A Survey of the Presence of Pharmaceutical Residues in Wastewaters. Evaluation of Their Removal using Conventional and Natural Treatment Procedures.

To encourage the reutilization of treated wastewaters as an adaptation strategy to climate change it is necessary to demonstrate their quality. If this is ensured, reclaimed waters could be a valuable resource that produces very little environmental impact and risks to human health. However, wastewaters are one of the main sources of emerging pollutants that are discharged in the environment. For this, it is essential to assess the presence of these pollutants, especially pharmaceutical compounds, in treated wastewaters. Moreover, the different treatment processes must be evaluated in order to know if conventional and natural treatment technologies are efficient in the removal of these types of compounds. This is an important consideration if the treated wastewaters are used in agricultural activities. Owing to the complexity of wastewater matrixes and the low concentrations of pharmaceutical residues in these types of samples, it is necessary to use sensitive analytical methodologies. In this study, the presence of 11 pharmaceutical compounds were assessed in three different wastewater treatment plants (WWTPs) in Gran Canaria (Spain). Two of these WWTPs use conventional purification technologies and they are located in densely populated areas, while the other studied WWTP is based in constructed wetlands which purify the wastewaters of a rural area. The sampling was performed monthly for two years. A solid phase extraction (SPE) coupled to ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was applied for the analysis of the samples, and the 11 pharmaceuticals were detected in all the studied WWTPs. The concentrations were variable and ranged from ng·L-1 in some compounds like diclofenac or carbamazepine to µg·L-1 in common pharmaceutical compounds such as caffeine, naproxen or ibuprofen. In addition, removal efficiencies in both conventional and natural purification systems were evaluated. Similar removal efficiencies were obtained using different purifying treatments, especially for some pharmaceutical families as stimulants or anti-inflammatories. Other compounds like carbamazepine showed a recalcitrant behavior. Secondary treatments presented similar removal efficiencies in both conventional and natural wastewater treatment plants, but conventional treatments showed slightly higher elimination ratios. Regarding tertiary system, the treatment with highest removal efficiencies was reverse osmosis in comparison with microfiltration and electrodialysis reversal.

Simultaneous and systematic analysis of cytostatic drugs in wastewater samples by ultra-high performance liquid chromatography tandem mass spectrometry.

Cytostatic drugs are compounds used to fight cancer, which may be excreted after administration to patients, and eventually reach wastewater. Given the high incidence of cancer and the properties of drugs, the drug concentrations in water systems should be evaluated. We present the optimization, development and application of a solid phase extraction (SPE) method for the determination of eight cytostatic compounds of different classes in wastewater and seawater samples. We compared four SPE cartridges prior to their determination by Ultra-High Performance Liquid Chromatography tandem Mass Spectrometry. For wastewater samples, the Oasis HLB cartridge gave the best recoveries, which were higher than 65% in most cases, achieving limit of detections (LODs) of 1.68-103.95 ng·L-1. In seawater samples, the Bond Elut cartridge afforded the best recoveries >70%, with LODs of 0.95-5.14 ng·L-1. The optimal procedure was applied in four hospital wastewater effluent samples taken during one year, and in different influents and effluents from wastewater treatment plants and seawater from marine outfalls taken in eight campaigns during two years, in Gran Canaria island (Spain). Results showed that etoposide was present in influents of wastewater treatment plants in several months and different wastewater treatment plants and hospital effluents in the range 375.8-5141 ng·L-1, while cyclophosphamide was present in some months in effluents from only one wastewater treatment plant and hospital effluents in the range 55.94-1212 ng·L-1. Vinblastine and vincristine were detected in one sample of hospital at concentrations of 1836 ng·L-1 and 1851 ng·L-1, respectively.

Preconcentration of pharmaceuticals residues in sediment samples using microwave assisted micellar extraction coupled with solid phase extraction and their determination by HPLC-UV.

