Local Transmembrane Research Articles

Alternating flow direction improves chemical cleaning efficiency in hollow fibre membranes following skim milk microfiltration

With efficient membrane cleaning remaining a challenge, novel approaches have promising potential to enhance cleaning efficiency. This study examined the concept of feed side cyclical and permanent flow reversal during membrane cleaning, to which so far it has not been applied to. Experiments were conducted in a hollow fibre microfiltration membrane with lengths of 0.5 m and 1.0 m using skim milk for deposit formation, NaOH as a cleaning agent, and flux recovery ratio and protein removal for cleaning evaluation. Overall, flow reversal was found to significantly increase cleaning success. Due to the length-dependent pressure drop and flow reversal, the local transmembrane pressure at the initial feed inlet ΔpTM, inlet and thus the ratio of flow velocity to ΔpTM, inlet was increased compared to steady forward flow cleaning. This proved beneficial as it combines high shear stress with low ΔpTM, inlet where fouling was most pronounced after filtration.

Intramodule pressure profiles and protein accumulation during tangential flow filtration.

Tangential flow filtration (TFF) through a 30 kDa nominal molecular weight cut-off (MWCO) ultrafiltration membrane is widely employed to concentrate purified monoclonal antibodies (mAbs) to levels required for their formulation into injectable biologics. While TFF has been used to remove casein from milk for cheese production for over 35 years, and in pharmaceutical manufacture of biotherapeutic proteins for 20 years, the rapid decline in filtration rate (i.e., flux) at high protein concentrations is a limitation that still needs to be addressed. This is particularly important for mAbs, many of which are 140-160 kDa immunoglobulin G (IgG) type proteins recovered at concentrations of 200 mg/mL or higher. This work reports the direct measurement of local transmembrane pressure drops and off-line confocal imaging of protein accumulation in stagnant regions on the surface of a 30 kDa regenerated cellulose membrane in a flat-sheet configuration widely used in manufacture of biotherapeutic proteins. These first-of-a-kind measurements using 150 kDa bovine IgG show that while axial pressure decreases by 58 psi across a process membrane cassette, the decrease in transmembrane pressure drop is constant at about 1.2 psi/cm along the 20.7 cm length of the membrane. Confocal laser scanning microscopy of the membrane surface at the completion of runs where retentate protein concentration exceeds 200 mg/mL, shows a 50 μm thick protein layer is uniformly deposited. The localized measurements made possible by the modified membrane system confirm the role of protein deposition on limiting ultrafiltration rate and indicate possible targets for improving membrane performance.

Wide-field fluorescence lifetime imaging of neuron spiking and subthreshold activity in vivo.

The development of voltage-sensitive fluorescent probes suggests fluorescence lifetime as a promising readout for electrical activity in biological systems. Existing approaches fail to achieve the speed and sensitivity required for voltage imaging in neuroscience applications. We demonstrated that wide-field electro-optic fluorescence lifetime imaging microscopy (EO-FLIM) allows lifetime imaging at kilohertz frame-acquisition rates, spatially resolving action potential propagation and subthreshold neural activity in live adult Drosophila. Lifetime resolutions of <5 picoseconds at 1 kilohertz were achieved for single-cell voltage recordings. Lifetime readout is limited by photon shot noise, and the method provides strong rejection of motion artifacts and technical noise sources. Recordings revealed local transmembrane depolarizations, two types of spikes with distinct fluorescence lifetimes, and phase locking of spikes to an external mechanical stimulus.

Efficacy optimization of low frequency microbubble-mediated sonoporation as a drug delivery platform to cancer cells

