Alkaline pH Range Research Articles

Partial purification and characterization of the quality deteriorating enzymes from Indian pink guava (Psidium guajava L.), var. Lalit.

Peroxidase (POD), polyphenol oxidase (PPO), ascorbate oxidase (AO) and pectin methylesterase (PME) from 'Lalit' guava variety have been purified using BioGel P-100 and UNOsphere Q resins. POD, PPO, AO, and PME were partially purified to 28, 30, 36 and 30 fold; single band on SDS-PAGE represented a molecular weight of 21, 70, 75, 50kDa, respectively. Optimum pH for POD and AO was 6.5 whereas, PPO and PME showed optimum pH in the alkaline range of pH 8.0 and 8.5, respectively. AO showed optimum activity at 25°C, POD and PPO showed optimum activity at 35°C; however, PME showed maximum activity at 55°C. AO was the most heat labile whereas, PME was the most heat stable enzyme studied and may carry an important role in the thermal processing of guava pulp as an indicator of the efficacy of treatment.

Back Cover: Optimized two‐dimensional gel electrophoresis in an alkaline pH range improves the identification of intracellular CFDA‐cisplatin‐protein adducts in ovarian cancer cells

DOI: 10.1002/elps.201700377 The cover picture shows the visualization of CFDA-cisplatin-protein adducts through 2DE. Human ovarian carcinoma cell lines A2780 and A2780cis cells were treated with CFDA-cisplatin and separated by 2DE. The use of fluorescent CFDA-cisplatin enables the visualization of CFDA-cisplatin-protein adducts. For the image analysis the fluorescence scans from CFDA-cisplatin-protein adducts and the Coomassie staining were fused into one image. The identification of nine previously unknown intracellular binding partners of CFDA-cisplatin by mass spectrometry provides new insights into the cellular processing of cisplatin.

Optimized two-dimensional gel electrophoresis in an alkaline pH range improves the identification of intracellular CFDA-cisplatin-protein adducts in ovarian cancer cells.

Intracellular binding of cisplatin to proteins has been associated with acquired resistance to chemotherapy. In our previous study we established an analytical method for the identification of intracellular cisplatin-binding proteins. The method used a fluorescent carboxyfluorescein-diacetate-labeled cisplatin analogue (CFDA-cisplatin), two-dimensional gel electrophoresis (2DE) and mass spectrometry, which allows detecting and identifying intracellular CFDA-cisplatin-containing protein adducts in the acidic pH range (pH 4-7). Based on this analytical method we extended the identification of intracellular cisplatin-protein adducts to the alkaline pH range (pH 6-10) giving chance to discover new important binding partners. 2DE analysis of alkaline proteins is challenging due to the difficult separation of basic proteins during the isoelectric focusing (IEF). The establishment of an optimized IEF protocol for basic proteins enabled us to identify several intracellular CFDA-cisplatin-binding proteins including enzymes of the glucose and serine metabolism like alpha enolase and D-3-phosphoglycerate 1-dehydrogenase.

A chemical/mineralogical investigation of the behavior of sulfoaluminate binders submitted to accelerated carbonation

The paper addresses the carbonation of CSA based binders from the mechanical, chemical and mineralogical point of view. Three different binders have been investigated, revealing that many hydrated phases can protect ettringite against carbonation. In particular, the presence of AFm phases gives a significant contribution to buffer the pH in alkaline range and to preserve the protective film of steel reinforcement bars. In a carbonated environment AFm phases act as ionic exchangers: carbonate ions replace sulfates in the interlayer and in these conditions the released sulfate ions contribute to the formation of new ettringite, with a beneficial effect on compressive strength performances. Experimental data are compared with models and carbonation mechanisms are described.

