Physicochemical characterization of experimental resin-based materials containing calcium orthophosphates or calcium silicate.

OBJECTIVE: To evaluate experimental dimethacrylate-based materials containing calcium orthophosphates or calcium silicate particles in terms of their optical, mechanical and Ca2+ release behaviour. METHODS: Dicalcium phosphate dihydrate (DCPD), hydroxyapatite (HAp), beta-tricalcium phosphate (ß-TCP) or calcium silicate (CaSi) particles were added to a photocurable BisGMA/TEGDMA resin (1:1 in mols) at a 30 vol% fraction. Materials containing silanized or non-silanized barium glass particles were used as controls. Degree of conversion (DC) at the top and base of 2-mm thick specimens was determined by ATR-FTIR spectroscopy (n = 5). Translucency parameter (TP) and transmittance (%T) were determined using a spectrophotometer (n = 3). Biaxial flexural strength (BFS) and flexural modulus (FM) were determined by biaxial flexural testing after 24 h storage in water (n = 10). Ca2+ release in water was determined during 28 days by inductively coupled plasma optical emission spectrometry (n = 3). Statistical analysis was performed using ANOVA/Tukey test (DC: two-way; TP, %T; BFS and FM: one-way; Ca2+ release: repeated measures two-way, α = 5 %). RESULTS: CaSi and ß-TCP particles drastically reduced DC at 2 mm, TP and %T (p < 0.001). Compared to both controls, all Ca2+-releasing materials presented lower BFS (p < 0.001) and only the material with DCPD showed significantly lower FM (p < 0.05). The material containing CaSi presented the highest Ca2+ release, while among materials formulated with calcium orthophosphates the use of DCPD resulted in the highest release (p < 0.001). SIGNIFICANCE: CaSi particles allowed the highest Ca2+ release. Notwithstanding, the use of DCPD resulted in a material with the best compromise between optical behaviour, DC, strength and Ca2+ release.

Effect of ultrasonic activation on setting time, pH and calcium ion release, solubility, and chemical structure of calcium silicate sealers.

This study evaluated the setting time, pH, calcium ion release, solubility, and chemical structure of four calcium silicate sealers after ultrasonic activation (UA). Five sealers were evaluated: Sealer Plus (SP - control); Sealer Plus BC (SPBC), Bio C Sealers (BCS), Endosequence BC Sealer (EBC), and BioRoot RCS (BR). Ten groups were created based on the use or not of ultrasonic activation: SP; SP/UA; SPBC; SPBC/UA; BCS; BCS/UA; EBC; EBC/UA; BR; and BR/UA. Setting time was performed based on ISO 6876:2012 and ASTM C266-07 specifications. Solubility at 24hs, based on ISO 6876:2012. pH and calcium release were evaluated at 1, 24, 72, and 168hs. Raman spectroscopy was used to evaluate structural changes. Quantitative data were analyzed using One-Way ANOVA and Tukey post-hoc test (α=5%). Raman spectroscopy results were qualitatively analyzed. Setting times and solubility of all sealers were not affected by UA (p>0.05). The highest solubility was found for BCS, BCS/UA; and BR, BR/UA (p<0.05). After 24hs, calcium silicate sealers had higher pH than SP and SP/UA (p<0.05). BR and BR/UA had the highest pH at all time points. SP and SP/UA had stable pH at all time points. SP and SP/UA had the lowest calcium release values at all time points (p<0.05). EBC and EBC/UA calcium release significantly differ at 24,72 and 168hs (p<0.05). No chemical changes were observed during Raman spectroscopy. In conclusion, ultrasonic activation affected calcium ion release only for EndoSequence BC Sealer. Ultrasonic activation did not influence the initial and final setting time, solubility, pH, and chemical structure of any investigated sealers.

Young bamboo flour as a substitute for emulsifying salts in requeijão cremoso processed cheese and the effect on the quality parameters.

