Liquid-liquid extraction of commercial and biosynthesized nisin by aqueous two-phase micellar systems

Nisin is a natural additive for conservation of food, and can also be used as a therapeutic agent. Nisin inhibits the outgrowth of spores, the growth of a variety of Gram-positive and Grain-negative bacteria. In this paper we present a potentially scalable and cost-effective way to purify commercial and biosynthesized in bioreactor nisin, including simultaneously removal of impurities and contaminants, increasing nisin activity. Aqueous two-phase micellar systems (ATPMS) are considered promising for bioseparation and purification purposes. Triton X-114 was chosen as the as phase-forming surfactant because it is relatively mild to proteins and it also forms two coexisting phases within a convenient temperature range. Nisin activity was determined by the agar diffusion assay utilizing Lactobacillus sake as a sensitive indicator microorganism. Results indicated that nisin partitions preferentially to the micelle rich-phase, despite the surfactant concentration tested, and its antimicrobial activity increases. The successful implementation of this peptide partitioning, from a suspension containing other compounds, represents an important step towards developing a separation method for nisin, and more generally, for other biomolecules of interest. (C) 2007 Elsevier Inc. All rights reserved.

Two-Phase Mapping for Projecting Massive Data Sets

Most multidimensional projection techniques rely on distance (dissimilarity) information between data instances to embed high-dimensional data into a visual space. When data are endowed with Cartesian coordinates, an extra computational effort is necessary to compute the needed distances, making multidimensional projection prohibitive in applications dealing with interactivity and massive data. The novel multidimensional projection technique proposed in this work, called Part-Linear Multidimensional Projection (PLMP), has been tailored to handle multivariate data represented in Cartesian high-dimensional spaces, requiring only distance information between pairs of representative samples. This characteristic renders PLMP faster than previous methods when processing large data sets while still being competitive in terms of precision. Moreover, knowing the range of variation for data instances in the high-dimensional space, we can make PLMP a truly streaming data projection technique, a trait absent in previous methods.

A Robust, Fully Adaptive Hybrid Level-Set/Front-Tracking Method for Two-Phase Flows with an Accurate Surface Tension Computation

We present a variable time step, fully adaptive in space, hybrid method for the accurate simulation of incompressible two-phase flows in the presence of surface tension in two dimensions. The method is based on the hybrid level set/front-tracking approach proposed in [H. D. Ceniceros and A. M. Roma, J. Comput. Phys., 205, 391400, 2005]. Geometric, interfacial quantities are computed from front-tracking via the immersed-boundary setting while the signed distance (level set) function, which is evaluated fast and to machine precision, is used as a fluid indicator. The surface tension force is obtained by employing the mixed Eulerian/Lagrangian representation introduced in [S. Shin, S. I. Abdel-Khalik, V. Daru and D. Juric, J. Comput. Phys., 203, 493-516, 2005] whose success for greatly reducing parasitic currents has been demonstrated. The use of our accurate fluid indicator together with effective Lagrangian marker control enhance this parasitic current reduction by several orders of magnitude. To resolve accurately and efficiently sharp gradients and salient flow features we employ dynamic, adaptive mesh refinements. This spatial adaption is used in concert with a dynamic control of the distribution of the Lagrangian nodes along the fluid interface and a variable time step, linearly implicit time integration scheme. We present numerical examples designed to test the capabilities and performance of the proposed approach as well as three applications: the long-time evolution of a fluid interface undergoing Rayleigh-Taylor instability, an example of bubble ascending dynamics, and a drop impacting on a free interface whose dynamics we compare with both existing numerical and experimental data.

Discontinuous Galerkin approximation of two-phase flows in heterogeneous porous media with discontinuous capillary pressures

We design and investigate a sequential discontinuous Galerkin method to approximate two-phase immiscible incompressible flows in heterogeneous porous media with discontinuous capillary pressures. The nonlinear interface conditions are enforced weakly through an adequate design of the penalties on interelement jumps of the pressure and the saturation. An accurate reconstruction of the total velocity is considered in the Raviart-Thomas(-Nedelec) finite element spaces, together with diffusivity-dependent weighted averages to cope with degeneracies in the saturation equation and with media heterogeneities. The proposed method is assessed on one-dimensional test cases exhibiting rough solutions, degeneracies, and capillary barriers. Stable and accurate solutions are obtained without limiters. (C) 2010 Elsevier B.V. All rights reserved.

