The variability of nitrifying activity in field soils

The effect of pH on the activity of nitrifying organisms was examined in pasture soils ranging in pH from 4.9 to 7.3, using a short-term nitrification assay (SNA). The optimum pH for nitrifier activity (pH_opt) was generally close to the soil pH, suggesting that the indigenous nitrifier populations adjusted to the prevailing soil pH. A consequence was that the SNA at the soil pH (SNA_pH) bore a near 1:1 relationship with the SNA at the pH_opt (SNA_opt) over a wide range of SNA values. The effect of soil moisture tension on the SNA was less pronounced than that of pH, but an optimum occurred around pF 3.4.     

pH Tolerance in Freshwater Bacterioplankton: Trait Variation of the Community as Measured by Leucine Incorporation

pH is an important factor determining bacterial community composition in soil and water. We have directly determined the community tolerance (trait variation) to pH in communities from 22 lakes and streams ranging in pH from 4 to 9 using a growth-based method not relying on distinguishing between individual populations. The pH in the water samples was altered to up to 16 pH values, covering in situ pH ± 2.5 U, and the tolerance was assessed by measuring bacterial growth (Leu incorporation) instantaneously after pH adjustment. The resulting unimodal response curves, reflecting community tolerance to pH, were well modeled with a double logistic equation (mean R² = 0.97). The optimal pH for growth (pH_opt) among the bacterial communities was closely correlated with in situ pH, with a slope (0.89 ± 0.099) close to unity. The pH interval, in which growth was ≥90% of that at pH_opt, was 1.1 to 3 pH units wide (mean 2.0 pH units). Tolerance response curves of communities originating from circum-neutral pH were symmetrical, whereas in high-pH (8.9) and especially in low-pH (<5.5) waters, asymmetric tolerance curves were found. In low-pH waters, decreasing pH was more detrimental for bacterial growth than increasing pH, with a tendency for the opposite for high-pH waters. A pH tolerance index, using the ratio of growth at only two pH values (pH 4 and 8), was closely related to pH_opt (R² = 0.83), allowing for easy determination of pH tolerance during rapid changes in pH.     

Anaerobic alkalithermophiles, a novel group of extremophiles

Although some anaerobic and aerobic mesophiles have long been known to grow at alkaline pH (above 9.5), little was known until recently about thermophilic alkaliphiles, termed now alkalithermophiles. This minireview describes presently known and recently validly described anaerobic alkalithermophilic bacteria (pH_opt ^55C > 8.5; T_opt > 55°C) and alkalitolerant thermophiles (pH_opt ^55C < 8.5 but pH_max ^55C above 9.0). Some of these are widely distributed, but others have been isolated (thus far) only from one specific location. This novel group of anaerobic bacteria is comprised of physiologically different genera and species which, so far, all belong to the Gram-type positive Bacillus-Clostridium phylogenetic subbranch. An interesting feature of these anaerobic alkalithermophiles is that most of the isolates have short doubling times. The fastest growing among them are strains of Thermobrachium celere, with doubling times as short as 10 min while growing above pH 9.0 and above 55°C. Received: January 22, 1998 / Accepted: February 16, 1998     

Rational pH-engineering of the thermostable xylanase based on computational model

Locating sites of amino acids related to enzyme properties is still a challenge for rational engineering. Based on the strategy that sites of amino acids can be located by dipeptides, a computational model was made for pH-related dipeptides of xylanase. According to the dipeptide model, pH of a thermostable xylanase B from Thermotoga maritima was rationally designed by locating pH-related amino acids in its sequence and structure. In agreement with expectation, the optimum pH (pH_opt) of the xylanase was improved by five amino acids substitutions: E70Q, E74Q, E77Q, G85Q and T87Q. In parallel assay reactions, at 90 °C, its pH_opt increases to 5.5 from 5.1, and its whole pH profile also shifts 0.5 units towards alkaline area; at 80 °C, the relative activity decreases very little over a wide pH range from 5.25 to 6.0. This result demonstrated that the bioinformatics model is useful for pH rational design and engineering of xylanase, a model molecular of a large family of ～10% proteins with (β/α)₈-barrel structure.     

