Production of N-glycolylneuraminic acid derived from chemically modifiedE. coli Kl bacterial polysaccharide by membrane enclosed enzymatic catalysis

Summary N-Glycolylneuraminic acid (Neu5Gc) has been prepared by enzymatic hydrolysis of its -(28) linked homopolymer. The rate of hydrolysis of the natural poly -(28)-(Neu5Ac) and the semi-synthetic poly -(28)-(Neu5Gc) were compared with the neuraminidases fromClostridium perfringens andVibrio cholerae. The natural Neu5Ac polysaccharide was a better substrate for both enzymes. For comparison, acid hydrolysis of the two polysaccharides showed extensive degradation.     

Alkyl-phenyl cleavage of the lignin model compounds guaiacylglycol and glycerol-β-guaiacyl ether by Phanerochaete chrysosporium

The white rot basidiomycete Phanerochaete chrysosporium metabolized guaiacylglycol--guaiacyl ether (I) in high nitrogen, shaking and stationary cultures. 2-(o-Methoxyphenoxy) ethanol (X), 2-(o-methoxyphenoxy) acetic acid (IX) and methoxy-phydroquinone (MHQ) were identified as products of the metabolism of (I). P. chrysosporium also metabolized guaiacylglycerol--guaiacyl ether (IV) in high nitrogen stationary cultures. 2-(o-Methoxyphenoxy)-1,3 propanediol (XII) and 3-hydroxy, 2-(o-methoxy-phenyxy) propionic acid (XIV) were identified as products of the metabolism of (IV). Finally, P. chrysosporium metabolized -deoxyguaiacylglycol--guaiacyl ether (VI) and -deoxyguaiacylglycerol--guaiacyl ether (VII) in limiting nitrogen cultures. 2-(o-Methoxyphenoxy) ethanol (X) and 2-(o-methoxyphenoxy)-1,3 propanediol (XII) were identified as products of the metabolism of VI and VII respectively indicating hydroxylation of those substrates with subsequent alkyl-phenyl bond cleavage. Metabolites were identified after comparison with chemically synthesized standards by GLC-mass spectrometry.Abbreviations GLC Gas liquid chromatography - TMSi trimethylsilyl - TLC thin layer chromatography - MHQ methoxyhydroquinone     

Parameters influencing transient and stable transformation of barley (Hordeum vulgare L.) protoplasts

Cat gene expression has been investigated following PEG-mediated plasmid uptake into barley protoplasts. The uptake conditions optimised for transient expression were employed for stable transformation. Transformed protoplast-derived calli of the cvs. Dissa and Igri, were selected on medium containing G418 at 40 g ml^–1 or kanamycin sulphate at 250 g ml^–1. Absolute transformation frequencies of 28.9×10^–5 and 21.3×10^–5 were recorded for Dissa with kanamycin sulphate and G418 selection, respectively. The frequency for Igri was 11.5×10^–5 with G418 selection. Antibiotic resistant protoplast-derived colonies expressed NPTII activity; Southern hybridisation confirmed integration of the nptII gene into barley genomic DNA.Abbreviations ABA abscisic acid - AC-CAP acetylated chloramphenicol - BAP 6-benzylaminopurine - cat chloramphenicol acetyltransferase gene - CAT chloramphenicol acetyltransferase activity - CaMV cauliflower mosaic virus - CAP chloramphenicol, 2,4-d-2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - G418 Geneticin - gus -glucuronidase gene - HEPES (N[2-hydroxyethyl] piperazine-N-[2-ethanesulphonic acid]) - IAA indole acetic acid - MES 2-N-morpholinoethane sulphonic acid - NAA -naphthaleneacetic acid - npt II neomycin phosphotransferase gene - NPTH neomycin phosphotransferase activity - PEG polyethylene glycol - SCV settled cell volume     

