pH and calcium ion release evaluation of pure and calcium hydroxide-containing Epiphany for use in retrograde filling

Objective: Hydroxyl (OH(-)) and calcium (Ca(++)) ion release was evaluated in six materials: G1) Sealer 26, G2) White mineral trioxide aggregate (MTA), G3) Epiphany, G4) Epiphany + 10% calcium hydroxide (CH), G5) Epiphany + 20% CH, and G6) zinc oxide and eugenol. Material and Methods: Specimens were placed in polyethylene tubes and immersed in distilled water. After 3, 6, 12, 24, and 48 h, 7, 14, and 28 days, the water was assessed for pH with a pH meter and for Ca++ release by atomic absorption spectrophotometry. Results: G1, G2, G4, and G5 had the highest pH until 14 days (p < 0.05). G1 presented the highest Ca(++) release until 6 h, and G4 and G5, from 12 h through 14 days. Ca(++) release was greater for G1 and G2 at 28 days. G6 released the least Ca(++). Conclusions: MTA, Sealer 26, Epiphany, and Epiphany + CH release OH-and Ca(++) ions. Epiphany + CH may be an alternative as retrofilling material.

pH, Calcium Ion Release, and Setting Time of an Experimental Mineral Trioxide Aggregate-based Root Canal Sealer

Introduction: An experimental mineral trioxide aggregate sealer (MTAS) has been developed for use as a root canal sealer. The aim of this study was to evaluate the setting time, pH, and calcium ion release of MTAS compared with white Portland cement (CPB-40; Votorantin Cimentos, Camargo Correa SA, Pedro Leopoldo, MG, Brazil), white MTA Angelus (MTA; Angelus, Londrina, PR, Brazil), and AH Plus (Dentsply DeTrey, Konstanz, Germany). Methods: For the evaluation of setting time, each material was analyzed using Gilmore-type needles. Polyethylene tubes with the materials were immersed in distilled water for the measurement of pH (digital pH meter) and calcium release (atomic absorption spectrophotometry). The evaluations were performed at 3, 6, 12, 24, and 48 hours and 7, 14, and 28 days. Data were analyzed by analysis of variance and the Tukey test at 5% significance level. Results: MTAS showed higher calcium release at all experimental periods, a greater increase in pH up to 48 hours and the longest setting time. Conclusions: MTAS presented favorable properties for its indication as a root canal sealer. (J Endod 2011;37:844-846)

Evaluation of pH and Calcium Ion Release of Calcium Hydroxide Pastes Containing Different Substances

Introduction: The objective of this study was to evaluate the pH and calcium ion release of calcium hydroxide pastes associated with different substances. Methods: Forty acrylic teeth with simulated root canals were divided into 4 groups according to the substance associated to the calcium hydroxide paste: chlorhexidine (CHX) in 2 formulations (1% solution and 2% gel), Casearia sylvestris Sw extract, and propylene glycol (control). The teeth with pastes and sealed coronal accesses were immersed in 10 mL deionized water. After 10 minutes, 24 hours, 48 hours, and 7, 15, and 30 days, the teeth were removed to another container, and the liquid was analyzed. Calcium ion release was measured by atomic absorption spectrophotometry, and pH readings were made with a pH meter. Data were analyzed statistically by analysis of variance and Tukey test (alpha = 0.05). Results: Calcium analysis revealed significant differences (P < .05) for 1% CHX solution and 2% CHX gel at 10 minutes. After 24 hours, 2% CHX gel x Control and 2% CHX gel x 1% CHX solution differed significantly (P < .05). After 48 hours, there were significant differences (P < .05) for 2% CHX gel x Control and Extract x Control. No differences (P > .05) were observed among groups in the other periods. Regarding the pH, there were significant differences (P < .05) for 2% CHX gel x Control and 2% CHX gel x 1% CHX solution after 48 hours and for 2% CHX gel x Control after 15 days. In the other periods, no differences (P > .05) were observed among groups. Conclusions: All pastes behaved similarly in terms of pH and calcium ion release in the studied periods. (J Endod 2009;35:1274-1277)

Evaluation of pH and Calcium Ion Release of Root-end Filling Materials Containing Calcium Hydroxide or Mineral Trioxide Aggregate

