Estimation of intracellular pH in muscle of fishes from different thermal environments

A technique based on homogenisation of rapidly frozen tissue was used to investigate the regulation of intracellular pH (pH_i) in freshwater and marine fish from diverse environmental temperatures. The following species were held at ambient temperatures of ca. 1°C (Notothenia coriiceps; Antarctica), 5°C (Pleuronectes platessa, Myoxocephalus scorpius; North Sea), and 26°C (Oreochromis niloticus; African lakes). The effects of seasonal acclimatisation to 4, 11 and 18°C were also examined in rainbow trout in the winter, autumn and summer, respectively. Extracellular (whole blood) pH (pH_e) did not follow the constant relative alkalinity relationship, where pH⁺=pOH⁻ for any particular temperature, over a range of 1–26°C (overall δpH_e/δT=0.009±0.002 U °C⁻¹; P<0.001), apparently being regulated by ionic fluxes and ventilation. Intracellular pH (pH_i) was also regulated independently of pN(=0.5 pK water) in all species of fish examined. The inverse relationship between pH_i and environmental temperature gave an overall δpH_i/δT of −0.010±0.001 U °C⁻¹ (for both white and red muscle) and −0.004±0.003 U °C⁻¹ (cardiac muscle). However, between 1 and 11°C δpH_i/δT was much higher (P<0.001), −0.022±0.003 U °C⁻¹ (white muscle) and −0.022±0.004 U °C⁻¹ (red muscle). The possible adaptive roles for these different acid–base responses to environmental temperature variation among tissues and species, and the potential difficulties of estimating pH_i, are discussed.     

An alkali tolerant α-l-rhamnosidase from Fusarium moniliforme MTCC-2088 used in de-rhamnosylation of natural glycosides

Analkali tolerant α-l-rhamnosidase has been purified to homogeneity from the culture filtrate of a new fungal strain, Fusarium moniliforme MTCC-2088, using concentration by ultrafiltration and cation exchange chromatography on CM cellulose column. The molecular mass of the purified enzyme has been found to be 36.0 kDa using SDS-PAGE analysis. The K_m value using p-nitrophenyl-α-l-rhamnopyranoside as the variable substrate in 0.2 M sodium phosphate buffer pH10.5 at50 °C was 0.50 mM. The catalytic rate constant was15.6 s⁻¹giving the values of k_cat/K_m is 3.12 × 10⁴M⁻¹ s⁻¹. The pH and temperature optima of the enzyme were 10.5 and 50 °C, respectively. The purified enzyme had better stability at 10 °C in basic pH medium. The enzyme derhamnosylated natural glycosides like naringin to prunin, rutin to isoquercitrin and hesperidin to hesperetin glucoside. The purified α-l-rhamnosidase has potential for enhancement of wine aroma.     

The membraneless bioelectrochemical reactor stimulates hydrogen fermentation by inhibiting methanogenic archaea

The membraneless bioelectrochemical reactor (Ml-BER) is useful for dark hydrogen fermentation. The effect of the electrochemical reaction on microorganisms in the Ml-BER was investigated using glucose as the substrate and compared with organisms in a membraneless non-bioelectrochemical reactor (Ml-NBER) and bioelectrochemical reactor (BER) with a proton exchange membrane. The potentials on the working electrode of the Ml-BER and BER with membrane were regulated to ?0.9 V (versus Ag/AgCl) to avoid water electrolysis with a carbon electrode. The Ml-BER showed suppressed methane production (19.8?±?9.1 mg-C·L^?1·day^?1) and increased hydrogen production (12.6?±?3.1 mg-H·L^?1·day^?1) at pH_out 6.2?±?0.1, and the major intermediate was butyrate (24.9?±?2.4 mM), suggesting efficient hydrogen fermentation. In contrast, the Ml-NBER showed high methane production (239.3?±?17.9 mg-C·L^?1·day^?1) and low hydrogen production (0.2?±?0.0 mg-H·L^?1·day^?1) at pH_out 6.3?±?0.1. In the cathodic chamber of the BER with membrane, methane production was high (276.3?±?20.4 mg-C·L^?1·day^?1) (pH_out, 7.2?±?0.1). In the anodic chamber of the BER with membrane (anode-BER), gas production was low because of high lactate production (43.6?±?1.7 mM) at pH_out 5.0?±?0.1. Methanogenic archaea were not detected in the Ml-BER and anode-BER. However, Methanosarcina sp. and Methanobacterium sp. were found in Ml-NBER. Prokaryotic copy numbers in the Ml-BER and Ml-NBER were similar, as were the bacterial community structures. Thus, the electrochemical reaction in the Ml-BER affected hydrogenotrophic and acetoclastic methanogens, but not the bacterial community.     

