Determination of nonylphenol ethoxylates in the aquatic environment by normal phase liquid chromatography-electrospray mass spectrometry

A comprehensive analytical method based on normal-phase liquid chromatography–electrospray ionization mass spectrometry (NPLC–ESI-MS) has been established for determination of nonylphenol ethoxylates (NPEOs) in the aquatic environment. Extraction and cleanup of samples were performed on graphitized carbon black (GCB) solid-phase extraction cartridges. Complete separation between each individual NPEOs was achieved by combining a C₁₈ pre-column with a silica analytical column and using acetonitrile–water as eluent. Quantitative determination by LC–ESI-MS was achieved in the positive ionization (PI) mode at a ramped cone voltage for NPEOs using selected ion monitoring. Recoveries for NPEOs ranged between 91.9 and 117.5%, and the limits of detection varied between 0.5 and 2 ng/l for individual NPEOs with n longer than 2, and between 5 and 0.5 μg/l for NP1EO and NP2EO. This method was successfully applied to the investigation of residual NPEOs with n>2 in the Chongqing area of the Changjiang river. NPEOs with n ranging from 1 to 22 were found to vary between 0.1 and 2900 ng/l with a distribution depending on the depth of water.     

Simultaneous sonication-assisted extraction, and determination by gas chromatography-mass spectrometry, of di-(2-ethylhexyl)phthalate, nonylphenol, nonylphenol ethoxylates and polychlorinated biphenyls in sludge from wastewater treatment plants

Di-(2-ethyl-hexyl)phthalate (DEHP), nonylphenol, nonylphenol mono- and diethoxylates (NPEs) and polychlorinated biphenyls (PCBs) are organic pollutants in sewage sludge which have to be monitored in the European Union according to a future Sludge Directive. In the present work, an analytical method for the simultaneous extraction and determination of DEHP, NPEs and PCBs is proposed for the routine analysis of these compounds in sludge from wastewater treatment plants. All the compounds were simultaneously extracted by sonication with hexane and analysed by gas chromatography-mass spectrometry (GC-MS) in electronic impact mode. Recoveries achieved were 105% for DEHP, 61.4-88.6% for NPEs and 55.8-108.3% for PCBs with relative standard deviation bellow 10%. Limits of quantification were 65 μg kg⁻¹ for DEHP, from 630 to 2504 μg kg⁻¹ for NPEs and from 5.4 to 10.6 μg kg⁻¹ for PCBs in dried sludge. The applicability of the proposed method was evaluated by the determination of these compounds in sludge from wastewater treatment plants in Seville (South Spain).     

Validation of an enzyme-linked immunosorbent assay (ELISA) for the determination of 4-nonylphenol and octylphenol in surface water samples by LC-ESI-MS

4-Nonylphenol (NP) and octylphenol (OP) were measured by direct ELISA in both laboratory-fortified and surface water samples collected monthly from 10 rivers. In this procedure, samples were concentrated by solid phase extraction (SPE) using Lichrolut RP-18 sorbent with good recoveries obtained for both LC-MS and ELISA, giving a low level of detection (LOD) at the range of low μg L⁻¹ and good reproducibility. Analysis of 40 surface water samples demonstrated that the ELISA may be a useful screening tool for the determination of the alkylphenols in surface water matrices. The concentration of NP and OP in surface waters ranged from 0.11 to 6.58 μg L⁻¹. A good correlation of results obtained by ELISA and LC-MS within the concentration range of 0.08-6.86 μg L⁻¹ was found in the river samples [R² = 0.924, n = 39]. The influence of various factors on assay determination was also discussed.     

Quantification of 5′-monophosphate cytosine arabinoside (Ara-CMP) in cell extracts using liquid chromatography-electrospray mass spectrometry

A LC/MS/ESI method is described to monitor 5′-monophosphate cytosine arabinoside (Ara-CMP) formed after incubation of an Ara-C prodrug in cell extracts. After addition of an internal standard and off-line treatment with perchloric acid, Ara-CMP is separated from Ara-C and endogeneous matrix components on a porous graphitic carbon stationary phase using gradient elution with a water/acetonitrile mobile phase containing formic acid. The results of method performances (accuracy, linearity, limit of quantification and repeatability) show that the method is acceptable for its intended use.     

