Analytica Chimica Acta

Published by Elsevier Masson
Salt-tolerant yeast Saccharomyces cerevisiae ARIF KD-003 was applied to highly sensitive and reproducible absorbance-based biochemical oxygen demand (BOD(AB-ScII)) measurement for seawater. In the previous work, we have studied the BOD(AB-ScI) method using normal Baker's yeast S. cerevisiae, and the excellent feature of the Baker's yeast as uniformly sustainable in solution could successfully be utilized. However, the BOD(AB-ScI) responses were disappeared by the existence of chloride ion as well as seawater. In the present method, uniformity in solution was also observed with S. cerevisiae ARIF KD-003, and salt-tolerance of the yeast was observed even in saturate concentration of sodium chloride. Next, characterizations of the influences of pH and incubation temperature were investigated. After optimum conditions were obtained, two calibration curves were made between 0.33 and 22 mg O2 L(-1) BOD using standard solution of glucose glutamic acid (GGA) or mixture of GGA and artificial seawater. Then, excellent reproducibility as the averages of relative standard deviation (R.S.D.(av)) in two calibration curves (nine points each) was successfully obtained at 1.10% at pure water or 1.03% at artificial seawater standard, respectively. In addition, the 3 sigma lower detection limit was calculated to be 0.07 mg O2 L(-1) BOD, and 0.11 mg O2 L(-1) BOD was experimentally detected by increase of the sample volume at 1.5-folds. The storage stability of the S. cerevisiae ARIF KD-003 was obtained at least 4 weeks.
1,1-Diphenyl-2-picrylhydrazyl (DPPH·) is a stable nitrogen centred radical widely used to evaluate direct radical scavenging properties of various synthetic or natural antioxidants (AOs). The bleaching rate of DPPH· absorbance at 515nm is usually monitored for this purpose. In order to avoid the interference of complex coloured natural products used as antioxidant supplements or cosmetics, HPLC systems have been reported as alternative techniques to spectrophotometry. They also rely upon measurement of DPPH· quenching rate and none of them permits to identify and measure 1,1-diphenyl-2-picryl-hydrazine (DPPH-H), the reduced product of DPPH· resulting from hydrogen atom transfer (HAT), which is the main mechanism of the reaction between DPPH· and AOs. We presently report an HPLC method devoted to the simultaneous measurement of DPPH· and DPPH-H. Both were fully separated on a C18 column eluted with acetonitrile-10 mM ammonium citrate buffer pH 6.8 (70:30, v/v) and detected at 330 nm. Adsorption process of DPPH· onto materials of the HPLC system was pointed out. Consequently, the linearity range observed for DPPH· was restricted, thus a much lower limit of detection was obtained for DPPH-H than for DPPH· using standards (0.02 and 14 μM, respectively). The method was applied to three commonly used AOs, i.e. Trolox(®), ascorbic acid and GSH, and compared with spectrophotometry. Further application to complex matrices (cell culture media, vegetal extracts) and nanomaterials demonstrated (i) its usefulness because of higher selectivity than colorimetry, and (ii) its help to investigate the mechanisms occurring with the free radical.
Two chiral stationary phases (CSPs) based on optically active (3,3'-diphenyl-1,1'-binaphthyl)-20-crown-6 covalently bonded to silica gel were utilized for the first time for the resolution of racemic beta-amino acids using high performance liquid chromatography. All of the 10 beta-amino acids tested were resolved on the CSP containing residual silanol-protecting n-octyl groups, while only five beta-amino acids were resolved on the CSP containing residual silanol groups. The superiority of the CSP containing residual silanol-protecting n-octyl groups and the characteristic retention behaviors of the two enantiomers on the CSP were rationalized to stem from the removal of the residual silanol groups, which can otherwise induce the non-enantioselective retention of the analytes, and the improved lipophilicity of the CSP. The elution orders of the two enantiomers of beta-amino acids were identical on the two CSPs and, consequently, it was concluded that the two CSPs were concluded to utilize identical chiral recognition mechanisms. The different elution orders of the analytes were proposed to be attributed to the presence or absence of pi-pi interactions between the CSP and analytes.
In the presence of a small amount of 1,2-dibromopropane (1,2-DBP), 1,1'-binaphthol (BINOL) displays strong room temperature phosphorescence in gamma-cyclodextrin (gamma-CD) solution without deoxygenation. The phosphorescence intensity as well as the phosphorescence lifetime of (S)-BINOL is greater than that of (R)-BINOL, indicating a distinct chiral discrimination of gamma-CD toward this pair of enantiomers. Both (R)-BINOL and (S)-BINOL exhibit a double exponential phosphorescence decay with lifetimes of 5.89 ms and 17.3 ms for (R)-BINOL and 7.58 ms and 23.6 ms for (S)-BINOL, respectively. The association constant obtained for (S)-BINOL/gamma-CD/1,2-DBP ternary complex is larger than that for (R)-BINOL/gamma-CD/1,2-DBP complex. Thus, the observation of RTP lifetime differences between (R)-BINOL and (S)-BINOL can be attributed to their different ability to form complexes with chiral gamma-CD, which is further supported by an analysis of the proton NMR chemical shift differences between (R)-BINOL and (S)-BINOL.
Second-order global hard-modelling was applied to resolve the complex formation between Co(2+), Ni(2+), and Cd(2+) cations and 1,10-phenantroline. The highly correlated spectral and concentration profiles of the species in these systems and low concentration of some species in the individual collected data matrices prevent the well-resolution of the profiles. Therefore, a collection of six equilibrium data matrices including series of absorption spectra taken with pH changes at different reactant ratios were analyzed. Firstly, a precise principle component analysis (PCA) of different augmented arrangements of the individual data matrices was used to distinguish the number of species involved in the equilibria. Based on the results of PCA, the equilibria included in the data were specified and second-order global hard-modelling of the appropriate arrangement of six collected equilibrium data matrices resulted in well-resolved profiles and equilibrium constants. The protonation constant of the ligand (1,10-phenantroline) and spectral profiles of its protonated and unprotonated forms are the additional information obtained by global analysis. For comparison, multivariate curve resolution-alternating least squares (MCR-ALS) was applied to the same data. The results showed that second-order global hard-modelling is more convenient compared with MCR-ALS especially for systems with completely known model. It can completely resolve the system and the concentration profiles which are closer to correct ones. Moreover, parameters showing the goodness of fit are better with second-order global hard-modelling.
