Salt effects on the denaturation of DNA. V. Preferential interactions of native and denatured calf thymus DNA in Na2SO4 solutions of varying ionic strength.
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Hydrolysis of steroid glucuronides with beta-glucuronidase preparations from bovine liver, Helix pomatia, and E. coli.
We determined the enzymic activity of beta-glucuronidase preparations from bovine liver, Helix pomatia, and Escherischia coli with steroid glucuronides and nonsteroid glucuronides as substrates. We also studied the effect of Na2SO4 on the enzymic hydrolysis of several substrates with the three preparations of beta-glucuronidase. Na2SO4 increases the rate of hydrolysis of all substrates with beta-glucuronidase from bovine liver. Hydrolysis of a steroid glucoronide with beta-glucuronidase from Helix pomatia and E. coli is inhibited by Na2SO4. None of the three enzyme preparations gives complete hydrolysis of urinary steroid conjugates, because urine contains inhibitors, which can be removed by absorption chromatography of the urine on a column of neutral polystyrene resin Amberlite XAD-2. But when Amberlite XAD-2 is not used, hydrolysis of urinary glucuronides of androsterone, etiocholanolone, pregnanediol, estriol, and 17-hydroxycorticosteroids proves that, given an incubation time of 24 h, the beta-glucuronidase preparation from bovine liver, in the presence of Na2SO4, is suited for determining all of the above steroids except esriol; the preparation from Helix pomatia is good for determining estriol and 17-hydroxycorticosteroids; the preparation from E. coli is good for determining androsterone, 17-hydroxycorticosteroids, and especially estriol, the glucuronide, of which is maximally hydrolyzed in 2 h.
17-Hydroxycorticosteroids↗
Salt effects on the denaturation of DNA. IV. A calorimetric study of the helix-coil conversion of the alternating copolymer poly[d(A-T)].
The enthalpy deltaH, entropy deltaS, and the temperature Tm of the conformational transition of poly[d (A-T)] from the ordered to the randomly oriented state have been determined at pH 6.8 with the help of an adiabatic differential scanning calorimeter in Na2SO4 solutions of increasing ionic strength. Spectrophotometric denaturation experiments supplemented the calorimetric measurements. All thermodynamic parameters were found to vary strongly with salt concentration: both deltaH and Tm increase linearly with the logarithm of the mean molal activity alpha plus or minus of Na2SO4. However, whereas the dependence of Tm on salt activity remains linear over the entire salt concentration range employed deltaH decreases abruptly in the most concentrated Na2SO4 solutions. The entropy of melting changes with salt concentration in a pattern similar to that displayed by deltaH. The data on deltaH as well as data derived from the maximum slopes of the calorimetric heat denaturation curves were used to calculate the cooperative length Lh, the stacking free energy epsilon, and the cooperativity parameter sigma of poly[d(A-T)] as a function of ionic strength. Lh decreases with increasing salt concentration whereas sigma increases. Epsilon assumes more positive values with increasing salt molality. These changes then are in agreement with the generally held belief that an increase in salt concentration leads to an increase in the "loop" content of the copolymer.
Adenine Nucleotides↗
Absorption, serum levels and urinary excretion of inorganic sulfate after oral administration of sodium sulfate in the conscious rat.
The absorption of inorganic sulfate after ingestion was investigated in rats. After oral administration of Na235SO4, 35S radioactivity was measurable in plasma already after 15 min and its plasma concentration reached a peak after about 1.5--2 h. The 35S-radioactivity excreted in urine during 24 h after ingestion of Na235SO4 together with varying amounts of unlabelled Na2SO4 (0.25--5.0 mmol Na2SO4 per rat) indicated an almost complete absorption of inorganic sulfate from the gastrointestinal tract. Determination of the inorganic sulfate concentration in rat serum 2 h after oral administration of 5.0 mmol Na2SO4 revealed a three-fold increase in serum sulfate concentration. The data suggest a rapid and almost complete absorption of inorganic sulfate after oral administration in the rat. Its importance in relation to the sulfate availability for sulfate conjugation of drugs is discussed.
