The effect of detrital addition on the development of nanoflagellates and bacteria in Lake Kinneret

The effect of adding dissolved substrates derived from algalcells on the patterns of nutrient cycling and growth of bacteria,heterotrophic nanoflagellates (HNAN) and photoautotrophs wasdetermined in samples of near-surface waters from Lake Kinneret.Supplementation of substrates always resulted in an increasedpeak of HNAN numbers and had little effect on bacterial numbers.HNAN-mediated nutrient remineralization of nitrogen and phosphoruswas also stimulated. In light-incubated samples the remineralizednutrients were taken up by photoautotrophic cells. Maximum growthrates observed for HNAN ranged from 0.03 to 0.11 h^–1,clearance rates for bacteria 1.1–7.3 nl HNAN^–1 h^–1and remineralization rates 6.4–8.4 µg N mg dry wt^–1h^–1 and 0.37–0.99 µg P mg dry wt^–1 h^–1.     

Relationship between biomass and enzymatic activity of a bloom-forming dinoflagellate (Dinophyceae) in southern Chile (41{degrees} S): a field approach

In a coastal area of southern Chile (41° S), the major ammoniumassimilating enzyme glutamine synthetase (GS) was detected ina green dinoflagellate bloom during April 2003. High chlorophylla concentrations (1000 µg L^–1) attributable to Gymnodiniumcf. chlorophorum in surface waters were associated with highand very low nitrate reductase activities. Coincident with thebloom, dissolved inorganic nitrogen concentrations were nearthe detection limit (NO₃^– + NH₄⁺ <0.5 µM). SinceGS correlates with the use of ammonium as an external nitrogensource, we suggest that GS activity seems to be a good indicatorof ammonium utilization in a period dominated by a single dinoflagellatespecies.     

Accurate and Simple Measurement of Phytic Acid Contents in Cereal Grains

We developed an accurate and simple method for measuring thephytic acid contents in cereal grains on the basis of closeexamination of various factors affecting the accuracy and reproducibilityof the measurement. Our conclusions were: (1) As extracting medium for phytic acid,HCl was better than trichloroacetic acid or H₂SO₄. (2) The suitablepH range for extracting phytic acid was 0.3 to 1.0. (3) Completeformation of Fe-phytate required four to nine times as muchFe^3$ as phytic acid. (4) No definite effect of Na₂SO₄ on therecovery of phytic acid was observed, though it increased theextracting efficiency of trichloroacetic acid. (5) The determinationlimit of phytic acid by the iron precipitation method was 3.3µmol per 20 ml. Our new method gave reliable results, and a linear relationshipof y=0.3163x$0.0597 was obtained up to 0.025 µmol µ^–1between the concentration of phytic acid and the zone lengthin isotachophoresis. The phytic acid contents of two kinds ofrice, two kinds of rye, and one kind of wheat and of barleywere determined with this method. (Received March 19, 1985; Accepted August 2, 1985)     

Fatty acid composition in photosynthetic products of natural phytoplankton population in Lake Biwa, Japan

Fatty acid composition of phytoplankton photosynthetic productswas determined by a ¹³C tracer and gas chromatography-mass spectrometry(¹³C-GC-MS) method from August 1985 to June 1986 in Lake Biwa,Japan. The total fatty acid production rate varied from 2.8to 10.9 µg C l^–1 day^–1 at the water surfaceand accounted for 9.1–30% of photosynthetic productionof particulate organic carbon. A high contribution of fattyacid to the particulate organic carbon production rate was noticedduring winter time, and an increase in the fatty acid contributionresulted in an increase in the C/N value in the photosyntheticproducts. The fatty acid composition varied throughout the year,mainly depending on the seasonal change in the dominant phytoplanktonspecies. The contribution of polyunsaturated fatty acids tototal fatty acids was low during the summer period, probablydue to nitrogen limitation of phytoplankton growth.     