Introduction: To evaluate calcium ion release and pH of Sealer 26 (S26) (Dentsply, Rio de Janeiro, RJ, Brazil), white mineral trioxide aggregate (MTA), Endo CPM Sealer (CPM1) (EGEO SRL Bajo licencia MTM Argentina SA, Buenos Aires, Argentina), Endo CPM Sealer in a thicker consistency (CPM 2), and zinc oxide and eugenol cement (ZOE). Methods: Material samples (n = 10) were placed in polyethylene tubes and immersed in 10 mL of distilled water. After 3, 6,12,24, and 48 hours and 7,14, and 28 days, the water pH was determined with a pH meter, and calcium release was assessed by atomic absorption spectrophotometry. An empty tube was used as the control group. Results: The control group presented a pH value of 6.9 at all studied periods and did not show the presence of calcium ion. S26 presented greater hydroxyl ion release up to 12 hours (p < 0.05). From 24 hours until 28 days, S26, MTA, CPM1, and CPM2 had similar results. in ail periods, ZOE presented the lowest hydroxyl ion release. CPM1, followed by CPM2, released the most calcium ions until 24 hours (p < 0.05). Between 48 hours and 7 days, CPM1 and CPM2 had the highest release. A greater calcium ion release was observed for CPM2, followed by CPM1 at 14 days and for S26, CPM1, and CPM2 at 28 days. ZOE released the least calcium ions in all periods. Conclusion: Sealer 26, MTA, and Endo CPM sealer at normal or thicker consistency release hydroxyl and calcium ions. Endo CPM sealer may be an alternative as root-end filling material. (J Endod 2009;35:1418-1421)

Crystal structure of an acidic platelet aggregation inhibitor and hypotensive phospholipase A(2) in the monomeric and dimeric states: insights into its oligomeric state

Phospholipases A(2) belong to the superfamily of proteins which hydrolyzes the sn-2 acyl groups of membrane phospholipids to release arachidonic acid and lysophospholipids. An acidic phospholipase A(2) isolated from Bothrops juraracussu snake venom presents a high catalytic, platelet aggregation inhibition and hypotensive activities. This protein was crystallized in two oligomeric states: monomeric and dimeric. The crystal structures were solved at 1.79 and 1.90 Angstrom resolution, respectively, for the two states. It was identified a Na+ ion at the center of Ca2+-binding site of the monomeric form. A novel dimeric conformation with the active sites exposed to the solvent was observed. Conformational states of the molecule may be due to the physicochemical conditions used in the crystallization experiments. We suggest dimeric state is one found in vivo. (C) 2004 Elsevier B.V. All rights reserved.

Comparison between apo and complexed structures of bothropstoxin-I reveals the role of Lys122 and Ca2+-binding loop region for the catalytically inactive Lys49-PLA(2)s

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Crystal structure of a myotoxic Asp49-phospholipase A(2) with low catalytic activity: Insights into Ca2+ -independent catalytic mechanism

A myotoxic Asp49-phospholipase A(2) (Asp49-PLA(2)) with low catalytic activity (BthTX-II from Bothrops jararacussu venom) was crystallized and the molecular-replacement solution has been obtained with a dimer in the asymmetric unit. The quaternary structure of BthTX-II resembles the myotoxic Asp49-PLA2 PrTX-III (piratoxin III from B. pirajai venom) and all non-catalytic and myotoxic dimeric Lys49-PLA(2)s. Despite of this, BthTX-II is different from the highly catalytic and non-myotoxic BthA-I (acidic PLA(2) from B. jararacussu) and other Asp49-PLA(2)s. BthTX-II structure showed a severe distortion of calcium-binding loop leading to displacement of the C-terminal region. Tyr28 side chain, present in this region, is in an opposite position in relation to the same residue in the catalytic activity Asp49-PLA(2)s, making a hydrogen bond with the atom 0 delta 2 of the catalytically active Asp49, which should coordinate the calcium. This high distortion may also be confirmed by the inability of BthTX-II to bind Na+ ions at the Ca2+-binding loop, despite of the crystallization to have occurred in the presence of this ion. In contrast, other Asp49-PLA(2)s which are able to bind Ca2+ ions are also able to bind Na+ ions at this loop. The comparison with other catalytic, non-catalytic and inhibited PLA(2)s indicates that the BthTX-II is not able to bind calcium ions; consequently, we suggest that its low catalytic function is based on an alternative way compared with other PLA(2)s. (c) 2008 Elsevier B.V All rights reserved.

