Removal of 17β Estradiol and Fluoranthene by Nanofiltration and Ultrafiltration

With the recent emergence of endocrine disrupting compounds as an important potable drinking water and reclaimed wastewater quality issue, the removal of two estrogenic compounds (17β-estradiol and fluoranthene) by nanofiltration and ultrafiltration membranes was investigated. A less hydrophobic organic compound model species [parachlorobenzoic acid (PCBA)] was tested. 17β-estradiol (E2), fluoranthene, and PCBA were applied to the membrane in the presence and absence of natural organic matter (NOM). Both batch adsorption and dead-end stirred-cell filtration experiments indicated that adsorption is an important mechanism for transport/removal of relatively hydrophobic compounds, and is related to the octanol-water partition coefficient (KOW) values. All filtration measurements were performed approximately the same permeate flow rate in order to minimize artifacts from concentration polarization varied with different hydrodynamic operating conditions at the membrane interface. The percent removal by dead-end stirred-cell filtration ranged from 10 to >95% depending upon membrane pore size/hydrophobicity and presence/absence of NOM at an initial concentration ranging from 0.1 to 0.5 μM. Additional batch adsorption experiments with radio-label (3H) E2 at lower concentrations ranging 0.025 to 5 nM showed that E2 removal due to adsorption was independent of its initial concentration. Adsorption occurs both on the membrane surface and interior membrane pore surfaces. However, adsorption was insignificant for PCBA (log?KOW = 2.7), but removal presumably occurred due to electrostatic exclusion. Partition coefficients (log?K) of 0.44 to 4.86 measured in this study increased with log?KOW and membrane pore size.     

Molten globule intermediate of recombinant human growth hormone: stabilization with surfactants

We demonstrate that a surfactant-stabilized molten globule intermediate exists for recombinant human growth hormone (rhGH), is very hydrophobic, and tends to form aggregates. Characterization of this intermediate included equilibrium denaturation measured by electron paramagnetic resonance (EPR) and CD spectroscopy, assessment of aggregation during refolding, and fluorescence studies of its binding to the hydrophobic probe, 1-anilinonapthalene-8-sulfonate (1,8-ANS). We have found that at 4.5 M guanidinium hydrochloride (GuHCl), a molten globule intermediate of rhGH is stabilized and results in significant aggregation upon refolding. This intermediate is populated by the addition of the nonionic surfactant, Tween. This surfactant also reduces the extent of aggregation during refolding of rhGH from 4.5 M GuHCl. Overall, our studies reveal that rhGH forms a molten globule-like intermediate during folding and this intermediate self-associates. This self-association is reduced upon formation of a Tween-rhGH complex. Tween also binds to the native protein. Thus, nonionic surfactants such as Tween may act like molecular chaperones in facilitating protein folding while not altering the native conformation.     

Impact of Membrane Surface Modification on the Treatment of Surface Water

New polyethersulfone (PES) based membranes for ultrafiltration (UF) were developed by blending a surface-modifying macromolecule (SMM) in the casting solution, in an attempt to minimize the impact of fouling. Fouling was evaluated using concentrated Ottawa River water (CORW), either unfractionated or fractionated via UF. These membranes also included some polyvinylpyrrolidone (PVP), a pore forming additive. A statistical analysis was conducted to evaluate the impact of some variables on the treatment of the surface water. The independent variables included PVP/PES ratio in the casting solution, with and without SMM, and the nature of the feed CORW [low molecular weight (LMW) fraction, unfractionated, high molecular weight (HMW) fraction]. The performance variables studied were total organic carbon (TOC) removal, the foulant accumulation at the membrane surface after filtration, the flux reduction, and the final permeate flux. The most important variable was the feed water. Filtration of LMW had a higher final flux, less fouling, but slightly lower TOC removal. The SMM did not significantly impact the membrane performance. TOC removal was high, compared with results reported in the literature for UF membranes.     