In this study, an electrochemical DNA biosensor was developed based on the recognition of target DNA by hybridization detection. The study was carried out using glassy carbon electrode (GCE) modified with lable-free 21-mer single-stranded oligonucleotides related to hepatitis B virus sequence via covalent immobilization and [Cu(dmp)(H(2)O)Cl(2)] (dmp=2,9-dimethyl-1,10-phenanthroline) as an electrochemical indicator, whose sizes are comparable to those of the small groove of native double-duplex DNA. The method, which is simple and low cost, allows the accumulation of copper complex within the DNA layer. Electochemical detection was performed by cyclic voltammetry and differential pulse voltammetry over the potential range where the [Cu(dmp)(H(2)O)Cl(2)] was active. Numerous factors affecting the probe immobilization, target hybridization, and indicator binding reactions were optimized to maximize the sensitivity and speed the assay time. With this approach, a sequence of the hepatitis B virus could be quantified over the ranges from 8.82 x 10(-8) to 8.82 x 10(-7) M with a linear correlation of r=0.9937 and a detection limit of 7.0 x 10(-8) M. The [Cu(dmp)(H(2)O)Cl(2)] signal observed from probe sequence before and after hybridization with four bases mismatch containing sequence is lower than that observed after hybridization with complementary sequence.
A new nano-sized silver(I) ion-imprinted polymer (IIP) was prepared via precipitation copolymerization using ethyleneglycol dimethacrylate, as a cross-linking agent in the presence of Ag(+) and an aza-thioether crown containing a 1,10-phenanthroline subunit as a highly selective complexing agent. The imprint silver(I) ion was removed from the polymeric matrix using a 1.0M HNO3 solution. The resulting powder material was characterized using IR spectroscopy and scanning electron microscopy. The SEM micrographs showed colloidal nanoparticles of about 52nm and 75nm in diameter and slightly irregular in shape for leached and unleached IIPs, respectively. The optimal pH for quantitative enrichment was 6.0 and maximum sorbent capacity of the prepared IIP for Ag(+) was 18.08μmolg(-1). The relative standard deviation and limit of detection (LOD=3Sb/m) for flame atomic absorption spectrometric determination of silver(I) ion, after its selective extraction by the prepared IIP nanobeads, were evaluated as 2.42% and 2.2×10(-8)M, respectively. The new Ag(+)-IIP was also applied as a suitable sensing element to the preparation of highly selective and sensitive voltammetric and potentiometric sensors for ultra trace detection of silver(I) ion in water samples, with limits of detection of 9.0×10(-10) and 1.2×10(-9)M, respectively. Copyright © 2014. Published by Elsevier B.V.
Electrochemiluminescence (ECL) of ruthenium complexes has broad applications and the immobilization of Ru(bpy)(3)(2+) has received extensive attention. In comparison with Ru(bpy)(3)(2+), Ru(phen)(3)(2+) can be immobilized more easily because of its better adsorbability. In this study, immobilization of Ru(phen)(3)(2+) for ECL analysis has been demonstrated for the first time by using graphene oxide (GO) as an immobilization matrix. The immobilization of Ru(phen)(3)(2+) is achieved easily by mixing Ru(phen)(3)(2+) with GO without using any ion exchange polymer or covalent method. The strong binding of Ru(phen)(3)(2+) with GO is attributed to both the π-π stacking interaction and the electrostatic interaction. The Ru(phen)(3)(2+)/GO modified electrode was characterized by using tripropylamine (TPA) as the coreactant. The linear range of TPA is from 3×10(-7) to 3×10(-2) mol L(-1) with the detection limit of 3×10(-7) mol L(-1). The ECL sensor demonstrates outstanding long-term stability. After the storage in the ambient environment for 90 days, the ECL response remains comparable with its original signal.
We have conducted a comprehensive comparative study of Ru(bipy)(3)(2+), Ru(bipy)(2)(phen)(2+), Ru(bipy)(phen)(2)(2+), and Ru(phen)(3)(2+) as chemiluminescence and electrochemiluminescence (ECL) reagents, to address several previous conflicting observations and gain a greater insight into their potential for chemical analysis. Clear trends were observed in many of their spectroscopic and electrochemical properties, but the relative chemiluminescence or ECL intensity with a range of analytes/co-reactants is complicated by the contribution of numerous (sometimes opposing) factors. Significantly, the reversibility of cyclic voltammetric responses for the complexes decreased as the number of phenanthroline ligands was increased, due to the lower stability of their ruthenium(III) form in the aqueous solvent. This trend was also evident over a longer timescale when the ruthenium(III) form was spectrophotometrically monitored after chemical oxidation of the ruthenium(II) complexes. In general, the greater stability of Ru(bipy)(3)(3+) resulted in lower blank signals, although this effect was less pronounced with ECL, where the reagent is oxidised in the presence of the co-reactants. Nevertheless, this shows the need to compare signal-to-blank ratios or detection limits, rather than the more common comparisons of overall signal intensity for different ruthenium complexes. Furthermore, our results support previous observations that, compared to Ru(bipy)(3)(2+), Ru(phen)(3)(2+) provides greater ECL and chemiluminescence intensities with oxalate, which in some circumstances translates to superior detection limits, but they do not support the subsequent generalised notion that Ru(phen)(3)(2+) is a more sensitive reagent than Ru(bipy)(3)(2+) for all analytes.
The optimum performance of an optical oxygen sensor based on polysulfone (PSF)/[Ru(II)-Tris(4,7-diphenyl-1,10-phenanthroline)] octylsulfonate (Ru(dpp)OS) was checked by carefully tuning the parameters affecting the membrane preparation. In particular, membranes having thickness ranging between 0.2 and 8.0 microm with various luminophore concentrations were prepared by dip-coating and tested. The membrane thickness was controlled by tuning the solution viscosity, and was measured both by secondary ion mass spectrometry (SIMS) and by visible spectroscopy (Vis). Luminescence-quenching-based calibration was a single value of the Stern-Volmer constant (K'SV) for membranes containing up to 20 mmol Ru(dpp) g-1 PSF (1.35 microm average thickness). The K'SV value decreased for larger concentration. The highest sensitivity was obtained with membrane thickness around 1.6 microm, having a response time close to 1 s. Thicker membranes exhibited an emission saturation effect and were characterized by longer response time. The K'SV behavior was interpreted on the basis of a mathematical approach accounting for the contribution of luminescence lifetime (tau0), oxygen diffusion coefficient (DO2) and oxygen solubility inside the membrane (sO2) establishing the role of all of them and allowing their experimental determination. Moreover, a simple experimental way to estimate K'SV without needing calibration was proposed. It was based either on the light emission asymmetry or on the percent variation of light emission on passing from pure nitrogen to pure oxygen.