Separation and determination of homogenous fatty alcohol ethoxylates by liquid chromatography with mulitstage mass spectrometry

Alcohol ethoxylates (AEs) are a significant component of a stream of surfactants directed to the aquatic environment. The aim of this work was the investigation of the dependence of the analytical signals of homogeneous AE homologues on liquid chromatography with tandem mass spectrometry conditions, as well as the separation of AEs from the water matrix and, on this basis, the development of an analytical procedure suitable for the determination of AEs in environmental samples. Homogeneous homologues containing dodecyl moiety and 2–9 oxyethylene subunits were investigated. The analytical signals of the investigated homologues were optimized in terms of concentration of ammonium acetate in the mobile phase (optimum 5 mM) and a column temperature (optimum 35°C) of the liquid chromatography with tandem mass spectrometry system. A separation of AEs from the water matrix by liquid–liquid extraction (ethyl acetate, chloroform) or solid‐phase extraction (C₁₈, styrene divinylbenzene, H‐RX) was investigated. In a model investigation, the best recoveries (>90%) were obtained with a styrene divinylbenzene cartridge eluted with a 1:1 mixture of chloroform and methanol. However, much worse recoveries were obtained from the river water sample. Better results were obtained for liquid–liquid extraction with ethyl acetate. Recoveries of 62–80% were obtained for homologues having 4–9 oxyethylene subunits, at the lowest spike.     

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.     

Simultaneous analysis of fourteen tertiary amine stimulants in human urine for doping control purposes by liquid chromatography-tandem mass spectrometry and gas chromatography-mass spectrometry

A method for the simultaneous screening and confirmation of the presence of fourteen tertiary amine stimulants in human urine by gas chromatography-mass spectrometry (GC-MS) in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been developed and validated. Solid phase extraction (SPE) and liquid-liquid extraction (LLE) approaches were utilized for the pre-treatment of the urine samples. The study indicated that the capillary temperature played a significant role in the signal abundances of the protonated molecules of cropropamide and crotethamide under positive ion electrospray ionization (ESI) conditions. In addition, comparison studies of two different pre-treatment approaches as well as the two ionization modes were conducted. The LODs of the developed method for all the analytes were lower than the minimum required performance limit (MRPL) as set forth in the World Anti-Doping Agency (WADA) technical document for laboratories. The human urine sample obtained after oral administration of prolintane·HCl was successfully analyzed by the developed method, which demonstrated the applicability and reliability of the method for routine doping control analysis.     

A rapid and environmental friendly determination of the dithiocarbamate metabolites ethylenethiourea and propylenethiourea in fruit and vegetables by ultra high performance liquid chromatography tandem mass spectrometry

Previous published methods for the analysis of ETU and PTU are time-consuming and furthermore use dichloromethane (DCM) for extraction or clean-up. This study details the development and validation of a rapid method that combines a simple extraction step with UHPLC-ESI(+)-MS/MS. This is the first application of UHPLC-MS/MS to analyse these compounds. Besides that, we replaced DCM with a more environmental-friendly solvent. The analytical performance was evaluated with the analysis of spiked celery samples at 50 μg kg(-1) (LOQ) and 300 μg kg(-1). The recoveries were between 65% and 90% for ETU and between 71% and 127% for PTU with RSDs in repeatability and reproducibility conditions below 10% for ETU. This method is rapid (a chromatographic run time of 2 min) and can easily be performed (no laborious clean-up). The presented method is environmental friendly with significant reduction in solvent consumption.     

Presence of nonylphenol, octyphenol and bisphenol a in two aquifers close to agricultural, industrial and urban areas

Active endocrine disruptors (nonylphenol, octylphenol and bisphenol A) were analysed in 2 aquifers and the corresponding surface waters. They are compounds widely used in industrial processes. The objective of this study was to determine the leaching potential of these compounds in groundwaters and to eventually correlate these levels with surface water samples. The areas sampled were agricultural, close to large cities and with an important industrial activity in the surrounding area. Samples (200 mL) were extracted using off-line SPE with polymeric OASIS 60 mg cartridges. Analyses were performed by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring (SIM) and full scan for quantification and unequivocal identification, respectively. This paper reports the detection limit for the compounds studied (from 0.001 to 0.030 μg L⁻¹), and method performance as regards to linearity (0.01–1.3 μg L⁻¹), reproducibility (less than 9%) and recovery (84 to 95%). The results from a monitoring program revealed the presence of the target compounds in all samples analysed, at levels of 0.07 and 1.9 μg L⁻¹. The presence of these compounds in groundwater was attributed basically to degradation of inert ingredients present in the formulation of many pesticides or to the increasing application of sludge in agricultural practice, although the infiltration of industrial run-off and wastewater disposal cannot be disregarded.     