Use of Iron Oxides to Enhance Metal Removal in Crossflow Microfiltration

This research investigated whether iron oxyhydroxides used in conjunction with microfiltration could improve the removal of metals from a waste while maintaining adequate flux. Filtration of individual metals, a mixture of metals, and the mixture with two iron oxides were investigated. The research indicated that use of a coagulant (an iron-containing salt) might not be as important in microfiltration systems as in conventional gravity separation systems, because of the small pore size of microfilters. In some cases, filtration efficiency was relatively poor at the beginning of a treatment cycle, but in almost all cases it became excellent once a layer had built up on the membrane surface. The greatest benefit provided by iron oxides might be to reduce membrane fouling. A crystalline iron oxide such as goethite is more attractive than ferrihydrite. The flux improvement with goethite is greatest if a thin layer is deposited on the membrane surface before the contaminant metals are injected into the system, so that the goethite can trap the potentially foulant metal hydroxide particles away from the membrane surface.     

A Model of Foam Filters

Filtration with foam filters is reported as a successful method to remove inclusions from top-cut silicon scrap. Inclusions in top-cut silicon scrap are needle-like Si₃N₄ particles and round SiC inclusions. A high filtration efficiency of more than 99 pct for 30-ppi SiC filters is achieved. The inclusions that remain are mainly SiC particles smaller than 10 μm. Possibly these particles are primarily secondary inclusions. The filtration efficiency increases with decreasing filter pore size. The main factor that plays a role in deep-bed filtration seems to be interception. Various models are considered to estimate the removal efficiency of foam filters by this mechanism. Here, we propose a new model called “the branch” model. This model gives a high filtration efficiency and agrees the best with the experimental results.     

Aggregation of recombinant human interferon gamma: kinetics and structural transitions

Protein aggregation is a complex phenomenon that can occur in vitro and in vivo, usually resulting in the loss of the protein's biological activity. While many aggregation studies focus on a mechanism due to a specific stress, this study focuses on the general nature of aggregation. Recombinant human interferon-gamma (rhIFN-gamma) provides an ideal model for studying protein aggregation, as it has a tendency to aggregate under mild denaturing stresses (low denaturant concentration, temperature below the Tm, and below pH 5). All of the aggregates induced by these stresses have a similar structure (high in intermolecular beta-sheet content and a large loss of alpha-helix) as determined by infrared and circular dichroism spectroscopy. Thermally induced and denaturant-induced aggregation processes follow first-order kinetics under the conditions of this study. Spectroscopic and kinetic data suggest that rhIFN-gamma aggregates through an intermediate form possessing a large amount of residual secondary structure. In contrast to the aggregates formed under denaturing stresses, the salted-out protein has a remarkably nativelike secondary structure.     

Simulations of reversible protein aggregate and crystal structure

We simulated the structure of reversible protein aggregates as a function of protein surface characteristics, protein-protein interaction energies, and the entropic penalty accompanying the immobilization of protein in a solid phase. These simulations represent an extension of our previous work on kinetically irreversible protein aggregate structure and are based on an explicit accounting of the specific protein-protein interactions that occur within reversible aggregates and crystals. We considered protein monomers with a mixture of hydrophobic and hydrophilic surface regions suspended in a polar solvent; the energetic driving force for aggregation is provided by the burial of solvent-exposed hydrophobic surface area. We analyzed the physical properties of the generated aggregates, including density, protein-protein contact distributions, solvent accessible surface area, porosity, and order, and compared our results with the protein crystallization literature as well as with the kinetically irreversible case. The physical properties of reversible aggregates were consonant with those observed for the irreversible aggregates, although in general, reversible aggregates were more stable energetically and were more crystal-like in their order content than their irreversible counterparts. The reversible aggregates were less dense than the irreversible aggregates, indicating that the increased energetic stability is derived primarily from the optimality rather than the density of the packing in the solid phase. The extent of hydrophobic protein-protein contacts and solvent-exposed surface area within the aggregate phase depended on the aggregation pathway: reversible aggregates tended to have a greater proportion of hydrophobic-hydrophobic contacts and a smaller fraction of hydrophobic solvent-exposed surface area. Furthermore, the arrangement of hydrophobic patches on the protein surface played a major role in the distribution of protein contacts and solvent content. This was readily reflected in the order of the aggregates: the greater the contiguity of the hydrophobic patches on the monomer surface, the less ordered the aggregates became, despite the opportunities for rearrangement offered by a reversible pathway. These simulations have enhanced our understanding of the impact of protein structural motifs on aggregate properties and on the demarcation between aggregation and crystallization.     