Ion Transport and the Effects of Acetic Acid in White Clover: II. POTASSIUM ABSORPTION

Dunlop, J., Knighton, M. V. and White, D. W. R. 1988. Ion transportand the effects of acetic acid in white clover. II. Potassiumabsorption.—J. exp. Bot. 39: 89–96. Acetic acid stimulated K⁺ influx into cells of white cloverin suspension culture and net K⁺ influx by roots of intact whiteclover plants. At concentrations of acetic acid up to 2·0mol m^–3 the stimulation continued unabated for at least2 h. However, at higher concentrations the rate of absorptiondeclined to near zero values after 2 h. When acetic acid wasremoved from the solution there was a net efflux of K⁺. Thestimulation was pH dependent with a maximum at pH 5·0.There was maximum stimulation at an acetic acid concentrationof 2·0 mol m^–3 Net H⁺ efflux was reduced by 1·0 mol m^–3 aceticacid. When Rb⁺ uptake and H⁺ efflux were measured at a rangeof RbCl concentrations Rb⁺ uptake increased with concentrationwhereas H⁺ efflux was maximum in the absence of Rb⁺ (and K⁺)and decreased as RbCl concentration was increased Acetic acid caused a hyperpolarization of the membrane electricalpotential difference (E) of about 25 mV. In 1·0 mol m^–3acetic acid the hyperpolarization persisted for at least 2 hwhereas at 10 mol m^–3 d E subsequently depolarized tovalues around –80 mV. With a slight lag, the time courseof the stimulation of the rate of K⁺ absorption followed thepolarization and depolarization of E. These results imply thatthe linkage between K⁺ and H⁺ movements is probably throughE. Key words: Proton efflux, membrane electrical potential difference, Trifolium repens     

Analysis of abscission responses to photodecomposition products of 1-naphthaleneacetic acid

This paper presents an analysis of abscission reponses of cottonexplants to (a) 1-naphthaneneacetic acid; (b) photodecompositionproducts of 1-naphthaleneacetic acid: 1-methylnaphthalene, 1-naphthaldehyde,1-naphthoic acid, naphthalene, and phthalic acid; and (c) arelated compound: naphthaleneacetyl aspartate. Abscission wasaccelerated by low amounts and retarded by high amounts of 1-naphthaleneaceticacid and 1-naphthoic acid. No significant effect on abscissionwas observed from 1- methylnaphthalene, 1-naphthaldehyde, orphthalic acid applied in amounts from 10^–8 to 10.0 µgper petiole; or with naphthalene from 10^–3 to 10.0µgper petiole. Naphthaleneacetyl aspartate had no effect at 5?10⁴to 5?10^–3 µg per petiole, but completely inhibitedabscission at 5 ? 10^–1 and 5.0 µg per petiole. Thedata are analyzed on part by a previously described mechanicalmethod for the determination of abscission indexes, and in partby a new method described herein, using a digital computer forthe analysis of the abscission time-course data. The resultshave significance to the understanding of the variability encounteredin fruit thinning by 1-naphthaleneacetic acid and related substances,and are discussed in relation to the known intermediate effectsof 1-naphthaleneacetic acid in fruit thinning. ¹Present address: Department of Biology, Univesity of California,Riverside, California 92502, U.S.A. (Received August 26, 1972; )     

