[Show abstract][Hide abstract] ABSTRACT: AimsTo perform wastewater analyses to assess spatial differences and temporal changes of illicit drug use in a large European population.DesignAnalyses of raw wastewater over a 1-week period in 2012 and 2013.Setting and ParticipantsCatchment areas of wastewater treatment plants (WWTPs) across Europe, as follows: 2012: 25 WWTPs in 11 countries (23 cities, total population 11.50 million); 2013: 47 WWTPs in 21 countries (42 cities, total population 24.74 million).MeasurementsExcretion products of five illicit drugs (cocaine, amphetamine, ecstasy, methamphetamine, cannabis) were quantified in wastewater samples using methods based on liquid chromatography coupled to mass spectrometry.FindingsSpatial differences were assessed and confirmed to vary greatly across European metropolitan areas. In general, results were in agreement with traditional surveillance data, where available. While temporal changes were substantial in individual cities and years (P ranging from insignificant to <10−3), overall means were relatively stable. The overall mean of methamphetamine was an exception (apparent decline in 2012), as it was influenced mainly by four cities.Conclusions
Wastewater analysis performed across Europe provides complementary evidence on illicit drug consumption and generally concurs with traditional surveillance data. Wastewater analysis can measure total illicit drug use more quickly and regularly than is the current norm for national surveys, and creates estimates where such data does not exist.
[Show abstract][Hide abstract] ABSTRACT: The s-triazine herbicide terbuthylazine (TERB) has been used as the main substitute of atrazine in many EU countries for more than 10 years. However, the ecological consequences of this substitution are still not fully understood. Since the fate of triazine herbicides is primarily dependent on microbial degradation, in this paper, we investigated the ability of a mixed bacterial culture, M3-T, originating from s-triazine-contaminated soil, to degrade TERB in liquid culture and soil microcosms. The M3-T culture grown in mineral medium with TERB as the N source and citrate as the C source degraded 50 mg L(-1) of TERB within 3 days of incubation. The culture was capable of degrading TERB as the sole C and N source, though at slower degradation kinetics. A thorough LC-MS analysis of the biodegradation media showed the formation of hydroxyterbuthylazine (TERB-OH) and N-t-butylammelide (TBA) as major metabolites, and desethylterbuthylazine (DET), hydroxydesethylterbuthylazine (DET-OH) and cyanuric acid (CA) as minor metabolites in the TERB degradation pathway. TBA was identified as a bottleneck in the catabolic pathway leading to its transient accumulation in culture media. The supplementation of glucose as the exogenous C source had no effect on TBA degradation, whereas citrate inhibited its disappearance. The addition of M3-T to sterile soil artificially contaminated with TERB at 3 mg kg(-1) of soil resulted in an accelerated TERB degradation with t 1/2 value being about 40 times shorter than that achieved by the native microbial community. Catabolic versatility of M3-T culture makes it a promising seed culture for accelerating biotransformation processes in s-triazine-contaminated environment.
Applied Microbiology and Biotechnology 05/2014; · 3.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Stability of the selected urinary biomarkers of six illicit drugs and two therapeutic opioids in municipal wastewater was studied in order to determine errors associated with their possible transformation in the sewer. The stability was assessed in experiments conducted at 10°C and 20°C in order to simulate typical winter and summer temperature conditions in the sewer system. Among fourteen substances tested, the most unstable compounds were morphine-3-β-D glucuronide (MG), 6-acetyl morphine (6-AM), cocaine (COC) and 6-acetyl codeine (6-AC), while all other investigated compounds appeared to be relatively stable over a period of 72h. The transformation of all degradable compounds followed pseudo-first order kinetics with significantly longer half-times (t1/2) at winter conditions. At 20°C, t1/2 of MG, 6-AM, COC and 6-AC was 7h, 87h, 35h and 58h, respectively, while the corresponding t1/2 values at 10°C were 18h, 139h, 173h and 87h. The main transformation mechanism of MG, 6-AM and 6-AC was most probably their enzymatic hydrolysis to morphine (MOR) and codeine (COD), while COC transformation to benzoylecgonine (BE) was primarily governed by chemical hydrolysis. The results indicate that the effect of the observed transformation of urinary biomarkers of COC and 6-AM on the estimates of COC and heroin consumption are relatively small (<10%) if the in-sewer hydraulic retention time is lower than 12h. Acidification of the wastewater samples proved to be the good way to stabilise the wastewater samples for the analysis of all selected compounds, except for 11-nor-9-carboxy-∆9-tetrahydrocannabinol (THC-COOH). This finding should be taken into account when selecting the preservation technique for multiresidual analyses of different groups of illicit drugs.
