Fluorescence imaging of macromolecule transport in high molecular weight cut-off microdialysis

When microdialysis (MD) membrane exceeds molecular weight cut-off (MWCO) of 100 kDa, the fluid mechanics are in the ultrafiltration regime. Consequently, fluidic mass transport of macromolecules in the perfusate over the membrane may reduce the biological relevance of the sampling and cause an inflammatory response in the test subject. Therefore, a method to investigate the molecular transport of high MWCO MD is presented. An in vitro test chamber was fabricated to facilitate the fluorescent imaging of the MD sampling process, using fluoresceinylisothiocyanate (FITC) dextran and fluorescence microscopy. Qualitative studies on dextran behavior inside and outside the membrane were performed. Semiquantitative results showed clear dextran leakage from both 40 and 250 kDa dextran when 100 kDa MWCO membranes were used. Dextran 40 kDa leaked out with an order of magnitude higher concentration and the leakage pattern resembled more of a convective flow pattern compared with dextran 250 kDa, where the leakage pattern was more diffusion based. No leakage was observed when dextran 500 kDa was used as a colloid osmotic agent. The results in this study suggest that fluorescence imaging could be used as a method for qualitative and semiquantitative molecular transport and fluid dynamics studies of MD membranes and other hollow fiber catheter membranes. Graphical Abstract

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The Nonreversibility of Adsorption-Desorption of Dieldrin and Lindane on Chitin in Seawater

Abstract

Adsorption and desorption of dieldrin and lindane on chitin were investigated in seawater batch tests as a function of chitin concentration, temperature, pH and salinity. For chitin concentrations ranging from 0.5 g l^?1 to 12.5 g l^?1, the pesticide concentrations were varied from 4 μgl^?1 to 65 μg^?1 for dieldrin and from 40 μl^?1 to 680 μg l^?1 for lindane. Both dieldrin and lindane show adsorption-desorption hysteresis at low chitin concentration. At high chitin concentrations (m > 6.25 g l^?1 for dieldrin and m > 10 gl^?1 for lindane) both pesticides exhibit reversible behaviour. However, only lindane adsorption is affected by chitin concentration. These types of behaviour remain fixed in prewashed chitin. However, an increase in the temperature and a decrease in the salinity made the process become reversible. A resistant-reversible two component model has been applied to account for these types of behaviour and provides a way to explain most of the observed effects by defining mass independent distribution coefficients.     

A laser light-scattering study of the structure of agarose gels

Diffusion coefficients of dextran fractions within agarose gels surrounded by dextran solution have been measured by laser light scattering using the autocorrelation method. Plots were made of the diffusion coefficient relative to that in dilute solution, D/D₀, against the logarithm of hydrodynamic diameter logd for each concentration of agarose, and superimposed by displacing horizontally to produce a unified plot. In this way it was shown that D/D₀ is a function of C^bd, where C is agarose concentration, with b = 1/3 and 1/2 for the cases in which the dextrans were mixed in before gelation and allowed to diffuse in afterwards, respectively, the plots being the same for a reference concentration of 0.7%. A value of b = 1/2 is that which would be expected if the molecular weight per unit length of the gel fibers were independent of concentration, and a value of 25 kg mol^?1 nm^?1 is calculated. Mobile concentrations of dextran within the gels relative to those in the surrounding solutions were found by determining the scattered intensity associated with the diffusing dextran molecules from the zero-time value of the autocorrelation function. All results and calculations are discussed in terms of current theories, and compared with earlier work on calcium alginate gels for which a molecular weight per unit length of gel fiber of 0.59 kg mol^?1 nm^?1 was calculated. The nature of the spectral broadening of the light scattered from agarose gels in the absence of dextran is described.     

Crystallization of X-type zeolite from concentrated sodium silicate and aluminate solutions

Crystallization of an X-type zeolite from concentrated solutions of sodium aluminate (Al₂O₃ 90–330 g l^?1) and silicate (SiO₂ 110–180 g l^?1) was studied.     

Flow-injection spectrophotometric determination of palladium in catalysts and dental alloys with 2-(5-bromo-2-pyridylazo)-5-(N-propyl-N-sulfopropyl-amino)aniline

2-(5-Bromo-2-pyridylazo)-5-(N-propyl-N-sulfopropylamino)aniline rapidly forms a water-soluble complex with palladium in an acetate-buffered medium at pH 3.2.The molar absorptivity of the complex is 9.84×10⁴l mol^?1 at 612 nm. The calibration graph is linear over the range of 10–100 μg l^?1 palladium; the detection limit is 2 μg l^?1 and the relative standard deviation is 0.6% for 100 μg l^?1 palladium. The sample throughput is 50 h^?1. Divalent transition metals (Fe, Ni, Co) do not interfere at levels from 2 to 10 mg l^?1. Interference from copper is prevented by adding 10^?3 M EDTA solution to the carrier stream. Palladium in solutions of catalysts and dental alloys can be determined selectively, sensitively and rapidly.     

