Intermediate pH Values Research Articles

Physico-chemical description of a condensed form of DNA.

AbstractWhen exposed to a low pH, in various ionic strength conditions and in sufficiently dilute solutions, DNA undergoes a transition, revealed by an increase in optical density. A careful analysis shows that, associated with this transition, there is an effective decrease in absorbance, overcompensated by an increase in scattering. The conditions for the new transition can be summarized conveniently by a graph in a pH–Na+ molarity diagram. If the pH of a DNA solution is progressively lowered at constant Na+ concentration, one finds first the melting transition (I), and at lower pH values, the new transition (II). If the same experiment is performed on pre‐denatured DNA, only transition II will be found. If native DNA is brought directly to the low pH conditions, without allowing it to denature irreversibly at intermediate pH values, transition II is reversible (with a small hysteresis effect). DNA, initially native, neutralized after prolonged exposures to the low pH, recovers the buoyant density value of native DNA, along with the absorption and scattering properties of the native state. The experiments are consistent with the interpretation that a new state exists in which DNA, still double stranded, assumes a very compact shape (of the order of 1500 Å in diameter for T2 DNA), with a hyperchromicity value of 10–14% above the native value. Nearly monodisperse suspensions of DNA molecule in this apparent state may be obtained only at very low concentrations (∼0.25 μg/ml). At 1 μg DNA/ml aggregation is noticeable. The possible connection with the condition of intraphage DNA is discussed.

Comparison of Myoglobins from Harbor Seal, Porpoise, and Sperm Whale: VII. MECHANISM AND CATALYSIS FOR ADDITION OF CYANIDE TO MYOGLOBINS

Abstract The pH rate profile for uncatalyzed formation of a complex between cyanide and sperm whale ferrimyoglobin at 25° and total ionic strength 0.11 has been established over the pH range 5 to 11 utilizing an integrated reversible second order rate equation of general applicability. The profile may be interpreted as reflecting three important routes for complex formation: (a) addition of cyanide ion to an unprotonated myoglobin, (b) addition of hydrocyanic acid to the unprotonated myoglobin (or the kinetically indistinguishable addition of cyanide ion to a protonated myoglobin), and (c) addition of hydrocyanic acid to the protonated myoglobin. Complex formation is strongly subject to catalysis by certain buffers: catalysis is most evident at intermediate values of pH and becomes negligible below pH 6 and above pH 10. Effective buffers include phosphate, glycine, and cyanide itself. Other buffers with forms capable of simultaneously accepting 2 protons through hydrogen bonding are also effective. Addition of cyanide to ferrimyoglobins from pilot whale, porpoise, and harbor seal is also subject to buffer catalysis. Guanidination of sperm whale ferrimyoglobin and carboxymethylation in the crystalline state have only small effects on the reactivity of the protein toward cyanide or on the sensitivity of this reaction to buffer catalysis. Carboxymethylation in solution, in contrast, markedly reduces reactivity toward cyanide. The hypothesis is developed that the buffer-binding site may lie between arginine-45 and histidine-64. A reasonable mechanism for the buffer catalysis is proposed.

Electrochemical reduction of parabanic acid: Products and mechanism

The polarographic reduction of parabanic acid, methylparabanic acid and dimethylparabanic acid has been investigated over a wide range of pH. The primary electrochemical reaction involves overall two-electrons and two-protons to give the corresponding 5-hydroxyhydantoin. At low pH and possibly at moderately high pH, the rate controlling step involves two-electrons and two-protons; at intermediate pH values a two-electron, one-proton rate controlling process is operative. 5-Hydroxyhydantoin appears to be slowly hydrolyzed to glyoxylic acid and urea so that prolonged electrolysis (in 1 M HOAc) of parabanic acid gives a mixture of 5-hydroxyhydantoin, urea and the reduction product of glyoxylic acid, namely glycolic acid. In phosphate-containing buffers between approximately pH 6 and 8 all the parabanic acids give a very weak complex with phosphate which is reduced at more negative potentials than the free acids in an essentially pH independent process. On the basis of the phosphate species present in solution and the pH independence of the complex wave, a tentative structure has been proposed for the complex.