Science of The Total Environment 01/2014; · 3.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: P-glycoprotein (P-gp, ABCB1) is an important part of the multixenobiotic resistance (MXR) defense system in aquatic organisms. The main goal of this study was identification of P-gp inhibitors in contaminated sediments using the effect-directed analysis (EDA) approach. The samples were collected from the Gorjak creek (Zagreb, Croatia), a recipient of wastewater effluents from the pharmaceutical industry. Sediment samples were extracted and fractionated using two-tiered approach. Resulting non-polar, medium polar and polar fractions were tested on the inhibition of P-gp activity using P-gp overexpressing PLHC-1/dox cells and calcein-AM as model substrate. The obtained EC50 values - up to 757 μg/g, expressed in toxicity equivalents of model P-gp inhibitor cyclosporine A - revealed high inhibitory potential of polar fractions of investigated sediments and clearly reflected the impact of pharmaceutical wastewater. P-gp specific ATPase assay and the cytotoxicity modulation experiments with colchicine indicated that most of the observed P-gp inhibition was due to the presence of non-competitive inhibitors. A detailed chemical analysis by ultrahigh-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry (UPLC-QToFMS) revealed nonionic surfactants, including alcohol polyethoxylates (LAEOs) and polypropylene glycols (PPGs), as the major components of the most active sub-fractions. Testing of several LAEO and PPG commercial mixtures confirmed their potential to inhibit the fish P-glycoprotein and modulate toxicity of other xenobiotics present in complex environmental samples.
[Show abstract][Hide abstract] ABSTRACT: For the first time, an integrated procedure for a quantitative multiresidue analysis of dissolved and particulate illicit drug target residues was developed and validated in three different wastewater matrices. The procedure consists of a comprehensive sample enrichment, fractionation and cleanup followed by the determination of target analytes by triple quadrupole liquid chromatography-tandem mass spectrometry in both positive and negative ionisation polarities. The enrichment of illicit drugs from suspended solids and aqueous samples was performed using pressurised liquid extraction and solid phase extraction (SPE), respectively. The performance of different SPE cartridges was investigated in order to optimise the overall recovery and to reduce the matrix effects. The optimal results were obtained by combining mixed cation exchange (Oasis MCX) cartridges for fractionated enrichment, weak anion exchange for an additional extract cleanup and optimised chromatographic separation to minimise the impact from co-extracted interferences. The method was applied for the analysis of raw wastewater (RW), activated sludge (AS) and secondary effluent (SE) samples collected at four different wastewater treatment plants. The average contributions of the particulate drugs in the RW and AS were 1-28 and 23-65 %, respectively. This suggested that the total mass loads of some drugs might be underestimated by neglecting the particulate fraction. Moreover, relatively high distribution coefficients, determined for 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (RW = 1211 L/kg) and 11-hydroxy-Δ(9)-tetrahydrocannabinol (RW = 1,786 L/kg) implied that adsorption might play a significant role in their overall removal during wastewater treatment.
