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Biochar, which contains abundant nutrients like phosphorus (P), is considered a potential source of nutrients for plant use after its application in soil. However, the localized distribution of P in the vicinity of biochar (that is, the charosphere) is poorly understood. In this study, the biochars derived from pinewood (PWB), maize straw (MSB), and chicken manure (CMB) were granulated and investigated for their short-term charosphere effect, using an integrated imaging technique in situ. The results showed that biochars significantly elevated the soil pH and P concentration around their granules, leading to the development of a charosphere over several days. Both pH and P were distributed in a gradient in the charosphere: the values were highest where the biochars were located and declined with increasing distance from the biochar. A highly positive correlation (p < 0.01) was found between the pH, electric conductivity (EC), ash content of the biochar, and the scope of the pH gradient in the charosphere. Moreover, the scope of the P gradient was positively related to the pH and EC values of the biochar as well (p < 0.05). The charosphere effect was prominent around the CMB and MSB granules because of their high ash content, an especially soluble component. The P diffusion in the charosphere was inferred to be influenced by the soil pH as well as the endogenous P species in the biochars. These findings provide new insight into the localized effect of biochars in soil, which is conducive to understanding the nutrient availability to plants after biochar application. [ABSTRACT FROM AUTHOR]

Softwater lakes with specific, rare, and protected aquatic plant vegetation are very sensitive to increased trophic and water chemical changes, especially alkalization. These changes might be reflected in the stable carbon and nitrogen isotopes of the organic matter (OM) of those plants (δ13CORG and δ15NORG) and sediments (δ13CORG and δ15NTN) which they cocreated. To recognize the relationship between OM in the aquatic plants of softwater lakes and the cocreated sediments, we analyzed the plants δ13CORG and δ15NORG in light of four ecological groups (as well as the sediments δ13CORG and δ15NTN in which these plants thrived). Studies were performed in July 2020 on 14 softwater lakes in northern Poland with varying pH gradients from 4.86 to 9.20. For each lake, a single stand was examined for each species detected. The goal was to investigate each species at 10 sites, but this target was not reached for several species. Among the investigated ecological groups, isoetids and mosses showed the least variation in obtained values. In contrast, elodeids exhibited the highest variation in δ13C results due to their diverse carbon uptake strategies, involving both CO2 and HCO3− forms. Moreover, the δ13C values of charophytes and elodeids were highly related to the increasing pH of water. The slight differences between the δ13C results of plants and sediments in isoetid stands further support this finding. Furthermore, we noticed an increase in sediment δ13CORG values along the alkalization gradient, suggesting that macrophytes were the primary source of OM for the sediments. The positive correlation between the δ13C values of plants and sediments (r = 0.69, p < 0.05) might also confirm that the macrophytes were a significant source of OM in sediments. Regarding δ15N values, we did not find significant differentiation between plants and sediments across ecological groups. However, sediment consistently exhibited 15N-enrichment compared to plant material. This enrichment is likely attributed to the accumulation of 15N during the decomposition of the deposited material. This study confirms the possibility of tracking changes in the aquatic vegetation of softwater lakes based primarily on the sediment δ13CORG values along the alkalization gradient. [ABSTRACT FROM AUTHOR]

Soil pH is a critical environmental parameter strongly correlating with microbial communities under various biogeochemical conditions. However, little is known about the co-occurrence pattern of microbial membership in compartments associated with different pH ranges of saline-alkaline grassland soils. To address this gap, we performed a network-based analysis using bacterial and fungal community datasets to characterize co-occurrence patterns and network topological features in four soil pH ranges in a saline-alkaline grassland. Our results indicate that the network topological features of the bacterial and fungal communities differ significantly between different pH ranges in the saline-alkaline grassland. The bacterial co-occurrence pattern in the pH range of 8.0-8.5 was more complex but less stable than that in the other pH ranges, while the fungal co-occurrence pattern had the opposite trend, being less complex but more stable in the pH range of 8.0-8.5. Our co-occurrence analysis identified 14 bacterial species (mainly Proteobacteria and Chloroflexi) and 5 fungal species (mainly in Ascomycota) as key taxa. Overall, our study provides insights into the effects of soil pH on microbial co-occurrence structure and identifies key microbial species in the saline-alkaline grassland of North China. These findings have practical implications for adjusting soil pH and core species to mediate soil microbial co-occurrence structure in saline-alkaline grasslands. [ABSTRACT FROM AUTHOR]

This study marks the exploration into the production of ectoine, a valuable compound with significant potential as an antioxidant, osmoprotectant, anti-inflammatory agent, and stabilizer of cell membranes, proteins, and DNA integrity. Our focus centred on investigating the presence of ectoine and optimizing its production by the novel ectoine producer bacterial strain, Piscibacillus halophilus. For the optimization of ectoine production the effects of carbon and nitrogen sources, salt, pH, agitation and incubation period were optimized by one-factor-at-a-time. We started with an initial ectoine content of 46.92 mg/L, and through a series of optimization processes, we achieved a remarkable increase, resulting in an ectoine content of 1498.2 mg/L. The bacterial species P. halophilus achieved its highest ectoine production after 48 h of incubation, with conditions set at 10 % (w/v) salinity, pH of 7.50, and an agitation speed of 160 rpm. These precise conditions were found to be the most favourable for maximizing ectoine production by this strain. Besides, we have achieved successful purification of ectoine from the crude extract through a streamlined single-step process. This purification method has delivered an exceptional level of purity, surpassing 99.15 %, and an impressive yield of over 99 %. Importantly, we accomplished this using readily available and cost-effective strong acids (HCl) and strong bases (NaOH) to arrange pH gradients. The use of acid and base in the purification process of ectoine reflects an innovative and sustainable methodology. • Piscibacillus halophilus as an alternative ectoine producer. • Ectoine purification with readily available and cost-effective strong acid (HCl) and strong base (NaOH). • Obtaining ectoine with high purity (99.15 %) and yield (99 %) using an innovative and sustainable methodology. [ABSTRACT FROM AUTHOR]