A genetically related response to iron deficiency stress in muskmelon

A mutant muskmelon (Cucumis melo L.) with characteristic Fe-deficiency chlorosis symptoms was compared to related cultivars in its ability to obtain Fe via the widely known Fe-stress response mechanisms of dicotyledonous plants. The three cultivars (fefe, the Fe-inefficient mutant; Mainstream and Edisto, both Fe efficient plants) were grown in nutrient solution in either 0 or 3.5 mg L^-1 Fe as FeCl₃. None of the three cultivars released reductants or phytosiderophores, but both Edisto and Mainstream produced massive amounts of H+ ions to reduce and maintain the pH of nutrient solutions below pH 4.0. The roots of these two Fe-efficient cultivars were also capable of reducing Fe³⁺ to Fe²⁺. These responses maintained green plants, resulted in high leaf Fe in both Edisto and Mainstream, and produced Mn toxicity in Mainstream. The lack of Fe-deficiency stress response in fefe not only affected leaf Fe concentration and chlorosis, but also resulted in reduced uptake of Mn. The importance of reduced Fe (Fe²⁺) to the Fe-efficient cultivars was confirmed by growing the cultivars with BPDS (4, 7-diphenyl-1, 10-phenanthroline disulfonic acid, a ferrous chelator) and EDDHA [ethylene-diamine di (0-hydroxphenylacetic acid)] (a ferric chelator), and observing increased chlorosis and reduced Fe uptake in BPDS grown plants. The Fe-deficiency response observed in these cultivars points out the diversity of responses to Fe deficiency stress in plants. The fefe mutant has a limited ability to absorb Fe and Mn and perhaps could be used to better understand Mn uptake in plants.     

Lactic acid excretion via carrier-mediated facilitated diffusion in Lactobacillus helveticus

Summary During growth on a complex medium containing 2% (w/v) lactose, Lactobacillus helveticus produced about 180 mm lactate. Due to the acidification, the external pH decreased to 3.7. The pH remained constant at a level of 0.5–0.7 units (40 mV), and µ_Lac decreased gradually from –60 to 0 mV. The mechanism of lactate extrusion was studied with resting cells. Upon dilution of lactate-loaded cells in a buffer containing [¹⁴C]-lactate, a typical counterflow was observed, suggesting that a carrier system was employed in lactate excretion. Influx of lactate could not be driven by an artificial membrane potential, indicating that lactate was electroneutrally transported. By examining efflux under various lactate anion and lactic acid concentrations, the undissociated form of the acid was shown to influence the velocity of the transport process. A pH-dependent apparent K _m value of the carrier system was observed in efflux experiments with increasing internal lactate concentrations. It was concluded that the mode of end-product excretion can be defined as a carrier-mediated facilitated diffusion with the undissociated lactic acid or the lactate anion in symport with one proton, respectively, as the object of transport.Abbreviations L _tota total lactate - L _undissb free lactic acid - L _dissc lactate anion - pH_ed external pH - pH_ie internal pH - pH transmembrane H⁺ gradient - µ_Lacf transmembrane gradient of total lactate - µ_HLg transmembrane gradient of the free lactic acid - µ_Lh transmembrane gradient of the lactate anion - V _Effii efflux velocity Offprint requests to: G. Gottschalk     

Denaturation behavior of α-chymotrypsinogen A in urea and alkylurea solutions: Fluorescence studies

The solvent denaturation of-chymotrypsinogen (-ctg A) in aqueous solution of urea, methyl-,N,N-dimethyl-, ethyl-, propyl- and butylurea was studied by fluorescence measurements. Data were analyzed on the assumption of a two-state approximation to obtain the apparent equilibrium constant,K and the apparent Gibbs free energy of transition G ⁰ . It has been observed that alkylsubstitution of urea significantly lowers the denaturant concentration needed to denature-ctg A at 25°C. Denaturation was accompanied by the red shift of emission maxima, the increase of the half-width of the fluorescence spectra, the increase of the fluorescence intensity, and the decrease of the fluorescence polarization. The differences of these fluorescence parameters observed for-ctg A in alkylureas and urea can be ascribed to different unfolded states of the protein in different denaturant solutions. Minor differences in the extent of unfolding were confirmed by size-exclusion chromatography.     

