Methylmercury determination in sediments and fish tissues from the Nerbioi-Ibaizabal estuary (Basque Country, Spain)

The analysis of methylmercury in extracts from environmental solid samples by gas chromatography coupled to microwave induced plasma atomic emission spectrometry (GC-MIP/AES) after the ethylation of the extract and the preconcentration of the volatile products in hexane has been critically investigated. In order to correct potential sources of random error along the analytical procedure affecting the overall repeatability of the analysis, the use of the inorganic mercury naturally occurring in the sample as internal standard in the analysis of methylmercury is proposed. A study to establish the best conditions to achieve a quantitative recovery of methylmercury without damaging its chemical structure has also been carried out. Magnetic stirring (without heating) of the sediment or fish tissue with 2 mol dm⁻³ HNO₃ or 10% methanolic KOH, respectively, during 90 min has been considered as the most effective procedure to release methylmercury preserving its structure. The proposed method has been validated using certified reference materials (CRM-580, CRM-463 and DOLT-2), assessing its quality in terms of accuracy, repeatability and detection limit. Finally, several sediment and fish samples collected in the estuary of the Nerbioi-Ibaizabal (Bilbao, Basque Country) have been analyzed following the procedures proposed. The results obtained show the validity of the proposed method to analyze real samples.     

Determination and comparison of heavy metals in selected seafood, water, vegetation and sediments by inductively coupled plasma-optical emission spectrometry from an industrialized and pristine waterway in Southwest Louisiana

Samples of crab, fish, sediments, vegetation and waters were collected from a pristine (Rockefeller Wildlife Refuge) and historically polluted (Bayou d'Inde) waterway in Southwest Louisiana. After sample preparation via microwave digestion to obtain a solution, cadmium, mercury, nickel and lead were determined by inductively coupled plasma-optical emission spectrometry. A comparison of results showed that there was very little difference in concentrations for the four metals between the two areas, typically in the 10 μg/g range for crabs and fish, around 0.05 μg/mL or lower for waters, and somewhat higher for sediments and vegetation of around two to three times.     

Trace metal concentrations in single specimens of the intestinal broad flatworm (Diphyllobothrium latum), compared to their fish host (Oncorhynchus mykiss) measured by total reflection X-ray fluorescence spectrometry

The aim of this study was to investigate (1) whether intestine endoparasites (Diphyllobothrium latum) accumulate trace elements related to its body size and (2) whether parasites bioconcentrate more trace elements than their host. Freshwater fish (rainbow trout Oncorhynchus mykiss) were sampled in the deep, oligotrophic and uncontaminated Lake Riñihue in Southern Chile. The element concentration of different organs (intestine, muscle, liver) and of the intestine endoparasites were analyzed using total reflection X-ray fluorescence spectrometry. The results showed that the mass fraction for Mn, Fe, Ni, Cu, and Pb decreased significantly with the body size (dry weight) of the endoparasite. Only Zn did not reveal such a relationship. Small parasites accumulated up to 80 times more Fe, Ni, Mn, Pb, and Cu than large parasites. Compared to the fish organs, small parasites accumulated in maximum 35 to 307 times more Mn, 5 to 255 times more Fe, 98 to 220 times more Ni, 3 to 175 times more Cu, and 0.4 to 12 times more Zn than the fish. Lead was only found in the endoparasite, but not in the fish organs. We conclude that (1) D. latum is a good indicator for trace element accumulation in fishes and that (2) small endoparasites are more sensitive as bioindicators because they showed higher bioconcentrations of trace metals than larger parasites.     

Optical enzyme sensor for determining <Emphasis Type="SmallCaps">l</Emphasis>-lysine content using <Emphasis Type="SmallCaps">l</Emphasis>-lysine oxidase from the rockfish <Emphasis Type="BoldItalic">Sebastes schlegeli</Emphasis>