Transport systems for carbonate in the extremely natronophilic cyanobacterium Euhalothece sp.

The effect of carbonate concentration, pH of the medium, and illumination intensity on the major physiological characteristics (growth rate and the intensities of CO₂ assimilation and oxygen photoproduction) of the natronophilic cyanobacterium Euhalothece sp. Z-M001 have been studied. It was established that the investigated microorganism has at least two transport systems (TS) for CO₂, which differ in both the pH optimum and substrate affinity: TS I has a pH_opt 9.4–9.5 and a K _{S 0.5} of 13–17 mM, whereas TS II has a pH_opt 9.9–10.2 and a K _{S 0.5} of 600–800 mM. The substrate affinity of these transport systems is several orders of magnitude lower than the substrate affinity of the transport systems of freshwater cyanobacteria. It is suggested that they are unique for extremely alkaliphilic cyanobacteria and reflect their adaptation to the seasonal cycles of the lake hydrochemistry.     

Surface Acidity and Solid-State Compatibility of Excipients with an Acid-Sensitive API: Case Study of Atorvastatin Calcium

The objectives of this study were to measure the apparent surface acidity of common excipients and to correlate the acidity with the chemical stability of an acid-sensitive active pharmaceutical ingredient (API) in binary API-excipient powder mixtures. The acidity of 26 solid excipients was determined by two methods, (i) by measuring the pH of their suspensions or solutions and (ii) the pH equivalent (pH_eq) measured via ionization of probe molecules deposited on the surface of the excipients. The chemical stability of an API, atorvastatin calcium (AC), in mixtures with the excipients was evaluated by monitoring the appearance of an acid-induced degradant, atorvastatin lactone, under accelerated storage conditions. The extent of lactone formation in AC-excipient mixtures was presented as a function of either solution/suspension pH or pH_eq. No lactone formation was observed in mixtures with excipients having pH_eq > 6, while the lactone levels were pronounced (> 0.6% after 6 weeks at 50°C/20% RH) with excipients exhibiting pH_eq < 3. The three pH_eq regions (> 6, 3–6, and < 3) were consistent with the reported solution pH-stability profile of AC. In contrast to the pH_eq scale, lactone formation did not show any clear trend when plotted as a function of the suspension/solution pH. Two mechanisms to explain the discrepancy between the suspension/solution pH and the chemical stability data were discussed. Acidic excipients, which are expected to be incompatible with an acid-sensitive API, were identified based on pH_eq measurements. The incompatibility prediction was confirmed in the chemical stability tests using AC as an example of an acid-sensitive API.

Electronic supplementary material

The online version of this article (doi:10.1208/s12249-014-0231-7) contains supplementary material, which is available to authorized users.KEY WORDS: acidity, atorvastatin, excipients, pH indicators, solid-state stability     

A new model for the effect of pH on microbial growth: an extension of the Gamma hypothesis

Aims: To investigate the appropriateness of the extended Lambert–Pearson model (ELPM) to model the effect of pH (as hydrogen and hydroxyl ions) over the whole biokinetic pH range in comparison with other available models. Methods and Results: Data for the effect of pH on microbial growth were obtained from the literature or in‐house. Data were examined using several models for pH. Models were compared using the residual mean of squares. Using the ELPM, pH was modelled as hydrogen ions and hydroxyl ions; hence, the model was monotonic in each. The ELPM was able to model data more successfully than the cardinal pH model (CPM) and other models in the majority of cases. Conclusions: Examining the effect of pH as hydrogen and hydroxyl ions has the advantage that the basic form of the ELPM can be retained as each is treated as a distinct antimicrobial effect. With the ELPM, each inhibitor is described by two parameters; from these parameters, the pH_min, pH_opt and pH_max can be obtained. Furthermore, the idea of a dose response, absent from other models, becomes important. Significance and Impact of the Study: The CPM is an excellent model for certain situations – where there is a high degree of symmetry between the suboptimal pH and superoptimal pH response and where there are few data points available. The ELPM is more amenable to highly asymmetric behaviour, especially where plateaus of effect around the pH optimum are observed and where the number of data points is not restrictive.     

