Modelling and optimization of inactivation of Lactobacillus plantarum by pulsed electric field treatment

AIMS: The effect of critical pulsed electric field (PEF) process parameters, such as electric field strength, pulse length and number of pulses, on inactivation of Lactobacillus plantarum was investigated. METHODS AND RESULTS: Experiments were performed in a pH 4.5 sodium phosphate buffer having a conductivity of 0.1 S m-1, using a laboratory-scale continuous PEF apparatus with a co-linear treatment chamber. An inactivation model was developed as a function of field strength, pulse length and number of pulses. Based on this inactivation model, the conditions for a PEF treatment were optimized with respect to the minimum energy required to obtain a certain level of inactivation. It was shown that the least efficient process parameter in the range investigated was the number of pulses. The most efficient way to optimize inactivation of Lact. plantarum was to increase the field strength up to 25.7 kV cm-1, at the shortest pulse length investigated, 0.85 micros, and using a minimum number of pulses. The highest inactivation of Lact. plantarum at the lowest energy costs is obtained by using the equation: E=26.7tau0.23, in which E is the field strength and tau the pulse length. An optimum is reached by substituting tau with 5.1. CONCLUSIONS: This study demonstrates that the correct choice of parameters, as predicted by the model described here, can considerably improve the PEF process. SIGNIFICANCE AND IMPACT OF THE STUDY: The knowledge gained in this study improves the understanding of the limitations and opportunities of the PEF process. Consequently, the advantage of the PEF process as a new option for non-thermal decontamination can be better utilized.     

Molecular monitoring of inactivation efficiencies of bacteria during pulsed electric field treatment of clinical wastewater

Aims: The applicability of an alternative wastewater disinfection concept based on the pulsed electric field (PEF) treatment is tested with molecular biology techniques using clinical wastewaters. Methods and Results: Hospital wastewater was treated with the PEF technology. The inactivation efficiencies of bacteria were successfully monitored with real‐time polymerase chain reaction (PCR). As the differentiation between living and dead bacterial cells is important for the determination of the disinfection efficiency, propidium monoazide (PMA) was applied. PMA selectively penetrates cells with compromised membranes and intercalates into the DNA inhibiting a subsequent PCR amplification. The rates of reduction were examined for specific pathogens and wastewater populations using PCR‐denaturing gradient gel electrophoresis. The results showed that the main part of the bacterial population could be inactivated efficiently with the PEF treatment. Moreover, it was demonstrated that naturally occurring nuclease activities were not affected by the PEF treatment in contrast to a thermal treatment. Conclusions: The results indicated that the PEF treatment is an appropriate alternative disinfection concept for the treatment of clinical wastewaters and surpass the disadvantages of other disinfection methods. Significance and Impact of the Study: With the use of propidium monoazide for live–dead distinction, a new concept could be developed for the evaluation of disinfection methods.     

Comparison of membrane electroporation and protein denature in response to pulsed electric field with different durations

In this paper, we compared the minimum potential differences in the electroporation of membrane lipid bilayers and the denaturation of membrane proteins in response to an intensive pulsed electric field with various pulse durations. Single skeletal muscle fibers were exposed to a pulsed external electric field. The field‐induced changes in the membrane integrity (leakage current) and the Na channel currents were monitored to identify the minimum electric field needed to damage the membrane lipid bilayer and the membrane proteins, respectively. We found that in response to a relatively long pulsed electric shock (longer than the membrane intrinsic time constant), a lower membrane potential was needed to electroporate the cell membrane than for denaturing the membrane proteins, while for a short pulse a higher membrane potential was needed. In other words, phospholipid bilayers are more sensitive to the electric field than the membrane proteins for a long pulsed shock, while for a short pulse the proteins become more vulnerable. We can predict that for a short or ultrashort pulsed electric shock, the minimum membrane potential required to start to denature the protein functions in the cell plasma membrane is lower than that which starts to reduce the membrane integrity. Bioelectromagnetics 34:253–263, 2013. © 2012 Wiley Periodicals, Inc.     