Electrochemical Investigation of Stainless Steel Cladded Carbon Steel Manufactured Via Powder Bed Selective Laser Melting

This study aims to investigate the electrochemical and corrosion properties of 316L-stainless steel cladded carbon steel manufactured using powder bed the selective laser melting (PB-SLM) technique in various environments. The PB-SLM technique involves localized melting of a thin layer of 316L-SS powder bed with Yb - fiber laser. Main parameters of the PB-SLM process such as particle size, particle size distribution, laser power, laser-scanning speed and layer thickness were selected to optimize the density of the cladded layer and the quality of metallurgical bond to the substrate. Using the optimized process parameters, carbon steel substrates were cladded with stainless steel of various thicknesses. Several mechanical and microscopic tests (viz. nano-indentation, X-ray diffraction, Scanning electron microscopy) were performed to characterize both the bulk properties of the stainless steel layer and the interface between the cladding and the substrate. Electrochemical and corrosion properties were tested via. open-circuit potential monitoring, electrochemical impedance spectroscopy (EIS) and cyclic polarization (CP) in electrolytes with acidic to alkaline pH range and with increasing levels of chloride, simulating different application conditions. Scanning vibrating electrode technique was employed to study the galvanic corrosion properties of the stainless steel and carbon steel at the clad-substrate interface.

Co-complexation effects during incineration bottom ash leaching via comparison of measurements and geochemical modeling

Abstract Under the proper setting of boundaries, leaching behaviors of incineration bottom ashes (IBA) may be predicted using geochemical models. The extent of agreement between measurements and the prediction is often affected by the model development. On the other hand, the inconsistency may also be caused by the complexity of the system for modeling. As such, discrepancies between measurements and geochemical modeling on IBA leaching would indicate uncovered yet significant mechanisms which may not be fully understood by the existing setting scenarios. pH-static leaching tests of IBA were performed, followed by simulations with two geochemical models. Simulations based on the surface area calculated with all leachable minerals provided a better matching with experimental data as compared to that calculated with surface area of Fe/Al oxides only. Yet, certain elements remained highly overestimated (up to 3 orders of magnitude, e.g. Cd, Ni, Sr, Zn, Fe, Al and Mg) or underestimated (e.g. Cr) in alkaline pH range. Significant co-complexation during leaching may be susceptible and responsible for the differences. Multivariate factor analysis testified that pH, Fe and Al in acidic environment tended to be clustered and highly loaded under the dominant correlation factor (accounting for 33.3–49.0% of variance). Contrastively, pH, Ca and cationic strength in alkaline environment tended to be clustered and highly loaded under the dominant correlation factor (accounting for 39.9–43.3% of variance). In both cases, co-complexation was evidenced. As the natural pHs of IBA usually fall within the high alkaline range, the comparison study offered quantitative discrimination of leaching potential between measurements and simulation, which would be crucial for decision making during IBA applications. In light of limited leaching potential of IBA under the alkaline pH range, its application with cement (with high alkalinity) seems to be one of the best options.

A highly efficient magnetic chitosan “fluid” adsorbent with a high capacity and fast adsorption kinetics for dyeing wastewater purification

A novel magnetic fluid adsorbent (magnetic chitosan “fluid”, MCF) was prepared via a one-step in situ co-precipitation procedure using inexpensive and environmentally friendly chitosan (CS) and iron precursor. The resultant ultrafine colloid was characterized using transmission electron microscopy (TEM), X-ray powder diffraction (XRD), thermogravimetric analysis (TGA), and Fourier transform infrared spectroscopy (FTIR). The sorption and desorption characteristics of the composite were studied using a common Congo Red (CR) anionic dye to evaluate the suitability of the MCF for anionic dye wastewater purification. As an important advantage, the dye adsorption could be achieved in a wide alkaline pH range and reached the equilibrium within 5 min demonstrating the superior properties of our material compared the previously reported CS-based adsorbents. The adsorption obeyed the Langmuir isotherm model with the remarkable maximum adsorption capacity of approx. 1700 mg/g for CR under neutral pH. NaOH treatment was utilized to desorb and regenerate MCF, while the removal rate still remained above 90% after recycling five times, demonstrating that the MCF could be employed for repeated use without significant capacity loss. MCF could be readily separated from water within 30 s under a low magnetic field (<0.035 T). Overall, the ultrafine colloidal state of the synthesized MCF enabled a rapid pollutant uptake, high adsorption capacity, and reusability. The above characteristics of MCF and its environmentally friendly composition make it a very promising material for practical dyeing wastewater purification.