Improved cesium sensitivity in electrothermal atomic absorption spectrometry

The sensitivity for cesium determination by electrothermal atomic absorption spectrometry is improved four-fold by the addition of a large excess of potassium nitrate. Zeeman background corrections is used to compensate for the large non-specific absorption signal resulting from the potassium. The characteristic concentration and detection limit are 0.44 and 2 μg l^?1, respectively, and the coefficient of variation is 2% at the 50 μg l^?1 level. The procedure is suitable for the rapid determination of cesium in leach solutions from nuclear waste fixation experiments.     

A method of determining and removing sulphide to allow the determination of sulphate,phosphate, nitrite and ammonia by conventional methods in small volumes of anoxic waters

Mercury(II) chloride is used to precipitate free sulphide from <10-ml samples of anoxic water. The sulphide-free supernatant solution can be used for estimation of sulphide by measuring the concentration of unreacted mercury(II) ion and for determinations of sulphate, inorganic phosphate, ammonia and nitrite by spectrophotometric methods which normally cannot be used because of sulphide interference. Concentrations that can be determined lie within the ranges: sulphide 0.5–180 000 μg S l^?1, sulphate 0.024–2.77 g S l^?1, ammonia 1–70 000 μg N l^?1, nitrite 1–3000 μg N l^?1, inorganic phosphate 1–4000 μg P l^?1. Interstitial waters from estuarine sediments, tidal flats, mangrove swamps, and an anoxic estuarine basin were examined.     

Determination of gold by atomic absorption spectrometry after extraction into the non-desulphurized fraction of crude oil distillate

The extraction of gold from aqueous chloride solutions into non-desulphurized fractions of crude oil distillates, especially paraffin oil, is described. The fraction boiling at 150–220°C exhibited optimum properties. The extraction is tested for solutions containing chlorides, dissolved chlorine and 0.1–3 M hydrochloric acid. Nitric acid should be absent. The distribution coefficient of gold varies from 400 to 900. Extracts containing ? 0.3 g l^?1 gold are stable for at least 12 months. The organic extract is sprayed into a lean acetylene/air flame with measurement at 242.8 nm (background correction). The calibration graph has linear portions over the ranges 0–2.5 and 2.5–16 mg l^?1. The limit of detection is 0.03 mg l^?1 gold in the extract (0.001 mg l^?1 in the aqueous phase). The minimum measurable concentration for gold in auriferous rocks and ores is 0.018 g per ton with 25-g samples.     

Determination of lead in whole blood with a simple flow-injection system and computerized stripping potentiometry

An automated (24 samples/hour) procedure is described for the determination of lead (0–1000 μg l^?1) in human blood based on flow-injection stripping potentiometry. The samples are diluted 20-fold with 0.5 M hydrochloric acid containing 100 mg l^?1 mercury and 40 μg l^?1 cadmium (II), and a 1.1 ml aliquot is injected into the flow system. With a mercury-coated carbon fibre as working electrode, lead (II) is determined by using cadmium (II) as internal standard and a calibration graph prepared from bovine blood. Analyses of two human blood reference samples yielded results of 335±37 and 691±24 μg l^?1 lead, the certified values being 332 and 663 μg l^?1, respectively.     

Large-scale purification of Leuconostoc mesenteriodes NRRL B512F dextransucrase for use in the biosynthesis of dextran by batch and continuous chromatography

The extracellular enzyme dextransucrase was produced from Leuconostoc mesenteriodes NRRL B512F and purified by ultracentrifugation and cross-flow ultrafiltration for use in the biosynthesis of the macromolecule dextran by ion exchange chromatographic reaction—separation techniques. The two-stage purification process yielded over 90% pure dextransucrase with overall enzyme recovery of over 60%. A second stage of centrifugation was required to achieve complete cell removal. The purified enzyme contained 1–2 g l⁻¹ of solute ions, which affected the operation of the chromatographic system. Gel filtration removed over 93% of the remaining ions but resulted in high activity losses. Two-phase separation with polyethylene glycol (PEG) and purification by ion exchange chromatography were less successful in desalting the enzyme. PEG precipitation was successful in concentrating the enzyme, but the ions remained predominantly with the enzyme portion of the two phases. The purified enzyme was found to be unstable during storage.Use of the enzyme in chromatographic reactor—separators for the production of dextran resulted in over 33% more high molecular weight dextran (the desired product) and a useful pure fructose byproduct being obtained than for a conventional reactor. Sodium and potassium ions in the enzyme hampered continuous operation by displacing calcium ions from the resin and thus reducing the separation efficiency of the system. Partial regeneration of the resin with calcium nitrate rather than complete enzyme desalting, which was very expensive and resulted in high activity losses, helped overcome this effect.     