The influence of plant cover on surfperch abundance at an offshore temperate reef

Synopsis At an offshore reef near Santa Barbara, southern California, young-of-year (young) of five surfperch species (Embiotocidae: Embiotoca jacksoni, E. lateralis, Hypsurus caryi, Rhacochilus toxotes, Damalichthys vacca) once thrived in a dense kelp understory of Pterygophora californica and Laminaria farlowii, but disappeared after a severe storm in February, 1980 deforested their habitat. Measurements of fish density and kelp cover made before deforestation indicated that the young surfperch appeared in the spring and disappeared in the fall as cover increased and declined. Adult surfperch and large kelp bass (Paralabrax clathratus), which can eat young but not adult surfperch, remained all year. We tested to see if kelp cover was an essential refuge for the young by pruning back (thinning) kelp blades from one of two transects. The thinning caused a significant redistribution of young which clearly avoided open spaces, but not of adults which are less vulnerable to predation. Measurements of fish density made after the storm-induced deforestation showed that adult surfperch and kelp bass still remained abundant even after the young surfperch were gone. Only after an abrupt reforestation in 1983, more than a year after the present study was terminated, were young once again seen on the reef. Although young surfperch may seek tiny prey living on kelp blades, most lines of evidence indicated that the distribution of the young is more a response to risk of predation. Hence the extent of kelp understory was probably the main determinant of the survival of young surfperch on the reef.     

Purification and properties of a novel β-galactosidase or exo-(1 → 4)-β-d-galactanase from the cotyledons of germinated Lupinus angustifolius L. seeds

The main polysaccharide component of the thickened cell walls in the storage parenchyma of Lupinus angustifolius L. cotyledons is a linear (1 4)--linked d-galactan, which is mobilised after germination (L.A. Crawshaw and J.S.G Reid, 1984, Planta 160, 449–454). The isolation from the germinated cotyledons of a -d-galactosidase or exo-(1 4)--d-galactanase with a high specificity for the lupin galactan is described. The enzyme, purified using diethylaminoethyl-cellulose, carboxymethyl-cellulose and affinity chromatography on lactose-agarose, gave two bands (major 60 kDa, minor 45 kDa) on sodium dodecyl sulphate-gel electrophoresis, and two similar bands on isoelectric focusing (major, pI 7.0, minor pI 6.7, both apparently possessing enzyme activity). The minor component cross-reacted with an antiserum raised against, and affinity-purified on, the major band. Both components had a common N-terminal sequence. The minor component was probably a degradation product of the major one. The enzyme had limited -galactosidase action, catalysing the hydrolysis of p-nitrophenyl--d-galactopyranoside and (1 4)- and (1 6)--linked galactobioses. Lactose [-d-galactopyranosyl-(1 4)-d-glucose] was hydrolysed only very slowly and methyl--d-galactopyranoside not at all. Lupin galactan was hydrolysed rapidly and extensively to galactose, whereas other cell-wall polysaccharides (xyloglucan and arabinogalactan) with terminal non-reducing -d-galactopyranosyl residues were not substrates. A linear (1 4)--linked galactopentaose was hydrolysed efficiently to the tetraose plus galactose, but further sequential removals of galactose to give the tetraose and lower homologues occurred more slowly. Galactose, -galactonolactone and Cu⁺² were inhibitory. No endo--d-galactanase activity was detected in lupin cotyledonary extracts, whereas exo-galactanase activity varied pari passu with galactan mobilisation. Exo-galactanase protein was detected, by Western immunoblotting of cotyledon extracts, just before the activity could be assayed and then increased and decreased in step with the enzyme activity. The exo-galactanase is clearly a key enzyme in galactan mobilisation and may be the sole activity involved in depolymerising the dominant (1 4)--galactan component of the cell wall.Abbreviations CM carboxymethyl - DEAE diethylaminoethyl - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - TLC thin-layer chromatography We wish to thank CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) for the award of a studentship to M.S. Buckeridge, and the Government of São Paulo State, Brazil for granting him leave of absence. We are grateful to Dr. Amanda Heyller (Unilever Research Laboratory, Colworth House, Bedford, UK) for N-terminal sequence determinations, to Dr. Stuart Wilson (Stirling) for preparing gelatin SDS-gels and to Cristina Fanutti (Stirling) for purifying the xyloglucan oligosaccharide.     

