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During intervertebral disc (IVD) degeneration, microenvironmental challenges such as decreasing levels of glucose, oxygen, and pH play crucial roles in cell survival and matrix turnover. Antacids, such as Mg(OH)2 and CaCO3, entrapped in microcapsules are capable of neutralizing acidic microenvironments in a controlled fashion and therefore may offer the potential to improve the acidic niche of the degenerated IVD and enhance cell‐based regeneration strategies. The objectives of this work were, first, to develop and characterize antacid microcapsules and assess their neutralization capacity in an acidic microenvironment and, second, to combine antacid microcapsules with cellular microcapsules in a hybrid gel system to investigate their neutralization effect as a potential therapeutic in a disc explant model. To achieve this, we screened five different pH‐ neutralizing agents (Al(OH)3, Mg(OH)2, CaCO3, and HEPES) in terms of their pH neutralization capacities, with Mg(OH)2 or CaCO3 being carried forward for further investigation. Antacid‐alginate microcapsules were formed at different concentrations using the electrohydrodynamic spraying process and assessed in terms of size, buffering kinetics, cell compatibility, and cytotoxicity. Finally, the combination of cellular microcapsules and antacid capsules was examined in a bovine disc explant model under physiological degenerative conditions. Overall, CaCO3 was found to be superior in terms of neutralization capacities, release kinetics, and cellular response. Specifically, CaCO3 elevated the acidic pH to neutral levels and is estimated to be maintained for several weeks based on Ca2+ release. Using a disc explant model, it was demonstrated that CaCO3 microcapsules were capable of increasing the local pH within the core of a hybrid cellular gel system. This work highlights the potential of antacid microcapsules to positively alter the challenging acidic microenvironment conditions typically observed in degenerative disc disease, which may be used in conjunction with cell therapies to augment regeneration. [ABSTRACT FROM AUTHOR]

Haia Mohammed Kamihan Alenazi, Mohammad Abdul Baseer, Osamah Mohammed AlMugeiren, Navin Anand Ingle Preventive Dentistry Department, College of Medicine and Dentistry, Riyadh Elm University, Riyadh, Saudi ArabiaCorrespondence: Mohammad Abdul Baseer, Preventive Dentistry Department, College of Medicine and Dentistry, Riyadh Elm University, P.Box 84891, Riyadh, 11681, Saudi Arabia, Email ma_baseer@riyadh.edu.saObjective: This study aimed to assess and compare the salivary secretion, pH, and buffer capacity between COVID-19 vaccinated and unvaccinated child patients visiting the clinics of private university dental hospitals in Riyadh, Saudi Arabia.Methods: This is the first comparative assessment of salivary parameters between unvaccinated and COVID-19-vaccinated child patients. The study sample comprised COVID-19 unvaccinated (n = 66) and vaccinated (n = 66) pediatric dental patients aged 4– 12 years seeking dental care in clinics of private university hospitals. Paraffin-stimulated saliva was collected from unvaccinated and vaccinated study participants, and the amount of saliva secreted per minute was noted. Salivary pH and buffering capacity (by 5 mmol Hydrochloric acid titration) were measured using a benchtop digital pH meter. The data obtained were compared between unvaccinated and vaccinated participants using an independent t-test.Results: The results showed a significantly higher salivary secretion rate in unvaccinated than vaccinated study participants (0.83 ± 0.24 mL/min vs 0.67 ± 0.24 mL/min, p = 0.001). Similarly, unvaccinated subjects compared to vaccinated subjects exhibited a significantly higher pH (7.33 ± 0.39 vs 7.04 ± 0.46, p < 0.001) and buffering capacity (6.31 ± 1.55 vs 5.40 ± 1.22, p < 0.001). Moreover, unvaccinated females demonstrated a significantly higher salivary secretion (0.87 ± 0.23 vs 0.71 ± 0.25, t = 2.627, p = 0.011) and buffering capacity 6.19 ± 1.52 vs 5.34 ± 1.25, t = 2.404, p = 0.019) than vaccinated females. Similarly, unvaccinated male exhibited significantly higher salivary secretion (0.80 ± 0.25 vs 0.64 ± 0.23, t = 2.670, p = 0.009), salivary pH (7.39 ± 0.45 vs 6.94 ± 0.41, t=4.309, p< 0.001) and buffering capacity (6.42 ± 1.60 vs 5.45 ± 1.21, t = 2.875, p = 0.005) than the vaccinated male subjects.Conclusion: The vaccinated subjects showed a significantly lower mean salivary secretion, pH, and buffering capacity than unvaccinated participants. Hence, COVID-19 vaccination is likely to affect salivary parameters among pediatric patients.Keywords: COVID-19, vaccine, salivary secretion, pH, buffer, pediatric, patients

ABSTRACT Biofilms provide individual bacteria with many advantages, yet dense cellular proliferation can also create intrinsic metabolic challenges including excessive acidification. Because such pH stress can be masked in buffered laboratory media—such as MSgg commonly used to study Bacillus subtilis biofilms—it is not always clear how such biofilms cope with minimally buffered natural environments. Here, we report how B. subtilis biofilms overcome this intrinsic metabolic challenge through an active pH regulation mechanism. Specifically, we find that these biofilms can modulate their extracellular pH to the preferred neutrophile range, even when starting from acidic and alkaline initial conditions, while planktonic cells cannot. We associate this behavior with dynamic interplay between acetate and acetoin biosynthesis and show that this mechanism is required to buffer against biofilm acidification. Furthermore, we find that buffering-deficient biofilms exhibit dysregulated biofilm development when grown in minimally buffered conditions. Our findings reveal an active pH regulation mechanism in B. subtilis biofilms that could lead to new targets to control unwanted biofilm growth.IMPORTANCEpH is known to influence microbial growth and community dynamics in multiple bacterial species and environmental contexts. Furthermore, in many bacterial species, rapid cellular proliferation demands the use of overflow metabolism, which can often result in excessive acidification. However, in the case of bacterial communities known as biofilms, these acidification challenges can be masked when buffered laboratory media are employed to stabilize the pH environment for optimal growth. Our study reveals that B. subtilis biofilms use an active pH regulation mechanism to mitigate both growth-associated acidification and external pH challenges. This discovery provides new opportunities for understanding microbial communities and could lead to new methods for controlling biofilm growth outside of buffered laboratory conditions.