The effect of the substitution of emulsifying salt by the young bamboo flour (BF) (0, 25, 50, 75, 100 % w/w) on requeijão cremoso processed cheese [REQ, REQ 25, REQ 75 REQ 100]) processing was investigated. Gross composition, calcium and sodium values, functional properties (melting rate), color parameters (L, a*, b*, C*, and Whiteness Index, WI), texture profile, fatty acid profile, volatile organic compounds (VOCs), and sensory profiling were evaluated. No effect was observed on the gross composition; however, sodium and melting rate values were decreased, and calcium values presented the opposite behavior. BF could modify the optical parameters, observing an increase in WI values. Higher BF addition increased hardness and lowered elasticity, and regarding the fatty acid profile, there is no significant difference. Different volatile compounds were noted in a proportional form with the BF addition, which was reflected in similar sensory acceptance for REQ 25 and control samples. Although some aspects require further in-depth studies, using BF as a substitute for emulsifying salt in requeijão cremoso processed cheese appears to be a viable option, especially when considering partial replacements.

Ion release mechanisms in composites containing CaP particles and hydrophilic monomers.

OBJECTIVE: To investigate the effect of hydrophilic/permeable polymer matrices on water sorption/solubility (WS/SL), Ca2+ release, mechanical properties and hydrolytic degradation of composites containing dicalcium phosphate dihydrate (DCPD) particles. METHODS: Six composites were tested, all with 10 vol% of glass particles and either 30 vol% or 40 vol% DCPD. Composites containing 1BisGMA:1TEGDMA in mols (at both inorganic levels) were considered controls. Four materials were formulated where 0.25 or 0.5 of the BisGMA/TEGDMA was replaced by pyromellitic dianhydride glycerol dimethacrylate (PMGDM)/ polyethylene glycol dimethacrylate (PEGDMA). Composites were tested for degree of conversion (FTIR spectroscopy), WS/SL (ISO 4049) and Ca2+ release (inductively coupled plasma optical emission spectroscopy). Fracture toughness (FT) and biaxial flexural strength/modulus (BFS/FM) were determined after 24 h and 60 days in water. The contributions of diffusional and relaxational mechanisms to Ca2+ release kinetics were analyzed using the semi-empirical Salim-Peppas model. Data were analysed by ANOVA/Tukey test (alpha: 0.05). RESULTS: WS/SL was higher for composites containing PMGDM/PEGDMA compared to the controls (p < 0.001). Only at 40% DCPD the 0.5 PMGDM/PEGDMA composite showed statistically higher Ca2+ release than the control. Relaxation diffusion was the main release mechanism. Initial FT was not negatively affected by matrix composition. BFS (both DCPD fractions) and FM (30% DCPD) were lower for composites with hydrophilic/permeable networks (p < 0.01). After 60 days in water, composites with PMGDM/PEGDMA presented significant reductions in FT, while all composites had reductions in BFS/FM. SIGNIFICANCE: Increasing matrix hydrophilicity/permeability significantly increased Ca2+ release only at a high DCPD fraction.

Calcium complexation by steroids involved in the steroidogenesis.

Steroids that take part in the pathways of human steroidogenesis are involved in many biological mechanisms where they interact with calcium. In the present work, the binding selectivities and affinities for calcium of progestagens, mineralocorticoids, androstagens, and estrogens were studied by Electrospray Ionization-Mass Spectrometry (ESI-MS). The adduct profile of each steroid was characterized by high resolution and tandem mass spectrometry. The relative stability of the most important adducts was studied by threshold collision induced dissociation, E1/2. Doubly-charged steroid-calcium complexes [nM + Ca]2+ with n = 1-6 were predominant in the mass spectra. The adduct [5M + Ca]2+ was the base peak for most 3-keto-steroids, while ligands bearing hindered ketones or α-hydroxy-ketones also yielded [nM + Ca + mH2O]2+ with n = 3-4 and m = 0-1. Principal component analysis allowed us to spot the main differences and similarities in the binding behavior of these steroids. The isomers testosterone and dehydroepiandrosterone, androstanolone and epiandrosterone, and 17-α-hydroxyprogesterone and 11-deoxycorticosterone showed remarkable differences in their adduct profiles. Computational modeling of representative adducts was performed by density functional theory methods. The possible binding modes at low and high numbers of steroid ligands were determined by calcium Gas Phase Affinity, and through modeling of the complexes and comparison of their relative stabilities, in agreement with the experimental results.

