Ion pair-based liquid-phase microextraction combined with cuvetteless UV-vis micro-spectrophotometry as a miniaturized assay for monitoring ammonia in waters

A miniaturized method based on liquid-phase microextraction (LPME) in combination with microvolume UV-vis spectrophotometry for monitoring ammonia in waters is proposed. The methodology is based on the extraction of the ion pair formed between the blue indophenol obtained according to the Berthelot reaction and a quaternary ammonium salt into a microvolume of organic solvent. Experimental parameters affecting the LPME performance such as type and concentration of the quaternary ammonium ion salt required to form the ion pair, type and volume of extractant solvent, effect of disperser solvent, ionic strength and extraction time, were optimized. A detection limit of 5.0 μg L⁻¹ ammonia and an enrichment factor of 30 can be attained after a microextraction time of 4 min. The repeatability, expressed as relative standard deviation, was 7.6% (n = 7). The proposed method can be successfully applied to the determination of trace amounts of ammonia in several environmental water samples.     

Dispersive liquid–liquid microextraction followed by gas chromatography–mass spectrometry for the rapid and sensitive determination of UV filters in environmental water samples

The performance of the dispersive liquid–liquid microextraction (DLLME) technique for the determination of eight UV filters and a structurally related personal care species, benzyl salicylate (BzS), in environmental water samples is evaluated. After extraction, analytes were determined by gas chromatography combined with mass spectrometry detection (GC-MS). Parameters potentially affecting the performance of the sample preparation method (sample pH, ionic strength, type and volume of dispersant and extractant solvents) were systematically investigated using both multi- and univariant optimization strategies. Under final working conditions, analytes were extracted from 10 mL water samples by addition of 1 mL of acetone (dispersant) containing 60 μL of chlorobenzene (extractant), without modifying either the pH or the ionic strength of the sample. Limits of quantification (LOQs) between 2 and 14 ng L⁻¹, inter-day variability (evaluated with relative standard deviations, RSDs) from 9% to 14% and good linearity up to concentrations of 10,000 ng L⁻¹ were obtained. Moreover, the efficiency of the extraction was scarcely affected by the type of water sample. With the only exception of 2-ethylhexyl-p-dimethylaminobenzoate (EHPABA), compounds were found in environmental water samples at concentrations between 6 ± 1 ng L⁻¹ and 26 ± 2 ng mL⁻¹.     

Comparative study of screening methodologies for ochratoxin A detection in winery by-products

The worldwide contamination of winery by-products by mycotoxins may present a serious hazard to human and animal health. Mycotoxins are secondary metabolites of fungi with possible adverse effects on humans, animals, and crops that result in illnesses and economic losses. Mycotoxins are under continuous survey in Europe, but the regulatory aspects still need to be set up for winery by-products, which may be used in animal feed. The aim of this study was to implement a simple but reliable analytical methodology for ochratoxin A (OTA) quantification in grape pomaces in order to perform a survey of samples from the Douro Demarcated Region, Portugal. The method involved a unique preparation step, solvent extraction, followed by high-performance liquid chromatography (HPLC) with fluorescence (FL) detection. A comparative study was performed with two extraction solvents (ethyl acetate and methanol) as well as using extraction on an immunoaffinity column. The linearity range for OTA analysis was 0.05–23.5 μg L⁻¹ with a detection limit of 0.05 μg L⁻¹ and a precision (expressed by the coefficient of variation under repeatability conditions) of 0.4–14.7%. The percentage of recovery was on average 23.5 ± 3.6% (extraction with ethyl acetate) or 70.1 ± 2.5% (extraction with 70% methanol). Accounting for the recovery factor and the chromatographic detection limit, as well as the preconcentration factor, the limit of detection in grape pomaces is 0.04 μg kg⁻¹ (ethyl acetate extraction) and 0.33 μg kg⁻¹ (methanol extraction). Samples from 12 out of 13 sites in the Douro Demarcated Region showed OTA presence with concentrations not exceeding 0.4 μg kg⁻¹. Both developed methods for evaluation of OTA in grape pomace are simple but efficient. Figure Extraction of ochratoxin A (OTA) from grape pomaces allows simple but efficient quantification of OTA in winery by-products by HPLC-FL     