Introduction: Commercially available local anaesthetics are acidic solutions associated with the vasopressor sting on injection, relatively slower onset of action and pain during palatal injections. The above drawbacks can be addressed by anaesthetic buffering. This prospective study was aimed at comparing the efficacy of buffered and non-buffered local anaesthesia in the extraction of grossly decayed maxillary molar teeth in relation to pain on local infiltration, onset and duration of action of local anaesthesia. Materials and Methods: This is a prospective randomised controlled trial done on 100 patients who required bilateral extraction of maxillary molar teeth. In the study group, patients were given buffered local anaesthesia (which was prepared by mixing 2% lignocaine with 1:80,000 adrenaline and 8.4% sodium bicarbonate) before extraction. In the control group, non-buffered local anaesthesia (2% lignocaine with 1:80,000 adrenaline) was given before extraction. Results: Statistical data confirmed that buffering reduces pain on infiltration, decreases the onset and increases the duration of action of the local anaesthesia compared to non-buffered local anaesthesia. All the parameters measured were statistically significant (P = 0.001). Discussion: The study concludes that buffered local anaesthesia was more beneficial than non-buffered local anaesthesia in reducing pain on injection, providing a quicker onset of local anaesthesia and increasing the duration of action of the local anaesthesia. Buffering is a safe, easy and efficient process and should be routinely followed to provide a better experience to the patients. [ABSTRACT FROM AUTHOR]

Asphyxia, a leading cause of illness and death in newborns, can be improved by early detection and management. Arterial blood gas (ABG) analysis is commonly used to diagnose and manage asphyxia, but it is invasive and carries risks. Dermal interstitial fluid (ISF) is an alternative physiological fluid that can provide valuable information about a person's health. ISF is more sensitive to severe hypoxia and metabolic disorders compared to blood, making it an attractive option for minimally invasive asphyxia detection using biosensors. However, obtaining ISF samples from humans is challenging due to ethical concerns and sampling difficulties. To address this, researchers are developing ISF-mimicking solutions as substitutes for early testing and evaluation of biosensors. This paper focuses on the development of these solutions for bench-based testing and validation of continuous asphyxia-monitoring biosensors. With an understanding of the factors influencing system quality and performance, these solutions can aid in the design of biosensors for in vivo monitoring of dermal ISF. Monitoring interstitial fluid pH levels can provide valuable insights into the severity and progression of asphyxia, aiding in accurate diagnosis and informed treatment decisions. In this study, buffer solutions were prepared to mimic the pH of ISF, and their electrical properties were analyzed. The results suggest that certain buffers can effectively mimic metabolic acidosis associated with asphyxia (pH < 7.30), while others can mimic metabolic alkalosis (pH > 7.45). Overall, this research contributes to the development of ISF-mimicking solutions and lays the groundwork for biosensor systems that monitor dermal ISF in real time. [ABSTRACT FROM AUTHOR]

Biocompatible gold nanoparticles (AuNPs) are used in wound healing due to their radical scavenging activity. They shorten wound healing time by, for example, improving re-epithelialization and promoting the formation of new connective tissue. Another approach that promotes wound healing through cell proliferation while inhibiting bacterial growth is an acidic microenvironment, which can be achieved with acid-forming buffers. Accordingly, a combination of these two approaches appears promising and is the focus of the present study. Here, 18 nm and 56 nm gold NP (Au) were prepared with Turkevich reduction synthesis using design-of-experiments methodology, and the influence of pH and ionic strength on their behaviour was investigated. The citrate buffer had a pronounced effect on the stability of AuNPs due to the more complex intermolecular interactions, which was also confirmed by the changes in optical properties. In contrast, AuNPs dispersed in lactate and phosphate buffer were stable at therapeutically relevant ionic strength, regardless of their size. Simulation of the local pH distribution near the particle surface also showed a steep pH gradient for particles smaller than 100 nm. This suggests that the healing potential is further enhanced by a more acidic environment at the particle surface, making this strategy a promising approach. [ABSTRACT FROM AUTHOR]

Torrefaction is known to reduce the logistical costs of biomass. Torrefied biomass’ prospects as feedstock for fermentation to liquid biofuel are largely unknown. This study investigated the interactions of torrefaction and alkaline pretreatment of wheat straw on glucose yields from enzymatic hydrolysis, including treatment order and the potential for pH to play a role in the process. Across a range of torrefaction severities with temperatures of 200 °C, 220 °C, and 240 °C and durations of 20, 40, and 60 min, torrefaction had a negative impact on glucose yield of wheat straw with or without alkaline pretreatment and regardless of the order of alkaline pretreatment. Alkaline pretreatment after torrefaction results in higher glucose yield than alkaline pretreatment before torrefaction, or by torrefaction alone. Hence, there is the prospect for adding logistical benefits of torrefaction to the bioethanol system if alkaline pretreatment is coupled with torrefaction. Decreasing trend in pH with increase in torrefaction severities was observed for trials without buffer, indicating chemical changes causing the decrease in pH might be associated with the reduction in yield.

For many acidic foods, including fermented and acidified vegetables, salsas, salad dressings, and others, maintaining a pH below 4.6 is a critical control to prevent botulism. The pH of acidic foods is controlled by acid content, low acid ingredients, and buffering; however, buffering of foods remains largely uncharacterized. A Matlab GUI program “BufferCapacity3” was developed to automate the process of quantifying the buffers present in foods utilizing acid/base titration data. The BufferCapacity3 program may be used to aid product development and help assure pH control and safety of acidic foods.

Torrefaction is known to reduce the logistical costs of biomass. Torrefied biomass' prospects as feedstock for fermentation to liquid biofuel are largely unknown. This study investigated the interactions of torrefaction and alkaline pretreatment of wheat straw on glucose yields from enzymatic hydrolysis, including treatment order and the potential for pH to play a role in the process. Across a range of torrefaction severities with temperatures of 200 °C, 220 °C, and 240 °C and durations of 20, 40, and 60 min, torrefaction had a negative impact on glucose yield of wheat straw with or without alkaline pretreatment and regardless of the order of alkaline pretreatment. Alkaline pretreatment after torrefaction results in higher glucose yield than alkaline pretreatment before torrefaction, or by torrefaction alone. Hence, there is the prospect for adding logistical benefits of torrefaction to the bioethanol system if alkaline pretreatment is coupled with torrefaction. Decreasing trend in pH with increase in torrefaction severities was observed for trials without buffer, indicating chemical changes causing the decrease in pH might be associated with the reduction in yield. [ABSTRACT FROM AUTHOR]

