A stable liquid-liquid extraction system for clavulanic acid using polymer-based aqueous two-phase systems

The partitioning of Clavulanic Acid (CA) in a novel inexpensive and stable aqueous two-phase system (ATPS) composed by poly(ethylene glycol) (PEG) and sodium polyacrylate (NaPA) has been studied. The aqueous two-phase systems are formed by mixing both polymers with a salt (NaCl or Na2SO4) and an aqueous solution of CA. The stability of CA on the presence of both polymers was investigated and it was observed that these polymers do not degrade the biomolecule. The effect of PEG-molecular size, polymer concentrations on the commercial CA partitioning has been studied, at 25 degrees C. The data showed that commercial CA was preferentially partitioned for the PEG-rich phase with a partition coefficient (K-CA) between 1 and 12 in the PEG/NaPA aqueous two phase systems supplemented with NaCl and Na2SO4. The partition to the PEG phase was increased in the systems with high polymer concentrations. Furthermore, Na2SO4 caused higher CA preference for the PEG-phase than NaCl. The systems having a composition with 10 wt.% of PEG4000, 20 wt.% of NaPA8000 and 6 wt.% of Na2SO4 were selected as the optimal ones in terms of recovery of CA from fermented broth of Streptomyces clavuligerus. The partitioning results (K-CA = 9.15 +/- 1.06) are competitive with commercial extraction methods of CA (K-CA = 11.91 +/- 2.08) which emphasizes that the system PEG/NaPA/Na2SO4 can be used as a new process to CA purification/concentration from fermented broth. (C) 2012 Elsevier B.V. All rights reserved.

Plasmid DNA partitioning and separation using poly(ethylene glycol)/poly(acrylate)/salt aqueous two-phase systems

Phase diagrams of poly(ethylene glycol)/polyacrylate/Na2SO4 systems have been investigated with respect to polymer size and pH. Plasmid DNA from Escherichia coil can depending on pH and polymer molecular weight be directed to a poly(ethylene glycol) or to a polyacrylate-rich phase in an aqueous two-phase system formed by these polymers. Bovine serum albumin (BSA) and E. coil homogenate proteins can be directed opposite to the plasmid partitioning in these systems. Two bioseparation processes have been developed where in the final step the pDNA is partitioned to a salt-rich phase giving a total process yield of 60-70%. In one of them the pDNA is partitioned between the polyacrylate and PEG-phases in order to remove proteins. In a more simplified process the plasmid is partitioned to a PEG-phase and back-extracted into a Na2SO4-rich phase. The novel polyacrylate/PEG system allows a strong change of the partitioning between the phases with relatively small changes in composition or pH. (C) 2012 Elsevier B.V. All rights reserved.

Quality characteristics and fertilizing ability of ram sperm subpopulations separated by partition in an aqueous two-phase system

Centrifugal countercurrent distribution (CCCD) in an aqueous two-phase system (TPS) is a resolute technique revealing sperm heterogeneity and for the estimation of the fertilizing potential of a given semen sample. However, separated sperm subpopulations have never been tested for their fertilizing ability yet. Here, we have compared sperm quality parameters and the fertilizing ability of sperm subpopulations separated by the CCCD process from ram semen samples maintained at 20 degrees C or cooled down to 5 degrees C. Total and progressive sperm motility was evaluated by computer-assisted analysis using a CASA system and membrane integrity was evaluated by flow cytometry by staining with CFDA/Pl. The capacitation state, staining with chlortetracycline, and apoptosis-related markers, such as phosphatidylserine (PS) translocation detected with Annexin V. and DNA damage detected by the TUNEL assay, were determined by fluorescence microscopy. Additionally, the fertilizing ability of the fractionated subpopulations was comparative assessed by zona binding assay (ZBA). CCCD analysis revealed that the number of spermatozoa displaying membrane and DNA alterations was higher in samples chilled at 5 degrees C than at 20 degrees C. which can be reflected in the displacement to the left of the CCCD profiles. The spermatozoa located in the central and right chambers (more hydrophobic) presented higher values (P<0.01) of membrane integrity, lower PS translocation (P<0.05) and DNA damage (P<0.001) than those in the left part of the profile, where apoptotic markers were significantly increased and the proportion of viable non-capacitated sperm was reduced. We have developed a new protocol to recover spermatozoa from the CCCD fractions and we proved that these differences were related with the fertilizing ability determined by ZBA, because we found that the number of spermatozoa attached per oocyte was significantly higher for spermatozoa recovered from the central and right chambers, in both types of samples. This is the first time, to our knowledge that sperm recovered from a two-phase partition procedure are used for fertilization assays. These results open up new possibilities for using specific subpopulations of sperm for artificial insemination or in vitro fertilization, not only regarding better sperm quality but also certain characteristics such as subpopulations enriched in spermatozoa bearing X or Y chromosome that we have already isolated or any other feature. (C) 2011 Elsevier B.V. All rights reserved.