The nucleotide sequence of a transposable haloalkanoic acid dehalogenase regulatory gene (dehR I) fromPseudomonas putida strain PP3 and its relationship with σ54-dependent activators

The mobile genetic element,DEH found inPseudomonas putida PP3 carries a 2-haloalkanoic acid dehalogenase structural gene,dehI, and its associated regulatory gene,dehR _I. The nucleotide sequence ofdehR _I was determined. The gene had an open reading frame putatively encoding for a 64 kDa protein containing 571 amino acid residues. The protein was similar to previously published sequences of several other ⁵⁴-dependent activator proteins. Amino acid sequence analysis showed that the deduced DehR_I protein clustered with the NifA nitrogenase regulatory activator family, and was most closely related, with 47.7% similarity, to a NifA-like deduced partial sequence from a plasmid-encoded ORF inPseudomonas sp. strain NS671, associated with L-amino acid production. The domain structure of DehR_I was analysed by alignment with other NifA-like and NtrC-like sequences and showed a highly conserved central region of approximately 230 amino acids, and a potential DNA-binding domain. No homology was detected between the deduced DehR_I and other ⁵⁴-dependent activator sequences at the N-terminus, a result which was consistent with that region being the domain which recognised inducer.     

Evaluation of laboratory indexes of absorption of soil phosphorus by plants: II

Summary A comparison was made of several laboratory methods for estimating the yield of phosphorus in plants grown in greenhouse cultures on samples of 22 soils from different parts of the United States. The methods investigated and their rank in order of increasing precision of the estimates of the yield of phosphorus were as follows: extraction with lactic acid, calcium lactate buffer solution extraction with 2 per cent citric acid solution < extraction with 0.03N NH₄F, 0.025N HCl solution < percentage phosphorus saturation (found as follows: 100 × labile phosphorus by isotopic dilution/ phosphorus adsorption capacity according to Langmuir adsorption equation) labile phosphorus by isotopic dilution phosphorus extracted by water.Journal Paper No. J-3747 of the Iowa Agricultural and Home Economics Experiment Station, Ames, Iowa. Project No. 1183. Contribution from the Department of Agronomy.     

Fermentation of high-protein plant biomass by introduction of lactic acid bacteria

Lactic acid bacteria displaying increased ability to produce lactic acid, medium proteolytic activity, and tolerance to osmotic stress were isolated under selective conditions from phyllosphere and rhizosphere of registered and raised cultivars of legumes. Lactic fermentation of poorly ensilable leguminous plants (red clover and Caucasian goats rue) was performed by introduction of rifampin-resistant homofermenting representatives of the genus Lactobacillus (selected according to a set of technologically important characteristics). The results demonstrate that introduction of active local strains of lactobacteria, as well as the collection strain Lactobacillus plantarum BS933, enhances activation of ensiling and increases the quality of fodder, as assessed according to the standard criteria (a decrease in pH of the medium, the ratio of lactic acid to fatty acid homologues, and the composition of silage microflora).Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 1, 2005, pp. 79–89.Original Russian Text Copyright © 2005 by Shurkhno, Gareev, Abulkhanov, Validov, Boronin, Naumova.     

Kinetics of growth and lactic acid production in continuous and batch culture

Summary In a lactic acid fermentation by Streptococcus faecalis, the specific consumption rate of glucose (v) and the specific production rate of lactic acid () were represented by the following simple equations as functions of the specific growth rate (): 1/=(1/) + 1/ = (1/) + By use of data from a batch culture, these two equations were derived from enzyme kinetics of the product inhibition. These equations were successfully applied to the results of batch culture and chemostat culture. In addition, calculation of ATP yield by these equations agreed with the experimental results better than the conventional Leudeking-Piret type equation, which includes two terms associated with growth and not with growth. Correspondence to: H. Ohara     

Abbau von 14C-, 3H- und 36Cl-markiertem γ-Hexachlorcyclohexan durch anaerobe Bodenmikroorganismen

