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Physiochemical and functional characterization of antigen proteins eluted from aluminum hydroxide adjuvant.

We have characterized protein antigens after quantitative dissociation from aluminum hydroxide adjuvant. Bovine serum albumin (BSA) and a multi-antigen vaccine for Group A Streptococcus (GrAS Vaccine) were formulated on aluminum hydroxide, stored for > or =10 days then eluted with a 48-h treatment at 4 degrees C with 0.85% H(3)PO(4) plus 4M guanidine HCl (GnHCl). BSA is recovered from adjuvant at 92+/-2%. GrAS antigens are equally recovered from GrAS Vaccine (95+/-11% of total protein expected using multiple lots stored for up to 12 months). Recovery after elution is similar when determined by RP-HPLC, SEC-HPLC, UV absorbance, or Bradford methods. Eluted antigens are structurally and functionally intact as judged relative to both treated and untreated antigen controls by SDS-PAGE, RP-HPLC, SEC-HPLC, and after desalting by circular dichroism, bis-ANS binding, and antigenicity determined by ELISA. When formulated and stored for a few weeks, BSA has more dimer (31+/-5%) relative to the elution control (9% dimer) as detected by SEC-HPLC, suggesting that BSA microaggregation is promoted on aluminum. Antigens eluted from very aged GrAS Vaccine (>12 months) show marked changes by RP-HPLC. Structural changes in the antigens under elution conditions were evaluated using bis-ANS, a fluorescent probe of protein structure. Binding of bis-ANS increases fluorescence approximately 100-fold and is significantly diminished with increasing GnHCl concentrations indicating a progressive denaturing of the proteins. At 4M GnHCl (with or without 0.85% H(3)PO(4)) the GrAS antigens are fully denatured and BSA is partially denatured. Interestingly, the addition of 0.85% H(3)PO(4) increases bis-ANS binding on GrAS antigens and reduces the denaturing of GrAS antigens and BSA by chaotropes. Desalting or diluting the eluted antigens results in renaturing of the proteins as judged by bis-ANS fluorescence, circular dichroism and antigenicity testing. The elution method provides a novel approach for high recovery and characterization of GrAS Vaccine antigens and may be applicable to the study of many aluminum hydroxide-bound vaccines.

Adjuvants, Immunologic↗

The properties of enzyme-hydrolyzed cellulose in aqueous sodium hydroxide.

Pure natural cellulose (softwood pulp) modified with cellulase is allowed to react with sodium hydroxide in a muller, and changes in structure and properties are investigated by FTIR and DSC. The reactivity of cellulose for some dissolving and derivatization processes is shown to be improved by an enzymatic hydrolysis and admixture with sodium hydroxide. The modified cellulose dissolved at 9% (wt) sodium hydroxide at -10 degrees C at 6% pulp consistency, while the DP of cellulose is >350.

Ascomycota↗

Prediction of retention times for anions in linear gradient elution ion chromatography with hydroxide eluents using artificial neural networks.

The feasibility of using an artificial neural network (ANN) to predict the retention times of anions when eluted from a Dionex AS11 column with linear hydroxide gradients of varying slope was investigated. The purpose of this study was to determine whether an ANN could be used as the basis of a computer-assisted optimisation method for the selection of optimal gradient conditions for anion separations. Using an ANN with a (1, 10, 19) architecture and a training set comprising retention data obtained with three gradient slopes (1.67, 2.50 and 4.00 mM/min) between starting and finishing conditions of 0.5 and 40.0 mM hydroxide, respectively, retention times for 19 analyte anions were predicted for four different gradient slopes. Predicted and experimental retention times for 133 data points agreed to within 0.08 min and percentage normalised differences between the predicted and experimental data averaged 0.29% with a standard deviation of 0.29%. ANNs appear to be a rapid and accurate method for predicting retention times in ion chromatography using linear hydroxide gradients.

Anions↗

Relationship of mercury with aluminum, iron and manganese oxy-hydroxides in sediments from the Alto Pantanal, Brazil.