Bioremediation potential of Chondrus crispus (Basin Head) and Palmaria palmata: effect of temperature and high nitrate on nutrient removal

To evaluate the nutrient removal capabilities of two red macroalgae, apical blades were cultured in the lab for 4?weeks at either 6, 10, or 17°C and nitrate at either 30 or 300?μM, typical of the seasonal range of conditions at a land-based Atlantic halibut farm. Stocking density was 2.0?g?L^?1, irradiance 125?μmol?photons?m^?2?s^?1, photoperiod 16:8 (L:D), and nitrogen to phosphorus ratio 10:1. For both species, the highest growth rate was at 300?μM NO ₃ ^? with Palmaria palmata growing fastest at 6°C, 5.8%?day^?1, and Chondrus crispus growing best at 17°C, 5.5%?day^?1. Nitrogen and carbon removal by P. palmata was inversely related to temperature, the highest rate at 6°C and 300?μM NO ₃ ^? of 0.47?mg N and 6.3?mg C per gram dry weight per day. In contrast, C. crispus removal of N was independent of temperature, with mean removal of 0.49?mgN?gDW^?1?day^?1 at 300?μM NO ₃ ^? . The highest carbon removal by C. crispus was 4.4?mgC?gDW^?1?day^?1 at 10°C and 300?μM nitrate, though not significantly different from either 6 or 17°C and 300?μM nitrate. Tissue carbon:nitrogen ratios were >20 in both species at 30?μM nitrate, and all temperatures indicating nitrogen limitation in these treatments. Phycoerythrin content of P. palmata was independent of temperature, with means of 23.6?mg?gFW^?1 at 300?μM nitrate. In C. crispus, phycoerythrin was different only between 6°C and 17°C at 300?μM nitrate, with the highest phycoerythrin content of 12.6?mg?gFW^?1 at 17°C. Morphological changes were observed in P. palmata at high NO ₃ ^? concentration as curling of the fronds, whilst C. crispus exhibited the formation of bladelets as an effect of high temperature.     

A study on intra-particle diffusion effects in enzymatic reactions: glucose-fructose isomerization

In this work different aspects of the glucose-fructose enzymatic isomerization, using immobilized glucose isomerase, are studied and quantified. Reaction temperatures range from 40?°C to 60?°C. Intra-particle effective diffusivities (D _e), determined after uptake experiments, are between 1.20?×?10^?6?cm²/s, at 40?°C, and 2.52?×?10^?6?cm²/s, at 60?°C. The estimated energy of activation for diffusion (E _aD) is 7.71?kcal/mol. No significant adsorption of the sugars on the support gel matrix is observed. Crushed particles (φ = 150–350?μ) are used during kinetic experiments. For this range of particle diameters, inherent kinetics is approached. A reversible Michaelis–Menten rate equation is fitted to the data, providing the following parameters at pH = 7.0: k ₀ = 2.15?×?10^?6?g/IU/s; E _a/R = 8998?K. Glucose (K _G) and fructose (K _F) affinity constants are essentially the same, ranging from 0.190?M, at 40?°C to 0.305?M, at 60?°C. The thermodynamic equilibrium constant is determined for the three temperatures, and the heat of reaction, estimated from a Van't Hoff plot, is ΔH = 1682?cal/mol. Independent experiments, where the reaction occurs in the presence of significant intra-particle mass transfer resistance, are used as validation tests.     