Ultrasound insonation of microbubbles can be used to form pores in cell membranes and facilitate the local trans-membrane transport of drugs and genes. An important factor in efficient delivery is the size of the delivered target compared to the generated membrane pores. Large molecule delivery remains a challenge, and can affect the resulting therapeutic outcomes. To facilitate large molecule delivery, large pores need to be formed. While ultrasound typically uses megahertz frequencies, it was recently shown that when microbubbles are excited at a frequency of 250 kHz (an order of magnitude below the resonance frequency of these agents), their oscillations are significantly enhanced as compared to the megahertz range. Here, to promote the delivery of large molecules, we suggest using this low frequency and inducing large pore formation through the high-amplitude oscillations of microbubbles. We assessed the impact of low frequency microbubble-mediated sonoporation on breast cancer cell uptake by optimizing the delivery of 4 fluorescent molecules ranging from 1.2 to 70 kDa in size. The optimal ultrasound peak negative pressure was found to be 500 kPa. Increasing the pressure did not enhance the fraction of fluorescent cells, and in fact reduced cell viability. For the smaller molecule sizes, 1.2 kDa and 4 kDa, the groups treated with an ultrasound pressure of 500 kPa and MB resulted in a fraction of 58% and 29% of fluorescent cells respectively, whereas delivery of 20 kDa and 70 kDa molecules yielded 10% and 5%, respectively. These findings suggest that low-frequency (e.g., 250 kHz) insonation of microbubbles results in high amplitude oscillation in vitro that increase the uptake of large molecules. Successful ultrasound-mediated molecule delivery requires the careful selection of insonation parameters to maximize the therapeutic effect by increasing cell uptake.

Investigation of instantaneous and local transmembrane pressure in rotating and vibrating filtration (RVF) module: Comparison of three impellers

The instantaneous and local pressure at membrane surface was experimentally investigated in a dynamic filtration module, named Rotating and Vibrating Filtration (RVF) module. The present paper focuses mainly on the pressure fluctuations in turbulent regime. To this end, the instantaneous pressure is decomposed into its time-averaged and fluctuating quantities using Statistical Analysis (SA), Probability Distribution Function (PDF) and Fast Fourier Transform (FFT). The effects of back pressure, flowrate, rotation frequency and radial position at the membrane on the magnitude of the pressure fluctuations are studied for three different impellers (Imp 1, 2 and 3). For mixing pressure, Imp 2 (6 blades) exhibits a larger core velocity coefficient than Imp 1 and Imp 3 (3 blades). For pressure fluctuation, the extracted variables from SA (standard deviations), PDF (peak-to-peak values) and FFT (amplitudes) confirm that the magnitude of Imp 1 > Imp 3 > Imp 2. Considering SA at 20 Hz, standard deviation of Imp 1 exceeds 100 mbar (up to 25% of TMP), while these values are negligible (<10%) for Imp 2 and 3. After FFT, the dominant frequency identified with Imp 1 is equal to 3 times the rotation frequency (3 N). Conversely, different frequencies (6 N, 3 N and N) exhibiting low amplitude are observed for Imp 2 and 3. Based on the PDF modelling, periodic and random contributions are extracted by deconvolution. Then, the empirical correlations are established to estimate their intensities as a function of rotation frequency and radial position. A “resonance frequency” of 21.1 Hz is clearly identified with Imp 1.

Centrifugal reverse osmosis (CRO) − a novel energy-efficient membrane process for desalination near local thermodynamic equilibrium

Seawater reverse osmosis (SWRO) can address the global water crisis that is being exacerbated by climate change, industrialization, pollution, and population growth. However, conventional single-stage RO (SSRO) incurs a high energy cost owing to pressurizing the feed to that required for the desired recovery. This cost could be reduced if SWRO were operated at a progressively increasing pressure dictated by thermodynamic equilibrium for the local transmembrane salt-concentration difference. This is approached by the semi-batch closed-circuit RO (CCRO) process that progressively increases the transmembrane pressure (TMP); however, it incurs cumulative energy losses until the pressure is reached for the desired recovery. The centrifugal reverse osmosis (CRO) process uses rotation of a module to create a centrifugal force that increases the TMP differentially with increasing radial distance from the axis-of-symmetry. Unlike CCRO, CRO is a continuous desalination process. A mathematical model and preliminary design are advanced for a CRO module. CRO can reduce the net specific energy consumption relative to SSRO by 31% for 50% recovery at 56 bar from a 35 g/L seawater feed. A TMP progressively increasing from 28 to 56 bar can be achieved with a CRO module having a radius of 0.72 m rotating at 1000 rpm.

