Partitioning behavior and enrichment of proteins with reversed micellar extraction: I. forward extraction of proteins from aqueous to reversed micellar phase

Protein extractions using aerosol OT (AOT)-isooctane reverse micelle solutions have been studied to explore the potential for separating and enriching proteins with the reversed micellar extraction. The effects of pH, ionic strength, and different cations of chlorides in a bulk aqueous phase and of AOT concentration in an organic phase on the partitioning of lysozyme and myoglobin and the solubilization of water are presented in detail. The extraction of lysozyme was affected by the concentration of potassium or barium but was almost independent of that of sodium or calcium, whose ionic diameter is smaller than that of potassium and barium. For the extraction of myoglobin, however, the effect of barium concentration was not appreciable. Lysozyme could be enriched into the reversed micellar phase up to 30 times the aqueous feed concentration. (c) 1993 John Wiley & Sons, Inc.     

An ecosystems approach to base-rich freshwater wetlands,with special reference to fenlands

A survey of base-rich wetlands in The Netherlands is presented. The main area of their occurrence is the low-lying Holocene part of the country, until some thousand years ago a large and coherent wetland landscape: the Holland wetland. The development of various parts of the Holland wetland into marshes, fens and bogs can be understood from hydrological relations in mire basins, as recognized in the distinction of primary, secondary and tertiary mire basin stages. Presently, the remnants of the Holland wetland are separate base-rich wetlands. The succession of their vegetation reflects various abiotic conditions and human influences. Three main developmental periods are distinguished as regards these factors. The first, geological period of mire development is seen as a post-glacial relaxation, with the inertia due to the considerable mass of wetland as a stabilizing factor. Biological “grazing” influences, as an aspect of utilization by humans, converted base-rich wetlands to whole new types in the second, historical period. Presently, mass and harvesting have decreased in importance, and actual successions in terrestrializing turbaries seem to reflect rapidly changing environmental conditions. Human control could well become the most important factor in the future development of wetland nature. The present value of open fen vegetation strongly depends on the continuation of the historical harvesting. The development of wooded fen may help to increase the mass of wetland in the future. Best results in terms of biodiversity are expected when their base state is maintained through water management. The vegetation and hydrology of floating fens in terrestrializing turbaries is treated in some more detail. Various lines and phases in the succession are distinguished. Open fen vegetation at base-rich, yet nutrient-poor sites is very rich in species threatened elsewhere. The fast acidification of certain such fens is attributed to hydrological and management factors. This acidification is illustrated in the profile of a floating raft sample. At the scale of these small fens, the elemental structure comprising base-rich fen, transitional fen and bog vegetation, is not as stable as it was in the large Holland wetland. A critical role seems to be played by the supply of bases with the water influx. The changing base state is supposed to change the nutrient cycling to such an extent that it would be correct to call this trophic excitation of the ecosystem, rather than just eutrophication. Eutrophication indicates a quantitative reaction to an increased nutrient supply, the internal system being unaltered. The drainage of fens, resulting in an increased productivity of the vegetation, provides another example of excitation, to the effect that the functional system is dramatically changed internally.     

Culture of rotifers with reference to some physico-chemical properties of water in nursery pond

Rotifers were cultured with five different organic and inorganic fertilizers in nursery ponds. Of the fertilizers used, mustard oil cake gave significantly (p < 0.01) higher production of rotifers than that of mohua oil cake followed by cow-dung, wheat bran, mixture of NPK and control. The higher production of rotifers was directly related with the higher doses of fertilizers. Among the rotifer species identified, the abundance of Brachionus caudatus and B. forficula were significantly (p < 0.01) higher than others. Available N, available P, exchangeable K and exchangeable Ca and exchangeable Mg were generally higher in ponds where organic fertilizers were used. Proximate composition of rotifers varied depending on the kinds of fertilizers. The multiple correlations of physico-chemical properties were highly significant (p < 0.01) with the growth and production of B. caudatus (R = 0.995), B. forficula (R = 0.932), Trichocerca capucina (R = 0.917), B. patulus (R = 0.901) and B. angularis (R = 0.892) and simply significant (p < 0.05) in the case of Keratella tropica (R = 0.880), Hexarthra intermadia (R = 0.875), B. calyciflorus (R = 0.864) and Filinia spp. (R = 0.856) contributing 91.20%, 86.86%, 84.09%, 81.18%, 79.57%, 77.44%, 76.56%, 74.65% and 73.27% of total effect of water properties on the growth of these species, respectively. The residual effect of nine different physico-chemical properties of water on the production of rotifers was 78.92% which indicates that these properties of water had only 21.08% influence on the production of rotifers.     