Interspecific variation in thermoregulation among three sympatric bats inhabiting a hot, semi-arid environment

Bats in hot roosts experience some of the most thermally challenging environments of any endotherms, but little is known about how heat tolerance and evaporative cooling capacity vary among species. We investigated thermoregulation in three sympatric species (Nycteris thebaica, Taphozous mauritianus and Sauromys petrophilus) in a hot, semi-arid environment by measuring body temperature (T _b), metabolic rate and evaporative water loss (EWL) at air temperatures (T _a) of 10?C42?°C. S. petrophilus was highly heterothermic with no clear thermoneutral zone, and exhibited rapid increases in EWL at high T _a to a maximum of 23.7?±?7.4?mg?g^?1?h^?1 at T _a????42?°C, with a concomitant maximum T _b of 43.7?±?1.0?°C. T. mauritianus remained largely normothermic at T _as below thermoneutrality and increased EWL to 14.7?±?1.3?mg?g^?1?h^?1 at T _a????42?°C, with a maximum T _b of 42.9?±?1.6?°C. In N. thebaica, EWL began increasing at lower T _a than in either of the other species and reached a maximum of 18.6?±?2.1?mg?g^?1?h^?1 at T _a?=?39.4?°C, with comparatively high maximum T _b values of 45.0?±?0.9?°C. Under the conditions of our study, N. thebaica was considerably less heat tolerant than the other two species. Among seven species of bats for which data on T _b as well as roost temperatures in comparison to outside T _a are available, we found limited evidence for a correlation between overall heat tolerance and the extent to which roosts are buffered from high T _a.     

The dehalogenation of the 5-halo-5,6-dihydrouracils: Uracil formation from 5-iodo- and 5-bromo-5,6-dihydrouracil

The elimination of halide ion from either 5-bromo- or 5-iodo-5,6-dihydrouracil to yield uracil is a slow reaction which, in the case of 5-iodo-5,6-dihydrouracil, is 400 times slower than the enzymatic release of ¹²⁵I^? from 5-[¹²⁵I]iodouracil. The elimination of HBr from 5-bromo-5,6-dihydrouracil is subject to general base catalysis by tris(hydroxymethyl)aminomethane (k₂^{Tris base} = 11 × 10^?4M^?1 min^?1, 37°C, ionic strength 1.0 M). At pH values near and above physiological, both the bromo- and iododihydropyrimidines are subject to hydrolysis of the dihydropyrimidine ring, a reaction which parallels halide elimination to yield uracil. The resulting 2-halo-3-ureidopropionate then cyclizes via intramolecular attack of the ureido oxygen atom to yield halide ion and 2-amino-2-oxazoline-5-carboxylic acid as final products. In dilute hydroxide ion, the kinetics of 5-bromo-5,6-dihydrouracil hydrolysis (25°C, ionic strength 1.0 M) show a change in rate-determining step as a function of increasing hydroxide ion concentration, a result which, as in the case of 5,6-dihydrouracil, can be explained in terms of the formation of a tetrahedral addition intermediate. The data are discussed relative to enzymatically catalyzed halopyrimidine dehalogenation.     

Respiratory properties of the blood of the burrowing red band fish Cepola rubescens L.

The respiratory properties of the whole blood of the burrowing red band fish Cepola rubescens L. were investigated. Oxygen dissociation curves constructed at 15°C were found to be close to hyperbolic in shape with a mean value for the cooperativity coefficient at half-saturation (n₅₀) of 1.56. Half-saturation oxygen tension (P₅₀) for pH = 7.56 (mean in vivo pH of venous blood) was 27 Torr. The blood showed a marked Bohr effect ($\text{Δ log}\text{P}{}_{50}\text{ΔpH}\text{= ?1.19}$) and also a Root effect which at the in vivo pH reduced oxygen carrying capacity by 20%. The Pv_CO2 was 3.2 Torr and the buffering power of the blood was low, the buffer value of true plasma averaging 5.43 mmol · 1^?1 · pH^?1. It is suggested that the large Bohr effect coupled with the low buffer value confers on the haemoglobin a flexibility, in terms of oxygen affinity, to withstand changes which occur in environmental oxygen tensions.     