Use of ZnSO4 and SDD mixture as sphalerite depressant in copper flotation

The depression performance and mechanism of the zinc sulfate (ZnSO4) and sodium dimethyl dithiocarbamate (SDD) combined depressant with butyl xanthate (BX) as collector in a mixed Cu-Zn sulfide floation system were fully investigated using micro-flotation experiments, zeta-potential determination, and local electrochemical impedance spectroscopy (LEIS) analysis. The micro-flotation experimental results indicated that the combined depressant has a significant selective depression effect on sphalerite but has slight effect on chalcopyrite. The depression performance of the combined depressant strictly depends on the dosage of ZnSO4 + SDD and flotation pH. However, excessive SDD is detrimental to chalcopyrite flotation. The optimum conditions for flotation separation of chalcopyrite from the Cu-Zn sulfide were as follows: combined depressant dosage of 10−4 mol/L (ZnSO4: SDD = 3:1), BX dosage of 10−5 mol/L, and flotation pH of 10. Under such conditions, a copper concentrate assaying 30.21% Cu with 86.79% recovery was achieved from the Cu-Zn sulfide flotation system, and this concentrate had a very low Zn grade of 4.20% with 5.48% recovery. Both the zeta-potential and LEIS results confirmed that the presence of the combined depressant can significantly prevent the adsorption of BX onto the sphalerite surface, especially for an alkaline pH range of 9–12; however, the adsorption of BX onto the chalcopyrite surface is not affected. The LEIS provides a powerful tool for observing the adsorption of flotation reagents on mineral surface.

Molecular structure and catalytic mechanism of fungal family G acidophilic xylanases.

Industrial applications of xylanases have made this enzyme an important subject of applied research work. Function of this particular enzyme is to degrade or hydrolyze the plentiful polysaccharide xylan, an important component of hemicellulose. It mainly cleaves the backbone of xylan that is made up of a number of xylose residues connected with β-1,4-glycosidic linkages. Fungi with mycelia are regarded as the best producer of xylanases. These varied xylanases not only differ in their sizes and shapes but also differ in their physicochemical properties. Depending on the optimum pH in which they work best, they have been classified into (1) acidophilic xylanases active at low pH or acidic pH range, (2) alkaliphilic xylanases that are active at high or alkaline pH range and (3) neutral xylanases having pH optima in the neutral range between pH 5 and 7. Other researchers have classified the xylanases also on the basis of their structural properties, kinetic parameters, etc. This review discusses the molecular structures of some acidophilic xylanases and the molecular basis of low pH optima observed for their activities. It also discusses their unique catalytic mechanism and actual role of the catalytic residues found in them. Apart from these, the review also discusses different applications of these acidophilic xylanases in different industries. The article concludes with brief suggestions about how these acidophilic xylanases can be created employing the techniques of genetic engineering and concepts of synthetic evolution, using the traits of the known acidophilic xylanases discussed in the review.

Immobilization of a Mesophilic Lipase on Graphene Oxide: Stability, Activity, and Reusability Insights.

Catalyst-mediated bioprocessing at an industrial scale is dependent on the sustained stability and activity of the biocatalyst. Here, we demonstrate that strategy employed to increase the stability and activity of a mesophilic lipase immobilized on graphene oxide (GO). The protein adsorption capacity of GO is much higher than of other large surface area carbonaceous materials. Its structure and physicochemical properties are reported beneficial for enzymatic activity modifications. A purified lipase from Brevibacillus borstelensis NLIP05 immobilized on GO showed remarkable increase in thermostability (at 95°C) over a broad alkaline pH range (pH 7-12) compared to the free enzyme. Thermodynamic analysis of the GO-lip showed decreases in Km and activation energy (Ea) with increased Vmax and deactivation energy (Ed) at both 45 and 95°C. The decrease in decay constant (k) coupled with the increase in t1/2 value with temperature increase were salient features of the GO-lip system. This implies that GO-based immobilization conferred structural stability to the enzyme at higher temperature indicating chaperone like activity. Our findings support suitability of immobilized lipase on GO nanosupport for possible production of value-added materials with varied biological applications in the form of active pharmaceutical substances, synthetic building blocks, and effective synthesis of surfactants. The enhanced stability and activity of lipase indicate major application of GO on an industrial scale.