Sediments from nine floodplain lakes in Pantanal were analyzed for a large-scale (300 km) survey of mercury (Hg) load in sediments and soils of the Alto Pantanal and to study the relationship between Hg and reactive aluminum, iron, and manganese oxy-hydroxides. The results were compared with the Hg content in river and stream sediments from the Poconé gold mining area, where Hg has been extensively used and still is in use. The results indicate that the Hg concentrations were elevated in river sediment close to the mining area in Bento Gomes river basin (average in the < 74-microm fraction 88.9 ng Hg g(-1) dry wt.; interquartile range 50.3-119.5), but there was no clear indication that the local Hg emissions have contaminated the remote floodplain lakes, where concentrations were surprisingly low (average in the < 74-microm fraction 33.2 ng Hg g(-1) dry wt. sediment; interquartile range 18.4-46.8), in particular when considering geochemical characteristics of the sediment. The sediment from the floodplain lakes contained less Hg-tot and more reactive iron oxy-hydroxides than soils from the Tapajós area in the Amazon basin. This resulted in a mass ratio between Hg and amorphous oxy-hydroxides of only 5 x 10(-6) for Hg-tot/Fe-oxa (interquartile range 3-7 x 10(-6).

Aluminum↗

Effect of calcium hydroxide paste dressing on uninstrumented root canal wall.

Dissolution of soft tissues on the uninstrumented root canal wall by an intracanal dressing of calcium hydroxide paste was examined by scanning electron microscopy (SEM). The roots of human extracted teeth were split longitudinally into halves, and the pulp tissue was removed by forceps. Then, the halves were divided randomly into three groups. Specimens in the control group were given no treatment. Specimens of the 1-wk group were stored for 7 days in calcium hydroxide-saline paste under the condition of 100% humidity and 37 degrees C. Specimens of the 4-wk group were stored for 28 days in the same manner, but washed and replaced into new paste every week. Root canal walls of these specimens were compared under a scanning electron microscope. A layer of odontoblasts was observed in the control group, whereas it had disappeared completely in the 1- and 4-wk groups. However, the predentin remained almost intact in the 1-wk group, and although the surface was eroded, in the 4-wk group. It was found that calcium hydroxide paste dressing could dissolve the odontoblastic cell layer, but had little effect on predentin.

Calcium Hydroxide↗

Evaluation of diagnostic radiopaque contrast materials used with calcium hydroxide.

An evaluation of three different radiopaque materials used in combination with calcium hydroxide powder for diagnostic contrast was made. Diatrizoate meglumine (65%) and iothalamate meglumine (60%) compared favorably with barium sulfate (35%). Since barium sulfate has demonstrated problems by altering the manipulative properties of calcium hydroxide as well as demonstrating a residual radiopacity, there may be advantages to the use of resorbable diatrizoate or iothalamate compounds as alternatives. These soluble iodine compounds also make possible an excellent aqueous medium in which to mix calcium hydroxide.

Barium Sulfate↗

Intracanal pH changes of calcium hydroxide pastes exposed to carbon dioxide in vitro.

It has been suggested that the main benefit of using calcium hydroxide as an intracanal medicament lies in its bactericidal effect, provided that the pH of the paste is above 12.5. The purpose of this study was to measure changes in the pH of several calcium hydroxide pastes sealed in root canals for 30 days. Sixty-two extracted, single-rooted human teeth were endodontically prepared using K-files up to size 60. The teeth were separated at random into six equal groups to be filled with either Calxyl, Hydrocalcium, or a paste made by mixing calcium hydroxide powder with either distilled water, camphorated p-monochlorophenol, local anesthetic solution, or Solvidont. Cavidentin was used to seal the coronal orifice of the teeth that were placed individually in vials containing 10 mL distilled water. Five vials of each group were exposed to air at room temperature, whereas the other five vials were exposed to carbon dioxide in a closed container. The pH of the paste in the root canal was measured after 30 days. There was no significant (p > 0.01) change in the pH (mean 13.11) of the pastes placed in teeth before and after exposure to air, whereas the pH of the pastes in teeth exposed to carbon dioxide was significantly (p < 0.01) reduced (mean 12.54). There was no significant difference in pH between the six preparations. After 30 days of exposure to carbon dioxide, they still maintained a purportedly bactericidal pH within the root canal.

Analysis of Variance↗

Antimicrobial evaluation of calcium hydroxide in infected dentinal tubules.