Proton Fluxes and the Activity of a Stelar Proton Pump in Onion Roots

The xylem vessels of excised adventitious roots of onion, Alliumcepa, were perfused with unbuffered nutrient solution adjustedinitially to either pH 9·3 or 3·9; the pH of thesolution after passage through the xylem, at rates not lessthan 2 xylem volume changes min^–1, was close to pH 6·5in both instances. The flux of H⁺ across the xylem/symplastboundary into mildly alkaline, phosphate-buffered solutionsperfusing the vessels could be increased greatly with increasingbuffer strength, up to a maximum value between 0·5–1·0pmol H⁺ mm^–2 s^–1. The apparent neutralization ofacidic malic acid buffers had a slightly lower maximum capacity,equivalent to –0·3 to –0·5 pmol H⁺mm^–2 s^–1. The addition of 5·0 pmol m^–3fusicoccin (FC) to the xylem perfusion solution stimulated theentry of H⁺ into the xylem; in unbuffered perfusion solutionsthe pH fell to pH 3·6 after a lag of 25–35 min.FC additions to phosphate-buffered solutions also stimulatedthe H⁺ flux to an extent similar to that in unbuffered solution,viz. 0·2–0·4 pmol mm^–2 s^–1. The release of K⁺ (³⁶Rb-labelled) into xylem sap transientlyincreased as the [K⁺] in weakly buffered perfusion solutionswas raised stepwise; a very marked increase being seen whenthe concentration was raised to 100 mol m^–3 from 40 molm^–3. The addition of 5·0 mmol m^–3 FC to theperfusing solution containing 100 mol m^–3 K⁺ rapidly decreasedthe K⁺ flux to the xylem as the H⁺ flux increased. Fusicoccinalso inhibited the flux of K⁺ into unbuffered perfusion solutionsbut the effect appeared reversible. Addition of 10 mmol m^–3abscisic acid (ABA) to the perfusion solution quickly producedtransient increases in both K⁺ and H⁺ fluxes into the xylem.In this and other experiments using weakly phosphate-bufferedperfusing solutions, H⁺ fluxes were comparable in size to thoseof K⁺ The results are consistent with the idea that the stele of onionroots contains a proton trarislocating ATPase whose activityresponds to the pH of the xylem sap. It is evident that theactivity of the proton secreting and proton neutralizing mechanismsin the xylem parenchyma control the movement of other ions acrossthe xylem/symplast boundary. Key words: Xylem perfusion, fusicoccin, abscisic acid, pH gradient     

Callus, Organogenesis and Plantlet Formation in Tissue Cultures of Clitoria ternatea

Decoated seeds of Clitoria ternatea L. germinated on Murashigeand Skoog (Physiologia Plantarum 1962, 15, 473–97) basalmedium (BM) and differentiated callus and bipolar embryoids(two-step method) in low frequency. Calluses developed on lateralroots [BM+KN(0.1 mg 1^–1)], on roots and hypocotyls [BM+KN(0.5mg 1^–1)], and on roots [BM+KN+IAA (0.5 mg 1^–1 ofeach)]. On basal medium with KN (0.5 mg 1^–1) and withKN+IAA (0.5 mg1^–1 of each), multiple shoot buds and embryoids(one-step method) were differentiated directly on split hypocotylsand roots. In the former, shoot buds developed even on unsplithypocotyls. Rhizogenesis on isolated shoot buds occurred efficientlyin BM+indole butyric acid (IBA 0.1 mg 1^–1) and BM+IAA(0.1 mg 1^–1 and 0.5 mg 1^–1). Profuse direct embryoidsand shoot buds developing on root systems are interesting morphogeneticphenomena rarely reported. Clitoria ternatea L., callus, embryoids, multiple shoot buds, regeneration     

Assemblage of Ciliated Protozoan Community in a Polluted and Non-polluted Environment in a Tropical Lake of Central Himalaya: Lake Naini Tal, India