Physicochemical properties of calcium silicate cement based endodontic sealers.

AIM: To mensure the physicochemical properties of three ceramic cement endodontic sealers AH Plus Bioceramic, Bio-C Sealer and Bio-C Sealer Ion+ with an epoxy resin sealer, AH Plus. MATERIAL AND METHODS: These properties were measured: hardening time (HT), dimensional change (DC), solubility (SL), flow (FL) and radiopacity (RD). The distilled water obtained from the SL test was analyzed with atomic absorption spectrometry. A sample calculation was made considering n = 5 repetitions for each experimental sealer evaluated. Statistical analysis was performed using one-way ANOVA and post hoc Tukey tests (p < 0.05). RESULTS: For the HT, AH Plus (484 ± 2.76 min) and AH Plus Bioceramic (424 ± 1.23 min) set more slowly than of Bio-C Sealer (370 ± 4.50 min) and Bio-C Sealer Ion+ (380 ± 1.42 min) (p < 0.05). AH Plus Bioceramic (12.56 ± 2.71 %) was more soluble than Bio-C Sealer (6.69 ± 1.67 %), Bio-C Sealer Ion+ (5.67 ± 2.16 %) and AH Plus (0.15 ± 0.01 %) (p < 0.05). AH Plus (0.03 ± 0.01 %) had slight expansion while the cement-based sealers had shrinkage: AH Plus Bioceramic (-1.60 ± 0.63 %) and Bio-C Sealer (-1.38 ± 0.69 %), Bio-C Sealer Ion+ (-5.19 ± 1.23 %) (p < 0.05). Bio-C Sealer Ion+ (59.80 ± 0.86 mm) and Bio-C Sealer (58.60 ± 0.98 mm) had the highest flow compared with AH Plus (56.90 ± 0.56 mm) and AH Plus Bioceramic (49.50 ± 0.63 mm) (p < 0.05). AH Plus (9.17 ± 0.06 mmAl) and AH Plus Bioceramic (8.27 ± 0.84 mmAl) showed radiopacity values when compared with those of Bio-C Sealer (4.90 ± 0.08 mmAl) and Bio-C Sealer Ion+ (4.14 ± 0.05 mmAl) (p > 0.05). CONCLUSION: Ion release is inhered to these cement-based sealers and this result in calcium ion release.

Optimization of enzymatic saccharification of industrial wastes using a thermostable and halotolerant endoglucanase through Box-Behnken experimental design.

This study describes the production, characterization and application of an endoglucanase from Penicillium roqueforti using lignocellulosic agro-industrial wastes as the substrate during solid-state fermentation. The endoglucanase was generated after culturing with different agro-industrial wastes for 96 h without any pretreatment. The highest activity was obtained at 50 °C and pH 4.0. Additionally, the enzyme showed stability in the temperature and pH ranges of 40-80 °C and 4.0-5.0, respectively. The addition of Ca2+, Zn2+, Mg2+, and Cu2+ increased enzymatic activity. Halotolerance as a characteristic of the enzyme was confirmed when its activity increased by 35% on addition of 2 M NaCl. The endoglucanase saccharified sugarcane bagasse, coconut shell, wheat bran, cocoa fruit shell, and cocoa seed husk. The Box-Behnken design was employed to optimize fermentable sugar production by evaluating the following parameters: time, substrate, and enzyme concentration. Under ideal conditions, 253.19 mg/g of fermentable sugars were obtained following the saccharification of wheat bran, which is 41.5 times higher than that obtained without optimizing. This study presents a thermostable, halotolerant endoglucanase that is resistant to metal ions and organic solvents with the potential to be applied in producing fermentable sugars for manufacturing biofuels from agro-industrial wastes.

Direct Determination of Ca, K, and Mg in Soy Leaf Samples Using Laser-Induced Breakdown Spectroscopy.