Methodology to investigate instantaneous and local transmembrane pressure within Rotating and Vibrating Filtration (RVF) module

• Semi-empirical correlations to estimate mixing and pumping powers. • Determination of local core velocity coefficients and mixing pressure. • Comparison of methods to analysis local fluctuating pressure. • Reconstruction of pressure distribution with periodic and random functions. Dynamic filtration exhibits high performances by generating wall shear stress (tangential to membrane) and additional pressure (normal to membrane) by the mechanical movements, such as rotating, oscillating or vibrating systems. Rotating and Vibrating Filtration (RVF) module includes rotating flat blades impellers which generate high and fluctuating shear stress and pressure at the membrane surface. To understand performances in turbulent regime and to optimise the operating conditions, global parameters (power consumption, pressure drop) and driving forces (mean, instantaneous and local pressure at the membrane surface) were characterised. For global approach, friction and mixing power in the RVF module were described by semi-empirical correlations. Euler number correlations were integrated based on feeding and mixing conditions. The balance between nominal power and thermal dissipation was reported. On the other hand, the mechanical power calculated with the empirical correlation of local shear stress was underestimated. For semi-local and local approaches, the local pressure at the membrane surface was measured with a specially designed and instrumented porous substrate. Mean radial pressure and core velocity coefficients were quantified versus flowrate and mixing rate. The core velocity coefficient decreases with mixing rate and radius up to a plateau value close to 0.6. For fluctuating component, pressure oscillation and its amplitude were treated by statistical analysis, probability distribution function and Fast Fourier transform. These methods show similar results with maximum fluctuating intensity for a mixing rate between 15 and 30 Hz, which increases with radius. The maximum value can be obtained at the outer edge of the impeller with a relative standard deviation of over 25%. It indicates that the influence of pressure fluctuations should be carefully considered to enhance filtration performances. Pressure fluctuation distributions were accurately modelled by the convolution of periodic (sinusoid wave) and random (normal) functions. The area of intensive fluctuation was identified, in which periodic component accounts for 60% up to 97% of total energy input.

The role of fouling in optimizing direct-flow filtration module design

The local transmembrane pressure (TMP) distribution plays a crucial role in the performance of hollow-fiber microfiltration and ultrafiltration membrane devices. A direct-flow filter comprises an array of hollow fibers encased within a single module. The TMP in each hollow fiber is dependent on the spacing between neighboring fibers, and evolves with time due to pore clogging by contaminants (fouling). We consider an idealized set-up in which the fibers are undeformable and equally spaced within the device, and study the impact of the pore-blocking phenomena on the TMP during the filtration process. The model is used to evaluate the optimum inter-fiber spacing that maximizes the fluid processed either after a prescribed time or before the filter blocks and its dependence on the membrane permeability and the fouling rate. We show that significant improvements can be made on the operating efficiency of a direct-flow device through careful choice in the fiber spacing during fabrication.

Optimal Design of Dialyzers.

Several types of synthetic dialysis membranes, including polysulfone, polyethersulfone, and polyester polymer alloy membranes, have asymmetrical structures. Dialyzers with these membranes show higher water and solute transport performance because the actual membrane thickness, which is related to the water and solute transfer resistance, is quite small compared with that in membranes with a homogeneous structure. The performance of a dialyzer depends not only on membrane permeability to water and solutes, but also on flow conditions of the blood and dialysate, which are determined during dialyzer fabrication. Many types of high-flux dialyzers with high-performance membranes have a high internal filtration/backfiltration (IF/BF) flow rate. In the enhanced IF/BF dialyzer, membrane fouling occurs more readily than with the conventional dialyzer because of the high local transmembrane pressure needed to enhance the IF/BF flow rate. To select the optimal enhanced IF/BF dialyzer for individual patients, we need to balance the disadvantage of membrane fouling with the advantage of increased convective transport. Key Messages: The following principles should guide dialyzer development in the near term. (1) Dialyzers should show high performance for the removal of low-molecular-weight proteins related to certain complications under conditions of low albumin and amino acid loss. (2) Dialyzers with biocompatible membranes are required to prevent severe adverse reactions, even though the causal relationship between these reactions and some complications remains to be clarified.

High Transmembrane Voltage Raised by Close Contact Initiates Fusion Pore.