Electrochemical reduction of parabanic acid: products and mechanism

The polarographic reduction of parabanic acid, methylparabanic acid and dimethylparabanic acid has been investigated over a wide range of pH. The primary electrochemical reaction involves overall two-electrons and two-protons to give the corresponding 5-hydroxyhydantoin. At low pH and possibly at moderately high pH, the rate controlling step involves two-electrons and two-protons; at intermediate pH values a two-electron, one-proton rate controlling process is operative. 5-Hydroxyhydantoin appears to be slowly hydrolyzed to glyoxylic acid and urea so that prolonged electrolysis (in 1 M HOAc) of parabanic acid gives a mixture of 5-hydroxyhydantoin, urea and the reduction product of glyoxylic acid, namely glycolic acid. In phosphate-containing buffers between approximately pH 6 and 8 all the parabanic acids give a very weak complex with phosphate which is reduced at more negative potentials than the free acids in an essentially pH independent process. On the basis of the phosphate species present in solution and the pH independence of the complex wave, a tentative structure has been proposed for the complex.

Stability of colloidal silica: I. Effect of simple electrolytes

The stability of colloidal silica sols (Ludox HS and Ludox AM) in the presence of various concentrations of NaCl, LiCl, KCl, CsCl, NaBr, NaI, CaCl 2, and Ca(NO 3) 2 has been studied as a function of pH. With increasing pH the stability of the silica sols towards electrolytes markedly decreases except for KCl and CsCl which show a minimum in stability at intermediate pH values. A moving boundary electrophoresis apparatus is described and the mobilities of the Ludox sols as a function of pH and the neutral electrolyte concentration are reported. No correlation was observed between the c.c.c. and the mobilities. Titration curves are given for the exchange of calcium ion for silanolic hydrogen on the sol surface. Ion exchange appears to be the primary reason for the destabilization of silica sols.

Effect of pH on the haemolysis of rabbit erythrocytes by staphylococcus alpha--toxin.

Summary The effect of toxin concentration on the kinetics of haemolysis of rabbit erythrocytes has been studied at pH values differing by about 0·5 pH units over the range 5·5 to 8·5 for both crude and partially purified toxin at concentrations of 2·5, 5·0, 10, 20, 30, 40, 60 MHD per ml and at 80 and 120 MHD per ml for the partially purified toxin. Increased concentration of toxin caused a decrease in the prelytic lag time (·) of the sigmoid haemolysis curves, a decrease in the time to reach 50 per cent. haemolysis (t1/2), and an increase in the maximum rate of haemolysis (Rm). Plateaux in Rm-toxin concentration, ·-toxin concentration and t1/2-toxin concentration curves were found at all pH values: these were displaced to higher concentrations in plots drawn for the partially purified toxin. The effect of pH on haemolysis is very marked, and pH profiles for the maximum rate of haemolysis were constructed: for the crude toxin, the shape of the profile changed as the toxin concentration was increased, while for the partially purified toxin the shape was independent of toxin concentration. The pattern of the profiles indicates that the crude toxin contains a small concentration of impurity that is strongly lytic at pH 5·5 and less markedly so at 8·5; the impurity has no significant effect at intermediate pH values. The reaction is first order with respect to toxin over a very small range of concentrations only, and the maximum rate of haemolysis is independent of toxin concentration at concentrations greater than 120 MHD per ml. The reaction is first order with respect to erythrocytes at all pH values, and over all measurable concentrations.

The co-precipitation of manganese by iron(III) hydroxide

Radioactive manganese and iron were used to study the precipitation of iron(III) and managanese hydroxides at differentpH values from their pure solutions as well as in the presence of each other. It was noticed that there was always some entrainment of mangenese by iron(III) hydroxide even at lowpH values at which the iron itself was incompletely precipitated. The amount of manganese entrained at various concentrations of manganese and iron at the intermediatepH values from 3 to 7 were determined. There is a criticalpH value (i.e. between 4.5 and 4.7) at which there is a change in the course of entrainment from adsorption process to the incorporation phenomena. The effect of aging of the precipitate in its mother liquor to reduce the entrainment was also investigated.