The concept of solubility, and its relation to molecular structure, is important for understanding the properties of biological molecules in solution as they interact with the solvent and with each other. To demonstrate the relation between structure, solubility, and buffering capacity, we report a low-cost laboratory exercise using monosodium glutamate (MSG) as a model solute. Students observe how glutamate solution initially resists acidification and how glutamate solubility is first decreased and then restored with acidification. Students should interpret these observations relative to the structure of the solute. This exercise has been employed in introductory level courses for both biology majors and nonmajors and would also be appropriate for AP Biology. [ABSTRACT FROM AUTHOR]

Strategies that alter the pH of wounds to improve healing outcomes are an emerging area of interest. Currently, there is limited understanding of the effect of hydrogen (H+) on the functionality of skin cells during proliferation and migration, highlighting the need for research to determine the effect of pH during wound healing. This study aimed to determine the effect of acidification on the metabolic activity and migration of human immortalized keratinocytes (HaCaT) and human foreskin fibroblasts (HFF). In vitro models were used with phosphoric and citric acid buffers at a pH range between 3 and 7. Our results showed that cells were more viable in buffers with low rather than high ionic strength. A time-dependent effect of the acidification treatment was also observed with cell metabolic activity varying with treatment duration and frequency. Our results showed that a 24 h treatment and subsequent resting phase significantly improved cell proliferation and migration. This in vitro study is the first to establish a correlation between the role of acidic pH, molarity and treatment regimen in cellular activity. Our data demonstrated a positive effect of acidic pH on cell metabolic activity and migration rate, suggesting a clinical potential in indications such as wound healing. [ABSTRACT FROM AUTHOR]

Abstract Background Fungi are premier hosts for the high-yield secretion of proteins for biomedical and industrial applications. The stability and activity of these secreted proteins is often dependent on the culture pH. As yeast acidifies the commonly used synthetic complete drop-out (SD) media that contains ammonium sulfate, the pH of the media needs to be buffered in order to maintain a desired extracellular pH during biomass production. At the same time, many buffering agents affect growth at the concentrations needed to support a stable pH. Although the standard for biotechnological research and development is shaken batch cultures or microtiter plate cultures that cannot be easily automatically pH-adjusted during growth, there is no comparative study that evaluates the buffering capacity and growth effects of different media types across pH-values in order to develop a pH-stable batch culture system. Results We systematically test the buffering capacity and growth effects of a citrate-phosphate buffer (CPB) from acidic to neutral pH across different media types. These media types differ in their nitrogen source (ammonium sulfate, urea or both). We find that the widely used synthetic drop-out media that uses ammonium sulfate as nitrogen source can only be effectively buffered at buffer concentrations that also affect growth. At lower concentrations, yeast biomass production still acidifies the media. When replacing the ammonium sulfate with urea, the media alkalizes. We then develop a medium combining ammonium sulfate and urea which can be buffered at low CPB concentrations that do not affect growth. In addition, we show that a buffer based on Tris/HCl is not effective in maintaining any of our media types at neutral pH even at relatively high concentrations. Conclusion Here we show that the buffering of yeast batch cultures is not straight-forward and addition of a buffering agent to set a desired starting pH does not guarantee pH-maintenance during growth. In response, we present a buffered media system based on an ammonium sulfate/urea medium that enables relatively stable pH-maintenance across a wide pH-range without affecting growth. This buffering system is useful for protein-secretion-screenings, antifungal activity assays, as well as for other pH-dependent basic biology or biotechnology projects.

The present research was conducted aimed at using the approach involving the in vitro tests to evaluate effect by various inorganic buffers used in the diets of Holstein dairy cow on the dietary buffering capacity, ruminal acidogenecity value (AV), methane emission and assess the relationship between them. The buffers were sodium bicarbonate (SB), sodium sesquicarbonate (SSc), a potassium carbonate (PC). Basal diets were low forage [30% forage and 70% concentrate, (FC30:70)], mid forage [35% forage and 65% concentrate, (FC35:65)], and high forage [40% forage and 60% concentrate, (FC40:60)]. The buffers were added to the diets in the concentrations of 0.0, 8.0 and 12.0 g/kg dry matter (DM). In vitro pH, AV, and methane emission of the experimental diets were determined using the gas production technique. Results showed that buffering capacity was significantly the highest for the PC, followed by BH, SB, and SSc (143.3, 138.3, 136.6, and 135, respectively). Analysis of the acid load revealed that adding 8 g/kg of DM of SB in the FC40:60 diet led to the lowest AV (9.6 mg Ca g-1 DM). In addition, adding 12 g/kg of DM of BH in the FC30:70 and 8 g/kg of DM of SB in the FC35:65 diet caused the lowest (5.27) and highest (5.43) pH compared to the other treatments, respectively. The FC40:60 diet containing 8 g/kg DM of PC had the lowest level of methane emission (1.01 mL/0.20 g DM). Our findings demonstrated that the rumen acid load and methane emission may alter when the dietary buffering capacities are changed using the inorganic buffers. [ABSTRACT FROM AUTHOR]

Background: Fungi are premier hosts for the high-yield secretion of proteins for biomedical and industrial applications. The stability and activity of these secreted proteins is often dependent on the culture pH. As yeast acidifies the commonly used synthetic complete drop-out (SD) media that contains ammonium sulfate, the pH of the media needs to be buffered in order to maintain a desired extracellular pH during biomass production. At the same time, many buffering agents affect growth at the concentrations needed to support a stable pH. Although the standard for biotechnological research and development is shaken batch cultures or microtiter plate cultures that cannot be easily automatically pH-adjusted during growth, there is no comparative study that evaluates the buffering capacity and growth effects of different media types across pH-values in order to develop a pH-stable batch culture system. Results: We systematically test the buffering capacity and growth effects of a citrate-phosphate buffer (CPB) from acidic to neutral pH across different media types. These media types differ in their nitrogen source (ammonium sulfate, urea or both). We find that the widely used synthetic drop-out media that uses ammonium sulfate as nitrogen source can only be effectively buffered at buffer concentrations that also affect growth. At lower concentrations, yeast biomass production still acidifies the media. When replacing the ammonium sulfate with urea, the media alkalizes. We then develop a medium combining ammonium sulfate and urea which can be buffered at low CPB concentrations that do not affect growth. In addition, we show that a buffer based on Tris/HCl is not effective in maintaining any of our media types at neutral pH even at relatively high concentrations. Conclusion: Here we show that the buffering of yeast batch cultures is not straight-forward and addition of a buffering agent to set a desired starting pH does not guarantee pH-maintenance during growth. In response, we present a buffered media system based on an ammonium sulfate/urea medium that enables relatively stable pH-maintenance across a wide pH-range without affecting growth. This buffering system is useful for protein-secretion-screenings, antifungal activity assays, as well as for other pH-dependent basic biology or biotechnology projects. [ABSTRACT FROM AUTHOR]

