Cysteine enhances clastogenic activity of dimethylarsinic acid

The effects of cysteine on dimethylarsinic acid (DMA)‐induced cytotoxicity and chromosomal aberration were studied using Chinese hamster V79 cells. The IC 50 of DMA, i.e. the concentration resulting in a 50% decrease in cell population of viable cells, was 130 µg ml^?1 (0.94 mM ), whereas that in the presence of 50 µg ml^?1 (0.28 mM ) cysteine was 20 µg ml^?1 (0.14 mM ). The mitotic index with co‐administration of 50 µg ml^?1 (0.36 mM ) DMA and 50 µg ml^?1 cysteine was 1.4 times that with 50 µg ml^?1 DMA alone. Whereas 82% of cells divided twice with 25 µg ml^?1 (0.18 mM ) DMA alone, most cells divided only once with co‐administration of 25 µg ml^?1 DMA and 50 µg ml^?1 cysteine. These results indicated that the increase in mitotic index by cysteine was due to enhancement of mitotic arrest by DMA. With co‐administration of 25 µg ml^?1 DMA and 50 µg ml^?1 cysteine, tetraploidy was 14.3% higher and fivefold by that with 25 µg ml^?1 DMA only. Cysteine at 50 µg ml^?1 enhanced induction of chromosomal aberrant cells by DMA. 100 µg ml^?1 (0.72 mM ) DMA induced 91% chromosomal aberrant cells in the presence of cysteine, and 12% in the absence of cysteine. Chromatid breaks and chromatid gaps were the most frequent types of aberration induced by co‐administration of DMA and cysteine or DMA alone. Co‐administration of DMA and cysteine produced many attenuated chromosomal figures. The attenuated chromosomal figures always had several chromatid gaps and chromatid breaks. Our findings may provide clues to arsenic carcinogenesis in humans. Copyright © 2002 John Wiley & Sons, Ltd.     

Synthetic and antimicrobial studies on new gold(I) complexes of imidazolidin‐2‐ylidenes

Six new 1,3‐diorganylimidazolidin‐2‐ylidene (NHC) gold(I) complexes of the type [Au(NHC)₂]⁺ (1–6), were synthesized by reacting [AuCl(PPh)₃] with 1,3‐dimesitylimidazolidin‐2‐ylidene or bis(1,3‐dialkylimidazolidin‐2‐ylidene). The complexes 1–6 were fully characterized by elemental analyses and spectroscopic data. The placement of mesityl or para‐substituted benzyl groups on the nitrogen atoms of the ring of the complexes leads to the particularly active antibacterial agents evaluated in this work. It is worth noting that the p‐methoxybenzyl derivative (2) inhibited the growth of Pseudomona aeruginosa, Staphylococcus epidermidis, Staphylococcus aureus and Enterococcus faecalis with minimum inhibitory concentration (MIC) values of 3.12 µg ml^?1, 6.25 µg ml^?1, 3.12 µg ml^?1 and 3.12 µg ml^?1 respectively. In contrast, the analogous p‐dimethylaminobenzyl derivative (3) is effective only against Escherichia coli (MIC = 3.12 µg ml^?1). Copyright © 2004 John Wiley & Sons, Ltd.     

Synthesis and cytotoxic activity of silacycloalkyl‐substituted heterocyclic aldehydes and their thiosemicarbazones

A series of 5‐[1‐methylsilacyclo‐pentyl/‐hexyl]‐2‐furfural, 5‐[1‐methylsilacyclo‐pentyl/‐hexyl]‐2‐thiophene carbaldehyde and 1,1‐bis(5‐formyl‐2‐furyl)silacyclo‐pentane/‐hexane and their thiosemicarbazones has been synthesized and subjected to antitumour assay. The effects of the substituents and the heterocycle were examined to establish structure–activity relationships. Thiosemicarbazones of 5‐(1‐methylsilacyclohexyl)furfural and 5‐(1‐methylsilacyclopentyl)furfural were very active (1.0–4.0 µg ml^?1) in vitro against human fibrosarcoma HT‐1080 and mouse hepatoma MG‐22A cells. At the same time, they were less toxic for normal fibroblasts 3T3. All compounds synthesized exhibited low or moderate toxicity (LD₅₀ 152–1904 mg kg^?1). Copyright © 2005 John Wiley & Sons, Ltd.     