Tetracycline residues in royal jelly and honey by liquid chromatography tandem mass spectrometry: validation study according to Commission Decision 2002/657/EC

A specific, sensitive and robust liquid chromatography tandem mass spectrometry method for determining oxytetracycline, tetracycline, chlortetracycline and doxycycline in royal jelly and honey samples is presented. Extraction of drug residues was performed by ammonium acetate buffer as extractant followed by a clean-up with metal chelate affinity chromatography and solid-phase extraction. Tetracycline analysis was performed using liquid chromatography–electrospray ionisation–tandem mass spectrometry. The presented method is the first validated for royal jelly and in accordance with the requirements set by Commission Decision 2002/657/EC. Recoveries of the methods, calculated spiking the samples at 5.0, 10.0, 20.0 and 30.0 μg kg⁻¹, were 79% to 90% for honey and 77% to 90% for royal jelly. The intra-day precision (RSD) ranged between 8.1% and 15.0% for honey and from 9.1% to 16.3% for royal jelly, while inter-day precision values were from 10.2% to 17.6% and from 10.6% to 18.4% respectively for honey and royal jelly. Linearity for the four analytes was calculated from 5.0 to 50.0 μg kg⁻¹. The decision limits (CCα) ranged from 6.2 to 6.4 μg kg⁻¹ and from 6.1 to 6.5 μg kg⁻¹ for honey and royal jelly, respectively. Detection capabilities values (CCβ) ranged between 7.2 and 7.7 μg kg⁻¹ and from 7.3 to 7.9 μg kg⁻¹ respectively for honey and royal jelly. The developed method is currently in use for confirmation of the official control analysis of honey and royal jelly samples.     

Ultrasound-enhanced surfactant-assisted dispersive liquid–liquid microextraction and high-performance liquid chromatography for determination of ketoconazole and econazole nitrate in human blood

A simple and efficient method, based on ultrasound-enhanced surfactant-assisted dispersive liquid–liquid microextraction (UESA-DLLME) followed by high-performance liquid chromatography (HPLC) has been developed for extraction and determination of ketoconazole and econazole nitrate in human blood samples. In this method, a common cationic surfactant, cetyltrimethylammonium bromide (CTAB), was used as dispersant. Chloroform (40 μL) as extraction solvent was added rapidly to 5 mL blood containing 0.068 mg mL⁻¹ CTAB. The mixture was then sonicated for 2 min to disperse the organic chloroform phase. After the extraction procedure, the mixture was centrifuged to sediment the organic chloroform phase, which was collected for HPLC analysis. Several conditions, including type and volume of extraction solvent, type and concentration of the surfactant, ultrasound time, extraction temperature, pH, and ionic strength were studied and optimized. Under the optimum conditions, linear calibration curves were obtained in the ranges 4–5000 μg L⁻¹ for ketoconazole and 8–5000 μg L⁻¹ for econazole nitrate, with linear correlation coefficients for both >0.99. The limits of detection (LODs, S/N = 3) and enrichment factors (EFs) were 1.1 and 2.3 μg L⁻¹, and 129 and 140 for ketoconazole and econazole nitrate, respectively. Reproducibility and recovery were good. The method was successfully applied to the determination of ketoconazole and econazole nitrate in human blood samples.     

An improved high-performance liquid chromatography method for the determination of homocysteine in human plasma