Enteric coatings have shown in vivo dissolution rates that are poorly predicted by traditional in vitro tests, with the in vivo dissolution being considerably slower than in vitro. To provide a more mechanistic understanding of this, the dependence of the release properties of various enteric-coated (EC) products on bulk pH and bicarbonate molarity was investigated. It was found that, at presumably in vivo -relevant values, the bicarbonate molarity is a more significant determinant of the dissolution profile than the bulk pH. The findings also indicate that this steep relationship between the dissolution of enteric coatings and bicarbonate molarity limits those coatings' performance in vivo. This is attributed to the relatively low bicarbonate molarities in human intestinal fluids. Further, the hydration and dehydrations kinetics of carbonic acid and carbon dioxide are not sufficiently rapid to reach equilibrium in the diffusion layer surrounding a dissolving ionizable solid. This results in the effective pKa of bicarbonate in the diffusion layer being lower than that determined potentiometrically at equilibrium in the bulk surrounding fluid. These results demonstrate the importance of thoroughly investigating the intestinal bicarbonate concentrations and using bicarbonate buffers or properly designed surrogates (if possible) when evaluating enteric drug products during product development and quality control. [ABSTRACT FROM AUTHOR]

pH is a central environmental factor influencing CH4 production from organic substrates, as every member of the complex microbial community has specific pH requirements. Here, we show how varying pH conditions (5.0−8.5, phosphate buffered) and the application of a phosphate buffer per se induce shifts in the microbial community composition and the carbon flow during nine weeks of thermophilic batch digestion. Beside monitoring the methane production as well as volatile fatty acid concentrations, amplicon sequencing of the 16S rRNA gene was conducted. The presence of 100 mM phosphate resulted in reduced CH4 production during the initial phase of the incubation, which was characterized by a shift in the dominant methanogenic genera from a mixed Methanosarcina and Methanoculleus to a pure Methanoculleus system. In buffered samples, acetate strongly accumulated in the beginning of the batch digestion and subsequently served as a substrate for methanogens. Methanogenesis was permanently inhibited at pH values ≤5.5, with the maximum CH4 production occurring at pH 7.5. Adaptations of the microbial community to the pH variations included shifts in the archaeal and bacterial composition, as less competitive organisms with a broad pH range were able to occupy metabolic niches at unfavorable pH conditions.

The use of buffers that mimic biological solutions is a foundation of biochemical and biophysical studies. However, buffering agents have both specific and nonspecific interactions with proteins. Buffer molecules can induce changes in conformational equilibria, dynamic behavior, and catalytic properties merely by their presence in solution. This effect is of concern because many of the standard experiments used to investigate protein structure and function involve changing solution conditions such as pH and/or temperature. In experiments in which pH is varied, it is common practice to switch buffering agents so that the pH is within the working range of the weak acid and conjugate base. If multiple buffers are used, it is not always possible to decouple buffer induced change from pH or temperature induced change. We have developed a series of mixed biological buffers for protein analysis that can be used across a broad pH range, are compatible with biologically relevant metal ions, and avoid complications that may arise from changing the small molecule composition of buffers when pH is used as an experimental variable.

During the production of biofuels and/or bioproducts, wood and other lignocellulosic materials are frequently exposed to buffers during enzyme treatments. Buffer pH varies according to the activity profiles of the enzyme(s) used. Time‐of‐flight secondary ion mass spectrometry (ToF‐SIMS) is an increasingly valuable analytical tool for the surface analysis of lignocellulosic solids, allowing for characterization of the lignin and polysaccharides at the surface, along with other components such as protein and inorganic salts. Despite the use of ToF‐SIMS to characterize dilute acid and alkali pretreatments of wood, the exposure of wood to buffers of intermediate pH range has not been studied as it relates to ToF‐SIMS analysis. This leads to the question: “How does soaking wood in various pH buffers impact ToF‐SIMS spectra?” Accordingly, a softwood (spruce) and hardwood (birch) were soaked in universal buffers ranging from pH 5 to 10, and then positive ion ToF‐SIMS spectra were acquired from the washed wood. Deacetylation was evident for both wood species above pH 8. Additionally, at higher pH, birch ToF‐SIMS spectra revealed a relative loss in polysaccharide peaks attributed to hemicellulose and an increase in lignin peaks. This study provides a basis for understanding the pH‐dependent alteration of wood solids in aqueous solution, which is important for understanding the controls in enzyme treatments. [ABSTRACT FROM AUTHOR]

Oncolytic measles virus (MV) is a promising treatment for cancer but titers of up to 1011 infectious particles per dose are needed for therapeutic efficacy, which requires an efficient, robust, and scalable production process. MV is highly sensitive to process conditions, and a substantial fraction of the virus is lost during current purification processes. We therefore conducted forced degradation studies under thermal, pH, chemical, and mechanical stress to determine critical process parameters. We found that MV remained stable following up to five freeze−thaw cycles, but was inactivated during short-term incubation (< 2 h) at temperatures exceeding 35 °C. The infectivity of MV declined at pH < 7, but was not influenced by different buffer systems or the ionic strength/osmolality, except high concentrations of CaCl2 and MgSO4. We observed low shear sensitivity (dependent on the flow rate) caused by the use of a peristaltic pump. For tangential flow filtration, the highest recovery of MV was at a shear rate of ~5700 s−1. Our results confirm that the application of forced degradation studies is important to identify critical process parameters for MV purification. This will be helpful during the early stages of process development, ensuring the recovery of high titers of active MV particles after purification.