Zusammenfassung Durch eine anaerobe Mischflora aus Ackerboden wurde -Hexachlorcyclohexan (-HCH) in 4–5 Tagen zu 90% abgebaut. Dabei erfolgte eine schnelle Abspaltung des Chlors in Form von Chloridionen und danach eine Freisetzung des C- und H-Anteiles in Form flüchtiger Verbindungen, in denen kein Chlor und auch kein CO₂ nachzuweisen war.Die Verwendung von ¹⁴C/³H- und ³⁶Cl/³H-doppelmarkiertem -HCH zeigte, daß die Cl- und H-Abspaltung nicht im Verhältnis von 1:1 erfolgte, sondern mehr Cl als H abgespalten wurde. Die flüchtigen Verbindungen enthielten andererseits höhere ¹⁴C- als ³H-Anteile. Gaschromatographische Untersuchungen zeigten ebenfalls eine rasche Verminderung des -HCH und die Bildung verschiedener Metabolite. Es wurde jedoch kein -Pentachlorcyclohexen nachgewiesen. Bei steigenden O₂-Gehalten in der Gasphase verminderte sich der -HCH-Abbau. Jedoch fanden auch noch bei 5% O₂ Chlorabspaltung und die Freisetzung flüchtiger Metabolite statt.-HCH wurde ebenfalls, jedoch langsamer, durch die anaerobe Mischflora abgebaut. Auch hier wurde Chlorid abgespalten, und es traten ebenfalls flüchtige Verbindungen auf, die kein Chlor enthielten.

---

Degradation of ¹⁴C-, ³H- and ³⁶Cl-labelled -hexachlorocyclohexane by anaerobic soil microorganisms

Up to 90% of the -Hexachlorocyclohexane (-HCH) applied to an anaerobic mixed bacterial flora enriched from an arable soil were degraded within 4–5 days. Degradation resulted in a rapid release of chloride and in formation of chlorine-free volatile metabolites. CO₂ formation from the molecule was not detected.Investigations with ¹⁴C/³H- and ³⁶Cl/³H double-labelled -HCH indicated that the release of Cl and H did not occur in the ratio of 1:1. More Cl than H was split off. The volatile compounds contained more ¹⁴C than ³H. Gas chromatographic studies also showed the rapid decrease of -HCH and the formation of several metabolites. -Pentachlorocyclohexene was not detected. Increasing O₂-contents in the gas phase of cultures resulted in decreases of the compound's degradation. Release of chloride and of volatile metabolites were observed with O₂ contents in the gas phase up to 5%.-HCH was also, but more slowly as with -HCH, degraded by the anaerobic mixed flora. Chloride was released and volatile, chlorine-free metabolites were found.

     

Effect of pH on<Emphasis Type="Italic"> Lactobacillus fermentum</Emphasis> growth,raffinose removal, α-galactosidase activity and fermentation products

In this study, the behaviour of Lactobacillus fermentum CRL 722 and CRL 251 were evaluated under different pH conditions (pH 6.0, 5.5, 5.0, 4.5) and without pH control. Growth was similar under all conditions assayed except at pH 4.5. These microorganisms were able to eliminate raffinose, a nondigestible -oligosaccharide (NDO) found in soy products, showing a consumption rate of 0.25 g l^–1 h^–1 (pH 6.0–5.0). The removal of raffinose was due to the high -galactosidase (-gal) activities of these lactic acid bacteria, which was highest at pH 5.5 (5.0 U/ml). The yield of organic acids produced during raffinose consumption was also highest at this pH. The results of this study will allow selection of the optimum growth conditions of L. fermentum with elevated levels of -gal to be used in the reduction of NDO in soy products when used as starter cultures.     

Plant regeneration from isolated microspores of linseed (Linum usitatissimum L.)