Summary Elevated plasma homocysteine is, a known risk factor in arteriosclerotic vascular disease. To measure homocysteine in a large number of samples, we have developed a rapid, simple, robust and inexpensive reversed-phase HPLC method for routine analysis. Mercaptopro-pionylglycine was used as the internal standard and an external calibration in plasma was performed. Improvement was achieved by the use of gradient elution (using a sodium acetate buffer and methanol) resulting in a higher number of samples analyzed per day. Plasma samples were reduced with tributylphosphine and the proteins were precipitated with perchloric acid before addition of internal standard. The analytes were derivatized by use of 7-fluorobenzofurazone-4-sulfonic acid ammonium salt. For calibration human plasma was spiked with nine different concentrations of homocysteine (range 2–50 μmol L⁻¹). The inter-assay precision of replicate (n=29) analysis of the concentration of homocysteine in a sample of pooled plasma was 3.0%. The limit of detection, defined as three times the signal-to-noise ratio, was 0.25 μmol L⁻¹. The linearity of the assay was confirmed for a plasma concentration range of 2–2000 μmol L⁻¹. The variation of duplicate analyses of 842 plasma samples was 2.6±1.7%.     

Immunosensors for pollutants working in organic media. Study of performances of different tracers with luminescent detection

A comparative study of enzymatic and non-enzymatic labels combined with luminescence detection, developed for immunosensing of pesticide residues (carbaryl, 1-naphthol, irgarol 1051) in organic media, is presented. Peroxidase and alkaline phosphatase enzymes with fluorogenic (3-p-hydroxyphenylpropanoic acid) and luminogenic (AMPPD derivative) substrates, respectively, were assessed as enzymatic markers. As an alternative, terbium(III) chelate, with time-resolved fluorescence detection, was evaluated as a non-enzymatic label. The best sensitivity was achieved by use of alkaline phosphatase in an immunocomplex capture assay format (I ₅₀ values 0.06, 0.27, and 7.45 μg L⁻¹ in buffer, 1:1 methanol–buffer, and methanol, respectively). Results were also good (I ₅₀ 1.00 and 6.30 μg L⁻¹ for water and aqueous–organic mixture, respectively) for Tb(III) chelate in an immobilized conjugate assay format. Use of alkaline phosphatase label to measure carbaryl (100 ng L⁻¹) in different spiked river water samples, after solid-phase extraction and analyte elution with an ethyl acetate–methanol mixture, resulted in recoveries ranging from 81 to 98%, with acceptable precision (CV 4–14%, n=4).     

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.     

LC–MS–MS determination of ibuprofen, 2-hydroxyibuprofen enantiomers,and carboxyibuprofen stereoisomers for application in biotransformation studies employing endophytic fungi

The purpose of this study was the development and validation of an LC–MS–MS method for simultaneous analysis of ibuprofen (IBP), 2-hydroxyibuprofen (2-OH-IBP) enantiomers, and carboxyibuprofen (COOH-IBP) stereoisomers in fungi culture medium, to investigate the ability of some endophytic fungi to biotransform the chiral drug IBP into its metabolites. Resolution of IBP and the stereoisomers of its main metabolites was achieved by use of a Chiralpak AS-H column (150 × 4.6 mm, 5 μm particle size), column temperature 8 °C, and the mobile phase hexane–isopropanol–trifluoroacetic acid (95: 5: 0.1, v/v) at a flow rate of 1.2 mL min⁻¹. Post-column infusion with 10 mmol L⁻¹ ammonium acetate in methanol at a flow rate of 0.3 mL min⁻¹ was performed to enhance MS detection (positive electrospray ionization). Liquid–liquid extraction was used for sample preparation with hexane–ethyl acetate (1:1, v/v) as extraction solvent. Linearity was obtained in the range 0.1–20 μg mL⁻¹ for IBP, 0.05–7.5 μg mL⁻¹ for each 2-OH-IBP enantiomer, and 0.025–5.0 μg mL⁻¹ for each COOH-IBP stereoisomer (r ≥ 0.99). The coefficients of variation and relative errors obtained in precision and accuracy studies (within-day and between-day) were below 15%. The stability studies showed that the samples were stable (p > 0.05) during freeze and thaw cycles, short-term exposure to room temperature, storage at −20 °C, and biotransformation conditions. Among the six fungi studied, only the strains Nigrospora sphaerica (SS67) and Chaetomium globosum (VR10) biotransformed IBP enantioselectively, with greater formation of the metabolite (+)-(S)-2-OH-IBP. Formation of the COOH-IBP stereoisomers, which involves hydroxylation at C3 and further oxidation to form the carboxyl group, was not observed.     