Background: Drought stress during flowering of maize (Zea mays L.) frequently results in decreased kernel setting, leading to grain yield depressions. Plasma membrane (PM) H+‐ATPase was identified as a key enzyme responsible for supply of assimilates to the developing maize kernels shortly after pollination. The activity of this enzyme was strongly inhibited under salt stress, pointing to an involvement in kernel abortion. Aims: This study aimed to determine whether also drought stress causes inhibition of PM H+‐ATPase in developing maize kernels shortly after pollination, leading to diminished hexose uptake and finally kernel abortion. The key questions are as follows: What are the limiting factors for grain yield production of maize plants facing drought? Are physiologically relevant parameters, quantified at flowering, reflected by yield determinants at maturity? Methods: Maize plants were cultivated using the container technique, and drought stress was imposed during 3 weeks bracketing flowering compared to well‐watered conditions throughout the entire growth period. The developing kernels were harvested 2 days after pollination, and PM vesicles were isolated and purified using two‐phase partitioning. Results: Water deficit caused a significant decrease in grain yield at maturity (−35%), which was determined by a reduced kernel number (−42%). Source limitation in the developing kernels under stress could be excluded. Acid invertase activity was unaffected by water deficit. Hexose availability was also no limiting factor for kernel setting and development. However, Vmax of in vitro hydrolytic activity of PM H+‐ATPase was significantly decreased in the developing maize kernels under drought stress and the maximal pH gradient at the PM was also significantly reduced. The observed inhibiting effects on PM H+‐ATPase were mainly of quantitative nature, as a lower number of proton pumps was present in the kernel PM. Qualitative changes of the enzyme (activation energy Ea, Michaelis constant Km) due to drought were not observed. Conclusions: The lower pH gradient probably decreased the proton‐driven transport of hexoses by carriers into the cytosol of the kernel cells, leading to kernel starvation and eventually contributing to kernel abortion. [ABSTRACT FROM AUTHOR]

A novel bioreactor simulating human colonic conditions for in vitro cultivation of intestinal microbiota is presented. The PEristaltic mixed Tubular bioReactor (PETR) is modular designed and periodically kneaded to simulate intestinal peristalsis. The reactor is introduced, characterized from a bioprocess engineer's perspective and discussed in its ability to mimic colon conditions. PETR provides physiological temperature and appropriate anaerobic conditions, simulates intestinal peristalsis, and has a mean residence time of 32.8 ± 0.8 h comparable to the adult human colon. The single‐tube design enables a time‐constant and longitudinally progressive pH gradient from 5.5 to 7.0. Using a dialysis liquid containing high molecular weight polyethylene glycol, the integrated dialysis system efficiently absorbs short chain fatty acids (up to 60%) and water (on average 850 mL d−1). Cultivation of a typical gut bacterium (Bifidobacterium animalis) was performed to demonstrate the applicability for controlled microbiota cultivation. PETR is unique in combining simulation of the entire colon, peristaltic mixing, dialytic water and metabolite absorption, and a progressive pH gradient in a single‐tube design. PETR is a further step to precise replication of colonic conditions in vitro for reliable and reproducible microbiota research, such as studying the effect of food compounds, prebiotics or probiotics, or the development and treatment of infections with enteric pathogens, but also for further medical applications such as drug delivery studies or to study the effect of drugs on and their degradation by the microbiota. [ABSTRACT FROM AUTHOR]

Newly synthesized secretory proteins are exported from the endoplasmic reticulum (ER) at specialized subcompartments called exit sites (ERES). Cargoes like procollagen are too large for export by the standard COPII-coated vesicle of 60 nm average diameter. We have previously suggested that procollagen is transported from the ER to the next secretory organelle, the ER-Golgi intermediate compartment (ERGIC), in TANGO1-dependent interorganelle tunnels. In the theoretical model presented here, we suggest that intrinsically disordered domains of TANGO1 in the ER lumen induce an entropic contraction, which exerts a force that draws procollagen toward the ERES. Within this framework, molecular gradients of pH and/or HSP47 between the ER and ERGIC create a force in the order of tens of femto-Newtons. This force is substantial enough to propel procollagen from the ER at a speed of approximately 1 nm ⋅ s-1. This calculated speed and the quantities of collagen secreted are similar to its observed physiological secretion rate in fibroblasts, consistent with the proposal that ER export is the rate-limiting step for procollagen secretion. Hence, the mechanism we propose is theoretically adequate to explain how cells can utilize molecular gradients and export procollagens at a rate commensurate with physiological needs. [ABSTRACT FROM AUTHOR]

Dielectrophoresis (DEP), a precision nonlinear electrokinetic tool utilized within microfluidic devices, can induce bioparticle polarization that manifests as motion in the electric field; this phenomenon has been leveraged for phenotypic cellular and biomolecular detection, making DEP invaluable for diagnostic applications. As device operation times lengthen, reproducibility and precision decrease, which has been postulated to be caused by ion gradients within the supporting electrolyte medium. This research focuses on characterizing pH gradients above, at, and below the electrode charging frequency (0.2–1.4 times charging frequency) in an aqueous electrolyte solution in order to extend the parameter space for which microdevice-imposed artifacts on cells in clinical diagnostic devices have been characterized. The nonlinear alternating current (AC) electric fields (0.07 Vpp/μm) required for DEP were generated via planar T-shaped and star-shaped microelectrodes overlaid by a 70 μm high microfluidic chamber. The experiments were designed to quantify pH changes temporally and spatially in the two microelectrode geometries. In parallel, a 50 nm hafnium oxide (HfO2) thin film on the microelectrodes was tested to provide insights into the role of Faradaic surface reactions on the pH. Electric field simulations were conducted to provide insights into the gradient shape within the microelectrode geometries. Frequency dependence was also examined to ascertain ion electromigration effects above, at, and below the electrode charging frequency. The results revealed Faradaic reactions above, at, and below the electrode charging frequency. Comparison experiments further demonstrated that pH changes caused by Faradaic reactions increased inversely with frequency and were more pronounced in the star-shaped geometry. Finally, HfO2 films demonstrated frequency-dependent properties, impeding Faradaic reactions. [ABSTRACT FROM AUTHOR]

High energy density was achieved through pH gradient creation in an amidic cellulose incorporated with gold nanoparticles. Three different pH gradients (6–8, 4–10, and 2–12) showed respective specific capacitances of 218, 423 and 603 Fg−1 energy densities of 34, 67 and 96 WhKg −1. The pH gradient overpotential were 0.011, 0.014 and 0.020 V for the three respective pH gradients, where the electrode having the highest overpotential had the highest energy density. The electrode with a pH gradient of 2–12 had the lowest diffusion-based activation energy of 1.358 KJmol −1, and the highest diffusion current. This work successfully show that pH gradient contributes to an additional diffusive current in supercapacitor electrodes and the diffusion current improves the overall specific capacitance. The work also show that pH gradient creates an overpotential that consequently improves the energy density of the electrodes. The retention capacities of 94%, 92% and 91% were obtained for the 6–8, 4–10 and 2–12 respective pH gradients, to establish that pH gradient had no obvious effect on the retention capacity of the electrodes.