Because ionically stabilized colloids in aqueous dispersions have net surface charges that depend on pH, it is potentially possible to separate mixtures of nanospheres having identical radii, yet different types of stabilizing surface charge groups, efficiently using passivated gel electrophoresis (gel-EP). To demonstrate this, we separate a binary dispersion of polystyrene nanospheres that have nearly identical radii and surface group densities, yet different types of anionic stabilizing surface charge groups: sulfate and carboxylate. We achieve an efficient separation by adjusting the pH of the running buffer to lie between the pKavalues of these charge groups, resulting in significantly different protonation and, consequently, different electrophoretic propagation velocities of the nanospheres. The measured steady-state propagation velocities of both types of anionic nanoparticles as a function of pH can be fit well by an equilibrium model of pH-dependent protonation of anionic surface charge groups. Thus, pH-controlled passivated gel-EP opens a route for separating similarly sized charged colloidal objects that are stabilized by a variety of different surface charge groups. [ABSTRACT FROM AUTHOR]

Local anesthesia is widely used in dermatological surgery. But a major concern with their use is the pain experienced by the patients at the time of injection. Various tips and tricks have been published in various studies to minimize this pain. The present review article is written to revisit those tips in the light of the recent literature. [ABSTRACT FROM AUTHOR]

Salmonella is a human pathogen that can accompany live broilers to the slaughter plant, contaminating fully processed carcasses. Feed is one potential source of Salmonella to growing broilers. Monitoring feed for the presence of Salmonella is part of good agricultural practice. The first step in culturing feed for Salmonella (which may be at low numbers and sub-lethally stressed) is to add it to a pre-enrichment broth which is incubated for 24 h. During the course of pre-enrichment, extraneous bacteria metabolize carbohydrates in some feed and excrete acidic byproducts, ausing the pH to drop dramatically. An acidic pre-enrichment pH can injure or kill Salmonella resulting in a failure to detect, even if it is present and available to infect chickens. The objective of this study was to test an array of buffering chemistries to prevent formation of an injurious acidic environment during pre-enrichment of feed in peptone water. Five grams of feed were added to 45 mL of peptone water buffered with carbonate, Tris pH 8, and phosphate buffering ingredients individually and in combination. Feed was subjected to a pre-enrichment at 35°C for 24 h; pH was measured at 0, 18, and 24 h. Standard phosphate buffering ingredients at concentrations up to 4 times the normal formulation were unable to fully prevent acidic conditions. Likewise, carbonate and Tris pH 8 were not fully effective. The combination of phosphate, carbonate, and Tris pH 8 was the most effective buffer tested. It is recommended that a highly buffered pre-enrichment broth be used to examine feed for the presence of Salmonella. [ABSTRACT FROM AUTHOR]

Effects of the types of sugars and buffers, and pH on the formation of Maillard reaction pigments were investigated using the xylose-lysine and glucose-lysine model systems. Sugars (13.3 mM) and lysine (34 mM) were dissolved in acetate, phosphate, and tris(hydroxymethyl)aminomethane (Tris) buffers and heated at 100°C for 60 min. The profiles of browning, reversed-phase HPLC, sugar-decrease, and gel permeation chromatography of each sample were examined. Browning was stimulated by phosphate and inhibited by Tris. With the rise in pH value, browning occurred more intensively, more sugars decreased, and more melanoidins were formed. Xylose samples turned darker brown than glucose ones. The ratio of absorbance at 400 nm to that at 450 or 500 nm increased in the xylose system with the rise in pH value, while that in the glucose system decreased. Low-molecular-weight pigments, dilysyldipyrrolones, were formed only in the xylose system, while melanoidins were formed in both systems. Lysine stimulated the degradation of sugars, polymerization, and browning in both systems, although more intensively in the xylose system than in the glucose one. [ABSTRACT FROM AUTHOR]

Triclosan, an antimicrobial agent commonly used in down-the-drain consumer products, is toxic to freshwater microalgae. However, the rapid photolysis and pH-dependent dissociation of this compound may give rise to uncertainty in growth inhibition tests with freshwater microalgae, if these are not well characterized. Methods are presented to minimize these uncertainties by stabilizing pH with an organic buffering agent (Bis-Tris) and by the application of ultraviolet (UV) covers to remove UV wavelengths. Toxicity tests with these methods were in compliance with the validity criteria of the Organisation for Economic Co-operation and Development test 201, and no negative effects were seen in controls relative to the unmodified method. The methods were used for toxicity tests with triclosan at pH levels of 7.0, 8.0, and 8.5, yielding effective concentration, 10% values of 0.5 µg/L, 0.6 µg/L, and 12.1 µg/L, respectively. The observed change in toxicity with pH was proportional to the change in bioconcentration factor (BCF) as calculated using the cell model (a dynamic flux model based on the Fick-Nernst-Planck equations, in this case parameterized for an algal cell). Effect concentrations produced with the methods presented in the present study offer robust data on which to base risk assessment, and it is suggested that similar approaches be used to minimize uncertainty when other compounds that dissociate and photolyse are tested. Environ Toxicol Chem 2014;33:2551-2559. © 2014 SETAC [ABSTRACT FROM AUTHOR]

Purpose: The purpose of this study is to determine the immediate effects of pH change on the metabolism of bovine chondrocytes. Methods: Bovine osteochondral explants were pre-cultured and placed in Ringer's lactate solution. Thirty explants were randomly divided into 3 groups. Buffered Ringer's lactate, pH 7.2, was used in Group I; buffered Ringer's lactate, pH 7.4 (normal bovine knee joint pH), was used in Group II; and this group was also set as the control group. Buffered Ringer's lactate, pH 7.6, was used in Group III. All specimens were soaked for 2 h. RNA yield analyses were performed to evaluate the changes in cartilage metabolism at different pH levels. Results: Mean RNA yields of (hypoxia-induced factor) 1α that were immersed at pH 7.2 and 7.6 were 10.5- and 9.9-fold from base, respectively, which were lower compared to explants at pH 7.4 which was found as 15.2-fold. Mean RNA yields of aggrecan that were immersed at pH 7.2 and 7.6 were 12.2- and 13.6-fold from base, respectively, which were lower compared to explants at pH 7.4 which was found as 20-fold. RNA yields of collagen type II at pH 7.2 and 7.6 were 16.1- and 16.6-fold from base, respectively, which were lower compared to explants immersed at pH 7.4 which was found as 22.1-fold. Conclusion: The findings of the presented study suggest that short-term exposures to both acidic and basic pH may have effects on chondrocyte function. Our findings also indicate that exposures to solutions with a pH different from normal by only 0.2 unit could suppress chondrocyte metabolism and RNA synthesis. Clinical relevance: Using buffered irrigation solutions with a pH closer to the normal joint pH could be more physiologic and causes less ultra-structural damage than regular irrigation solutions. Level of evidence: II. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: The purpose of this study was to evaluate the influence of dentin on the pH levels of different concentrations of sodium hypochlorite (NaOCl) solutions over time and to evaluate if preconditioning of dentin with 17% EDTA or agitation of the NaOCl solution influences these pH levels. Methods: A novel clinically representative model that scales with the ratio of the irrigant volume to the dentin surface area of a human root canal was used. Three standardized bovine dentin bars (2 × 2 × 10 mm) were placed in a plastic test tube. A total of 150 tubes were distributed in 29 groups. In the first experiment, the pH of various NaOCl solutions, with different concentrations (3%, 6%, and 9%) and starting pH levels (5 and 12), was monitored during exposure to dentin between 10 and 300 seconds. In a second experiment, the effect of agitation (45 Hz) and pretreatment of dentin with 17% EDTA on the pH levels of various NaOCl solutions was studied after 30 seconds of exposure to dentin. The short-term chemical stability of the tested solutions was assessed for both the concentration and the pH. Results: The exposure time (P < .001) and concentration of the NaOCl solution (P < .011) significantly influence the pH level after exposure to dentin. However, the change in pH is too small to induce a change in the irrigant antimicrobial/tissue dissolution capacity. Conclusions: Agitation of the irrigant and preconditioning of the dentin did not alter the pH (P > .05). Both the pH 5 and pH 12 solutions were chemically stable for 1 hour. [Copyright &y& Elsevier]

