Analysis of endocrine disruptor compounds in marine sediments by in cell clean up-pressurized liquid extraction-liquid chromatography tandem mass spectrometry determination

A less time-, solvent- and sorbent-consuming analytical methodology for the determination of bisphenol A and alkylphenols (4-tert-octylphenol, 4-octylphenol, 4-n-nonylphenol, nonylphenol) in marine sediment was developed and validated. The method was based on selective pressurized liquid extraction (SPLE) with a simultaneous in cell clean up combined with liquid chromatography-electrospray ionization tandem mass spectrometry in negative mode (LC-ESI-MS/MS). The SPLE extraction conditions were optimized by a Plackett–Burman design followed by a central composite design. Quantitation was performed by standard addition curves in order to correct matrix effects. The analytical features of the method were satisfactory: relative recoveries varied between 94 and 100% and repeatability and intermediate precision were <6% for all compounds. Uncertainty assessment of measurement was estimated on the basis of an in-house validation according to EURACHEM/CITAC guide. Quantitation limits of the method (MQL) ranged between 0.17 (4-n-nonylphenol) and 4.01 ng g⁻¹ dry weight (nonylphenol). Sensitivity, selectivity, automaticity and fastness are the main advantages of this green methodology. As an application, marine sediment samples from Galicia coast (NW of Spain) were analysed. Nonylphenol and 4-tert-octylphenol were measured in all samples at concentrations between 20.1 and 1409 ng g⁻¹ dry weight, respectively. Sediment toxicity was estimated and no risk to aquatic biota was found.     

Application of SPME to the determination of alkylphenols and bisphenol A in cyanobacteria culture media

In order to survey the influence of estrogenic compounds on cyanobacteria, solid-phase microextraction (SPME) with a carbowax-divinylbenzene fibre was used for the determination of tert-octylphenol (tert-OP), n-nonylphenol (n-NP), technical nonylphenol (tech-NP) and bisphenol A (BPA) in cyanobacteria culture media by gas chromatography with flame ionization detection. Determinations were carried out without derivatization in deionized water and filtered culture media. A comparison between f2 and Fraquil culture media was performed, which showed that only f2 allowed quantitative recoveries. Headspace SPME with salting out, requiring only 10 mL of sample, was suitable for tert-OP, n-NP, and tech-NP determination with limits of detection (LOD) of <0.05 μg L⁻¹. For BPA, direct immersion SPME could provide a LOD of 1 μg L⁻¹. Automated sampling allowed reproducible extraction. No exudate substances overlapped with the studied compounds during the chromatographic separation and no matrix effects were observed. Ecotoxicity tests can be performed by single spiking of tert-OP and tech-NP and multiple spiking of n-NP due to its lower stability.     

Polar herbicides,pharmaceutical products,perfluorooctanesulfonate (PFOS), perfluorooctanoate (PFOA), and nonylphenol and its carboxylates and ethoxylates in surface and tap waters around Lake Maggiore in Northern Italy

A survey of contamination of surface and drinking waters around Lake Maggiore in Northern Italy with polar anthropogenic environmental pollutants has been conducted. The target analytes were polar herbicides, pharmaceuticals (including antibiotics), steroid estrogens, perfluorooctanesulfonate (PFOS), perfluoroalkyl carboxylates (including perfluorooctanoate PFOA), nonylphenol and its carboxylates and ethoxylates (NPEO surfactants), and triclosan, a bactericide used in personal-care products. Analysis of water samples was performed by solid-phase extraction (SPE) then liquid chromatography–triple-quadrupole (tandem) mass spectrometry (LC–MS–MS). By extraction of 1-L water samples and concentration of the extract to 100 μL, method detection limits (MDLs) as low as 0.05–0.1 ng L⁻¹ were achieved for most compounds. Lake-water samples from seven different locations in the Southern part of Lake Maggiore and eleven samples from different tributary rivers and creeks were investigated. Rain water was also analyzed to investigate atmospheric input of the contaminants. Compounds regularly detected at very low concentrations in the lake water included: caffeine (max. concentration 124 ng L⁻¹), the herbicides terbutylazine (7 ng L⁻¹), atrazine (5 ng L⁻¹), simazine (16 ng L⁻¹), diuron (11 ng L⁻¹), and atrazine-desethyl (11 ng L⁻¹), the pharmaceuticals carbamazepine (9 ng L⁻¹), sulfamethoxazole (10 ng L⁻¹), gemfibrozil (1.7 ng L⁻¹), and benzafibrate (1.2 ng L⁻¹), the surfactant metabolite nonylphenol (15 ng L⁻¹), its carboxylates (NPE₁C 120 ng L⁻¹, NPE₂C 7 ng L⁻¹, NPE₃C 15 ng L⁻¹) and ethoxylates (NPE _n Os, n = 3-17; 300 ng L⁻¹), perfluorinated surfactants (PFOS 9 ng L⁻¹, PFOA 3 ng L⁻¹), and estrone (0.4 ng L⁻¹). Levels of these compounds in drinking water produced from Lake Maggiore were almost identical with those found in the lake itself, revealing the poor performance of sand filtration and chlorination applied by the local waterworks.     