In-line solid-phase extraction–capillary electrophoresis coupled with mass spectrometry for determination of drugs of abuse in human urine

In-line solid-phase extraction–capillary electrophoresis coupled with mass spectrometric detection (SPE–CE–MS) has been used for determination of 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), codeine (COD), hydrocodeine (HCOD), and 6-acetylmorphine (6AM) in urine. The preconcentration system consists of a small capillary filled with Oasis HLB sorbent and inserted into the inlet section of the electrophoresis capillary. The SPE–CE–MS experimental conditions were optimized as follows: the sample (adjusted to pH 6.0) was loaded at 930 mbar for 60 min, elution was performed with methanol at 50 mbar for 35 s, 60 mmol L⁻¹ ammonium acetate at pH 3.8 was used as running buffer, the separation voltage was 30 kV, and the sheath liquid at a flow rate of 5.0 μL min⁻¹ was isopropanol–water 50:50 (v/v) containing 0.5% acetic acid. Analysis of urine samples spiked with the four drugs and diluted 1:1 (v/v) was studied in the linear range 0.08–10 ng mL⁻¹. Detection limits (LODs) (S/N = 3) were between 0.013 and 0.210 ng mL⁻¹. Repeatability (expressed as relative standard deviation) was below 7.2%. The method developed enables simple and effective determination of these drugs of abuse in urine samples at the levels encountered in toxicology and doping.     

Determination of phenolic compounds in water samples by on-line solid-phase extraction—supercritical-fluid chromatography with diode-array detection

Summary The eleven Environmental Protection Agency (EPA) priority phenolic compounds have been determined by solid-phase extraction (SPE) coupled on-line to supercritical-fluid chromatography (SFC) with diodearray detection. The variables affecting chromatographic separation were optimized and the analytes were separated at 40 °C in two diol columns connected in series; a gradient of methanol, as modifier, and CO₂ was used as mobile phase. Under these conditions, all the compounds studied were separated to baseline in less than 13 min. PLRP-S and LiChrolut EN were tested as sorbents in a 10×3 mm i.d. laboratory-packed precolumn for solid-phase extraction. An ion-pair reagent, tetrabutylammonium bromide (TBA), was used in the extraction process to increase break-through volumes. The performance of the method was checked with tap and river waters and the pre-concentration of 20 mL of sample in a PLRP-S pre-column enabled phenolic compounds to be determined at low μg L⁻¹ levels with limits of detection ranging between 0.4 and 2 μg L⁻¹. The repeatability and reproducibility between days (n=3) for real samples spiked at 10 μg L⁻¹ were lower than 10%.     

Micelle-mediated extractions using nonionic surfactant mixtures and HPLC-UV to determine endocrine-disrupting phenols in seawaters

An environmentally friendly method to extract endocrine-disrupting phenols (EDPs) from seawaters was realized using nonionic surfactant mixtures and micelle-mediated extractions. The preconcentration step was achieved directly in the seawater matrix, and was followed by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection without any clean-up steps to remove the surfactant mixture prior to injection. Various nonionic surfactant mixtures were used, and polyoxyethylene-10-laurylether (POLE) with polyoxyethylene-4-laurylether (Brij 30) was found to be the best to work with. Method optimization involved maximizing the preconcentration factor using the studied mixtures. The proposed method gave extraction recoveries ranging from 83.3 to 114.4% for an EDP spiking level of 46.7 μg L⁻¹, and from 63.4 to 112.4% for a spiking level of 4.7 μg L⁻¹ for EDPs studied in real seawater matrices, with relative standard deviations of <12.1%. The detection limits of the method varied from 0.18 μg L⁻¹ for bisphenol A (BPA) to 1.17 μg L⁻¹ for 4-cumylphenol (4-CP). The method was applied to seawaters from the Canary Islands with successful results. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Multi-component analysis of tetracyclines, sulfonamides and tylosin in swine manure by liquid chromatography–tandem mass spectrometry