Temperature and pH optima of extremely halophilic archaea: a mini-review

Archaeal microorganisms that grow optimally at Na⁺ concentrations of 1.7 M, or the equivalent of 10% (w/v) NaCl, and greater are considered to be extreme halophiles. This review encompasses extremely halophilic archaea and their growth characteristics with respect to the correlation between the extent of alkaline pH and elevated temperature optima and the extent of salt tolerance. The focus is on poly-extremophiles, i.e., taxa growing optimally at a Na⁺ concentration at or above 1.7 M (approximately 10% w/v NaCl); alkaline pH, at or above 8.5; and elevated temperature optima, at or above 50°C. So far, only a very few extreme halophiles that are able to grow optimally under alkaline conditions as well as at elevated temperatures have been isolated. The distribution of extremely halophilic archaea growing optimally at 3.4 M Na⁺ (approximately 20% w/v NaCl) is bifurcated with respect to pH optima, either they are neutrophilic, with a pH_opt of approximately 7, or strongly alkaliphilic, with pH_opt at or above 8.5. Amongst these extreme halophiles which have elevated pH optima, only four taxa have an optimum temperature above 50°C: Haloarcula quadrata (52°C), Haloferax elongans (53°C), Haloferax mediterranei (51°C) and Natronolimnobius ‘aegyptiacus’ (55°C).     

Hysteresis in poly‐2′‐deoxycytidine i‐motif folding is impacted by the method of analysis as well as loop and stem lengths

In DNA, i‐motif (iM) folds occur under slightly acidic conditions when sequences rich in 2′‐deoxycytidine (dC) nucleotides adopt consecutive dC self base pairs. The pH stability of an iM is defined by the midpoint in the pH transition (pH_T) between the folded and unfolded states. Two different experiments to determine pH_T values via circular dichroism (CD) spectroscopy were performed on poly‐dC iMs of length 15, 19, or 23 nucleotides. These experiments demonstrate two points: (1) pH_T values were dependent on the titration experiment performed, and (2) pH‐induced denaturing or annealing processes produced isothermal hysteresis in the pH_T values. These results in tandem with model iMs with judicious mutations of dC to thymidine to favor particular folds found the hysteresis was maximal for the shorter poly‐dC iMs and those with an even number of base pairs, while the hysteresis was minimal for longer poly‐dC iMs and those with an odd number of base pairs. Experiments to follow the iM folding via thermal changes identified thermal hysteresis between the denaturing and annealing cycles. Similar trends were found to those observed in the CD experiments. The results demonstrate that the method of iM analysis can impact the pH_T parameter measured, and hysteresis was observed in the pH_T and T_m values.     

Effects of transmembrane potential and pH gradient on the cytochrome c-promoted fusion of mitochondrial mimetic membranes

The present study investigated the effects of ΔΨ and ΔpH (pH gradient) on the interaction of cytochrome c with a mitochondrial mimetic membrane composed of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cardiolipin (CL) leading to vesicle fusion. ΔpH generated by lowered bulk pH (pH_out) of PCPECL liposomes, with an internal pH (pH_in) of 8.0, favored vesicle fusion with a titration sigmoidal profile (pK _a?~?6.9). Conversely, ΔpH generated by enhanced pH_in of PCPECL at a pH_out of 6.0 favored the fusion of vesicles with a linear profile. We did not observe a significant amount of liposome fusion when ΔpH was generated by lowered pH_in at a pH_out of 8.0. At bulk acidic pH, ΔΨ generated by Na⁺ gradient also favored cyt c-promoted vesicle fusion. At acidic and alkaline pH_out, the presence of ΔpH and ΔΨ did not affect cytochrome c binding affinity measured by pyrene quenching. Therefore, cytochrome c-mediated PC/PE/CL vesicle fusion is dependent of ionization of the protein site L (acidic pH) and the presence of transmembrane potential. The effect of transmembrane potential is probably related to the generation of defects on the lipid bilayer. These results are consistent with previous reports showing that cytochrome c release prior to the dissipation of the ΔΨ_M blocks inner mitochondrial membrane fusion during apoptosis.     