Development of a highly precise ID-ICP-SFMS method for analysis of low concentrations of lead in rice flour reference materials

Microwave digestion and isotope dilution inductively coupled plasma mass spectrometry (ID-ICP-SFMS) has been applied to the determination of Pb in rice flour. In order to achieve highly precise determination of low concentrations of Pb, the digestion blank for Pb was reduced to 0.21 ng g⁻¹ after optimization of the digestion conditions, in which 20 mL analysis solution was obtained after digestion of 0.5 g rice flour. The observed value of Pb in a non-fat milk powder certified reference material (CRM), NIST SRM 1549, was 16.8 ± 0.8 ng g⁻¹ (mean ± expanded uncertainty, k = 2; n = 5), which agreed with the certified value of 19 ± 3 ng g⁻¹ and indicated the effectiveness of the method. Analytical results for Pb in three brown rice flour CRMs, NIST SRM 1568a, NIES CRM 10-a, and NIES CRM 10-b, were 7.32 ± 0.24 ng g⁻¹ (n = 5), 1010 ± 10 ng g⁻¹ (n = 5), and 1250 ± 20 ng g⁻¹ (n = 5), respectively. The concentration of Pb in a candidate white rice flour reference material (RM) sample prepared by the National Metrology Institute of Japan (NMIJ) was observed to be 4.36 ± 0.28 ng g⁻¹ (n = 10 bottles). Figure Digestion blank of Pb was carefully reduced to approximately 0.2 ng g^-1 which permitted the highly precise determination of Pb at low ng g^-1 level in foodstuff samples by ID-SFMS     

Occurrence of endocrine-disrupting compounds in reclaimed water from Tianjin,China

Continuous disposal of endocrine-disrupting compounds (EDCs) into the environment can lead to serious human health problems and can affect plants and aquatic organisms. The determination of EDCs in water has become an increasingly important activity due to our increased knowledge about their toxicities, even at low concentration. The EDCs in water samples from the reclaimed water plant of Tianjin, northern China, were identified by gas chromatography (GC)–mass spectrometry (MS). Important and contrasting EDCs including estrone (E1), 17β-estradiol (E2), 17α-ethynylestradiol (EE2), 4-tert-octylphenol (OP), 4-nonylphenol (NP), bisphenol A (BPA), di-n-butyl phthalate (DnBP), diisobutyl phthalate (DIBP), and di(2-ethylhexyl)phthalate (DEHP) were selected as the target compounds. Concentrations of steroid hormones, alkylphenolic compounds and phthalates ranged from below the limit of detection (LOD) to 8.1 ng L⁻¹, from <LOD to 14.2 ng L⁻¹, and from 1.00 μg L⁻¹ to 23.8 μg L⁻¹, respectively. The average removal efficiencies for target EDCs varied from 30% to 82%. These results indicate that environmental endocrine disrupting compounds are not completely removed during reclaimed water treatment and may be carried over into the general aquatic environment.     

Determination of the food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in human hair by solid-phase extraction and gas chromatography-mass spectrometry

Summary A method for the determination of the food mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in human hair has been developed and validated. Hair samples (200 mg) were dissolved in NaOH (1 M) and PhIP was isolated by successive solid-phase extraction on a polystyrene-divinylbenzene column and on a silica-based mixed-mode column with C₈ and-SO₃ ⁻ functional groups. Quantification was performed by gas chromatography-electron-impact ionization high-resolution mass spectrometry in selected-ion-monitoring mode. The method was validated for determination of PhIP in the concentration range 0.5–25 ng g⁻¹ hair with [²H₃]PhIP as internal standard. The limit of quantification was 0.26 ng g⁻¹ hair. Within-day and between-day precision were in the ranges 1–27% and 2–15% relative standard deviation, respectively. The hair sample used for method validation was found to contain 0.26 ng PhIP g⁻¹ hair.     