Spectrophotometric Determination of Some Fluoroquinolone Derivatives in Dosage Forms and Biological Fluids Using Ion‐Pair Complex Formation

Abstract

A simple, rapid, sensitive, and reproducible procedure for assaying norfloxacin (NOR), ciprofloxacin (CIP), and ofloxacin (OFL) was investigated. The procedure is based on the reaction of selected drugs with Sudan II (I), Congo red (II), and Gentian violet (III) in universal buffer to give soluble ion‐pair complexes. The effects of various parameters have been studied. Beer's law plots were obeyed in the concentration ranges 0.5–11 µg ml^?1, whereas Ringbom optimum ranges were 0.7–9.5 µg ml^?1. The apparent molar absorptivity (6.4×10⁴ L mol^?1 cm^?1), Sandell sensitivity (4.99 ng cm^?2), detection (0.13 µg ml^?1), and quantification (0.44 µg ml^?1) limits were calculated. The relative standard deviation for ten determinations, for samples containing 4.0 µg ml^?1, was found to be 1.40%. The influence of commonly employed excipients in the determination of the studied drugs was examined. There was no interference from degradate product results from thermal and hydrolytic treatments. The results obtained by the proposed procedure were statistically validated. The developed procedure was successfully applied to the determination of the studied drugs in dosage forms and biological fluids.     

Antineoplastic activity of polyaspartamide–ferrocene conjugates

The ferrocene/ferricenium redox system plays a significant role in biological oxidation, reduction and free-radical reactions. Of particular interest are the findings of earlier investigations which showed certain water-soluble ferricenium salts to possess appreciable antiproliferative activity against various murine tumor lines and a xenografted human colorectal adenocarcinoma. Solubility in water, a prerequisite for efficacious transport and dissipation in central circulation, was then proposed as a principal requirement for the ferrocene complex system to exert antineoplastic activity irrespective of the oxidation state in which it is administered. In order to shed more light on this question, we decided to investigate the antiproliferative properties of polymer–ferrocene conjugates containing the metal complex in the non-oxidized (ferrocene) form while fulfilling the critical requirement of water solubility. To this end, five selected, water-soluble conjugates, synthesized by reversible coupling of 4-ferrocenylbutanoic acid to variously structured polyaspartamides featuring pendant primary amino groups as coupling sites, were tested in vitro against cultured HeLa cells at concentrations up to 50 µg Fe ml⁻¹. Optimal antiproliferative activities, with IC₅₀ in the range of 2–7 µg Fe ml⁻¹, were determined for three compounds possessing tertiary-amine functions susceptible to protonation at physiological pH. Lower activities (IC₅₀ = 45–60 µg Fe ml⁻¹) were demonstrated for two poly(ethylene oxide)-containing conjugates. However, no reasonable structure–performance relationships can be derived at this stage from the small number of compounds tested. Copyright © 1998 John Wiley & Sons, Ltd.     

Comparison of Ion-Pairing and Reversed Phase Liquid Chromatography in Determination of Sulfamethoxazole and Trimethoprim