Simultaneous control of cadmium release and acidic pH neutralization in excavated sedimentary rock with concurrent oxidation of pyrite using steel slag

The aim of this study was to determine whether steel slag potentially could be utilized as an amendment for the reuse of excavated sedimentary rock naturally containing pyrite and the toxic metal cadmium. For this purpose, this study elucidated the mechanisms for simultaneous pH neutralization and cadmium immobilization of excavated sedimentary rock using steel slag under conditions of concurrent pyrite oxidation. A batch leaching test was conducted using excavated sedimentary rock (mudstone) that was well mixed with steel slag in combinations ranging from 0 to 50 %wt to evaluate the relationship between the dosage of steel slag and both cadmium release and the pH value after 30 days of the treated rock. Another batch leaching test was performed using excavated sedimentary rock combined with steel slag from 0 to 1.5 %wt to monitor cadmium release and the pH as pyrite oxidation proceeded. Cadmium was sorbed onto the steel slag to evaluate cadmium sorption ability at pH values of 6.5 and 9.0. Using the cadmium-sorbed steel slag, the crystalline structure and the level of cadmium release from the steel slag at different pH values were determined. The pH of excavated sedimentary rock without steel slag decreased, while that with more than 0.5 %wt steel slag remained within a neutral to alkaline pH range after 90 days, when potential pyrite oxidation was complete. The presence of steel slag did not inhibit the process of pyrite oxidation. These findings demonstrate that both pyrite oxidation and pH neutralization independently occurred in the excavated sedimentary rock, even in the presence of steel slag. The addition of more than 0.5 %wt steel slag maintained the level of cadmium released at less than 0.1 mg kg−1 throughout the 90-day period. The cadmium sorption test and X-ray diffraction analysis showed that cadmium immobilization using the steel slag was mainly governed by the (co)precipitation of carbonate minerals. This study indicates that an addition of 0.5 %wt steel slag to the excavated sedimentary rock can achieve pH neutralization and suppress cadmium release simultaneously, even with the occurrence of pyrite oxidation. Thus, this study suggests that steel slag potentially could be utilized as an amendment to allow the reuse of excavated sedimentary rock.

A Novel Cold Active Esterase from a Deep Sea Sponge Stelletta normani Metagenomic Library

Esterases catalyze the hydrolysis of ester bonds in fatty acid esters with short-chain acyl groups. Due to the widespread applications of lipolytic enzymes in various industrial applications, there continues to be an interest in novel esterases with unique properties. Marine ecosystem has long been acknowledged as a significant reservoir of microbial biodiversity and in particular of bacterial enzymes with desirable characteristics for industrial use, such as for example cold adaptation and activity in the alkaline pH range. We employed a functional metagenomic approach to exploit the enzymatic potential of one particular marine ecosystem, namely the microbiome of the deep sea sponge Stelletta normani. Screening of a metagenomics library from this sponge resulted in the identification of a number of lipolytic active clones. One of these encoded a highly, cold-active esterase 7N9, and the recombinant esterase was subsequently heterologously expressed in Escherichia coli. The esterase was classified as a type IV lipolytic enzyme, belonging to the GDSAG subfamily of hormone sensitive lipases. Furthermore the recombinant 7N9 esterase was biochemically characterized and and was found to be most active at alkaline pH (8.0) and displays salt tolerance over a wide range of concentrations. In silico docking studies confirmed the enzyme’s activity towards short-chain fatty acids while also highlighting the specificity towards certain inhibitors. Furthermore structural differences to a closely related mesophilic E40 esterase isolated from a marine sediment metagenomics library are discussed.