Membrane fusion lies at the heart of neuronal communication but the detailed mechanism of a critical step, fusion pore initiation, remains poorly understood. Here, through atomistic molecular dynamics simulations, a transient pore formation induced by a close contact of two apposed bilayers is firstly reported. Such a close contact gives rise to a high local transmembrane voltage that induces the transient pore formation. Through simulations on two apposed bilayers fixed at a series of given distances, the process in which two bilayers approaching to each other under the pulling force from fusion proteins for membrane fusion was mimicked. Of note, this close contact induced fusion pore formation is contrasted with previous reported electroporation under ad hoc applied external electric field or ionic charge in-balance. We show that the transmembrane voltage increases with the decrease of the distance between the bilayers. Below a critical distance, depending on the lipid composition, the local transmembrane voltage can be sufficiently high to induce the transient pores. The size of these pores is approximately 1~2 nm in diameter, which is large enough to allow passing of neurotransmitters. A resealing of the membrane pores resulting from the neutralization of the transmembrane voltage by ions through the pores was then observed. We also found that the membrane tension can either prolong the lifetime of transient pores or cause them to dilate for full collapse. This result provides a possible mechanism for fusion pore formation and regulation of pathway of fusion process.

Direct Monolayer Packing Imbalance and Phospholipid Flip-Flop: Two Mechanisms of Local Bilayer Deformation. Application to Mitochondrial Cristae of Wild-Type and Cardiolipin-Deficient Mutant

We have shown that a spatially localized transmembrane pH gradient, produced by HCl micro-injection near the external surface of cardiolipin(CL)-containing giant unilamellar vesicles (GUVs), promotes the formation of tubules that retract after collapse of this gradient. Such tubules have morphologies similar to mitochondrial cristae. The tubulation effect is due to direct phospholipid packing modification in the outer leaflet, that is associated with protonation of CL headgroups (1). We have also compared the case of CL-containing GUVs with that of phosphatidylglycerol (PG)-containing GUVs. Local HCl micro-injection also promotes formation of tubules in the latter. However, compared to CL-containing GUVs the tubules are longer, undergo a visible pearling and have a much longer retraction time after acid micro-injection is stopped (2). We attribute these differences to an additional mechanism that increases monolayer lipid density imbalance, namely inward PG flip-flop promoted by the local transmembrane pH-gradient. Simulations using a fully non-linear membrane model, that describes the bilayer by its midsurface shape and by a lipid density field for each monolayer (3), confirm this hypothesis. Interestingly, among yeast mutants deficient in CL biosynthesis, only the crd1-null mutant, which accumulates PG, displays significant mitochondrial activity, in agreement with our data. Our work emphasizes the importance of the transmembrane proton gradient in cristae morphology as well as the salient role of specific lipids in mitochondrial function.1. Khalifat, N., N. Puff, S. Bonneau, J. B. Fournier, and M. I. Angelova. 2008. Biophys. J. 95:4924-4933.2. Khalifat, N.; Rahimi, M.; Bitbol, A. F.; Seigneuret, M.; Fournier, J. B.; Puff, N.; Arroyo, M.; Angelova, M. I. 2014 Biophys. J. 107: 879-8903. Rahimi, M. and Arroyo, M., 2012. Physical Review E, 86: 011932.

Peak I of the human auditory brainstem response results from the somatic regions of type I spiral ganglion cells: Evidence from computer modeling

Early neural responses to acoustic signals can be electrically recorded as a series of waves, termed the auditory brainstem response (ABR). The latencies of the ABR waves are important for clinical and neurophysiological evaluations. Using a biophysical model of transmembrane currents along spiral ganglion cells, we show that in human (i) the non-myelinated somatic regions of type I cells, which innervate inner hair cells, predominantly contribute to peak I, (ii) the supra-strong postsynaptic stimulating current (400 pA) and transmembrane currents of the myelinated peripheral axons of type I cells are an order smaller; such postsynaptic currents correspond to the short latencies of a small recordable ABR peak I’, (iii) the ABR signal involvement of the central axon of bipolar type I cells is more effective than their peripheral counterpart as the doubled diameter causes larger transmembrane currents and a larger spike dipole-length, (iv) non-myelinated fibers of type II cells which innervate the outer hair cells generate essentially larger transmembrane currents but their ABR contribution is small because of the small ratio type II/type I cells, low firing rates and a short dipole length of spikes propagating slowly in non-myelinated fibers. Using a finite element model of a simplified head, peaks In and II (where In is the negative peak after peak I) are found to be stationary potentials when volleys of spikes cross the external electrical conductivity barrier at the bone&dura/CSF and at the CSF/brainstem interface whereas peaks I’ and I may be generated by strong local transmembrane currents as postsynaptic events at the distal ending and the soma region of type I cells, respectively. All simulated human inter-peak times (I–I′, II–I, In–I) are close to published data.