In order to obtain information on the assemblage of Protozoain the changing environment in Lake Naini Tal, this study wascarried out for a period of one year from November 1995 to October1996. Samplings were done from the mud–water interfaceat two stations which differed considerably in their magnitudeof pollution. Station I was moderately polluted while StationII was highly polluted. Some physico-chemical parameters ofwater such as temperature, dissolved oxygen, free CO₂, pH, nitrate-nitrogen,phosphate-phosphorus and BOD were also measured. A total of23 ciliates were found at the mud–water interface of thetwo stations during the sampling period. The annual mean ofspecies richness at Station I was significantly greater (19species) than that of Station II (13 species). The species compositionwas also different at the two stations. In general, StationI supported larger ciliated species (1249 x 10³ µm³ taxon^–1)than Station II (348 x 10³ µm³ taxon^–1). Among differentfeeding groups of ciliates, the groups ‘Algivore-Bacterivore’and ‘Bacterivore’ were about twice as common atStation II (116 x 10³ cells^–1) than at Station I (55 x10³ cells^–1). The annual average ciliate community abundancewas more diverse at Station I than Station II. The annual averagebiomass in terms of carbon content for both stations was almostthe same (6.0 mg Cl^–1 for Station I and 6.1 mg Cl^–1Station II). However, different species were responsible forthe contribution to the biomass at the two stations. The valuesfor Shannon-Weiner's diversity indices at Staion I were higherthat those for Station II.     

Contribution of heterotrophic plankton to nitrogen regeneration in the upwelling ecosystem of A Coruna (NW Spain)

The contribution of heterotrophic plankton to nitrogen (N) regenerationin the water column, and its significance for the requirementsof phytoplankton, were studied at the seasonal scale in thecoastal upwelling ecosystem of A Coruña (Galicia, NWSpain). During 1995–1997, monthly measurements were takenof hydrographic conditions, dissolved nutrients, and abundanceand biomass of microplanktonic heterotrophs (bacteria, flagellatesand ciliates), phytoplankton and mesozooplankton (>200 µm).Additionally, series of experiments were conducted to quantifyN fluxes, including primary production (¹⁴C method), phytoplanktonuptake of nitrate, ammonium and urea (¹⁵N-labelling techniques),microheterotrophic regeneration of ammonium, mesozooplanktongrazing (chlorophyll gut-content method) and excretion of ammoniumby mesozooplankton. Two N budgets were built for the averagesituations of high (>100 mg C m^-2 h^-1) and low (<100 mgC m^-2 h^-1) primary production. The results revealed that phytoplanktonrelied strongly on regenerated ammonium all year round (33 and43% of total N uptake in high and low production situations,respectively). This demand for ammonium was closely matchedby regeneration rates of microplankton (0.14–0.25 mmolN m^-2 h^-1), whereas zooplankton contributed on average <10%to N regeneration. Likewise, zooplankton grazing had littledirect control on phytoplanktonic biomass. The results obtainedindicate that in the A Coruña upwelling system, N biomassof heterotrophic plankton is generally higher than phytoplanktonN biomass. The high rates of N regeneration measured also suggestthat a large proportion of the organic matter produced afteran upwelling pulse is recycled in the water column through themicrobial food web.     

Contrasting Incorporation of 2H from 2H2O into ABA, Xanthoxin and Carotenoids in Tomato Shoots

Nonhebel, H. M. and Milborrow, B. V. 1987. Contrasting incorporationof ²H from ²H²O into ABA, xanthoxin and carotenoids in tomatoshoots.—J. exp. Bot. 38: 980–991. The incorporation of ²H into abscisie acid, xanthoxin, ß-carotene,lutein, lutein epoxide and violaxanthin in tomato shoots incubatedfor 6 d in 70% ²H₂O was compared to investigate whether thesecompounds are precursors of abscisie acid. On average, 5% ofabscisie acid molecules became labelled with a single ²H atomand 21% with from 3 to 14 atoms of ²H. However, mass spectralanalysis of endogenous xanthoxin extracted from the same plants,in darkness, under nitrogen, and derivatized to the pentafluorobenzyloximeshowed incorporation of only single ²H atoms, ruling out xanthoxinas an abscisie acid precursor. Normal-phase HPLC analysis oftomato shoot extracts showed four major carotenoid peaks; ß-carotene,lutein, lutein epoxide and violaxanthin. Calculations basedon the measured carotenoid pool sizes and on the calculatedminimum pool size of the ABA precursor predicted that at least6·8% of violaxanthin molecules or 7·9% of luteinepoxide molecules should become labelled with from 3 to 14 ²Hatoms if these molecules are precursors of abscisie acid. However,mass spectral analysis of xanthoxin derived from purified violaxanthinand lutein epoxide showed no molecules with more than a single²H atom, with detection limits of less than 1% and 0·2%respectively. Similarly, mass spectra of ß-caroteneand lutein did not show any ²H. We conclude that these carotenoidsare not precursors of abscisie acid. Key words: Abscisic acid, xanthoxin, carotenoids     