Administration, Oral↗
Active hydrogen excretion and sodium absorption through isolated frog skin.
The in vitro skin of Rana esculenta was studied in open-circuit conditions. It was shown that when the external face is bathed in a 2-meq solution of NaCl, sodium is absorbed at a significantly higher rate than chloride. The ionic balance is maintained by excretion of hydrogen. With a mucosal solution of 2 meq Na2SO4 the equation relating sodium absorption to proton excretion is JnH+ = (-25 +/- 7) - (0.73 +/- 0.04) JnNa+. The correlation between the two variables is highly significant. Hydrogen excretion obeys saturation kinetics in relation to the sodium concentration of the mucosal solution. Maximum excretion occurs at a sodium concentration of 4 meq. When the mucosal solution is a 115-meq solution of Na2SO4 the net flux of sodium is 2.3 times higher than that of hydrogen. The balance is maintained by absorption of SO42-. The effects of various substances on the Na+ext/H+int exchange were studied. With a mucosal solution of 2 meq Na2SO4 and short-circuit conditions it was shown that the hydrogen excretion is active and nearly the same as in open circuit, the short-circuit current is equal (to within 8%) to the sum of the sodium and hydrogen net fluxes, and the correlation between the movements of the two ions is low. A model relating the active proton excretion with the sodium transport mechanism is proposed.
Acetazolamide↗
Untargeted metabolomics reveals anion and organ-specific metabolic responses of salinity tolerance in willow.
Willows can alleviate soil salinisation while generating sustainable feedstock for biorefinery, yet the metabolomic adaptations underlying their tolerance remain poorly understood. Salix miyabeana was treated with two environmentally abundant salts, NaCl and Na2SO4, in a 12-week pot trial. Willows tolerated salts across all treatments (up to 9.1 dS m-1 soil ECe), maintaining biomass while selectively partitioning ions, confining Na+ to roots and accumulating Cl- andin the canopy and adapting to osmotic stress via reduced stomatal conductance. Untargeted metabolomics captured >5000 putative compounds, including 278 core willow metabolome compounds constitutively produced across organs. Across all treatments, salinity drove widespread metabolic reprogramming, altering 28% of the overall metabolome, with organ-tailored strategies. Comparing salt forms at equimolar sodium, shared differentially abundant metabolites were limited to 3% of the metabolome, representing the generalised salinity response, predominantly in roots. Anion-specific metabolomic responses were extensive. NaCl reduced carbohydrates and tricarboxylic acid cycle intermediates, suggesting potential carbon and energy resource pressure, and accumulated root structuring compounds, antioxidant flavonoids, and fatty acids. Na2SO4 salinity triggered accumulation of sulphur-containing larger peptides, suggesting excess sulphate incorporation leverages ion toxicity to produce specialised salt-tolerance-associated metabolites. This high-depth picture of the willow metabolome underscores the importance of capturing plant adaptations to salt stress at organ scale and considering ion-specific contributions to soil salinity.
Salix↗
Evidence for electronogenic sodium pumping in the ductal epithelium of rabbit salivary gland and its relationship with (Na+ plus K+)-ATPase.