A Wet‐Chemical and Phosphorus K‐edge X‐ray Absorption Near Edge Structure Investigation of Phosphate Adsorption on Binary Mixtures of Ferrihydrite and Calcite: Implications for Phosphorus Bioavailability

Core Ideas Understanding P sorption complexity is critical to managing alkaline soil‐P. Molecular and macro‐scale sorption are characterized in representative systems. Preferential P sorption on ferrihydrite is seen in ferrihydrite–calcite mixtures. Nonlinear P sorptive behavior in mineral–mixtures indicates mineral interaction. An increase in P bioavailability is expected with soil reduction and pH increase. A fundamental understanding of phosphorus (P) bioavailability in alkaline soils is critical to its better management in agricultural and environmental applications. The main objective of this study is to characterize phosphate speciation in the alkaline pH range (pH 8–9.5) in binary mixtures of ferrihydrite and calcite using a combination of wet‐chemical and P K‐edge X‐ray absorption near edge structure spectroscopy. Our results showed that ferrihydrite and calcite sorbed phosphate as an inner‐sphere surface complex with binuclear‐bidentate mode of phosphate bonding on ferrihydrite in both single‐ and mixed‐mineral systems of ferrihydrite and calcite. Phosphate sorbed preferentially on ferrihydrite in binary mixtures of ferrihydrite and calcite particularly at higher concentrations of sorbed phosphate, probably due to greater concentration of metal‐OH surface sites in ferrihydrite than in calcite. In addition, phosphate sorption in mineral mixtures did not behave as a linear combination of phosphate sorption in their single‐mineral components, indicating mineral interactive effects. Molecular‐scale insights from this study have important implications for macro‐scale behavior of P bioavailability in alkaline soils, such as an increase in P bioavailability and subsequent P release with soil reduction and pH increase.

In situ surface transfer process of Cry1Ac protein on SiO2： The effect of biosurfactants for desorption

Genetically modified Bacillus thuringiensis (Bt) crops, which have been widely used in agricultural transgenic plants, express insecticidal Cry proteins and release the toxin into soils. Taking into consideration the environmental risk of Cry proteins, biosurfactant—rhamnolipids were applied to desorb Cry proteins from soil environment, which has not been elucidated before. Quartz crystal microbalance with dissipation (QCM-D) was used in this article to investigate the adsorption and desorption behaviors of Cry1Ac on SiO2 surface (model soil). Results showed that patch-controlled electrostatic attraction (PCEA) governed Cry1Ac adsorption to SiO2, and the solution pH or ionic strength can affect PCEA. The adsorption kinetics could be fitted by the pseudo-second-order model, and the adsorption isotherm was fitted to Langmuir model with correlation coefficients higher than 0.999. The desorption characteristics of Cry1Ac from SiO2 were assessed in the presence of mono-rhamnolipid, di-rhamnolipid or complex-rhamnolipid. Mono-rhamnolipid exhibited the most significant positive effect on desorption performance. With a complete removal of Cry1Ac reached when mono-rhamnolipid concentration was up to 50mgL−1. Additionally, the desorption was enhanced at alkaline pH range, and Cry1Ac can be completely and rapidly desorbed by rhamnolipids from SiO2 at ionic strength of 5×10−2M.

Equilibria and kinetics of chromium(VI) speciation in aqueous solution – A comprehensive study from pH 2 to 11

