Trace determination of triclosan and triclocarban in environmental water samples with ionic liquid dispersive liquid-phase microextraction prior to HPLC–ESI-MS–MS

A novel and environmentally friendly microextraction method, termed ionic liquid dispersive liquid-phase microextraction (IL-DLPME), has been developed for rapid enrichment of triclosan and triclocarban before analysis by high-performance liquid phase chromatography–electrospray tandem mass spectrometry (HPLC–ESI-MS–MS). Instead of using toxic organic solvents, an ionic liquid was used as a green extraction solvent. This also avoided the instability of the suspending drop in single-drop liquid-phase microextraction, and the heating and cooling step in temperature-controlled ionic liquid dispersive liquid phase microextraction. Factors that may affect the enrichment efficiency, for example volume of ionic liquid, type and volume of dispersive solvent, pH, extraction time, and NaCl content were investigated in detail and optimized. Under optimum conditions, linearity of the method was observed over the range 0.2–12 μg L⁻¹ for triclocarban and 1–60 μg L⁻¹ for triclosan with correlation coefficients ranging from 0.9980 to 0.9990, respectively. The sensitivity of the proposed method was found to be excellent, with limits of detection in the range 0.040–0.58 μg L⁻¹ and precision in the range 7.0–8.8% (RSD, n = 5). This method has been successfully used to analyze real environmental water samples and satisfactory results were achieved. Average recoveries of spiked compounds were in the range 70.0–103.5%. All these results indicated that the developed method would be a green method for rapid determination of triclosan and triclocarban at trace levels in environmental water samples.     

Enzyme-linked immunosorbent assays for the sensitive analysis of 2,4-dinitroaniline and 2,6-dinitroaniline in water and soil

The development and characterization of one rat monoclonal antibody (mAb) for 2,4-dinitroaniline and of two rat mAbs for 2,6-dinitroaniline are described. With the immunization of rats with 2,4,6-trinitrophenyl-glycylglycine–keyhole limpet hemocyanine (KLH) conjugate one mAb (PK 5H6) has been developed and formatted into a competitive enzyme-linked immunosorbent assay (ELISA). This assay no. 1 is very sensitive for 2,4-dinitroaniline with a test midpoint of 0.24 ± 0.06 μg L⁻¹ (n = 19) in 40 mM phosphate-buffered saline (PBS). A second hapten, 3-(4-amino-2,6-dinitrophenyl)propionic acid, which was also conjugated to KLH and used for the immunization of rats, led to two sensitive ELISAs for 2,6-dinitroaniline in 40 mM PBS with test midpoints of 0.61 ± 0.08 μg L⁻¹ (n = 15; mAb DNT4 3C6; assay no. 2) and 0.94 ± 0.29 μg L⁻¹ (n = 17; mAb DNT4 1A7, assay no. 3). Selectivities of all mAbs were checked with more than 20 compounds, including nitroaromatic compounds, 2,6-dinitroaniline pesticides, and other substituted derivatives of aniline. As very noticeable cross-reactivities, all mAbs recognize 2-chloro-4,6-dinitroaniline, 4-chloro-2,6-dinitroaniline and 2-bromo-4,6-dinitroaniline, the last of these being a major metabolite of the azo dye Disperse Blue 79. As first demonstrations of applications, two ELISAs (assays no. 1 and 2) were used for the analysis of 2,4- or 2,6-dinitroaniline in spiked water and soil samples. Recovery data were determined and the majority of these data were in the range of 90–120%. These assays can contribute to a very cost-effective and environmentally friendly immunochemical surveillance monitoring of environmental samples for contaminations with these compounds. To the best of the authors’ knowledge, these are the first antibodies described for 2,4-dinitroaniline and for 2,6-dinitroaniline.     