Determination of 5-n-Butyl-4-{4-[2-(1H-tetrazole-5-yl)-1H-pyrrol-1-yl]phenylmethyl}-2,4-dihydro-2-(2,6-dichloridephenyl)-3H-1,2,4-triazol-3-one, a New Angiotensin Type 1 Receptor Antagonist in Rat Plasma by LC-ESI-MS: Application to Pharmacokinetic Studies

A simple and sensitive reversed-phase LC-ESI-MS method to identify and quantitate 5-n-butyl-4-{4-[2-(1H-tetrazole-5-yl)-1H-pyrrol-1-yl]phenylmethyl}-2,4-dihydro-2-(2,6-dichloridephenyl)-3H-1,2,4-triazol-3-one (1b), a new Angiotensin II type 1 receptor antagonist in rat plasma has been developed and validated. Sample preparation used a simple liquid–liquid extraction with ethyl acetate. Separation was achieved by gradient elution on a C₁₈ column. The mobile phase consisted of acetonitrile and water (0.05% triethylamine and 0.05% acetic acid) at a flow rate of 0.2 mL min⁻¹. The detection utilized selected ion monitoring (SIM) in the negative mode at m/z 507.1 and m/z 407.2 for the deprotonated molecular ions of 1b and the internal standard irbesartan, respectively. The lower limit of quantification was reproducible at 5 ng mL⁻¹ with 100 μL of plasma and the good linear was observed in the 5–500 ng mL⁻¹ range. This concentration range corresponded well with the plasma concentrations of 1b in pharmacokinetic studies. Recoveries of 1b in rat plasma were 76.1, 74.6 and 79.0% at 5, 50 and 500 ng mL⁻¹. The RSD of intra-assay and inter-assay variations were all less than 5%. This validated LC-ESI-MS assay is an economic, quick, precise and reliable method for the analysis of 1b in pharmacokinetic studies.     

Uncertainty of determination of palladium in road dust sample by inductively coupled plasma mass spectrometry

The determination of palladium in a road dust sample taken close to the highway (w _Pd=450 ng g⁻¹) was carried out by the ICP-MS method after sample decomposition by aqua-regia. Analyses were evaluated by two methods: external calibration accompanied with mathematical correction of spectral interferences (EC) and isotope dilution measurement after separation of Pd by extraction to dibutyl sulfide solution (ID). In both cases, the uncertainties and accuracy of results were investigated. Although in the case of ideally homogeneous sample the repeatability of EC results (11 ng g⁻¹ Pd) was somewhat lower than those of ID results (16 ng g⁻¹ Pd), the uncertainties of results of both techniques were almost the same and they reached the level of 19 ng g⁻¹ Pd. The main uncertainty source of the EC method is represented by the correction of spectral interferences. In case of real non-homogeneous sample, the main uncertainty component represents the soil sampling. The uncertainty of results (approx. 75 ng g⁻¹ Pd) only slightly exceeded the repeatability (approx. 70 ng g⁻¹ Pd). The accuracy of results was proven by analyses of CRM TDB–1 Diabas Rock (in case of ID) and by the standard addition method (in case of EC).     

Determination of bisphenol A, 4-n-nonylphenol, and 4-tert-octylphenol by temperature-controlled ionic liquid dispersive liquid-phase microextraction combined with high performance liquid chromatography-fluorescence detector

Present study described a simple, sensitive, and viable method for the determination of bisphenol A, 4-n-nonylphenol and 4-tert-octylphenol in water samples using temperature-controlled ionic liquid dispersive liquid-phase microextraction coupled to high performance liquid chromatography-fluorescence detector. In this experiment, 1-octyl-3-methylimidazolium hexafluorophosphate ([C₈MIM][PF₆]) was used as the extraction solvent, and bisphenol A, 4-n-nonylphenol and 4-tert-octylphenol were selected as the model analytes. Parameters affecting the extraction efficiency such as the volume of [C₈MIM][PF₆], dissolving temperature, extraction time, sample pH, centrifuging time and salting-out effect have been investigated in detail. Under the optimized conditions, good linear relationship was found in the concentration range of 1.0-100 μg L⁻¹ for BPA, 1.5-150 μg L⁻¹ for 4-NP, and 3-300 μg L⁻¹ for 4-OP, respectively. Limits of detection (LOD, S/N = 3) were in the range of 0.23-0.48 μg L⁻¹. Intra day and inter day precisions (RSDs, n = 6) were in the range of 4.6-5.5% and 8.5-13.3%, respectively. This method has been also successfully applied to analyze the real water samples at two different spiked concentrations and excellent results were obtained.     