Seasonal succession of phytoplankton in a lake of the Paraná river floodplain,Argentina

Phytoplankton biomass, morphological and taxonomic composition, species diversity and productivity were analyzed in a shallow lake of the Middle Paraná River floodplain (El Tigre, 31 ° 41 S and 60° 42 W), between November 1986 and July 1988. Lake inundation (filling and through-flow phases) constituted an intense long-term perturbation in the physical and chemical environment. As the lake filled with river water, K-selected species (netplanktonic filamentous bluegreens, > 37 µm, with low surface area/volume (SA/V) ratios) that had existed prior to filling (late spring 1986) were replaced in summer-fall by r-selected species (nannoplanktonic chlorophytes and cryptophytes, < 37 µm, mainly stout forms with high SA/V ratios). During the through-flow phase, lentic phytoplankton was replaced by lotic flagellate populations due to the direct flushing by river water. During the period of falling water (drainage and isolation phases), nanoplanktonic algae with similar characteristics to those of the filling phase dominated in late winter-spring. Later in the isolation phase, these were succeeded by K-selected species (netplanktonic algae, mainly motile spherical dinoflagellates and filamentous bluegreens with low SA/V ratios). Simultaneously, primary production per unit biomass decreased and total biomass and specific diversity increased. Seasonal changes of phytoplankton in floodplain lakes can be interpreted as the interaction between true successional development (as observed in the drainage and isolation phases) and intermediate disturbance. Using Reynolds' terminology, short-term disturbance (slight inflow of nutrient-rich river water) caused reversion to an earlier stage in the former succession, and long-term disturbance (lake inundation) truncated the successional progression and a new (or shifted) succession was initiated.     

Retention of nitrogen in small streams artificially polluted with nitrate

A simple method was developed to test hypotheses on nitrogen retention in first-order streams in an agricultural region near Oslo, SE Norway. A gravity-operated system added a nitrate solution to the streams continuously at a constant rate. Water samples were collected at fixed intervals downstream to follow the rate of decline in streamwater nitrate. Repeated sampling allowed calculation of regression lines from experiments with different levels of additions of nitrate.The experiments showed that removal of nitrate generally increased with higher initial nitrate concentration, regardless of temperature (range 8–16 °C). Higher nitrate removal rates were found in a stream polluted by easily degradable organic matter than in a similar stream fed by groundwater.Experiments conducted in indoor channels lined with a layer of stream sediment gave reproducible, exponential rates of nitrate decrease in the recirculated water.The results are discussed in the framework of first-order streams as protective ecotones between agricultural areas and higher-order parts of the watersheds.     

Seed development in Pisum

The Ih and lh _i alleles have been shown previously to reduce the level of endogenous gibberellin A₁ (GA₁) in shoots of pea (Pisum sativum L.), resulting in a dwarf phenotype compared with the wild type, cv. Torsdag (Lh). In addition, plants homozygous for the lh ⁱ allele have reduced seed yield compared with Lh (tall, wild type) and lh (dwarf) plants. In this paper we show that the lh ⁱ mutation is expressed in developing seeds and pods. Comparison of GA levels in young shoots and developing seeds of genotypes lh and lh ⁱ demonstrates that the relative severity of the two mutations varies in different tissues. Homozygous h ⁱ seeds have reduced GA levels, weigh less, and are less likely to develop to maturity when compared with Lh seeds. However, fertilization of lh ⁱ plants with Lh pollen increases seed GA levels, seed weight and seed survival, indicating that an increase in seed GA levels due to the presence of the Lh allele can restore normal seed growth. Pods developing on self-pollinated lh ⁱ plants are shorter than pods on Lh (wild type) plants, although this may be an indirect effect of the increased seed abortion of lh _i plants. Based on these results we suggest that endogenous GAs play an important role in the development of seeds of P. sativum L.Abbreviations GA_(n) gibberellin A_n We wish to thank Katherine McPherson, Peter Newman, Leigh Johnson and Peter Bobbi for technical assistance, Professor L. Mander (ANU, Canberra) and Professor B.O. Phinney (UCLA, USA) for labelled GA standards, and the Australian Research Council for financial support.     