A multi-component method focussing on thorough sample preparation has been developed for simultaneous analysis of swine manure for three classes of antibiotic—tetracyclines, sulfonamides, and tylosin. Liquid manure was initially freeze-dried and homogenised by pulverization before extraction by pressurised liquid extraction. The extraction was performed at 75°C and 2,500 psig in three steps using two cycles with 0.2 mol L⁻¹ citric acid buffer (pH 4.7) and one cycle with a mixture of 80% methanol with 0.2 mol L⁻¹ citric acid (pH 3). After liquid–liquid extraction with heptane to remove lipids, the pH of the manure was adjusted to 3 with formic acid and the sample was vacuum-filtered through 0.6 μm glass-fibre filters. Finally the samples were pre-concentrated by tandem SPE (SAX-HLB). Recoveries were determined for manure samples spiked at three concentrations (50–5,000 μg kg⁻¹ dry matter); quantification was achieved by matrix-matched calibration. Recoveries were >70% except for oxytetracycline (42–54%), sulfadiazine (59–73%), and tylosin (9–35%) and did not vary with concentration or from day-to-day. Limits of quantification (LOQ) for all compounds, determined as a signal-to-noise ratio of 10, were in the range 10–100 μg kg⁻¹ dry matter. The suitability of the method was assessed by analysis of swine manure samples from six different pig-production sites, e.g. finishing pigs, sows, or mixed production. Residues of antibiotics were detected in all samples. The largest amounts were found for tetracyclines (up to 30 mg kg⁻¹ dry matter for the sum of CTC and ECTC). Sulfonamides were detected at concentrations up to 2 mg kg⁻¹ dry matter (SDZ); tylosin was not detected in any samples.      

Application of molecularly imprinted polymer solid-phase extraction for salivary cotinine

A method constituted by molecularly imprinted solid-phase extraction (MISPE) with high-performance liquid chromatography coupled to diode array detector (HPLC-DAD) was developed for cotinine analysis in saliva samples. For this purpose, the separation was carried out with a C18 reversed-phase column at 20 °C. The mobile phase which was composed of a mixture of 09:91 (v/v) acetonitrile/phosphate buffer, pH 6.3, was delivered with isocratic flow rate at 1.4 mL min⁻¹. Employing MISPE, the best conditions were achieved with 1.5 mL of saliva plus 1.5 mL of 0.1 mol L⁻¹ of acetate buffer, pH 5.5, which were then passed through a cartridge previously conditioned with 2 mL acetonitrile, 2 mL methanol, and 2 mL of 0.1 mol L⁻¹ sodium acetate buffer, pH 5.5. The washing was carried out with 1 mL deionized water, 1 mL of 0.1 mol L⁻¹ sodium hydroxide, and 1 mL hexane; finally; the cotinine elution was carried out with 3 mL methanol/water (97.5: 2.5, v/v). Linearity ranged from 30 to 500 ng mL⁻¹ with r > 0.99. Intra-assay, interassay precision, and accuracy ranged from 3.1% to 10.1%, 5.2% to 15.9%, and 99.22% to 111.17%, respectively. The detection and quantification limits were 10 and 30 ng mL⁻¹, respectively. This investigation has provided a reliable method for routine cotinine determination in saliva, and it is an important tool for monitoring cigarette smoke exposure in smokers. The method was applied in five smokers’ samples who consumed around five to 20 cigarettes per day and the values of cotinine in saliva were from 66.7 to 316.16 ng mL⁻¹.     

On-line determination of bipyridylium herbicides in water by HPLC

Summary Selective on-line solid phase extraction (SPE) and liquid chromatography determination (HPLC) of diquat, paraquat and difenzoquat from environmental water samples has been accomplished with Graphitized Carbon Black (GCB) as both extraction and analytical columns. The method involved passing of 50 mL of water through a cartridge filled with Carbograph. In the elution step, the herbicides were transferred from the cartridge to the analytical column (Hypercarb) by mean of a gradient of pH 3 aqueous solution of tetramethylammonium hydroxide (TMAOH) and ammonium sulphate and methanol. Hypercarb columns were found to give a low probability of false positives for bypiridylium herbicides and are very selective for polar compounds. Recovery was better than 80 %. The breakthrough volume was studied with distilled water spiked with the herbicides at various concentration levels (from 0.1 to 20 μg L⁻¹). The limits of quantification of the method were lower than 0.1 μg L⁻¹. The total analytical method was applied to surface waters from Torreblanca Nature Park (Castelló, Spain). Presented at the 21^st ISC held in Stuttgart, Germany, 15^th–20^th September, 1996.     