In this paper, the synthetic route of a potential antitumor reagent, benzo[b][1,10] phenanthrolin-7(12H)-one (BPO), was improved. A sulfonic group was introduced to BPO to form a new compound, 7-oxobenzo[b][1,10]phenan-throline-12(7H)-sulfonic acid (OPSA), in order to enhance its water-solubility. The molecular structure of OPSA has been confirmed by IR, UV, MS, (1)H NMR and elements analysis. It was proved in our experiments that DNA could quench the fluorescence of OPSA and the maximum quenched intensity appeared at 408 nm (lambda(ex)=284 nm). The quenched fluorescence intensity was proportional to the concentration of DNA. Based on this phenomenon, OPSA had been used as the fluorescent probe for detection of calf thymus DNA (ct-DNA) and the corresponding linear response range was from 1.0 to 150.0 microg mL(-1) and the limit of detection (LOD) was 3.8 ng mL(-1). Its interaction with ct-DNA was investigated by fluorescence, absorption and viscosity measurements. When binding to ct-DNA, OPSA showed obvious fluorescence quenching and the quenched intensity was stable with the presence and absence of NaCl. The absorption spectra of OPSA had no evidence of increasing or decreasing when ct-DNA was added. The viscosity of OPSA and ct-DNA mixture showed no obvious change comparing with the viscosity of ct-DNA along. The results suggested that the interaction between OPSA and ct-DNA was groove binding in nature. Scatchard plots constructed from fluorescence titration data gave a binding constant of 8.9 x 10(5) L mol(-1) and a binding site size of 0.35 base pairs per bound drug molecule.
Previous studies have suggested that tris(4,7-diphenyl-1,10-phenanthrolinedisulfonate)ruthenium(II) (Ru(BPS)(3)(4-)) has great potential as a chemiluminescence reagent in acidic aqueous solution. We have evaluated four different samples of this reagent (two commercially available and two synthesised in our laboratory) in comparison with tris(2,2'-bipyridine)ruthenium(II) (Ru(bipy)(3)(2+)) and tris(1,10-phenanthroline)ruthenium(II) (Ru(phen)(3)(2+)), using a range of structurally diverse analytes. In general, Ru(BPS)(3)(4-) produced more intense chemiluminescence, but the oxidised Ru(BPS)(3)(3-) species is less stable in aqueous solution than Ru(bipy)(3)(3+) and produced a greater blank signal than Ru(bipy)(3)(3+) or Ru(phen)(3)(3+), which had a detrimental effect on sensitivity. Although the complex is often depicted with the sulfonate groups of the BPS ligand in the para position on the phenyl rings, NMR characterisation revealed that the commercially available BPS material used in this study was predominantly the meta isomer.
This study proposes a method for identifying 1,3-dichloro-2-propanol and 3-chloro-1,2-propandiol in aqueous matrices by using headspace on-fiber derivatization following solid-phase microextraction combined with gas chromatography-mass spectrometry. The optimized SPME experimental procedures for extracting 1,3-dichloro-2-propanol and 3-chloro-1,2-propandiol in aqueous solutions involved a 85 microm polyacrylate-coated fiber at pH 6, a sodium chloride concentration of 0.36 g mL(-1), extraction at 50 degrees C for 15 min and desorption of analytes at 260 degrees C for 3 min. Headspace derivatization was conducted in a laboratory-made design with N-methyl-N-(trimethylsilyl)-trifluoroacetamide vapor following solid-phase microextraction by using 3 microL N-methyl-N-(trimethylsilyl)-trifluoroacetamide at an oil bath temperature of 230 degrees C for 40 s. This method had good repeatability (R.S.D.s < or = 19%, n = 8) and good linearity (r2 > or = 0.9972) for ultrapure water and soy sauce samples that were spiked with two analytes. Detection limits were obtained at the ng mL(-1). The result demonstrated that headspace on-fiber derivatization following solid-phase microextraction was a simple, fast and accurate technique for identifying trace 1,3-dichloro-2-propanol and 3-chloro-1,2-propandiol in soy sauce.
A new method for the simultaneous determination of 1,4-dichlorobenzene (p-DCB), naphthalene and 1,2-dibromoethane (1,2-DBE) residues in honey has been developed. Analysis is carried out using gas chromatography-mass spectrometry (GC/MS) in selected ion monitoring mode (SIM), after extraction and preconcentration of target analytes by headspace solid-phase microextraction (HS-SPME), with a 100 microm film thickness polydimethylsiloxane (PDMS) fiber. Several parameters affecting the extension of the adsorption process (i.e., addition of salt, extraction time, extraction temperature) were studied. The optimal conditions for the determination of these analytes were established. The proposed HS-SPME method showed good sensitivity, without carryover between the samples. Linearity was studied from 5 to 2500 microg kg(-1) for p-DCB, 0.5 to 500 microg kg(-1) for naphthalene and 5 to 500 microg kg(-1) honey for 1,2-DBE with correlation coefficients (r(2)) ranging from 0.9901 to 0.9999. Precision was assessed and both intra and inter-day R.S.D.s (%) were below 6.3%. The detection limits were found to be 1, 0.1 and 2 microg kg(-1) honey for p-DCB, naphthalene and 1,2-DBE, respectively. The percentage recoveries that were evaluated with the proposed HS-SPME method and the standard addition calibration technique gave values among 72.8 and 104.3% for measurements in samples spiked with one target analyte or mixtures of the three. This method has been applied for the analysis of unknown honey samples. The results showed an excellent applicability of the proposed method for the determination of the target compounds in honey samples.