Interference of some aqueous two-phase system phase-forming components in protein determination by the Bradford method

The interference of some specific aqueous two-phase system (ATPS) phase-forming components in bovine serum albumin (BSA) determination by the Bradford method was investigated. For this purpose, calibration curves were obtained for BSA in the presence of different concentrations of salts and polymers. A total of 19 salts [Na2SO4, (NH4)(2)SO4, MgSO4, LiSO4, Na2HPO4, sodium phosphate buffer (pH 7.0), NaH2PO4, K2HPO4, potassium phosphate buffer (pH 7.0), KH2PO4, C6H8O7, Na3C6HSO7, KCHO2, NaCHO2, NaCO3, NaHCO3, C2H4O2, sodium acetate buffer (pH 4.5), and NaC2H3O2] and 7 polymers [PEG 4000, PEG 8000, PEG 20000, UCON 3900, Ficoll 70000, PES 100000, and PVP 40000] were tested, and each calibration curve was compared with the one obtained for BSA in water. Some concentrations of salts and polymers had considerable effect in the BSA calibration curve. Carbonate salts were responsible for the highest salt interference, whereas citric and acetic acids did not produce interference even in the maximum concentration level tested (5 wt%). Among the polymers, UCON gave the highest interference, whereas Ficoll did not produce interference when used in concentrations up to 10 wt%. It was concluded that a convenient dilution of the samples prior to the protein quantification is needed to ensure no significant interference from ATPS phase-forming constituents. (C) 2011 Elsevier Inc. All rights reserved.

Partitioning and extraction of collagenase from Penicillium aurantiogriseum in poly(ethylene glycol)/phosphate aqueous two-phase system

Purification of collagenase produced by Penicillium aurantiogriseum URM4622 was carried using a PEG/phosphate aqueous two-phase system (ATPS). A 2(3)-full experimental design was used to investigate the influence of PEG molar mass, PEG concentration and phosphate concentration on the selected responses, namely partition coefficient, activity yield and purification factor. The ATPS was composed of PEG (molar mass of 550, 1500 and 4000 g/mol) at concentrations of 15.0, 17.5 and 20.0% (w/w) and phosphate at concentrations of 12.5, 15.0 and 17.5% (w/w). The best results of one-step extraction of collagenase from the fermentation broth (partition coefficient of 1.01, activity yield of 242% and purification factor of 23.5) were obtained at pH 6.0 using 20.0% (w/w) PEG 550 and 17.5% (w/w) phosphate. The results of this preliminary study demonstrate that the selected ATPS is satisfactorily selective for the extraction of such a collagenase. (C) 2012 Elsevier B.V. All rights reserved.

