Effect Of Osmotic Pressure Research Articles

Study of the effect of osmotic pressure on the water permeability of carbon-based two-dimensional materials

Two-dimensional materials such as graphene and carbon nanotube have promising applications in the preparation of permeable membranes and the field of environmental protection due to their unique structures and components. This paper conducts a study on graphene and nanotubes in salt solution using molecular dynamics computational method, and ascertains the quantitative relations between the ionic concentration, water flux and reverse osmosis pressure by setting up theoretical porous models of the two materials. This fundamental research provides much guidance in the determination of critical parameters like pressure and water flux in the actual application of membranes.

Computational Modeling of In Vitro Swelling of Mitochondria: A Biophysical Approach.

Swelling of mitochondria plays an important role in the pathogenesis of human diseases by stimulating mitochondria-mediated cell death through apoptosis, necrosis, and autophagy. Changes in the permeability of the inner mitochondrial membrane (IMM) of ions and other substances induce an increase in the colloid osmotic pressure, leading to matrix swelling. Modeling of mitochondrial swelling is important for simulation and prediction of in vivo events in the cell during oxidative and energy stress. In the present study, we developed a computational model that describes the mechanism of mitochondrial swelling based on osmosis, the rigidity of the IMM, and dynamics of ionic/neutral species. The model describes a new biophysical approach to swelling dynamics, where osmotic pressure created in the matrix is compensated for by the rigidity of the IMM, i.e., osmotic pressure induces membrane deformation, which compensates for the osmotic pressure effect. Thus, the effect is linear and reversible at small membrane deformations, allowing the membrane to restore its normal form. On the other hand, the membrane rigidity drops to zero at large deformations, and the swelling becomes irreversible. As a result, an increased number of dysfunctional mitochondria can activate mitophagy and initiate cell death. Numerical modeling analysis produced results that reasonably describe the experimental data reported earlier.

Effect of Osmotic Pressure on Cellular Stiffness as Evaluated Through Force Mapping Measurements

Atomic force microscopy (AFM) has been used to measure cellular stiffness at different osmolarities to investigate the effect of osmotic pressure on cells. However, substantial direct evidence is essential to clarify the phenomena derived from the experimental results. This study used both the single-point and force mapping methods to measure the effective Young's modulus of the cell by using temporal and spatial information. The single-point force measurements confirmed the positive correlation between cellular stiffness and osmolarity. The force mapping measurements provided local stiffness on the cellular surface and identified the cytoskeleton distribution underneath the plasma membrane. At hyper-osmolarity, the cytoskeleton was observed to cover most of the area underneath the plasma membrane, and the effective Young's modulus on the area with cytoskeleton support was determined to be higher than that at iso-osmolarity. The overall increase in cellular Young's modulus confirmed the occurrence of cytoskeleton compression at hyper-osmolarity. On the other hand, although the average Young's modulus at hypo-osmolarity was lower than that at iso-osmolarity, we observed that the local Young's modulus measured on the areas with cytoskeleton support remained similar from iso-osmolarity to hypo-osmolarity. The reduction of the average Young's modulus at hypo-osmolarity was attributed to reduced cytoskeleton coverage underneath the plasma membrane.

Osmotic Pressure Alters Time-dependent Recovery Behavior of the Intervertebral Disc.

Disc recovery behavior under hypo- and hyperosmotic pressure. To evaluate the effect of osmotic pressure on the unloaded recovery response of healthy discs. The intervertebral disc is a poroviscoelastic material that experiences large fluctuations in water composition throughout a diurnal loading cycle. Fluid flow out of the disc occurs through mechanical loading, whereas fluid flow into the disc occurs through passive diffusion because of an imbalance of ions between the disc and its surrounding environment. Osmotic pressure has been used to alter water uptake and tissue hydration. Motion segments were prepared from the caudal spine sections of the skeletally mature bovines. A 300-N compressive load was applied for 2 hours before unloaded recovery for 12 hours. Hypo- and hyperosmotic pressure was used to alter the rate of water uptake and disc height recovery during unloaded recovery. A 5-parameter rheological model was used to describe the disc's time-dependent recovery behavior. The elastic response was not altered by changes in osmotic pressure; however, viscoelastic recovery was highly dependent on saline osmolarity and recovery time. The fast response of viscoelastic recovery was not dependent on osmotic pressure. The time constant for the slow response decreased whereas the slow response stiffness increased as osmotic pressure increased. The fast response of viscoelastic recovery is governed by flow-independent recovery, whereas the slow response is related to flow-dependent recovery. The rate and magnitude of flow-dependent recovery are highly sensitive to changes in osmotic pressure of the saline bath. There is an osmotic pressure that reduces disc recovery behavior to an elastic response or flow-independent recovery. N/A.