5,11,17,23-Tetrakis(1,1-dimethylethyl)-25,26-dihydroxy-27,28-crown-4-calix[4]arene in the cone conformation was synthesized. This p-tert-butylcalix[4]arene-1,2-crown-4 compound was then anchored with Merrifield chloromethylated resin beads. The modified polymeric resin was characterized by (1)H NMR, FT-IR and elemental analysis and used successfully for the separation and preconcentration of Cu(II), Cd(II), Co(II), Ni(II) and Zn(II) prior to their determination by FAAS. Effective extraction conditions were optimized in both batch and column methods. The resin exhibits good separating ability with maximum between pH 6.0-7.0 for Cu(II), pH 6.0 for Cd(II), pH 5.0 for Co(II), pH 4.0-4.5 for Ni(II), and pH 4.5 for Zn(II). The elution studies were carried out with 0.5 mol L(-1) HCl for Cu(II), Co(II) and Co(II), 1.0 mol L(-1) HCl for Cd(II) and Zn(II). The sorption capacity, preconcentration factor and distribution coefficient of each metal ion were determined. The detection limits were 1.10, 1.25, 1.83, 1.68 and 2.01 microg L(-1) for Cu(II), Cd(II), Co(II), Ni(II) and Zn(II). The influence of several ions on the resin performance was also investigated. The validity of the proposed method was checked for these metal ions in NIST standard reference material 2709 (San Joaquin Soil) and 2711 (Montana Soil).
A new method has been established for the determination of dopamine hydrochloride (DPH) using sodium 1,2-naphthoquinone-4-sulfonate (NQS) and tetradecyl benzyl dimethyl ammonium chloride (Zeph). This method is based on the formation of a pink compound from the reaction of DPH, sodium 1,2-naphthoquinone-4-sulfonate and Zeph. The condensation reaction proceeds quantitatively in pH 9.40 buffer solution. The maximum absorption wavelength and the value of epsilon491 were 491 nm, and 7.51x10(3) l mol(-1) cm(-1), respectively, when the stoichiometric ratio of the reaction was 1:1:1. Beer's law was obeyed in the range of 0.16-40 mg l(-1) of DPH. The data have been filled to a linear regression equation A=0.5781+0.0254 C (mg l(-1)), with a correlation coefficient of 0.9993. The detection limit is 0.12 mg l(-1), R.S.D. is 0.64% (40 mg l(-1), n=11), and average recovery is over 99.7%. This paper further improves the determination of DPH compared to the previous methods. The kinetic property and reaction mechanism have also been discussed. This proposed method has been successfully applied to the determination of DPH in injection and biological samples with satisfactory results.
A novel and simple spectrophotometric method for the determination of Captopril with sodium 1,2-naphthoquinone-4-sulfonate is established in this paper. The detailed reaction mechanism is proposed and discussed. It is based on the fact that captopril can catalyze the reaction between sodium 1,2-naphthoquinone-4-sulfonate and hydroxyl ion to form 2-hydroxy-1,4-naphthoquinone in buffer solution of pH 13.00. Beer's law is obeyed in a range of 0.64-80 mugmL(-1) at the maximal absorption wavelength of 442 nm. The equation of linear regression is A=0.05815+0.00589C (mugmL(-1)), with a linear regression correlation coefficient of 0.9981. The detection limit is 0.3 mugmL(-1), R.S.D. is 0.77% and the recovery rate is in range of 96.0-103.8%. Furthermore, the method has been validated and successfully applied to the determination of captopril in pharmaceutical samples.
Both 3-monochloropropane-1,2-diol (3-MCPD) and acrylamide are contaminants found in heat-processed foods and their related products. A quantitative method was developed for the simultaneous determination of both contaminants in food by gas chromatography-triple quadrupole mass spectrometry (GC-MS/MS). The analytes were purified and extracted by the matrix solid-phase dispersion extraction (MSPDE) technique with Extrelut NT. A coupled column (a 3m Innowax combined with a 30m DB-5ms) was developed to separate both compounds efficiently without derivatization. Triple quadrupole mass spectrometry in multiple reaction monitoring mode (MRM) was applied to suppress matrix interference and obtain good sensitivity in the determination of both analytes. The limit of detection (LOD) in the sample matrix was 5μg kg(-1) for 3-MCPD or acrylamide. The average recoveries for 3-MCPD and acrylamide in different food matrices were 90.5-107% and 81.9-95.7%, respectively, with the intraday relative standard deviations (RSDs) of 5.6-13.5% and 5.3-13.4%, respectively. The interday RSDs were 6.1-12.6% for 3-MCPD and were 5.0-12.8% for acrylamide. Both contaminants were found in samples of bread, fried chips, fried instant noodles, soy sauce, and instant noodle flavoring. Neither 3-MCPD nor acrylamide was detected in the samples of dairy products (solid or liquid samples) and non-fried instant noodles.
Highly selective and sensitive poly(vinyl chloride) (PVC) membrane electrodes based on 6-methyl-4-{[1-(2-thienyl)methylidene]amino}3-thioxo-3,4-dihydro-1,2,4-triazin-5-(2H)-one (MATDTO) as new carriers for gadolinium ion-selective electrode was reported. The membrane solutions containing PVC, o-nitrophenyl octyl ether (NPOE) as plasticizer, sodium tetraphenyl borate (NaTPB) as lipophilic ionic additive, and MATDTO, displays a calibration response for Gd3+ ions over a wide concentration range of 1.0x10(-6)-1.0x10(-1) M with Nernstian slopes of 19.8+/-0.2 mV per decade and a detection limit as 5.8x10(-7) M. The sensor has a relatively fast response time of <10 s and can be used in the pH range 3.2-8.7 for at least 2 months without any significant divergence in potentials. The selectivity coefficients for mono-, di-, and trivalent cations indicate good selectivity for Gd(III) ions over a large number of interfering cations. The membrane sensor was used as an indicator electrode in the potentiometric titration of Gd(III) ions with EDTA. The proposed electrode was also applied to the determination of concentration of Gd(III) ions in soil and sediment samples and validation with CRMs.
The Raman and mass spectra of 1,4-diphenylbuta-1,3-diene and several of its monomethyl and dimethyl homologues are reported and discussed, with a view to developing a spectroscopic protocol for detecting the presence and position of a methyl group in these compounds. Raman spectroscopy and mass spectrometry are shown to provide complementary information, by which the four available monomethyl homologues may be readily distinguished from each other and 1,4-diphenylbuta-1,3-diene itself. The utility of these 1,4-diarylbutadienes as model compounds for carotenoids and related materials, which may serve as indicators of extinct or extant extraterrestrial life, is considered.