Abstract

Two simple, rapid, and sensitive HPLC methods have been developed for the simultaneous determination of sulfamethoxazole and trimethoprim in their pure and dosage forms, one utilizing reversed phase HPLC and the other ion-pair HPLC. In the reversed phase HPLC method (A) the mobile phase consists of 0.05% aqueous solution of formic acid with pH adjusted to 4.5±0.2 with triethylamine : acetonitrile:tetrahydrofuran 50 : 49 : 1 (v/v), and the mobile phase pumped at flow rate of 1.0 ml min^?1. An Appolo LC18 column (5.0 µm), 250 mm length × 4.6 mm diameter, was utilized as the stationary phase. Detection was affected spectrophotometrically at 254 nm. In the ion-pair HPLC method (B) the mobile phase consisted of methanol : buffer 35 : 65 (v/v) with the buffer composed of potassium dihydrogen phosphate 0.3 M and sodium heptan sulfonic acid 5.0 mM. To 500 ml of buffer was added 2.0 ml triethylamine, and then the pH was adjusted to 5.0 with phosphoric acid, and the mobile phase was pumped at a flow rate of 1.2 ml min^?1. A Hypersil C₁₈ column (5.0 µm), 150 mm length × 4.6 mm diameter, was utilized as the stationary phase. Detection was affected spectrophotometrically at 254 nm. Linearity ranges for sulfamethoxazole and trimethoprim were 1.0–110 and 1.5–98 µg ml^?1, respectively, with method A and 0.5–100 and 1.0–125 µg ml^?1, respectively, with method (B). Minimum detection limits obtained were 0.1969 and 0.3451 µg ml^?1 for sulfamethoxazole and trimethoprim, respectively, with method A, and 0.1377 and 0.2454 µg ml^?1 with method (B). The proposed methods were further applied to the analysis of tablets containing the two drugs, and the results were satisfied.     

Biomethylation and biotransformation of arsenic in a freshwater food chain: Green alga (chlorella vulgaris)→shrimp (neocaridina denticulata)→killifish (oryzias iatipes)

Tolerance, bioaccumulation, biotransformation and excretion of arsenic compounds by the fresh–water shrimp (Neocaridina denticulata) and the killifish (Oryzias latipes) (collected from the natural environment) were investigated. Tolerances (LC₅₀) of the shrimp against disodium arsenate [abbreviated as As(V)], methylarsonic acid (MAA), dimethylarsinic acid (DMAA), and arsenobetaine (AB) were 1.5, 10, 40, and 150μg As ml^?1, respectively. N. denticulata accumulated arsenic from an aqueous phase containing 1 μg As ml^?1 of As(V), 10 μg As ml^?1 of MAA, 30 μg As ml^?1 of DMAA or 150 μg As ml^?1 of AB, and biotransformed and excreted part of these species. Both methylation and demethylation of the arsenicals were observed in vivo. When living N. denticulata accumulating arsenic was transferred into an arsenic–free medium, a part of the accumulated arsenic was excreted. The concentration of methylated arsenicals relative to total arsenic was higher in the excrement than in the organism. Total arsenic accumulation in each species via food in the food chain Green algae (Chlorella vulgaris) → shrimp (N. denticulata) → killifish (O. latipes) decreased by one order of magnitude or more, and the concentration of methylated arsenic relative to total arsenic accumulated increased successively with elevation in the trophic level. Only trace amounts of monomethylarsenic species were detected in the shrimp and fish tested. Dimethylarsenic species in alga and shrimp, and trimethylarsenic species in killifish, were the predominant methylated arsenic species, respectively.     

Synthesis,spectral investigation and development of tetrahedral copper(II) complexes as artificial metallonucleases and antimalarial agents