Internal fit of two all-ceramic systems and metal-ceramic crowns

Objectives: The aim of this study was to investigate the internal fit (IF) of glass-infiltrated alumina (ICA - In-Ceram Alumina), yttria-stabilized tetragonal zirconia polycrystals (Y-TZP - IPS e.max ZirCAD), and metal-ceramic (MC - Ni-Cr alloy) crowns. Material and Methods: Sixty standardized resin-tooth replicas of a maxillary first molar were produced for crown placement and divided into 3 groups (n=20 each) according to the core material used (metal, ICA or Y-TZP). The IF of the crowns was measured using the replica technique, which employs a light body polyvinyl siloxane impression material to simulate the cement layer thickness. The data were analyzed according to the surfaces obtained for the occlusal space (OS), axial space (AS) and total mean (TM) using two-way ANOVA with Tukey's multiple comparison test (p<0.05). Results: No differences among the different areas were detected in the MC group. For the Y-TZP and ICA groups, AS was statistically lower than both OS and TM. No differences in AS were observed among the groups. However, OS and TM showed significantly higher values for ICA and Y-TZP groups than MC group. Comparisons of ICA and Y-TZP revealed that OS was significantly lower for Y-TZP group, whereas no differences were observed for TM. Conclusions: The total mean achieved by all groups was within the range of clinical acceptability. However, the metal-ceramic group demonstrated significantly lower values than the all-ceramic groups, especially in OS.

Two-phase flow patterns and pressure drop inside horizontal tubes containing twisted-tape inserts

This paper presents an experimental study on two-phase flow patterns and pressure drop of R134a inside a 15.9 mm ID tube containing twisted-tape inserts. Experimental results were obtained in a horizontal test section for twisted-tape ratios of 3, 4, 9 and 14, mass velocities ranging from 75 to 250 kg/m(2) s and saturation temperatures of 5 and 15 degrees C. An unprecedented discussion on two-phase flow patterns inside tubes containing twisted-tape inserts is presented and the flow pattern effects on the frictional pressure drop are carefully discussed. Additionally, a new method to predict the frictional pressure drop during two-phase flow inside tubes containing twisted-tape inserts is proposed. (C) 2012 Elsevier Ltd. All rights reserved.

Internal fit of two all-ceramic systems and metal-ceramic crowns

OBJECTIVES: The aim of this study was to investigate the internal fit (IF) of glass-infiltrated alumina (ICA - In-Ceram Alumina), yttria-stabilized tetragonal zirconia polycrystals (Y-TZP - IPS e.max ZirCAD), and metal-ceramic (MC - Ni-Cr alloy) crowns. MATERIAL AND METHODS: Sixty standardized resin-tooth replicas of a maxillary first molar were produced for crown placement and divided into 3 groups (n=20 each) according to the core material used (metal, ICA or Y-TZP). The IF of the crowns was measured using the replica technique, which employs a light body polyvinyl siloxane impression material to simulate the cement layer thickness. The data were analyzed according to the surfaces obtained for the occlusal space (OS), axial space (AS) and total mean (TM) using two-way ANOVA with Tukey s multiple comparison test (p<0.05). RESULTS: No differences among the different areas were detected in the MC group. For the Y-TZP and ICA groups, AS was statistically lower than both OS and TM. No differences in AS were observed among the groups. However, OS and TM showed significantly higher values for ICA and Y-TZP groups than MC group. Comparisons of ICA and Y-TZP revealed that OS was significantly lower for Y-TZP group, whereas no differences were observed for TM. CONCLUSIONS: The total mean achieved by all groups was within the range of clinical acceptability. However, the metal-ceramic group demonstrated significantly lower values than the all-ceramic groups, especially in OS.