Application of a pulsed electric field to cross-flow ultrafiltration of protein solution

The application of pulsed electric field was investigated in the crossflow ultrafiltration of BSA (bovine serum albumn) to economize the application time of electric current as well as to avoid inherent problems of long-term application of electric field. During the application of various cyclic patterns of pulsed electric current, the averaged filtration flowrate and the degree of concentration were maintained higher than those obtained in the absence of electric current application. The temperature increase, pH change, and BSA loss by electrodeposition were all negligible during the operation. The averaged filtration flowrate increased as the ON/OFF duration ratio of electric current was higher and as the period of ON/OFF cycle was shorter. The re-establishment of concentration polarization was dependent to the duration of current OFF state and, therefore, a longer duration of OFF state was not favorable in maintaining higher filtration flow rate. Although the averaged filtration flowrate was enhanced as the magnitude of electric current increased, the flowrate enhancement became smaller as the magnitude of current increased because there exists a current value above which the degree of electrokinetic depolarization is no further improved.     

Changes in biosynthesis of exopolysaccharide in Lactococcus lactis subspecies cremoris treated by moderate pulsed electric field treatment

Metabolome analysis and physicochemical analyses were executed with cell extracts of a Lactococcus lactis subspecies cremoris strain treated by moderate pulsed electric field (PEF) to elucidate the mechanism of enhanced production of exopolysaccharide (EPS) by the treatment. Metabolome analysis by capillary electrophoresis time of flight mass spectrometry annotated 224 metabolites from the cytoplasmic extract of the strain, which, however, showed no significant changes in metabolites related to the EPS production. Electron microscopic observation and chemical analysis of undecaprenoids as carrier of EPS biosynthetic intermediates suggested that PEF treatment dissociated immature EPSs from the intermediates due to the focal electro-condensation of hydrogen ions at the cell surface. Thus, liberated undecaprenyl phosphates were recycled efficiently, which resulted in mass increase of EPS with smaller molecular weight. The study suggested the feasibility of moderate PEF treatment as a food processing technique and revealed the mechanism of enhanced production of EPS by the treatment.     

Variation in resistance of natural isolates of Staphylococcus aureus to heat, pulsed electric field and ultrasound under pressure

AIMS: To study and compare the resistance of 15 Staphylococcus aureus isolates to heat, pulsed electric field (PEF) and ultrasound (UW) under pressure (manosonication, MS). METHODS AND RESULTS: Survival curves to heat (58 degrees C), to PEF (22 kV cm(-1), 2 micros square wave pulses) and to UW under pressure (117 microm, 20 kHz, 200 kPa) were obtained and inactivation parameters (decimal reduction times for heat and UW under pressure, and b-values for PEF) were calculated. A wide resistance variation to heat treatment, but not to PEF and MS, was observed amongst the 15 strains. CONCLUSIONS: There was no relationship between the resistances to the three physical agents studied. Staphylococcus aureus was relatively resistant to MS but sensitive to PEF. Heat resistance varied with strain and was positively correlated to carotenoid pigment content. SIGNIFICANCE AND IMPACT OF THE STUDY: Results would help in defining safe food preservation processes. Care should be taken to choose the most adequate strain of S. aureus to model food preservation processing.     

Antimicrobial effect and shelf-life extension by combined thermal and pulsed electric field treatment of milk

Aims:  The impact of a combined hurdle treatment of heat and pulsed electric fields (PEF) was studied on native microbiota used for the inoculation of low-fat ultra-high temperature (UHT) milk and whole raw milk. Microbiological shelf-life of the latter following hurdle treatment or thermal pasteurization was also investigated.
Methods and Results:  UHT milk was preheated to 30°C, 40°C or 50°C over a 60-s period, pulsed for 50  μ s or 60  μ s at a field strength of 40 kV cm⁻¹ or for 33  μ s at 50 kV cm⁻¹. Heat and PEF reduced the microbial count by a maximum of 6·4 log in UHT milk (50°C; 50 kV cm⁻¹, 33  μ s) compared to 6·0 log ( P  ≥ 0·05) obtained by thermal pasteurization (26 s, 72°C). When raw milk was treated with a combination of hurdles (50°C; 40 kV cm⁻¹, 60  μ s) a 6·0 log inactivation of microbiota was achieved and microbiological milk shelf-life was extended to 21 days under refrigeration (4°C) vs 14 days in thermally pasteurized milk. Native microbiota was decreased by 6·7 log following conventional pasteurization.
Conclusions:  The findings suggest that heat and PEF achieved similar inactivation of native microbiota in milk and longer stabilization of microbiological shelf-life than thermal pasteurization.
Significance and Impact of the Study:  A hurdle approach of heat and PEF could represent a valid milk processing alternative to conventional pasteurization. Hurdle treatment might also preserve native milk quality better due to less thermal exposure.     