l-Lysine (l-Lys) in living bodies is critical for metabolism; therefore, determination of its levels in food is important. Most enzymatic methods for l-Lys analysis are performed using l-lysine oxidase (LyOx), but commercially manufactured LyOx is generally not highly selective for l-Lys among amino acids. We previously isolated LyOx as an antibacterial protein secreted from the skin of the rockfish Sebastes schlegeli. In the present study, we developed an optical enzyme sensor system for rapid and continuous determination of l-Lys using this LyOx. The system comprised an immobilized LyOx membrane, an optical oxygen probe, a flow system, and a personal computer. The amount of l-Lys was detected as a decrease in the oxygen concentration due to the LyOx reaction. The specificity of the sensor was examined against various amino acids. The sensor response was specific for l-Lys. Good reproducibility was obtained in 58 assays. The response of the sensor using commercially prepared LyOx was unstable compared with the response using LyOx isolated in our laboratory. Our sensor system could be used for 5 weeks without our having to change the enzyme membrane. The calibration curve for a standard l-Lys solution was linear from 0.1 to 3.0 mmol L⁻¹. One assay could be completed within 2 min. The sensor was applied to determine the l-Lys content in food samples such as bonito cooking water and scallop hepatopancreas. The values obtained using the sensor and conventional high-performance liquid chromatography methods were well correlated.     

Development of a group selective molecularly imprinted polymers based solid phase extraction of malachite green from fish water and fish feed samples

A group selective molecularly imprinted solid phase extraction (MISPE) for malachite green (MG) from fish water and fish feed samples was developed. Using MG as template molecule, methacrylic acid as functional monomer, ethylene glycoldimethacrylate as linking agent and bulk polymerization as synthetic method, the molecularly imprinted polymers (MIPs) were synthesized and characterized with rebinding experiment. The Scatchard polt's analysis revealed that the template-polymer system showed the two-site binding behavior with dissociation constants of 0.3194 μmol L⁻¹ and 15.70 μmol L⁻¹, respectively. MG and two structurally related compounds, leucomalachite green (LMG) and crystal violet (CV) were employed for selectivity test. The MIPs exhibited the highest selective rebinding to MG, but also displayed 83.0% and 87.5% of cross-reactivity with LMG and CV, demonstrating that MIPs could be used as group recognition sorbents in solid phase extraction. The extraction conditions of MISPE column for MG were optimized. Tap water samples spiked with MG at concentration of 0.5-10 ng mL⁻¹ were extracted by MISPE column and analyzed by high performance liquid chromatography. The recoveries of MISPE column for MG extraction were found to be 76.8-93.7% with the relative standard deviations of 2.12-10.09%, indicating the feasibility of the prepared MIPs for MG extraction. No detectable MG was observed in one fish farming water sample and two fish feed samples; while the MG concentrations in two pet fishpond water samples were found at 1.50 ng mL⁻¹ and 0.67 ng mL⁻¹, respectively.     

Environmentally friendly solid‐phase microextraction coupled with gas chromatography and mass spectrometry for the determination of biogenic amines in fish samples

In this work, a facile and environmentally friendly solid‐phase microextraction assay based on on‐fiber derivatization coupled with gas chromatography and mass spectrometry was developed for determining four nonvolatile index biogenic amines (putrescine, cadaverine, histamine, and tyramine) in fish samples. In the assay, the fiber was firstly dipped into a solution with isobutyl chloroformate as derivatization reagent and isooctane as extraction solvent. Thus, a thin organic liquid membrane coating was developed. Then the modified fiber was immersed into sample solution to extract four important bioamines. Afterwards, the fiber was directly inserted into gas chromatography injection port for thermal desorption. 1,7‐Diaminoheptane was employed as internal standard reagent for quantification of the targets. The limits of detection of the method were 2.98–45.3 μg/kg. The proposed method was successfully applied to the detection of bioamines in several fish samples with recoveries ranging 78.9–110%. The organic reagent used for extraction was as few as microliter that can greatly reduce the harm to manipulator and environment. Moreover, the extraction procedures were very simple without concentration and elution procedures, which can greatly simplify the pretreatment process. The assay can be extended to the in situ screening of other pollutant in food safety by changing the derivatization reagent.     