Analytical and Bioanalytical Chemistry 02/2013; · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Comprehensive study on the occurrence and fate of several classes of antimicrobials, including sulfonamides, trimethoprim, fluoroquinolones and macrolides, in Croatian municipal wastewaters was performed using an integrated approach, which comprised analysis of both dissolved and particulate fractions. A nation-wide screening showed ubiquitous occurrence of human-use antimicrobials in raw wastewater samples with the total concentrations ranging from 2 to 20 μg/L, while veterinary antimicrobials were typically present in much lower concentrations (<100 ng/L). The percentage of the particulate fraction in raw wastewater varied significantly depending on the type of the antimicrobial and the load of suspended solids. A detailed study of the mass flows of dissolved and particulate antimicrobials, performed in the wastewater treatment plant of the city of Zagreb, allowed an improved assessment of the biological and physico-chemical removal mechanisms of investigated compounds during the conventional activated sludge treatment. The overall removal efficiencies of antimicrobials from the water phase were rather variable, ranging from 0% for trimethoprim to 85% for norfloxacin. A significant percentage of fluoroquinolones (norfloxacin and ciprofloxacin) and macrolides (azithromycin and clarithromycin) was associated with the primary and excess secondary sludge, explaining 14-77% of the total removal. The removal, which could be attributed to biological transformation, was relatively poor for all antimicrobials, exceeding 30% only for SMX (32%) and clarithromycin (55%).
[Show abstract][Hide abstract] ABSTRACT: The analysis of sewage for urinary biomarkers of illicit drugs is a promising and complementary approach for estimating the use of these substances in the general population. For the first time, this approach was simultaneously applied in 19 European cities, making it possible to directly compare illicit drug loads in Europe over a 1-week period. An inter-laboratory comparison study was performed to evaluate the analytical performance of the participating laboratories. Raw 24-hour composite sewage samples were collected from 19 European cities during a single week in March 2011 and analyzed for the urinary biomarkers of cocaine, amphetamine, ecstasy, methamphetamine and cannabis using in-house optimized and validated analytical methods. The load of each substance used in each city was back-calculated from the measured concentrations. The data show distinct temporal and spatial patterns in drug use across Europe. Cocaine use was higher in Western and Central Europe and lower in Northern and Eastern Europe. The extrapolated total daily use of cocaine in Europe during the study period was equivalent to 356 kg/day. High per capita ecstasy loads were observed in Dutch cities, as well as in Antwerp and London. In general, cocaine and ecstasy loads were significantly elevated during the weekend compared to weekdays. Per-capita loads of methamphetamine were highest in Helsinki and Turku, Oslo and Budweis, while the per capita loads of cannabis were similar throughout Europe. This study shows that a standardized analysis for illicit drug urinary biomarkers in sewage can be applied to estimate and compare the use of these substances at local and international scales. This approach has the potential to deliver important information on drug markets (supply indicator).
Science of The Total Environment 07/2012; 432:432-9. · 3.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Region-specific contaminant prioritisation is an important prerequisite for sustainable and cost-effective monitoring due to the high number of different contaminants that may be present. Surface water and sediment samples from the Sava River, Croatia, were collected at four locations covering a 150-km-long river section characterised by well-defined pollution gradients. Analysis of contaminant profiles along the pollution gradients was performed by combining toxicity screening using a battery of small-scale or in vitro bioassays, which covered different modes of action, with detailed chemical characterisation based on gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/quadrupole time-of-flight mass spectrometry (LC-QTOF-MS). A large number of contaminants, belonging to different toxicant classes, were identified in both analysed matrices. Analyses of water samples showed that contaminants having polar character occurred in the highest concentrations, while in sediments, contributions from both non-polar and amphiphilic contaminants should be taken into account. Estimated contributions of individual contaminant classes to the overall toxicity indicated that, besides the classical pollutants, a number of emerging contaminants, including surfactants, pharmaceuticals, personal care products and plasticizers, should be taken into consideration in future monitoring activities. This work demonstrates the importance of the integrated chemical and bioanalytical approach for a systematic region-specific pollutant prioritisation. Finally, the results presented in this study confirm that hazard assessment in complex environmental matrices should be directed towards identification of key pollutants, rather than focusing on a priori selected contaminants alone.
