Aptamer-based optical biosensor for rapid and sensitive detection of 17β-estradiol in water samples

Required routine monitoring of endocrine disrupting compounds (EDCs) in water samples, as posed by EPA Unregulated Contaminant Regulation (UCMR3), demands for cost-effective, reliable and sensitive EDC detection methods. This study reports a reusable evanescent wave aptamer-based biosensor for rapid, sensitive and highly selective detection of 17β-estradiol, an EDC that is frequently detected in environmental water samples. In this system, the capture molecular, β-estradiol 6-(O-carboxy-methyl)oxime-BSA, was covalently immobilized onto the optical fiber sensor surface. With an indirect competitive detection mode, samples containing different concentrations of 17β-estradiol were premixed with a given concentration of fluorescence-labeled DNA aptamer, which highly specifically binds to 17β-estradiol. Then, the sample mixture is pumped to the sensor surface, and a higher concentration of 17β-estradiol leads to less fluorescence-labeled DNA aptamer bound to the sensor surface and thus to lower fluorescence signal. The dose-response curve of 17β-estradiol was established and a detection limit was determined as 2.1 nM (0.6 ng mL(-1)). The high specificity and selectivity of the sensor were demonstrated by evaluating its response to a number of potentially interfering EDCs. Potential interference of real environmental sample matrix was assessed by spiked samples in several tertiary wastewater effluents. The sensor can be regenerated with a 0.5% SDS solution (pH 1.9) over tens of times without significant deterioration of the sensor performance. This portable sensor system can be potentially applied for on-site real-time inexpensive and easy-to-use monitoring of 17β-estradiol in environmental samples such as effluents or water bodies.     

Microwave-assisted extraction and determination of cyanuric acid residue in infant formula samples by liquid chromatography–tandem mass spectrometry

In the present work, microwave-assisted extraction method in combining with liquid chromatography–tandem mass spectrometry (LC–MS/MS) was proposed for the determination of cyanuric acid (CYA) in infant formula samples. The separation was performed on a MERCK ZIC HILIC column (150 × 2.1 mm i.d., 5 μm) with gradient elution of 20 mM ammonium acetate solution – acetonitrile. The method could respond linearly with cyanuric acid at concentrations from 1.0 to 50 ng mL⁻¹ with a quantification limit of 0.25 mg kg⁻¹. The intra- and inter-day precision was less than 4% and the recovery of the assay was in the range of 86.7–93.1%. In the analysis of practical spiked infant formula samples, the new method yielded satisfactory results. Due to its simplicity and accuracy the straightforward method is particularly suitable for routine cyanuric acid detection.     

Proteomic analysis of 17β-estradiol degradation by Stenotrophomonas maltophilia

Microbial degradation plays a critical role in determining the environmental fate of steroid hormones, such as 17β-estradiol (E2). The molecular mechanisms governing the microbial transformation of E2 and its primary degradation intermediate, estrone (E1), are largely unknown. The objective of this study was to identify metabolism pathways that might be involved in microbial estrogen degradation. To achieve the objective, Stenotrophomonas maltophilia strain ZL1 was used as a model estrogen degrading bacterium and its protein expression level during E2/E1 degradation was studied using quantitative proteomics. During an E2 degradation experiment, strain ZL1 first converted E2 to E1 stoichiometrically. At 16 h E1 reached its peak concentration, and microbial growth started. At the same time, enzymes involved in certain catabolic and anabolic pathways were most highly expressed compared to the other time points tested. Among those enzymes, the ones involved in protein and lipid biosyntheses were observed to be particularly active. Based on the metabolite information from a previous study and the proteomic data from this study, we hypothesized that S. maltophilia strain ZL1 was able to convert E1 to amino acid tyrosine through ring cleavage on a saturated ring of the E1 molecule and then utilize tyrosine in protein biosynthesis.     

Simultaneous determination of kasugamycin and validamycin-A residues in cereals by consecutive solid-phase extraction combined with liquid chromatography–tandem mass spectrometry

Two polar aminoglycosides, kasugamycin and validamycin-A, were determined in cereals (brown rice, wheat and corn) by high-performance liquid chromatography–tandem mass spectrometry. The analytes were extracted from samples using methanol and water (70:30, v/v) at pH 5.5, purified using both a hydrophilic–hydrophobic-balanced cartridge and a strong cation-exchange cartridge, and then analysed using multiple reaction monitoring in positive electrospray ionisation mode with a special ReproSil 100 C₁₈ high-performance liquid chromatography column. This newly proposed method yielded good sensitivity and excellent chromatographic performance. The limits of quantification for kasugamycin and validamycin-A were 4.1 µg/kg and 1.0 µg/kg, respectively. The recoveries for both compounds at three fortification levels (4, 100 and 500 µg/kg for kasugamycin; 1, 10 and 100 µg/kg for validamycin-A) ranged from 75% to 110%, and the relative standard deviations were below 15%.     

Comparison of estrone and 17β-estradiol levels in commercial goat and cow milk

Increased levels of estrogen metabolites are believed to be associated with cancers of the reproductive system. One potential dietary source of these metabolites that is commonly consumed worldwide is milk. In North America, dairy cows are the most common source of milk; however, goats are the primary source of milk worldwide. In this study, the absolute concentrations of unconjugated and total (unconjugated plus conjugated) estrone (E(1)) and 17β-estradiol (E(2)) were compared in a variety of commercial cow milks (regular and organic) and goat milk. A lower combined concentration of E(1) and E(2) was found in goat milk than in any of the cow milk products tested. The differences in E(1) and E(2) levels between regular and organic cow milks were not as significant as the differences between goat milk and any of the cow milk products. Goat milk represents a better dietary choice for individuals concerned with limiting their estrogen intake.     