The temperature dependence of the transepithelial potential difference (PD) of the main duct of the submaxillary gland has been measured during in vitro perfusion studies. The magnitude of the PD depends strongly on the anion composition of the perfusing and bathing fluids. The following combinations of perfusion and bathing fluids respectively were used: (1) Na2SO4/NaCl, (2) Na2SO4, (3) NaCl/-NaCl, (4) NaCl/Na2SO4. The mean transepithelial potential differences at 35 degrees C with these four sets of conditions were respectively: 144, 148, 10 and - 15 mV, serosal side positive with respect to lumen. From the data obtained it was possible to construct Arrhenius plots of temperature dependence of the PD for the four sets of experimental conditions. They all show a breakpoint between 16 and 19 degrees C. The apparent activation energies in the four situations above the breakpoint are 4.2, 1.4, 12.0 and 10.6 kcal/mol, respectively. Below the breakpoint they are 29.9, 37.5, 29.0 and 31.3 kcal/mol, respectively. The rapid change in the PD as a function of temperature (which can also be achieved by the addition of ouabain), the effects of the removal of K+ on the serosal side on the PD, the decrease in the PD after the addition of ouabain or CN-, and the activation energies and breakpoints all lead to the conclusion that a large part of the PD is caused by an electrogenic sodium pump which is very probably the enzyme (Na+ plus K+)-ATPase. When the duct is perfused with Na2SO4 we find, above the breakpoint in the Arrhenius plots, a lower activation energy than is found when perfusing with NaCl.
Adenosine Triphosphatases↗
The steady-state kinetics of yeast phosphoglycerate kinase. Anomalous kinetic plots and the effects of salts on activity.
1. A re-investigation of the kinetics of yeast phosphoglycerate kinase in the direction of 1,3-bisphosphoglycerate formation has been carried out, covering a 1000-fold range in substrate concentrations. A variety of improved spectrophotometric and fluorimetric assay procedures have been used. 2. Kinetic plots proved to be non-linear for each variable substrate. A variety of checks have been carried out to show that this is not due to artifacts in the assay procedures or heterogeneity of the enzyme preparation. 3. The effects of a variety of salts on the activity of the enzyme have been examined. Most salts, especially those with multivalent anions, can cause activation of the enzyme, but inhibit at high concentration. 4. The salt effect is shown to be principally due to anions rather than cations, and not to ionic strength changes. Sulphate, as one of the most effective anions has been used in most comparisons. 5. Salt activation is steepest when the substrate concentrations are low; maximum activation has been about 5-fold with 0.2 mM MgATP and 0.2 mM 3-phosphoglycerate. Inhibition at the higher salt concentrations is strongest at the same substrate concentrations as when activation is steepest, indicating a link between the two effects. 6. The presence of 20 mM or more Na2SO4 converted non-linear kinetic plots to linear ones. A study of the kinetics in the presence of 40 mM Na2SO4 was interpreted in terms of a random sequential binding mechanism, with sulphate acting as a competitive inhibitor. 7. Possible explanations for these anomalous results are discussed in terms of several mechanisms which have been shown to apply in other systems.
Anions↗
Inter-relationship of sodium, chloride, bicarbonate and acetate transport by the colon of the pig.
1. Net transport of Na, Cl, HCO3 and acetate was examined in the temporarily isolated colon of conscious pigs weighing 46 +/- 8 kg. 2. The entire colon absorbs 4.1 ml. H2O, 0.8 m-equiv Na, 1.3 m-equiv acetate and secretes 0.5 m-equiv HCO3/min with a solution comparable to the normal contents. The absorptive capacity of the proximal and distal halves of the colon was comparable per unit dry weight of mucosa when each segment was presented with the same solution. 3. A series of studies using ion replacement solutions showed that net Na absorption and net HCO3 accumulation in the lumen solution were both increased in the presence of acetate. Cl absorption was independent of Na absorption and was accompanied by an equivalent net secretion of HCO3 in the absence of Na. When NaCl in the perfusion solution was replaced with Na2SO4, Na and HCO3 were absorbed at equal rates. 4. Final pCO2 values observed in NaCl and Na2SO4 solutions were greater than those observed in plasma while the pCO2 of the Na acetate solution after perfusion was reduced to values below plasma concentrations. 5. Results are consistent with the hypothesis that hydration of CO2 in the lumen solution or mucosal cell provides a continuous source of H ions for absorption of the more permeable undissociated acid. The evidence also suggests an additional source of H ions may be provided by a Na-H exchange process located in one of the limiting cell membranes.
Acetates↗
Metabolic requirements for anaerobic active Cl and Na transport in the bullfrog cornea.