Environmental Science and Pollution Research 07/2012; · 2.76 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Removal of numerous classes of pharmaceuticals from the municipal and industrial wastewater, using conventional wastewater treatment, is incomplete and several studies suggested that improvement of this situation would require the application of advanced treatment techniques. This is particularly important for the treatment of industrial effluents, released from pharmaceutical industries, which can contain rather high concentrations of antimicrobials. The aim of this work was to evaluate membrane bioreactors (MBRs), nanofiltration, reverse osmosis and ozonation, as well as their combinations, for the removal of antimicrobials from a synthetic wastewater which simulated highly contaminated industrial effluents. The study was performed using a mixture of four important classes of antimicrobials, including sulfonamides (SA), fluoroquinolones (FQ), macrolides (MAC) and trimethoprim (TMP). Performance of two different types of MBRs, Kubota and Zenon, was evaluated under different regimes regarding hydraulic retention time, total organic load and total nitrogen load. It was shown that elimination of SA in MBR treatment was very efficient, while the elimination of MAC, FQ, and TMP was incomplete. A mass balance of these contaminants in MBR suggested that microbial transformation represented the main mechanism, while only a small percentage was eliminated from the aqueous phase by adsorption onto sludge particles. Nanofiltration and reverse osmosis achieved high elimination rates however produced highly contaminated concentrate. High removal was achieved using ozonation, but further research is needed to characterize formed ozonation products.
Journal of hazardous materials 05/2011; 192(1):319-28. · 4.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ultrahigh-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry was applied for the identification of transformation products (TPs) of fluoroquinolone (norfloxacin and ciprofloxacin) and macrolide (azithromycin, erythromycin, and roxitromycin) antimicrobials in wastewater effluents from a Zenon hollow-fiber membrane bioreactor (MBR). The detected TPs were thoroughly characterized using the accurate mass feature for the determination of the tentative molecular formulae and MS-MS experiments for the structural elucidation of unknowns. Several novel TPs, which have not been previously reported in the literature, were identified. The TPs of azithromycin and roxithromycin, identified in MBR effluent, were conjugate compounds, which were formed by phosphorylation of desosamine moiety. Transformation of fluoroquinolones yielded two types of products: conjugates, formed by succinylation of the piperazine ring, and smaller metabolites, formed by an oxidative break-up of piperazine moiety to form the 7-[(2-carboxymethyl)amino] group. A semi-quantitative assessment of these TPs suggested that they might have contributed significantly to the overall balance of antimicrobial residues in MBR effluents and thus to the overall removal efficiency. Determination of TPs during a period of 2 months indicated a conspicuous dynamics, which warrants further research to identify microorganisms involved and treatment conditions leading to their formation.
Analytical and Bioanalytical Chemistry 05/2011; 401(1):353-63. · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The hazardous chemical contamination of untreated wastewater and secondary effluent from the wastewater treatment plant (WWTP) of the city of Zagreb, Croatia was comprehensively characterized using large-volume solid-phase extraction (SPE) and silica gel fractionation, followed by a detailed analysis of the resulting extracts by a combination of chemical and bioassay methods. Over 100 individual contaminants or closely related-contaminant groups were identified by high-resolution gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/quadrupole time-of-flight mass spectrometry (LC-QTOF). Ecotoxicity profiling of the investigated samples, including cytotoxicity, chronic toxicity and EROD activity; inhibition of the multixenobiotic resistance (MXR), genotoxicity and estrogenic potential, revealed the most significant contribution of toxic compounds to be present in polar fractions. Wastewater treatment using conventional activated sludge process reduced the initial toxicity of raw wastewater to various extents, ranging from 28% for algal toxicity to 73.2% for an estrogenic activity. The most efficient toxicity removal was observed for the polar compounds.