Trastuzumab is known to be heterogeneous in terms of charge. Stressing trastuzumab under physiological conditions (pH 7.4 and 37 °C) increases charge heterogeneity further. Separation of charge variants of stressed trastuzumab at the intact protein level is challenging due to increasing complexity making it difficult to obtain pure charge variants for further characterization. Here we report an approach for revealing charge heterogeneity of stressed trastuzumab at the subunit level by pH gradient cation-exchange chromatography. Trastuzumab subunits were generated after limited proteolytic cleavage with papain, IdeS, and GingisKHAN®. The basic pI of Fab and F(ab)2 fragments allowed to use the same pH gradient for intact protein and subunit level analysis. Baseline separation of Fab subunits was obtained after GingisKHAN® and papain digestion and the corresponding modifications were determined by LC–MS/MS peptide mapping and middle-down MALDI-ISD FT-ICR MS. The described approach allows a comprehensive charge variant analysis of therapeutic antibodies that have two or more modification sites in the Fab region. [ABSTRACT FROM AUTHOR]

The local pH values in the close vicinity of the cathode/anode were experimentally determined to be strongly acidic/weakly basic, respectively, during the operation of a high-voltage aqueous sodium-ion battery with Prussian blue-type electrodes and a concentrated aqueous electrolyte. The observed pH gradient was ascribed to O2/H2 evolution at the cathode/anode in aqueous cells, which should contribute to the expansion of the electrochemical window if the pH gradient is maintained. An increase in the distance between the cathode and anode proved to be one of possible solution to suppress the undesired pH neutralization. On the other hand, the reduction of water as well as the dissolved O2 in the electrolyte may diminish the capacity of the anode by a self-discharge. This became more pronounced if the full cell was left to stand for a longer time after charging.

Dielectrophoresis (DEP), a precision nonlinear electrokinetic tool utilized within microfluidic devices, can induce bioparticle polarization that manifests as motion in the electric field; this phenomenon has been leveraged for phenotypic cellular and biomolecular detection, making DEP invaluable for diagnostic applications. As device operation times lengthen, reproducibility and precision decrease, which has been postulated to be caused by ion gradients within the supporting electrolyte medium. This research focuses on characterizing pH gradients above, at, and below the electrode charging frequency (0.2–1.4 times charging frequency) in an aqueous electrolyte solution in order to extend the parameter space for which microdevice-imposed artifacts on cells in clinical diagnostic devices have been characterized. The nonlinear alternating current (AC) electric fields (0.07 Vpp/μm) required for DEP were generated via planar T-shaped and star-shaped microelectrodes overlaid by a 70 μm high microfluidic chamber. The experiments were designed to quantify pH changes temporally and spatially in the two microelectrode geometries. In parallel, a 50 nm hafnium oxide (HfO2) thin film on the microelectrodes was tested to provide insights into the role of Faradaic surface reactions on the pH. Electric field simulations were conducted to provide insights into the gradient shape within the microelectrode geometries. Frequency dependence was also examined to ascertain ion electromigration effects above, at, and below the electrode charging frequency. The results revealed Faradaic reactions above, at, and below the electrode charging frequency. Comparison experiments further demonstrated that pH changes caused by Faradaic reactions increased inversely with frequency and were more pronounced in the star-shaped geometry. Finally, HfO2 films demonstrated frequency-dependent properties, impeding Faradaic reactions.

Self-propelled droplets are of considerable interest as an appropriate model for understanding the self-propulsion of objects in the fields of nonequilibrium physics and nonlinear science. Several research groups have reported the monodirectional motion of droplets, that is, chemotaxis, using stimuli-responsive materials. However, the precise control of chemotaxis remains challenging from the perspective of synthetic chemistry because chemotactic motion is primarily induced by the consumption of reactive oil or surfactants. Herein, we report a chemical system containing pH-responsive fumaric acid derivatives, in which the oil droplet exhibited positive chemotaxis over a wide pH range-from basic to acidic conditions. From the measurements of the interfacial tension between the oil and aqueous phases, it was deduced that the positive chemotaxis was due to heterogeneity in the interfacial tension of the droplet surface, which was accompanied by the production of surface-active compounds in the pH gradient in a linear-type channel. [ABSTRACT FROM AUTHOR]

An overarching challenge in the development of supramolecular sensor systems is to enhance their sensitivity, which commonly involves the synthesis of refined receptors with increased affinity to the analyte. We show that a dramatic sensitivity increase by 1–2 orders of magnitude can be achieved by encapsulating supramolecular chemosensors inside liposomes and exposing them to a pH gradient across the lipid bilayer membrane. This causes an imbalance of the influx and efflux rates of basic and acidic analytes leading to a significantly increased concentration of the analyte in the liposome interior. The utility of our liposome‐enhanced sensors was demonstrated with various host–dye reporter pairs and sensing mechanisms, and we could easily increase the sensitivity towards multiple biologically relevant analytes, including the neurotransmitters serotonin and tryptamine. [ABSTRACT FROM AUTHOR]

Eine umfassende Herausforderung in der Entwicklung supramolekularer Sensorsysteme ist die Verbesserung ihrer Sensitivität, was für gewöhnlich die Synthese verbesserter Rezeptoren mit erhöhter Affinität für den Analyten beinhaltet. Wir zeigen, dass eine ausgeprägte Sensitivitätsverbesserung um ein bis zwei Größenordnungen dadurch erreicht werden kann, dass supramolekulare Chemosensoren in Liposomen eingeschlossen werden, die einem pH‐Gradienten über die Lipiddoppelschicht ausgesetzt sind. Dies hat ein Ungleichgewicht der Zu‐ und Abflussgeschwindigkeiten von basischen und sauren Analyten in das Liposom zur Folge und führt zu einer signifikant erhöhten Analytkonzentration im Inneren des Liposoms. Der Nutzwert unserer mit Liposomen verbesserten Sensoren wurde mit verschiedenen Wirt‐Farbstoff‐Reporterpaaren und Sensormechanismen gezeigt, und wir konnten sehr einfach die Sensitivität für mehrere biologisch wichtige Analyten, inklusive der Neurotransmitter Serotonin und Tryptamin, signifikant erhöhen. [ABSTRACT FROM AUTHOR]