Embryogenesis and plant regeneration were induced in isolated microspore culture of linseed (oilflax, Linum usitatissimum). Microspores underwent cell divison which led to either microcallus or embryoid formation, when they were cultured in a modified liquid Nitsch-Lichter-Nitsch medium (Lichter 1985) at two different incubation temperatures (30 and 35 °C); some embryoids and microcalli further developed to larger calli. After transfer of the microspore derived calli to a solid medium containing zeatin (Img 1^–1) shoot induction was achieved from 36 to 66% of the calli. The highest frequency of regenerated plants was obtained in microspore cultures of the hybrid Atalante x Szegedi 62 (F₁) at 30 °C, whereas for the second genotype Pedigree 2 x Kiszombori 41 (F₂) the higher incubation temperature seemed to be more efficient. Shoots could be successfully rooted on an indole acetic acid containing medium and then transplanted to vermiculite and finally to soil. Most of the plants survived the transfer into soil in the greenhouse, where they could be successfully grown to maturity.Abbreviations BAP 6-benzylaminopurine - 2,4D dichlorophenoxyacetic acid - IAA indole acetic acid - N6 Chu (1978) medium - NAA naphthaleneacetic acid - NLN Nitsch-Lichter-Nitsch (1985) - MS Murashige and Skoog (1962) medium - ZEA zeatin     

Microbial population and biochemical changes in fermenting finger millet (Eleusine coracana)

Natural fermentation of finger millet (Eleusine coracana) was carried out for 48 h. Microbiological and chemical analysis was performed throughout the fermentation process. The fermentation was heterolactic dominated by lactic acid bacteria accompanied by the production of lactic and acetic acids with decrease in pH and increase in titratable acidity. The microbial population increased until 18 to 24 h accompanied by a rapid decrease in total and reducing sugars. The microflora stabilized between 24 and 48 h, during which time the total and -amylase activities increased with accumulation of sugars. Total free amino acids also increased. Yeast counts were low and moulds and coliforms were absent. Repeated fermentations showed consistency in the qualitative and quantitative changes in microflora. Five predominant types of bacteria, strains belonging to Leuconostoc, Pediococcus and Lactobacillus were identified. Of these only one type, Pediococcus, dominated (>80%) in the latter half of fermentation.     

Comparison of the kinetic properties,inhibition and labelling of the phosphate translocators from maize and spinach mesophyll chloroplasts

The kinetic properties of the phosphate translocator from maize (Zea mays L.) mesophyll chloroplasts have been determined. We have used a double silicone-oil-layer centrifugation system in order to obtain true initial uptake rates in forward-reaction experiments. In addition, it was possible to perform back-exchange experiments and to study the effects of illumination and of preloading the chloroplasts with different substrates on transport. It is shown that the phosphate translocator from mesophyll chloroplasts of maize, a C₄ plant, transports inorganic phosphate and phosphorylated C₃ compounds in which the phosphate group is linked to the C₃ atom (e.g. 3-phosphoglycerate and triose phosphate). The affinities of the transported metabolites towards the translocator protein are about one order of magnitude higher than in mesophyll chloroplasts from the C₃ plant, spinach. In contrast to the phosphate translocator from C₃-mesophyll chloroplasts, that of C₄-mesophyll chloroplasts catalyzes in addition the transport of C₃ compounds where the phosphate group is attached to the C₂ atom (e.g. 2-phosphoglycerate, phosphoenolpyruvate). The phosphate translocator from both chloroplast types is strongly inhibited by pyridoxal-5-phosphate (PLP), 2,4,6-trinitrobenzenesulfonic acid and 4,4-diisothiocyanostilbene-2,2-disulfonic acid (DIDS). In the case of the spinach translocator protein these inhibitors were shown to react with the same amino-acid residue at the substrate binding site, and one molecule of either DIDS or PLP is obviously required per substrate binding site for the inactivation of the translocation process. In the functionally active dimeric translocator protein only one substrate-binding site appears to be accessible at a particular time, indicating that the site might be exposed to each side of the membrane in turn. Using [³H]-H₂DIDS for the labelling of maize mesophyll envelopes the radioactivity was found to be associated with two polypeptides of about 29 and 30 kDa. Since Western-blot analysis showed that only the 30 kDa polypeptide reacted with an antiserum directed against the spinach phosphate translocator protein it is suggested that this polypeptide presumably represents the phosphate translocator from maize mesophyll chloroplasts.Abbreviations DIDS 4,4-diisothiocyanostilbene-2,2-disulfonic acid - PEP phosphoenolpyruvate - 2-,3-PGA 2-,3-phosphoglycerate - PLP pyridoxal-5-phosphate - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis - TNBS 2,4,6-trinitrobenzenesulfonic acid - triose P triose phosphate This work was supported by the Deutsche Forschungsgemeinschaft     