稀土尾矿库周边地下水中稀土元素含量测定与分布规律的研究

现行的地下水监测方法中,常利用水溶性稀土元素的含量判断其生态效应,而吸附在固体颗粒物、胶体及微生物上的稀土元素在检测中往往被忽略。本实验采用不同孔径的滤膜对尾矿库周边地下水进行连续过滤,测定滤上物及滤液中稀土元素含量,并对其进行了归一化处理,进而分析地下水中稀土元素的分布模式。结果表明,0.45μm滤膜滤上物中稀土元素含量远高于0.45μm滤液中稀土元素含量。滤后水样中稀土元素含量随滤膜孔径减小而降低,其分布规律均表现为Ce负异常,Eu正异常,但水体中稀土元素与理化指标间的相关性发生了改变。说明经过不同孔径滤膜处理后,水体中稀土元素与固体颗粒物的络合性质发生改变。因此,只分析水溶性稀土元素含量来判断地下水中稀土元素生态效应是有失偏颇的。     

Annexin V and vesicle membrane electroporation

The method of membrane electroporation (ME) has been used as an analytical tool to quantify the effect of membrane curvature on transient electric pore formation, and on the adsorption of the protein annexin V (M(r)= 35,800) to the outer surface of unilamellar lipid vesicles (of radii 25 < or = a/nm < or = 200). Relaxation kinetic studies using optical membrane probes of the diphenylhexatriene type suggest that electric pore formation is induced by ionic interfacial polarization causing entrance of the (more polarizable) water into the lipid bilayer membrane yielding (hydrophobic and hydrophilic) pore states with a mean stationary pore radius rp = 0.35 (+/-0.05) nm. Extent and rate of ME, compared at the same induced transmembrane voltage, were found to decrease both with increasing vesicle radius and with increasing protein concentration. This 'inhibitory' effect of annexin V is apparently allosteric and saturates at about [ANT]sat = 4 microM annexin V for vesicles of a = 100 nm at 1 mM total lipid concentration, 0.13 mM total Ca2+ concentration and at T = 293 K. Data analysis in terms of Gibbs area-difference-elasticity energy suggests that the bound annexin V reduces the gradient of the lateral pressure across the membrane. At [ANT]sat, about 20% of the vesicle surface is covered by the bound protein, but it is only 0.01% of the surface of the outer lipid leaflet in which a part of the protein, perhaps the aromatic residue of the tryptophan (W 187), is inserted. Insertion leads to a denser packing of the lipid molecules in the outer membrane leaflet. As a consequence, the radius of the electropores in the remaining membrane part, not covered by annexin V decreases (rp/nm = 0.37, 0.36 and 0.27) with increasing adsorption of the protein ([ANT] = 0, 2 and 4 microM, respectively).     

Effect of excipients on the stability and structure of lyophilized recombinant human growth hormone

We have investigated the effect of mannitol, sorbitol, methyl alpha-D-mannopyranoside, lactose, trehalose, and cellobiose on the stability and structure of the pharmaceutical protein recombinant human growth hormone (rhGH) in the lyophilized state. All excipients afforded significant protection of the protein against aggregation, particularly at levels to potentially satisfy water-binding sites on the protein in the dried state (i.e., 131:1 excipient-to-protein molar ratio). At higher excipient-to-protein ratios, X-ray diffraction studies showed that mannitol and sorbitol were prone to crystallization and afforded somewhat less stabilization than at lower ratios where the excipient remained in the amorphous, protein-containing phase. The secondary structure of rhGH was determined using Fourier transform infrared (FTIR) spectroscopy. rhGH exhibited a decrease in alpha-helix and increase in beta-sheet structures upon drying. Addition of excipient stabilized the secondary structure upon lyophilization to a varying extent depending on the formulation. Samples with a significant degree of structural conservation, as indicated by the alpha-helix content, generally exhibited reduced aggregation. In addition, prevention of protein-protein interactions (indicated by reduced beta-sheet formation) also tended to result in lower rates of aggregation. Therefore, in addition to preserving the protein structure, bulk additives that do not crystallize easily and remain amorphous in the solid state can be used to increase protein-protein distance and thus prevent aggregation.     