In the bullfrog cornea, the relationships between the rates of aerobic and anaerobic glycolysis and active Cl and Na transport were studied. In NaCl Ringer (glucose-free), the short-circuit current (SCC) declined much more slowly under aerobic than under anaerobic conditions. The aerobic lactate effluxes in glucose-free and glucose-rich NaCl Ringer were 0.08 and 0.23 micromol/h.cm2, respectively. The transition to anoxia caused these values to increase significantly and was accompanied by depletion of endogenous glycogen in glucose-free Ringer. In Na2SO4 Ringer, amphotericin B (10(-5) M) stimulation of the aerobic SCC was not dependent on the presence of glucose but under anoxia, SCC stimulation required glucose. In Na2SO4 (glucose-rich) Ringer, amphotericin B stimulated the aerobic lactate efflux from 0.26 to 0.36 mumol/h.cm2 and anoxia increased it to 0.55 micromol/h.cm2. In NaCl Ringer, the addition of either 0.5 mM adenosine or 1 mM ATP with 26 mM glucose restored the anaerobic-inhibited SCC and lactate efflux of glucose-depleted corneas. The results show that the reactions of glycolysis are a sufficient energy source for supporting active Na and Cl transport.
Amphotericin B↗
Renal hydrogen ion secretion after release of unilateral ureteral obstruction.
The effect of 24 h of unilateral ureteral obstruction on HCO3 reabsroption and urinary acidification was studied in dogs. The postobstructed kidney (EK) had a significantly lower glomerular filtration rate and renal plasma flow than the contralateral kidney (CK). Urinary pH prior to HCO3 loading was significantly higher in the EK as was maximal HCO3 reabsorption. Saline loading depressed HCO3 reabsorption to the same degree in both kidneys. Urinary PCO2, during HCO3 loading, and during phosphate infusion, was significantly lower in the EK than the CK. Fractional Na excretion was significantly higher in the EK than the CK after deoxycorticosterone acetate administration. Na2SO4 administration enhanced acid excretion only in the CK. K excretion was significantly lower in the EK than the CK both during HCO3 loading and Na2SO4 administration. There was redistribution of cortical blood flow from the outer cortex toward the inner cortex in the EK as compared to the CK. There was no difference in plasma renin activity from both renal veins. These data demonstrate enhanced proximal H+ secretion (which is abolished by volume expansion) and impaired distal H+ secretion by the postobstructed kidney. The distal defect is likely an effect of a generalized disorder of distal transport in that both K secretion and steroid-responsive Na reabsorption were impaired in the postobstructed kidney.
Animals↗
[Solubility of adenosine in concentrated salt solutions].
The solubility of adenosine has been measured in concentrated neutral salt solutions of NaClO4 (salt-destabilizer) and Na2SO4 (salt-stabilizer) as a function of salt concentration and temperature. The thermodynamic functions of adenosine transfer from water to salt solutions have been estimated. The values of delta Ftr, delta Htr, delta Str adenosine transfer in NaClO4 solutions are negative, in Na2SO4--positive. The results are considered in connection with the mechanism of high concentrations of salt action on DNA in the water solutions.
Adenosine↗
Ionic effects on the uptake of chloromercuribenzene-p-sulphonic acid by pancreatic islets.
Effects of inorganic ions on the uptake of chloromercuribenzene-p-sulphonic acid (CMBS) were studied in microdissected pancreatic islets of non-inbred ob/ob-mice. Na2SO4 stimulated the total islet cell uptake of CMBS but decreased the amount of CMBS remaining in islets after brief washing with L-cysteine. CaCl2 stimulated both the total and the cysteine-nondisplaceable uptake; the stimulatory effect of CaCl2 on the cysteine-non-displaceable CMBS uptake was counteracted by Na2SO4. NaCl, KCl or choline chloride had no significant effect on the total islet cell uptake of CMBS, whereas LiCl was stimulatory. It is concluded that beta-cells resemble erythrocytes in having a permeation path for CMBS that is inhibited by SO4(2-). By analogy with existing models of the erythrocyte membrane, it is suggested that the SO4(2-) sensitive path leads to sulphydryl groups controlling monovalent cationic permeability in beta-cells.