Rapid determination of copper,nickel, lead and cadmium in small samples of estuarine and coastal waters by liquid/liquid extraction and electrothermal atomic absorption spectrometry

A rapid liquid/liquid extraction of 1.25-ml samples is used with graphite-furnace atomic absorption spectrometry for the determination of dissolved trace metals in saline waters. The metals are chelated with ammonium pyrrolidine dithiocarbamate and extracted into 1,1,1-trichloroethane; 20–40 μl of extract is injected into the furnace. Sample manipulation and overall time are greatly decreased compared to other similar large-scale extraction methods; all the chemical steps are done in the sample cups of an auto-sampler for graphite-furnace a.a.s. Detection limits (Cu 0.3 μg l^?1, Cd 0.02 μg l^?1, Pb 0.7 μg l^?1, Ni 0.5 μg l^?1 are low enough for applications in routine monitoring of filterable trace metal concentrations in coastal and estuarine waters to check for compliance with Environmental Quality Standards that apply in the European Community.     

Flow-injection technique for the determination of low levels of phosphorus in natural waters

An inexpensive flow-injection instrument for determining low concentrations of dissolved reactive phosphorus in natural waters is reported. The unique feature is the use of an inexpensive detector consisting of a flow cell and a simple photometer that incorporates a super-bright light-emitting diode as the source and a photodiode as the detector. The tin(II) chloride-molybdate method was optimized using a modified simplex optimization method. Silicate interference up to 5 mg Si l^?1 was removed by addition of 0.10% (w/v) tartaric acid. Using the tartaric acid-modified optimized reagents, a detection limit of 0.6 μg P l^?1 was achieved. The method was linear over the range 0–100 μg P l^?1 with an excellent precision (r.s.d. 2.9% at 2.0 and 0.5% at 50 μg P l^?1). An in-line pre-concentration anion-exchange column was used to obtain an even lower detection limit of 0.1 μg P l^?1 and applied to the analysis of real samples.     

Preconcentration of palladium(II) from water with thionalide loaded on silica gel

2-Mercapto-N-2-naphthylacetamide (thionalide) on silica gel is used for rapid preconcentration of μg l^?1 levels of palladium(II) from aqueous solution, followed by atomic absorption spectrometric measurement. In batch experiments, palladium was quantitatively retained on the gel from solutions 5 M in acid to pH 8; equilibrium was achieved within 10 s. The chelating capacity of the gel was 7.5 μmol Pd g^?1 at pH < 4. The effect of flow rate on retention was studied. Palladium retained on the column was completely eluted with 20 ml of 0.2 M thiourea in 0.1 M hydrochloric acid. The palladium concentration in sea water is shown to be < 0.3 μg l^?1.     

Determination of selenium in human body fluids by hydride-generation atomic absorption spectrometry : Optimization of sample decomposition

The accuracy of the determination of selenium in human body fluids by hydride-generation a.a.s. depends critically upon the sample decomposition-method used. Digestion with HNO₃ alone gave low selenium recoveries, but with nitric, sulfuric and perchloric acids at a final temperature of 310°C gave results that agreed with those obtained by other techniques. The recovery of selenomethionine added to whole blood and of trimethylselenonium iodide added to urine was 97–104%. The average selenium values found for 6 healthy individuals were 88 μg l^?1 in whole blood, 75 μg l^?1 in blood plasma and 307 μg (kg Hb)^?1 in erythrocytes. A detection limit of 5 μg l^?1 Se in body fluids was found under routine conditions.     

Differential preconcentration of arsenic(III) and arsenic(V) with thionalide loaded on silica gel

2-Mercapto-N-2-naphtylacetamide (thionalide) on silica gel is used for differential preconcentration of μg l^?1 levels of arsenic(III) and arsenic(V) from aqueous solution. In batch experiments, arsenic(III) was quantitatively retained on the gel from solutions of pH 6.5–8.5, but arsenic(V) and organic arsenic compounds were not retained. The chelating capacity of the gel was 5.6 μmol g^?1 As(III) at pH 7.0. Arsenic retained on teh column was completely eluted with 25 ml of 0.01 M sodium borate in 0.01 M sodium hydroxide containing 10 mg l^?1 iodine (pH 10). The arsenic was determined by silver diethyldithiocarbamate spectrophotometry. Arsenic(V) was subsequently determined after reduction to arsenic(III) with sulphite and iodide. Arsenic(III) and arsenic(V) in sea water are shown to be < 0.12 and 1.6 μg l^?1, respectively.     