Ecotoxicology and Environmental Safety 05/2011; 74(4):844-51. · 2.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The anthropogenic impact on the pristine karst lakes was investigated using combination of specific parameters, including multielemental analysis of major inorganic constituents (Al, K, Fe) and trace metals (Li, Ag, Cd, Sn, Pb, Bi, Cr, Co, Ni, Cu, Zn and Sb), polycyclic aromatic hydrocarbons (PAHs) and anionic surfactants of linear alkylbenzene sulfonate (LAS) type. The study was performed in the Plitvice Lakes National Park, situated in a sparsely populated area of the northwestern Dinarides, central Croatia. Dated cores of recent sediments from the two biggest lakes, Lake Prosce and Lake Kozjak, were analysed for the selected contaminants using highly specific methods, involving inductively coupled plasma mass spectrometry (ICP/MS), gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/tandem mass spectrometry (LC/MS/MS). The concentration of inorganic constituents reflected primarily the geological background of the area as well as geomorphological and geochemical characteristics of the Plitvice Lakes. Due to the higher terrigenous input, the concentration of all elements was significantly higher in the Lake Prosce. The concentration of toxic metals was relatively low in both lakes, except for Cd (>1 mg kg(-1)) and Pb (up to 40 mg kg(-1)). The vertical profiles of these metals suggested that elevated concentrations of Cd were of natural origin, derived from the erosion of the Jurassic dolomite bedrock, while Pb was predominately of recent anthropogenic origin. A similar distribution pattern, suggesting the same prevailing mechanism of input, was observed for pyrolytic PAHs. The characteristic diagnostic PAH ratios revealed that higher PAHs prevailingly originated from the combustion of biomass and fossil fuels. LAS, which represent highly specific indicators of untreated wastewaters, were found in rather high concentrations in the recent sediment layers (up to 4.7 mg kg(-1)), suggesting that contaminated household and hotel wastewaters reach the Lakes, very probably by leaking through the porous karst rocks.
[Show abstract][Hide abstract] ABSTRACT: Picoplankton distribution at the boundary zone of the southern Adriatic in May 2009 on a 75 km long shelf-continental slope transect was assessed by combining epifluorescence microscopy, flow cytometry and high-performance liquid chromatography data with hydrographic observations. The picoplankton distribution was greatly influenced by the hydrographic conditions prevailing in the southern Adriatic because of the influence of the Levantine Intermediate Water (LIW) and East Adriatic Current (EAC) forcing. Heterotrophic bacteria numerically dominated the picoplankton community through the entire transect with no significant accumulation. By contrast, picophytoplankton accumulated in the 50–75 m layer, forming a pronounced deep chlorophyll maximum. Synechococcus dominated the photosynthetic picoplankton, whereas picoeukaryotes were the least abundant. The intrusion of warm LIW observed in the layer between 100 and 350 m was followed by Prochlorococcus and Synechococcus peaks (10 × 103 cells mL−1 and 90 × 103 cells mL−1, respectively), as well as by the appearance of two Synechococcus ecotypes. Most picoeukaryotes were observed at the offshore stations, where geostrophic current calculation revealed the strongest EAC influence. A strong EAC spread over the central and eastern basin created a barrier for Prochlorococcus, whereas the picoeukaryote maxima coincided with the core of the EAC, suggesting its persistence to hydrological instabilities.Research highlights► We examined the influence of hydrographic conditions on picoplankton distribution. ► Picophytoplankton accumulated from 50 to 75 m, forming a deep chlorophyll maximum. ► LIW intrusion was followed by the appearance of two Synechococcus ecotypes. ► The East Adriatic Current (EAC) acted like a barrier for Prochlorococcus. ► The picoeukaryotes maxima coincided with the core of the EAC.
[Show abstract][Hide abstract] ABSTRACT: A comprehensive analytical procedure for a reliable identification of nontarget polar contaminants in aquatic sediments was developed, based on the application of ultra-high-pressure liquid chromatography (UHPLC) coupled to hybrid quadrupole time-of-flight mass spectrometry (QTOFMS). The procedure was applied for the analysis of freshwater sediment that was highly impacted by wastewater discharges from the pharmaceutical industry. A number of different contaminants were successfully identified owing to the high mass accuracy of the QTOFMS system, used in combination with high chromatographic resolution of UHPLC. The major compounds, identified in investigated sediment, included a series of polypropylene glycols (n=3-16), alkylbenzene sulfonate and benzalkonium surfactants as well as a number of various pharmaceuticals (chlorthalidone, warfarin, terbinafine, torsemide, zolpidem and macrolide antibiotics). The particular advantage of the applied technique is its capability to detect less known pharmaceutical intermediates and/or transformation products, which have not been previously reported in freshwater sediments.