Abstract A coherent and comprehensive speciation study was conducted in the aqueous Cr(VI) system in order to clarify several discrepancies in the literature. Using pH potentiometric UV–vis titrations and stopped-flow kinetic measurements, it was shown that the dominant Cr(VI) species are Cr2O72−, HCrO4− and CrO42− in an aqueous solution of Cr(VI) under ambient conditions ([Cr(VI)] = 7.7 mM–155 mM, pH = 2–11, I = 1.00 M NaClO4, T = 25.0 °C). This paper provides a comprehensive interpretation of the kinetic features of the dimerization equilibria of Cr(VI) in the acidic – alkaline pH range from pH 2 to 11. Under acidic conditions, the conversion of HCrO4− and Cr2O72− is catalyzed by H+. This observation is consistent with the formation of a protonated intermediate, which is presumably HCr2O7−. The forward and backward rate constants for the 2 HCrO4− ⇌ Cr2O72− + H2O reaction were determined at various pH values in concentration jump experiments. In the absence of catalysts, the slowest dimerization reaction was observed at pH 7 with a lifetime of a few minutes. At basic pH, OH− reacts with Cr2O72− to give CrO42− on stopped-flow timescale. The basic hydrolysis of Cr2O72− is irreversible, and the reaction is first order with respect to both Cr2O72− and OH−. The results presented here are consistent with some of the previously published experimental observations, and resolve the inconsistencies in the literature of Cr(VI) speciation.

Molecular characterization and application of lipase from Bacillus sp. PU1 and investigation of structural changes based on pH and temperature using MD simulation

A gene coding lipase from Bacillus sp. PU1 was cloned and expressed in E. coli BL21(DE3) pLysS. The purified lipase has a molecular weight of 23kDa, is highly alkaline (pH range 8–10) and mesophilic (20–50°C). Three dimensional structure of the lipase was modeled by comparative homology and identified as a typical serine lipase by the presence of conserved Ser77, Asp133, His156. The molecular stability and behavior of the lipase was carried out using MD simulation studies at different pH and temperature was performed in comparison with biochemical analysis. Structural modifications of the lipase under these conditions were trapped by dihedral based FEL analysis and the functional loops (loop-H5/B4 and loop-H6/B5 of lipase) are identified which would cause the catalytic behavior of the lipase by high flexibility. Further characteristic feature of lipase are observed as follows; SDS completely inhibits the lipase activity and enzyme activity is enhanced with non-ionic surfactants. The lipase was highly stable in different organic solvents and also it could tolerate NaCl (0.4–0.8M). This enzyme was found to disrupt the biofilm of tested pathogenic bacterial strains.

Upgradation of Recycled Pulp Using Endoglucanase Enzyme Produced by Pycnoporus sanguineus NFCCI-3628

The importance of recycling and utilization of recycled fiber has recently been recognized because of concerns about environmental issues and economic aspects. In spite of several advantages offered by paper recycling such as wood savings through substitution of virgin pulp and reduction in chemical usage, recycled pulp also contains plenty of fines having high relative surface area and high amount of amorphous cellulose. These fines adsorb water to a large extent and dewatering rate is lower compared to virgin pulp. Selective hydrolysis of these excess ultra-fines through eco-friendly cellulase enzymes can be utilized for enhancing the drainage rate leading to increased paper production. The present study deals with the isolation, production, biochemical characterization of alkali stable endoglucanase enzyme from Pycnoporus sanguineus NFCCI-3628 and its potential application for the improvement of pulp drainage. The enzyme produced by the fungus was stable even in neutral to alkaline pH range 7-8. In the experiments carried out, significant improvement of ~9-14% in drain ability of pulp treated with endoglucanase enzyme was achieved along with better paper properties such as tensile index & smoothness.

A novel heterogeneous Fenton photocatalyst prepared using waste wool fiber combined with Fe3+ ions for dye degradation

Wool fiber Fe complex (Fe-Wool) was prepared with waste wool fiber and Fe3+ ions by a simple exhaust method, and characterized using SEM, FTIR, XRD, XPS, and diffuse reflectance UV-vis spectroscopy. The activity of Fe-Wool was tested as a novel heterogeneous Fenton catalyst for dye degradation in a wide pH range. Effect of Fe content, incorporation of Cu2+ ions, and light irradiation on its catalytic activity was also examined. The results indicated that amino and carboxylic groups or disulfide crosslinks from wool fiber as the ligand sites could react with Fe3+ ions to form Fe-Wool, and the Fe content in Fe-Wool was highly dependent on Fe3+ initial concentration and temperature. Fe-Wool showed a better catalytic activity on the dye degradation under light irradiation than in the dark because it was activated in the UV and visible regions. Incorporation of Cu2+ ions could significantly increase catalytic activity of Fe-Wool for the dye degradation, especially in neutral and alkaline pH range.