Determination of four phenolic endocrine disrupting chemicals in Dianchi Lake,China

An efficient derivatization method using phenyltrimethylammonium (PTA-OH) has been developed to determine simultaneously four phenolic endocrine disrupting chemicals, 4-n-nonylphenol (4-n-NP), 4-tert-octylphenol (4-t-OP), bisphenol A (BPA) and 4-cumylphenol (4-CP) in surface water of Dianchi Lake (China) by solid-phase extraction (SPE) and gas chromatography-mass spectrometry (GC-MS). Compared with silylation of target phenols using N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA)?+?1% trimethylchlorosilane (TMCS), methylation by PTA-OH displayed a higher response and stability based on the investigations of various derivatization conditions, including derivatization solvent, amount of derivatization reagent, reaction temperature and time. Experiments were carried out to examine the performance of the proposed method based on the correlation coefficient, the method quantification limit (MQL), mean recovery rate and relative standard deviation (RSD). Under optimum derivatization conditions, MQLs of the methylated target compounds were all below 1?ng?L^?1. Results revealed that the proposed method exhibited a satisfactory precision and reproducibility for the separation and determination of target phenols. The proposed method had been applied to determine four phenols in surface water of Dianchi Lake located in southwest of China. The concentrations of 4-n-NP, 4-t-OP, BPA and 4-CP were determined to be 13.6-141.6?ng?L^?1, N.D.-56.5?ng?L^?1, N.D.- 4713.6?ng?L^?1 and 23.3-48.5?ng?L^?1, respectively.     

Speciation analysis of inorganic antimony in soil using HPLC-ID-ICP-MS

Speciation analysis of Sb(III) and Sb(V) in a soil sample was performed through extraction and on-line isotope dilution concentration determination after a chromatographic separation. The total Sb concentration found in a through traffic contaminated soil sample was (4.17 μg g⁻¹, 0.3 μg g⁻¹ SD, n=6). It was determined using ICP-MS after soil digestion using the sodium peroxide sintering method. The optimized extraction procedure for speciation analysis was carried out using 100 mmol L⁻¹ citric acid at pH 2.08 by applying an ultrasonic bath for 45 min at room temperature. The effects of citric acid concentration (0–500 mmol L⁻¹), pH (1–6), and temperature (30–60°C) on inorganic antimony species distribution in the examined sample were studied and optimized. The separation of Sb(III) and Sb(V) was achieved using an anion exchange column (PRP-X100) and 10 mmol L⁻¹ EDTA and 1 mmol L⁻¹ phthalic acid at pH 4.5 as a mobile phase. The eluent from the HPLC was mixed with an enriched (94.2%) ¹²³Sb spike solution that was pumped by a peristaltic pump with a constant flow rate (0.5 mL min⁻¹) in a three-way valve. The blend passed directly to the Conikal nebulizer of the ICP-MS. By using the above extraction procedure and methodology, 43.2% Sb(V) (2.9% RSD, n=3) and 6.0% Sb(III) (1.3% RSD, n=3) of total Sb found in the sample could be detected. The detection limits achieved by the proposed method were 20 ng L⁻¹ and 65 ng L⁻¹ for Sb(V) and Sb(III), respectively. The precision, evaluated by using RSD with 100 ng L⁻¹ calibration solutions, was 2.7% and 3.2% (n=6) for Sb(V) and Sb(III), respectively, in aqueous solutions.     

Determination of total Sb,Se, Te,and Bi and evaluation of their inorganic species in garlic by hydride-generation–atomic-fluorescence spectrometry

A sensitive and simple analytical method has been developed for determination of Sb(III), Sb(V), Se(IV), Se(VI), Te(IV), Te(VI), and Bi(III) in garlic samples by using hydride-generation–atomic-fluorescence spectrometry (HG–AFS). The method is based on a single extraction of the inorganic species by sonication at room temperature with 1 mol L⁻¹ H₂SO₄ and washing of the solid phase with 0.1% (w/v) EDTA, followed by measurement of the corresponding hydrides generated under two different experimental conditions directly and after a pre-reduction step. The limit of detection of the method was 0.7 ng g⁻¹ for Sb(III), 1.0 ng g⁻¹ for Sb(V), 1.3 ng g⁻¹ for Se(IV), 1.0 ng g⁻¹ for Se(VI), 1.1 ng g⁻¹ for Te(IV), 0.5 ng g⁻¹ for Te(VI), and 0.9 ng g⁻¹ for Bi(III), in all cases expressed in terms of sample dry weight.     