Aluminum + water reaction

During the aluminum + water reaction, aluminum ions and electrons are removed in separate steps at different sites on the surface. Ions are removed nearly uniform over the entire surface, which is covered at all times with a thin amorphous oxide film. The outer surface of this oxide is first hydrolyzed and then dissolves to yield soluble species which either remain in solution or, at intermediate values of pH, precipitate as a porous hydroxide of extremely small particle size. The hydroxide appears to be identical to pseudoboehmite. The overall rate of the corrosion reaction is controlled by this dissolution of the film and by the disposition of the soluble products. The corrosion rate is nearly independent of specimen potential, solution pH below 10, and of the presence in solution of many salts at concentrations as high as 1 mole/1. The rate is strongly dependent on temperature, on the presence of specific inhibitive salts, and increases rapidly at high pH. In environments which preclude oxide dissolution, e.g., water + dioxane mixtures or water vapour, the corrosion rate is drastically reduced. The corrosion rate is constant when no solid hydroxide is formed, and is otherwise a strong function of the amount of reaction. At high temperatures the rate decreases with time as the precipitated hydroxide hinders transport. At lower temperatures the rate may first increase with time as nucleation of hydroxide provides sinks for soluble species close to the interface, and then at long times the rate decreases as the hydroxide layer thickens. Electrons are removed more readily at special sites, and in our specimens these sites were primarily at the grain boundaries. Electron removal results in increased hydroxyl ion concentration, and this in turn results in more rapid attack on the protective oxide. At such cathodes the oxide film is maintained at a small constant thickness at which there is a balance between the rates of its dissolution by the basic solution and growth because of the great affinity of aluminum for oxygen. At cathodic sites there is thus anodic activity as well, but electron removal predominates. The metal is corroded more rapidly at cathodes, producing pronounced grain boundary attack in our specimens. Application of anodic potentials eliminates grain boundary attack.

Allosteric Effects in Nicotinamide Adenine Dinucleotide Phosphate-specific Glutamate Dehydrogenase from Neurospora

Abstract Neurospora nicotinamide adenine dinucleotide phosphate-specific glutamate dehydrogenase is predominantly in an inactive form at pH 7.2, is fully active above pH 8.0, and is partially active at intermediate pH values. Between pH 7.0 and 7.4, and in the absence of NADPH, the enzyme is activated by the substrates 2-oxoglutarate and glutamate, as well as by the nonsubstrates citrate, isocitrate, succinate, malate, fumarate, oxalacetate, or ethylenediaminetetraacetate. Curves relating activation to activator or substrate concentration were found to be sigmoid, indicating cooperation between activating molecules. At lower pH values the addition of NADPH results in nearly complete inactivation. Under these conditions activation requires 2-oxoglutarate specifically; glutamate or nonsubstrate acids have no activating effect. However, 2-oxoglutarate activates even better than in the absence of NADPH. NADP+ causes some activation by itself and enhances activation by 2-oxoglutarate, glutamate, and succinate. It is concluded that the enzyme has allosteric properties, the possible regulatory significance of which is briefly discussed. Properties previously reported for certain mutant varieties of this enzyme obtained from amination-deficient (am) mutants and their partial revertants are now seen as distorted manifestations of the normal allosteric interactions reported here for the enzyme obtained from a wild type strain.

THE DISTRIBUTION CONSTANT FOR EXCHANGE OF CALCIUM AND MAGNESIUM IN WYOMING BENTONITE

The distribution constant for exchange of calcium and magnesium ions was determined in Wyoming bentonite suspensions with pH values varying from 3.5 to 9.2. Using a neutral salt to extract the exchangeable cations, the mean value of the distribution constant using ion activities in solution was found to be 1.06 and there was no significant difference in the constant at different pH values. Thus, the presence of Al ions in the acid preparations, the precipitation of hydroxy aluminum polymers on the clay surfaces at intermediate pH values, and the precipitation of excess base in the alkaline region possibly as double hydroxides of Al and Mg or Ca had no significant effect on the exchange distribution of Mg and Ca ions.Under comparable conditions, the ion product (Al) (OH)3 had the same value in the presence of Ca or Mg. Because of this and the fact that the distribution constant for exchange was approximately 1, the two ions may be considered as a single ionic species for practical purposes in suspensions of Wyoming bentonite.