Seven new copper(II) complexes of type [Cu(A)(L)]?H₂O (A = sparfloxacin, ciprofloxacin, levofloxacin, gatifloxacin, pefloxacin, ofloxacin, norfloxacin; L = 5‐[(3‐chlorophenyl)diazenyl]‐4‐hydroxy‐1,3‐thiazole‐2(3H)‐thione) were synthesized and characterized using elemental and thermogravimetric analyses, and electronic, electron paramagnetic resonance (EPR), Fourier transform infrared and liquid chromatography–mass spectroscopies. Tetrahedral geometry around copper is assigned in all complexes using EPR and electronic spectral analyses. All complexes were investigated for their interaction with herring sperm DNA utilizing absorption titration (K_b = 1.27–3.13 × 10⁵ M^?1) and hydrodynamic volume measurement studies. The studies suggest the classical intercalative mode of DNA binding. The cleavage reaction on pUC19 DNA was monitored by agarose gel electrophoresis. The results indicate that the Cu(II) complexes can more effectively promote the cleavage of plasmid DNA. The superoxide dismutase mimic activity of the complexes was evaluated by nitroblue tetrazolium assay, and the complexes catalysed the dismutation of superoxide at pH = 7.8 with IC₅₀ values in the range 0.597–0.900 μM. The complexes were screened for their in vitro antibacterial activity against five pathogenic bacteria. All the complexes are good cytotoxic agents and show LC₅₀ values ranging from 5.559 to 11.912 µg ml^?1. All newly synthesized Cu(II) complexes were also evaluated for their in vitro antimalarial activity against Plasmodium falciparum strain (IC₅₀ = 0.62–2.0 µg ml^?1). Copyright © 2015 John Wiley & Sons, Ltd.     

Catalyzed Determination of Glucose with a Mimic Enzyme Constructed of Bis‐[‐6‐O‐(‐β‐ethyloic‐butanedioic Acid‐1,4‐ester‐4‐)]β‐Cyclodextrin · Fe3+

Abstract

Catalyzed determination of glucose with mimic glucose oxidase is constructed by the reaction of β‐cyclodextrin, maleic anhydride, and chloroacetic acid with iron trichloride in hydrogen peroxide. The method is simple and convenient, and sensitivity and repeatability are ideal. Beer's law is obeyed in a concentration range of 30–197 µg · ml^?1 glucose with an excellent correlation coefficient (r=0.9994), while the detection limit is 4.10 µg · ml^?1, the RSD is 0.98% (n=8). The recovery of sample is 95.8–103.1%.     

Chlorodesmethyldiazepam Fluorimetric Determination in Pharmaceutical Formulations and in Aqueous Media

Abstract

A direct fluorimetric method for the determination of chlorodesmethyldiazepam was proposed. The fluorescence process was pH dependent. The dynamic range for the method was 1.6–12 µg ml^–1. A linear relationship between the fluorescence intensity and the concentration of chlorodesmethyldiazepam solution was obtained with r ² of 0.9958. The detection limit for the method was found to be 0.224 µg ml^–1. The method has successfully applied to the determination of chlorodesmethyldiazepam in pure, authentic, and aqueous samples.     

Response surface optimisation applied to a headspace-solid phase microextraction-gas chromatography-mass spectrometry method for the analysis of volatile organic compounds in water matrices

A new method for the simultaneous determination of 12 volatile organic compounds (trans-1,2-dichloroethene, 1,1,1-trichloroethane, benzene, 1,2-dichloroethane, trichloroethene, toluene, 1,1,2-trichloroethane, tetrachloroethene, ethylbenzene, m-, p-, o-xylene) in water samples by headspace solid phase microextraction (HS–SPME)–gas chromatography mass spectrometry (GC–MS) was described, using a 100?µm PDMS (polydimethylsiloxane) coated fibre. The response surface methodology was used to optimise the effect of the extraction time and temperature, as well as the influence of the salt addition in the extraction process. Optimal conditions were extraction time and temperature of 30?min and ?20°C, respectively, and NaCl concentration of 4?mol?L^?1. The detection limits were in the range of 1.1?×?10^?3–2.3?µg?L^?1 for the 12 volatile organic compounds (VOCs). Global uncertainties were in the range of 4–68%, when concentrations decrease from 250?µg?L^?1 down to the limits of quantification. The method proved adequate to detect VOCs in six river samples.     