Conformational flexibility of domain III of annexin V studied by fluorescence of tryptophan 187 and circular dichroism: the effect of pH

The conformation and dynamics of domain III of annexin V was studied by steady-state and time-resolved fluorescence of its single tryptophan residue (Trp187) as a function of pH in the absence of calcium. At neutral pH, the maximum of emission occurs at 326 nm, in agreement with the hydrophobic location of the tryptophan residue seen in the three-dimensional structure. Upon decreasing the pH, a progressive red-shift by about 12 nm of the fluorescence emission spectrum is observed. The effect is complete between pH 6 and 4.5, and most likely involves at least one and maybe two carboxylic group(s). Circular dichroism mesurements give evidence for a preservation of the native folding of the protein in these mild acidic conditions. A fluorescence red-shift of smaller amplitude is also observed at high pH (approximately 11). The aggregation state of the protein is affected by pH: while at neutral pH, the protein is monomeric (rotational correlation time = 14 ns); it forms aggregates larger than a dimer (rotational correlation time > 40 ns) in acidic pH conditions. These results suggest that electrostatic interactions are probably important for the stabilization of the folding of domain III without calcium. The conformational change may be related to the aggregation state of the molecule. Examination of the protein crystal structures with and without calcium ion in domain III shows an interplay of salt bridges implying charged amino acid side chains at the molecule surface of domain III. These observations may provide a further clue to the mechanism of the conformational change of domain III of annexin V induced by high calcium concentrations and interaction at the membrane/water interface.     

NOM Accumulation at NF Membrane Surface: Impact of Chemistry and Shear

The effects of solution chemistry, surface shear, and composition of natural organic matter (NOM) were investigated for their impact on accumulation of foulant material at the surface of charged polymeric nanofiltration membranes. The source of NOM was the Suwannee River. A bench-scale, batch recycle system was used with 20 hollow fiber, nanofiltration membranes. Membrane flux decline and foulant accumulation increased at low pH and high ionic strength as a result of neutralization of charge, electric double layer compression, and the apparent shift in conformation of charged NOM macromolecules. The rate of NOM accumulation decreased with operating time, suggestive of an eventual steady state between adsorption and desorption. The effect of NOM composition on membrane fouling could not be discerned by a standard technique to isolate hydrophobic and hydrophilic NOM fractions, quite possibly because of the fractionation methodology's failure to recover a small but important fouling fraction or because of NOM interactions that are lost when individual fractions are separately tested. However, a greater percentage of the hydrophilic than hydrophobic fraction permeated the membrane, in agreement with prior observations by others. Increasing the cross flow velocity from 85 to 255 cm∕s reduced the extent of flux decline, presumably due to hydrodynamic disruption of cake layer formation.     

Constriction-Based Retention Criterion for Granular Filter Design

The filter design criteria in practice are currently based on laboratory tests that were carried out on uniform base soil and filter materials. These criteria mostly involve specific particle size ratios, where the system of base soil and filter is represented by some characteristic particle sizes. Consequently, these criteria have limitations when applied to nonuniform materials. In filters, it is the constriction size rather than the particle size that affects filtration. In this paper, a mathematical procedure to determine the controlling constriction size is introduced, and subsequently, a constriction-based retention criterion for granular filters is presented. The model also incorporates the effect of nonuniformity of base soil in terms of its particle size distribution, considering the surface area of the particles. The proposed retention criterion is verified based on experimental data taken from past studies plus large-scale filtration tests carried out by the authors. The model successfully and distinctly demarcates the boundary between effective and ineffective filters in the case of cohensionless base soils.     