Modeling Two-Oscillator Circadian Systems Entrained by Two Environmental Cycles

Several experimental studies have altered the phase relationship between photic and non-photic environmental, 24 h cycles (zeitgebers) in order to assess their role in the synchronization of circadian rhythms. To assist in the interpretation of the complex activity patterns that emerge from these ""conflicting zeitgeber'' protocols, we present computer simulations of coupled circadian oscillators forced by two independent zeitgebers. This circadian system configuration was first employed by Pittendrigh and Bruce (1959), to model their studies of the light and temperature entrainment of the eclosion oscillator in Drosophila. Whereas most of the recent experiments have restricted conflicting zeitgeber experiments to two experimental conditions, by comparing circadian oscillator phases under two distinct phase relationships between zeitgebers (usually 0 and 12 h), Pittendrigh and Bruce compared eclosion phase under 12 distinct phase relationships, spanning the 24 h interval. Our simulations using non-linear differential equations replicated complex non-linear phenomena, such as ""phase jumps'' and sudden switches in zeitgeber preferences, which had previously been difficult to interpret. Our simulations reveal that these phenomena generally arise when inter-oscillator coupling is high in relation to the zeitgeber strength. Manipulations in the structural symmetry of the model indicated that these results can be expected to apply to a wide range of system configurations. Finally, our studies recommend the use of the complete protocol employed by Pittendrigh and Bruce, because different system configurations can generate similar results when a ""conflicting zeitgeber experiment'' incorporates only two phase relationships between zeitgebers.

Multiphase flow in the vascular system of wood: From microscopic exploration to 3-D Lattice Boltzmann experiments

This paper provides insights into liquid free water dynamics in wood vessels based on Lattice Boltzmann experiments. The anatomy of real wood samples was reconstructed from systematic 3-D analyses of the vessel contours derived from successive microscopic images. This virtual vascular system was then used to supply fluid-solid boundary conditions to a two-phase Lattice Boltzmann scheme and investigate capillary invasion of this hydrophilic porous medium. Behavior of the liquid phase was strongly dependent on anatomical features, especially vessel bifurcations and reconnections. Various parameters were examined in numerical experiments with ideal vessel bifurcations, to clarify our interpretation of these features. (c) 2010 Elsevier Ltd. All rights reserved.

Aqueous two-phasemicellar systems in an oscillatory flowmicro-reactor: study of perspectives and experimental performance

BACKGROUND: Aqueous two-phase micellar systems (ATPMS) are micellar surfactant solutions with physical properties that make them very efficient for the extraction/concentration of biological products. In this work the main proposal that has been discussed is the possible applicability and importance of a novel oscillatory flow micro-reactor (micro-OFR) envisaged for parallel screening and/or development of industrial bioprocesses in ATPMS. Based on the technology of oscillatory flow mixing (OFM), this batch or continuous micro-reactor has been presented as a new small-scale alternative for biological or physical-chemical applications. RESULTS: ATPMS experiments were carried out in different OFM conditions (times, temperatures, oscillation frequencies and amplitudes) for the extraction of glucose-6-phosphate dehydrogenase (G6PD) in Triton X-114/buffer with Cibacron Blue as affinity ligand. CONCLUSION: The results suggest the potential use of OFR, considering this process a promising and new alternative for the purification or pre-concentration of bioproducts. Despite the applied homogenization and extraction conditions have presented no improvements in the partitioning selectivity of the target enzyme, when at rest temperature they have influenced the partitioning behavior in Triton X-114 ATPMS. (C) 2011 Society of Chemical Industry

Phase diagrams of a CTAB/organic solvent/buffer system applied to extraction of enzymes by reverse micelles

A partial pseudo-ternary phase diagram has been studied for the cethyltrimethylammonium bromide/isooctane:hexanol:butanol/potassium phosphate buffer system, where the two-phase diagram consisting of the reverse micelle phase (L-2) in equilibrium with the solvent is indicated. Based on these diagrams two-phase systems of reverse micelles were prepared with different compositions of the compounds and used for extraction and recovery of two enzymes, and the percentage of enzyme recovery yield monitored. The enzymes glucose-6-phosphate dehydrogenase (G6PD) and xylose redutase (XR) obtained from Candida guilliermondii yeast were used in the extraction procedures. The recovery yield data indicate that micelles having different composition give selective extraction of enzymes. The method can thus be used to optimize enzyme extraction processes. (c) 2007 Elsevier B.V. All rights reserved.