Influence of salts and pH on growth and activity of a novel facultatively alkaliphilic,extremely salt-tolerant,obligately chemolithoautotrophic sufur-oxidizing Gammaproteobacterium <Emphasis Type="Italic">Thioalkalibacter halophilus</Emphasis> gen. nov., sp. nov. from South-Western Siberian soda lakes

A chemolithoautotrophic sulfur-oxidizing bacterium (SOB) strain ALCO 1 capable of growing at both near-neutral and extremely alkaline pH was isolated from hypersaline soda lakes in S-W Siberia (Altai, Russia). Strain ALCO 1 represents a novel separate branch within the halothiobacilli in the Gammaproteobacteria, which, so far, contained only neutro-halophilic SOB. On the basis of its unique phenotypic properties and distant phylogeny, strain ALCO 1 is proposed as a new genus and species Thioalkalibacter halophilus gen. nov. sp. nov. ALCO 1 was able to grow within a broad range of salinity (0.5–3.5 M of total sodium) with an optimum at around 1 M Na⁺, and pH (7.2–10.2, pH_opt at around 8.5). Na⁺ was required for sulfur-dependent respiration in ALCO 1. The neutral (NaCl)-grown chemostat culture had a much lower maximum growth rate (μ_max), respiratory activity and total cytochrome c content than its alkaline-grown counterpart. The specific concentration of osmolytes (ectoine and glycine-betaine) produced at neutral pH and 3 M NaCl was roughly two times higher than at pH 10 in soda. Altogether, strain ALCO 1 represents an interesting chemolithoautotrophic model organism for comparative investigations of bacterial adaptations to high salinity and pH. Nucleotide sequence accession number: The GenBank/EMBL accession number of the 16S rRNA gene sequence of strain ALCO1^T is EU124668.     

Separate Gating Mechanisms Mediate the Regulation of K2P Potassium Channel TASK-2 by Intra- and Extracellular pH

TASK-2 (KCNK5 or K_2P5.1) is a background K⁺ channel that is opened by extracellular alkalinization and plays a role in renal bicarbonate reabsorption and central chemoreception. Here, we demonstrate that in addition to its regulation by extracellular protons (pH_o) TASK-2 is gated open by intracellular alkalinization. The following pieces of evidence suggest that the gating process controlled by intracellular pH (pH_i) is independent from that under the command of pH_o. It was not possible to overcome closure by extracellular acidification by means of intracellular alkalinization. The mutant TASK-2-R224A that lacks sensitivity to pH_o had normal pH_i-dependent gating. Increasing extracellular K⁺ concentration acid shifts pH_o activity curve of TASK-2 yet did not affect pH_i gating of TASK-2. pH_o modulation of TASK-2 is voltage-dependent, whereas pH_i gating was not altered by membrane potential. These results suggest that pH_o, which controls a selectivity filter external gate, and pH_i act at different gating processes to open and close TASK-2 channels. We speculate that pH_i regulates an inner gate. We demonstrate that neutralization of a lysine residue (Lys²⁴⁵) located at the C-terminal end of transmembrane domain 4 by mutation to alanine abolishes gating by pH_i. We postulate that this lysine acts as an intracellular pH sensor as its mutation to histidine acid-shifts the pH_i-dependence curve of TASK-2 as expected from its lower pK_a. We conclude that intracellular pH, together with pH_o, is a critical determinant of TASK-2 activity and therefore of its physiological function.     