Anabolic, doping, and lifestyle drugs, and selected metabolites in wastewater—detection, quantification, and behaviour monitored by high-resolution MS and MS n before and after sewage treatment

Municipal wastewater has been examined for steroids, β₂-agonists, stimulants, diuretics, and phosphodiesterase type V inhibitors (PDE type V inhibitors), which are “dual-use-drugs” applied either as anabolic, doping, and lifestyle drugs or for treatment of diverse diseases. To identify their origin, fitness centre discharges under suspicion of being point sources and sewage-treatment plant feed and effluents were sampled and concentrations determined. Sensitive and selective methods for determination and quantification based on solid-phase extraction (SPE) followed by high-performance liquid chromatography–high resolution mass and tandem mass spectrometry (HPLC–(HR)MS and HPLC–MS–MS) were developed and established for analysis of these compounds in wastewater and to assess their effect on the environment. The methods developed enabled quantification at trace concentrations (limit of quantification (LOQ): 5 ng L⁻¹). Of the steroids and stimulants under investigation, testosterone, methyltestosterone, and boldenone or ephedrine, amphetamine, and MDMA (3,4-methylendioxy-N-methylamphetamine) were observed at up to 5 μg L⁻¹ (ephedrine). Of the β₂-agonists salbutamol only, and of the diuretics furosemide and hydrochlorothiazide were confirmed in the extracts. Quite high concentrations of the PDE type V inhibitors sildenafil, tadalafil, and vardenafil and their metabolites were confirmed in fitness centre discharges (sildenafil: 1,945 ng L⁻¹) whereas their concentrations in municipal wastewater did not exceed 35 ng L⁻¹. This study identified anabolic and doping drugs in wastewater for the first time. Results obtained from wastewater treatment plant effluents proved that these “dual-use-drugs”, with the exception of hydrochlorothiazide, were mostly eliminated.     

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.     

A novel and rapid method for determination of natamycin in wines based on ultrahigh-performance liquid chromatography coupled to tandem mass spectrometry: validation according to the 2002/657/EC European decision

A novel, simple, and rapid reversed-phase liquid chromatography–tandem mass spectrometric methodology was developed for the analysis of natamycin in wine samples. Natamycin was protonated to form singly charged ions in an electrospray positive ion mode. Data acquisition under MS/MS was achieved by applying multiple reaction monitoring (MRM) of three fragment ion transitions (666.3 → 648.2, 666.3 → 503.3, and 666.3 → 485.2) to provide a high degree of sensitivity and specificity. Chromatographic separation was performed on a rapid resolution column using a mobile phase consisting of an acetonitrile/water mixture with a total run time of 5.0 min. After only filtration as pretreatment, the sample was injected into the chromatographic system. The proposed method was validated in terms of selectivity, trueness, precision, decision limit (CCα), and detection capability (CCβ) according to 2002/657/EC Commission decision. The values for trueness, reported as bias (%), agreed with those established by the aforementioned document. Repeatability (intraday variability) values were 12.37% at a concentration of 1.0 μg L⁻¹ and 8.99–4.19% at concentrations between 2.5 and 10 μg L⁻¹. The overall within-laboratory (interday variability) reproducibility was 15.47% at a concentration of 1.0 μg L⁻¹, which was significantly lower than the indicative value reported in the EU decision. The results indicated that the proposed approach is a sensitive, fast, reproducible, and robust methodology suitable for the analysis of natamycin levels in wine samples.     

Production and analytical characterization of neopterin immunoreagents for biosensor developments