Hollow fiber-based liquid-phase microextraction (HF-LPME) of isradipine and its main metabolite followed by chiral HPLC analysis: application to an in vitro biotransformation study

An enantioselective liquid chromatographic method using two-phase hollow fiber liquid-phase microextraction (HF-LPME-HPLC) was developed for the determination of isradipine (ISR) enantiomers and its main metabolite (pyridine derivative of isradipine, PDI) in microsomal fractions isolated from rat liver. The analytes were extracted from 1 mL of microsomal medium using a two-phase HF-LPME procedure with hexyl acetate as the acceptor phase, 30 min of extraction, and sample agitation at 1,500 rpm. For the first time, ISR enantiomers and PDI were resolved. For this separation, a ChiralpakA (R) AD column with hexane/2-propanol/ethanol (94:04:02, v/v/v) as the mobile phase at a flow rate of 1.5 mL min(-1) was used. The column was kept at 23 A +/- 2 A degrees C. The drug and metabolite detection was performed at 325 nm and the internal standard oxybutynin was detected at 225 nm. The recoveries were 23% for PDI and 19% for each ISR enantiomer. The method presented quantification limits (LOQ) of 50 ng mL(-1) and was linear over the concentration range of 50-5,000 and 50-2,500 ng mL(-1) for PDI and each ISR enantiomer, respectively. The validated method was employed to an in vitro biotransformation study of ISR using rat liver microsomal fraction showing that (+)-(S)-ISR is preferentially biotransformed.

Determination of beta-artemether and its main metabolite dihydroartemisinin in plasma employing liquid-phase microextraction prior to liquid chromatographic-tandem mass spectrometric analysis

A method for the determination of artemether (ART) and its main metabolite dihydroartemisinin (DHA) in plasma employing liquid-phase microextraction (LPME) for sample preparation prior to liquid chromatography-tandem mass spectrometry (LC-MS-MS) was developed. The analytes were extracted from 1 nil, of plasma utilizing a two-phase LPME procedure with artemisinin as internal standard. Using the optimized LPME conditions, mean absolute recovery rates of 25 and 32% for DHA and ART, respectively, were achieved using toluene-n-octanol (1:1, viv) as organic phase with an extraction time of 30 min. After extraction, the analytes were resolved within 5 min using a mobile phase consisting of methanol-ammonium acetate (10 mmol L(-1) pH 5.0, 80:20. v/v) on a laboratory-made column based on poly(methyltetradecylsiloxane) attached to a zirconized-silica support. MS-MS detection was employed using an electrospray interface in the positive ion mode. The method developed was linear over the range of 5-1000 ng mL(-1) for both analytes. Precision and accuracy were within acceptable levels of confidence (<15%). The assay was applied to the determination of these analytes in plasma from rats treated with ART. The two-phase LPME procedure is affordable and the solvent consumption was very low compared to the traditional methods of sample preparation. (C) 2010 Elsevier B.V. All rights reserved.

A phase II study of UFT with leucovorin administered as a twice daily schedule in the treatment of patients with metastatic colorectal cancer

Prolonged infusions have been shown to be safer and potentially more effective than bolus regimens of 5-fluorouracil (5-FU) as treatment for metastatic colorectal cancer (mCRC). However, infusional 5-FU requires central venous access and costly infusion pumps. Oral fluoropyrimidines enable longer exposures to 5-FU with increased convenience. Tegafur - uracil (UFT) with leucovorin (LV) given thrice daily has improved safety plus comparable survival and response rates to bolus 5-FU/LV. We conducted a phase II clinical study in 98 patients with mCRC to evaluate if UFT with LV given twice daily provided comparable time to progression ( TTP), efficacy and tolerability to that reported for thrice daily in two phase III clinical studies. Secondary objectives included overall response rate ( ORR) and overall survival ( OS). Median TTP was 3.8 months, when compared with 3.5 months for thrice daily. The ORR ( 11%) and median OS ( 12.8 months) with twice daily administration were similar to that of thrice daily administration ( 12% and 12.4 months). The incidence of grade 3/4 treatment-related diarrhoea was 30% on the twice daily and 21% on the thrice daily schedule. These results suggest that twice daily administration has similar efficacy and tolerability to thrice daily administration and is an acceptable alternative for patients who would benefit from UFT with LV therapy.