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     

The influence of irradiance on growth, photosynthesis and respiration of Gyrodinium cf. aureolum

The bloom-forming marine dinoflagellate Gyrodinium cf. aureolumwas grown in batch cultures over a range of irradiances (35–380µmolm^–2 s^–1 and growth, photosynthesis and respirationrates determined. Saturation of growth occurred at irradiancesof 100µmol m^–2 s^–1 Below this light level,decreases in growth rates and cell size, and a relative increasein carbon specific respiration rates, were observed. On theother hand, photosynthesis-irradiance relationships determinedfrom dissolved oxygen incubations showed that on a cellularand carbon basis, cultures grown at low irradiances had higherrates of light-limited and light-saturated photosynthesis, mainlyas a result of large increases in cell chlorophyll content.This adaptation strategy enables low-light-grown organisms toexploit available high irradiance through a relatively highphotosynthetic capacity. In cells grown at higher light levels(>100µmol m^–2 s^–1), excess photosynthatemay be diverted to storage rather than used for growth.     

Three distinct mechanisms of HCO3- secretion in rat distal colon

HCO₃^– secretion has long been recognized in the mammalian colon, but it has not been well characterized. Although most studies of colonic HCO₃^– secretion have revealed evidence of lumen Cl^– dependence, suggesting a role for apical membrane Cl^–/HCO₃^– exchange, direct examination of HCO₃^– secretion in isolated crypt from rat distal colon did not identify Cl^–-dependent HCO₃^– secretion but did reveal cAMP-induced, Cl^–-independent HCO₃^– secretion. Studies were therefore initiated to determine the characteristics of HCO₃^– secretion in isolated colonic mucosa to identify HCO₃^– secretion in both surface and crypt cells. HCO₃^– secretion was measured in rat distal colonic mucosa stripped of muscular and serosal layers by using a pH stat technique. Basal HCO₃^– secretion (5.6 ± 0.03 µeq·h^–1·cm^–2) was abolished by removal of either lumen Cl^– or bath HCO₃^–; this Cl^–-dependent HCO₃^– secretion was also inhibited by 100 µM DIDS (0.5 ± 0.03 µeq·h^–1·cm^–2) but not by 5-nitro-3-(3-phenylpropyl-amino)benzoic acid (NPPB), a Cl^– channel blocker. 8-Bromo-cAMP induced Cl^–-independent HCO₃^– secretion (and also inhibited Cl^–-dependent HCO₃^– secretion), which was inhibited by NPPB and by glibenclamide, a CFTR blocker, but not by DIDS. Isobutyrate, a poorly metabolized short-chain fatty acid (SCFA), also induced a Cl^–-independent, DIDS-insensitive, saturable HCO₃^– secretion that was not inhibited by NPPB. Three distinct HCO₃^– secretory mechanisms were identified: 1) Cl^–-dependent secretion associated with apical membrane Cl^–/HCO₃^– exchange, 2) cAMP-induced secretion that was a result of an apical membrane anion channel, and 3) SCFA-dependent secretion associated with an apical membrane SCFA/HCO₃^– exchange. chloride/bicarbonate exchange; short-chain fatty acid/bicarbonate exchange; anion channel; pH stat     

Plankton in the River Rhine: structural and functional changes observed during downstream transport