Multi-Residue Analysis of Avermectins in Bovine Liver and Muscle by Liquid Chromatography–Fluorescence Detector

A simple multi-residue analysis method for the quantitative determination of eprinomectin, abamectin, doramectin and ivermectin in bovine tissues was developed. The tissue sample was extracted with acetonitrile, followed by clean-up on a C₁₈ solid phase extraction cartridge. The eluate was derivatised before being analyzed by HPLC coupled to a fluorescence detector. The method was validated using bovine liver and muscle fortified with the drugs at 0, 5, 10 and 50 ng g⁻¹. The mean recoveries of the four drugs were 70.31–87.11% in liver and 79.57–93.65% in muscle, with relative standard deviations below 17.84% in liver and 14.68% in muscle. The limits of detection were between 0.5 and 1.0 ng g⁻¹ and the limits of quantification were 1–2 ng g⁻¹ in bovine tissues for the four drugs.     

Analysis of sulfamerazine in pond water and several fishes by high-performance liquid chromatography using molecularly imprinted solid-phase extraction

The synthesis and evaluation of a molecularly imprinted polymer (MIP) used as a selective solid-phase extraction sorbent and coupled to high-performance liquid chromatography (HPLC) for the efficient determination of sulfamerazine (SMR) in pond water and three fishes are reported. The polymer was prepared using SMR as the template molecule, methacrylic acid as the functional monomer and ethylene glycol dimethacrylate as the crosslinking monomer in the presence of tetrahydrofuran as the solvent. The SMR-imprinted polymers and nonimprinted polymers were characterized by FT-IR and static adsorption experiments. The prepared SMR-imprinted material showed a high adsorption capacity, significant selectivity and good site accessibility. The maximum static adsorption capacities of the SMR-imprinted and nonimprinted materials for SMR were 108.8 and 79.6 mg g⁻¹, respectively. The relative selectivity factor of this SMR-imprinted material was 1.6. Several parameters influencing the solid-phase extraction process were optimized. Finally, the SMR-imprinted polymers were used as the sorbent in solid-phase extraction to determine SMR in pond water and three fishes with satisfactory recovery. The average recoveries of the MIP-SPE method were 94.0% in ultrapure water and 95.8% in pond water. Relative standard deviations ranging from 0.3% to 5.2% in MIP were acquired. The results for the SMR concentrations in crucian, carp and wuchang fish were 66.0, 127.1 and 51.5 ng g⁻¹, respectively. The RSDs (n = 5) were 3.51%, 0.53% and 5.08%, respectively. The limit of detection (LOD) for SMR was 1 ng g⁻¹ and the limit of quantitation (LOQ) was 3.5 ng g⁻¹.     

An analytical method for the simultaneous trace determination of acidic pharmaceuticals and phenolic endocrine disrupting chemicals in wastewater and sewage sludge by gas chromatography-mass spectrometry