Protein A affinity chromatography has been widely used for initial product capture in recombinant antibody/Fc-fusion purification. However, in general Protein A lacks the capability of separating aggregates (unless the aggregates are too large to enter the pores of resin beads or have their Protein A binding sites buried, in which case the aggregates do not bind). In the current work, we demonstrated that CaptureSelect FcXP affinity medium exhibited strong aggregate separation capability and effectively removed aggregates under pH or conductivity gradient elution in two bispecific antibody (bsAb) cases. For these two cases, aggregate contents were reduced from >16% and >22% (in the feed) to <1% and <5% (in the eluate) for the first and second bsAbs, respectively. While more case studies are required to further demonstrate FcXP's superiority in aggregate removal, findings from the current study suggest that FcXP can potentially be a better alternative than Protein A for product capture in cases where aggregate content is high. • In general, Protein A affinity chromatography lacks the capability of separating aggregates. • FcXP, a CH3-specific affinity medium, exhibits good aggregate separation potential under linear pH gradient elution. • Linear conductivity gradient, which enables improved resolution, facilitates convenient conversion into stepwise elution. • FcXP can potentially be a better alternative than Protein A for product capture in cases where aggregate content is high. [ABSTRACT FROM AUTHOR]

Post-translational modifications ("PTMs") in monoclonal antibodies (mAbs) contribute to charge variant distribution, which will affect biological efficacy and safety. For the characterization of mAbs, charge variants are used as a critical quality attributes for product quality, stability consistency and effectiveness. Charge variants in mAbs are characterized by a time-consuming and a multistep process starting from cation/anion exchange chromatography, acidic/basic fractions collection and subsequent reverse phase (RP) liquid chromatography, coupled with mass spectrometry (MS) analysis. Hence, an alternative characterization approach that would be highly selective for ion exchange chromatography-based charge variant analysis, which is compatible with on-line MS detection, is needed in the biopharma industry. Against this backdrop, multiple studies are being conducted to develop a simple straight on-line charge variant analysis method. In this regard, we apply the current study, which aims to develop a charge variant analytical method, based on volatile buffers with low ionic strength that can be used for on-line MS detection of charge variants of mAbs. This would enable the detection on "PTMs" using low ionic strength mobile phase compatible with MS. Hence, fruitful data can be obtained with a single chromatography run without any test sample preparation, eliminating the need for multiple steps of analysis, time-consuming process and multiple sample preparation steps. Thus, Charge Variant Analysis-MS technique will allow the characterization of charge-related PTMs on the intact protein stage. In this regard, this study is about development of a method having combination of chromatography and volatile mobile phase for mass spectrometry detection of mAbs being analyzed in native form. The method is qualified considering pharmacopeia guidelines because the ultimate aim is to transfer this method for Quality Control (QC) release testing of a monoclonal antibody, which is critical for batch release and the regulatory point of view. Acidic and basic variants have been separated with high resolution peak profile. Furthermore, there was no matrix interference and good separation selectivity in terms of specificity was obtained using this method. The experimental data suggested for the linearity of the method are 2.4 mg/mL to 3.6 mg/mL with % RSD below 2.0%. Additionally, Limit of Quantitation is found to be 0.15 mg/mL, which is 5% of loading amount. Consistently, the data show that the method is precise under the same operating conditions with a short time interval. Overall a simple, accurate, robust and precise pH gradient cation exchange chromatography method was developed and qualified for the characterization of a therapeutic native mAb. Additionally, this method can be used to claim a biosimilar product profile of an in-house product compare to an innovator. [ABSTRACT FROM AUTHOR]

The electrocatalytic nitrate‐to‐ammonia reduction reaction route (NARR) is one of the emerging routes toward green ammonia synthesis, and its conversion efficiency is controlled mainly by the hydrogenation selectivity. This study proposed a likely NARR route feasible and effective even in a neutral condition. Its high catalytic selectivity and efficiency were achieved by a switch of the sulfate solution to the phosphate buffer solution (PBS), while conditions of NO3− concentration, pH, and applied potential were maintained unchanged. Specifically, the faradaic efficiencies toward NH3 (FENH3) in Na2SO4 were as low as 9.8, 19.8, and 11.4 % versus remarkably jumping to 82.8, 90.5, and 89.5 % in PBS under −0.75, −1.0, and −1.25 V, respectively. The corresponding faradaic efficiencies toward NO2− (FENO2-), 77.0, 69.2, and 73.7 % in Na2SO4, significantly dropped to10.8, 7.4, and 4.4 % in PBS, evidencing an unexpected selectivity reversal of the nitrate reduction from NO2− to NH3. This insight was further revealed by the visualization of the pH gradient near the electrode surface during NARR and confirmed by density functional theory calculations; PBS notably facilitated the proton transport and active mitigation over the proton transfer barrier. The use of PBS resulted in a maximal partial current density toward NH3 (JNH3) and NH3 formation rate (rNH3) up to 133.5 mA cm−2 and 1.74×10−7 mol s−1 cm−2 in 1.0 m KNO3 at −1.25 V. [ABSTRACT FROM AUTHOR]

There is considerable interest in developing drugs and probes targeted to mitochondria in order to understand and treat the many pathologies associated with mitochondrial dysfunction. The large membrane potential, negative inside, across the mitochondrial inner membrane enables delivery of molecules conjugated to lipophilic phosphonium cations to the organelle. Due to their combination of charge and hydrophobicity, quaternary triarylphosphonium cations rapidly cross biological membranes without the requirement for a carrier. Their extent of uptake is determined by the magnitude of the mitochondrial membrane potential, as described by the Nernst equation. To further enhance this uptake here we explored whether incorporation of a carboxylic acid into a quaternary triarylphosphonium cation would enhance its mitochondrial uptake in response to both the membrane potential and the mitochondrial pH gradient (alkaline inside). Accumulation of arylpropionic acid derivatives depended on both the membrane potential and the pH gradient. However, acetic or benzoic derivatives did not accumulate, due to their lowered pKa. Surprisingly, despite not being taken up by mitochondria, the phenylacetic or phenylbenzoic derivatives were not retained within mitochondria when generated within the mitochondrial matrix by hydrolysis of their cognate esters. Computational studies, supported by crystallography, showed that these molecules passed through the hydrophobic core of mitochondrial inner membrane as a neutral dimer. This finding extends our understanding of the mechanisms of membrane permeation of lipophilic cations and suggests future strategies to enhance drug and probe delivery to mitochondria.