The Nature of the Amino Acid Residues Involved in the Inactivation of Ribonuclease by Iodoacetate

SUMMARY Ribonuclease is inactivated by iodoacetate at 40”, the rate depending upon the pH at which reaction is carried out. It, is most rapid between pH 5.5 and 6, and at, pH 2.8, less rapid at intermediate pH values and under more alkaline conditions (pH 8.5 to 10). The variation in rate is a consequence of the fact that different types of amino acid residues in the ribonuclease molecule react with iodoacetate at the different pH values. Amino acid analyses showed that at pH 5.5 to 6, a histidine residue is involved, at pH 8.5 to pH 10 reaction occurs with the e-amino groups of lysine residues, whereas at pH 2.8, the thio- ether sulfur of one or more methionine residues is alkyla,ted. After removal of iodoacetate, the various reaction mixtures were chromatographed on columns of IRC-50 in an attempt to isolate purified carboxymethylated derivatives. From the mixture present after reaction at pH 5.5, it was possible to isolate a chromatographically homogeneous protein that was totally in- active and differed from ribonuclease A only in that the imidaz- ole group of a single histidine residue had been carboxymethyl- ated. The fact, that the pH optimum for the reaction leading to the formation of imidazole carboxymethyl ribonuclease is around pH 5.5 to 6, instead of at the more alkaline pH values observed for the alkylation of cy-N-acetylhistidine, suggests that inactivation takes place as a result of substitution on a particular one of the four histidine residues present in ribonuclease.

PH standards of high acidity and high alkalinity and the practical scale of pH

The practical scale of pH is defined in terms of the electromotive force of the galvanic Pt; H2 (g), solution | satd. KC1 | standard, H2 (g); Pt. This potential is usually obtained as the difference of two electromotive force values for a cell with glass and calomel electrodes, one of which is a calibration with a standard of known pH. Appropriate corrections must be applied if the glass electrode does not respond to changes in hydrogen-ion activity in exactly the same manner as the hydrogen electrode. However, there is no simple means of correcting pH measurements for the potential differences at the junctions of the solution and the standard with the solution of potassium chloride. These errors are sufficiently large in highly acid and highly alkaline solutions to render uncertain the interpretation of measured pH in these regions, in spite of the fact that reasonably accurate standards of hydrogen-ion activity are available at intermediate pH values. The purpose of this study was twofold: (a) to determine the extent of aberration of the practical pH scale near its ends, and (b) to select new standards that might improve the accuracy of pH measurements and facilitate their interpretation over the entire practical scale, with particular attention to the regions of high acidity and high alkalinity. The results indicate that pH obtained by adjustment of the meter with the present standards (phthalate, pH 4.01 at 25° C; phosphate, pH 6.86; borax, pH 9.18) will usually be low by at least 0.02 to 0.05 unit above pH 11, while errors as great as 0.03 unit, either positive or negative, are not uncommon below pH 2.5. The following additional standards were selected to supplement the three presently available: (1) 0.01-M potassium tetroxalate— pH 2.15 at 25° C; (2) potassium hydrogen tartrate (saturated at room temperature)—pH 3.56; (3) 0.025-M sodium acid succinate, 0.025-M sodium succinate—pH 5.40; (4) 0.025-M sodium bicarbonate, 0.025-M sodium carbonate—pH 10.02; and (5) 0.01 M trisodium phosphate—pH 11.72. The choice was based on a comparison of pH derived from cells with and without liquid junction in a study of 41 promising standard solutions. The pH on the practical scale was determined at 25° C, and electromotive force measurements of hydrogen-silver chloride cells without liquid junction were made at 0°, 10°, 25°, and 38° C.

The Dyeing Properties of Chlorinated Wool I–Qualitative Experiments using Wool chlorinated at Various pH Values1

A preliminary qualitative investigation has been carried out into the dyeing properties of wool treated in solutions of sodium hypochlorite at various pH values. It has been confirmed that, when using selected equalising acid, neutral–dyeing acid, direct cotton, and chrome dyes, wool treated in acid solutions of hypochlorite absorbs more dye than untreated wool when samples of both are dyed togethor in the same solution, whereas when treated under neutral or alkaline conditions it absorbs less dye and a resist effect is obtained. Treatment in hypochlorite solutions at certain intermediate pH values yields material with dyeing properties similar to untreated wool. Using basic dyes, the chlorinated wool absorbs more dye than the unchlorinated material irrespective of the pH at which the treatment is carried out.