The growth dynamics of phytoplankton, zooplankton and bacterioplanktonin the River Rhine were analysed simultaneously with a numberof environmental factors in order to identify environmentalsteering factors and to describe some of their interrelations.Observations on the metabolic activity (for algae and bacteria)and density (for all organisms) were carried Out three timesin 1990 using successive sampling of the same water parcel duringits transport in the lower 660 km reach of the river. High algaldensities (up to 170.5 µg chlorophyll a l^–1), rotifers(up to 1728 l^–1), crustaceans (up to 65 l^–1) andbacteria (up to 16x10⁹ l^–1) were found. Algae and rotifersshowed a rapid successive development during transport, whilecrustaceans were only abundant in the tidal reach of the river.In May, a vigorous growth of phytoplankton, zooplankton andbacteria was found. The diatom-dominated phytoplankton depletedthe dissolved silicate in the river water and this led to acollapse of the populations, indicated by a decreased specificrate of photosynthesis (measured by the ¹⁴C method) and vigorousbacterial growth (measured by [³H]thymidine incorporation).Subsequently, the remaining phytoplankton diminished to verylow levels near the river mouth. In July and September, it seemedthat biological interactions within the plankton populationsor between plankton and benthos balance the population densitiesso that separate developmental stages, as in spring, were lessprominent. Estimates of the growth rates and loss rates of thephytoplankton were made. Phytoplankton exerted a substantialinfluence on the partitioning of nutrients (nitrogen, phosphorus,silicate) over water and suspended matter (as analysed by elementanalysis). It seems likely that only the reduction of phosphate,as planned under the Rhine Action Programme, and not that ofnitrogen, may restrict the peaks of plankton growth describedhere.     

The Effect of Selected Growth-Promoting and Growth-Inhibiting Substances on the Extension of Segments of Etiolated Tomato Seedling Hypocotyls

The effects of a number of growth-promoting and growth-inhibitingsubstances, including two fungal toxins, were studied on theextension of segments of etiolated tomato seedling hypocotyls.The bioassay was sensitive to small quantities of NaF, coumarinand ₂, ₄-DNP and inhibition was observed at all concentrations.₂, ₄-DNP or Iodoacetate stimulated growth at concentrationsbetween 1? 10^–4 and 5 ? 10^–6M. or 1 ? 10^–6and 1 ? 10^–7M. respectively. Inhibitor experiments inbuffered nutrient solution were approximately 10 per cent. moresensitive than those in deionized water. By means of paper partition chromatography small quantitiesof two fungal toxins, fusaric and alternaric acid were chromatographedand bioassayed. The effect of fussric acid (5, n-butyI picolinicacid) on hypocoty1 growth was detected at concentrations aslow as 1 ? 10^–5M. Experiments with recongnized growth-promoting substances showedthat Kinetin inhibited growth at concentrations up to 1 ?10^–8M.in both light and dark. IAA inhibited growth up to 1 ? 10^–6M.At 1 ? 10^–7 and 1 ? 10^–8 only small increases occurredwith IAA and the effect of light was negligible. Gibberellicacid (GA₂)stimulated growth at concentrations from 10^–3to 10^–7M. and significant increases up to 17 per cent.were recorded in the light. Since the light induced inhibitionwas only partly restored, the existence of some other naturallight sensitive growth substance is suggested. The value ofthe bioassay as a method for estimating natural growth-inhibitingand growth-promoting substances is discussed.     