Chemical monitoring and occurrence of alkylphenols, alkylphenol ethoxylates, alcohol ethoxylates, phthalates and benzothiazoles in sewage treatment plants and receiving waters along the Ter River basin (Catalonia, N. E. Spain)

This study presents a quantitative estimation of the analysis and fate of several emerging pollutants, some of them endocrine-disrupting compounds, in surface water samples collected at several locations along the Ter River and two of its tributaries. Influent and effluent waters and particulate matter from five sewage treatment plants (STP) that discharge into these rivers were also studied. The target compounds analyzed were: nonylphenol ethoxylates (NPEO), nonylphenol (NP), octylphenol (OP), bisphenol A (BPA), phthalates, alcohol ethoxylates (AEO) and benzothiazoles. Chemical analysis by liquid chromatography–mass spectrometry using an electrospray interface (LC–ESI–MS) revealed the presence of low amounts (between 0.06 and 17.5 μg L⁻¹) of the target compounds NPE₁₊₂O and NP, which were detected in 100% and 84% of the samples respectively. Maximum concentrations occurred in the STPs associated with the municipalities of Vic and Girona. From the fate and behavior data obtained for the various compounds analyzed in the STP influent and effluent, we can conclude that the STPs are effective at removing large amounts (more than 70%) of the compounds studied from the water.      

Simultaneous determination of imidacloprid, carbendazim, methiocarb and hexythiazox in peaches and nectarines by liquid chromatography-mass spectrometry

A liquid chromatographic (LC) atmospheric pressure chemical ionization mass spectrometric (MS) method is described for determining imidacloprid, carbendazim, methiocarb and hexythiazox in peaches and nectarines. The samples were extracted with ethyl acetate and anhydrous sodium sulfate. Recoveries yield for spiking samples were ranged from 64±9% (R.S.D) for carbendazim to 108±14% (R.S.D.) for hexythiazox at the concentration of 0.1 mg kg⁻¹ (n=5). The correlation coefficients were greater than 0.998 over the range between 0.02 and 2 mg kg⁻¹. The limits of quantitation (LOQ) were 0.02 mg kg⁻¹ for all the pesticides. The applicability of the method to detect and quantify imidacloprid, carbendazim, methiocarb and hexythiazox was demonstrated successfully in 159 peach and nectarine samples obtained from an agricultural co-operative. The calculation of the estimated daily intakes (EDI) from these data showed that contribution of fruits to dietary intakes were much lower than acceptable daily intakes (ADI) proposed by international agencies.     

Simultaneous determination of bisphenols,benzophenones and parabens in human urine by using UHPLC-TQMS

An analytical method for the simultaneous determination of six bisphenols, five benzophenones and seven parabens by using ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry was developed and applied for human urine sample analysis.     

Identification of phenyl-N-methylcarbamates and their transformation products in Tunisian surface water by solid-phase extraction liquid chromatography-tandem mass spectrometry

Liquid chromatography-pneumatically assisted electrospray mass spectrometry with both negative and positive ionization has been used for the determination of carbamates pesticides and their transformation products in Tunisian surface water. Eight pesticides and four of their hydrolysis products were covered in this study.Optimization of electrospray inlet conditions is described as well as results from investigations of the linearity of the detector response. Conditions for tandem mass spectrometry (MS-MS) detection of characteristic daughter ions formed by collision induced dissociation (CID) of the parent ion are described. Detection limits using MS in the selected ion monitoring (SIM) mode were generally in the order of 0.5 μg L⁻¹ or below. A principle of analysis is proposed based on triple quadrupole MS as a method for quantitative determination followed by verification of positive findings by CID-MS-MS. Application of the method for detecting carbamates residues in surface water is demonstrated.     

Identification of anthocyanins in wines by liquid chromatography, liquid chromatography-mass spectrometry and nuclear magnetic resonance

Liquid chromatography (LC) was used for the fractionation of particular anthocyanins in glycoside form from methanol extracts of red grape skins and solid phase extracts of red wine. By the combination of nuclear magnetic resonance spectroscopy and LC-mass spectroscopy the identification of 13 anthocyanins in a particular LC fraction and hence the in particular peaks in chromatograms were obtained. Peaks areas in the chromatograms obtained under the semi-quantitaive conditions of the solid phase extracts of red wines Pinot Noir, Cabernet Sauvignon and Merlot from the Coastal wine-growing region in Slovenia, produced in 1999, were used as input data in chemometric analysis. The chemometric methods used were hierarchical clustering analysis and regularised discriminant analysis. The results of both methods give 100% correct classification of wines regarding the vine variety.     