Neopterin is a valuable biomarker of cellular immunity associated with various pathological situations such as viral and bacterial infections, autoimmune, cardiovascular, neurodegenerative and malignant disorders. To produce specific antibodies against neopterin for a rapid multi-biomarker-based diagnosis, a novel hapten derivative was synthesized and attached to carrier proteins. The thoroughly characterized conjugates were used for immunization of BALB/c mice and rabbits. The produced monoclonal antibody reached in both direct and indirect enzyme-linked immunosorbent assay (ELISA) format LoD of 0.18 and 0.45 μg L⁻¹, respectively, and was a superior immunoreagent for further biosensor developments with regard to assay sensitivity and material availability. The best polyclonal antibody was somewhat more sensitive in direct ELISA with LoD of 0.05 μg L⁻¹. The optimized ELISA method was evaluated with blood samples collected from patients with renal insufficiency, patients with sepsis, patients without confirmed clinical diagnosis, and healthy volunteers. In plasma samples, neopterin concentrations ranging from 3.2 to 103 μg L⁻¹ could be determined with the monoclonal ELISA whereas twofold lower results were obtained with the polyclonal ELISA. A satisfactory correlation of results was found between the polyclonal ELISA and IBL Neopterin ELISA kit within the concentration range 0.5–16 μg L⁻¹ (R = 0.874; n = 40), and slightly lower correlation was found for monoclonal-based ELISA (R = 0.819; n = 40). These data show that the generated antibodies may be used as functional analytical reagents for the integration into multianalyte biochip detection systems.     

Determination of enzyme activity inhibition by FTIR spectroscopy on the example of fructose bisphosphatase

A mid-infrared enzymatic assay for label-free monitoring of the enzymatic reaction of fructose-1,6-bisphosphatase with fructose 1,6-bisphosphate has been proposed. The whole procedure was done in an automated way operating in the stopped flow mode by incorporating a temperature-controlled flow cell in a sequential injection manifold. Fourier transform infrared difference spectra were evaluated for kinetic parameters, like the Michaelis–Menten constant (K _M) of the enzyme and V _max of the reaction. The obtained K _M of the reaction was 14 ± 3 g L⁻¹ (41 μM). Furthermore, inhibition by adenosine 5′-monophosphate (AMP) was evaluated, and the K _M^App value was determined to be 12 ± 2 g L⁻¹ (35 μM) for 7.5 and 15 μM AMP, respectively, with V _max decreasing from 0.1 ± 0.03 to 0.05 ± 0.01 g L⁻¹ min⁻¹. Therefore, AMP exerted a non-competitive inhibition.     

Rapid residue analysis of four triazolopyrimidine herbicides in soil, water, and wheat by ultra-performance liquid chromatography coupled to tandem mass spectrometry

A sensitive and effective method for simultaneous determination of triazolopyrimidine sulfonamide herbicide residues in soil, water, and wheat was developed using ultra-performance liquid chromatography coupled with tandem mass spectrometry. The four herbicides (pyroxsulam, flumetsulam, metosulam, and diclosulam) were cleaned up with an off-line C18 SPE cartridge and detected by tandem mass spectrometry using an electrospray ionization source in positive mode (ESI+). The determination of the target compounds was achieved in <2.0 min. The limits of detection were below 1 μg kg⁻¹, while the limits of quantification did not exceed 3 μg kg⁻¹ in different matrices. Quantitation was determined from calibration curves of standards containing 0.05–100 μg L⁻¹ with r ² > 0.997. Recovery studies were conducted at three spiked levels (0.2, 1, and 5 μg kg⁻¹ for water; 5, 10, and 100 μg kg⁻¹ for soil and wheat). The overall average recoveries for this method in water, soil, wheat plants, and seeds at three levels ranged from 75.4% to 106.0%, with relative standard deviations in the range of 2.1–12.5% (n = 5) for all analytes.     

Miniaturized liquid–liquid extraction coupled with ultra-performance liquid chromatography/tandem mass spectrometry for determination of topramezone in soil,corn, wheat,and water