An analytical method combining microwave assisted micellar extraction (MAME) and solid phase extraction (SPE) has been developed to extract and preconcentrate a selected group of eight pharmaceutical compounds in sediment samples prior to their determination using liquid chromatography with an UV-DAD detector. A non-ionic surfactant, Polyoxyethylene 10 lauryl ether (POLE) was used for the MAME extraction and the different parameters for the optimization process were studied. Then, SPE was used to clean-up and preconcentrate the target analytes in the extract, prior to their determination using HPLC-UV. The method was applied to the determination of the selected pharmaceuticals compounds in several kinds of sediment samples with different characteristics. Relative recoveries for spiked sediment samples were over 70% and relative standard deviations (RSDs) were under 11% for all recoveries tested. Detection limits between 4 and 167ngg(-1) were obtained. The method was validated using Soxhlet extraction procedure.

Implementation of solid-phase microextraction with micellar desorption method for priority phenolic compound determination in natural waters.

Eleven phenolic compounds considered by the Environmental Protection Agency to be priority pollutants are extracted and determined in different water samples. The method involves the extraction and clean-up step of target compounds by solid-phase microextraction and micellar desorption (SPME-MD) and a second step of determination by liquid chromatography with diode array detection. Different fibers and surfactants are evaluated for the analysis of these target analytes in water samples. In the optimum conditions for the SPME process, recoveries for the target compounds are between 80% and 109%; relative standard deviations are lower than 10%, and detection limits are in the range 0.3-3.5 ng/mL. The main advantages of this method are the combination of time and efficiency, safety, and an environmentally friendly process for sample extraction prior to instrumental determination. This demonstrates that SPME-MD can be used as an alternative to traditional methods for the extraction and determination of priority phenolic compounds in natural waters from different origins.

Occurrence and environmental impact of pharmaceutical residues from conventional and natural wastewater treatment plants in Gran Canaria (Spain).

The presence and fate of pharmaceutical residues in environmental samples are of great interest. There is a vast number of studies published regarding their input, presence, effects and risks in ecosystems. Moreover, it has been demonstrated that the primary source of input of these contaminants in the environment is from Wastewater Treatment Plants (WWTPs). It is therefore essential to evaluate the efficiency of commonly used treatments and the necessity of applying novel purification processes in order to eliminate or reduce the concentration of pharmaceuticals from wastewater or from the effluent of WWTPs. The aim of this work was to quantify twenty-three pharmaceutical compounds in the aqueous phase at different stages of a conventional and a natural WWTP situated in Gran Canaria (Spain). The results indicate concentration levels in the range of 0.004±0.001 to 59.2±11.7µgL-1 and 0.018±0.001 to 148±14.7µgL-1 from conventional and natural WWTPs, respectively. Better efficiency was, however, offered by the conventional WWTP with a removal median of 99.7%. In addition, the impact on different aquatic organisms (algae, daphnids and fish) was assessed in terms of risk quotients. The results reveal a possible highly harmful effect towards organisms by gemfibrozil, ibuprofen and ofloxacin.

Simplified solid-phase extraction procedure combined with liquid chromatography tandem-mass spectrometry for multiresidue assessment of pharmaceutical compounds in environmental liquid samples.

To follow the twelve "green analytical chemistry" (GAC) principles, it is necessary to continuously develop analytical extraction and determination methodologies to assess the presence of micropollutants, such as pharmaceuticals, in environmental samples. A reduction in the analysis time and solvent quantity, which is one of the GAC principles, has been achieved through a simplified solid-phase extraction (SPE) procedure combined with high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the determination of twenty-three pharmaceuticals in liquid environmental samples using N-vinylpyrrolidone-divinylbenzene copolymer (OASIS HLB) cartridges. The optimal SPE conditions were studied. In these optimized conditions, 82.6% of the data have a median recovery above 70% for all compounds in each sample. The relative standard deviations (RSDs) were below 14.4% and 22.0% for intra- and inter-day repeatability, respectively. Method detection limits (MDLs) and method quantification limits (MQLs) ranged from 0.011 to 188ngL-1 and from 0.033 to 628ngL-1, respectively. The applicability of the method was evaluated in real samples from natural and conventional wastewater treatment plants (WWTPs), and results were obtained in concentration ranges from 0.013 to 91.5µgL-1 and from 0.004 to 49.1µgL-1, respectively.

Sample extraction method combining micellar extraction-SPME and HPLC for the determination of organochlorine pesticides in agricultural soils.