Chemical composition and antibacterial activity of Tussilago farfara (L.) essential oil from Quebec,Canada

Abstract

The chemical composition of Tussilago farfara L. essential oil from the Saguenay-Lac-St-Jean region of Quebec, Canada was analyzed by gas chromatography–flame ionisation detector (GC-FID) and gas chromatography–mass spectrometry (GC-MS), and the antibacterial activity of the oil was tested against Escherichia coli and Staphylococcus aureus. Forty-five (45) compounds were identified from the GC profile. The main components were 1-nonene (40.1%), α-phellandrene (26.0%) and ρ-cymene (6.6%). The essential oil demonstrated antibacterial activity against E. coli (MIC₅₀ = 468 µg·mL^?1; MIC₉₀ = 6869 µg·mL^?1) and S. aureus (MIC₅₀ = 368 µg·mL^?1; MIC₉₀ = 773 µg·mL^?1). Dodecanoic acid was found to be active against both bacteria having a MIC₅₀ and MIC₉₀ of 16.4 µg·mL^?1 and 95 µg·mL^?1, respectively for E. coli and a MIC₅₀ and MIC₉₀ of 9.8 µg·mL^?1 and 27.3 µg·mL^?1, respectively for S. aureus. In addition, 1-decene and (E)-cyclodecene were also found to be active against E. coli.     

Flow-Injection Analysis Based on Extraction and Spectrophotometric Determination of Penicillins with Thiazine Dyes

Abstract

A new method for flow-injection analysis (FIA) for the determination of penicillins based on the extraction and spectrophotometric determination of ion associates with selected thiazine dyes (methylene blue, azure A, and azure B) is proposed. The reaction conditions (c_dye = 2 × 10^?4 mol l^?1, c_KCl = 1 mol l^?1, pH ? 6, λ = 635 nm) were found. The factorial design has been carried out to determine the optimum flow conditions. A wide linear dynamic range of calibration curves (5.1–700 µg ml^?1 for penicillin V with all dyes, R = 0.9985) and good repeatability (e.g., relative standard deviation [RSD] = 4.6–0.6% in this concentration range for the reaction with azure B) were found. The detection limit for penicillin V is 1.5 µg ml^?1, and the determination limit is 5.1 µg ml^?1. The maximum analysis rate is 35 samples per h. The practical samples of pharmaceutics were tested. There are no interferences from the additives in pharmaceutics.     

Levels of polycyclic aromatic hydrocarbons,polychlorinated byphenyls,methylmercury and butyltins in the natural UNESCO reserve of the biosphere of Urdaibai (Bay of Biscay,Spain)

Polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), methylmercury (MeHg⁺) and butyltins (mono-, di- and tri-butyltin, MBT, DBT and TBT) were monitored in oysters (Crassostrea sp.) and sediments collected in different sampling points of the UNESCO reserve of the biosphere of Urdaibai (Bay of Biscay) from March 2006 to June 2007. In the case of oyster samples, concentrations in the 290–1814 µg kg^?1 (PAHs), 70–475 µg kg^?1 (PCBs), 75–644 µg kg^?1 (MeHg⁺) and 200–1300 µg kg^?1 (as a sum of the three butyltins) ranges were obtained. In most samples TBT was the most abundant butyltin, followed by DBT and MBT. It should be highlighted that most samples exceeded the highest range (367 µg kg^?1) found in the last mussel watch programme carried out by the National Oceanic and Atmospheric Administration (NOAA) for butyltins in oyster samples. This could be due to the presence of a shipyard in the estuary. Sediment concentrations ranged as follows: total PAHs (856–3495 µg kg^?1) and total PCBs (58–220 µg kg^?1). Organometallic species were always below the limits of detection (LODs) (0.24 µg kg^?1 for MeHg⁺, 0.6 µg kg^?1 for MBT, 0.48 µg kg^?1 for DBT and 1.1 µg kg^?1 for TBT). In both sediment and oyster PAH sources were mostly combustion. In the case of PCBs, 4-6 chlorine-atom congeners were the most abundant ones. Slight differences in the profile of PAHs as well as PCBs can be detected when the matrices were compared with each other. Finally, in the case of PAHs, sediment and water column played the main role in the accumulation pathway into the organisms in all the sampling stations.     