Pathogenicity and reproductive fitness of Pratylenchus brachyurus on cowpea

The lesion nematode Pratylenchus brachyurus is widespread in cowpea plantations throughout the tropics and sub-tropics. However, the pathogenicity of P. brachyurus on cowpea has been scarcely studied. In this work, it was demonstrated in two glasshouse experiments that an isolate (Pb-20) of P brachyurus was pathogenic to cowpea cv. IPA-206, adversely affecting the plant growth and pod formation and filling. Initial population levels of 5000 and 15 000 nematodes per plant caused reduction of root growth and typical decay of root tissue. The third experiment demonstrated that all six cowpea cultivars selected for evaluation supported reproduction of three isolates of P. brachyurus (Pb-20, Pb-21 and Pb-23) in their roots, although the reproduction factor values obtained indicated that they were dissimilar in their reproductive fitness. Low resistance to R brachyurus was reported for at least one tested cultivar, but apparently of an insufficient degree to be effective for field management of the nematode.

Changes in temporomandibular joint disc position and form following Herbst and fixed orthodontic treatment

Objective: To determine the changes in the position and form of the temporomandibular joint articular disc in adolescents with Class II division 1 malocclusion and mandibular retrognathism treated with the Herbst appliance (phase I) and fixed orthodontic appliance (phase II). Materials and Methods: Thirty-two consecutive adolescents went through phase I of treatment and 23 completed phase II. The temporomandibular joints were evaluated qualitatively by means of magnetic resonance images at the beginning of treatment (T1), during phase I (T2), at the end of phase I (T3), and at the end of phase II (T4). Results: Significant changes in disc position were not observed with the mouth closed between T1 X T3 (P = .317), T3 X T4 (P = .287), or T1 X T4 (P = .261). At T2, on average, the disc was positioned regressively. With the mouth open, no difference was observed between T1 X T3 (P = .223) or T1 X T4 (P = .082). We did observe a significant difference between T3 X T4 (P < .05). Significant changes in the disc form were found with the mouth closed between T1 X T2 (P < .001) and T2 X T3 (P < .001). Conclusions: At the end of the two-phase treatment, in general terms, the position and form of the initial articular discs were maintained; however, in some temporomandibular joints some seemingly adverse effects were observed at T4. (Angle Orthod. 2010;80:843-852.)

Harvest-ironman: heavy armature, and not its defensive secretions, protects a harvestman against a spider

Natural selection has caused prey species to evolve distinct defensive mechanisms. One of such mechanisms was the evolution of noxious or distasteful chemicals, which have appeared independently in a number of vertebrates and invertebrates. In detailed analyses of arthropod behaviour, scent gland secretions have consistently been shown to be responsible for repelling specific predators. Because using such chemicals is costly, animals with alternative cheaper defences are expected not to release such secretions when alternative options exist. In this study, we sought to determine the defensive mechanisms of the harvestman Discocyrtus invalidus, a heavy bodied species that bears a pair of repugnatorial glands. The spider Enoploctenus cyclothorax was used as the predator, and the cricket Gryllus sp. was used as a control. In a first set of experiments, the harvestmen were preyed upon significantly less than the crickets. In two other experiments, we found that harvestmen did not use their scent gland secretions to deter the predator. Moreover, results of a fourth experiment revealed that these spiders are not repelled by defensive secretions. Discocyrtus invalidus has a thick cuticle on the entire body: scanning electron micrographs revealed that only the mouth, the articulations of appendages and the tips of the legs are not covered by a hard integument. In a fifth experiment, we found that these spiders had difficulty piercing the harvestmen body. This is the first experimental evidence that a chemically defended arachnid does not use its scent gland secretions to repel a much larger predator but instead relies on its heavily built body. (c) 2010 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.

Microstructural characterization of rapidly solidified and heat-treated Ti(92)B(8) eutectic alloy

In this work, Ti(92)B(8) alloy was processed via rapid solidification (splat-cooling) and then heat-treated at 700 degrees C and 1000 degrees C. A careful microstructural characterization indicated that, after rapid solidification, a very fine two-phase microstructure was produced with no significant saturation of B in alpha/beta Ti. There was no indication of amorphous formation in the rapidly solidified splats. Both alpha Ti and TiB were observed in the microstructures of the splats after heat-treatment at 700 degrees C and 1000 degrees C, confirming the stability of the alpha Ti+TiB two-phase region in this temperature range. (C) 2008 Elsevier Inc. All rights reserved.