This study was dedicated to developing analytical methods for determining macronutrients (Ca, K, and Mg) in soy leaf samples with and without petioles. The study's primary purpose was to present Laser-induced breakdown spectroscopy (LIBS) as a viable alternative for directly analyzing leaf samples using chemometric tools to interpret the data obtained. The instrumental condition chosen for LIBS was 70 mJ of laser pulse energy, 1.0 µs of delay time, and 100 µm of spot size, which was applied to 896 samples: 305 of soy without petioles and 591 of soy with petioles. The reference values of the analytes for the proposition of calibration models were obtained using inductively coupled plasma optical emission spectroscopy (ICP-OES) technique. Twelve normalization modes and two calibration strategies were tested to minimize signal variations and sample matrix microheterogeneity. The following were studied: multivariate calibration using partial least squares and univariate calibration using the area and height of several selected emission lines. The notable normalization mode for most models was the Euclidean norm. No analyte showed promising results for univariate calibrations. Micronutrients, P and S, were also tested, and no multivariate models presented satisfactory results. The models obtained for Ca, K, and Mg showed good results. The standard error of calibration ranged from 2.3 g/kg for Ca in soy leaves without petioles with two latent variables to 5.0 g/kg for K in soy leaves with petioles with two latent variables.

Assessment of calcium alginate gels as wall materials for encapsulation systems.

BACKGROUND: Calcium alginate gels are widely used to encapsulate active compounds. Some characteristic parameters of these gels are necessary to describe the release of active compounds through mechanistic mathematical models. In this work, transport and kinetics properties of calcium alginate gels were determined through simple experimental techniques. RESULTS: The weight-average molecular weight ( M ¯ w = 192 × 103 Da) and the fraction of residues of α-l-guluronic acid ( F G = 0.356) of sodium alginate were determined by capillary viscometry and 1 H-nuclear magnetic resonance at 25 °C, respectively. Considering the half egg-box model, both values were used to estimate the molecular weight of calcium alginate as M g = 2.02 × 105 Da. An effective diffusion coefficient of water ( D eff , w = 2.256 × 10-9 m2 s-1 ) in calcium alginate was determined using a diffusion cell at 37 °C. Finally, a kinetics constant of depolymerization ( k m = 9.72 × 10-9 m3 mol-1 s-1 ) of calcium alginate was obtained considering dissolution of calcium to a medium under intestinal conditions. CONCLUSION: The experimental techniques used are simple and easily reproducible. The obtained values may be useful in the design, production, and optimization of the alginate-based delivery systems that require specific release kinetics of the encapsulated active compounds. © 2023 Society of Chemical Industry.

Effects of pharmaceuticals compounds and calcium on granulation, microbiology, and performance of anaerobic granular sludge systems.

Granular sludge is a promising biotechnology to treat sewage contaminated with pharmaceuticals due to its increased toxicity resistance. In this context, this study evaluated the potential of Ca2+ as a granulation precursor and how pharmaceutical compounds (loratadine, prednisone, fluconazole, fenofibrate, betamethasone, 17α-ethinyl estradiol, and ketoprofen) affect granulation. Continuous and intermittent dosages of Ca2+ in the presence and absence of pharmaceuticals were evaluated. The results showed that intermittent addition of Ca2+ reduces the time for anaerobic sludge granulation, and pharmaceuticals presence did not impair granulation. 10% of the granules presented mean diameters greater than 2.11 mm within 93 days with intermittent Ca2+ dosage in the pharmaceuticals' presence. In contrast, no granules higher than 2.0 mm were observed with no precursor addition. The pharmaceuticals' toxicity may have created a stress condition for the microbial community, contributing to more EPS production and a greater potential for granulation. It was also verified that pharmaceuticals' presence did not decrease organic matter, total alkalinity, and volatile fatty acids removals. The 16S rRNA gene analysis revealed taxa resistance to recalcitrant compounds when pharmaceuticals were added. Besides, the efficiency of a granular sludge bioreactor (EGSB) was evaluated for pharmaceuticals removal, and betamethasone, fenofibrate, and prednisone were effectively removed.