We investigated the production of hydrogen peroxide ( HOOH) in illuminated seawater media containing a variety of zwitterionic buffers. Production rates varied extensively among buffers, with 4-(2-hydroxyethyl)1-piperazineethanesulfonic acid ( HEPES) highest and N-Tris(hydroxymethyl)methyl-3-aminopropanesulfonic acid ( TAPS) among the lowest. The rate of HOOH accumulation was remarkably consistent over many days, and increased linearly with buffer concentration, natural seawater concentration, and light level. Concentrations of HEPES commonly used in culture media (1-10 mM) generated enough HOOH to kill the axenic Prochlorococcus strain VOL1 during growth in enriched seawater media at lower, environmentally realistic cell concentrations and/or under high light exposure. We also demonstrated that HEPES can be used experimentally to study the biological effects of chronic exposure to sublethal levels of HOOH such as may be experienced by light-exposed microorganisms. [ABSTRACT FROM AUTHOR]

Abstract: Cooled stallion semen has a short viable life, which ranges with acceptable motility and viability from 24 up to 48 hours. The purpose of this study was to compare the effects of storage pH, the ability of three different zwitterionic buffers, and cholesterol-loaded cyclodextrins (CLC) to preserve the motility and integrity of stallion sperm cooled to 5°C for 48 hours. Fourteen ejaculates were collected and split to receive CLC or not (control group). After incubation, each sample was split into six subsamples and diluted in KMT extender containing 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES), N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (BES), or 2-(N-morpholino)ethanesulfonic acid (MES) buffers, and the final pH was adjusted to either 7.0 or 6.6, totalizing 12 experimental groups as a function of CLC, buffer, and pH variables (2 × 3 × 2 factorial). The motility parameters and integrity of plasma and acrosome membranes (live cell index) were determined using computer-automated semen analysis and epifluorescence microscopy at 3, 6, 24, and 48 hours of cooling period. According to results, pH was not a significant source of variation for motility and live sperm over different cooling periods. However, samples diluted in BES exhibited higher progressive motility within 3 hours and higher percentages of total motile cells after 48 hours of incubation at 5°C (P < .05). After 24 hours of storage, CLC-treated sperm samples presented higher motility than control group (P < .05), and after 48 hours of incubation, CLC-treated sperm exhibited higher percentages of live, motile, and progressively motile sperms (P < .05). We inferred that equine semen diluted in KMT containing BES as buffer and CLC treatment improve the equine sperm survival during storage at 5°C for 48 hours. [Copyright &y& Elsevier]

Abstract: Carbon dioxide sequestration using brines has emerged as a promising technology to mitigate the adverse impacts of climate change due to its large storage capacity and favourable chemistries. However, the permanent storage (mineral trapping) of CO2 in brines takes significantly long periods of time as the formation of carbonates is very slow. The main parameters (brine composition, brine pH, system temperature and pressure) have been reported to affect significantly mineral trapping of CO2 sequestration in brines. The precipitation of mineral carbonates is mostly dependent on brine pH. This study aims to promote the formation of carbonates by using additives that can enhance the brine pH. The pH effect of mixtures of a typical buffer solution (0.3M Tris) and coal fly ash (Class C) on promoting the precipitation of mineral carbonates (mainly calcium carbonates) for above-ground carbonation was evaluated in this study. The CO2 binding capacity of the combination of tris buffer and fly ash was around 5.1 times larger than when only tris buffer was used and the combination had a synergistic effect on promoting the formation of mineral carbonates.. [Copyright &y& Elsevier]

The deamidation of asparagine into aspartate and isoaspartate moieties is a major pathway for the chemical degradation of monoclonal antibodies (mAbs). It can affect the shelf life of a therapeutic antibody that is not formulated or stored appropriately. A new approach to detect deamidation using ion exchange chromatography was developed that separates papain-digested mAbs into Fc and Fab fragments. From this, deamidation rates of each fragment can be calculated. To generate kinetic parameters useful in setting shelf life, buffers prepared at room temperature and then placed at the appropriate stability temperatures. Solution pH was not adjusted to the same at different temperatures. Deamidation rate at 40°C was faster in acidic buffers than in basic buffers. However, this trend is reversed at 5°C, attributed to the change in hydroxide ion concentration influenced by buffer and temperature. The apparent activation energy was higher for rates generated in an acidic buffer than in a basic buffer. The rate-pH profile for mAb1 can be deconvoluted to Fc and Fab. The Fc deamidation showed a V-shaped profile: deamidation of PENNY peptide is responsible for the rate at high-pH, whereas deamidation of a new site, Asn323, may be responsible for the rate at low-pH. The profile for Fab is a straight line without curvature. © 2013 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:1712-1723, 2013 [ABSTRACT FROM AUTHOR]