A method for the determination of organochlorine pesticides in soil samples combining microwave assisted micellar extraction (MAME) with solid-phase microextraction (SPME) and high-performance liquid chromatography-UV has been developed. A mixture of two nonionic surfactants (polyoxyethylene 10 lauryl ether and polyoxyethylene 10 stearyl ether) was used for the extraction of pesticides from agricultural soils, and different types of SPME fibers were compared. The different parameters which affect extraction efficiency in the SPME procedure were optimized such as extraction time and temperature. The method developed involves extraction and preconcentration for the target analytes in soil samples. The analytical parameters were also studied and good recoveries obtained, RSD being lower than 10% and detection limits ranging between 36 and 164 ng g(-1) for the pesticides studied. The proposed method was successfully applied to the determination of some organochlorine pesticides in several kinds of agricultural soil samples with different characteristics.

Combination of microwave assisted micellar extraction and liquid chromatography for the determination of organophosphorous pesticides in soil samples.

A new methodology based on the microwave assisted micellar extraction (MAME) technique has been optimised, using soil samples, to extract and determine a mixture of the eight organophosphorous pesticides mainly used in agriculture. The pesticides under study have been extracted using the non-ionic surfactants polyoxyethylene 10 lauryl ether (POLE) and oligoethylene glycol monoalkyl ether (Genapol X-080). The optimal extraction variables, such as surfactant concentration, pH, radiation time and microwave power were determined for each surfactant. The results show the advantage of using POLE instead of Genapol X-080 for the extraction of the organophosphorus pesticides with recoveries higher than 70% for most of the compounds and relative standard deviations (RSD) below 2.6%. This method was successfully applied to fresh samples as well as to aged samples for the analysis of soils with different characteristics and compared with the traditional Soxhlet technique.

Evaluation of the presence of endocrine-disrupting compounds in dissolved and solid wastewater treatment plant samples of Gran Canaria Island (Spain).

Liquid and solid samples from two wastewater treatment plants (WWTPs) on Gran Canaria Island (Spain) have been tested for the presence of compounds with endocrine-disrupting properties. The selected degradation stages were sampled bimonthly from each WWTP over the 12-month period from July 2010 to July 2011. The analytical methods used for the determination of the endocrine-disrupting compounds (EDCs) were based on on-line solid phase extraction, microwave-assisted extraction (MAE), and ultrasonic-assisted extraction (UAE) coupled to UHPLC-MS/MS. All of the hyphenated methodologies employed in this work showed good recoveries (72-104%) and sensitivities, with LODs lower than 7.0 ng L(-1) and 6.3 ng g(-1) for the dissolved and solid fractions, respectively. We have also evaluated the estrogenicity of the samples in terms of their estradiol equivalent concentrations (EEQs). The chemical analysis of the selected EDCs revealed fairly low concentrations for both natural and synthetic oestrogens, alkylphenolic compounds, and bisphenol-A in each of the dissolved, particulate, and sludge samples (ng L(-1) or ng g(-1)). However, the estimated estrogenic activity indicated that the majority of samples could represent an important environmental risk, clearly surpassing the threshold to exert deleterious consequences on living beings.

Development and optimisation of an on-line solid phase extraction coupled to ultra-high-performance liquid chromatography-tandem mass spectrometry methodology for the simultaneous determination of endocrine disrupting compounds in wastewater samples.

A process using a fully automated on-line solid-phase extraction system combined with ultra-high-performance liquid chromatography and tandem mass spectrometry detection (SPE-UHPLC-MS/MS) has been developed and applied to the determination of 27 endocrine disrupting compounds (EDCs) in sewage samples. This technology allows separation and detection of all substances in a single chromatographic run that requires less than 4 min. In order to obtain a more sensitive method for the quantification of these compounds in sewage samples, an on-line solid-phase extraction step with Oasis HLB columns was performed prior to chromatographic determination. The complete analysis of each sample by this process requires only 9 min, provides satisfactory recoveries (72-110%) and limits of detection on the order of a few nanograms per liter (0.3-2.1 ng L(-1)), demonstrating their potential for analyses of environmental samples. Thus, this methodology has been applied to samples collected from two wastewater treatment plants (WWTPs) located in Las Palmas de Gran Canaria (Canary Islands, Spain). One of these plants utilised conventional activated sludge treatment (CAS), while the other employed biomembrane reactor treatment (MBR).