Modification of amaranth (Amaranthus viridis) starch, identification of functional groups, and its potentials as fat replacer

In this study, effects of oxidation, esterification, osmotic pressure, and cross-linking modification methods on functional groups in Amaranthus viridis starch and feasibility of utilizing both native and derived starches as fat replacers were evaluated. The functional groups in native and modified starches were found to consist of OH stretch, CH stretch, CO stretch, NH bend, CH2 and CH3 bend, CO stretch, and CH bend. The results showed that oxidation, osmotic pressure, and cross-linking increased the presence of alkenyl group in starch samples, which enhanced their characteristics as fat replacers. Moreover, modification methods affected the functional groups, which could lead to variations in performance when used as food ingredient. The ability of native and modified A. viridis starches as fat replacers could be ranked as oxidized starch > osmotic pressured starch > cross-linked starch > native starch > esterified starch. Practical applications Amaranthus viridis grains contain a significant amount of starch. The functionality of starch in food industry could be enhanced through modifications. Modified A. viridis starches have functional group (alkenyl group), which enhances their characteristics as fat replacers. Consequently, the modified starches could be used to partially replace fat in fat rich foods such as mayonnaise and salad cream. Thus, minimizes health risks associated with consumption of fatty foods

Effects of osmotic pressure and pH on citric acid and erythritol production from waste cooking oil by Yarrowia lipolytica.

Erythritol and citric acid could be produced from waste cooking oil (WCO) by Yarrowia lipolytica under different medium conditions, and osmotic pressure together with pH were considered to be the critical factors in this process. High osmotic pressure (2.76osmol/L) combined with low pH (pH 3.0) promoted the highest yield of erythritol (21.8g/L) accompanied by low-producing citric acid (2.5g/L). By contrast, the highest citric acid biosynthesis (12.6g/L) was detected under a pH of 6.0 and an osmotic pressure of 0.75osmol/L, when only 4.0g/L of erythritol was yielded. Moreover, lipase activities in these two media were also detected, and pH 3.0-OP 2.76 was supposed to be more beneficial to lipase activity. Biochemical pathways involved in the biosynthesis of erythritol and citric acid were subsequently investigated, and the products yielded from WCO were assumed to be correlated with the activities of transketolase, erythrose reductase, citrate synthase, and glycerol kinase. However, RT-PCR analysis revealed that mRNA levels of these enzymes did not significantly differ, confirming that metabolic flux regulations of erythritol and citric acid mostly took place at the post-transcriptional level.

An Accelerated Release Method of Risperidone Loaded PLGA Microspheres with Good IVIVC.

A long release period lasting several days or several weeks is always needed and thereby it is tedious and time consuming to screen formulations of such microspheres with so long release period and evaluate their release profiles in vitro with conventional long-term or "real-time" release method. So, an accelerated release testing of such system is necessary for formulation design as well as quality control purpose. The purpose of this study is to obtain an accelerated release method of risperidone loaded poly(lactic-co-glycolic acid) (PLGA) microspheres with good in vitro/in vivo correlation (IVIVC). Two formulations of risperidone loaded PLGA microspheres used for evaluating IVIVC were prepared by O/W method. The accelerated release condition was optimized by investigating the effect of pH, osmotic pressure, temperature and ethanol concentration on the release of risperidone from microspheres and the in vitro accelerated release profiles of risperidone from PLGA microspheres were obtained under this optimized accelerated release condition. The plasma concentration of risperidone were also detected after subcutaneous injection of risperidone loaded microspheres to rats. The in vivo cumulative absorption profiles were then calculated using Wagner-Nelson model, Loo- Riegelman model and numerical convolution model, respectively. The correlation between in vitro accelerated release and in vivo cumulative absorption were finally evaluated with Least Square Method. It was shown that temperature and ethanol concentration significantly affected the release of risperidone from the microspheres while pH and osmotic pressure of release media slightly affected the release behavior of risperidone. The in vitro release of risperidone from microspheres were finally undergone in PBS (pH7.0, 300mosm) with 20% (V/V) ethanol at 45°C. The sustained and complete release of risperidone was observed in both formulations under the accelerated release condition although these two release profiles were dissimilar. The correlation coefficients (R2) of IVIVC were all above 0.95 and the slopes were all between 0.9564 and 1.1868 in spite of fitted model and microsphere formulation. An in vitro accelerated release method of risperidone microspheres with good IVIVC was established in this paper and this accelerated release method was supposed to have great potential in both in vivo performance prediction and quality control for risperidone loaded PLGA microspheres.