Transmembrane acid-base fluxes affect the intracellular pH and unstirred layer pH around a superfused biological preparation. In this paper the factors influencing the unstirred layer pH and its gradient are studied. An analytical expression of the unstirred layer pH gradient in steady state is derived as a function of simultaneous transmembrane fluxes of (weak) acids and bases with the dehydration reaction of carbonic acid in equilibrium. Also a multicompartment computer model is described consisting of the extracellular bulk compartment, different unstirred layer compartments and the intracellular compartment. With this model also transient changes and the influence of carbonic anhydrase (CA) can be studied. The analytical expression and simulations with the multicompartment model demonstrate that in steady state the unstirred layer pH and its gradient are influenced by the size and type of transmembrane flux of acids and bases, their dissociation constant and diffusion coefficient, the concentration, diffusion coefficient and type of mobile buffers and the activity and location of CA. Similar principles contribute to the amplitude of the unstirred layer pH transients. According to these models an immobile buffer does not influence the steady-state pH, but reduces the amplitude of pH transients especially when these are fast. The unstirred layer pH provides useful information about transmembrane acid-base fluxes. This paper gives more insight how the unstirred layer pH and its transients can be interpreted. Methodological issues are discussed. [ABSTRACT FROM AUTHOR]

A common technique for assaying lipase/esterase activity in vitro is the enzymatic cleavage of para-nitrophenyl esters, with spectrophotometric monitorization at 405-410 nm of the released p-nitrophenol ( pNP). This method has its limitations, since the extinction coefficient of pNP is strongly pH-dependent. Despite this, the method is being frequently used for investigating lipolytic activity over a pH range without any pH-related corrections, which may bring along false results. We show to what extent the results may be altered by this approach and we review alternative strategies allowing the method to become usable both in the acidic range and with varying pH. [ABSTRACT FROM AUTHOR]

Using modeling and simulation, we quantify the influence of spatiotemporal dynamics on the accuracy of data obtained from sensors placed in microscaled reaction volumes. The model refers to cellular reaction (i.e. proton extrusion and oxygen consumption) in complex, buffering solutions. Whole cells or viable tissues cultured in such devices are monitored in real time with integrated sensors for pH and dissolved oxygen. A 3D finite element model of diffusion and metabolic reaction was set up. With respect to pH, the effect of buffering species on proton diffusion is analysed in detail. To account for the delayed time response of real sensors, the sensor impulse response time was implemented by linear convolution. A validation of the model has been achieved by an electrochemical approach. The model reveals significant deviations of measured pH and O, and values of these parameters actually occurring at different sites of the cell culture volume. It is applicable to any setting of (bio-) sensors involving reaction and diffusion of dissolved gases and particularly H ions in buffered solutions. [ABSTRACT FROM AUTHOR]

Abstract: Ecotoxicological tests based on soil enzyme activity are widely used to assess the terrestrial ecotoxicology of metals in soils. However, several standard enzymatic methods use buffers that may alter the chemical pseudoequilibrium of soils and affect metal speciation, and, in turn, the metal effects on enzymes and enzyme kinetics. Researchers have suggested the use of H2O as a solvent rather than chemical buffers, but opponents are concerned about pH fluctuations during incubation and the resulting difficulty in comparing enzymatic studies. Enzyme assays were conducted on 10 pairs of Zn-contaminated soils to evaluate 1) the buffer effect on Zn lability 2) the pH fluctuation during enzymatic assays conducted in water and 3) the comparison of enzymatic results obtained using chemical buffers versus water. Four standard enzymatic methods covering the major biogeochemical cycles were targeted: arylsulfatase (acetate pH 5.8), urease (borate pH 10), acid phosphatase (modified universal buffer pH 6.5) and protease (THAM pH 8.1). Furthermore, deionized water was tested in parallel as a surrogate solvent for these four methods. With the exception of the acetate buffer, the tested solvents did not significantly change the labile Zn concentration in the soil samples. The pH slightly fluctuated by +0.57 pH unit, corresponding to the intrinsic variability of soils. Enzymatic methods using buffers showed similar results compared to those using water, except for urease. These observations suggest that enzymatic methods setting alkaline conditions should be used with caution and that H2O could be used as surrogate solvent in this context. [Copyright &y& Elsevier]

The aim of this study was to compare tooth surface pH after drinking orange juice or water in 39 patients with dental erosion and in 17 controls. The following investigations were carried out: measurement of pH values on selected tooth surfaces after ingestion of orange juice followed by ingestion of water (acid clearance), measurement of salivary flow rate and buffering capacity. Compared with the controls, patients with erosion showed significantly greater decreases in pH after drinking orange juice, and the pH stayed lower for a longer period of time (p < 0.05). Saliva parameters showed no significant differences between the two patient groups except for a lower buffering capacity at pH 5.5 in the erosion group. Copyright © 2012 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Using modeling and simulation, we quantify the influence of spatiotemporal dynamics on the accuracy of data obtained from sensors placed in microscaled reaction volumes. The model refers to cellular reaction (i.e. proton extrusion and oxygen consumption) in complex, buffering solutions. Whole cells or viable tissues cultured in such devices are monitored in real time with integrated sensors for pH and dissolved oxygen. A 3D finite element model of diffusion and metabolic reaction was set up. With respect to pH, the effect of buffering species on proton diffusion is analysed in detail. To account for the delayed time response of real sensors, the sensor impulse response time was implemented by linear convolution. A validation of the model has been achieved by an electrochemical approach. The model reveals significant deviations of measured pH and O2, and values of these parameters actually occurring at different sites of the cell culture volume. It is applicable to any setting of (bio-) sensors involving reaction and diffusion of dissolved gases and particularly H+ ions in buffered solutions. [ABSTRACT FROM AUTHOR]

Abstract: Mucosal immunization offers various advantages over parenteral vaccination, but typically requires potent delivery systems and/or adjuvants to result in protective immunity. Here we report on the preparation of trimethylated chitosan (TMC) and chitosan (CHT) nanoparticles (NPs) loaded with hepatitis B surface antigen (HB), by a simple and scalable method. TMC:HB and CHT:HB NPs were prepared by direct coating of antigen by polymer. The impact of buffer, pH and tonicity of the dispersion medium on NPs’ polydispersity, zeta potential and association percentage of polymer with antigen was evaluated. Moreover, biological properties of both NPs were addressed in vitro by studying their effect on cell viability, transepithelial electrical resistance (TEER) and dendritic cell (DC) maturation. Finally, immunogenicity was assessed by evaluating IgG, IgG1, IgG2a, IgA titers and sIgA after both mucosal (nasal) as well intramuscular (i.m.) vaccination in a murine model. TMC:HB and CHT:HB NPs, prepared in acetate buffer pH 6.7 of three different tonicities, had comparable size, polydispersity, zeta potential and association percentage. TMC:HB NPs, but not CHT:HB NPs, had a mild negative effect on cell viability and TEER, and a considerable positive effect on DC maturation. After nasal and i.m. immunization, TMC:HB NPs in hypotonic medium and CHT:HB NPs in all media induced higher serum and nasal antibody titers compared with HB solution (P <0.001). After i.m. injection, both TMC:HB and CHT:HB NPs induced higher IgG and IgG2a titers compared with alum adsorbed HB (P <0.001). For CHT:HB NPs, the tonicity of the dispersion medium did not affect the mucosal and systemic immune responses. In conclusion, TMC NPs and CHT NPs are similarly potent mucosal immunoadjuvants for HB. Moreover, both polymers are potent immunoadjuvants for i.m. administered isotonic HB, resulting in higher IgG2a/IgG1 ratios compared with alum adjuvanted HB. [Copyright &y& Elsevier]