Determination of phosphate in freshwater samples by flow-injection with lucigenin chemiluminescence

Lucigenin chemiluminescence (CL) in conjunction with flow-injection analysis (FIA) is used for the determination of phosphate in freshwater samples. The procedure is based on the formation of molybdophosphoric heteropoly acid (MoP–HPA) by the reaction of phosphate and ammonium molybdate under acidic conditions. CL emission was observed as a result of oxidation of lucigenin in aqueous sodium hydroxide solution in the presence of MoP–HPA. Calibration was linear up to 500?µg?L^?1 (r ^2?=?0.9998; n?=?8), with a detection limit (S/N?=?3) of 0.95?µg?L^?1. An injection throughput of 120 h^?1, and relative standard deviation (RSD; n?=?4) of 1.3–3.2% were achieved in the concentration range studied. An on-line chelating column was used to remove interfering cations. The method was applied to freshwater samples, and the results (51?±?1.0 – 107?±?2.0?µg?L^?1) did not differ significantly from results obtained using a spectrophotometric method (52.5?±?1.0 – 102?±?2.0?µg?L^?1) at 95% confidence level (t-test).     

Flow‐Injection Spectrophotometric Determination of N‐acetylcysteine in Pharmaceutical Formulations with On‐Line Solid‐Phase Reactor Containing Zn(II) Phosphate Immobilized in a Polyester Resin

Abstract

A flow‐injection spectrophotometric procedure was developed for determining N‐acetylcysteine in pharmaceutical formulations. The sample was dissolved in deionized water and 400 µl of the solution was injected into a carrier stream of 1.0×10^?2 mol l^?1 sodium borate solution. The sample flowed through a column (70 mm length×2.0 mm i.d.) packed with Zn₃(PO₄)₂ immobilized in a polymeric matrix of polyester resin and Zn(II) ions were released from the solid‐phase reactor because of the formation of the Zn(II) (N‐acetylcysteine)₂ complex. The mixture merged with a stream of borate buffer solution (pH 9.0) containing 5.0×10^?4 mol l^?1 Alizarin red S and the Zn(II)Alizarin red complex formed was measured spectrophotometrically at 540 nm. The analytical curve was linear in the N‐acetylcysteine concentration range from 3.0×10^?5 to 1.5×10^?4 mol l^?1 (4.9 to 24.5 µg ml^?1) with a detections limit of 8.0×10^?6 mol l^?1 (1.3 µg ml^?1). The relative standard deviations (RSDs) were smaller than 0.5% (n=10) for solutions containing 5.0×10^?5 mol l^?1 (8.0 µg ml^?1) and 8.0×10^?5 mol l^?1 (13.0 µg ml^?1) of N‐acetylcysteine, and the analytical frequency was 60 determinations per hour. A paired t‐test showed that all results obtained for N‐acetylcysteine in commercial formulations using the proposed flow‐injection procedure and a comparative procedure agreed at the 95% confidence level.     

A New Spectrophotometric Method for the Determination of Arsenic in Environmental and Biological Samples

Abstract

A simple and selective spectrophotometric method has been developed for the determination of trace amounts of arsenic using azure B as a chromogenic reagent. The proposed method is based on the reaction of arsenic(III) with potassium iodate in acid medium to liberate iodine. The liberated iodine bleaches the violet color of azure B and is measured at 644 nm. This decrease in absorbance is directly proportional to the As(III) concentration, and Beer's law is obeyed in the range 0.2–10 µg ml^?1 of As(III). The molar absorptivity, Sandell's sensitivity, detection limit, and quantitation limit of the method were found to be 1.12×10⁴ l mol^?1cm^?1, 6.71×10^?3 µg cm^?2, 0.02 µg ml^?1 and 0.08 µg ml^?1, respectively. The optimum reaction conditions and other analytical parameters were evaluated. The proposed method has been successfully applied for the determination of arsenic in various environmental and biological samples.     