A new catalytic oxidative coupling reaction of N,N-dimethyl-p-phenylenediamine (DPD) with 1,3-phenylenediamine (mPD) in the presence of hydrogen peroxide has been developed for trace metals analysis. The rate of the oxidation/coupling reaction can be enhanced significantly by iron, copper and cobalt. These metal ions can catalyze the oxidation reaction of DPD to form an oxidized product; the oxidized DPD was then coupled with mPD to give a blue-colored product which was measured spectrophotometrically at 650 nm. On the basis of such a reaction scheme, two simple flow injection analysis methods for the determination of copper and iron have been developed. Detailed studies on chemical and FIA variables affecting the sensitivity of the detection were carried out. Interferences from several ionic species were examined for the determination of copper: the interference effect by Fe(III) and Fe(II) up to 1.5 mg L(-1) was successfully suppressed by pretreating sample with ammonium acetate buffer solution (pH 8.4). Good linearity of a standard calibration graph was obtained over the ranges of 0-8 and 0-2 microg L(-1) of copper and iron, respectively, and the detection limits were 0.05 and 0.02 microg L(-1) for copper and iron, respectively. The precision of the methods in terms of relative standard deviation were 1.4 and 1.5% of R.S.D. which were obtained from 10 injections of 2.0 and 1.0 microg L(-1) of standard copper and iron, respectively. The proposed methods were successfully applied to the determination of copper and iron in tap and river water samples. The accuracy of the proposed methods was assessed by the analysis of certified reference material of river water.
N,N'-bis-(α-methylsalicylidene)-2,2-dimethyl-1,3-propanediimine (SBTD) modified silica gel was prepared and used as sorbent for solid phase extraction of Cr(III) ions from aqueous solution. This sorbent showed a high sorption affinity for Cr(III) while recovery of Cr(VI) was very low. The analyte ion retained on the column was eluted with 1 mol L(-1) HNO(3). The chromium ion in the eluent was determined by graphite furnace atomic absorption spectrometry. The effects of different parameters such as pH, eluent type and volume, Schiff's base concentration, sample and eluent flow rate, interfering ions and adsorbent amount were investigated.
A simple, fast, low cost and sensitive direct beta-correction spectrophotometric assay of cyanide ions based on its reaction with the reagent 4-hydroxy-3-(2-oxoindolin-3-ylideneamino)-2-thioxo-2H-1,3-thiazin-6(3H)-one, abbreviated as HOTT in aqueous media of pH 7-10 is described. The electronic spectrum of the produced brown-red colored species showed well defined and sharp peak at lambda(max)=466 nm. The effective molar absorptivity for the produced cyano compound was 2.5 x 10(4) L mol(-1) cm(-1). Beer's law and Ringbom's plots were obeyed in the concentration range 0.05-2.0 and 0.30-1.5 microg mL(-1) cyanide ions, respectively. The proposed method offers 16.0 and 50.3 microg L(-1) lower limits of detection (LOD) and quantification (LOQ) of the cyanide ion, respectively. The analytical utility of the method for the analysis of cyanide ions in tap and drinking water samples was demonstrated and the results were compared successfully with the conventional cyanide ion selective electrode. The short time response and the detection by the naked eye make the method available for the detection and quantitative determination of cyanide in a variety of samples e.g. fresh and drinking water. Moreover, the structure of the produced colored species was determined with the aid of spectroscopic measurements (UV-Vis, IR, 1H and 13C NMR) and elemental analysis.
This paper describes the simultaneous determination of epinephrine (EP), uric acid (UA) and xanthine (XN) in the presence of ascorbic acid (AA) using electropolymerized ultrathin film of 5-amino-1,3,4-thiadiazole-2-thiol (p-ATT) modified glassy carbon (GC) electrode in 0.2 M phosphate buffer solution (pH 5). Although bare GC electrode resolves the voltammetric signals of AA and XN, it fails to resolve the voltammetric signals of EP and UA in a mixture. However, the p-ATT modified electrode not only separates the voltammetric signals of AA, EP, UA and XN with potential difference of 150, 120 and 400 mV between AA-EP, EP-UA and UA-XN, respectively but also shows higher oxidation current for these molecules. The p-ATT modified electrode exhibits excellent selectivity towards the oxidation of EP, UA and XN in the presence of 40-fold higher concentration of AA. Further, the p-ATT modified electrode was also used for the selective determination of EP in the presence of 40-fold higher concentrations of AA, UA and XN. Using amperometric method, we achieved the lowest detection of 40 nM EP and 60nM each UA and XN. The amperometric current response was increased linearly with increasing EP concentration in the range of 4.0 x 10(-8) to 4.0 x 10(-5) M and the detection limit was found to be 27 x 10(-11) M (S/N = 3). The practical application of the present modified electrode was demonstrated by determining the concentration of EP in epinephrine tartrate injection and XN in human urine samples.
This work reports the development of a selective, sensitive and rapid spectrofluorimetric method for the determination of reduced glutathione (GSH) in the presence of relatively high levels of cysteine (Cys) in clinical and biological samples using 1,3,5,7-tetramethyl-8-phenyl-(2-maleimide)-difluoroboradiaza-s-indacene (TMPAB-o-M). The fluorescence from TMPAB-o-M is strongly quenched by its maleimide moiety, but after reaction with thiol, the fluorescence is restored with a 350-fold intensity increase (fluorescence quantum yield from 0.002 to 0.73). In H(3)Cit-Na(2)HPO(4) buffer (pH 7.40), the derivatization is completed in just 5 min under 37 degrees C. The linear range is 0.005-0.2 micromol L(-1), with detection limit of 1.1 x 10(-10)mol L(-1) (signal-to-noise ratio=3). Almost all amino acids, including Cys, impose no interference even if present at relatively high concentrations (amino acids:GSH=100:1, Cys:GSH=1:1, molar ratio, C(GSH)=3 x 10(-7)mol L(-1)). The sample can be used directly without further treatment after the protein is removed. The developed method is precise with a relative standard deviation (R.S.D.) lower than 5.0% (n=6) and has been applied to the determination of GSH in human blood and pig's liver with recoveries between 94.4 and 105.6%.