Electrochemical behavior and differential pulse voltammetric detection of thiobencarb on 2-(4-((4-ethoxyphenyl)diazenyl)phenylamino)ethanol-modified carbon paste electrode

Cyclic voltammetry, chronoamperometry, and rotating disk electrode voltammetry were used to investigate the electrochemical behavior of thiobencarb (TB) at carbon paste electrode modified with an azo dye, 2-(4-((4-ethoxyphenyl)diazenyl)phenylamino)ethanol (EDPE), EDPE/modified carbon paste electrode (MCPE). The modified electrode showed high electrocatalytic activity toward thiobencarb. The current was enhanced significantly relative to the situation prevailing when a bare glassy carbon electrode was used. The kinetics parameters of this process were calculated, the apparent electron transfer rate constant k _s and α (charge transfer coefficient between electrode and EDPE) were 14.6 s⁻¹ and 0.48, respectively. The experimental parameters were optimized, and the mechanism of the catalytic process was discussed. The best defined cathodic peak was obtained with 0.1 M acetate buffer (pH 3.0). The response of the sensor was very quick, and response time was approximately 5 s. The differential pulse voltammetry response of the MCPE was linear against the concentration of TB in the range of 0.96 to 106 μg L⁻¹. The limit of detection was found to be 0.025 μg L⁻¹. The precision was examined by carrying out eight replicate measurements at a concentration of 25 μg L⁻¹ TB; the relative standard deviation was 2.9%.     

Analysis of Terbumeton and its Major Metabolites by SPE and DAD-HPLC in Soil Bulk Water

A rapid and inexpensive method for simultaneous quantification of terbumeton (TER), and its major potential metabolites (TED; terbumeton-desethyl, TOH; terbumeton-2-hydroxy and TID; terbumeton-deisopropyl) in soil bulk water (SBW) samples is proposed. The analytical method involves extraction–concentration from SBW samples using a graphitized carbon black (GCB) cartridge followed by their separation–detection by reversed-phase high-performance liquid chromatography analysis using a C₁₈ column and a diode array detector. A mobile phase of acetonitrile−0.005 mol L⁻¹ phosphate buffer (pH 7.0) (35:65, v/v) at a flow rate of 0.8 mL min⁻¹ in isocratic elution mode has been used. After optimization of the extraction and separation conditions, this method can be used for the simultaneous determination of investigated compounds in the range of the international limits of 0.1 μg L⁻¹. For TER the detection limit was 0.009 μg L⁻¹ and it was 0.100, 0.550, and 0.480 μg L⁻¹ for TED, TOH, and TID, respectively. The recoveries of TER, TED, TOH, and TID from SBW samples, measured at three levels of concentration range, were found to be between 48.0 and 102.0%. The intra-day precision measured by relative standard deviation (RSD) was always lower than 9.0%.     

Profiling of amine metabolites in human biofluids by micellar electrokinetic chromatography with laser-induced fluorescence detection