The objective of this study was to evaluate the antimicrobial activity of calcium hydroxide in infected dentinal tubules. Four microorganisms, strains of ATCC (Streptococcus faecalis (ATCC-29212), Staphylococcus aureus (ATCC-6538), Bacillus subtilis (ATCC-6633), and Pseudomonas aeruginosa (ATCC-27853)) and one mixture of these were used. These strains were inoculated in brain heart infusion (BHI) and incubated at 37 degrees C for 24 h. Sixty-three human maxillary central incisors were prepared and sterilized by autoclaving. Five groups of 12 teeth each were contaminated for 28 days using new 24-h cultures every 72 h, prepared and adjusted to tube 2 of the MacFarland scale (6 x 10(8) cells/ml). Root canals were then irrigated with 5 ml of saline, dried, and completely filled with calcium hydroxide paste. At intervals of 0, 48, and 72 h, and 7 days, dressings were removed and teeth were immersed in 5 ml of BHI and incubated at 37 degrees C for 48 h to observe the growth and multiplication of the microorganisms. Three uninoculated teeth were maintained in a humid environment as an aseptic control. These teeth were immersed in BHI and maintained at 37 degrees C for 7 days to determine microbial growth. Bacterial growth was shown by turbidity of the culture medium and confirmed by seeding these broths on BHI agar at 37 degrees C for 24 h. The positive BHI tubes were selected, and inoculum was spread on the surface of BHI agar, followed by the same incubation conditions. Gram stain was conducted from BHI growth and from colonies growing on solid medium. Calcium hydroxide in infected dentinal tubules showed no antimicrobial effect on S. faecalis, S. aureus, B. subtilis, P. aeruginosa, or on the bacterial mixture used throughout the experiment.

Bacillus subtilis↗

Ginseng extract in aluminium hydroxide adjuvanted vaccines improves the antibody response of pigs to porcine parvovirus and Erysipelothrix rhusiopathiae.

Ginseng, the dry extract prepared from the Panax ginseng C.A. Mayer-root contain immunomodulators named ginsenosides, which in the pig enhance the antibody response to viral and bacterial antigens. The enhancing effect of ginseng was demonstrated vaccinating pigs against porcine parvovirus (PPV) and Erysipelothrix rhusiopathiae infections, using commercially available vaccines. The potency of the licensed, aluminium hydroxide adjuvanted; vaccines were compared with those supplemented with ginseng. The antibody response to PPV was measured by the haemagglutination inhibition (HI) test whereas the mouse potency test and ELISA evaluated the immune response to E. rhusiopathiae. Antibodies to the 64-66 kDa glycoprotein of the E. rhusiopathiae were demonstrated by immunoblotting. The qualitative antibody responses were evaluated by means of ELISA(s) using monoclonal antibodies to swine IgG1 and IgG2. The addition of 2mg ginseng per vaccine dose, potentiate the antibody response of the commercial vaccines without altering their safety. Significantly higher (P<0.001) antibody titres were achieved to both PPV and to E. rhusiopathiae by the supplementation with ginseng. Aluminium hydroxide adjuvanted vaccines favoured the production of IgG1 antibodies. Interestingly, the vaccines supplemented with ginseng favoured IgG2. The vaccines used in the evaluations varied in their immunogenic potency. However, after the addition of ginseng the less immunogenic vaccine proved to be as potent as the better one without ginseng. Thus, the use of ginseng as a co-adjuvant provides a simple, safe and cheap alternative for improving the potency of aluminium hydroxide adjuvanted vaccines.

Adjuvants, Immunologic↗

Five methods of calcium hydroxide intracanal placement: an in vitro evaluation.