[Show abstract][Hide abstract] ABSTRACT: A comprehensive study of various psychoactive substances and their metabolites was performed in the wastewater treatment plant of the city of Zagreb (780 000 inhabitants) using liquid chromatography/tandem mass spectrometry (LC-MS-MS). The estimation of drug abuse for five different illicit drugs, including heroin, cocaine, marijuana, amphetamine and ecstasy, was made on the basis of their representative excretion rates, which were determined over a period of 8 months. Marijuana (1000 kg/year), heroin (75 kg/year) and cocaine (47 kg/year) were found to be the most frequently consumed illicit drugs, while the consumption of amphetamine-type drugs was much lower (1-3 kg/year). A comparison with other reports indicated that drug abuse profiles in transition countries might be different from those reported for Western Europe, in particular with respect to the comparatively increased consumption of heroin. Enhanced consumption of stimulating drugs (cocaine and ectasy) was systematically detected during weekends.
[Show abstract][Hide abstract] ABSTRACT: This paper reports on a comprehensive reconnaissance of over seventy individual wastewater contaminants in the region of Western Balkan (WB; Bosnia and Herzegovina, Croatia and Serbia), including some prominent classes of emerging contaminants such as pharmaceuticals and personal care products, surfactants and their degradation products, plasticizers, pesticides, insect repellents, and flame retardants. All determinations were carried out using a multiresidue analytical approach, based on the application of gas chromatographic and liquid chromatographic techniques coupled to mass spectrometric detection. The results confirmed a widespread occurrence of the emerging contaminants in municipal wastewaters of the region. The most prominent contaminant classes, determined in municipal wastewaters, were those derived from aromatic surfactants, including linear alkylbenzene sulphonates (LAS) and alkylphenol polyethoxylates (APEO), with the concentrations in raw wastewater reaching into the mg/l range. All other contaminants were present in much lower concentrations, rarely exceeding few microg/l. The most abundant individual compounds belonged to several classes of pharmaceuticals (antimicrobials, analgesics and antiinflammatories, beta-blockers and lipid regulators) and personal care products (fragrances). Due to the rather poor wastewater management practices in WB countries, with less than 5% of all wastewaters being biologically treated, most of the contaminants present in wastewaters reach ambient waters and may represent a significant environmental concern.
Science of The Total Environment 08/2008; 399(1-3):66-77. · 3.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Phytoplankton distribution and environmental characteristics were determined in a shallow, highly stratified and oligotrophic estuary (Zrmanja, eastern Adriatic). Samples were collected in two contrasting seasons; winter (February 2000), when river discharge was high, and in summer (July 2003), a period of drought. Phytoplankton distribution was closely related to salinity gradients, nutrient levels, and water residence time. Microscopic analysis revealed that phytoplankton was composed mainly of marine diatoms, dinoflagellates, cryptophytes, green flagellates, and coccolithophorids. The dominant biomarker pigments were fucoxanthin, alloxanthin and 19'-hexanoyloxyfucoxanthin, while lower, but indicative contributions of peridinin and chlorophyll b were also noted. Maximum abundance and biomass were found in the middle estuary in winter and in the upper estuary in summer. The estuary is mostly P-limited. Development of chain-forming marine diatoms was evident in winter. Due to the reduced nutrient input in summer, the biomass accumulated in the upper estuary (1,000 ng chlorophyll a l(-1)) was composed mostly of nanoplanktonic unicellular diatoms, nanoplanktonic marine dinoflagellates, cryptophytes, and chlorophytes. The concentrations of about 200 ng l(-1) hex-fuco, suggested that the contribution of prymnesiophytes to total biomass was comparable to that of diatoms and dinoflagellates. In the middle estuary and coastal sea, PO(4) and TIN were 3.5 times lower, resulting in a fivefold decrease in biomass (<100 ng chlorophyll a l(-1)). The oligotrophic Zrmanja and other karstic rivers discharging in the eastern Adriatic Sea, provide insufficient source of nutrients and low productivity of the eastern Adriatic Sea.
Environmental Monitoring and Assessment 07/2008; 142(1-3):199-218. · 1.59 Impact Factor