Surface structure of ionic liquids under an external electric field

Abstract

Surface structure and properties play an important role in many applications of ionic liquids (ILs). ILs can form unique surface structures that are very different from the bulk. In imidazolium-based ILs, for example, polar groups form ordered layer structure, while cationic alkyl chains are bundled together and point out from the surface. In many applications, ILs work under an external electric field. However, the effect of external electric field on the surface structure of ILs is still not clear. Here by using coarse-grained molecular dynamics simulation, we found that an electric field as strong as 1 V/nm is required to alter the surface structure of 1-dodecyl-3-methylimidazolium nitrate. Under a strong external electric field, layered structure disappears, and instead a sparser, more homogeneous and less orientationally ordered surface develops.     

A numerical analysis of the aortic blood flow pattern during pulsed cardiopulmonary bypass

In the modern era, stroke remains a main cause of morbidity after cardiac surgery despite continuing improvements in the cardiopulmonary bypass (CPB) techniques. The aim of the current work was to numerically investigate the blood flow in aorta and epiaortic vessels during standard and pulsed CPB, obtained with the intra-aortic balloon pump (IABP). A multi-scale model, realized coupling a 3D computational fluid dynamics study with a 0D model, was developed and validated with in vivo data. The presence of IABP improved the flow pattern directed towards the epiaortic vessels with a mean flow increase of 6.3% and reduced flow vorticity.     

Molecular dynamics simulation of liquid water under the influence of an external electric field

Molecular dynamics simulations of liquid water were performed at 258K and a density of 1.0?g/cm³ under various applied external electric field, ranging 0～10¹⁰?V/m. The influence of external field on structural and dynamical properties of water was investigated. The simple point charge (SPC) model is used for water molecules. An enhancement of the water hydrogen bond structure with increasing strength of the electric field has been deduced from the radial distribution functions and the analysis of hydrogen bonds structure. With increasing field strength, water system has a more perfect structure, which is similar to ice structure. However, the electrofreezing phenomenon of liquid water has not been detected since the self-diffusion coefficient was very large. The self-diffusion coefficient decreases remarkably with increasing strength of electric field and the self-diffusion coefficient is anisotropic.     

A simple fluid–structure coupling algorithm for the study of the anastomotic mechanics of vascular grafts

Vascular anastomoses constitute a main factor in poor graft performance due to mismatches in distensibility between the host artery and the graft. This work aims at computational fluid–structure investigations of proximal and distal anastomoses of vein grafts and synthetic grafts. Finite element and finite volume models were developed and coupled with a user-defined algorithm. Emphasis was placed on the simplicity of the coupling algorithm. An artery and vein graft showed a larger dilation mismatch than an artery and synthetic graft. The vein graft distended nearly twice as much as the artery while the synthetic graft displayed only approximately half the arterial dilation. For the vein graft, luminal mismatching was aggravated by development of an anastomotic pseudo-stenosis. While this study focused on end-to-end anastomoses as a vehicle for developing the coupling algorithm, it may serve as useful point of departure for further investigations such as other anastomotic configurations, refined modelling of sutures and fully transient behaviour.     

Dynamic changes of [Ca2+]i in cerebellar granule cells exposed to pulsed electric fields

Intracellular free Ca2+ concentration ([Ca2+]i) in embryonic chick cerebellar granule cells loaded with fluo-3/AM and exposed to a single pulsed electric field was investigated using a confocal laser scanning microscope and fluorescent microscope equipped with CCD video imaging system.The results showed that [Ca2+]i increased immediately and rose to the peak rapidly as the cells exposed to a single pulsed electric field.The amplitude and rate of the increases of [Ca2+]i depend on the intensity of external electric field.In the presence of Ca2+ chelant EGTA or Ca2+ channels blocker La3+ in the pulsation solutions,the increase of [Ca2+]i was still observable.It was also observed that [Ca2+]i of different intracellular areas in the cell elevated simultaneously while the peak of the increase of [Ca2+]i in the poles of the cell preceded to the peak in its somata and recovered to a plateau within a short time.     