A rapid and simple miniaturized liquid–liquid extraction method has been developed for the determination of topramezone in soil, corn, wheat, and water samples using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-electrospray ionization (ESI)/MS/MS). The established method for the extraction and purification procedure was based on liquid–liquid partitioning into an aqueous solution at a low pH (pH ≈ 2.5), followed by back-partitioning into water at pH > 9. Two precursor, product ion transitions for topramezone were measured and evaluated to provide the maximum degree of confidence in the results. Under negative ESI conditions, quantitation was achieved by monitoring the fragment at m/z = 334 and the qualitative fragment at m/z = 318, whereas also collecting the corresponding parent ion at m/z = 362. Chromatographic separation was achieved using gradient elution with a mobile phase consisting of methanol and a 0.01% aqueous ammonium hydroxide solution. Recovery studies for soil, corn, wheat, and water were conducted at four different topramezone concentrations (5 or 10, 50, 100, and 1,000 μg kg⁻¹); the overall average recoveries ranged from 79.9% to 98.4% with intra-day relative standard deviations (RSD) of 3.1~8.7% and inter-day RSD of 4.3~7.5%. Quantitative results were determined from calibration curves of topramezone standards containing 1–500 μg L⁻¹ with an R ² ≥ 0.9994. Method sensitivities expressed as limits of quantitation were typically 6, 8, 9, and 1 μg kg⁻¹ in soil, corn, wheat, and water, respectively. The results of the method validation confirmed that this proposed method was convenient and reliable for the determination of topramezone residues in soil, corn, wheat, and water.     

Analysis of captan, folpet, and captafol in apples by dispersive liquid–liquid microextraction combined with gas chromatography

A novel method was developed for the determination of captan, folpet, and captafol in apples by dispersive liquid–liquid microextraction (DLLME) coupled with gas chromatography–electron capture detection (GC–ECD). Some experimental parameters that influence the extraction efficiency, such as the type and volume of the disperser solvents and extraction solvents, extraction time, and addition of salt, were studied and optimized to obtain the best extraction results. Under the optimum conditions, high enrichment factors for the compounds were achieved ranging from 824 to 912. The recoveries of fungicides in apples at spiking levels of 20.0 μg kg⁻¹ and 70.0 μg kg⁻¹ were 93.0–109.5% and 95.4–107.7%, respectively. The relative standard deviations (RSDs) for the apple samples at 30.0 μg kg⁻¹ of each fungicide were in the range from 3.8 to 4.9%. The limits of detection were between 3.0 and 8.0 μg kg⁻¹. The linearity of the method ranged from 10 to 100 μg kg⁻¹ for the three fungicides, with correlation coefficients (r ²) varying from 0.9982 to 0.9997. The obtained results show that the DLLME combined with GC–ECD can satisfy the requirements for the determination of fungicides in apple samples. Figure Dispersive liquid–liquid microextraction (DLLME) coupled with gas chromatography–electron capture detection (GC–ECD) allows satisfactory determination of fungicides in apple 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.     

Stannum film electrode for square wave voltammetric determination of trace copper(II)

An anodic stripping voltammetric procedure for the determination of Cu(II) at an in situ-plated stannum film electrode (SnFE) was described. The results indicated that the SnFE had an attractive electroanalytical performance, with two distinct voltammetric stripping signals for copper and stannum, and showed the superior advantage for the determination of copper compared with the bismuth film electrode. Several experimental parameters were optimized. The SnFE exhibited highly linear behavior in the concentration range from 1.0 to 100.0 μg L⁻¹ of Cu(II) (r = 0.994) with the detection limit of 0.61 μg L⁻¹ (S/N = 3), and the relative standard deviation for a solution containing 40.0 μg L⁻¹ Cu(II) was 2.2% (n = 8). The procedure has been successfully applied for the determination of Cu(II) in lake water sample.     

Determination of butoxyacetic acid (biomarker of ethylene glycol monobutyl ether exposure) in human urine candidate reference material