OBJECTIVE: The purpose of this study was to evaluate root canal dressing with calcium hydroxide paste using 5 techniques of placement. STUDY DESIGN: After endodontic preparation, each of 50 single-rooted premolars was filled with calcium hydroxide paste through use of one of the following techniques: Gutta-Condensor, MecaShaper, K-type ultrasonic file, Lentulo, and Pastinject. Radiographs of all 3 third root zones were made after cleaning and shaping and obturation. Density measurements (in pixels) were registered by means of an image analyzer and then compared through use of analysis of variance and Newman-Keuls multiple means tests. RESULTS: With respect to average filling density, the techniques ranked as follows, in ascending order: Gutta-Condensor, MecaShaper, K-type ultrasonic file, Lentulo, and Pastinject. There was a significant difference (P <.0009) between the Gutta-Condensor and Pastinject techniques, between the MecaShaper and Pastinject techniques, and between the ultrasonic file and Pastinject techniques; all favored the Pastinject. There were no significant differences between the Gutta-Condensor, MecaShaper, ultrasonic file, and Lentulo techniques or between the Lentulo and Pastinject techniques. CONCLUSIONS: The use of a specially designed paste carrier (the Pastinject) improved calcium hydroxide placement in root canals.

Analysis of Variance↗

Direct capping of human pulps with a dentin bonding system or with calcium hydroxide cement.

OBJECTIVE: To study human pulp reactions to direct cappings with a dentin adhesive compared with a calcium hydroxide cement, especially with respect to formation of hard tissue in the exposure site. STUDY DESIGN: Direct pulp cappings were made in 34 caries-free premolars scheduled for extraction due to orthodontic treatment. Half of the teeth were capped with a dentin adhesive, the other half with a calcium hydroxide cement. After periods of from 7 to 70 days, the teeth were extracted and processed for light microscopical examination. RESULTS: No postoperative sensitivity was experienced. A slight inflammatory reaction was seen in both groups in the short observation period. In the long observation period, significantly more "dentin bridging" was seen in the calcium hydroxide group than in the dentin adhesive group (P < .05). CONCLUSION: The findings from the present limited material is in accordance with previous findings from human studies and contraindicate use of dentin adhesives for direct pulp capping.

Adolescent↗

Interaction of kappa-carrageenan with nickel, cobalt, and iron hydroxides.

In a previous paper, it was shown that the functional polysaccharide kappa-carrageenan acts as an efficient stabilizer to prevent the precipitation of iron oxides and hydroxides up to very high pH values (Jones, et al. Colloid Polym. Sci. 2000, 278, 491-501). Here, this process was investigated for its application to synthesize stable cobalt and nickel oxide particles. Nickel hydroxide nanoparticles were well stabilized for long periods, while the hydroxide of cobalt(II), although initially stable at pH 13, progressively showed some complex structure rearrangement, invoked by alkaline oxidation of the primary particles. The final product of this oxidation/self-assembly process are spherical, fluffy superstructures of cobalt(III) oxide platelets with an overall diameter of 12 microns, in coexistence with 65-75% of free kappa-carrageenan.

Carrageenan↗

Verdoheme reactivity. Remarkable paramagnetically shifted (1)H NMR spectra of intermediates from the addition of hydroxide or methoxide with Fe(II) and Fe(III) verdohemes.

Studies of the reaction of 5-oxaporphyrin iron complexes (verdohemes) with methoxide ion or hydroxide ion have been undertaken to understand the initial step of ring opening of verdohemes. High-spin [ClFe(III)(OEOP)] undergoes a complex series of reactions upon treatment with hydroxide ion in chloroform, and similar species are also detected in dichloromethane, acetonitrile, and dimethyl sulfoxide. Three distinct paramagnetic intermediates have been identified by (1)H NMR spectroscopy. These reactive species are formed by addition of hydroxide to the macrocycle and to the iron as an axial ligand. Treatment of low-spin [(py)(2)Fe(II)(OEOP)]Cl (OEOP is the monoanion of octaethyl-5-oxaporphyrin) with excess methoxide ion in pyridine solution produces [(py)(n)()Fe(II)(OEBOMe)] (n = 1 or 2) ((OEBOMe), dianion of octaethylmethoxybiliverdin), whose (1)H NMR spectrum undergoes marked alteration upon addition of further amounts of methoxide ion. An identical (1)H NMR spectrum, which is characterized by methylene resonances with both upfield and downfield paramagnetic shifts, is formed upon treatment of [Fe(II)(OEBOMe)](2) with methoxide in pyridine solution and results from the formation of [(MeO)Fe(II)(OEBOMe)](-).

Acetonitriles↗

Solvent effects on the structural and formyl substrate reactivity properties of a nitrogen/sulfur-ligated zinc hydroxide complex.