Pulsed electric field stimulates plant secondary metabolism in suspension cultures of Taxus chinensis

The effects of pulsed electric field (PEF) on growth and secondary metabolite production by plant cell culture were investigated by using suspension cultures of Taxus chinensis as a model system. Cultured cells in different growth phases were exposed to a PEF (50 Hz, 10 V/m) for various periods of time. A significant increase in intracellular accumulation of taxuyunnanine C (Tc), a bioactive secondary metabolite, was observed by exposing the cells in the early exponential growth phase to a 30-min PEF. The Tc content (i.e., the specific production based on dry cell weight) was increased by 30% after exposure to PEF, without loss of biomass, compared with the control. The combination of PEF treatment and sucrose feeding proved useful for improving secondary metabolite formation. Production levels of reactive oxygen species, extracellular Tc, and phenolics were all increased, whereas cell capacitance was decreased with PEF treatment. The results show that PEF induced a defense response of plant cells and may have altered the cell/membrane's dielectric properties. PEF, an external stimulus or stress, is proposed as a promising new abiotic elicitor for stimulating secondary metabolite biosynthesis in plant cell cultures.     

Influence of flow rate and scaffold pore size on cell behavior during mechanical stimulation in a flow perfusion bioreactor

Mechanically stimulating cell-seeded scaffolds by flow-perfusion is one approach utilized for developing clinically applicable bone graft substitutes. A key challenge is determining the magnitude of stimuli to apply that enhances cell differentiation but minimizes cell detachment from the scaffold. In this study, we employed a combined computational modeling and experimental approach to examine how the scaffold mean pore size influences cell attachment morphology and subsequently impacts upon cell deformation and detachment when subjected to fluid-flow. Cell detachment from osteoblast-seeded collagen-GAG scaffolds was evaluated experimentally across a range of scaffold pore sizes subjected to different flow rates and exposure times in a perfusion bioreactor. Cell detachment was found to be proportional to flow rate and inversely proportional to pore size. Using this data, a theoretical model was derived that accurately predicted cell detachment as a function of mean shear stress, mean pore size, and time. Computational modeling of cell deformation in response to fluid flow showed the percentage of cells exceeding a critical threshold of deformation correlated with cell detachment experimentally and the majority of these cells were of a bridging morphology (cells stretched across pores). These findings will help researchers optimize the mean pore size of scaffolds and perfusion bioreactor operating conditions to manage cell detachment when mechanically simulating cells via flow perfusion.     

正常乳腺温度分布的有限元分析

基于正常乳腺的解剖学结构和生理学特征,建立了一个乳腺组织多维热传递模型。该模型考虑了代谢产热、血液灌注和动静脉血管与组织间热的相互作用,采用有限元分析方法求解热传导方程,数值模拟正常乳腺的稳态温度分布,着重研究血液灌注和代谢产热对正常乳腺组织温度分布的影响。研究结果可为乳腺疾病的热图像分析提供重要参考。     

Water transport in xylem conduits with ring thickenings

Helical or annular wall thickenings are not only present in protoxylem, but may also he a feature of the tracheids or vessel elements of secondary wood. The frequency of their occurrence tends to be a function of climatic factors and conduit diameter. In order to obtain a functional explanation for these structures, the hydrodynamic behaviour of xylem conduits with various patterns of annular wall thickenings was investigated numerically using a commercial CFD (Computational Fluid Dynamics) package. The fluid flow phenomena are presented in detail. The calculations show that the developing pressure gradient of the structures with corrugated walls is in each case lower than that of a smooth pipe with a diameter corresponding to the distance between two opposite thickenings. Furthermore, complex flow patterns with circulation zones between the thickenings develop which are dependent on the geometry of the wall. It may be hypothesized that these circulation zones influence the lateral water flow. The results are discussed with regard to the relationships between the water conduction function of the xylem and ecological factors.     

Evaluation of dentinal fluid flow behaviours: a fluid-structure interaction simulation

This study uses the fluid-structure interaction (FSI) method to investigate the fluid flow in dental pulp. First, the FSI method is used for the biomechanical simulation of dental intrapulpal responses during force loading (50, 100 and 150 N) on a tooth. The results are validated by comparison with experimental outcomes. Second, the FSI method is used to investigate an intact tooth subjected to a mechanical stimulus during loading at various loading rates. Force loading (0–100 N) is applied gradually to an intact tooth surface with loading rates of 125, 62.5, 25 and 12.5 N/s, respectively, and the fluid flow changes in the pulp are evaluated. FSI analysis is found to be suitable for examining intrapulpal biomechanics. An external force applied to a tooth with a low loading rate leads to a low fluid flow velocity in the pulp chamber, thus avoiding tooth pain.