Hydrogenase activity and proton-motive force generation by Escherichia coli during glycerol fermentation

Proton motive force (Δp) generation by Escherichia coli wild type cells during glycerol fermentation was first studied. Its two components, electrical—the membrane potential (?φ) and chemical—the pH transmembrane gradient (ΔpH), were established and the effects of external pH (pH_ex) were determined. Intracellular pH was 7.0 and 6.0 and lower than pH_ex at pH 7.5 and 6.5, respectively; and it was higher than pH_ex at pH 5.5. At high pH_ex, the increase of ?φ (?130 mV) was only partially compensated by a reversed ΔpH, resulting in a low Δp. At low pH_ex ?φ and consequently Δp were decreased. The generation of Δp during glycerol fermentation was compared with glucose fermentation, and the difference in Δp might be due to distinguished mechanisms for H⁺ transport through the membrane, especially to hydrogenase (Hyd) enzymes besides the F₀F₁-ATPase. H⁺ efflux was determined to depend on pH_ex; overall and N,N’-dicyclohexylcarbodiimide (DCCD)-inhibitory H⁺ efflux was maximal at pH 6.5. Moreover, ΔpH was changed at pH 6.5 and Δp was different at pH 6.5 and 5.5 with the hypF mutant lacking all Hyd enzymes. DCCD-inhibited ATPase activity of membrane vesicles was maximal at pH 7.5 and decreased with the hypF mutant. Thus, Δp generation by E. coli during glycerol fermentation is different than that during glucose fermentation. Δp is dependent on pH_ex, and a role of Hyd enzymes in its generation is suggested.     

Caloric restriction controls stationary phase survival through Protein Kinase A (PKA) and cytosolic pH

Calorie restriction is the only physiological intervention that extends lifespan throughout all kingdoms of life. In the budding yeast Saccharomyces cerevisiae, cytosolic pH (pH_c) controls growth and responds to nutrient availability, decreasing upon glucose depletion. We investigated the interactions between glucose availability, pH_c and the central nutrient signalling cAMP‐Protein Kinase A (PKA) pathway. Glucose abundance during the growth phase enhanced acidification upon glucose depletion, via modulation of PKA activity. This actively controlled reduction in starvation pH_c correlated with reduced stationary phase survival. Whereas changes in PKA activity affected both acidification and survival, targeted manipulation of starvation pH_c showed that cytosolic acidification was downstream of PKA and the causal agent of the reduced chronological lifespan. Thus, caloric restriction controls stationary phase survival through PKA and cytosolic pH.     

Reactivity of trypsin in reverse micelles: pH-effects on the W0 versus enzyme activity profiles

pH-Dependence of hydrolytic activity of trypsin has been studied in cationic reverse micellar system of cetyltrimethylammonium bromide (CTAB) in (50% v/v) chloroform/isooctane using a positively charged substrate N_α-benzoyl-L-arginine ethyl ester (BAEE). The pH of the medium was varied from 4.0 to 8.5 with addition of 0.025 M citrate-phosphate buffer containing 1 mM CaCl₂. Optimum pH for maximum enzyme activity, pH_opt in reverse micelles is found to be similar to that observed in bulk aqueous solution (8.0–8.5). However, changes in activity of trypsin (k_cat) as a function of water content W₀ (W₀ = [H₂O]/[CTAB]) in reverse micelles are found to be pH dependent. At low pH (4.0) and low water content (W₀ = 5) the enzyme is more active in reverse micelles than in bulk aqueous solution by a factor of 2. This ‘superactivity’ is lost at higher W₀ values and the k_cat in reverse micelles is found to be similar to that observed in aqueous bulk. At pH 5, the enzyme activity is found to be independent of W₀ while at pH 6.0–6.5 the enzyme activity is low at W₀ 5 and increases with water content to a constant value which is still 50% lower than that in aqueous buffer. Above pH 7, the W_o-activity profile becomes distinctly bell shaped with W₀ optimum around 10–15. The enzyme activity at optimum W₀ is close to that observed in aqueous bulk.     