The solution structural and formyl substrate reactivity properties of a nitrogen/sulfur-ligated zinc hydroxide complex, [(bmnpaZn)2(mu-OH)2](ClO4)2 (1, bmnpa = N,N-bis-2-(methylthio)ethyl-N-((6-neopentylamino-2-pyridyl)methyl)amine), in acetonitrile and methanol are reported. In CH3CN, 1 has a binuclear cation [(bmnpaZn)2(mu-OH)2]2+ that is stabilized by secondary hydrogen bonding and CH/pi interactions involving the bmnpa chelate ligand. In CH3OH, 1 undergoes reaction with solvent to yield a zinc methoxide species, as determined by 1H NMR and electrospray mass spectral analysis. Treatment of 1 with methyl formate in CH3CN results in stoichiometric hydrolysis of the formyl ester to produce [(bmnpa)Zn(O2CH)]ClO4 (2) and methanol. The formate complex was identified via independent synthesis and characterization (X-ray crystallography, 1H and 13C NMR, FTIR, LRFAB-MS, conductance, and elemental analysis). In the solid state, 2n has a formate-bridged coordination polymer-type structure. However, in CH3CN, 2 behaves as 1:1 electrolyte, indicating cleavage of the polymer structure into mononuclear [(bmnpa)Zn(O2CH)]ClO4 species. Treatment of 1 with a stoichiometric amount of formanilide in CH3CN for 48 h at 45 degrees C results in decomposition of the zinc hydroxide complex to yield the free bmnpa ligand and an inorganic solid, presumably a zinc hydroxide or oxide species. Treatment of 1 with a stoichiometric amount of ethyl formate in CD3OD results in rapid, quantitative transesterification of the formyl carboxylate ester. A control reaction indicates that this transesterification reaction does not occur on the same time scale in the absence of the catalyst. Treatment of 1 with an excess of ethyl formate in CD3OD results in catalytic formyl carboxylate ester transesterification, with approximately 1000 turnovers in 60 min at 22(1) degrees C. Treatment of a CD3OD solution of 1 (0.5 equiv) with formanilide (1 equiv) results in the formation of aniline, d3-methyl formate, and the zinc formate complex 2. While aniline is produced stoichiometrically, the yield of d3-methyl formate varied from 30 to 50%, and the yield of 2 varied from 50 to 70% in repetitive experiments. Formation of both d3-methyl formate and 2 indicates that both methanolysis and hydrolysis reactions take place.

Crystallography, X-Ray↗

Mononuclear nitrogen/sulfur-ligated zinc methoxide and hydroxide complexes: investigating ligand effects on the hydrolytic stability of zinc alkoxide species.

The synthesis and properties of mononuclear zinc methoxide ([(ebnpa)Zn-OCH3]ClO4) (1) and hydroxide ([(ebnpa)Zn-OH]ClO4) (2) complexes of a new mixed nitrogen/sulfur ligand (ebnpa = N-2-(ethylthio)ethyl-N,N-bis(6-neopentylamino-2-pyridylmethyl)amine) are reported. The structures of 1 and 2 were determined by X-ray diffraction. Each possesses a single zinc-coordinated anion (methoxide or hydroxide) and exhibits an overall trigonal bipyramidal geometry. Structural and spectroscopic studies indicate the presence of two hydrogen-bonding interactions involving the oxygen atom of the zinc-bound anion in each complex. Treatment of [(ebnpa)Zn-OH]ClO4 with CH3OH results in the formation of an equilibrium mixture of 1 and 2. 1H NMR spectroscopic methods were used to examine the equilibrium as a function of temperature, yielding KMe (304 K) = 0.30(8), DeltaHMe = -0.9(1) kcal/mol, and DeltaSMe = -5(1) eu. The negative enthalpy indicates that spontaneous zinc alkoxide formation from a hydroxide precursor occurs in this system at low temperature. Using the experimentally determined DeltaHMe value, we found the homolytic Zn-O bond dissociation energy (BDE) in the Zn-OCH3 unit to be approximately -14 kcal/mol relative to the Zn-O BDE in the Zn-OH unit.