Use of Computerized Microtomography, Energy Dispersive Spectroscopy, Scanning Electron Microscopy, and Atomic Force Microscopy to Monitor Effects of Adding Calcium to Bleaching Gels.

OBJECTIVES: The aim of this study was to evaluate the mineral content, expressed by calcium (Ca) and phosphate (P), in dental enamel exposed to bleaching agents using micro-computed tomography (micro-CT), scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), and atomic force microscopy (AFM). METHODS: Sixty bovine dental enamel specimens were randomly divided into three groups (n=20): HP35ca (bleached using 35% hydrogen peroxide with Ca); HP35wca (bleached using 35% hydrogen peroxide without Ca); and control (without bleaching). Five specimens from each group were used for SEM and EDS analyses, 10 specimens were used for AFM analysis, and the remaining five specimens were used for micro-CT analysis. The pH of the gels was measured using a pH meter. The EDS and micro-CT data were analyzed using one-way ANOVA and Pearson's correlation test. The AFM data were analyzed using one-way ANOVA (α=0.05). RESULTS: The weight percentages of Ca and P obtained using EDS were similar between the bleached and control groups. Small, superficial changes were observed by SEM in the HP35wca group. The HP35ca group showed similar patterns to the control group. AFM results showed no significant changes in the enamel roughness in any of the tested groups. No significant difference in the volume or depth of structural enamel loss was found between gels with and without Ca. No mineral loss was observed in the dentin substrate. The EDS and micro-CT analysis data exhibited a high correlation (p<0.001). CONCLUSION: The addition of Ca to the bleaching gel had no beneficial effect on the bleached tooth enamel in terms of composition, mineral loss, and surface roughness. Micro-CT results exhibited a high correlation with the EDS results.

Mechanical properties of gellan gum beads prepared with potassium or calcium ions.

Biopolymer beads can be used as carrier and encapsulation system for a wide variety of materials in food, medical, pharmaceutical, cosmetics, agricultural, and environmental applications. Beads of low acyl gellan gum (0.4-1.2% w/w) were produced using extrusion technique (dripping) followed by an ionotropic gelation step with calcium or potassium chloride. In this methodology, gel formation is accomplished by cations diffusion at room temperature and, as a consequence, different structure and gel properties could be obtained. Gellan beads were subjected to uniaxial compression measurements. The force-displacement curves showed that the occurrence of structural failure under tested conditions depended on beads formulation and was only observed at polysaccharide concentration above 0.8% (w/w). Maximum force or force at failure was mainly dependent on the type (monovalent or divalent cation) and salt concentration. Moreover, at fixed salt amounts, higher values of maximum force were reached using a concentration of 1% (w/w) gellan. Young modulus, determined using Hertz approach, showed values between 445 and 840 kPa depending on polysaccharide concentration and salt type added. Mechanical properties are critical features of gel beads and can define their suitability for a specific application. Therefore, the results obtained, mainly intrinsic properties such as Young modulus, could be a tool for comparing and choosing polysaccharides for specific uses.

Computational Analyses of the AtTPC1 (Arabidopsis Two-Pore Channel 1) Permeation Pathway.

Two Pore Channels (TPCs) are cation-selective voltage- and ligand-gated ion channels in membranes of intracellular organelles of eukaryotic cells. In plants, the TPC1 subtype forms the slowly activating vacuolar (SV) channel, the most dominant ion channel in the vacuolar membrane. Controversial reports about the permeability properties of plant SV channels fueled speculations about the physiological roles of this channel type. TPC1 is thought to have high Ca2+ permeability, a conclusion derived from relative permeability analyses using the Goldman-Hodgkin-Katz (GHK) equation. Here, we investigated in computational analyses the properties of the permeation pathway of TPC1 from Arabidopsis thaliana. Using the crystal structure of AtTPC1, protein modeling, molecular dynamics (MD) simulations, and free energy calculations, we identified a free energy minimum for Ca2+, but not for K+, at the luminal side next to the selectivity filter. Residues D269 and E637 coordinate in particular Ca2+ as demonstrated in in silico mutagenesis experiments. Such a Ca2+-specific coordination site in the pore explains contradicting data for the relative Ca2+/K+ permeability and strongly suggests that the Ca2+ permeability of SV channels is largely overestimated from relative permeability analyses. This conclusion was further supported by in silico electrophysiological studies showing a remarkable permeation of K+ but not Ca2+ through the open channel.