Electron and proton transfers from P-430 to ferredoxin-NADP-reductase in Chlorella cells

After blocking Photosystem II on whole Chlorella cells, we measured the absorption changes between 0°C and ?10°C.The absorption changes measured 2 μs after the beginning of a Xenon Flash are the sum of changes due to P⁺-700 and changes due to P^?-430 (after the subtraction of the carotenoid triplet change and of the electrochromic effect).The reduction of P^?-430 is not resolved by our technique. Its reoxidation presents a half-time around 1 μs at 0°C and around 2 μs at ?10°C.The reduction and protonation of ferredoxin-NADP-reductase to its neutral semi-quinoid form FNRH° present a half-time of about 3 μs at 0°C and 6 μs at ?10°C.The presence of only one photoreducible ferredoxin-NADP-reductase per Photosystem I center is confirmed. The acceptor preceding ferredoxin-NADP-reductase is not ferredoxin, but is an acceptor X' the differential extinction coefficients of which are weak or null from 420 nm to 480 nm.Tentative explanations which would reconcile these results with what was already known about ferredoxin are proposed.     

Accumulation and turnover of abscisic acid in osmotically stressed cotton leaf tissue in relation to temperature

Leaf discs of cotton (Gossypium hirsutum L. cv. Deltapine 70) were osmotically stressed by floating them on solutions of polyethylene glycol 8000. The tissue produced copious amounts of abscisic acid (ABA) when stressed. Accumulation of ABA depended strongly upon temperature during the incubation, displaying a maximum at 20°C. At 35°C, the amount of ABA accumulated after 24 h was 45–80% less than at 20°C. Temperature did not affect leakage of ABA into the medium. Turnover rate of [¹⁴C]ABA was more than 3 times greater at 35°C than at 20°C. This rapd turnover at 35°C could account for the decreased ABA accumulation. Three ¹⁴C-containing metabolites of ABA were extracted from the tissue. At 20°C, two of these accumulated and retained substantial ¹⁴C over 16 h. At 35°C, though, the ¹⁴C in one of these compounds was almost completely lost during the last 8 h of the incubation. Although the metabolites are not identified, the results show some specific effects of temperature on ABA metabolism. The strong effect of temperature on ABA accumulation may contribute to patterns of ABA-dependent processes (such as stomatal closure) during water stress.     

Body temperature and respiratory dynamics in un-shaded beef cattle

In this study body temperature (BT, °C) and panting score (PS, 0–4.5; where 0?=?no panting/no stress and 4.5?=?catastrophic stress) data were obtained from 30 Angus steers housed outside over 120 days Steers were implanted with a BT transmitter on day ?31, BT was recorded at 30-min intervals to a data logger and downloaded each day to a database. The cattle were housed in ten outdoor un-shaded pens with an earthen floor, eight of which had a pen floor area of 144 m² (three transmitter steers plus five non-transmitter steers; 18 m²/steer) and two had an area of 168 m² (three transmitter steers and six non-transmitter steers; 18.7 m²/steer). Only data from the transmitter steers were used in this study. The PS of the steers was obtained daily (± 15 min) at 0600 hours (AM), 1200 hours (MD) and 1600 hours (PM). At the same times climate variables (ambient temperature, black globe temperature, solar radiation, relative humidity, wind speed and rainfall) were obtained from an on-site weather station. PS observations were made from outside the pens so as not to influence cattle responses. The two closest BT values to the time when PS was obtained were downloaded retrospectively from a logger and averaged. A total of 8,352 observations were used to generate second order polynomial response curves: (AM) y?=?39.08?+?0.009?x +?0.137x ² (R ²?=?0.94; P? y?=?39.09?+?0.914x ? 0.080x ² (R ²?=?0.89; P? y?=?39.52?+?0.790x ? 0.068x ² (R ²?=?0.83; P?x?PS. These data suggest that PS is a good indicator of body temperature. The BT at MD corresponded to slightly lower PS compared with PM, e.g., for PS 1; BT at MD?=?39.1?±?0.05 °C whereas BT at PM?=?39.5?±?0.05 °C. However during AM, BT was lower (P?相似文献   

The kinetics of light-induced changes of C-550, cytochrome b559 and fluorescence yield in chloroplasts at low temperature