Biophysical and Biochemical Regulation of Cytoplasmic pH in Chara corallina during Acid Loads

The contribution of membrane transport to regulation of cytoplasmicpH in Chara corallina has been measured during proton-loadingby uptake of butyric acid. In the short-term (i.e. up to 20min) uptake of butyric acid is not affected by removal of externalK⁺, Na⁺ or Cl^– but over longer periods uptake is decreased(by 20–50% in different experiments) in the absence ofexternal Na⁺ or, sometimes, K⁺. Influxes of both Na⁺ and K⁺increase temporarily after addition of butyrate, Na⁺ immediatelyand K⁺ after a lag. Effects on Cl^– influx are small butCl^– efflux increases enormously after a short lag. Anapproximate comparison of internal butyrate with changes inthe concentration of K⁺, Na⁺, and Cl^– suggests that initially(i.e. for a few min) cytoplasmic pH is determined by bufferingand possibly by some decarboxylation of organic acids (biochemicalpH regulation), and that biophysical pH regulation involvingefflux of H⁺ balanced by influxes of K⁺, Na⁺ and especiallyefflux of Cl^– progressively becomes dominant. When butyric acid is washed out of the cells, cytoplasmic pHis restored completely or partially (depending on the butyrateconcentration used) and this is independent of the presenceor absence of external Cl^–. Where Cl^– is present,its influx is relatively small. It is suggested that cytoplasmicpH is then controlled biochemically, involving the synthesisof an (unidentified) organic acid and the accumulation of acidicanions in place of butyurate lost from the cell. During thesecond application of butyrate, net Cl^– efflux is small:it is suggested that control of cytoplasmic pH then involvesdecarboxylation of the organic acid anions. The questions of the source of Cl^– lost from the cell(cytoplasm or vacuole) and of possible cytoplasmic swellingassociated with the accumulation of butyrate are discussed. Key words: Chara corallina, butyric acid, cytoplasmic pH, membrane transport     

Fatty acid composition of microsomal phospholipids and H+-ATPase activity in the roots of Scots pine seedlings grown at different root temperatures during flushing

The fatty acid composition of phospholipids in the microsomesand the vanadate-sensitive H⁺-ATPase activity of the roots ofone-year-old Scots pine (Pinus sylvestris L.) seedlings werestudied during flushing in spring. The seedlings in hydroponiccultures were subjected to different root temperatures (5, 12or 20°C). The shoot was maintained at 20/15° C (day/night)during the 35 d experiment. After 35 d at 5° C, root growthwas totally inhibited and shoot growth partly inhibited. In roots grown at 5° C the fatty acid composition of themicrosomal phospholipids and the degree of fatty acid unsaturation(bond index) were unchanged, while in roots grown at 12 and20° C the fatty acid composition changed and bond indexdecreased. At those root temperatures, the most obvious changewas a decline in the proportion of linolenic acid (C18:3). Inthe new white roots grown either at 12°C or 20°C theproportion of C18:2 was higher and the proportion of C18:3 lowerthan in 1-year-old roots. Independently of root temperature,H⁺-ATPase activity, determined on a fresh weight basis, declinedto half of the original activity during the experiment. Thedecline in H⁺ -ATPase activity was most rapid during the firstweek. In the old roots the decline in H⁺-ATPase activity followedclosely the decline in amount of membrane protein. In new rootsH⁺-ATPase activity was high and increased with increasing roottemperature. These results suggest that in the roots of Scotspine seedlings, vanadate-sensitive H⁺-ATPase activity is dependenton age, while changes in the microsomal fatty acid compositionof phospholipids are regulated mainly by root temperature. Key words: Fatty acids of phospholipids, microsomes, H⁺-ATPase, root temperature, Scots pine     

Biosynthesis of folic acid compounds in plants VI. The occurrence and properties of the dihydrofolate-synthesizing enzyme in pea seedlings