Combined effects of osmotic and hydrostatic pressure on multilamellar lipid membranes in the presence of PEG and trehalose.

We studied the interaction of lipid membranes with the disaccharide trehalose (TRH), which is known to stabilize biomembranes against various environmental stress factors. Generally, stress factors include low/high temperature, shear, osmotic and hydrostatic pressure. Small-angle X-ray-scattering was applied in combination with fluorescence spectroscopy and calorimetric measurements to get insights into the influence of trehalose on the supramolecular structure, hydration level, and elastic and thermodynamic properties as well as phase behavior of the model biomembrane DMPC, covering a large region of the temperature, osmotic and hydrostatic pressure phase space. We observed distinct effects of trehalose on the topology of the lipid's supramolecular structure. Trehalose, unlike osmotic pressure induced by polyethylene glycol, leads to a decrease of lamellar order and a swelling of multilamellar vesicles, which is attributable to direct interactions between the membrane and trehalose. Our results revealed a distinct biphasic concentration dependence of the observed effects of trehalose. While trehalose intercalates between the polar head groups at low concentrations, the effects after saturation are dominated by the exclusion of trehalose from the membrane surface.

Preserving the inflated structure of lyophilized sporopollenin exine capsules with polyethylene glycol osmolyte

Extracted from natural pollen grains, sporopollenin exine capsules (SECs) are robust, chemically inert biopolymer shells that posess highly uniform size and shape characteristics and that can be utilized as hollow microcapsules for drug delivery applications. However, it is challenging to extract fully functional SECs from many pollen species because pollen grains often collapse, causing the loss of architectural features, loading volume, and bulk uniformity. Herein, we demonstrate that polyethylene glycol (PEG) osmolyte solutions can help preserve the native architectural features of extracted SECs, yielding inflated microcapsules of high uniformity that persist even after subsequent lyophilization. Optimal conditions were first identified to extract SECs from cattail (Typhae angustfolia) pollen via phosphoric acid processing after which successful protein removal was confirmed by elemental (CHN), mass spectrometry (MALDI-TOF), and confocal laser canning microscopy (CLSM) analyses. The shape of SECs was then assessed by scanning electron microscopy (SEM) and dynamic image particle analysis (DIPA). While acid-processed SECs experienced high degrees of structural collapse, incubation in 2.5% or higher PEG solutions significantly improved preservation of spherical SEC shape by inducing inflation within the microcapsules. A theoretical model of PEG-induced osmotic pressure effects was used to interpret the experimental data, and the results show excellent agreement with the known mechanical properties of pollen exine walls. Taken together, these findings demonstrate that PEG osmolyte is a useful additive for preserving particle shape in lyophilized SEC formulations, opening the door to broadly applicable strategies for stabilizing the structure of hollow microcapsules.

Revisiting in vitro release test for topical gel formulations: The effect of osmotic pressure explored for better bio-relevance

Release test methods for topical dosage forms including pharmacopeial tests require a large volume of release media, with limited application for high throughput screening. In the present study, we evaluated Transwell assay to miniaturize the release test method for optimization of thermoreversible topical gel formulations. We also explored the osmotic effect on the in vitro release rates from gel formulations to understand the bio-relevance of release media. An extreme vertices type of mixture design in Minitab®16 generated eleven formulations composed of poloxamer 407, poloxamer 188, and phosphate buffered saline (PBS). A quadratic equation adequately described the composition dependence of gelation temperature. Epidermal growth factor (EGF) and trypan blue were used as model drugs for proteins and small molecules, respectively. Cumulative release in PBS containing 30% sucrose exhibited linear correlation with respect to the gel compositions, while PBS without sucrose did not differentiate various compositions. Higher release rates in PBS than in sucrose media are attributable to the osmotic water flow from PBS into the donor phase, and subsequent increase in diffusivity. The time course of in vivo near-infrared fluorescence imaging of topical EGF gels on the wound sites were consistent with the in vitro release profiles measured with PBS as the release media. To the best of our knowledge, this is the first study to propose a release test method suitable for high throughput screening of topical formulations with emphasis on the osmotic pressure effect. Bio-relevant release media composition for a topical formulation would vary depending on its clinical application because the osmotic water flow through the normal skin would be negligible compared to compromised skin.