The kinetics of the photoreduction of C-550, the photooxidation of cytochrome b₅₅₉ and the fluorescence yield changes during irradiation of chloroplasts at ?196 °C were measured and compared. The photoreduction of C-550 proceeded more rapidly than the photooxidation of cytochrome b₅₅₉ and the fluorescence yield increase followed the cytochrome b₅₅₉ oxidation. These results suggest that fluorescence yield under these conditions indicates the dark reduction of the primary electron donor to Photosystem II, P680⁺, by cytochrome b₅₅₉ rather than the photoreduction of the primary electron acceptor.The photoreduction of C-550 showed little if any temperature dependence over the range of ?196 to ?100 °C. The amount of cytochrome b₅₅₉ photooxidized was sensitive to temperature decreasing from the maximal change at temperatures between ?196 to ?160 °C to no change at ?100 °C. To the extent that the reaction occurred at temperatures between ?160 and ?100 °C the rate was largely independent of temperature. The rate of the fluorescence increase was dependent on temperature over this range being 3–4 times more rapid at ?100 than at ?160 °C. At ?100 °C the light-induced fluorescence increase and the photoreduction of C-550 show similar kinetics. The temperature dependence of the fluorescence induction curve is attributed to the temperature dependence of the dark reduction of P680⁺.The intensity dependence of the photoreduction of C-550 and of the photooxidation of cytochrome b₅₅₉ are linear at low intensities (below 200 μW/cm²) but fall off at higher intensities. The failure of reciprocity in the photoreduction of C-550 at the higher intensities is not explained by the simple model proposed for the Photosystem II reaction centers.     

Acute Toxicity of Arsenic under Different Temperatures and Salinity Conditions on the White Shrimp Litopenaeus vannamei

The aim of this study was to determine acute toxicity in the post larvae of the white shrimp Litopenaeus vannamei after 96 h of exposure to dissolved arsenic under three different temperatures and salinity conditions. Recent reports have shown an increase in the presence of this metalloid in coastal waters, estuaries, and lagoons along the Mexican coast. The white shrimp stands out for its adaptability to temperature and salinity changes and for being the main product for many commercial fisheries; it has the highest volume of oceanic capture and production in Mexican shrimp farms. Lethal concentrations (LC₅₀–96 h) were obtained at nine different combinations (3?×?3 combinations in total) of temperature (20, 25, and 30 °C) and salinity (17, 25, and 33) showing mean LC₅₀–96 h values (±standard error) of 9.13?±?0.76, 9.17?±?0.56, and 6.23?±?0.57 mgAs?L^?1(at 20 °C and 17, 25, and 33 salinity); 12.29?±?2.09, 8.70?±?0.82, and 8.03?±?0.59 mgAs?L^?1 (at 25 °C and 17, 25, and 33 salinity); and 7.84?±?1.30, 8.49?±?1.40, and 7.54?±?0.51 mgAs?L^?1 (at 30 °C and 17, 25, and 33 salinity), respectively. No significant differences were observed for the optimal temperature and isosmotic point of maintenance (25 °C–S 25) for the species, with respect to the other experimental conditions tested, except for at 20 °C–S 33, which was the most toxic. Toxicity under 20 °C–S 33 conditions was also higher than 25 °C–S 17 and 20 °C (S 17 or 25). The least toxic condition was 25 °C–S 17. All this suggests that the toxic effect of arsenic is not affected by temperature changes; it depends on the osmoregulatory pattern developed by the shrimp, either hyperosmotic at low salinity or hiposmotic at high salinity, as observed at least on the extreme salinity conditions here tested (17 and 33). However, further studies testing salinities near the isosmotic point (between 20 and 30 salinities) are needed to clarify these mechanisms.     

Nitrogen release from decomposing seaweeds: species and temperature effects

This study determined the rate at which nitrogen accumulated in seaweeds is released during decomposition and the effect of temperature on their rates of decomposition and nitrogen release. Gracilaria verrucosa and Ulva lactuca decomposed rapidly in outdoor mesocosms. Ulva, but not Gracilaria, became nitrogen-enriched during decomposition. Maximal weekly rates of nitrogen release were 5.91 ± 2.23 and 6.37 ± 2.59 g N m^?2 d^?1, respectively for Gracilaria and Ulva. Temperature had a significant effect on the decomposition rate of Gracilaria in a laboratory experiment: decomposition was greater at 30 °C than at 25 °C. No net decomposition was observed at 16 °C. Gracilaria became nitrogen enriched at 30 °C, but not at 16° or 25°. The release of stored nutrients from decaying seaweeds should be included in nutrient budgets and models when seaweed standing stocks are significant. Seaweed source-sink relationships are important ecologically and can be applied to attempts at using seaweeds as environmental monitors of anthropogenic eutrophication and to efforts of cultivating seaweeds for the improvement of water quality.     