K+ channels of stomatal guard cells: Abscisic-acid-evoked control of the outward rectifier mediated by cytoplasmic pH

The activation by abscisic acid (ABA) of current through outward-rectifying K⁺ channels and its dependence on cytoplasmic pH (pH_i) was examined in stomatal guard cells of Vicia faba L. Intact guard cells were impaled with multibarrelled and H⁺-selective microelectrodes to record membrane potentials and pH_i during exposures to ABA and the weak acid butyrate. Potassium channel currents were monitored under voltage clamp and, in some experiments, guard cells were loaded with pH buffers by iontophoresis to suppress changes in pH_i. Following impalements, stable pH_i values ranged between 7.53 and 7.81 (7.67±0.04, n = 17). On adding 20 M ABA, pH_i rose over periods of 5–8 min to values 0.27±0.03 pH units above the pH_i before ABA addition, and declined slowly thereafter. Concurrent voltage-clamp measurements showed a parallel rise in the outward-rectifying K⁺ channel current (I_{K, out}) and, once evoked, both pH_i and I_{K, out} responses were unaffected by ABA washout. Acid loads, imposed with external butyrate, abolished the ABA-evoked rise in I_{K, out}. Butyrate concentrations of 10 and 30 mM (pH₀ 6.1) caused pH_i to fall to values near 7.0 and below, both before and after adding ABA, consistent with a cytoplasmic buffer capacity of 128±12 mM per pH unit (n = 10) near neutrality. Butyrate washout was characterised by an appreciable alkaline overshoot in pH_i and concomitant swell in the steady-state conductance of I_{K, out}. The rise in pH_i and i_{K, out} in ABA were also virtually eliminated when guard cells were first loaded with pH buffers to raise the cytoplasmic buffer capacity four- to sixfold; however, buffer loading was without appreciable effect on the ABA-evoked inactivation of a second, inward-rectifying class of K⁺ channels (I_{K, in}). The pH_i dependence of I_{K, out} was consistent with a cooperative binding of at least 2H⁺ (apparent pK_a = 8.3) to achieve a voltage-independent block of the channel. These results establish a causal link previously implicated between cytoplasmic alkalinisation and the activation of I_{K, out} in ABA and, thus, affirm a role for H⁺ in signalling and transport control in plants distinct from its function as a substrate in H⁺-coupled transport. Additional evidence implicates a coordinate control of I_{K, in} by cytoplasmic-free [Ca²⁺] and pH_i.Abbreviations ABA abscisic acid - [Ca²⁺]_i cytoplasmic free [Ca²⁺]_i - E_K K⁺ equilibrium potential - I_{K, out}, I_{K, in} outward-, inward-rectifying K⁺ channel (current) - I-V current-voltage (relation) - Mes 2-(N-morpholino)ethanesulfonic acid - pH_i cytoplasmic pH - Tes 2-{[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]-amino}ethanesulfonic acid - V_m membrane potential We are grateful to G. Thiel (Pflanzenphysiologisches Institut, Universität Göttingen, Germany) for helpful discussions. This work was possible with equipment grants-in-aid from the Gatsby Charitable Foundation, the Royal Society and the University of London Central Research Fund. F.A. holds a Sainsbury Studentship.     