A precolumn-derivatization high performance liquid chromatographic method for biogenic amine analysis has been developed. Derivatization of biogenic amines was performed with a new fluorescent reagent, 8-phenyl-(4-oxy-acetic acid N-hydroxysuccinimide ester)-4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene (TMPAB-OSu), which was synthesized in our lab. The derivatization conditions and the influence of elution composition on the separation were investigated. In pH 8.5 H3BO3-Na2B4O7 media, the reaction of biogenic amines with TMPAB-OSu completed at 45 degrees C for 5 min. The labeled biogenic amines were separated on an ZORBAX Eclipse XDB-C8 column (150 mm x 4.6 mm, 5 microm) and detected with fluorescence at excitation and emission wavelengths of 497 and 509 nm, respectively. Detection limits of biogenic amines were 0.1-4 nmol L(-1), at a signal-to-noise ratio of 3. The proposed method has been applied to the quantitative determination of spermine, phenethylamine, spermidine, cadaverine and putrescine in pericarp and pulp of mature and immature apples and wine with recoveries of 95-102% and R.S.D. of 0.7-3.9.
The objective of this study was to create a thin film optode for fast pH measurements that meets the requirements for imaging pH-responses from cells as well as for a future hybrid design for detection of multiple analytes simultaneously. The sensor is based on the covalent attachment of 8-hydroxypyrene-1,3,6-trisulfonate (HPTS) to a film forming cellulose acetate material through a sulfonamide linkage. The synthesis routes of the cellulose material and regio-specific covalent attachment of the dye are described in detail. The sensor was sterilized in two different ways and showed excellent biocompatibility with Chinese hamster ovary cells. Imprints from cells and mum thin cell extensions were visualized when altering pH of the surrounding solution. The sensor was tested together with time-dependent sigmoidal calibrations giving pH determinations with an exceptional precision and accuracy during measurement within pH 6-8. The precision of the optode, calculated as pooled S.D. (n=8) according to IUPAC recommendations between pH 6.641 and 7.742 was 0.0029. The accuracy was significantly better than the electrode used as reference during the measurements. The response time (0-95%) was 100s between pH 6.641 and 7.742 and the reverse response (95-0%) was 80s. The sensor shows great potential for extra-cellular pH determination over time during cell growth and pharmacological exposure.
The data presented for the behavior of 3-hydroxy-1,4-benzodiazepines during non-aqueous titrations indicate that: (a) the First stage protonation during titration as a base with perchloric acid occurs at the N—4 position and is relatively unaffected by substitution at the 3-position, except when the substituent is electron-withdrawing; (b) deprotonation during titration as an acid with tetrabutylammonium hydroxide occurs at the N-1 position rather than at the 3-OH substituent; (c) the similarity of the u.v. spectra of neutral and basic solutions of lorazepam indicate that a tautomerism in basic solution postulated previously does not occur to any great extent; and (d) titration with tetrabutylammonium hydroxide should be acceptable as an assay method tor lorazepam.
The immobilization conditions and kinetic behaviour of trypsin, covalently immobilized via the 1,4-diisothiocyanatobenzene (DITC) linker onto aminopropylated controlled pore glass (CPG) particles, have been evaluated to establish a rapid and efficient protocol for fabrication of an immobilized enzyme microreactor (IMER) for protein hydrolysis and subsequent peptide mapping. Addition of calcium ions to either the immobilization reaction solution or hydrolysis assay was studied for a synthetic substrate. Activity was slightly higher when immobilization was carried out in the presence of Ca(2+) whereas more enzyme could be immobilized in its absence. A protocol requiring less than 3 h was devised to obtain maximal enzymatic activity with the lowest ratio of soluble trypsin to DITC-CPG particles. The resulting immobilized enzyme was found to retain an acceptable percentage (ca. 35%) of its activity after immobilization. The particles were dry-packed into a capillary to make a microscale IMER. Repeatability, reusability and digestion efficiency of the microIMER were investigated for the substrate beta-casein using capillary electrophoretic-based peptide mapping. In initial tests, a single device showed reproducible peptide maps for 21 digestions lasting 2 h each, carried out over a period of 2 months. Complete digestion of beta-casein could be achieved in a few minutes (86 s residence time in the microIMER followed by a wash step).
Uptake for lead, copper, cadmium, nickel and manganese from aqueous solution using the Moringa oleifera seeds biomass (MOSB) and amine-based ligand (ABL) was investigated. Experiments on two synthetic multi-solute systems revealed that MOSB performed well in the biosorption and followed the decreasing orders Pb(II)>Cu(II)>Cd(II)>Ni(II)>Mn(II) and Zn(II)>Cu(II)>Ni(II). The general trend of the heavy metal ions uptake by the amine-based ligand followed decreased in the order Mn>Cd>Cu>Ni>Pb, which is the reverse trend for what was observed for MOSB. Comparing the single- and multi-metal solutions, there was no clear effect in the biosorption capacity of MOSB suggesting the presence of sufficient active binding sites for all metal ions studied. The MOSB performance is also not affected by pH in the range 3.5-8.
Synthesis and application of 2-amino-3-(alpha-N-phenylmethyl-2'-amino-1',4'-naphthoquinonyl)-1,4 naphthoquinone (S) as a neutral ionophore for the determination of gallium(III) in PVC-based membrane sensors has been described. The sensor based on membrane composition (w/w, mg%); 5.0 (S):30.0 (PVC):5.0 (KTpClPB):60.0 (o-NPOE) is the best and showed a working range of 2.3x10(-7) to 1.0x10(-2) M with detection limit of 1.2x10(-7) M. It can tolerate non-aqueous media up to 15% with a slope of 19.7 mV decade(-1) of activity. The sensor has been used to assess the Ga(III) concentration in different natural samples (peach and tomato leaves, coal-fly-ash and river sediments). It can be used for 2.5 months without any distortion in results, after which, leaching of ionophore was observed from the membrane phase. The proposed sensor has shown a good dynamic response time of 11 s.
A simple, fast, sensitive and robust analytical method using gas chromatography (GC)-isotope dilution mass spectrometry (MS) was developed and validated for the identification and quantification of 1,4-dichlorobenzene (p-DCB) residues in honey samples. The proposed methodology is based on steam-distillation using a Clevenger-type apparatus followed by gas chromatography-mass spectrometry (GC-MS) in the selected ion monitoring (SIM) mode employing the isotopically labeled analogue d4-1,4-dichlorobenzene (d4-p-DCB) as internal standard (IS). Validation of the method was performed in two different GC-MS systems, using quadrupole MS (QMS) and ion-trap MS (ITMS) detectors, with no statistically significant differences between two. Recoveries were better than 91% with percent relative standard deviations lower than 12%. The instrumental limits of detection were 1 microg kg(-1) in the GC-ITMS system and 0.6 microg kg(-1) in the GC-QMS system. The expanded uncertainty was estimated as 17% at the currently accepted "action level" of 10 microg kg(-1). The method was applied to the analysis of 310 honey samples in an extensive national monitoring study. A quality control (QC) system applied during the assays has demonstrated a good performance and long-term stability over a period of more than 8 months of continuous operation.