Biochemical characterization of LysVpKK5 endolysin from a marine vibriophage.

Endolysins have been proposed as a potential antibacterial alternative for aquaculture, especially against Vibrio; the bacterial-agents that most frequently cause disease. Although multiple marine vibriophages have been characterized to date, research on vibriophage endolysins is recent. In this study, biochemical characterization of LysVpKK5 endolysin encoded by Vibrio parahaemolyticus-infecting VpKK5 phage was performed. In silico analysis revealed that LysVpKK5 possesses a conserved amidase_2 domain with a zinc-binding motif of high structural similarity to T7 lysozyme (RMSD = 0.107 Å). Contrary to expectations, the activity was inhibited with Zn2+ and was improved with other divalent cations, especially Ca2+. It showed optimal muralytic activity at pH 10, and curiously, no lytic activity at pH ≤ 7 was recorded. As for the thermal stability test, the optimal activity was recorded at 30 °C; the higher residual activity was recorded at 4 °C, and was lost at ≥ 50 °C. On the other hand, increasing NaCl concentrations reduced the activity gradually; the optimal activity was recorded at 50 mM NaCl. On the other hand, the enzymatic activity at 0.5 M NaCl was approx 30% and of approx 50% in seawater. LysVpKK5 endolysin exhibited a higher activity on V. parahaemolyticus ATCC-17802 strain, in comparison with AHPND + strains.

CaMn3IV O4 Cubane Models of the Oxygen-Evolving Complex: Spin Ground States S<9/2 and the Effect of Oxo Protonation.

We report the single crystal XRD and MicroED structure, magnetic susceptibility, and EPR data of a series of CaMn3IV O4 and YMn3IV O4 complexes as structural and spectroscopic models of the cuboidal subunit of the oxygen-evolving complex (OEC). The effect of changes in heterometal identity, cluster geometry, and bridging oxo protonation on the spin-state structure was investigated. In contrast to previous computational models, we show that the spin ground state of CaMn3IV O4 complexes and variants with protonated oxo moieties need not be S=9/2. Desymmetrization of the pseudo-C3 -symmetric Ca(Y)Mn3IV O4 core leads to a lower S=5/2 spin ground state. The magnitude of the magnetic exchange coupling is attenuated upon oxo protonation, and an S=3/2 spin ground state is observed in CaMn3IV O3 (OH). Our studies complement the observation that the interconversion between the low-spin and high-spin forms of the S2 state is pH-dependent, suggesting that the (de)protonation of bridging or terminal oxygen atoms in the OEC may be connected to spin-state changes.

A novel method for the in vitro assembly of virus-like particles and multimeric proteins.

OBJECTIVE: To develop a method for the efficient assembly of viral or multimeric proteins into virus-like particles (VLP) or other macro structures. RESULTS: Protein monomers were assembled by eliminating calcium ions through precipitation. The model protein, rotavirus VP6, assembled into stable, long nanotubes with better quality than the assemblies obtained directly from cell culture. Nanotube length was directly proportional to the initial concentration of VP6 monomers, in accordance with the classic nucleation theory of capsid assembly. The quality of the obtained assemblies was confirmed when the nanotubes were functionalized with metals, yielding unique nanobiomaterials. Assembly efficiency was improved in comparison with other previously proposed methods. CONCLUSIONS: The novel method presented here is simpler and faster than other reported methods for the assembly and disassembly of viral proteins, a step needed for most applications.