A new model for computing the evolution of the extracellular, innercellular and membrane potential simultaneously

In order to model extracellular potentials the Line-Source method provides [1] a very powerful and accurate approach. In this method transmembane fluxes are understood as sources for potential distributions which obey the Poission-equation with zero boundary conditions in the infinity. Its solutions reveal that the waveforms are proportional to local transmembrane net currents. The extracellular potentials are comparable small in amplitude and with the aid of their second special derivatives, it is possible to interpret them as additional fluxes to be included into the cable equation having an impact on the membrane potential of surrounding cells [2]. On this basis ephaptic interactions have been studied and have been considered to play a minor role in the network activity. This modeling study provides a new approach based on the first principle of the conservation of charges which leads to a generalized form of the cable equation taking into account the full three-dimensional detail of the cell’s geometry and the presence of the extracellular potential. So instead of coupling the compartment model and the model for extracellular potentials by means of the transmembrane currents, a non-linear system of partial differential equations is solved. Because the abstraction of deviding the cell’s geometry into compartments falls apart, it is possible to examine the contribution of the precise cell geometry to the signal processing while not neglecting the impact which could result from the extracellular potential. Some simulations of propagating action potentials on ramified geometries are going to be shown as well as the resulting distributions of extracellular action potentials.

Non-invasive imaging of cardiac activation and recovery.

The sequences of activation and recovery of the heart have physiological and clinical relevance. We report on progress made over the last years in the method that images these timings based on an equivalent double layer on the myocardial surface serving as the equivalent source of cardiac activity, with local transmembrane potentials (TMP) acting as their strength. The TMP wave forms were described analytically by timing parameters, found by minimizing the difference between observed body surface potentials and those based on the source description. The parameter estimation procedure involved is non-linear, and consequently requires the specification of initial estimates of its solution. Those of the timing of depolarization were based on the fastest route algorithm, taking into account properties of anisotropic propagation inside the myocardium. Those of recovery were based on electrotonic effects. Body surface potentials and individual geometry were recorded on: a healthy subject, a WPW patient and a Brugada patient during an Ajmaline provocation test. In all three cases, the inversely estimated timing agreed entirely with available physiological knowledge. The improvements to the inverse procedure made are attributed to our use of initial estimates based on the general electrophysiology of propagation. The quality of the results and the required computation time permit the application of this inverse procedure in a clinical setting.

Correlation between local TMP distribution and bio-cake porosity on the membrane in a submerged MBR

In a submerged membrane bioreactor (SMBR) the spatial distribution of bio-cake porosity on the membrane module is very important in the detection of local membrane biofouling, which is directly related to efficient membrane-cleaning and energy consumption. In this study, the local bio-cake porosity was determined experimentally and correlated with the local flux measured by a newly developed experimental method. This exercise made it possible to suggest the optimum position of an aerator in the reactor to obtain minimal membrane biofouling. The local transmembrane pressure (TMP) on the membrane module became bigger and bigger as the distance from the local position to the suction pump became nearer and nearer owing to the pressure drop inside the hollow fiber lumen. A good correlation was found between the local TMP distribution and the local porosity of bio-cake formed on the membrane surface which was measured by a confocal laser scanning microscope (CLSM) and image analysis technique. The bio-cake formed on a local position of the membrane near the suction pump was more easily compressed by higher TMP, and thus its porosity became lower. From this local porosity distribution on the membrane the effect of aerator positions on the membrane biofouling was studied. Under identical mixing intensity, the positioning of a diffused aerator near the suction pump was the most effective location for the alleviation of the membrane fouling because the porosity of bio-cake at that location was the smallest, and thus the worst biofouling took place.