Determination of various estradiol mimicking-compounds in sewage sludge by the combination of microwave-assisted extraction and LC-MS/MS.

In this work, we present the development and application of a microwave assisted extraction followed by liquid chromatography-tandem mass spectrometry methodology (MAE-LC-MS/MS) for the determination of various estradiol-mimicking compounds in sewage sludge samples. For the purification of the MAE extracts, we have employed a solid phase extraction (SPE) clean-up procedure, previously optimised. The entire method provides recoveries between 71.7% and 103.1%, with relative standard deviation lower than 11.1% and limits of detection ranging from 0.6 to 3.5 ng g(-1). The developed method was applied to samples from three wastewater treatment plants (WWTPs) located in Las Palmas of Gran Canaria (Spain), two of which had a conventional activated sludge treatment (AST), whereas the third treatment plant had an advanced membrane bioreactor treatment (MBR). All of the analytes in the study, including (nonylphenol (NP), octylphenol (OP), and some of their ethoxylated chains AP(n)EOs (n ≤ 7), 17ß-estradiol (E2), estriol (E3), 17α-ethynylestradiol (EE) and bisphenol-A (BPA)), were found in almost all samples in concentrations ranging from 0.9 to 710.2 ng g(-1).

Determination of alkylphenol polyethoxylates, bisphenol-A, 17α-ethynylestradiol and 17ß-estradiol and its metabolites in sewage samples by SPE and LC/MS/MS.

Recently, many chemicals released into the environment have been shown to mimic endogenous hormones such as estradiol. It has been demonstrated that these compounds cause several adverse effects on wildlife and humans, such as the feminization of animal species, development of physical abnormalities and birth defects, and reproductive failure. In an effort to model the behaviour of some endocrine-disrupting chemicals (EDCs) and to establish the level of contamination in sewage samples, a quantitative method for the simultaneous determination of nonylphenol, octylphenol and corresponding ethoxylates (1-12), 17α-ethynylestradiol, bisphenol-A, and 17ß-estradiol and two of its metabolites have been developed. Identification and quantification were achieved by high performance liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS). Satisfactory detection limits (between 0.5-6 ng L(-1) in the dissolved phase and 1.4-12.7 ng g(-1) in the particulate phase) and analyte recoveries (between 60% and 108%) were achieved for target compounds. The optimised method was applied to the determination of EDCs in liquid sewage samples collected from a wastewater treatment plant (WWTP) in Las Palmas de Gran Canaria (Spain). Concentrations of EDCs ranged from <10 ng L(-1) to nearly 1200 ng L(-1) in the dissolved phase, and from 0.005 µg g(-1) to 2.8 µg g(-1) in the suspended particulate matter.

Microwave-assisted micellar extraction coupled with solid-phase extraction for preconcentration of pharmaceuticals in molluscs prior to determination by HPLC.

A simple and specific analytical method was developed and tested for the determination of pharmaceuticals in mollusc samples. A combination of microwave-assisted micellar extraction (MAME) and solid-phase extraction (SPE) using a non-ionic surfactant, polyoxyethylene 10 lauryl ether, was examined to extract and determine simultaneously a group of pharmaceuticals such as carbamazepine, clorfibric acid, ketoprofen, naproxen, bezafibrate and ibuprofen by liquid chromatography using UV-diode array detector. The MAME extraction performance was evaluated by studying various parameters such as the volume and concentration of surfactant and microwave conditions. Finally, an OASIS HLB cartridge was used as an optimum SPE sorbent to clean up the extracts and preconcentrate the selected analytes. The proposed method showed satisfactory linearity and reproducibility (between 3 and 15%), as well as detection limits ranging from 30 to 220 ng/g. Finally, the method was successfully applied to the determination of the target pharmaceuticals in various kinds of mollusc samples. This study has demonstrated that microwave-assisted micellar extraction with solid-phase extraction may be used as a viable alternative to conventional methods for the extraction of pharmaceuticals in this type of matrices.