The feasibility of nanofiltration membrane bioreactor (NF-MBR)+reverse osmosis (RO) process for water reclamation: Comparison with ultrafiltration membrane bioreactor (UF-MBR)+RO process

This study examines the feasibility of a novel nanofiltration membrane bioreactor (NF-MBR) followed by reverse osmosis (RO) process for water reclamation at 90% recovery and using an ultrafiltration MBR (UF-MBR)+RO as baseline for comparison. Both MBRs adopted the same external hollow fiber membrane configurations and operating conditions. The collected permeates of the MBRs were subsequently fed to the respective RO systems. The results showed that the NF-MBR (operated at a constant flux of 10 L/m2h) achieved superior MBR permeate quality due to enhanced biodegradation and high rejection capacity of the NF membrane, leading to lower RO fouling rates (∼3.3 times) as compared to the UF-MBR. Further analysis indicated that the cake layer fouling that caused the cake-enhanced osmotic pressure (CEOP) effect contributed predominantly to the transmembrane pressure (TMP) increase in the NF-MBR, while irreversible pore fouling was the major reason for UF membrane fouling. Furthermore, it was found that the biopolymers (i.e., organics with MW > 10 kDa) were the main components present in the foulants of the NF/UF membranes and RO membranes. The analysis indicated that the NF-MBR + RO system at recovery of 90% has comparable energy consumption as the UF-MBR + RO system at recovery of 75%. Our findings proved the feasibility of the NF-MBR + RO for water reclamation at a high recovery rate.

Unpacking compaction: Effect of hydraulic pressure on alginate fouling

High pressure is often considered to be the cause of the high fouling propensity of reverse osmosis (RO) relative to forward osmosis (FO). Several experimental studies have shown that alginate fouling is more susceptible to cleaning in FO than in RO, but the theory that foulant compaction causes this disparity seems to be contradicted by the incompressibility of alginate hydrogels. In addition, the effect of hydraulic pressure on fouling in osmotic membrane desalination has never been experimentally isolated, because fixed-flux comparisons at different hydraulic pressures require different draw solution osmotic pressures. In this study, a new approach to isolating the effect of hydraulic pressure on alginate fouling and cleaning is introduced: operating FO with elevated but equal feed and draw hydraulic pressures. The same concentration of sodium chloride is used as the draw solution in all trials to eliminate possible effects of draw solution osmotic pressure on membrane fouling or cleaning. Theoretical modeling of the effect of alginate foulant compaction on flux reveals that foulant compaction should accelerate flux decline with low salinity feeds but retard flux decline at high salinity. However, in low-salinity alginate fouling trials, for which foulant compaction should accelerate flux decline, the measured flux decline rate was not affected by hydraulic pressure. Furthermore, when fouled membranes were cleaned by increasing the feed velocity and reducing the draw osmotic pressure, there was no apparent relationship between hydraulic pressure and cleaning effectiveness. Finally, in situ visualization of foulant removal during the cleaning process revealed no difference in foulant removal mechanisms between different hydraulic pressures. These findings demonstrate that alginate gel compaction by high feed hydraulic pressure does not occur and suggest that other explanations should be sought for FO's high fouling resistance relative to RO.

Effects of hydraulic retention time on adsorption behaviours of EPS in an A/O-MBR: biofouling study with QCM-D

Extra-cellular polymeric substances (EPS) are a major cause of membrane fouling in membrane bioreactors (MBRs). In this study, an anoxic–oxic membrane bioreactor (A/O-MBR) was run continuously for 98 days. The runs were divided into three stages according to hydraulic retention time (HRT) (11.8, 12.5 and 14.3 h, respectively). EPS were extracted from the reactor under the different HRTs. A quartz crystal microbalance with dissipation monitoring (QCM-D) and Fourier transform infrared (FT–IR) were used to study the adherence layer structures and the adsorption behaviours of EPS on the membrane surface. The results indicated that the removal rate of TN was more susceptible to HRT than NH3-N. The observations in the QCM-D suggested that at the lowest HRT (11.8 h), the structure of the adsorption layer is loose and soft and the fluidity was better than for HRTs of 12.5 or 14.3 h. It is likely one of the major reasons for the rapidly blocking of the membrane pores. Furthermore, the higher EPS adherence as analyzed in the QCM-D and EPS concentration could induce a higher osmotic pressure effect, leading to a rapid membrane-fouling rate.