This article presents an analytical method based on solid-phase extraction (SPE) and gas chromatography coupled with mass spectrometry for the simultaneous determination of the most frequently used acidic pharmaceutical residues, ibuprofen, diclofenac, naproxen and ketoprofen (KFN), and phenolic endocrine disruptors, bisphenol (BPA), triclosan (TCS), nonylphenol, nonylphenol monoethoxylate and nonylphenol diethoxylate, in wastewater and sewage sludge samples. In the first phase of the study, each compound has been characterized individually and afterwards in mixture as a trimethylsilyl derivative in order to identify the characteristic ions (m/z ratio) constituting the mass spectrum and to choose the ions for quantification and confirmation. Subsequently, derivatization was evaluated by testing different variables such as the volume of the derivatization solvent bis(trimethylsilyl)trifluoroacetamide and the effect of each catalyst, pyridine and 1% trimethyl chlorosilane, in the derivatized solution. For the analysis of wastewater samples, two commercial SPE cartridges, C18 and Oasis HLB, were compared for their extraction efficiency of the target compounds. The key parameter of extraction procedure included the effect of pH (2.5, 5.3 and 7) of the loading solution. For solid samples, parameters such as the extracted biomass, the volume of the extraction organic solvent and the effect of matrix interferences in chromatographic analysis were evaluated. By using C18 cartridges as purification procedure and ultrasound sonication, satisfactory mean relative recoveries with BPA-d16 and meclofenamic acid as surrogates were obtained ranging from 91% to 117% for wastewater and 84% to 107% for sewage sludge samples. Nine-point calibration of the standard mixture was performed by linear regression analysis with a correlation coefficient >0.99 for all the tested compounds. Limits of detection for the developed methods were established between 0.3 (KFN) and 14.8 (BPA) ng L⁻¹, and 15.0 (TCS) and 32.9 (BPA) ng g⁻¹ for wastewater and sewage sludge, respectively. Application to real samples of the wastewater treatment plant in Athens, the capital of Greece, demonstrated the presence of all tested compounds in most of the samples.     

A confirmatory method for the determination of phenolic endocrine disruptors in honey using restricted-access material–liquid chromatography–tandem mass spectrometry

The present work describes the development and validation of an analytical method based on liquid chromatography (LC), coupled with tandem mass spectrometry (MS/MS) that allows the determination and confirmation of several endocrine-disrupting chemicals (EDCs) in honey. The EDCs studied were nine phenols of different nature: chlorophenols (2,4-dichlorophenol, 2,4,5-trichlorophenol, and pentachlorophenol), alkylphenols (4-tert-butylphenol, 4-tert-octylphenol, and 4-n-octylphenol) bisphenols (bisphenol-A and bisphenol-F), and 4-tert-butylbenzoic acid. The method incorporates a restricted-access material (RAM), coupled on-line to the LC-MS/MS system, which allows direct injection of the matrix into the RAM-LC-MS/MS system. The optimized method developed, RAM-LC-MS/MS, was applied to fortified honey samples, affording detection limits in the 0.6–7.2 ng g⁻¹ range, calculated for a signal-to-noise ratio of 3. In addition, the method was validated as a quantitative confirmatory method according to European Union Decision 2002/657/EC. The validation criteria evaluated were linearity, repeatability, reproducibility, recovery, decision limits, detection capabilities, specificity, and ruggedness. Repeatability and within-laboratory reproducibility were evaluated at two concentration levels, being ±11% or below at 20 ng g⁻¹. The decision limits (CC_α) and detection capabilities (CC_β) were in the 1.7–12.6 and 2.8–21.6 ng g⁻¹ range, respectively.     

Determination of organotin compounds in biological samples using accelerated solvent extraction,sodium tetraethylborate ethylation,and multicapillary gas chromatography–flame photometric detection

A method has been developed for species-selective analysis of organotin compounds in solid, biological samples. The procedure is based on accelerated solvent extraction (ASE) of analytes and includes extraction of the tin species with a methanol–water (90% methanol) solution of acetic acid/sodium acetate containing tropolone (0.03% w/v), their ethylation with NaBEt₄, and separation and detection by GC–FPD. The analytical procedure was optimized with an unspiked sample of harbor porpoise (Phocoena phocoena) liver. Effects of ASE operational variables (extraction temperature and pressure, solvent composition, number of static extraction steps) are discussed. Method detection limits (MDL) were in the range 6–10 ng(Sn) g^–1 dry weight and 7–17 ng(Sn) g^–1 dry weight for butyl- and phenyltin compounds, respectively. Recoveries were comparable with or better than those obtained by use of other procedures reported in the literature. The analytical procedure was validated by analysis of NIES No. 11 (fish tissue) certified reference material.     

Simultaneous determination of enrofloxacin and its primary metabolite, ciprofloxacin, in chicken tissues

Summary An HPLC method with fluorescence detection is presented for the analysis of enrofloxcin (ENR) and ciprofloxacin (CIP) in chicken tissue using sarafloxacin (SAR) as internal standard. Tissue sample preparations were carried out by adding a phosphate buffer (pH 7.4, 0.1 M), followed by extraction with trichloromethane. Fluoroquinolones were separated on a reversed-phase column with a mobile phase of aqueous phosphate buffer-acetonitrile (80:20). The concentrations of CIP, ENR and SAR eluted off the column, with retention times of 2.28, 3.30 and 4.40, respectively, were monitored by fluorescence detection atλ _ex 338 andλ _em 425 nm. The detection limit was 32 ng g⁻¹ for CIP and 10 ng g⁻¹ for ENR. The standard curves were linearly related to concentration in the range of 1 to 2000 ng g⁻¹. Recovery was determinated as 91.3% and 78.3% for ENR and CIP, respectively. The measurement of the tissue levels of ENR and CIP in the chicken after oral administration confirmed the utility of the proposed analytical methodology.     