A rapid and sensitive capillary electrophoresis (CE) method has been developed for profiling organic metabolites containing amine functional groups in mammalian biofluids. Metabolites containing an amine group were derivatized with 4-fluoro-7-nitrobenzo-2,1,3-oxadiazol (NBD-F), separated by micellar electrokinetic chromatography (MEKC), and detected by argon-ion (488 nm) laser-induced fluorescence detection. The optimized MEKC background electrolyte conditions were: 50 mmol L⁻¹ sodium cholate, 5 mmol L⁻¹ β-cyclodextrin, and 20 mmol L⁻¹ Brij 35 in 20 mmol L⁻¹ aqueous borate buffer, pH 9.3, containing 7% methanol. Under these conditions all the amine compounds in mammalian biofluids, for example plasma, saliva, and urine, were derivatized directly, without extraction, in a minimum volume of 100 nL and the derivatives could be separated within 16 min. Up to 90% of the amine-containing metabolites in plasma and saliva could be identified by reference to standard compounds. For twenty amine standards linearity of calibration was better than R ² = 0.99. Migration-time and peak-area reproducibility were better than RSD 1.5% and 15% respectively. In replicate analysis of human plasma bioanalytical precision ranged between 0.7 and 3.8 RSD% for a 5.0-μL volume and between 1.7 and 5.5 RSD% for 100-nL volume. The concentrations measured were found to be in agreement with literature values.     

Technology of DTPA and immunoglobulins conjugation and their attachment to <Superscript>90</Superscript>Y and <Superscript>177</Superscript>Lu radionuclides

The aim of the study of labeling of ligand–antibody conjugates was to find optimal conditions of preparing of these conjugates and appropriate radioactivity of selected nuclide for applications in nuclear medicine. Conjugation of the γ-immunoglobulin G (human or bovine IgG, polyclonal antibodies) and bifunctional chelating agent, diethylenetriaminepentaacetic acid dianhydride (cDTPAA), was carried out. Various values of the cDTPAA/antibody ratio, the weight concentration of polyclonal or monoclonal antibodies (MEM-97) and buffers were used. Further, the labeling conditions of the DTPA–IgG conjugate by radionuclides ⁹⁰Y and ¹⁷⁷Lu were optimized, and the labeling yield and the conjugation ratio of prepared radionuclide–DTPA–IgG conjugates was determined. Optimal incubation time of the immunoglobulin conjugation was obtained at 30 min from mixing of individual components. The labeling yield of radionuclide–DTPA–antibody conjugate higher than 95% was achieved. Higher values of conjugation ratio of radionuclide–DTPA–antibody conjugate were achieved in 0.1 mol L⁻¹ carbonate buffer, pH 8.5, and the 0.1 mol L⁻¹ carbonate buffer is suitable for studied conjugation systems. This study showed that the labeling yield as well as the conjugation ratio of tested systems depend on the amount of antibody substance, bifunctional chelating agent/antibody molar ratio and pH value of the buffer used.     

Optimisation and comparison of several microextraction/methylation methods for determining haloacetic acids in water using gas chromatography

This article presents the different modes and configurations of liquid-phase microextraction (LPME) through comparison with headspace solid-phase microextraction (HS-SPME) for the simultaneous extraction/methylation of the nine haloacetic acids (HAAs) found in water. This is the first analytical case reported of solvent bar extraction–preconcentration–derivatisation assisted by an ion-pairing transfer for HAAs. In this method, 5 μL of the organic extractant, decane, was confined within a hollow-fibre membrane that was placed in a stirred aqueous sample containing the derivatising reagents (dimethylsulphate with a tetrabutylammonium salt). With heating at 45 °C in the HS-SPME method, some organic solvents (extractant, excess of derivatising reagent) are also volatilised and compete with the esters on the fibre (the fibre is damaged and it can be reused only 50−60 times). In addition, the HS-SPME method provides inadequate sensitivity (limits of detections between 0.3 and 5 μg/L) to quantify HAAs at the level usually found in drinking waters. Alternative headspace LPME methods for HAAs require heating (45 °C, 25 min) to derivatise and volatilise the esters but, by using solvent bar microextraction (SBME), the extraction/methylation takes place at room temperature without degradation of HAAs to trihalomethanes. Adequate precision (relative standard deviation of approximately 8%), linearity (0.1–500 μg/L) and sensitivity (10 times higher than the HS-SPME alternative) indicate that the SBME method can be a candidate for routine determination of HAAs in tap water. Finally, the SBME method was applied for the analysis of HAAs in tap and swimming pool water and the results were compared with those of a previous validated headspace gas chromatography–mass spectrometry method.