Tonoplast Anion Channel Activity Modulation by pH in Chara corallina

The patch-clamp technique was used to investigate regulation of anion channel activity in the tonoplast of Chara corallina in response to changing proton and calcium concentrations on both sides of the membrane. These channels are known to be Ca²⁺-dependent, with conductances in the range of 37 to 48 pS at pH 7.4. By using low pH at the vacuolar side (either pH_vac 5.3 or 6.0) and a cytosolic pH (pH_cyt) varying in a range of 4.3 to 9.0, anion channel activity and single-channel conductance could be reversibly modulated. In addition, Ca²⁺-sensitivity of the channels was markedly influenced by pH changes. At pH_cyt values of 7.2 and 7.4 the half-maximal concentration (EC ₅₀) for calcium activation was 100–200 μm, whereas an EC ₅₀ of about 5 μm was found at a pH_cyt of 6.0. This suggests an improved binding of Ca²⁺ ions to the channel protein at more acidic cytoplasm. At low pH_cyt, anion channel activity and mean open times were voltage-dependent. At pipette potentials (V _p) of +100 mV, channel activity was approximately 15-fold higher than activity at negative pipette potentials and the mean open time of the channel increased. In contrast, at pH_cyt 7.2, anion channel activity and the opening behavior seemed to be independent of the applied V _p. The kinetics of the channel could be further controlled by the Ca²⁺ concentration at the cytosolic membrane side: the mean open time significantly increased in the presence of a high cytosolic Ca²⁺ concentration. These results show that tonoplast anion channels are maintained in a highly active state in a narrow pH range, below the resting pH_cyt. A putative physiological role of the pH-dependent modulation of these anion channels is discussed. Received: 14 March 2001/Revised: 16 July 2001     

Models for relating pH measurements in water and calcium chloride for a wide range of pH,soil types and depths

Soil pH is commonly measured in water (pHw) or 0.01 M CaCl₂ (pH_Ca). The need to convert between these methods has led to the publication of linear, quadratic and cubic polynomial relationships for limited suites of soils. Concerns over the applicability of such relationships when mapping a wide range of soils and pH led to the establishment of a database of pH_W and pH_Ca values on each of 7894 samples from soil survey and field experimental sites in Queensland. The relationship between pH_W and pH_Ca across all soils was investigated and preliminary results examining the effect of soil depth and soil type on the relationship are presented.For all soils and depths, a linear regression accounted for 93.2% of the variation but did not predict pH_Ca well at very high or low pH_W values. The inclusion of second and third powers of pH_W accounted for significantly more of the variation (R²=0.94) in pH_Ca and the resultant curve matched the data better at high and low pH.Analysis of surface, sub-surface and subsoil groupings did not reveal any appreciable differences in the relationship between pH_W and pH_Ca attributable to depth. In contrast, differences in the relationship were evident between soil types. Generally, the mildly leached soils had linear relationships, while the weathered soils were distinctly curvilinear at low pH.     

Responses of coral gastrovascular cavity pH during light and dark incubations to reduced seawater pH suggest species-specific responses to the effects of ocean acidification on calcification