Abstract: Carbon nanoparticles of ca. 9–18nm diameter (Emperor 2000™) are surface-modified by covalently linking l-dopa-boc (boc-protected l-3,4-dihydroxyphenylalanine) with a surface coverage of approximately 100 per particle (or 3×1013 cm−2). In solution environments these redox-active nanoparticles provide chemically stable and pH-sensitive voltammetirc responses (reversible 2-electron 2-proton oxidation) over a pH range from 2 to 12. When mixed into Dowex 50 Wx4 cation exchanger or Dowex 50 1x2 anion exchanger and placed in contact with a glassy carbon electrode in a flow of humidified gas, the l-dopa-boc-modified carbon nanoparticles provide pH-sensitive surface probes to monitor the surface conditions. In a two-terminal cell it is demonstrated that gas flow measurements are possible with both modified cation and anion exchanger particles in contact to glassy carbon electrodes. The anion exchanger particles allow pH control after pre-conditioning in phosphate buffer. Loading-dependent sensitivity to ammonia gas is investigated and high sensitivity to ammonia is observed for Dowex 50 1x2 anion exchanger pre-equilibrated in phosphate buffer pH 3 and decorated with l-dopa-boc-modified carbon nanoparticles. Responses are observed with sequential injections of 1cm3 ammmonia into a gas flow-through device. [Copyright &y& Elsevier]

Abstract: This work investigated the effect of time on the carbonation reaction in synthetic saline solution, under mild conditions and with a controlled pH, aiming the mineral trapping of CO2 on saline aquifers. The experiments were performed in a glass reactor at 40  °C, 6 hours and atmospheric pressure. The saline solution and solid phase were characterized by Flame Atomic Absorption Spectroscopy (FAAS), Scanning Electron Microscopy (SEM) and Fourier Transform Infrared Spectroscopy (FTIR). This work demonstrated that the saline solution pH lowers to approximately 7-8 in the first hour of reaction (initial pH of approximately 10), remaining constant until the end of the reaction. Through the analysis by AAS, it was found that the concentration of Ca ions in the brine decreases with time. A greatest reduction in the Ca ions concentration occurs within the first two hours of reaction, remaining constant in the others hours. The Mg concentration showed the trend of the final concentration remains close to initial concentration, even with the carbonates precipitation. SEM analysis indicated that the precipitates obtained had morphology similar to that found for various forms of calcium carbonate (i.e calcite and aragonite). FTIR analysis shows characteristic bands of carbonates, which agrees with the results of the analysis by SEM. [Copyright &y& Elsevier]

Manipulation of the extracellular and/or intracellular pH of tumors may have considerable potential in cancer therapy. The extracellular space of most tumors is mildly acidic, owing to exuberant production of lactic acid. Aerobic glycolysis - attributable largely to chronic activation of hypoxia-inducible factor-1 (HIF-1) - as well as tumor hypoxia, are chiefly responsible for this phenomenon. Tumor acidity tends to correlate with cancer aggressiveness; in part, this reflects the ability of HIF-1 to promote invasiveness and angiogenesis. But there is growing evidence that extracellular acidity perse boosts the invasiveness and metastatic capacity of cancer cells; moreover, this acidity renders cancer cells relatively resistant to the high proportion of chemotherapeutic drugs that are mildly basic, and may impede immune rejection of tumors. Thus, practical strategies for raising the extracellular pH of tumors may have therapeutic utility. In rodents, oral administration of sodium bicarbonate can raise the extracellular pH of tumors, an effect associated with inhibition of metastasis and improved responsiveness to certain cytotoxic agents; clinical application of this strategy appears feasible. As an alternative approach, drugs that inhibit proton pumps in cancer cells may alleviate extracellular tumor acidity while lowering the intracellular pH of cancer cells; reduction of intracellular pH slows proliferation and promotes apoptosis in various cancer cell lines. Well-tolerated doses of the proton pump inhibitor esomeprazole have markedly impeded tumor growth and prolonged survival in nude mice implanted with a human melanoma. Finally, it may prove feasible to exploit the aerobic glycolysis of cancers in hyperacidification therapies; intense intracellular acidification of cancer cells achieved by induced hyperglycemia, concurrent administration of proton pump inhibitor drugs, and possibly dinitrophenol, may have the potential to kill cancer cells directly, or to potentiate their responsiveness to adjunctive measures. A similar strategy, but without proton pump inhibition, could be employed to maximize extracellular tumor acidity, enabling tumor- selective release of cytotoxic drugs encased in pH-sensitive nanoparticles. [ABSTRACT FROM AUTHOR]

Maintenance of stable pH is important for optimizing gamete and embryo culture. One method to stabilize pH entails using zwitterionic buffers in IVF handling media used outside the laboratory incubator. Current handling media utilize single buffers, such as MOPS or HEPES. However, the use of a single buffer limits the ability to adjust the range of buffering capacity. Furthermore, changes in temperature alter buffering of these compounds. Therefore, traditional IVF handling media utilizing a single buffer may not provide ideal pH buffering. This study reports that combining multiple buffers, such as HEPES, MOPS and DIPSO, into a single medium in various ratios gives the ability to shift the effective buffering range to cover a specific pH. Additionally, by combining various buffers, it is possible to expand pH buffering over a range of temperatures, while simultaneously reducing the absolute concentration of individual buffers, thereby reducing or alleviating toxicity concerns. This report verifies that DIPSO, MOPS and HEPES, and their combinations, support embryo development. Therefore, utilization of bi- and tri-buffered media, containing a mixture of HEPES, MOPS or DIPSO, offers advantages compared with media containing HEPES or MOPS alone, and may be used for procedures such as oocyte retrieval, intracytoplasmic sperm injection, embryo transfer and cryopreservation. [ABSTRACT FROM AUTHOR]