Development of a quantitative high-performance thin-layer chromatographic method for sucralose in sewage effluent, surface water, and drinking water

Sucralose, a persistent chlorinated substance used as sweetener, can already be found in waste water, and various countries focused on the release of sucralose into the aquatic environment. A quantitative high-performance thin-layer chromatography (HPTLC) method, which is orthogonal to existing methods, was developed to analyze sucralose in water. After sample preparation, separation of up to 17 samples was performed in parallel on a HPTLC plate silica gel 60 F(254) with a mixture of isopropyl acetate, methanol and water (15:3:1, v/v/v) within 15 min. Due to the weak native UV absorption of sucralose (≤200 nm), various post-chromatographic derivatization reactions were compared to selectively detect sucralose in effluent and surface water matrices. Thereby p-aminobenzoic acid reagent was discovered as a new derivatization reagent for sucralose. Compared to the latter and to β-naphthol, derivatization with aniline diphenylamine o-phosphoric acid reagent was slightly preferred and densitometry was performed by absorbance measurement at 400 nm. The limit of quantification (LOQ) of sucralose in drinking and surface water was calculated to be 100 ng/L for a given recovery rate of 80% and the extraction of a 0.5 L water sample. The sucralose content determined in four water samples obtained during an interlaboratory trial in 2008 was in good agreement to the mean laboratory values of that trial. According to the t-test, which compares the results with the target value, the means obtained by HPTLC were not significantly different from the respective means of six laboratories, analyzed by HPLC-MS/MS or HPLC-TOF-MS with the use of mostly isotopically labeled standards. The good accuracy and high sample throughput capacity proved HPTLC as a well suited method regarding quantification of sucralose in various aqueous matrices.     

Determination of pesticides and veterinary drug residues in food by liquid chromatography-mass spectrometry: A review

Monitoring of pesticides and veterinary drug residues is required to enforce legislation and guarantee food safety. Liquid chromatography-mass spectrometry (LC-MS) is the prevailing technique for assessing both types of residues because LC offers a versatile and universal separation mechanism suitable for non-gas chromatography (GC) amenable and the majority of GC-amenable compounds. This characteristic becomes more relevant when LC is coupled to MS because the high sensitivity and specificity of the detector allows to apply generic sample preparation procedures, which simultaneously extract a wide variety of residues with different physico-chemical properties. Determination of metabolites and degradation products, non-target suspected screening of an increasing number of residues, and even unknowns identification are also becoming inherent LC-MS advantages thanks to the latest advances. For routine analysis and, in particular, for official surveillance purposes in food control, analytical methods properly validated following strict guidelines are needed. After a brief introduction and an outline of the legislation applicable around the world, aspects such as improvement of specificity of high-throughput methods, resolution and mass accuracy of identification strategies and quantitative accuracy are critically reviewed in this article. In them, extraction, separation and determination are emphasized. The main objective is to offer an assessment of the state of the art and identify research needs and future trends in determining pesticide and veterinary drug residues in food by LC-MS.     

Benzylic rearrangement stable isotope labeling for quantitation of guanidino and ureido compounds in thyroid tissues by liquid chromatography-electrospray ionization mass spectrometry

Benzylic rearrangement stable isotope labeling (BRSIL) was explored to quantify the guanidino and ureido compounds (GCs and UCs). This method employed a common reagent, benzil, to label the guanidino and ureido groups through nucleophilic attacking then benzylic migrating. The use of BRSIL was investigated in the analysis of five GCs (creatine, l-arginine, homoarginine, 4-guanidinobutyric acid, and methylguanidine) and two UCs (urea and citrulline). The labeling was found simple and specific. The introduction of bi-phenyl group and the generation of nitrogen heterocyclic ring in the benzil-d0/d5 labeled GCs and UCs improved the retention behaviors in liquid chromatography (LC) and increased the sensitivity of electrospray ionization mass spectrometry (ESI MS) detection. The fragment ion pairs of m/z 182/187 and m/z 210/215 from the benzil-d0/d5 tags facilitated the discovery of potential GCs and UCs candidates residing in biological matrices. The use of BRSIL combined with LC-ESI MS was applied for simultaneously quantitation of GCs and UCs in thyroid tissues. It was demonstrated that nine GCs and UCs were detected, six of which were further quantified based on corresponding standards. It was concluded that five GCs and UCs (l-arginine, homoarginine, 4-guanidinobutyric acid, methylguanidine, and citrulline) were statistically significantly different (p < 0.05) between the para-carcinoma and carcinoma thyroid tissue samples.     