Univalent ions in phospholipid model membranes: Thermodynamic and hydration aspects

By means of differential scanning calorimetry, effects of systematic series of Group I and VII ions on the phase state of model multibilayer dimyristoylphosphatidylcholine (di(14:0)PC) membranes have been studied at a lipid/ion molar ratio of 3/1. The sign-changing correlations between the ionic radii of cations and temperature shifts of di(14:0)PC phase transition were obtained. For cosmotropic Li⁺ and Na⁺, the observed shifts were positive (LiCl: ΔT _m = 0.6°C; ΔT _p = 1.9°C), whereas chaotropic K⁺ and Rb⁺ presence resulted in negative shifts (RbCl: ΔT _m = ?0.3°C; ΔT _p = ?2.5°C). The anions (Cl^?, Br^?, I^?) showed a similar effect increasing with the ion chaotropicity. An essentially weaker effect of Cs⁺ as compared to other alkali metal ions (CsCl: ΔT _m ≈ 0°C; ΔT _p = ?0.1°C) can be one of the reasons of its accumulation in living organisms. Generalization of all available data allowed us to specify some important factors of lipid-ion interactions that should be taken into account in further investigations in this field.     

High-level expression of a xylanase gene from the thermophilic fungus Paecilomyces thermophila in Pichia pastoris

A xylanase gene from Paecilomyces thermophila was functionally expressed in Pichia pastoris. The recombinant xylanase (xynA) was predominantly extracellular; in a 5?l fermentor culture, the total extracellular protein was 8.1?g?l^?1 with an activity of 52,940?U?ml^?1. The enzyme was purified to homogeneity with a recovery of 48?%. The recombinant xynA was optimally active at 75?°C, as measured over 10?min, and at pH 7. The enzyme was stable up to 80?°C for 30?min. It hydrolyzed birchwood xylan, beechwood xylan and xylooligosaccharides to produce xylobiose and xylotriose as the main products.     

The enthalpy of oxidation of flavin mononucleotide

The oxidation enthalpy of reduced flavin mononucleotide at pH 7.0 in 0.2 m phosphate buffer has been studied by determining the heat associated with the reaction: FMNH₂ + 2 Fe(CN)^?3₆ ? FMN + 2 Fe(CN)^?4₆ + 2 H⁺. (a) (The quinone, semiquinone, and hydroquinone forms of FMN are represented as FMN, FMNH, and FMNH₂, respectively.) Calorimetric experiments were performed in a flow microcalorimeter which was modified to prevent sample contamination by oxygen. The enthalpy observed for reaction (a), after correction for dilution and buffer effects, was ?39.2 ± 0.4 kcal (mole FMNH₂)^?1 at 25 °C. The potential difference, ΔE′, developed by reaction (a) was determined potentiometrically and corresponded to a free energy change, ΔG′, of ?30.3 kcal (mole FMNH₂)^?1. The resulting entropy change, ΔS′, was thus calculated to be ?29.8 e.u. Reaction (a) was also studied at temperatures of 7 °C and 35.5 °C. ΔC_p′ for the reaction was calculated as ?155 ± 18 cal deg^?1 (mole FMNH₂)^?1 at 20 °C. ΔH′ for the reaction (b), FMNH₂ ? FMN + H₂, (b) was calculated as +14.2 ± 0.7 kcal mole^?1 at 25 °C, relative to the enthalpy of the hydrogen electrode being identically equal to zero at all values of pH and temperature. The free energy at pH 7.0 for reaction (b), calculated from the potential was found to be ?9.7 kcal mole^?1, which resulted in an entropy for reaction (b) of 80.2 e.u. A thermal titration of reaction (a) was used to calculate the thermodynamic parameters for the formation of semiquinone dimer according to the reaction FMNH₂ + FMN ? (·FMNH)₂. (c) The free energy, enthalpy, and entropy changes for reaction (c) were estimated to be ?6.1 kcal mole^?1, ?7 kcal mole^?1, and ?3 e.u., respectively.