Improving micropropagation of some grape cultivars via boron, calcium and phosphate

BACKGROUND: An efficient regeneration protocol is a priority for the successful application of plant biotechnology. Grape nodal explants were used to develop a micropropagation protocol for Thompson Seedless and Taify cvs. Explants were cultured on MS medium supplemented with Kinetin or benzylaminopurine (BA) and indolebutyric acid (IBA). RESULTS: For both cultivars, axillary buds were grown, only, on a medium enriched with kinetin, moreover, shoot tip necrosis and callus formation were observed on Thompson Seedless cv. cultures grown on a medium with BA. Supplementing the growth medium with 100 mM (boron) B and 2.5 mM (calcium) Ca successfully help overcome these phenomena. The highest regenerated shoot numbers (14 and 6.2 explant 1 ) for Taify and Thompson Seedless cvs., respectively, were on media supplemented with 13.2 mM BA + 4.9 mM IBA and BA 13.2 mM + 5.8 mM IBA, respectively. Moreover, these media supported the developing shoots to have the heaviest dry weights (1.46 and 0.72 mg explant 1 ) for Taify and Thompson Seedless cvs., respectively. Thompson Seedless cv. regenerated shoot numbers and their dry weights were significantly increased by increasing the MS medium PO4 concentration. However, these two parameters were significantly decreased for Taify cv. Developing shoots were elongated and rooted on MS medium enriched with 4.9 mM, IBA 100 mM B and 2.5 mM Ca. Plantlets were acclimatized and successfully transferred to the greenhouse conditions. CONCLUSIONS: A novel promising protocol for Thomson Seedless and Taify cvs. micropropagation using single nodes has been developed.

Biochemical, pharmacological and structural characterization of BmooMP-I, a new P-I metalloproteinase from Bothrops moojeni venom.

Snakebite envenoming is still a worrying health problem in countries under development, being recognized as a neglected disease by the World Health Organization. In Latin America, snakes from the genus Bothrops are widely spread and in Brazil, the Bothrops moojeni is a medically important species. The pharmacological effects of bothropic snake venoms include pain, blisters, bleeding, necrosis and even amputation of the affected limb. Snake venom metalloproteinases are enzymes abundantly present in venom from Bothrops snakes. These enzymes can cause hemorrhagic effects and lead to myonecrosis due to ischemia. Here, we present BmooMP-I, a new P-I class of metalloproteinase (this class only has the catalytic domain in the mature form) isolated from B. moojeni venom. This protein is able to express fibrinogenolytic and gelatinase activities, which play important roles in the prey's immobilization and digestion, and also induces weak hemorrhagic effect. The primary sequence assignment was done by a novel method, SEQUENCE SLIDER, which combines crystallographic, bioinformatics and mass spectrometry data. The high-resolution crystal structure reveals the monomeric assembly and the conserved metal binding site H141ExxH145xxG148xxH151 with the natural substitution Gly148Asp that does not interfere in the zinc coordination. The presence of a structural calcium ion on the surface of the protein, which can play an important role in the stabilization of hemorrhagic toxins, was observed in the BmooMP-I structure. Due to the relevant local and systemic effects of snake venom metalloproteinases, studies involving these proteins help to better understand the pathological effects of snakebite envenoming.

Spittlebug damage on tropical grass and its impact in pasture-based beef production systems.

Spittlebugs are the main pest of tropical pastures and Marandu palisade grass (Urochloa brizantha cv. Marandu) is the most representative cultivated pasture in the tropics. Our objective was to characterize Marandu palisade grass responses subjected to Mahanarva (Hemiptera: Cercopidae) attack and to estimate the losses in terms of beef production from pasture-based systems. A set of five experiments were carried out. Three consecutive years of monitoring showed that Mahanarva spittlebugs increased their abundance after first rains with three to four peaks throughout the wet season. A decrease of 66% on herbage yield was observed in the greenhouse trial, with an average decrease of 61% on pools of calcium, magnesium, phosphorus, sulfur, potassium, crude protein, neutral-detergent fiber and in vitro digestible dry matter of Marandu palisade grass. Results from field experiments corroborated with greenhouse trial showing decreases on herbage yield varying from 31 to 43% depending on level of fertilization and grazing severity of Marandu palisade grass. Finally, an unprecedented 154-ha field experiment indicated that Mahanarva decreases 74% the beef productivity (i.e. kg body weight ha-1) of Nellore heifers grazing Marandu palisade grass.