Coral polyps have a fluid-filled internal compartment, the gastrovascular cavity (GVC). Respiration and photosynthesis cause large daily excursions in GVC oxygen concentration (O₂) and pH, but few studies have examined how this correlates with calcification rates. We hypothesized that GVC chemistry can mediate and ameliorate the effects of decreasing seawater pH (pH_SW) on coral calcification. Microelectrodes were used to monitor O₂ and pH within the GVC of Montastraea cavernosa and Duncanopsammia axifuga (pH only) in both the light and the dark, and three pH_SW levels (8.2, 7.9, and 7.6). At pH_SW 8.2, GVC O₂ ranged from ca. 0 to over 400% saturation in the dark and light, respectively, with transitions from low to high (and vice versa) within minutes of turning the light on or off. For all three pH_SW treatments and both species, pH_GVC was always significantly above and below pH_SW in the light and dark, respectively. For M. cavernosa in the light, pH_GVC reached levels of pH 8.4–8.7 with no difference among pH_SW treatments tested; in the dark, pH_GVC dropped below pH_SW and even below pH 7.0 in some trials at pH_SW 7.6. For D. axifuga in both the light and the dark, pH_GVC decreased linearly as pH_SW decreased. Calcification rates were measured in the light concurrent with measurements of GVC O₂ and pH_GVC. For both species, calcification rates were similar at pH_SW 8.2 and 7.9 but were significantly lower at pH_SW 7.6. Thus, for both species, calcification was protected from seawater acidification by intrinsic coral physiology at pH_SW 7.9 but not 7.6. Calcification was not correlated with pH_GVC for M. cavernosa but was for D. axifuga. These results highlight the diverse responses of corals to changes in pH_SW, their varying abilities to control pH_GVC, and consequently their susceptibility to ocean acidification.

     

Characterization of intracellular buffering power in human induced pluripotent stem cells and the loss of pluripotency is delayed by acidic stimulation and increase of NHE1 activity

The homeostasis of intracellular pH (pH_i) affects many cellular functions. Our previous study has established a functional and molecular model of the active pH_i regulators in human induced pluripotent stem cells (hiPSCs). The aims of the present study were to further quantify passive pH_i buffering power (β) and to investigate the effects of extracellular pH and Na⁺–H⁺ exchanger 1 (NHE1) activity on pluripotency in hiPSCs. pH_i was detected by microspectrofluorimetry with pH‐sensitive dye‐BCECF. Western blot, immunofluorescence staining, and flow cytometry were used to detect protein expression and pluripotency. Our study in hiPSCs showed that (a) the value of total (β_tot), intrinsic (β_i), and CO₂‐dependent () buffering power all increased while pH_i increased; (b) during the spontaneous differentiation for 4 days, the β values of β_tot and changed in a tendency of decrease, despite the absence of statistical significance; (c) an acidic cultured environment retained pluripotency and further upregulated expression and activity of NHE1 during spontaneous differentiation; (d) inhibition on NHE1 activity promoted the loss of pluripotency. In conclusion, we, for the first time, established a quantitative model of passive β during differentiation and demonstrated that maintenance of NHE1 at a higher level was of critical importance for pluripotency retention in hiPSCs.     

Preferential intracellular pH regulation represents a general pattern of pH homeostasis during acid–base disturbances in the armoured catfish,Pterygoplichthys pardalis

Preferential intracellular pH (pH_i) regulation, where pH_i is tightly regulated in the face of a blood acidosis, has been observed in a few species of fish, but only during elevated blood PCO₂. To determine whether preferential pH_i regulation may represent a general pattern for acid–base regulation during other pH disturbances we challenged the armoured catfish, Pterygoplichthys pardalis, with anoxia and exhaustive exercise, to induce a metabolic acidosis, and bicarbonate injections to induce a metabolic alkalosis. Fish were terminally sampled 2–3 h following the respective treatments and extracellular blood pH, pH_i of red blood cells (RBC), brain, heart, liver and white muscle, and plasma lactate and total CO₂ were measured. All treatments resulted in significant changes in extracellular pH and RBC pH_i that likely cover a large portion of the pH tolerance limits of this species (pH 7.15–7.86). In all tissues other than RBC, pH_i remained tightly regulated and did not differ significantly from control values, with the exception of a decrease in white muscle pH_i after anoxia and an increase in liver pH_i following a metabolic alkalosis. Thus preferential pH_i regulation appears to be a general pattern for acid–base homeostasis in the armoured catfish and may be a common response in Amazonian fishes.