Hybrid modeling of convective laminar flow in a permeable tube associated with the cross-flow process

The confined flows in tubes with permeable surfaces are associated to tangential filtration processes (microfiltration or ultrafiltration). The complexity of the phenomena do not allow for the development of exact analytical solutions, however, approximate solutions are of great interest for the calculation of the transmembrane outflow and estimate of the concentration polarization phenomenon. In the present work, the generalized integral transform technique (GITT) was employed in solving the laminar and permanent flow in permeable tubes of Newtonian and incompressible fluid. The mathematical formulation employed the parabolic differential equation of chemical species conservation (convective–diffusive equation). The velocity profiles for the entrance region flow, which are found in the connective terms of the equation, were assessed by solutions obtained from literature. The velocity at the permeable wall was considered uniform, with the concentration at the tube wall regarded as variable with an axial position. A computational methodology using global error control was applied to determine the concentration in the wall and concentration boundary layer thickness. The results obtained for the local transmembrane flux and the concentration boundary layer thickness were compared against others in literature.

Neurons Skip a Beat during Fast Ripples

Fast ripples are EEG transients emanating from epileptic foci, but fast-ripple frequencies far exceed maximal neuronal firing rates. In this issue of Neuron, Menendez de la Prida and coworkers propose that out-of-phase firing of a subpopulation of neurons during physiological ripple activity effectively doubles the ripple frequency to produce fast ripples.

Enhancing permeate flux in a RO plant by controlling membrane fouling

Concentration polarization near the membrane surface increases local trans-membrane osmotic pressure and solute passage. It is governed by solute properties, membrane properties and hydrodynamics. Increasing the feed velocity confines the affected area to a thinner layer and usually results in reducing membrane fouling and scaling phenomena. Higher feed velocity reduces scaling and fouling. The results of fouling control by changing softening process from lime to caustic regime have been outlined in this work in the reverse osmosis (RO) units that treat brackish water for producing 400 m 3/h industrial water. Performances of these RO units were not satisfactory for many months due to high tendency in membrane fouling even following the recommendations of membrane supplier. Therefore, a new softening method has been applied successfully for enhancing the RO plant performance. This improvement in performance of RO units clearly justifies the importance of softening regime as a key parameter in fouling control.

Monophasic Action Potential Recordings in Humans

Bridging basic and clinical electrophysiology has been facilitated by monophasic action potential recordings. The electrocardiogram is a useful clinical approach in detecting abnormal repolarization, but falls short in depicting local repolarization details. The MAP waveform is a reflection of local transmembrane action potentials. We hope to convey a basic understanding of monophasic action potential recording and highlight the clinical utility in both ventricular and atrial arrhythmias.

Physiological Changes in Chronic Epileptic Rats Are Prominent in Superficial Layers of the Medial Entorhinal Area

We investigated whether the functional network properties of the medial entorhinal area (MEA) of the entorhinal cortex were altered in a rat model of chronic epilepsy that is characterized by extensive cell loss in MEA layer III. Responses were evoked in the entorhinal cortex by electrical stimulation of the subiculum in anesthetized chronic epileptic rats, 2-4 months after status epilepticus, induced by systemic kainate (KA) injections. Laminar field potentials were measured using a 16-channel silicon probe that covered all six layers of the MEA; an estimate of the local transmembrane currents was made using current source density analysis. Double-pulse stimulation of the subiculum evoked responses in deep and superficial layers of the MEA in control and KA rats. A current sink in layer I and at the border of layer I and II that was induced by antidromic activation of MEA-II, was much more prominent in KA rats with extensive neuronal loss in MEA-III than in control rats or KA rats with minor MEA-III loss. Furthermore, KA rats that displayed MEA-III loss presented a series of oscillations induced by subicular stimulation in the beta/gamma-frequency range (20-100 Hz), which were confined to superficial layers of MEA. These oscillations were never observed in control rats or KA rats with minor MEA-III loss. These results indicate that the observed alterations in the superficial MEA responses to subiculum stimulation and the occurrence of beta/gamma-oscillations are related phenomena, which are a consequence of altered and impaired inhibition within these MEA layers in chronic epileptic rats.