Ant colony optimization (ACO) is a meta-heuristic algorithm, which is derived from the observation of real ants. In this paper, ACO algorithm is proposed to feature selection in quantitative structure property relationship (QSPR) modeling and to predict lambda(max) of 1,4-naphthoquinone derivatives. Feature selection is the most important step in classification and regression systems. The performance of the proposed algorithm (ACO) is compared with that of a stepwise regression, genetic algorithm and simulated annealing methods. The average absolute relative deviation in this QSPR study using ACO, stepwise regression, genetic algorithm and simulated annealing using multiple linear regression method for calibration and prediction sets were 5.0%, 3.4% and 6.8%, 6.1% and 5.1%, 8.6% and 6.0%, 5.7%, respectively. It has been demonstrated that the ACO is a useful tool for feature selection with nice performance.
A differential pulse polarographic examination of 1,4-benzodiazepines structurally related to medazepam. chlordiazepoxide and N-desmethyldiazepam is presented. The data are used to correlate polarographic peak potentials (Ep) and Hammett substituent constants for the three series of compounds. Good correlations were found for the reduction of the 4,5-azomethine functional group for the three series of compounds in pH 3 and pH 7 supporting electrolyte. No correlation could be established for the reduction of the 1,2-azomethine and N4-oxide in the series of compounds structurally related to chlordiazepoxide. Polarographic data are also presented for two groups of benzodiazepines structurally related to medazepam and N-desmethyldiazepam with heterocyclic substituents in the 5-position.
Oxazepam is the major metabolite screened in urine samples for the evidence of the use of benzodiazepine drugs. The methods currently used, however, are laborious and time consuming. This paper proposes an oxazepam detection method based on its hydrolysis and cyclization--a reaction catalysed by cerium (IV) in an ortho-phosphoric acid-containing medium--to form 2-chloro-9(10H)-acridinone, a strongly fluorescent molecule. The variables involved in the hydrolysis and cyclization stages were optimised. Oxazepam was detectable in the 5-900 ng mL(-1) range, with a detection limit of 4.15 ng mL(-1) for k=3. The method was successfully used for the determination of oxazepam in urine samples collected at different times after the oral administration of Valium and Tranxilium.
The use of 2,7-dimethyl-3,8-dinitrodipyrazolo[1,5-a:1',5'-d]pyrazine-4,9-dione as pre-column reagent for LC analysis of amino acids is proposed. The compound reacts (30 min at 68 degrees C in presence of 0.04 M triethylamine in a dimethylsulfoxide-water mixture) with primary and secondary amino function and the stable resulting adducts can be chromatographed under reversed-phase conditions and detected at lambda=280 nm. The derivatization conditions were optimized by a series of experiments. The effect of temperature, triethylamine concentration and reagent on the reaction was investigated. The yield of the glycine derivative was found to be quantitative at a reagent amino acid molar ratio of about 6 by comparison with an authentic specimen of synthesized glycine adduct. Application of the method to quality control of commercially available oral polyaminoacid formulations is described.
A selective detection method for dopamine (DA) was developed by incorporating cibacron blue (F3GA) into poly-1,5-diaminonaphthalene (PDAN) layer. Scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), and cyclic voltammetry (CV) were employed to characterize the modified surfaces. The modified electrode was effective in selectively facilitating the electron transfer of DA and blocking the interferences of negatively charged species attributed to the sulfonate groups in the F3GA/PDAN composite film. This method enabled the determination of DA in the presence of various interfering species, including ascorbic acid (AA), in a phosphate buffer solution (pH 7.4). The modified electrode demonstrated good performance in the detection of DA in a concentration range of 5.0-100 microM, with a detection limit (k=3) of 0.1+/-0.01 microM. The application was conducted for the determination of DA in a human urine sample and the sensor was proven to be rapid, has excellent selectivity, and stable amperometric response.
In this paper, we report a double-receptor sandwich type fluorescence sensing method for the determination of fructose bisphosphates (FBPs) using fructose 1,6-bisphosphate (F-1,6-BP) as a model analyte based on uranyl-salophen complexes. The solid phase receptor is an immobilized uranyl-salophen (IUS) complex which is bound on the surface of glass slides by covalent bonds. The labeled receptor is another uranyl-salophen complex containing a fluorescence group, or uranyl-salophen-fluorescein (USF). In the procedure of determining F-1,6-BP in sample solution, F-1,6-BP is first adsorbed on the surface of the glass slide through the coordination reaction of F-1,6-BP with IUS. It then binds USF through another coordination reaction to form a sandwich-type structure of IUS-F-1,6-BP-USF. The amount of F-1,6-BP is detected by the determination of the fluorescence intensity of IUS-F-1,6-BP-USF bound on the glass slide. Under optimal conditions, the linear range for the detection of F-1,6-BP is 0.05-5.0nmolmL(-1) with a detection limit of 0.027nmolmL(-1). The proposed method has been successfully applied for the determination of F-1,6-BP in real samples with satisfactory results.
The effectiveness of a novel binary matrix composed of 1,8-bis(dimethylamino)naphthalene (DMAN; proton sponge) and 9-aminoacridine (9AA) for the direct lipid analysis of whole bacterial cells by matrix assisted laser desorption ionization mass spectrometry (MALDI MS) is demonstrated. Deprotonated analyte signals nearly free of matrix-related ions were observed in negative ion mode. The effect of the most important factors (laser energy, pulse voltage, DMAN/9AA ratio, analyte/matrix ratio) was investigated using a Box-Behnken response surface design followed by multi-response optimization in order to simultaneously maximize signal-to-noise (S/N) ratio and resolution. The chemical surface composition of single or mixed matrices was explored by X-ray photoelectron spectroscopy (XPS). Moreover, XPS imaging was used to map the spatial distribution of a model phospholipid in single or binary matrices. The DMAN/9AA binary matrix was then successfully applied to the analysis of intact Gram positive (Lactobacillus sanfranciscensis) or Gram negative (Escherichia coli) microorganisms. About fifty major membrane components (free fatty acids, mono-, di- and tri-glycerides, phospholipids, glycolipids and cardiolipins) were quickly and easily detected over a mass range spanning from ca. 200 to ca. 1600m/z. Moreover, mass spectra with improved S/N ratio (compared to single matrices), reduced chemical noise and no formation of matrix-clusters were invariably obtained demonstrating the potential of this binary matrix to improve sensitivity.