Ethylene glycol monobutyl ether (EGBE), an industrial solvent, is absorbed by the body not only by inhalation but also by dermal absorption (liquid or vapour). EGBE is metabolized to butoxyacetic acid (BAA). Pooled freeze-dried urine candidate reference material (RM) was prepared from urine obtained from persons occupationally exposed to EGBE. This material has the advantage of containing butoxyacetic acid in both the free and conjugated (glutamine and glycine) forms, as found in native urine. In all GC method modifications used, acid hydrolysis was used to release BAA from its conjugated form. The amount of butoxyacetic acid in homogeneity and stability testing was measured by GC after derivatisation with N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide. Detection was by MS in EI mode, in the authors’ laboratory. For interlaboratory comparison of the reference material GC methods with MS, FID, and ECD were used. Different extraction solvents (dichloromethane–isopropanol 2:1, ethyl acetate, or dichloromethane) and derivatisation reagents (trimethylsilyldiazomethane, N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide) were used. Using ANOVA (at the statistical level α = 0.05) no changes were found in the concentration of butoxyacetic acid during fifteen month isochronous stability testing, or in homogeneity testing. The uncertainty contributions were u _h = 8.8 mg L⁻¹ and u _s = 6.5 mg L⁻¹. The concentration of butoxyacetic acid in freeze-dried urine RM was evaluated from the results of eight laboratory data sets within an interlaboratory comparison by use of the interactive statistical software IPECA. The contribution to total uncertainty derived from interlaboratory comparison was u _i = 12.7 mg L⁻¹. The reference value (c = 273 ± 33 mg L⁻¹) is an unweighted arithmetic average of accepted results. The value is traceable to the pure butoxyacetic acid (98% w/w; Acros Organic #257760010) used as calibrant. The uncertainty given is combined expanded uncertainty derived from the results from interlaboratory comparison, and from homogeneity and stability tests (k = 2). The reference material will be used to verify method performance in the biological monitoring of occupational exposure to EGBE.     

Determination of low-level ink photoinitiator residues in packaged milk by solid-phase extraction and LC-ESI/MS/MS using triple-quadrupole mass analyzer

A confirmatory and quantitative method based on liquid chromatography–electrospray ionization tandem mass spectrometry (LC-ESI/MS/MS) has been developed for simultaneous determination of seven photoinitiator residues: benzophenone, (1-hydroxycyclohexyl)phenylketone (Irgacure 184), isopropylthioxanthone (ITX), 2-ethylhexyl-(4-dimethylamino)benzoate (EHA or EHDAB), 2-methyl-1-[4-(methylthio)phenyl]-2-(4-morpholinyl)-1-propanone (Irgacure 907), (2,4,6-trimethylbenzoyl)diphenylphosphine oxide (TPO) and 2-benzyl-2-(dimethylamino)-1-(4-morpholinophenyl)-1-butanone (Irgacure 369) in packaged milk and related packaging materials. Residues of photoinitiators were extracted from milk using acetonitrile, and further enriched and purified on HLB solid-phase extraction cartridges prior to being analyzed by LC-ESI/MS/MS with selected reaction monitoring mode, while photoinitiators in packaging materials were extracted using the same solvent. Satisfactory recovery (from 80 to 111%), intra- and inter-day precision (below 12%), and low limits of quantification (from 0.1 to 5.0 μg kg⁻¹) were evaluated from spiked samples at three concentration levels (5.0, 10.0 and 25.0 μg kg⁻¹ for Irgacure 184 and 2.5, 5.0 and 25.0 μg kg⁻¹ for others). These excellent validation data suggested the possibility of using the LC-ESI/MS/MS method for simultaneous determination of low-level photoinitiator residues migrating from printed food-packaging materials into milk. The method has been successfully applied to the analysis of real samples of different fat contents ranging from 8 to 30 g L⁻¹. The photoinitiator residues were revealed to be higher in milk with higher fat content and the most important contaminations were benzophenone and ITX in concentration ranges of 2.84–18.35 and 0.83–8.87 μg kg⁻¹, respectively.     