Identification of the inactivating factors and mechanisms exerted on MS2 coliphage in concentrated synthetic urine

Volume reduction (condensation) is a key for the practical usage of human urine as a fertilizer because it enables the saving of storage space and the reduction of transportation cost. However, concentrated urine may carry infectious disease risks resulting from human pathogens frequently present in excreta, though the survival of pathogens in concentrated urine is not well understood. In this study, the inactivation of MS2 coliphage, a surrogate for single-stranded RNA human enteric viruses, in concentrated synthetic urine was investigated. The infectious titer reduction of MS2 coliphage in synthetic urine samples was measured by plaque assay, and the reduction of genome copy number was monitored by reverse transcription-quantitative PCR (RTqPCR). Among chemical-physical conditions such as pH and osmotic pressure, uncharged ammonia was shown to be the predominant factor responsible for MS2 inactivation, independently of urine concentration level. The reduction rate of the viral genome number varied among genome regions, but the comprehensive reduction rate of six genome regions was well correlated with that of the infectious titer of MS2 coliphage. This indicates that genome degradation is the main mechanism driving loss of infectivity, and that RT-qPCR targeting the six genome regions can be used as a culture-independent assay for monitoring infectivity loss of the coliphage in urine. MS2 inactivation rate constants were well predicted by a model using ion composition and speciation in synthetic urine samples, which suggests that MS2 infectivity loss can be estimated solely based on the solution composition, temperature and pH, without explicitly accounting for effects of osmotic pressure.

Simultaneous determination of eight B-vitamins in rat intestinal perfusate to identify effects of osmotic pressures on absorptions.

A rapid and accurate HPLC-DAD method was developed and validated to simultaneously determine eight B-vitamins (VBs, namely thiamine, riboflavin, niacinamide, calcium pantothenic, pyridoxine, biotin, folic acid and cyanocobalamin) and phenolsulfonphthalein in rat intestinal perfusate. Chromatographic separation was achieved using an Inertsil ODS-3 column (250 × 4.6 mm i.d., 5 μm) at a temperature of 40°C. Gradient elution mode was applied at the flow rate of 1.0 mL/min with the mobile phase of acetonitrile-30 mm K2 HPO4 (pH 5.80). The method was successfully applied to identify the effects of osmotic pressures on the absorption of the VBs. The absorption profiles of single and mixed VBs were also compared. Histological section technology was applied to observe the microstructure of small bowel mucosa after perfusion. The results indicated that each compound possessed a better absorption profile under isotonic conditions than under hypotonic or hypertonic conditions for single or mixed solutions. Compared with single VBs, better absorptions in mixed VBs were observed. Pathological tissue slice test suggested that hypotonic and hypertonic solutions changed or damaged the microstructure of mucosa to varying degrees. Taken together, the investigations indicated that multi-VBs administered orally under isotonic condition could generate fast and complete absorption profiles for VBs.

Effect of osmotic pressure on spermatozoa characteristics of cryopreserved buffalo bull (Bubalus bubalis) semen

ABSTRACTThis study was designed to determine the effect of different osmotic pressures on spermatozoa characteristics of cryopreserved buffalo bull semen using tris egg yolk extender (TEYE). Semen of Nili-Ravi buffalo bulls (n = 4) housed at a semen production unit was collected at weekly intervals for 10 weeks. Three solutions of tris-citric acid-fructose with osmotic pressures of 255, 275 and 295 mOsm/kg were used in extender preparation. Semen straws containing 20 × 106 spermatozoa were processed and stored at −196°C in liquid nitrogen. Post-thaw analyses of spermatozoa included motility, viability, acrosomal integrity, plasma membrane integrity, DNA integrity and lipid per-oxidation. Significantly higher (P < .05) sperm motility, acrosomal and DNA integrity were recorded at osmotic pressures of 275 and 295 mOsm/kg compared to 255 mOsm/kg. However, differences in spermatozoa viability, plasma membrane integrity and lipid per-oxidation were non-significant among three osmotic pressures. It is concluded that osmotic pressure of the solution used in extender preparation plays an important role in post-thaw quality of cryopreserved buffalo bull semen.