4-Chloromercuribenzenesulfonate↗
Production of maximally acid urine by the isolated dog kidney.
The isolated kidney has not been reported to acidify urine maximally. To study this defect, kidneys from dogs fed NH4Cl were perfused with autologous blood. Perfusate pH was 7.20 +/- 0.03 [HCO3] was 14 +/- 1 mEq/L, and urine pH was abnormally high, 6.60 +/- 0.08. When corrected for difference in GFR, UNH4+V was similar to that seen in vivo, but UTAV and UNet H+V were low. FEHCO3- was 2.3% +/- 0.8% and HCO3- excretion persisted to a small degree at perfusate [HCO3-] of 8 to 9 mEq/L. In response to HCO3- infusions, large increases in excretion were not seen until perfusate values were over 24 to 26 mEq/L. HCO3- Tmax was 2.94 +/- 0.07 mEq/dl of glomerular filtrate. The isolated kidney failed to raise U-B PCO2 with HCO3- infusion secondary to low urine [HCO3-] and [Pi]. During perfusion in another group of kidneys from dogs fed NH4Cl and given DOC, perfusate pH and [HCO3-] were similar to those in the first group. Urine pH was also inappropriately high, 7.12 +/- 0.09, and there was no UNet H+V. In response to Na2SO4 infusion, urinary pH fell to 5.00 +/- 0.27. Log10UUAV was correlated to urine pH during the control perfusions in both groups and after Na2SO4 in the NH4Cl + DOC group. Thus production of a low urine pH in the isolated kidney may be mediated by changes in transtubular potential difference resulting from increased distal nephron delivery of Na+ and nonabsorbable anion. The defect in acidification is similar to that observed in incomplete forms of clinical type 1 (distal) renal tubular acidosis.
Ammonium Chloride↗
Sodium chloride stimulated respiration of Anacystis nidulans.
With certain salts a stimulation of respiration of the blue-green alga Anacystis nidulans was found in the dark. The stimulation was observed only at high concentrations (10(-2)M--10(-1)M). NaCl or LiCl are the most effective salts and on addition the increase of the respiration is about 2.5fold. Li is assumed to function as a substitute for Na. Potassium salts, except KCl, are ineffective. The order for the effectiveness is: NaCl greater than NaNO3, Na2SO4 greater than KCl greater than KNO3, K2SO4 (=zero). Accordingly, the cation Na+, and to a less degree the anion Cl- are responsible for the stimulatory effect. K, which is ineffective, is passively accumulated by Anacystis according to the membrane potential. Na is actively extruded. At 0.1 M external NaCl, the passive influx of Na is high, but even then it is balanced by an active efflux. This increases the energy consumption of the cells and leads to a stimulated respiration. With DCCD (N,N'-dicyclohexylcarbodiimide) or NEM (N-ethylmaleimide), the Na efflux is inhibited, simultaneously the stimulation of respiration is abolished and the passive influx of Na becomes detectable. At 0.1 M NaCl, the passive influx of Na measured in presence of DCCD is 5 x 10(-6) moles Na/min and ml packed cells. In absence of DCCD on addition of 0.1 M NaCl the extra oxygen consumption is 2 x 10(-6) moles O2/min and ml cells. This may prove that the stimulation of respiration is mainly caused by the active Na extrusion.
Chlorides↗
Adenosine-5'-phosphosulfate (APS) as sulfate donor for assimilatory sulfate reduction in Rhodospirillum rubrum.