化学发光酶免疫分析测定鱼肉中呋喃它酮代谢物方法研究

建立了呋喃它酮代谢物5-吗啉甲基-3-氨基-2-6唑烷基酮(AMOZ)间接竞争化学发光酶免疫分析(icCLEIA)检测方法,通过单因素实验优化了包被原浓度、抗体稀释倍数、反应缓冲体系及浓度、竞争反应时间等参数,结果表明icCLEIA最佳反应条件为:包被抗原浓度为10 ng/mL,抗体稀释60000倍,最佳竞争时间为50 min,体系缓冲液0.01 mol/L PBS(pH 7.4).在优化的条件下,本方法的线性检测范围为0.026 ～3.52 μg/L,IC50为0.29 μg/L,检出限(LOD,IC10)为0.012 μg/L.对鱼肉样品的平均添加回收率在101.4％～115.5％之间.建立的icCLEIA方法可用于实际样品中AMOZ残留检测.     

Effect of different lipid extraction methods on δ13C of lipid and lipid-free fractions of fish and different fish feeds

For many ecological applications of stable carbon isotope techniques, it is necessary to separate the lipid and lipid-free fractions. The effect of different lipid extraction methods on the isotope signature of the remaining lipid-free matter as well as the lipid fraction was tested. A hot extraction form of the Soxhlet method using petrol-ether was compared with two liquid-liquid extraction methods for lipid determination described by Bligh and Dyer and Smedes. Solid samples of fish and different natural food items were subjected to extraction and the carbon isotope ratios in lipid and lipid-free matter determined by IRMS. All methods were suitable for lipid extraction from all samples analysed here and did not cause biologically relevant differences (>1) in carbon isotopic ratios, except the Bligh and Dyer extraction method using chloroform which caused systematic errors for δ ¹³C when applied to diatoms.     

Optimisation of two-dimensional undulatory swimming at high Reynolds number

The propulsion performance of a flexible plate undergoing an arbitrary harmonic motion in a two-dimensional and inviscid fluid is addressed. This plate being free of external forces, heaving and pitching cannot be imposed and are the results of recoil conditions on the body. Linear unsteady airfoil theory is first used to calculate the average thrust and power required for swimming. The propulsive performance is then discussed in terms of hydrodynamic efficiency, energy consumption and average swimming speed and two different asymptotic regimes are identified: the low-velocity regime and the high-velocity regime. The optimal swimming gaits are calculated in the different regimes as a function of the plate mass ratio and leading-edge suction. Finally an empirical non-linear model is proposed to complement the linear model and the optimal swimming modes are calculated and discussed within this framework. For a fixed number of degrees of freedom, it is found that the Strouhal number of the optimal mode is almost constant, independently of the swimming regime.     

Simple, high throughput ultra-high performance liquid chromatography/tandem mass spectrometry trace analysis of perfluorinated alkylated substances in food of animal origin: milk and fish

The present study documents development and validation of a novel approach for determination of 23 perfluorinated alkylated substances (PFASs) in food of animal origin represented by milk and fish. The list of target analytes comprises four classes of PFASs, both ionic and non-ionic: 11 perfluorocarboxylic acids (PFCAs), 4 perfluorosulphonic acids (PFSAs), 5 perfluorosulphonamides (FOSAs) and 3 perfluorophosphonic acids (PFPAs). Fast sample preparation procedure is based on an extraction of target analytes with acetonitrile (MeCN) and their transfer (supported by inorganic salts and acidification) into the organic phase. Removing of matrix co-extracts by a simple dispersive solid phase extraction (SPE) employing ENVI-Carb and C18 sorbents is followed by an efficient sample pre-concentration performed by acetonitrile evaporation and subsequent dilution of residue in a small volume of methanol (matrix equivalent in the final extracts was 16 and 8 g mL(-1), for milk and fish respectively). Using modern instrumentation consisting of ultra-high performance liquid chromatography (UHPLC) hyphenated with a tandem mass spectrometer (MS/MS), limits of quantification (LOQs) as low as 0.001-0.006 μg kg(-1) for milk and 0.002-0.013 μg kg(-1) for fish can be achieved. Under these conditions, a wide spectrum of PFASs, including minor representatives, can be determined which enables collecting data required for human exposure studies. The pilot study employing the new method for examination of milk and canned fish samples was realized. Whereas in majority of canned fish products a wide spectrum of PFCAs, perfluorooctanesulphonic acid (PFOS) and perfluoro-1-octanesulphonamide (PFOSA) was detected, only in a few milk samples very low concentrations (LOQ levels) of PFOS and perfluorooctansulphonic acid (PFDS) were found.