Alcohol Dehydrogenase↗

Improved performance in dye-sensitized solar cells employing TiO2 photoelectrodes coated with metal hydroxides.

The performance of dye-sensitized solar cells (DSCs) was compared before and after processing the TiO(2) electrodes by minute-order electrochemical reactions with metal nitrates, where the metals were Mg, Zn, Al, and La, in 2-propanol. An overcoating of metal hydroxide was formed without the need for a sintering process, and magnesium hydroxide was found to give the largest improvement in photovoltage, fill factor, and eventually overall conversion efficiency of the DSCs. To analyze the nature of the improvement, the diffusion coefficient (D) and electron lifetime (tau) were determined. While little influence of overcoating on D was seen, a correlation between the increase in tau and V(oc) was observed for the metals examined here. The remarkable improvement in the electron lifetime of the DSCs suggests that an overcoating with magnesium hydroxide species function as the blocking layers at the fluorine-doped tin oxide and TiO(2) interfaces, thus contributing to the suppression of electron leakage, i.e., recombination processes between unidirectional transporting electrons and poly-iodides such as tri-iodide in the processed TiO(2) photoelectrode systems. The increase in V(oc) can be explained by the increased electron density caused by the increase in electron lifetime.

2-Propanol↗

Liposomal subunit vaccines: effects of lipid A and aluminum hydroxide on immunogenicity.

Protein and peptide antigens frequently are only slightly immunogenic when utilized alone in vaccines. Formulation of these antigens in a carrier vehicle, particularly when an adjuvant is included, often results in markedly enhanced immune responses. Encapsulation of peptide and protein antigens in liposomes generally results in a relatively slight enhancement of antibody production compared with that observed with the antigen alone. However, when lipid A is included in the liposomes, immunogenicity is markedly increased compared both with antigen alone and with antigen encapsulated in liposomes lacking lipid A. The enhancement of the immune response caused by lipid A is dependent on the liposomal lipid A dose. Aluminum salts, such as aluminum hydroxide and aluminum phosphate, act as adjuvants for some antigens and are used in a variety of human vaccines. When liposomes containing encapsulated protein or peptide antigens were adsorbed with aluminum hydroxide, an enhancement of the antibody response was observed with some antigens, whereas with other antigens the presence of aluminum hydroxide either had no effect or resulted in a diminished antibody response. Immunogenicity of protein and peptide antigens can be enhanced by formulation in liposomes containing lipid A and, depending on the antigen, can be enhanced further by adsorption of the liposomal antigen formulation with aluminum salts.

Adjuvants, Immunologic↗

A comparative study of hydroxide adsorption on the (111), (110), and (100) faces of silver with cyclic voltammetry, ex situ electron diffraction, and in situ second harmonic generation.

Hydroxide adsorption on the (111), (110), and (100) faces of silver electrodes from mixed NaOH/NaF solution is studied using cyclic voltammetry and in situ second harmonic generation (SHG). Cyclic voltammograms for the three low index silver planes in alkaline electrolytes are for the first time compared. They show two pairs of anodic and cathodic peaks in the potential interval below the equilibrium Ag/Ag(2)O potential. These are attributed to the specific adsorption of hydroxide ions followed by submonolayer oxide formation. The differences in the cyclic voltammograms for the (111), (110), and (100) planes are attributed to different (i) work functions, (ii) surface atomic densities, and (iii) corrugation potentials for these surfaces. Ex situ low energy electron diffraction (LEED) and reflection high energy electron diffraction (RHEED) show that disordered adlayers are formed on Ag(111) and Ag(100), in contrast to Ag(110), where ordered structures are produced in the region of the first pair of current peaks. In the region of the second pair of peaks, LEED indicates disordered oxide phases on each crystal plane and RHEED shows the presence of small islands of c(2 x 2) structure at some potentials on (110) and (100). SHG measurements were performed (i) in the potential scan mode at constant rotational angle and (ii) at constant potential as a function of the rotational angle. The isotropic (for the (111), (110), and (100) planes) and anisotropic (for the (110) and (111) planes) contributions to the SHG intensity were calculated by fitting the experimental data and are discussed in terms of their dependence on the charge density at the interface, on hydroxide adsorption, and on submonolayer oxide formation.

Adsorption↗