Mechanism of mixed dithiophosphate and mercaptobenzothiazole collectors for Cu sulfide ore minerals

The species adsorbed on Cu sulfide ore minerals from alkaline or near neutral solutions of blends of the flotation collectors dibutyl dithiophosphate (BDTP) and 2-mercaptobenzothiazole (MBT) have been established by X-ray photoelectron spectroscopy. Adsorption on air-exposed Cu metal, cuprite, chalcocite, covellite, chalcopyrite and pyrite was investigated from relatively high concentration solutions to simulate processing of variably oxidised ores or slug addition of collector under plant conditions, as well as from solutions of lower concentration. For air-exposed Cu metal, it was found that for a typical BDTP-MBT blend in which di-n-butyl DTP predominated, negligible BDTP was adsorbed at high concentration unless the pH was below 7. By contrast, at low concentrations but under non-starvation conditions, BDTP adsorption predominated even in the alkaline pH range. Both BDTP and MBT species were adsorbed following sequential conditioning in the two collectors. The dithiolate (MBT)2 was not adsorbed from high concentrations of the blend, whereas it was adsorbed from similar concentrations of MBT collector alone. Most of the observed behaviour can be explained by collector solution species reactivity with Cu decreasing in the order MBT−>BDTP−>HMBT. There was no evidence to suggest any surface chemical synergism between the two collectors. For the minerals, competitive chemisorption broadly similar to that for Cu metal was observed, but the formation and adsorption of multilayer molecular CuBDTP or CuMBT was lower, and that of (MBT)2 and HMBT was higher, when the surface availability of Cu was lower than at a native oxide layer. Open circuit potentials for Cu metal, chalcocite and covellite electrodes in 10−3M solution of the collector blend were lower than for the corresponding values in MBT collector alone. For all concentrations in the alkaline pH range, surfaces conditioned in either BDTP alone or the blend were obviously hydrophobic, whereas surfaces conditioned in high concentrations of MBT were not obviously hydrophobic. Some surfaces conditioned sequentially in the two collectors, or in high concentrations of the blend at a pH below 7, were only moderately hydrophobic. Thus, depending on the collector concentration and pH, direct (albeit ex-situ) surface chemical characterisation established the adsorption of different species from a BDTP-MBT blend compared with sequential conditioning in the individual collectors, and hence potentially different hydrophobicities.

Purification and characterization of a novel trypsin-like protease from green-seeded chickpea (Cicer arientum)

ABSTRACTThe present study describes the purification and physicochemical and biochemical characterization of trypsin-like protease from green-seeded chickpea (Cicer arientum). The crude extract of chickpea trypsin (CpT) was obtained by homogenization followed by differential ammonium sulfate precipitation. The CpT was purified by ion-exchange chromatography on diethylaminoethyl (DEAE) column, pre-equilibrated with 20 mM tris-CaCl2 buffer (pH 8.2) with a flow rate of 0.5 mL min−1. The molecular weight and purity of ∼23 kDa of CpT were determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Activity of protease was determined using Nα-benzoyl-DL-arginine-p-nitroanilide as chromogenic substrate and CpT purified showed a specific inhibitor activity of 26978.7697 U mg−1, fold purity of 9.8, and the yield of 70.2%. The characterization was performed for thermal stability, pH profile, and effect of various inhibitors on enzymatic activity. The protein isolated showed stability in the neutral to mild alkaline pH range and thermostability up to 50°C. CpT confirmed its serine nature as it was appreciably inhibited by serine protease inhibitors (maximum 6%), whereas metalloprotease inhibitors barely affected the activity of the enzyme (85%). To the best of our knowledge, it is first reported on purification of protease with trypsin-like properties, from this source.