Determination of soluble cyanide in soil samples by differential pulse polarography

Soluble cyanide can be determined ins oil samples by differential pulse polarography. Interference from sulphide is avoided by treating the alkali-stabilized sample solutions with lead carbonate prior to distillation of cyanide from the soil extract. Less than 10 μg l^?1 cyanide can be determined accurately, depending on teh weight of sample taken and the final collection volume. For a 100-g soil sample, the detection limit is 5 ng g^?1, which is similar to the limit of a standard spectrophotometric method. Relative standard deviations are 1–3%.     

Determination of total tin in silicate rocks by graphite furnace atomic absorption spectrometry

A method is described for the determination of total tin in silicate rocks utilizing a graphite furnace atomic absorption spectrometer with a stabilized-temperature platform furnace and Zeeman-effect background correction. The sample is decomposed by lithium metaborate fusion (3 + 1) in graphite crucibles with the melt being dissolved in 7.5% hydrochloric acid. Tin extractions (4 + 1 or 8 + 1) are executed on portions of the acid solutions using a 4% solution of trioctylphosphine oxide in methyl isobutyl ketone (MIBK). Ascorbic acid is added as a reducing agent prior to extraction. A solution of diammonium hydrogenphosphate and magnesium nitrate is used as a matrix modifier in the graphite furnace determination. The limit of detection is > 10 pg, equivalent to > 1 μg l^?1 of tin in the MIBK solution or 0.2–0.3 μg g^?1 in the rock. The concentration range is linear between 2.5 and 500 μg l^?1 tin in solution. The precision, measured as relative standard deviation, is < 20% at the 2.5 μg l^?1 level and < 7% at the 10–30 μg l^?1 level of tin. Excellent agreement with recommended literature values was found when the method was applied to the international silicate rock standards BCR-1, PCC-1, GSP-1, AGV-1, STM-1, JGb-1 and Mica-Fe. Application was made to the determination of tin in geological core samples with total tin concentrations of the order of 1 μg g^?1 or less.     

Continuous flow extraction of indium with bis(2-ethylhexyl)phosphoric acid in 4-methylpentane-2-one coupled on-line with flame atomic absorption spectrometry

Extraction in liquid-liquid segmented flow is used for preconcentration of indium from dilute nitric acid solutions into bis(2-ethylhexyl)phosphoric acid dissolved in 4-methylpentane-2-one. The extraction setup is coupled on-line with flame spectrometry to give a fully mechanized system. The detection limit of the method is 0.03 mg l^?1, the calibration plot is linear up to 1.75 mg l^?1. Repeatability is 1.5% RSD measured at 1 mg l^?1. Sample throughput is 60 h^?1.     

Ethanol Production from Starch Hydrolyzates using <Emphasis Type="Italic">Zymomonas mobilis</Emphasis> and Glucoamylase Entrapped in Polyvinylalcohol Hydrogel

The glucoamylase from Aspergillus niger, immobilized into poly(vinylalcohol) hydrogel lens-shaped capsules LentiKats®, was used for simultaneous saccharification and fermentation (SSF) with Zymomonas mobilis in free form. This system was stable in both the repeated batch and continuous mode of SSF. The microorganism was found to adsorb on the capsules with immobilized enzyme. This increased the ethanol productivity of the repeated batch system with 5% w/v of immobilized glucoamylase almost 2.1 times (7.2 g l^?1 h^?1) compared to free enzyme–free microorganism system (3.5 g l^?1 h^?1). The continuous SSF with the immobilized glucoamylase (11.5% w/v) tested for 15 days had productivity 10 g l^?1 h^?1, which is comparable to continuous experiments on semi-defined glucose medium (10 g l^?1 h^?1). These two systems were stable in both glucoamylase activity and microorganism productivity.     

An efficient flow-injection system with on-line ion-exchange preconcentration for the determination of trace amounts of heavy metals by atomic absorption spectrometry

A flow-injection system with on-line ion-exchange preconcentration on dual columns is described for the determination of trace amounts of heavy metals at μg l^?1 and sub-μg l^?1 levels by flame atomic absorption spectrometry. The degree of preconcentration ranges from 50- to 105-fold for different elements at a sampling frequency of 60 s h^?1. The detection limits for Cu, Zn, Pb and Cd are 0.07, 0.03, 0.5, and 0.05 μg l^?1, respectively. Relative standard deviations were 1.2–3.2% at μg l^?1 levels. The behaviour of the different chelating exchangers used was studied with respect to their preconcentration characteristics, with special emphasis on interferences encountered in the analysis of sea water.