Background and Aims: Plant-parasitic nematodes are able to sense and respond to gradients of chemical signals. How pH and inorganic salts in the rhizosphere affect nematode accumulation and host-seeking is poorly understood. We investigate the response of different groups of plant-parasitic nematodes to pH and salt concentration gradients. Methods: Responses of infective juveniles (J2) of the economically important plant-parasitic nematodes, soybean cyst nematodes (SCN; Heterodera glycines) and root-knot nematodes (RKN; Meloidogyne incognita and M. hapla) to pH and salt gradients were assessed using Pluronic F-127 gel-based assays. Microelectrodes were utilized to measure pH and ion concentration gradients in the gel. Key Results: Differences were found between the three nematode species in response to acid, base and salts. For SCN, maximum nematode accumulation was between pH range 4.98–5.46 in an acid gradient, while the preferred alkaline pH ranges were 8.40–8.78 and 9.52–9.99. The preferred Cl− concentration for SCN attraction was 171–256 mM. RKN showed weak attraction to base and salt at low J2 concentration but increasing attraction at a greater nematode concentration. Conclusions: The pH and inorganic salts affect nematode behavior, accumulation, and survival. These findings provide new considerations for strategies to manage plant-parasitic nematodes under field conditions. [ABSTRACT FROM AUTHOR]

Mitochondrial Ca2+ handling is accomplished by balancing Ca2+ uptake, primarily via the Ru360-sensitive mitochondrial calcium uniporter (MCU), Ca2+ buffering in the matrix and Ca2+ efflux mainly via Ca2+ ion exchangers, such as the Na+/Ca2+ exchanger (NCLX) and the Ca2+/H+ exchanger (CHE). The mechanism of CHE in cardiac mitochondria is not well-understood and its contribution to matrix Ca2+ regulation is thought to be negligible, despite higher expression of the putative CHE protein, LETM1, compared to hepatic mitochondria. In this study, Ca2+ efflux via the CHE was investigated in isolated rat cardiac mitochondria and permeabilized H9c2 cells. Mitochondria were exposed to (a) increasing matrix Ca2+ load via repetitive application of a finite CaCl2 bolus to the external medium and (b) change in the pH gradient across the inner mitochondrial membrane (IMM). Ca2+ efflux at different matrix Ca2+ loads was revealed by inhibiting Ca2+ uptake or reuptake with Ru360 after increasing number of CaCl2 boluses. In Na+-free experimental buffer and with Ca2+ uptake inhibited, the rate of Ca2+ efflux and steady-state free matrix Ca2+ [mCa2+]ss increased as the number of administered CaCl2 boluses increased. ADP and cyclosporine A (CsA), which are known to increase Ca2+ buffering while maintaining a constant [mCa2+]ss, decreased the rate of Ca2+ efflux via the CHE, with a significantly greater decrease in the presence of ADP. ADP also increased Ca2+ buffering rate and decreased [mCa2+]ss. A change in the pH of the external medium to a more acidic value from 7.15 to 6.8∼6.9 caused a twofold increase in the Ca2+ efflux rate, while an alkaline change in pH from 7.15 to 7.4∼7.5 did not change the Ca2+ efflux rate. In addition, CHE activation was associated with membrane depolarization. Targeted transient knockdown of LETM1 in permeabilized H9c2 cells modulated Ca2+ efflux. The results indicate that Ca2+ efflux via the CHE in cardiac mitochondria is modulated by acidic buffer pH and by total matrix Ca2+. A mechanism is proposed whereby activation of CHE is sensitive to changes in both the matrix Ca2+ buffering system and the matrix free Ca2+ concentration. [ABSTRACT FROM AUTHOR]

Paper-based analytical device (PAD) has received more and more attention in the field of point-of-care test (POCT) due to its low cost, portability and simple operation. Sensitivity and selectivity are two important aspects in clinical diagnostic analysis. However, low sensitivity of a PAD limits its wider application for POCT. Here we introduced a PAD that can clean and enrich the protein content from salty media with both electric field (E) and pH gradient (double E/pH gradients), with which 100-fold enrichment effect could be achieved within 70 s as demonstrated by fluorescein isothiocyanate labeled bovine serum albumin (FITC-BSA) from artificial urine media. With post staining of the protein stacking band with bromophenol blue (BPB), selective colorimetric detection of human serum albumin (HSA) was achieved simply with smartphone camera in the clinically significant range of 10–300 mg‧L−1 (R2 = 0.99) with a limit of detection (LOD) of 4.9 mg‧L−1. Detection of microalbuminuria (MAU) of diabetic patients was demonstrated with this method without difference (ɑ = 0.01) to that by the clinical method (immunoturbidimetry). This work demonstrated the potential of this PAD method in online sample pretreatment and detection of target component from complex physiological samples. Image 1 • Both electric field and pH gradient (E/pH double gradients) can be established and sustained on a paper fluidic channel. • Ampholytic content from salty sample media can be stacked/separated from other types of species. • Proteins from urine can be cumulatively stacked on the double gradient interface without intensive pre-treatment. • Sensitive and selective colorimetric detection of microalbuminuria (MAU) from clinical samples was successfully demonstrated. [ABSTRACT FROM AUTHOR]

Bone differs from other connective tissues; it is isolated by a layer of osteoblasts that are connected by tight and gap junctions. This allows bone to create dense lamellar type I collagen, control pH, mineral deposition, and regulate water content forming a compact and strong structure. New woven bone formed after degradation of mineralized cartilage is rapidly degraded and resynthesized to impart structural order for local bone strength. Ossification is regulated by thickness of bone units and by patterning via bone morphogenetic receptors including activin, other bone morphogenetic protein receptors, transforming growth factor- receptors, all part of a receptor superfamily. This superfamily interacts with receptors for additional signals in bone differentiation. Important features of the osteoblast environment were established using recent tools including osteoblast differentiation in vitro. Osteoblasts deposit matrix protein, over 90% type I collagen, in lamellae with orientation alternating parallel or orthogonal to the main stress axis of the bone. Into this organic matrix, mineral is deposited as hydroxyapatite. Mineral matrix matures from amorphous to crystalline hydroxyapatite. This process includes at least two-phase changes of the calcium-phosphate mineral as well as intermediates involving tropocollagen fibrils to form the bone composite. Beginning with initiation of mineral deposition, there is uncertainty regarding cardinal processes, but the driving force is not merely exceeding the calcium-phosphate solubility product. It occurs behind a epithelial-like layer of osteoblasts, which generate phosphate and remove protons liberated during calcium- phosphate salt deposition. The forming bone matrix is discontinuous from the general extracellular fluid. Required adjustment of ionic concentrations and water removal from bone matrix are important details remaining to be addressed. [ABSTRACT FROM AUTHOR]