Fate of 4-nonylphenol and 17β-estradiol in the Redwood River of Minnesota

The majority of previous research investigating the fate of endocrine-disrupting compounds has focused on single processes generally in controlled laboratory experiments, and limited studies have directly evaluated their fate and transport in rivers. This study evaluated the fate and transport of 4-nonylphenol, 17β-estradiol, and estrone in a 10-km reach of the Redwood River in southwestern Minnesota. The same parcel of water was sampled as it moved downstream, integrating chemical transformation and hydrologic processes. The conservative tracer bromide was used to track the parcel of water being sampled, and the change in mass of the target compounds relative to bromide was determined at two locations downstream from a wastewater treatment plant effluent outfall. In-stream attenuation coefficients (k(stream)) were calculated by assuming first-order kinetics (negative values correspond to attenuation, whereas positive values indicate production). Attenuation of 17β-estradiol (k(stream) = -3.2 ± 1.0 day(-1)) was attributed primarily due to sorption and biodegradation by the stream biofilm and bed sediments. Estrone (k(stream) = 0.6 ± 0.8 day(-1)) and 4-nonylphenol (k(stream) = 1.4 ± 1.9 day(-1)) were produced in the evaluated 10-km reach, likely due to biochemical transformation from parent compounds (17β-estradiol, 4-nonylphenolpolyethoxylates, and 4-nonyphenolpolyethoxycarboxylates). Despite attenuation, these compounds were transported kilometers downstream, and thus additive concentrations from multiple sources and transformation of parent compounds into degradates having estrogenic activity can explain their environmental persistence and widespread observations of biological disruption in surface waters.     

Determination of alkylphenol residues in breast and commercial milk by solid-phase extraction and gas chromatography–mass spectrometry

This study presents a feasible and sensitive method to determine alkylphenol residues (i.e., 4-t-octylphenol (4-t-OP) and the isomers of 4-nonylphenols (4-NPs)) in breast milk samples and commercial cow milk products. The matrix interference associated with the constituents in the milk was reduced by extraction with n-hexane and dilution with 50% methanolic solution (v/v, methanol/water), then followed by the Oasis-HLB SPE extraction. The analytes were determined by a GC–MS system in full-scan and selected ion monitoring modes simultaneously. Limit of quantitation was less than 0.05 ng/g in a 20 g (wet weight) milk sample. The 4-NPs were detected in 19 of the 20 breast milk samples at concentrations ranging from 1.7 to 11.6 ng/g, while 4-t-OP was detected in 8 samples at concentrations ranging from 0.4 to 1.1 ng/g. The 4-NPs were detected in all the testing commercial cow milk products at concentrations ranging from 2.9 to 8.8 ng/g.     

Mercury speciation in seafood samples by LC–ICP-MS with a rapid ultrasound-assisted extraction procedure: Application to the determination of mercury in Brazilian seafood samples

This paper describes a simple method for mercury speciation in seafood samples by LC–ICP-MS with a fast sample preparation procedure. Prior to analysis, mercury species were extracted from food samples with a solution containing mercaptoethanol, l-cysteine and HCl and sonication for 15 min. Separation of mercury species was accomplished in less than 5 min on a C8 reverse phase column with a mobile phase containing 0.05%-v/v mercaptoethanol, 0.4% m/v l-cysteine and 0.06 mol L⁻¹ ammonium acetate. The method detection limits were found to be 0.25, 0.20 and 0.1 ng g⁻¹ for inorganic mercury, ethylmercury and methylmercury, respectively. Method accuracy is traceable to Certified Reference Materials (DOLT-3 and DORM-3) from the National Research Council Canada (NRCC). With the proposed method there is a considerable reduction of the time of sample preparation. Finally, the method was applied for the speciation of mercury in seafood samples purchased from the Brazilian market.     

Selectivity of molecularly imprinted solid phase extraction for sterol compounds

MIPs (molecularly imprinted polymers) for 17β-estradiol were prepared using 2-(trifluoromethyl) acrylic acid (TFMAA) as functional monomer and trimethylolpropane trimethacrylate (TRIM) as cross-linker by precipitation polymerisation. Data from HPLC system were calculated, and separation properties of several MIPs were reported. MISPEs (molecularly imprinted solid phase extractions) were performed with MIPs as sorbent, and absorption properties of the MISPEs prepared by UV irradiation initiating polymerisation at a ratio of 1/4 (17β-estradiol/TFMAA) for 17α-estradiol, 17β-estradiol, 4-androstene-3, 17-dione and progesterone investigated by detecting filtrates at each stage of sampling, washing and elution. Elution trials proved that addition acid or alkali reagents might affect retention of the MISPE and CSPE (control/no molecularly imprinted solid phase extraction) for 17β-estradiol. A competition test implicates that the MISPE has the strongest specific retention and enrichment for progesterone. In the actual experiment for 17β-estradiol with extracting solution of milk powder as samples, the MISPE showed better selectivity and enrichment property than C18 and CSPE, and the recovery on the MISPE was up to 85.5%.