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Estrone (E1), a metabolite of the estrogenic hormones 17β-estradiol (β-E2) and 17α-estradiol (α-E2), is itself a potent estrogen which can have a significant impact on the hormonal balance. Due to ist high potential for adverse effects on human health and aquatic life even at pg/L to ng/L levels, its appearance in water should be monitored. E1 has also been considered a marker substance for the presence of other estrogens. This study presents a newly developed direct competitive enzymelinked immunosorbent assay (ELISA) for quantification of E1 in environmental water samples using new monoclonal antibodies.
The quantification range of the ELISA is 0.15 μg/L to 8.7 μg/L E1, and the limit of detection is around 60 ng/L for not pre-concentrated water samples. A pre-concentration step after careful selection of suitable phases for SPE was developed, too. The influence of organic solvents and natural organic matter on the ELISA was assessed. The high selectivity of the monoclonal antibody was demonstrated by determining the cross-reactivity against 20 structurally related compounds.
For the assessment of matrix effects, a concept (“LC–ELISA”) is thoroughly exploited, i.e., separating complex samples by HPLC into 0.3 min fractions and determination of the apparent E1 concentration. Furthermore, fractions with interferences for nontarget/suspected-target analysis can be assigned. A dilution approach was applied to distinguish between specific interferences (cross-reactants) and non-specific interferences (matrix effects). In the determination of 18 environmental samples, a good agreement of the E1 concentration in the respective fractions was obtained with mean recoveries of 103 % to 132 % comparing ELISA to LC–MS/MS.
Several works evaluated the toxicity of pharmaceutical drugs and climate related changes in invertebrates but few explored the combined effects of both stressors, namely considering their mode of action (MoA). Carbamazepine (CBZ) and cetirizine (CTZ) are pharmaceutical drugs detected in the environment and the toxicity derived from the combined effects of these drugs with ocean acidification (OA) is poorly explored. Thus, the present study investigated the biochemical parameters related to an oxidative stress response and the transcription of genes related to the MoA of CBZ (1.0 mg/L) and CTZ (0.6 mg/L) in the clam Ruditapes philippinarum chronically exposed (28 days) to control (7.8) and low (7.5) pH conditions. The results obtained showed that despite the clams accumulated both drugs, at low pH the clams exposed to CTZ decreased drug concentration and BCF values (CTZ uptake: 2.0 ± 0.5 ng/g fresh weight; BCF: 3.8 ± 0.9) in comparison with clams exposed to control pH (CTZ uptake: 2.9 ± 0.3 ng/g fresh weight; BCF: 5.5 ± 0.6). No oxidative stress was induced by the exposure to CBZ or CTZ at each pH level, but the transcription of several genes related with the MoA (neurotransmission, immunity and biomineralization) was altered by low pH, drug exposure and the combination of both stressors. At both pH conditions, CBZ increased the transcription of GABA receptor gene (neurotransmission) and CTZ led to a decrease of Perlucin gene (biomineralization) transcription. The transcription of MyD88 gene (immunity) decreased at low pH (7.5) combined with drug exposure (CBZ or CTZ). Thus, it was highlighted that the interaction of drug exposure and low pH conditions can change bivalves’ sensitivity to drugs or alter drugs toxicity.
In the aquatic environment, organisms are exposed to complex mixtures of contaminants which may alter the toxicity profile of each compound, compared to its toxicity alone. Pharmaceutical drugs (e.g. carbamazepine (CBZ) and cetirizine (CTZ)) and metals (e.g. cadmium (Cd)) are among those contaminants that co-occur in the environment. However, most studies concerning their toxicity towards aquatic species are based on single exposure experiments. Thus, the present study aimed to evaluate single and combined effects of Cd and CBZ or CTZ (single conditions: Cd, CTZ, CBZ; combined conditions: CTZ+Cd, CBZ+Cd) on biomarkers related to oxidative stress and energy metabolism in the edible clam Ruditapes philippinarum, by exposing the organisms for 28 days to environmentally relevant concentrations of these contaminants. The biomarkers studied were: i) the electron transport system activity, protein and glycogen contents (indicators of organisms’ metabolic status and energy reserves); ii) lipid peroxidation and the ratio between reduced and oxidized glutathione (indicators of oxidative stress); iii) superoxide dismutase and catalase activities (enzymes indicators of antioxidant defence) and iv) activity of glutathione S-transferases (family of enzymes indicators of biotransformation capacity). Results obtained showed that the uptake of Cd and CBZ was not affected by the combined presence of the contaminants.
However, for CTZ, the uptake was higher in the presence than in the absence of Cd. Concerning toxicity data, in general, the combined exposures (CTZ+Cd, CBZ+Cd) had lower biological effects than the contaminants alone. Nevertheless, our data showed that despite the low concentrations tested, they were enough to exert biological effects that differed between single and combined treatments, evidencing the need to conduct more co-exposure studies to increase the environmental relevance of the gathered data.
Microfluidic electrochemical immunosensor for the trace analysis of cocaine in water and body fluids
(2018)
Quick but accurate testing and on‐the‐spot monitoring of cocaine in oral fluids and urine continues to be an important toxicological issue. In terms of drug testing, a number of devices have been introduced into the market in recent decades, notably for workplace inspection or roadside testing. However, these systems do not always fulfill the requirements in terms of reliability, especially when low cut‐off levels are required. With respect to surface water, the presence of anthropogenic small organic molecules such as prescription and over‐the‐counter pharmaceuticals as well as illicit drugs like cannabinoids, heroin, or cocaine, has become a challenge for scientists to develop new analytical tools for screening and on‐site analysis because many of them serve as markers for anthropogenic input and consumer behavior. Here, a modular approach for the detection of cocaine is presented, integrating an electrochemical enzyme‐linked immunosorbent assay (ELISA) performed on antibody‐grafted magnetic beads in a hybrid microfluidic sensor utilizing flexible tubing, static chip and screen‐printed electrode (SPE) elements for incubation, recognition, and cyclic voltammetry measurements. A linear response of the sensor vs. the logarithm of cocaine concentration was obtained with a limit of detection of 0.15 ng/L. Within an overall assay time of 25 minutes, concentrations down to 1 ng/L could be reliably determined in water, oral fluids, and urine, the system possessing a dynamic working range up to 1 mg/L.