Rapid and sensitive liquid chromatography-tandem mass spectrometry for the quantitation of epirubicin and identification of metabolites in biological samples

A highly sensitive and selective liquid chromatography-mass spectrometry (LC-MS) method has been developed for the determination of epirubicin in serum and cell specimens using daunorubicin as an internal standard. Using atmospheric pressure chemical ionisation (APCI), the epirubicin metabolites were readily distinguishable by their fragmentation pattern in the mass spectrometer. Selected reaction monitoring (SRM) mode was employed for quantitation of epirubicin and the metabolites. Following extraction, chromatography was performed on a C18 column with a mobile phase consisting of water-acetonitrile-formic acid, pH 3.2, with a flow rate of 200 μl/min. The limit of detection (LOD) and the limit of quantitation (LOQ) of this method in serum were determined to be 1.0 and 2.5 ng/ml, respectively. Linearity of the method was verified over the concentration range of 2.5-2000 ng/ml, with a high correlation coefficient (R² ≥ 0.998). For the extraction procedure, an aliquot of 500 μl serum, spiked with internal standard, was extracted using a chloroform-2-isopropanol (2:1, v/v) mixture. The method has been applied to the analysis of epirubicin in cancer cell samples and the identification of known and unknown metabolites in clinical trial patient serum samples.     

Subcellular localization of docosahexaenoic acid and arachidonic acid ω-hydroxylation activity in the brain, liver and colonic adenocarcinoma

A homogenate of rat brain, rat liver or human colonic well differentiated adenocarcinoma was prepared in 250 mM sucrose isoosmolaric buffer (pH 7.6) and fractionated by differential centrifugation at 10³, 10⁴ and 10⁵ g. Each precipitate or supernatant was incubated with NADPH and docosahexaenoic acid or arachidonic acid as a substrate for 30 min at 37°C under aerobic conditions. ω-Hydroxydocosahexaenoic acid or ω-hydroxyeicosatetraenoic acid from an incubation mixture was detected by reversed-phase high-performance liquid chromatography-thermospray mass spectrometry with selected-ion monitoring. ω-Hydroxy polyunsaturated fatty acids were characterized by high intensity of the molecular ion (MH⁺) although common hydroxy polyunsaturated fatty acids were characterized by high intensity of the MH⁺ - H₂O ion. For the rat brain, ω-hydroxylation activity (the amount of ω-hydroxy product produced in 30 min) was concentrated to a 103 g precipitate although the specific activity (the activity per 1 mg of protein) in the 10³ g precipitate did not indicate superiority over other fractions. However, the specific activity of the rat brain increased on addition of a 10⁴ or 10⁵ g precipitate. For the rat liver, although ω-hydroxylation activity was concentrated to a 10³ g precipitate, the specific activity was concentrated to a 10⁵ g precipitate and the Subcellular localization differed from that of rat brain. In the human colonic well differentiated adenocarcinoma, although ω-hydroxylation activity was relatively high in the 10³ g supernatant, the specific activity was relatively high in the 10³ and 10⁵ g precipitates. These results suggest that there is a difference regarding subcellular localization of the ω-hydroxylation activity depending on the species of the organs.     

On-line LC-GC-AED and LC-GC-MS – multidimensional methods for the analysis of PAC in fuels,combustion emissions and atmospheric samples

The composition and concentration of polycyclic aromatic compounds (PAC) in fuels. Theier combustion products and in the atmosphere remains a topic of considerable interest. Despite the wealth of literature on the identification of PAC, speciation at low concentrations remains difficult due to instrument limitation and the complexity of fuel and environmental samples. Consequently on line sample preparation procedures (SPE, SFE, LC, etc.) are becomeing an increasingly important step in the analysis procedure particularly where sample clean-up and fractionation are essential for improving analytical resolution. In this study a normal phase high pressure analytical resolution. In this study a normal phase high pressure liquid chromatography-gas chromatography (LC-GC) system has been developed to provide quantitative analysis of samples, as diverse as coal liquids, petroleum fuels, diesel exhaust particulates, and urban air particulates. Separation and identification of parent and alkylated PAH, hetercycline nitro-and oxy-PAC can be achieved by direct coupling to an atomic emission detector and a bech top mass spectrometer. For both systems the primary LC separation combined with the large sample volume transferred to GC vastly improves detection limits. Furthermore the complimentary nature of the two detectors used enables the positive indentification of many unknowns.