Crude extracts of Rhodospirillum rubrum catalyzed the formation of acid-volatile radioactivity from (35S) sulfate, (35S) adenosine-5'-phosphosulfate, and (35S) 3'-phosphoadenosine-5'-phosphosulfate. An enzyme fraction similar to APS-sulfotransferases from plant sources was purified 228-fold from Rhodospirillum rubrum. It is suggested here that this enzyme is specific for adenosine-5'-phosphosulfate, because the purified enzyme fraction metabolized adenosine-5'-phosphosulfate; 3'-phosphoadenosine-5'-phosphosulfate, however, only at a rate of 1/10 of that with adenosine-5'-phosphosulfate. Further, the reaction with 3'-phosphoadenosine-5'-phosphosulfate was inhibited with 3'-phosphoadenosine-5'-phosphate whereas this nucleotide had no effect on the reaction with adenosine-5'-phosphosulfate. For this activity with adenosine-5'-phosphosulfate the name APS-sulfotransferase is suggested. This APS-sulfotransferase needs thiols for activity; good rates were obtained with either dithioerythritol or reduced glutathione; other thiols like cysteine, 2'-3'-dimercaptopropanol or mercaptoethanol are less effective. The electron donor methylviologen did not catalyze this reaction. The pH-optimum was about 9.0; the apparent Km for adenosine-5'phosphosulfate was determined to be 0.05 mM with this so far purified enzyme fraction. Enzyme activity was increased with K2SO4 and Na2SO4 and was inhibited by 5'-AMP. These properties are similar to assimilatory APS-sulfotransferases from spinach and Chlorella.
Adenine Nucleotides↗
Negative potential level in the outer layer of the toad skin.
The isolated skin of the toad Bufo marinus ictericus when impaled from the outer surface by glass microelectrodes filled with 3 M KCl shows a voltage profile which is a continuous function of the depth of impalement. The superficial intraepithelial potential difference measured with reference to the external solution (PDi) is negative with NaCl-Ringer's solution on both sides of the skin, displaying a minimum of -26.7+/-3.6 mV at 6+/-2 mum. Null value is obtained at 19+/-3 mum, with positive values for deeper impalements. Indications of cell impalements (abrupt voltage and resistance jumps) were frequently observed at sites deeper than 25 mum from the outer surface. Measurements of the electrical resistance between the microelectrode and the external solution, made with single- and double-barreled microelectrodes, showed great discrepancies, which may be attributed to distinct pathways of different resistances in the stratum corneum. PDi measured at a depth of 5 mum was a logarithmic function of Na2SO4 or K2SO4 concentration in the external solution, increasing in negativity with a reduction in concentration. Substitution of Na by K in the external solution had only minor effects on PDi. Acidification of the external solution from pH 9 is accompanied by a reduction in the negative value of PDi. At pH 3 PDi was positive. PDi was interpreted as a diffusion potential at the tip of the microelectrode due to KCl diffusion from the electrode into the matrix of the stratum corneum. Differences in K and Cl mobilities, responsible for the origin of PDi, were attributed to fixed charges in the matrix of the stratum corneum, with density and polarity determined by their degree of proponation, controlled by the hydrogen ion concentration of the external solution. Skin potential, short-circuit current and their relationship to PDI were discussed.
Animals↗
Reconstitution of a proton pump from gastric mucosa.
Purified vesicular fractions from hog gastric mucosa have been incorporated into phosphatidyl serine bilayers. In the presence of MgATP on one side and symmetrical Na2SO4 solutions, a short circuit current (SCC) away from that side is observed increasing exponentially with time, while the corresponding open circuit potential (OCP) is maintained constant for greater than 30 min. In K2SO4 solutions the SCC time course is essentially unchanged, but the OCP falls to almost zero after 15-20 min. In Na -- K gradient there is a similar SCC away from the K-side whose exponential rate is increased by ATP added to both sides. The time course of these events depends only on the time from the formation on the black film. These results are interpreted as showing: (1) There is an ATP-driven proton pump generating a constant potential EH in series with a time dependent conductance gHocekt. (2) There is a shunting K-conductance gKocek't. (3) In the presence of ATP k' greater than k. (4) This time dependence is due to thickness changes in the bilayer. A model relates these results to those obtained with the intact vesicles.
Adenosine Triphosphatases↗