A simplified method for simultaneous quantitation of alkylphenols and alkylphenol ethoxylates in meat and fish using high-performance liquid chromatography with fluorescence detection

Nonylphenol (NP), octylphenol (OP), nonylphenol monoethoxylate (NP₁EO) and nonylphenol diethoxylate (NP₂EO) are products of the biodegradation of alkylphenol polyethoxylates (AP _n EO) which are used worldwide as detergents and surfactants. NP and OP are categorized as definitely endocrine disruptors. 2,4-Tert-butylphenol (BP) is extensively used for anti-oxidant of rubber and plastics. This work proposed a simple and stable method for simultaneously determining the concentration of NP, OP, BP, n-NP₁EO and n-NP₂EO in meat and fish, without requiring the complex pretreatments of current methods. This study used liquid extraction with acetonitrile and hexane and solid extraction using Florisil, in that order to pretreat samples. The sample solutions were analyzed to identify NP, OP, BP, n-NP₁EO and n-NP₂EO by HPLC with fluorescence detection. The mean recoveries were 85.3?±?3.32% for OP, 87.5?±?6.01% for BP, 90.9?±?4.72% for NP, 86.4?±?4.81% for n-NP₂EO and 90.9?±?4.84% for n-NP₁EO. The average coefficients of variation were about 6%. The method's detection limits were 5.4?ng?g^?1 for OP, 5.2?ng?g^?1 for BP, 8.9?ng?g^?1 for NP, 8.7?ng?g^?1 for n-NP₂EO and 8.1?ng?g^?1 for n-NP₁EO. This work analyzed 5 kinds of usual foodstuffs of meat and fish that are frequently consumed by residents of Taiwan. All of these samples contained NP, but not detectable levels n-NP₁EO. Only salmon was contaminated with n-NP₂EO. The NP level was highest in cod (198.41?±?129.34?ng?g^?1, wet weight). The fried chicken had the highest BP level (48.0?±?41.3?ng?g^?1, wet weight), and the uncooked chicken had the highest OP level (66.6?±?53.0?ng?g^?1, wet weight).     

A highly sensitive and specific polyclonal antibody-based enzyme-linked immunosorbent assay for detection of antibiotic olaquindox in animal feed samples

The use of olaquindox (OLA) as an additive in animal feedstuffs has been prohibited in the European Union and many other countries. In this study, a highly sensitive and specific indirect competitive enzyme-linked immunosorbent assay (ELISA) for determination of OLA in animal feed samples was developed. OLA was activated by N′N-carbonyldiimidazole and coupled with bovine serum albumin (BSA) and ovalbumin (OVA). It was found that the sensitivity and specificity of the two antisera were very similar, with the IC₅₀ values of 16 ng mL⁻¹ and 19 ng mL⁻¹, respectively. Cross-reactivity was less than 35% for four structurally related compounds and no recognition of five other antibiotics was observed. The better antiserum I was selected for further experiments, for example testing stability, solvent effect, accuracy, and precision. The IC₅₀ value for eight standard curves was in the range 12–18 ng mL⁻¹ and the LOD at a signal-to-noise ratio of 3 (S/N = 3) was 0.31 ± 0.11 ng mL⁻¹. The ELISA tolerated 5% methanol without significant influence on IC₅₀ value. The recoveries of spiked OLA in five different animal feed types including auxin, pig complex feed, fish complex feed, broiler concentrated feed, and pig premix feed were in the range 88.3–119.0% and the intra-assay relative standard deviation (RSD) was within 4.7–33.5% (n = 3). The ELISA for unspiked feed samples was confirmed by high-performance liquid chromatography (HPLC), with a high correlation coefficient of 0.9862 (n = 5). The proposed ELISA could be a feasible quantitative/screening method for OLA analysis in feed samples with the properties of high sensitivity, specificity, simplicity of sample pretreatment, high sample throughput, and low expense. Figure Polyclonal antibody based ELISA for detection of olaquindox