Nocturnal water storage in plants having Crassulacean acid metabolism

Measurements of water uptake and transpiration, during the dark period of plants having Crassulacean acid metabolism (CAM) allow calculation of leaf-volume changes (V). Nocturnal leaf-volume changes of CAM plants have also been reported in the literature on the basis of waterdisplacement measurements. A third way of estimation is from measurements of turgor changes and cellular water-storage capacity using the pressure probe, cytomorphometry and the Scholander pressure chamber. An extension of the interpretation of results reported in the literature shows that for leaf succulent CAM plants the three different approaches give similar values of V ranging between 2.3 and 10.7% (v/v). It is evident that nocturnal malic-acid accumulation osmotically drives significant water storage in CAM leaf tissue.Abbreviations and symbols C_c water-storage capacity - E transporation (evaporational water loss) - P turgor pressure - U water uptake - V cell volume - cell-wall elastic modulus - osmotic pressure - CAM Crassulacean acid metabolism     

Mechanism of inhibition of cytoplasmic streaming by myrmicacin (Β-hydroxydecanoic acid) inChara andSpirogyra

Summary Myrmicacin (-hydroxydecanoic acid) applied externally inhibited the cytoplasmic streaming ofChara cells in acidic (pH 4.5) but not in neutral (pH 7.5) solution. Application by intracellular perfusion at neutral pH also did not inhibit the streaming. Other carboxylic acids could inhibit the streaming when given as an acidic external solution. Inhibition of the cytoplasmic streaming was concluded to occur due to acidification of the intracellular pH upon application of an acidic external medium of these carboxylic acids. The cytoplasmic streaming ofSpirogyra was also inhibited by myrmicacin and acetic acid in acidic solution.Abbreviations APW artificial pond water - CB cytochalasin B - DMGA 3.3-dimethylglutaric acid - DMSO dimethylsulfoxide - EGTA ethyleneglycol-bis-(\-aminoethyl ether)N - N tetraacetic acid - HEPES 2-hydroxyethylpiperazine - N ethanesulfonic acid - MES 2-(N-morpholino)ethanesulfonic acid - pH _i intracellular pH - Tris trishydroxyaminomethane     

Interpretations of gradients in δ13C value in thick photosynthetic tissues of plants with Crassulacean acid metabolism

In Ceropegia dichotoma, Crassula argentea, Esheveria colorata, Kalanchoë beharensis, Opuntia ficus-indica, Sansveria stuckyi and Opuntia inermis the carbon-isotope ratio ( ¹³C) of tissues close to the epidermis is 2–4.3 more negative than those in the centre of the leaf or cladode. The greatest change in ¹³C value occurs between the epidermal layer and the layer of mesophyll tissue immediately underneath. Analysis of major metabolic and structural components in successive layers of Crassula argentea grown under controlled environmental conditions conducive to Crassulacean acid metabolism confirmed that much of the variation in ¹³C values of bulk carbon is caused by differences in chemical composition. Thus the steep gradient in ¹³C value at the epidermis reflects, in part, the contribution of more-negative ¹³C values of lipids in these tissues. Moreover, during nocturnal CO₂ fixation the amount of malic acid synthesised decreases with depth and the ¹³C value of the methanol-soluble fraction is less negative with distance away from the upper epidermis. These results are consistent with diffusion limitation to CO₂ uptake in these thick leaf tissues, which also contributes to the observed gradients in ¹³C value.Abbreviation CAM Crassulacean acid metabolism - FW fresh weight - PEPCase phospoenolpyruvate carboxylase - Rubisco ribulose-1,5-bisphosphate carboxylase-oxygenase - ¹³C carbon-isotope ratio This work was supported by grants from the North Carolina Biotechnology Centre to Duke University, National Science Foundation (NSF) grant DCB90-06830, Department of Energy, Division of Energy Biosciences grant DE-FG05-89 ER 14005, and NSF grant BSR 87-06429 to Duke University Phytotron.