Two-Phase Frictional Pressure Drop and Flow Boiling Heat Transfer for R245fa in a 2.32-mm Tube

Experimental two-phase frictional pressure drop and flow boiling heat transfer results are presented for a horizontal 2.32-mm ID stainless-steel tube using R245fa as working fluid. The frictional pressure drop data was obtained under adiabatic and diabatic conditions. Experiments were performed for mass velocities ranging from 100 to 700 kg m−2 s−1 , heat flux from 0 to 55 kW m−2 , exit saturation temperatures of 31 and 41◦C, and vapor qualities from 0.10 to 0.99. Pressures drop gradients and heat transfer coefficients ranging from 1 to 70 kPa m−1 and from 1 to 7 kW m−2 K−1 were measured. It was found that the heat transfer coefficient is a strong function of the heat flux, mass velocity, and vapor quality. Five frictional pressure drop predictive methods were compared against the experimental database. The Cioncolini et al. (2009) method was found to work the best. Six flow boiling heat transfer predictive methods were also compared against the present database. Liu and Winterton (1991), Zhang et al. (2004), and Saitoh et al. (2007) were ranked as the best methods. They predicted the experimental flow boiling heat transfer data with an average error around 19%.

Multi-planet extrasolar systems - detection and dynamics

20 years after the discovery of the first planets outside our solar system, the current exoplanetary population includes more than 700 confirmed planets around main sequence stars. Approximately 50% belong to multiple-planet systems in very diverse dynamical configurations, from two-planet hierarchical systems to multiple resonances that could only have been attained as the consequence of a smooth large-scale orbital migration. The first part of this paper reviews the main detection techniques employed for the detection and orbital characterization of multiple-planet systems, from the (now) classical radial velocity (RV) method to the use of transit time variations (TTV) for the identification of additional planetary bodies orbiting the same star. In the second part we discuss the dynamical evolution of multi-planet systems due to their mutual gravitational interactions. We analyze possible modes of motion for hierarchical, secular or resonant configurations, and what stability criteria can be defined in each case. In some cases, the dynamics can be well approximated by simple analytical expressions for the Hamiltonian function, while other configurations can only be studied with semi-analytical or numerical tools. In particular, we show how mean-motion resonances can generate complex structures in the phase space where different libration islands and circulation domains are separated by chaotic layers. In all cases we use real exoplanetary systems as working examples.

Magneto-optical probe of quantum Hall states in a wide parabolic well modulated by random potential

Polarized photoluminescence from weakly coupled random multiple well quasi-three-dimensional electron system is studied in the regime of the integer quantum Hall effect. Two quantum Hall ferromagnetic ground states assigned to the uncorrelated miniband quantum Hall state and to the spontaneous interwell phase coherent dimer quantum Hall state are observed. Photoluminescence associated with these states exhibits features caused by finite-size skyrmions: dramatic reduction of the electron spin polarization when the magnetic field is increased past the filling factor nu = 1. The effective skyrmion size is larger than in two-dimensional electron systems.

Experimental investigation of gravitational gas separation in an inclined annular channel

The oil industry uses gas separators in production wells as the free gas present in the suction of the pump reduces the pumping efficiency and pump lifetime. Therefore, free gas is one of the most important variables in the design of pumping systems. However, in the literature there is little information on these separators. It is the case of the inverted-shroud gravitational gas separator. It has an annular geometry due to the installation of a cylindrical container in between the well casing and pioduction pipe (tubing). The purpose of the present study is to understand the phenomenology and behavior of inverted-shroud separator. Experimental tests were performed in a 10.5-m-length inclinable glass tube with air and water as working fluids. The water flow rate was in the range of 8.265-26.117 l/min and the average inlet air mass flow rate was 1.1041 kg/h, with inclination angles of 15 degrees, 30 degrees, 45 degrees, 60 degrees, 75 degrees, 80 degrees and 85 degrees. One of the findings is that the length between the inner annular level and production pipe inlet is one of the most important design parameters and based on that a new criterion for total gas separation is proposed. We also found that the phenomenology of the studied separator is not directly dependent on the gas flow rate, but on the average velocity of the free surface flow generated inside the separator. Maps of efficiency of gas separation were plotted and showed that liquid flow rate, inclination angle and pressure difference between casing and production pipe outlet are the main variables related to the gas separation phenomenon. The new data can be used for the development of design tools aiming to the optimized project of the pumping system for oil production in directional wells. (C) 2012 Elsevier Inc. All rights reserved.