• Macro-porous anode is effective for degrading PFOA. • Sulfate can enhance PFOA degradation at neutral pH. • Low localized pH in the pore is responsible for production of SO 4 − radicals and PFOA oxidation. The present study investigated the electrochemical degradation of perfluorooctanoic acid (PFOA) by macro-porous titanium suboxide (p-TiSO) anode in the presence of sulfate at pH-neutral condition (pH 6.9 ± 0.3). Based on initial PFOA concentration of 0.1 mmol L−1, current density of 10 mA cm−2 and electrolysis of 2 h, the removal efficiencies of PFOA were 97.1% and 90.8% for Na 2 SO 4 electrolyte (k app = 2.1 h−1) and NaNO 3 electrolyte (k = 0.84 h−1), corresponding to defluorination efficiency of 61.4% and 54.2%, respectively. The 2.3-fold enhancement of oxidation rate should be originated from the production of sulfate radicals (SO 4 −) by activation of sulfate via direct electron transfer with anode and indirect OH-mediated oxidation under the potential of 3.01–3.41 V versus standard hydrogen electrode (SHE). The production of OH and SO 4 − could be verified on a qualitative basis by using electron spin resonance (ESR) technology. Localized pH measurement achieved with a Pt/IrO x ultra-micro electrode revealed significantly lower pH inside the pores of p-TiSO anode (pH 2.6) compared to that in bulk electrolyte (pH 6.1). The formation of pH gradient should be attributed to surface interaction and diffusion restriction in the porous structure, and the resulting low pH was favorable for enhancing oxidizability of OH toward activation of bisulfate and oxidation of PFOA pollutant. This study offers a proof-in-concept demonstration of electrochemical activation of sulfate and PFOA degradation without need for chemical addition and pH adjustment, which can be realized by simply creating macro-porous structure of anode materials in electrochemical advanced oxidation process. [ABSTRACT FROM AUTHOR]

• New CEX stationary phases were systematically investigated for mAb separations. • Salt-, pH- and salt mediated pH gradient modes were studied. • The particle size of the columns impacted retention rather than efficiency. • A method was suggested to study the presence of secondary interactions. • Small CEX columns allowed for 5–6 min long mAb separations. In this second part of the series, recently commercialized cation exchanger stationary phases were systematically investigated for their capabilities to separate therapeutic monoclonal antibodies. It was demonstrated that the different combinations of stationary and mobile phases result in diverse retention, selectivity and efficiency. Hence, the whole phase system (combination of stationary and mobile phase) should be considered when developing a method. In addition, retention behavior is mAb dependent and should be individually optimized. Another interesting observation was that in cation exchange chromatographic separations of large proteins, the particle size of the columns probably impacts retention rather than efficiency, due to the non-porous particle structure - and therefore the higher specific surface area of smaller particles -. Particle size influences the specific surface area and total porosity. Therefore, columns packed with larger particles showed lower retention (when the ion exchanger group was the same e.g. strong exchanger sulfonic group) while no link was observed between efficiency and particle size. The retention, efficiency and selectivity of the studied columns were quite different and strongly dependent on the elution mode (i.e. salt gradient, pH gradient or combined salt/pH gradient mode). The columns can be considered to be complementary, suggesting that it is useful to have more than one type of column on hand while developing new charge variant assays. Moreover, this work shows that it is especially attractive to make use of short, narrow bore ion exchange columns that offer the possibility to perform 4–6 min long separations of both intact and partially digested antibodies. [ABSTRACT FROM AUTHOR]

Silk sericin is a globular protein whose resistance against fouling is important for applications in biomaterials and water-purification membranes. Here it is shown how sericin generates a water-exclusion zone that may facilitate antifouling behavior. Negatively charged microspheres were used to mimic the surface charge and hydrophobic domains in bacteria. Immersed in water, regenerated silk sericin formed a 100-µm-sized exclusion zone (for micron-size foulants), along with a proton gradient with a decrease of >2 pH-units. Thus, when in contact with sericin, water molecules near the surface restructure to form a physical exclusionary barrier that might prevent biofouling. The decreased pH turns the aqueous medium unviable for neutrophilic bacteria. Therefore, resistance to biofouling seems explainable, among other factors, on the basis of water-exclusionary phenomena. Furthermore, sericin may play a role in triggering the fibroin assembly process by lowering the pH to the required value. [ABSTRACT FROM AUTHOR]

Abstract A traditional immobilized pH gradient (IPG) has a high stability for isoelectric focusing (IEF) but suffers from time-consuming rehydration, focusing and staining-imaging as well as complex performance. To address these issues, an IEF system with an array of 24 IPG columns (10 mm × 600 μm × 50 μm) and dynamic scanning imaging (DSI) was firstly designed for protein focusing. Moreover, two IPG columns (pH 4–9 and pH 6.7–7.7 of 10 mm in length) were firstly synthesized for IEF. A series of experiments were carried out based on the IEF array. In contrast to a traditional IPG IEF with more than 10 h rehydration, 5–14 h IEF and ca 10 h stain-imaging, the IEF array had the following merits: 25 min rehydration for sample loading, 4 min IEF, and 2 min dynamics scanning of 24 columns, well addressing the issues of traditional IEF. Furthermore, the IEF array had fair sensitivity (LOD of 60 ng), good recovery (95%), and high stability (1.02% RSD for intra-day and 2.16% for inter-day). Finally, the developed array was successfully used for separation and determination of HbA 1c (a key biomarker for diabetes diagnosis) in blood samples. All these results indicated the applicability of the developed IEF array to diabetes diagnosis. Graphical abstract Image 1 Highlights • A novel IEF system with an array of 24 IPG columns and dynamic scanning imaging (DSI). • Merits of fast rehydration of 25 min, quick focusing of 4 min and 2 min DSI for 24 IPG column assay. • First report on efficient quantitation of HbA 1c in diabetes sample via IEF array but not HPLC. [ABSTRACT FROM AUTHOR]

