcentration when combined with 21 per cent oxygen.     

The Heterotrophic Nutrition of Chlorella vulgaris (Brannon No. 1 Strain): With two Figures In the Text

A range of sugars, sugar alcohols, sugar phosphates, organicacids, and monohydric alcohols have been tested as carbon sourcesfor growth and as respiratory substrates using Chlorella vulgaris,Brannon I, grown in darkness. Much higher rates of growth and respiration were obtained withd-glucose than with any other substance tested. Ethanol (at0·005 M.) sustained both growth and respiration at c.50 per cent, of the level with glucose (0·028 M. or higher).Evidence was obtained that the organism can become ‘adapted’to utilize d-galactose and sucrose as effective carbon sources.Sustained growth was not obtained with any of the other substancestested. The glucose monophosphates, methanol and certain organic acids(oxalacetate, -ketoglutarate, cis-aconitate, and pyruvate) clearlystimulated oxygen uptake but to a less extent than ethanol.The other substances tested were either inhibitory to respirationor inactive or of very low activity as substrates. The growth in darkness and in liquid culture of Chlorella whensupplied with d-glucose was insensitive to pH over the range4·5 to 7·0 and was markedly enhanced by a highlevel of aeration. Gains in cellular dry weight ranging from45 to 90 per cent, of the weight of d-glucose disappearing fromthe culture medium were recorded in growth experiments; measurementsof CO₂ evolution in the Warburg indicated retention of up totwo-thirds of the glucose-C in cell material.     

Glycollate Formation during the Photorespiration of Acetate by Chlorella

WhenChlorella pyrenoidosa photoassimilates ³H-¹⁴C-acetate theglycollic acid formed shows a high ³H/¹⁴C ratio, the only othercompounds showing similar ratios being glycerate and serine.The ³H/¹⁴C ratio of glycollate was unaffected by the TCA cycleinhibitors MFA, diethylmalonate and arsenite showing that ³Hin glycollate does not result from the oxidation of acetatevia the TCA cycle, the resulting NADP³H₂ or NAD³H₂ being usedfor the reduction of the glycollate precursor. Although DCMUdecreased the ³H/¹⁴C ratio, complete inhibition of glycollatelabelling was not observed with 10^–6 M DCMU, at whichconcentration complete inhibition of the Hill reaction is achieved.Although the ³H/¹⁴C ratio was unaltered, total dpm of both ¹⁴Cand ³H in glycollate were increased by INH. The ³H/¹⁴C ratiosof glycerate and serine were decreased by INH, as were the totaldpm of ³H and ¹⁴C incorporated into these compounds. Thus, INHinhibits the further metabolism of glycollate to glycerate andserine. The effect of INH on incorporation of ¹⁴C-I-acetateinto various cell fractions was investigated. The incorporationof ¹⁴C into polysaccharide and lipid was decreased, while theincorporation of ¹⁴C into the water-soluble fraction of cellsand therelease of ¹⁴CO₂ were little affected. Although glycollicacid was an early product of acetate photoassimilation in Chlorellapyrenoidosa, glycollate excretion does not take place undera wide range of environmental conditions shown to favour glycollateexcretion by other algae. However, small amounts of labelledglycollate were detected in the supernatant from the cells duringthe photoassimilation of ³H-¹⁴C-acetate, but this glycollatedid not show the high ³H/¹⁴C ratio of glycollate present withinthe cell. The failure of Chlorella pyrenoidosa to excrete appreciableamounts of glycollate when photoassimilating acetate or carbondioxide was considered to result from the presence of glycollateoxidase (EC 1.1.3.1 [EC] ) which allowed the further metabolism ofglycollate. Besides glycollate oxidase, glyoxylate reductasewas also demonstrated in Chlorella pyrenoidosa so that glycollatecould function in hydrogen transfer during the photoassimilationof acetate.     

Stimulation of Spore Discharge in Sordaria by Carbon Dioxide

The effect on the rate of spore discharge in Sordaria fimicolaof small concentrations of carbon dioxide has been studied.Substitution of an air-stream containing 0.2-2.5 per cent carbondioxide for air freed from this gas circulating over the peritheciaalways led to a marked increase in the rate of spore discharge.     

Studies of Glycollate Utilization and Some Associated Enzymes of C1 Metabolism in the Endosperm of Ricinus communis L.

Glycollate metabolism in 5-day-old endosperm tissues of Ricinuscommunis L. was examined by feeding micromolar quantities of[2-¹⁴C]glycollate to tissue slices. It was found that glycollatecarbon was rapidly incorporated into glyoxylate, glycine, serine,and carbon dioxide. Only small amounts of ¹⁴C were incorporatedinto the sugars. Changes in the distribution of ¹⁴C with timesuggested that glyoxylate was a primary product and that glycineand serine were secondary products of glycollate metabolism.The results of feeding experiments are interpreted as indicatingthat a glycollate pathway leading to sugar biosynthesis is ofminor importance compared to the rapid utilization of glycollatefor the biosynthesis of glycine and serine. Enzymes necessaryto catalyse the incorporation of glycollate into glycine andserine have been examined in castor-bean endosperm extracts.These included: glycollic acid oxidase, gloxylic acid reductase,glyoxylate transaminase, N¹⁰ formyltetrahydrofolate synthetase,N⁵,N¹⁰-methylenetetrahydrofolate dehydrogenase, and serine hydroxymethyltransferase.