Removal of Copper Ions from Waters Using Surfactant-Enhanced Hybrid PAC/MF Process

This paper deals with the removal of copper ions from aqueous solutions by using surfactant-enhanced powdered activated carbon (PAC)/microfiltration (MF) hybrid process, including the evaluation of process performance and fouling dynamics at various linear alkyl benzene sulfonate (LABS), PAC, and Cu2+ concentrations of feed solution. Although the use of surfactant as an additive material increased the adsorption efficiency in PAC/MF hybrid process, a considerable amount of the flux was lost for surfactant concentration above critical micelle concentration. The process could be employed with a performance of 74.7%, 97.2% and 87?L/m2?h for LABS rejection, Cu2+ rejection and permeate flux at the conditions of 2?g PAC/L, 5?mM LABS, 0.2?mM Cu2+, and 60-min process time. Cu2+ rejection, which increased with increasing of LABS, and PAC amounts decreased with the increase in Cu2+ concentration. It was understood that the increments in LABS, PAC, and Cu2+ concentrations being an indicator for the feed solution quality led to the occurrence of more fouling on the membrane. The analyses of dynamics concerning the fouling behaviors, which were carried out using single and combined pore blocking models, put forward that the cake formation was the main predominant mechanism in the process. It was also determined that the variation of feed contents deduced the presence of rather complex fouling behaviors as a simultaneous function of secondary membrane layer formation and clogging and narrowing of membrane pores by surfactants.     

Aggregation Patterns of Proteasome in Injured Neurons Induced by Transient Cerebral Ischemia

Proteasome activity reduction is all important pathological phenomenon,resulting in proteins aggregation and neuronal death in the injured neurons induced by transient ischemia.Our previous report showed that the trap of proteasome in the protein aggregates WaS a reason to lead to the reduction of proteasome activity.However, the patterns of proteasome entered into protein aggregates are not clear.In this study,we used a global ischemia model,Hematoxylin-Eosin staining,differential centrifuge,proteasome activity assay,sucrose gradient density centrifuge,and Western blot analysis to investigate this problem.Our results show that there are two aggregation pattems of proteasome after transient ischemia and reperfusion.One is that 26S proteasome is trapped by protein aggregates as a whole unit,and the other is that 19S or 20S is trapped in the protein aggregates,respectively,after 26S disassociates.     

Folding of beta-sheet membrane proteins: a hydrophobic hexapeptide model

Beta-sheets, in the form of the beta-barrel folding motif, are found in several constitutive membrane proteins (porins) and in several microbial toxins that assemble on membranes to form oligomeric transmembrane channels. We report here a first step towards understanding the principles of beta-sheet formation in membranes. In particular, we describe the properties of a simple hydrophobic hexapeptide, acetyl-Trp-Leu5 (AcWL5), that assembles cooperatively into beta-sheet aggregates upon partitioning into lipid bilayer membranes from the aqueous phase where the peptide is strictly monomeric and random coil. The aggregates, containing 10 to 20 monomers, undergo a relatively sharp and reversible thermal unfolding at approximately 60 degreesC. No pores are formed by the aggregates, but they do induce graded leakage of vesicle contents at very high peptide to lipid ratios. Because beta-sheet structure is not observed when the peptide is dissolved in n-octanol, trifluoroethanol or sodium dodecyl sulfate micelles, aggregation into beta-sheets appears to be an exclusive property of the peptide in the bilayer membrane interface. This is an expected consequence of the hypothesis that a reduction in the free energy of partitioning of peptide bonds caused by hydrogen bonding drives secondary structure formation in membrane interfaces. But, other features of interfacial partitioning, such as side-chain interactions and reduction of dimensionality, must also contribute. We estimate from our partitioning data that the free energy reduction per residue for aggregation is about 0.5 kcal mol-1. Although modest, its aggregate effect on the free energy of assembling beta-sheet proteins can be huge. This surprising finding, that a simple hydrophobic hexapeptide readily assembles into oligomeric beta-sheets in membranes, reveals the potent ability of membranes to promote secondary structure in peptides, and shows that the formation of beta-sheets in membranes is more facile than expected. Furthermore, it provides a basis for understanding the observation that membranes promote self-association of beta-amyloid peptides. AcWL5 and related peptides thus provide a good starting point for designing peptide models for exploring the principles of beta-sheet formation in membranes.     

Modeling Formation of Natural Organic Matter Fouling Layers on Ultrafiltration Membranes

Three simple mathematical models to describe fouling of an ultrafiltration membrane by natural organic matter (NOM) are developed and compared. These models attribute the fouling to: (1) an increase of the effective pore length by an amount equal to the thickness of the NOM gel layer that forms on the membrane surface; (2) formation of a uniform, microporous NOM gel layer on the membrane surface, made of primary particles comprising tens to hundreds of NOM molecules; or (3) narrowing of the membrane pores by sorption of a monolayer of NOM molecules along the full length of each pore. The key parameters characterizing each model are identified and estimated based on data for flux and film growth gathered in the same system. In each case, the estimated parameter values are plausible in light of the known physical properties of the membrane and NOM molecules.     