The electroreduction of 2-phenylindan-1.3-dione has been investigated by polarography, cyclic voltammetry and coulometry at controlled potential. In citrate buffer pH 4.9, the drug is reduced in two welI-defined 2-electron steps. The height of the first polarographic wave is independent of pH and proportional to the concentration. in the range 10-6 2.5 · lO-4 M. The drug is strongly adsorbed at the electrode surface, and can therefore be determined in the presence of surface-active substances like proteins. A procedure has been developed for the direct polarographic determination of microgram quantities of the drug in serum. The proposed method is very rapid and accurate and permits determination of 4– 500 μg per ml of the drug in serum.
This manuscript presents a response to a recently published article in Analytica Chimica Acta by J. Meija and Z. Mester (doi:10.1016/j.aca.2010.09.029) in which the authors comment on some aspects of our previous work entitled "Determination of the uncertainties in the theoretical mass isotopomer distribution of molecules" Anal. Chim. Acta 664 (2010) 68-76. We present here new arguments to support our previous findings and to clarify some aspects regarding to the calculation of the uncertainty of isotope patterns. The uncertainty calculations proposed by Meija and Mester for carbon-only clusters have been compared with our previously developed first-order model based on the spreadsheet approach described by Kragten (J. Kragten, Analyst 119 (1994) 2161). The results obtained in this comparison demonstrate that the Kragten procedure can be applied safely for the calculation of the uncertainties in the mass isotopomer distribution of molecules when the u average value is calculated. In addition the procedure provides good results even when the first-order error propagation law failed for some particular cases (C(93) and C(186) clusters).
A detailed analysis of the (35)Cl/(37)Cl isotope effects observed in the 19.11 MHz (103)Rh NMR resonances of [RhCl(n)(H(2)O)(6-n)](3-n) complexes (n=3-6) in acidic solution at 292.1K, shows that the 'fine structure' of each (103)Rh resonance can be understood in terms of the unique isotopologue and in certain instances the isotopomer distribution in each complex. These (35)Cl/(37)Cl isotope effects in the (103)Rh NMR resonance of the [Rh(35/37)Cl(6)](3-) species manifest only as a result of the statistically expected (35)Cl/(37)Cl isotopologues, whereas for the aquated species such as for example [Rh(35/37)Cl(5)(H(2)O)](2-), cis-[Rh(35/37)Cl(4)(H(2)O)(2)](-) as well as the mer-[Rh(35/37)Cl(3)(H(2)O)(3)] complexes, additional fine-structure due to the various possible isotopomers within each class of isotopologues, is visible. Of interest is the possibility of the direct identification of stereoisomers cis-[RhCl(4)(H(2)O)(2)](-), trans-[RhCl(4)(H(2)O)(2)](-), fac-[RhCl(3)(H(2)O)(3)] and mer-[RhCl(3)(H(2)O)(3)] based on the (103)Rh NMR line shape, other than on the basis of their very similar δ((103)Rh) chemical shift. The (103)Rh NMR resonance structure thus serves as a novel and unique 'NMR-fingerprint' leading to the unambiguous assignment of [RhCl(n)(H(2)O)(6-n)](3-n) complexes (n=3-6), without reliance on accurate δ((103)Rh) chemical shifts.
With the technique of metabolomics, gas chromatography/mass spectrometry (GC/MS), urine or serum metabolites can be assayed to explore disease biomarkers. In this work, we present a metabolomic method to investigate the urinary metabolic difference between hepatocellular carcinoma (HCC, n - 20) male patients and normal male subjects (n - 20). The urinary endogenous metabolome was assayed using chemical derivatization followed by GC/MS. After GC/MS analysis, 103 metabolites were detected, of which 66 were annotated as known compounds. By a two sample t-test statistics with p < 0.05, 18 metabolites were shown to be significantly different between the HCC and control groups. A diagnostic model was constructed with a combination of 18 marker metabolites or together with alphafetoprotein, using principal component analysis and receiver-operator characteristic curves. The multivariate statistics of the diagnostic model yielded a separation between the two groups with an area under the curve value of 0.9275. This non-invasive technique of identifying HCC biomarkers from urine may have clinical utility.
The mixed phenylpropanoid polymer lignin is one of the most abundant biopolymers on the planet and is used in the paper, pulp and biorenewable industries. For many downstream applications, the lignin monomeric composition is required, but traditional methods for performing this analysis do not necessarily represent the lignin composition as it existed in the plant. Herein, it is shown that Raman spectroscopy can be used to measure the lignin monomer composition. The use of 1064 nm excitation is needed for lignin analyses since high fluorescence backgrounds are measured at wavelengths as long as 785 nm. The instrument used for these measurements is a 1064 nm dispersive multichannel Raman spectrometer that is suitable for applications outside of the laboratory, for example in-field or in-line analyses and using remote sensing fiber optics. This spectrometer has the capability of acquiring toluene/acetonitrile spectra with 800 cm(-1) spectral coverage, 6.5 cm(-1) spectral resolution and 54 S/N ratio in 10s using 280 mW incident laser powers. The 1135-1350 cm(-1) and 1560-1650 cm(-1) regions of the lignin spectrum can be used to distinguish among the three primary model lignin monomers: coumaric, ferulic and sinapic acids. Mixtures of the three model monomers and first derivative spectra or partial least squares analysis of the phenyl ring breathing modes around 1600 cm(-1) are used to determine sugarcane lignin monomer composition. Lignin extracted from sugarcane is shown to have a predominant dimethoxylated and monomethoxylated phenylpropanoid content with a lesser amount of non-methoxylated phenol, which is consistent with sugarcane's classification as a non-woody angiosperm. The location of the phenyl ring breathing mode peaks do not shift in ethanol, methanol, isopropanol, 1,4 dioxane or acetone.
Top-cited authors
William Vetterling
  • ZINK Holdings
Brian Flannery
  • Resources for the Future
William Press
  • University of Texas at Austin
Elias Zagatto
  • University of São Paulo
Lo Gorton
  • Lund University