Determination of antibacterials in animal feed by pressurized liquid extraction followed by online purification and liquid chromatography-electrospray tandem mass spectrometry

A fully automated method has been developed for analysis of eighteen antibacterial compounds, including penicillins, cephalosporins and sulfonamides, in animal feed with limits of quantification in the range 0.25–5.79 μg kg⁻¹. The method is based on pressurized liquid extraction of 3 g homogenized feed with water and online clean-up of 500 μL of the extract with C₁₈HD cartridges. The purified sample was directly analysed by liquid chromatography–electrospray tandem mass spectrometry (SPE–LC–ESI-MS–MS). Chromatographic separation was achieved within 10 min by use of a C₁₂ Phenomenex Hydro-RP reversed-phase analytical column and a mobile phase gradient (water + 0.1% formic acid–methanol + 0.1% formic acid). The method was validated, revealing capability for detection of concentrations as low as 0.09 μg kg⁻¹, decision limits (CCα) and detection capabilities (CCβ) in the range 10–174 μg kg⁻¹ and 22–182 μg kg⁻¹, respectively, and inter-day precision ranging from 0.7 to 8.3%. Recovery, with internal standard correction, was in the range 93–134% for all analytes. The method was then applied to analysis of fifteen feed samples, nine of which contained at least one antimicrobial at concentrations between 0.006 and 1.526 mg kg⁻¹. The performance data and results from the method were compared with those from a previous method developed by our group, using offline SPE, by analyzing the same set of samples by both methods. The online SPE approach resulted in slightly improved sensitivity, with LODs of 0.09–2.12 μg kg⁻¹ compared with 0.12–3.94 μg kg⁻¹ by the offline approach. In general, better recovery was achieved by use of online purification (for 72% of the analytes) and the correlation between the two methods was good. The main advantages of the new online method are rapid and automated sample pre-treatment, and reduction of sample manipulation, enabling high-throughput analysis and highly accurate results. Because of all these characteristics, the proposed method is applicable and could be deemed necessary within the field of food control and safety.     

Preparation of antibodies and development of a sensitive immunoassay with fluorescence detection for triazine herbicides

Specific polyclonal antibodies against s-triazine herbicides were obtained by preparing immunogens coupling home-synthesized haptens derivatives of simazine (6-chloro-N-ethyl-N′-ethyl-1,3,5-triazine-2,4-diamine) to lysine groups of hemocyanin from keyhole limpets and bovine serum albumin carrier proteins. Three highly sensitive rabbit antisera were obtained and evaluated with a battery of six enzyme tracers derived from triazine structures in an optimized ELISA format. The antiserum As8 and the HRP-2f tracer, which yield the best assay sensitivity for simazine (detection limit 0.11 ± 0.02 μg L⁻¹, IC₅₀ 0.88 ± 0.04 μg L⁻¹), were applied to the development of a sensitive flow-through immunoassay for the analysis of this herbicide. The automated assay was based on a direct competitive immunosorbent assay and fluorescence detection. The optimized method presents an IC₅₀ value of 0.35 ± 0.04 μg L⁻¹ with a detection limit of 1.3 ± 0.9 ng L⁻¹ and a dynamic range from 0.010 to 7.5 μg L⁻¹ simazine. The generic nature of the antiserum was shown by good relative cross-reactivities with other triazines such as atrazine (420%) or propazine (130%) and a lower response to terbutylazine (6.4%) and desethyl-atrazine (2.2%). No cross-reactivity was obtained for nonrelated pesticides such as 2,4-dichlorophenoxyacetic acid or linuron and the assay could be applied as a screening method for triazine herbicides. The total analysis time was 30 min per determination and the immunosensor could be reused for more than 150 cycles without significant loss of activity. The immunosensor has been successfully applied to the direct analysis of simazine in surface water samples at the nanogram per liter level. The results obtained by comparative analysis of the immunosensor with a chromatographic procedure for triazines showed a close correspondence.     