Application of 32 Factorial D-Optimal Design in Formulation of Porous Osmotic Pump Tablets of Ropinirole; An Anti-Parkinson’s Agent

: Objective: In the present study, our objective was to develop porous osmotic pump tablets of an anti-Parkinson’s agent, ropinirole. It also aimed to demonstrate the applicability of factorial designs and there by a statistical optimization in developing a controlled drug releasing device. Methods: A 32 factorial D-optimal study design was adopted to identify an optimized formulation. Tablets were prepared by direct compression using varying amounts of microcrystalline cellulose (MCC) and sodium chloride, followed by coating with semipermeable membrane of cellulose acetate (CA) containing polyethylene glycol (PEG) 400 as a pore former. The plasticity of the membranes was adjusted using castor oil. All the formulations were evaluated for various physical parameters including in vitro drug release and the effect of osmogent and pore former were also studied. Drug release kinetics studies such as zero order, first order and Korsmeyer Peppas were carried out and compared. ANOVA in drug release of all the formulations were determined. Formulations were optimized by using numerical optimization technique to achieve a controlled zero order release of ropinirole for 12 hours. Results: Drug release from the optimized formulation containing 20%w/w of PEG, without osmogent was not significantly affected by change in pH or agitation of the dissolution medium. The mechanism of drug release was further confirmed by studying the effect of osmotic pressure on drug release. Conclusion: The porous osmotic pump tablets of ropinirole can provide prolonged, controlled and GI environment-independent drug release. A software based optimization is helpful and reliable in developing porous osmotic pump tablets.Key words: Osmotic pump, Ropinirole, Controlled release, Factorial design, D-optimal design, Optimization.

Starch-carrageenan interactions in aqueous media: Role of each polysaccharide chemical and macromolecular characteristics

In this study, physico-chemical interactions between carrageenan chains and starch granules were investigated through methylene blue spectrophotometric method by considering the chemical and macromolecular characteristics of each polysaccharide. For that purpose, various proportions of κ-, ι- or λ- carrageenan and different types of starches (native, chemically modified, “deproteinated” or not) were used. Blue dextran spectrophotometric method was also developed to evaluate the quantity of water absorbed by starch granules and hereby their swelling behavior allowing to determine the accurate amount of trapped carrageenan. It was demonstrated that the interactions between starch granules and carrageenan chains are not strongly impacted by starch chemical modifications. However, experiments with “deproteinated” starch showed that starch endogenous proteins can play a part in starch-carrageenan interactions. Three coexisting phenomena were found to occur when starch is pasted together with carrageenan. That consisted in: (i) partial penetration of carrageenan in starch granules, (ii) partial “exclusion” of carrageenan by starch granules and (iii) a predominant adsorption of carrageenan on starch granules due mainly to osmotic pressure effect. Moreover, the electrostatic interactions between starch endogenous proteins and carrageenan chains played a minor role. As expected, the interaction level appeared to be strongly depending from the carrageenan charge density: the lower the charge, the higher the interaction between starch and carrageenan, independently from starch chemical characteristics. Furthermore, it seemed that the rheological properties of the mixed systems are preferentially driven by the partial “exclusion” phenomenon as well as the role of starch granules as “filler”.

Stability of equilibria of a two-phase Stokes-osmosis problem

Within the framework of variational modelling we derive a two-phase moving boundary problem that describes the motion of a semipermeable membrane separating two viscous liquids in a fixed container. The model includes the effects of osmotic pressure and surface tension of the membrane. For this problem we prove that the manifold of steady states is locally exponentially attractive.

Investigating and modeling the combined effects of pH and osmotic pressure on seed germination for use in phytoactivity and allelopathic research

Phytoactivity and allelopathic studies are heavily dependent on germination bioassays of water solutions of allelochemical(s), which necessarily imply that pH and osmotic pressure vary among treatments and between treatments and controls and are therefore a confounding factor in the assessment of seed germination responses to allelochemical(s). When the contribution of pH and osmotic pressure to seed germination responses is considered in experimental designs their effects are almost without exceptions examined separately being assumed, without any evidences, that pH and osmotic pressure act independently on seed germination responses. The objectives of this work were to examine experimentally such assumption using wheat, lettuce, and subterranean clover cultivars to evaluate and model the combined effects on germination of pH and osmotic pressure in the range between 3.0–6.0 and 0–100 mOsmol kg−1, respectively. Empirical equations are fitted, discussed, and the need to consider the simultaneous effects of pH and osmotic pressure firmly established. Finally, the use of the equations fitted and its impact on conclusions is exemplified in a dose-response bioassay of water extracts of Cistus ladanifer on seed germination using subterranean clover as target species where hormesis was found before allelochemical effects were corrected for pH and osmotic pressure values of control and extracts.