Highlights • Alternative buffer systems are suggested for CEX separation of mAbs. • Benefits of salt mediated pH gradient are discussed. • The combination of a "true" pH- and salt gradients extends the possibilities of method development. • Complete method development is feasible within 6 h. Abstract Cation exchange chromatography (CEX) of therapeutic monoclonal antibodies is generally performed with either salt gradient (MES buffer + NaCl) or using commercial pH gradient buffer. The goal of this study was to find out some alternative buffer systems for CEX separation of mAbs, which may offer alternative selectivity, while maintaining similar peak shapes. Among the new buffers that were tested, (N-morpholino)ethanesulfonic acid (MES) / 1,3-diamino-2-propanol (DAP), and citric acid / 2-(cyclohexylamino)ethanesulfonic acid (CHES) systems were particularly promising, especially when combining them with a moderate salt gradient of NaCl. This two buffer system provides an equivalent or slightly better separation than the standard, mobile phases for therapeutic mAbs. It was also demonstrated that working with salt-mediated pH gradients, allows to extend the possibilities in method development, since the concentration of salt in the mobile phase has a significant impact on selectivity. Using HPLC modeling software (Drylab), it was possible to successfully develop CEX methods for authentic mAb samples within only 6 h, by optimizing the gradient steepness and salt concentration in the B eluent. [ABSTRACT FROM AUTHOR]

Global climate change may have a large impact on increased emission rates of carbon dioxide and methane to total greenhouse gas emissions from terrestrial wetlands. Methane consumption by soil microbiota in alpine wet meadows serves as a biofilter for the methane produced in the waterlogged soil below. Altered pH regimes change microbial community composition and structure by exerting selection pressure on soil microorganisms with different ecological strategies and thus affect greenhouse gas emissions resulting from the metabolic activity of soil microorganisms. However, responses of prokaryotic communities to artificial pH shift under elevated methane concentration remain unclear. In this study, we assessed diversity and relative abundance of soil prokaryotes in an alpine meadow under elevated methane concentration along an artificial pH gradient using laboratory incubation experiments. We established an incubation experiment treated with artificial pH gradient (pH 4.5–8.5). After 3 months of incubation, 300 ml of methane at a concentration of 20,000 ppm was added to stimulate potential methanothrophs in topsoil. Sequencing of 16S rRNA gene indicated increasing of relative abundances of Crenarchaeota, Chloroflexi, Bacteroidetes, and Planctomycetes in soil after addition of methane, while the relative abundances of Actinobacteria and Gemmatimonadetes did not significant change before and after methane treatment. Results of phylogenetic relatedness of soil prokaryotes showed that microbial community is mostly shaped by deterministic factors. Species indicator analysis revealed distinct OTUs among various pH and methane treatments. Network analysis revealed distinct co-occurrence patterns of soil prokaryotic community before and after methane addition, and different correlation patterns among various prokaryotic taxa. Linear regression model revealed significant decrease of methane oxidation along elevated pH gradient. Soil pH constituted a strong environmental filter in species assembly of soil prokaryotic community. Methane oxidation rates decreased significantly with elevated pH. The interactive effects of elevated methane concentration and pH are therefore promising topic for future research. [ABSTRACT FROM AUTHOR]

Chloride (Cl−) is essential for O 2 evolution during photosynthetic water oxidation. Two chlorides near the water-oxidizing complex (WOC) in Photosystem II (PSII) structures from Thermosynechococcus elongatus (and T. vulcanus) have been postulated to transfer protons generated from water oxidation. We monitored four criteria: primary charge separation flash yield (P* → P+Q A −), rates of water oxidation steps (S-states), rate of proton evolution, and flash O 2 yield oscillations by measuring chlorophyll variable fluorescence (P* quenching), pH-sensitive dye changes, and oximetry. Br-substitution slows and destabilizes cellular growth, resulting from lower light-saturated O 2 evolution rate (−20 %) and proton release (−36 % ΔpH gradient). The latter implies less ATP production. In Br- cultures, protonogenic S-state transitions (S 2 → S 3 → S 0 ') slow with increasing light intensity and during O 2 /water exchange (S 0 ' → S 0 → S 1), while the non-protonogenic S 1 → S 2 transition is kinetically unaffected. As flash rate increases in Cl− cultures, both rate and extent of acidification of the lumen increase, while charge recombination is suppressed relative to Br−. The Cl− advantage in rapid proton escape from the WOC to lumen is attributed to correlated ion-pair movement of H 3 O+Cl− in dry water channels vs. separated Br− and H+ ion movement through different regions (>200-fold difference in Bronsted acidities). By contrast, at low flash rates a previously unreported reversal occurs that favors Br− cultures for both proton evolution and less PSII charge recombination. In Br− cultures, slower proton transfer rate is attributed to stronger ion-pairing of Br− with AA residues lining the water channels. Both anions charge-neutralize protons and shepherd them to the lumen using dry aqueous channels. [Display omitted] • Whole-cell bromide substitution limits photosynthetic O 2 yield and proton gradient. • Bromide effects are specific to S-state transitions known to release protons. • Bromide substitution is beneficial to charge separation efficiency at low light. • We propose a mechanism of proton removal and attribute Br-effect to basicity. [ABSTRACT FROM AUTHOR]

Bipolar Membrane Reverse Electrodialysis (BMRED) can be used to produce electricity exploiting acid-base neutralization, thus representing a valuable route in reusing waste streams. The present work investigates the performance of a lab-scale BMRED module under several operating conditions. By feeding the stack with 1 M HCl and NaOH streams, a maximum power density of ~17 W m−2 was obtained at 100 A m−2 with a 10-triplet stack with a flow velocity of 1 cm s−1, while an energy density of ~10 kWh m−3 acid could be extracted by a complete neutralization. Parasitic currents along feed and drain manifolds significantly affected the performance of the stack when equipped with a higher number of triplets. The apparent permselectivity at 1 M acid and base decreased from 93% with the five-triplet stack to 54% with the 38-triplet stack, which exhibited lower values (~35% less) of power density. An important role may be played also by the presence of NaCl in the acidic and alkaline solutions. With a low number of triplets, the added salt had almost negligible effects. However, with a higher number of triplets it led to a reduction of 23.4–45.7% in power density. The risk of membrane delamination is another aspect that can limit the process performance. However, overall, the present results highlight the high potential of BMRED systems as a productive way of neutralizing waste solutions for energy harvesting.