Microwave-assisted hydrolysis and extraction of tricyclic antidepressants from human hair

The objective of this research was to develop, optimize, and validate a modern, rapid method of preparation of human hair samples, using microwave irradiation, for analysis of eight tricyclic antidepressants (TCADs): nordoxepin, nortriptyline, imipramine, amitriptyline, doxepin, desipramine, clomipramine, and norclomipramine. It was based on simultaneous alkaline hair microwave-assisted hydrolysis and microwave-assisted extraction (MAH–MAE). Extracts were analyzed by high-performance liquid chromatography with diode-array detection (HPLC–DAD). A mixture of n-hexane and isoamyl alcohol (99:1, v/v) was used as extraction solvent and the process was performed at 60°C. Application of 1.0 mol L⁻¹ NaOH and microwave irradiation for 40 min were found to be optimum for hair samples. Limits of detection ranged from 0.3 to 1.2 μg g⁻¹ and LOQ from 0.9 to 4.0 μg g⁻¹ for the different drugs. This enabled us to quantify them in hair samples within average therapeutic concentration ranges.     

Comparison of CIM discs and CPG glass as solid supports for bioanalytical columns used in allergen detection

This work presents a comparison of convective interaction media (CIM) and controlled pore glass (CPG) as solid supports for immunoglobulin antibodies used in bioanalytical detection of allergens in foodstuffs. A flow-injection manifold with highly sensitive thermal lens spectrometric detection was used for this purpose. Using beta-lactoglobulin, a milk allergen, as a model analyte, CIM disc supports had a higher linear range (0.2–3.5 μg L⁻¹), better reproducibility (intra-day RSD = 1%, inter-day RSD = 10%), lower consumption of reagents, and better immunocolumn stability (1 month, over 240 injections of substrate), while providing comparable LODs (0.1 μg L⁻¹). Application of CIM discs as solid supports in immunocolumns for allergen detection enables fast and sensitive screening of allergens in foodstuffs with sample throughput of up to eight samples per hour.     

Dispersive liquid-liquid microextraction based on the solidification of a floating organic droplet for simultaneous analysis of diethofencarb and pyrimethanil in apple pulp and peel

A method for analysis of diethofencarb and pyrimethanil in apple pulp and peel was developed by using dispersive liquid–liquid microextraction based on solidification of a floating organic droplet (DLLME-SFO) and high-performance liquid chromatography with diode-array detection (HPLC–DAD). Acetonitrile was used as the solvent to extract the two fungicides from apple pulp and peel, assisted by microwave irradiation. When the extraction process was finished, the target analytes in the extraction solvent were rapidly transferred from the acetonitrile extract to another extraction solvent (1-undecanol) by using DLLME-SFO. Because of the lower density of 1-undecanol than that of water, the finely dispersed droplets of 1-undecanol collected on the top of aqueous sample and solidified at low temperature. Meanwhile, the tiny particles of apple cooled and precipitated. Recovery was tested for a concentration of 8 μg kg⁻¹. Recovery of diethofencarb and pyrimethanil from apple pulp and peel was in the range 83.5–101.3%. The repeatability of the method, expressed as relative standard deviation, varied between 4.8 and 8.3% (n = 6). Detection limits of the method for apple pulp and peel varied from 1.2–1.6 μg kg⁻¹ for the two fungicides. Compared with conventional sample preparation, the method has the advantage of rapid speed and simple operation, and has high enrichment factors and low consumption of organic solvent.     

Sampling of benzene in environmental and exhaled air by solid-phase microextraction and analysis by gas chromatography–mass spectrometry

Benzene is classified as a Group I carcinogen by the International Agency for Research on Cancer (IARC). The risk assessment for benzene can be performed by monitoring environmental and occupational air, as well as biological monitoring through biomarkers. The present work developed and validated methods for benzene analysis by GC/MS using SPME as the sampling technique for ambient air and breath. The results of the analysis of air in parks and avenues demonstrated a significant difference, with average values of 4.05 and 18.26 μg m⁻³, respectively, for benzene. Sampling of air in the occupational environment furnished an average of 3.41 and 39.81 μg m⁻³. Moreover, the correlations between ambient air and expired air showed a significant tendency to linearity (R ² = 0.850 and R ² = 0.879). The results obtained for two groups of employees (31.91 and 72.62 μg m⁻³) presented the same trend as that from the analysis of environmental air.