Differential pulse polarographic behaviour of thiazopyr herbicide and application to its determination in fruit juice and soil samples

A differential pulse polarographic method for the determination of the herbicide thiazopyr has been developed. The polarographic study of thiazopyr exhibited two well-defined cathodic peaks within the pH range of 1.0 to 8.0. The variation of pH and polarographic parameters indicated that the optimum conditions under which thiazopyr could be reduced were a pH 7.0 BR buffer solution, a reduction peak potential of ?1270 mV (vs. SCE), scan rate of 5 mV s^?1, pulse amplitude of 50 mV with pulse duration of 50 ms at an ambient temperature of 25 ± 3°C. The main reduction peak was characterised by cyclic voltammetry as being irreversible and diffusion-controlled. A linear relationship between the peak current and the concentration of thiazopyr was obtained in the range of 0.43–38.6 µg mL^?1, with a detection limit of 0.127 µg mL^?1. The proposed method was successfully applied to the determination of thiazopyr in spiked fruit juice and soil samples. The mean recoveries of the 19.8 µg g^?1 and 3.96 µg mL^?1 thiazopyr spiked to soil and orange juice were 20.2 ± 1.0 µg g^?1 and 3.84 ± 0.12 µg mL^?1, at 95% confidence level, respectively. The sufficiently good recoveries and low relative standard deviation (RSD) data confirm the high accuracy and precision of the proposed method. The interferences effects of several commonly used pesticides and inorganic species were also studied. Interfering effects were eliminated either by providing selectivity with pH, or using EDTA as complexing agent.     

Determination of arsenic species in a freshwater crustacean Procambarus clarkii

The arsenic species present in samples of the crayfish Procambarus clarkii caught in the area affected by the toxic mine‐tailing spill at Aznalcóllar (Seville, Southern Spain) were analyzed. The total arsenic contents ranged between 1.2 and 8.5 µg g^?1 dry mass (DM). With regard to the different species of arsenic, the highest concentrations were for inorganic arsenic (0.34–5.4 µg g^?1 DM), whereas arsenobetaine, unlike the situation found in marine fish products, was not the major arsenic species (0.16 ± 0.09 µg g^?1 DM). Smaller concentrations were found of arsenosugars 1a (0.18 ± 0.11 µg g^?1 DM), 1b (0.077 ± 0.049 µg g^?1 DM), 1c (0.080 ± 0.089 µg g^?1 DM), and 1d (0.14 ± 0.13 µg g^?1 DM). The presence of two unknown arsenic species was revealed (U1: 0.058 ± 0.058 µg g^?1 DM; U2: 0.12 ± 0.12 µg g^?1 DM). No significant differences were seen with respect to the total arsenic contents between the sexes. However, significant differences in the total arsenic contents were revealed between the area affected by the spill and the area not affected, the contents being greater in the affected area. Copyright © 2002 John Wiley & Sons, Ltd.     

Label-free immunosensor for monitoring vitellogenin as a biomarker for exogenous oestrogen compounds in amphibian species

A label-free, optical waveguide lightmode spectroscopy based immunosensor was developed for frog (Bombina orientalis) vitellogenin (Vtg) determination in biological samples as a biomarker for exogenous oestrogen compounds. Antibody against Vtg was produced in rabbits immunised with purified lipovitellin (Lpv), a precursor of Vtg, from the homogenised ovary of oriental fire-bellied toads (B. orientalis). The purified protein and Lpv/Vtg-specific serum were applied in both competitive and direct immunoassay formats using optical waveguide lightmode spectroscopy immunosensor. When measuring Vtg in direct manner, the Lpv antibody (1.76 µg mL^?1) was immobilised on the sensor surface, and the linear measuring range for Vtg was 0.1–10 µg mL^?1. During the competitive measurement, 100 ng mL^?1 Lpv was applied for the immobilisation. The linear measuring range for Vtg was 0.5–50 ng mL^?1. We studied the relative substrate specificity of the antibody, and it was concluded that the method is suitable for the sensitive and selective determination of Vtg levels in toads. Heart, liver and gonad samples from male animals were spiked with Vtg and were analysed using the newly developed method, and female toads and spawn samples were tested and compared to the calibration curve obtained by the spiked samples.