An enzyme, which catalyzes the formation of dihydrofolate fromdihydropteroic acid and L-glutamic acid, was found in pea seedlings.The enzyme was purified approximately 25-fold from the crudeextracts of pea seedlings, and its some properties were investigated.Optimum pH for the enzyme activity was found to be 8.8. Pteroicand tetrahydropteroic acids were not active as substrate. Theenzymatic reaction required as cofactors ATP, divalent (Mg²⁺or Mn²⁺) and univalent (K⁺, NH₄⁺ or Rb⁺) cations. The productwas characterized as dihydrofolic acid by bioautography. MICHAELIS constants for L-glutamic acid, ATP, dihydropteroicacid and Mg²⁺ were 7.0x10^–4, 9.0x10^–5, 3.5x10^–6and 1.2x10^–3 M, respectively. The MICHAELIS constant forMn²⁺ was 3.0x10^–4. The enzyme was inhibited by PCMB orsilver nitrate and, to some extent, by L-aspartic acid. Inhibitionby PCMB was completely reversed by addition of 2-mercaptoethanol.Enzyme activity was distributed widely among plants. The importanceof magnesium and potassium ions for enzyme catalysis is discussed. ¹For the previous paper, Part V, see Reference (30). (Received March 28, 1970; )     

Ageing of Cucumber and Onion Seeds: Phospholipase D, Lipoxygenase Activity and Changes in Phospholipid Content

Cucumber (Cucumis sativus L. cv. Mesa) and onion (Allium cepaL. cv. Rijnsburger Heldis) seeds were rapidly aged at 40 °Cand 74% relative humidity. Onion seeds were also slowly agedat 40 °C with 15% relative humidity for 11 months and onemore month at 28% relative humidity. Significant loss of totaland individual phospholipids was an early event during bothstorage treatments. With slow ageing of onion, loss of phosphatidylcholineoccurred several months before loss of viability and vigourwas detected. Phosphatidic acid, the lipid product of phospholipaseD action, increased during rapid ageing of both cucumber andonion. Phosphatidic acid was present in onion seeds before theageing treatments and its content remained unchanged in theslowly aged seeds. There was 1600 (cucumber) and 2000 (onion)times more phospholipase D activity (6 x 10⁵ and 2·9x 10⁵ nmol g^–1 d^–1 in cucumber and onion, respectively)in crude extracts from non-aged seeds than was required to accountfor the fastest fall in phospholipids (72, 372 and 144 nmolg^–1 d^–1 for cotyledons and radicles of cucumberand onion, respectively, over the first 9 d [cucumber] or 1d [onion] of ageing) and fastest increase in phosphatidic acid(7, 162 and 37 nmol g^–1 d^–1). How accurate a guidethe in vitro activity of phospholipase D was to the in vivoactivity was unclear. However, the considerable excess activityseen with the formation of phosphatidic acid supports the proposalthat hydrolysis of phospholipids by phospholipase D is a firststep in deterioration during ageing. Substantial lipoxygenaseactivity was also detected (58 x 10³ and 54 x 10³ nmol g^–1d^–1 respectively, for non-aged cucumber and onion seeds).However, the increase in conjugated dienes (an early productof peroxidation) in ageing cucumber seeds was comparativelysmall (90 nmol g^–1 d^–1 over 21 d ageing), and increasein malondialdehyde could not be detected, indicating that peroxidationmay not have been a major factor in cucumber. The increase inconjugated dienes during rapid ageing of onion seeds was larger(1·5 x 10³ nmol g^–1 d^–1 over days 0–2of rapid ageing), much greater than the decrease in phospholipidacyl groups (260 nmol g^–1 d^–1 over days 0–2of rapid ageing) indicating the occurrence of peroxidation offatty acids released from reserve as well as from membrane lipids.This higher level of conjugated dienes during onion ageing wasthe main difference between cucumber and onion, indicating thatthe level of peroxidation could be an important difference betweenfast and slow ageing seeds. However, peroxidation is not theonly possible deleterious process since hydrolysis of the normalmembrane phospholipids to phosphatidic acid increased the contentof non-bilayer-forming lipids and this too could be a directmembrane-destabilizing consequence of phospholipase D actionduring ageing. Key words: Cucumis sativus L. (cucumber), Allium cepa L. (onion), seed ageing, phospholipase D, lipoxygenase, phospholipids