Solid-phase microextraction of benzimidazole fungicides in environmental liquid samples and HPLC-fluorescence determination.

Solid-phase microextraction (SPME) coupled with high-performance liquid chromatography (HPLC) with fluorescence detection was optimized for extraction and determination of four benzimidazole fungicides (benomyl, carbendazim, thiabendazole, and fuberidazole) in water. We studied extraction and desorption conditions, for example fiber type, extraction time, ionic strength, extraction temperature, and desorption time to achieve the maximum efficiency in the extraction. Results indicate that SPME using a Carboxen-polydimethylsiloxane 75 microm (CAR-PDMS) fiber is suitable for extraction of these types of compound. Final analysis of benzimidazole fungicides was performed by HPLC with fluorescence detection. Recoveries ranged from 80.6 to 119.6 with RSDs below 9% and limits of detection between 0.03 and 1.30 ng mL-1 for the different analytes. The optimized procedure was applied successfully to the determination of benzimidazole fungicides mixtures in environmental water samples (sea, sewage, and ground water).

Environmental analysis based on luminescence in organized supramolecular systems.

The use of organized supramolecular systems-including micellar media and cyclodextrin inclusion complexes-combined with luminescence techniques in the study and determination of compounds and elements of environmental interest from 1990 to 2005 is reviewed. Analyses of environmental samples performed using fluorescence, photochemically induced fluorescence and phosphorescence spectroscopy as well as liquid chromatography, capillary electrophoresis and flow injection with luminescence detection in the presence of these organized media are described in detail.

Optimisation of solid-phase microextraction coupled to HPLC-UV for the determination of organochlorine pesticides and their metabolites in environmental liquid samples.

A solid-phase microextraction (SPME) procedure using two commercial fibers coupled with high-performance liquid chromatography (HPLC) is presented for the extraction and determination of organochlorine pesticides in water samples. We have evaluated the extraction efficiency of this kind of compound using two different fibers: 60-mum polydimethylsiloxane-divinylbenzene (PDMS-DVB) and Carbowax/TPR-100 (CW/TPR). Parameters involved in the extraction and desorption procedures (e.g. extraction time, ionic strength, extraction temperature, desorption and soaking time) were studied and optimized to achieve the maximum efficiency. Results indicate that both PDMS-DVB and CW/TPR fibers are suitable for the extraction of this type of compound, and a simple calibration curve method based on simple aqueous standards can be used. All the correlation coefficients were better than 0.9950, and the RSDs ranged from 7% to 13% for 60-mum PDMS-DVB fiber and from 3% to 10% for CW/TPR fiber. Optimized procedures were applied to the determination of a mixture of six organochlorine pesticides in environmental liquid samples (sea, sewage and ground waters), employing HPLC with UV-diode array detector.

Optimization of the separation selectivity of PCBs in a hydroorganic reverse-phase liquid chromatography in the presence of cetyltrimethylammonium bromide with fluorescence detection.

The retention of different PCBs of toxicological and environmental interest was studied in a hydro-alcoholic reverse-phase liquid chromatography in the presence of cetyltrimethylammonium bromide (CTAB). The influence of variables like type and percentage of alcohol, concentration of CTAB, temperature and mobile phase flow was determined. According to the aforementioned studies, the appropriate conditions for separation of PCBs in presence of CTAB in a hydro-alcoholic medium are: percentage of 1-Propanol 55% (v/v), concentration of CTAB 5.0 x 10(-3) M, temperature 50 +/- 1 degrees C and mobile phase flow 1 ml min-1.

Determination of free amino acids in microalgae by high-performance liquid chromatography using pre-column fluorescence derivatization.

An analytical method for a rapid reverse-phase liquid chromatography of amino acids is presented. The total analysis time was 19 min. The average relative deviation over the measured peak area was lower than 10% and the fluorescent response was linear with concentration for OPA derivatives (r > 0.99) in the range 100-1000 nM. The proposed method was applied to the determination of free amino acids in two microalgae species (Chlorella pyrenoidosa and Chlamydomonas rinhardtii). The principal aminoacids found were alanine, arginine leucine, serine and glutamic acid.