Effect of substance P and receptor antagonists on secretion of lingual lipase and amylase from rat von Ebner''s gland

Electron donor acceptor gels based on cyanocarbons have been tested for human serum protein adsorption in the absence of salt-promotion by water-structuring salt. This phenomenon was compared with a normal adsorption process in the presence of salt. The tricyanoaminopropene-divinyl sulfone-agarose displayed unusual protein adsorption properties as binding could occur both independently or dependently of the salt-promotion. The absence of hydrophobic or ionic character of the salt-independent interaction suggests an electron donor acceptor adsorption mechanism which is shown, for the first time, to occur independently of salt-promotion in aqueous solution. Study of the protein adsorption specificity showed similar protein selectivity for the fractions adsorbed in both conditions.     

Conformation and ion-channeling activity of a 27-residue peptide modeled on the single-transmembrane segment of the IsK (minK) protein

IsK (minK) protein, in concert with another channel protein KVLQT1, mediates a distinct, slowly activating, voltage-gated potassium current across certain mammalian cell membranes. Site-directed mutational studies have led to the proposal that the single transmembrane segment of IsK participates in the pore of the potassium channel [Takumi, T. (1993) News Physiol. Sci. 8, 175-178]. We present functional and structural studies of a short peptide (K27) with primary structure NH2-1KLEALYILMVLGFFGFFTLGIMLSYI27R-COOH, corresponding to the transmembrane segment of IsK (residues 42-68). When K27 was incorporated, at low concentrations, into phosphatidylethanolamine, black-lipid membranes, single-channel activity was observed, with no strong ion selectivity. IR measurements reveal the peptide has a predominantly helical conformation in the membrane. The atomic resolution structure of the helix has been established by high-resolution 1H NMR spectroscopy studies. These studies were carried out in a solvent comprising 86% v/v 1,1,1,3,3,3-hexafluoro-isopropanol-14% v/v water, in which the IR spectrum of the peptide was found to be very similar to that observed in the bilayer. The NMR studies have established that residues 1-3 are disordered, while residues 4-27 have an alpha-helical conformation, the helix being looser near the termini and more stable in the central region of the molecule. The length (2. 6 nm) of the hydrophobic segment of the helix, residues 7-23, matches the span of the hydrocarbon chains (2.3 +/- 0.25 nm) of fully hydrated bilayers of phosphatidylcholine lipid mixture from egg yolk. The side chains on the helix surface are predominantly hydrophobic, consistent with a transmembrane location of the helix. The ion-channeling activity is believed to stem from long-lived aggregates of these helices. The aggregation is mediated by the pi-pi stacking of phenylalanine aromatic rings of adjacent helices and favorable interactions of the opposing aliphatic-like side chains, such as leucine and methionine, with the lipid chains of the bilayer. This mechanism is in keeping with site-directed mutational studies which suggest that the transmembrane segment of IsK is an integral part of the pore of the potassium channel and has a similar disposition to that in the peptide model system.     

Volume regulation of spermatozoa by quinine-sensitive channels

Bovine spermatozoa were shown to exhibit rapid regulatory volume decrease (RVD) when exposed to hypotonic saline media. This quinine- and quinidine-sensitive regulatory volume decrease was coincident with K+ release due to stretch-activation of inhibitor-specific presumptive K+ channels. The regulatory volume decrease response was much faster than a similar phenomenon observed in human peripheral blood lymphocytes. Studies on volume changes in different electrolyte and nonelectrolyte media suggested that: (1) this inhibitor-specific channel could also be a nonspecific pore in the spermatozoal membrane for nonelectrolytes below 150 daltons; (2) subpopulations (of nearly equal size) of the spermatozoa differ in the expression of the pore; (3) capacitation abolishes this distinction between subpopulations of spermatozoa; and (4) the general case of RVD for other mammalian spermatozoa was also established.