Severe hypoxia decreases oxygen uptake relative to intensity during submaximal graded exercise

The effect of severe acute hypoxia (fractional concentration of inspired oxygen equalled 0.104) was studied in nine male subjects performing an incremental exercise test. For power outputs over 125 W, all the subjects in a state of hypoxia showed a decrease in oxygen consumption ( O₂) relative to exercise intensity compared with normoxia (P < 0.05). This would suggest an increased anaerobic metabolism as an energy source during hypoxic exercise. During submaximal exercise, for a given O₂, higher blood lactate concentrations were found in hypoxia than in normoxia (P < 0.05). In consequence, the onset of blood lactate accumulation (OBLA) was shifted to a lower O₂ ( O₂ 1.77 l·min^–1 in hypoxia vs 3.10 l·min^–1 in normoxia). Lactate concentration increases relative to minute ventilation ( _E) responses were significantly higher during hypoxia than in normoxia (P < 0.05). At OBLA, _E during hypoxia was 25% lower than in the normoxic test. This study would suggest that in hypoxia subjects are able to use an increased anaerobic metabolism to maintain exercise performance.     

Sequence and structural homology among membrane-associated domains of CFTR and certain transporter proteins

A sequence comparison of the two membrane-associated (MA) domains of the cystic fibrosis transmembrane conductance regulator (CFTR), multidrug resistance transporter (MDR), and -factor pheromone export system (STE6) proteins, each of which are believed to contain a total of 12 transmembrane (TM) segments, reveals significant amino acid homology and length conservation in the loop regions that connect individual TM sequences. Similar structural homology is observed between these proteins, hemolysin B (HLYB) and the major histocompatibility-linked peptide transporter, HAM1, the latter two which contain a single MA domain composed of six TM segments. In addition, there are specific sequences that are conserved within the TM segments of the five different membrane proteins. This observation suggests that the folding topologies of the MA domains of MDR, STE6, and CFTR in the plasma membrane are likely to be very similar. The sequence analysis also reveals that there are three characteristic motifs (a pair of aromatic residues, LTLXXXXXXP and GXXL) that are conserved in MDR, STE6, HLYB, HAM1, but not in CFTR. We propose that although CFTR may be evolutionarily related to these other membrane proteins, it belongs to a separate subclass.     

The ecosystem wilting point defines drought response and recovery of a Quercus-Carya forest

Soil and atmospheric droughts increasingly threaten plant survival and productivity around the world. Yet, conceptual gaps constrain our ability to predict ecosystem-scale drought impacts under climate change. Here, we introduce the ecosystem wilting point (Ψ_EWP), a property that integrates the drought response of an ecosystem's plant community across the soil–plant–atmosphere continuum. Specifically, Ψ_EWP defines a threshold below which the capacity of the root system to extract soil water and the ability of the leaves to maintain stomatal function are strongly diminished. We combined ecosystem flux and leaf water potential measurements to derive the Ψ_EWP of a Quercus-Carya forest from an “ecosystem pressure–volume (PV) curve,” which is analogous to the tissue-level technique. When community predawn leaf water potential (Ψ_pd) was above Ψ_EWP (=−2.0 MPa), the forest was highly responsive to environmental dynamics. When Ψ_pd fell below Ψ_EWP, the forest became insensitive to environmental variation and was a net source of carbon dioxide for nearly 2 months. Thus, Ψ_EWP is a threshold defining marked shifts in ecosystem functional state. Though there was rainfall-induced recovery of ecosystem gas exchange following soaking rains, a legacy of structural and physiological damage inhibited canopy photosynthetic capacity. Although over 16 growing seasons, only 10% of Ψ_pd observations fell below Ψ_EWP, the forest is commonly only 2–4 weeks of intense drought away from reaching Ψ_EWP, and thus highly reliant on frequent rainfall to replenish the soil water supply. We propose, based on a bottom-up analysis of root density profiles and soil moisture characteristic curves, that soil water acquisition capacity is the major determinant of Ψ_EWP, and species in an ecosystem require compatible leaf-level traits such as turgor loss point so that leaf wilting is coordinated with the inability to extract further water from the soil.