Human activity is generating an excess of atmospheric CO 2 , resulting in what we know as ocean acidification, which produces changes in marine ecosystems. Until recently, most of the research in this area had been done under small-scale, laboratory conditions, using few variables, few species and few life cycle stages. These limitations raise questions about the reproducibility of the environment and about the importance of indirect effects and synergies in the final results of these experiments. One way to address these experimental problems is by conducting studies in situ, in natural areas where expected future pH conditions already occur, such as CO 2 vent systems. In the present work, we compile and discuss the latest research carried out in these natural laboratories, with the objective to summarize their advantages and disadvantages for research to improve these investigations so they can better help us understand how the oceans of the future will change., (Copyright © 2024 Elsevier Ltd. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Glyphosate is currently the most widely used herbicide in the world, yet screening of environmental waters for this chemical is limited by the need for specialized derivatization and measurement methods that can be tedious and time-consuming. In this work, we present a novel method for the detection and quantification at trace levels of glyphosate and aminomethylphosphonic acid (AMPA) in environmental water samples. The detection and quantification of the analytes was performed by liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS). Chromatographic separation was achieved with an ion-exchange column and a pH-gradient elution of a solution of ammonium hydroxide and ammonium acetate. The limit of detection for glyphosate and AMPA was 0.25 μg L-1 and the limit of quantification was 0.5 μg L-1with a 20-μL injection. The method was used to investigate the levels of glyphosate and AMPA in surface water samples from the Yarra River catchment area and urban constructed stormwater wetlands. The results indicate that at the time of sampling, no glyphosate or AMPA was present in the samples from the Yarra River catchment area (n = 10). However, glyphosate was detected above the limit of quantification in 33% of the wetland samples (n = 12), with concentrations ranging from 1.95 to 2.96 μg L-1. Similarly, AMPA was quantified in 83% of the wetland samples, with concentrations ranging from 0.55 to 2.42 μg L-1. To our knowledge, this is the first report of a pH-gradient LC-MS/MS method for glyphosate and AMPA analysis at ultratrace levels, with minimal sample processing, avoiding costly, time-consuming derivatization and preconcentration steps.ᅟ [ABSTRACT FROM AUTHOR]

Various types of boron delivery for boron neutron capture therapy (BNCT) have been studied. To selectively destroy cancer cells, high accumulation and selective delivery of 10B into tumor tissue are required. In this study, we developed polyboranes from 1,2-dicarba-closo-dodecaborane as boron carriers, and they were encapsulated into liposomes using either the pH gradient or reverse phase evaporation. The encapsulation efficiency of the liposome prepared using the pH gradient was twice as high as that prepared using reverse-phase evaporation. These liposomes, with diameters of either 50 or 100 nm, were injected into the tail veins of tumor-bearing mice to evaluate their biodistribution at 4, 12, and 24 h post-administration. Boron concentration of the polyborane encapsulated liposomes prepared using the pH gradient achieved 110-150 μg/g of tumor tissue, and the liposomes prepared using the pH gradient were able to achieve an intratumoral 10B concentration of 20-30 μg/g without replacing 11B with 10B. Moreover, this liposome maintained a high 10B level in the tumor for at least 20 h. Average tumor/blood ratios of liposomes reached 5-15 at 4-24 h after injection. From these results, use of polyborane encapsulated liposomes prepared using the pH gradient for BNCT was suggested. [ABSTRACT FROM AUTHOR]

Altered levels of polyamines in biological specimens have been suggested as potential biomarkers for cancer. Difluoromethylornithine (DFMO, an irreversible inhibitor of ornithine decarboxylase) is reported to modulate polyamines to potentially attenuate proliferation of neuroblastoma cells. A clinical trial is being conducted to evaluate DFMO in various cancers. To determine the pharmacodynamics effect of DFMO, an analytical assay is needed to accurately measure the changes in polyamines in cancer cells. In this study, a novel pH gradient LC-ESI-MS/MS method was developed and validated for the quantitation of polyamines (putrescine, spermidine and spermine) in cancer cells. To separate polar and basic polyamines, a multi-mode column composed of ODS and weak ionic ligands was used. The pH gradient was generated from pH 5.3 to pH 2.7 with 2 mM ammonium acetate and 0.4% acetic acid in 10% acetonitrile as mobile phase. The detection of polyamines was performed utilizing multiple reaction monitoring on electrospray ionization mass spectrometry operated in positive ion mode. A pH gradient method increased resolution and decreased peak width of conventional analytical assays, resulting in 10–250-fold higher detection limits. Mobile phases without ion-pairing reagents were LC-MS compatible and eliminated possible signal suppression and MS contamination. The developed method was successfully applied to the analysis of polyamines in neuroblastoma and leukemia cells treated with DFMO. Putrescine levels were significantly ( p  < 0.001) decreased in CCRF-CEM (3.68 vs 1.23 ng/mg protein), SK-N-BE(2) (1.98 vs 1.31 ng/mg protein) and CHLA-20 (2.06 vs 0.90 ng/mg protein) cells treated with DFMO relative to vehicle control. The assay will provide a useful tool in determining the pharmacodynamic effect of DFMO in cancer clinical trials. [ABSTRACT FROM AUTHOR]

Human activity is generating an excess of atmospheric CO2, resulting in what we know as ocean acidification, which produces changes in marine ecosystems. Until recently, most of the research in this area had been done under small-scale, laboratory conditions, using few variables, few species and few life cycle stages. These limitations raise questions about the reproducibility of the environment and about the importance of indirect effects and synergies in the final results of these experiments. One way to address these experimental problems is by conducting studies in situ, in natural areas where expected future pH conditions already occur, such as CO2 vent systems. In the present work, we compile and discuss the latest research carried out in these natural laboratories, with the objective to summarize their advantages and disadvantages for research to improve these investigations so they can better help us understand how the oceans of the future will change. [ABSTRACT FROM AUTHOR]

Soil pH plays an important role in shaping the structure and diversity of prokaryotic community. Altered pH regimes may change prokaryotic community composition by selecting species or groups with different ecological strategies to optimize their fitness. However, it remains unresolved whether prokaryotic communities exhibit deterministic (phylogenetically conserved) or stochastic (phylogenetically overdispersed) responses to pH. In this study we investigated the responses of greenhouse gas emissions and prokaryotic community structure to pH using three-month incubation experiments by adjusting an artificial pH gradient from 4.5 to 8.5. We found decreasing OTUs richness after three months of incubation. Phylogenetic clustering of the prokaryotic community was observed at earlier incubation times whereas greater phylogenetic distance of the prokaryotic community was found at later incubation time. Our results evidenced differential responses of various soil bacterial taxa to the changes in pH. Relative abundances of bacterial phyla and classes of main ecological groups of soil prokaryotes, oligotrophs and copiotrophs, changed significantly along an artificial pH gradient at various incubation times. Relative abundance of Acidobacteria significantly increased with pH at the start of experiment, while opposite trend was observed after 90 days of incubation. In contrast, the relative abundance of Bacteroidetes showed opposite response as Acidobacteria to elevated pH gradient during various incubation time. Methane emissions increased with pH as well as with incubation time, but carbon dioxide and nitrous oxide only increased with incubation time. [ABSTRACT FROM AUTHOR]