Higher respiratory activity decreases mitochondrial reactive oxygen release and increases life span in Saccharomyces cerevisiae

Increased replicative longevity in Saccharomyces cerevisiae because of calorie restriction has been linked to enhanced mitochondrial respiratory activity. Here we have further investigated how mitochondrial respiration affects yeast life span. We found that calorie restriction by growth in low glucose increased respiration but decreased mitochondrial reactive oxygen species production relative to oxygen consumption. Calorie restriction also enhanced chronological life span. The beneficial effects of calorie restriction on mitochondrial respiration, reactive oxygen species release, and replicative and chronological life span could be mimicked by uncoupling agents such as dinitrophenol. Conversely, chronological life span decreased in cells treated with antimycin (which strongly increases mitochondrial reactive oxygen species generation) or in yeast mutants null for mitochondrial superoxide dismutase (which removes superoxide radicals) and for RTG2 (which participates in retrograde feedback signaling between mitochondria and the nucleus). These results suggest that yeast aging is linked to changes in mitochondrial metabolism and oxidative stress and that mild mitochondrial uncoupling can increase both chronological and replicative life span.

Using post-release monitoring data to optimize avian reintroduction programs: a 2-year case study from the Brazilian Atlantic Rainforest

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Modelling post-release survival of reintroduced Red-billed Curassows Crax blumenbachii

Modelling post-release survival probabilities of reintroduced birds can help inform 'soft-release' strategies for avian reintroductions that use captive-bred individuals. We used post-release radiotelemetry data to estimate the survival probabilities of reintroduced captive-bred Red-billed Curassow Crax blumenbachii, a globally threatened Cracid endemic to the Brazilian Atlantic Rainforest. Between August 2006 and December 2008, 46 radiotagged Curassows from the Crax Brazil breeding centre were reintroduced to the Guapiacu Ecological Reserve (REGUA), Rio de Janeiro state, Brazil, in seven different cohorts. Reintroduced birds were most vulnerable during the first 12 months post-release from natural predation, domestic dogs and hunting. Annual post-release survival probability was high (75%) compared with published estimates for other Galliform species. However, when considering survival in all birds transported to REGUA (some birds died before release or were retained in captivity) and not only post-release survival, phi in this study was closer to estimates for other species (60%). The duration of the pre-release acclimatization period within the soft-release enclosure and the size of the released cohorts both positively influenced post-release survival of reintroduced Curassows. Our results are relevant to future Cracid reintroductions and highlight the importance of utilizing post-release monitoring data for evidence-based improvements to soft-release strategies that can significantly enhance the post-release survival of captive-bred birds.

Refolding and purification of bothropstoxin-1, a Lys49 - Phospholipase A(2) homologue, expressed as inclusion bodies in Escherichia coli

Hydrolysis of phospholipids by Group II phospholipase A(2) enzymes involves a nucleophilic attack on the sn-2 ester bond by the His48 residue and stabilization of the reaction intermediate by a Ca2+ ion cofactor bound to the Asp49 residue in the protein active site region, Bothropstoxin-I (BthTX-I) is a PLA, variant present in the venom of the snake Bothrops jararacussu which shows a Asp49 to Lys substitution and which lacks hydrolytic activity yet damages artificial membranes by a noncatalytic Ca2+-independent mechanism. In order to better characterize this unusual mechanism of membrane damage, we have established an expression system for BthTX-I in Escherichia coli. The DNA-coding sequence for BthTX-I was subcloned into the vector pET11-d, and the BthTX-I was expressed as inclusion bodies in E, coli BL21(DE3). The native BthTX-I contains seven disulfide bonds, and a straightforward protocol has been developed to refold the recombinant protein at high protein concentration in the presence of surfactants using a size-exclusion chromatography matrix. After refolding, recovery yields of 2.5% (corresponding to 4-5 mg of refolded recombinant BthTX-I per liter of bacterial culture) were routinely obtained. After refolding, identical fluorescent and circular dichroism spectra were obtained for the recombinant BthTX-I compared to those of the native protein. Furthermore, the native and refolded recombinant protein demonstrated identical membrane-damaging properties as evaluated by measuring the release of an entrapped fluorescent marker from liposomes, (C) 2001 Academic Press.

Insights into metal ion binding in phospholipases A(2): ultra high-resolution crystal structures of an acidic phospholipase A(2) in the Ca2+ free and bound states

The electrophile Ca2+ is an essential multifunctional co-factor in the phospholipase A(2) mediated hydrolysis of phospholipids. Crystal structures of an acidic phospholipase A(2) from the venom of Bothrops jararacussu have been determined both in the Ca2+ free and bound states at 0.97 and 1.60 angstrom resolutions, respectively. In the Ca2+ bound state, the Ca2+ ion is penta-coordinated by a distorted pyramidal cage of oxygen and nitrogen atoms that is significantly different to that observed in structures of other Group I/II phospholipases A(2). In the absence of Ca2+, a water molecule occupies the position of the Ca2+ ion and the side chain of Asp49 and the calcium-binding loop adopts a different conformation. (c) 2005 Elsevier SAS. All rights reserved.

Competitive release and area effects

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)