Occurrence of common plastic additives and contaminants in mussel samples: Validation of analytical method based on matrix solid-phase dispersion.

A new matrix solid-phase dispersion (MSPD) extraction methodology, combined with high-performance liquid chromatography equipped with a diode-array detector, was developed and validated for the simultaneous determination of 10 compounds in mussels from Galician Rias (Spain). These pollutants are compounds commonly used for plastic production as additives, as well as common plastic contaminants. The compounds selected were bisphenol-A, bisphenol-F, bisphenol-S, nonylphenol-9, nonylphenol, diethyl phthalate, dibutyl phthalate, di-2-ethylhexyl phthalate, dichlorodiphenyltrichloroethane, dichlorodiphenyldichloroethane, and dichlorodiphenyldichloroethylene. The parameters affecting the MSPD extraction efficiency such as the type of sorbent, mass sample-sorbent ratio, and extraction solvent were optimised. The proposed method provided satisfactory quantitative recoveries (80-100%), with relative standard deviations lower than 7%. In all cases, the matrix-matched calibration curves were linear in the concentration range of 0.32-120.00 µg/kg, with quantification limits of 0.25-16.20 µg/kg. The novel developed MSPD-high-performance liquid chromatography methodology provided good sensitivity, accuracy, and repeatability for quality control analysis in mussels.

Effective determination of ampicillin in cow milk using a molecularly imprinted polymer as sorbent for sample preconcentration.

The aim of this study was to prepare molecularly imprinted polymers (MIPs) with ampicillin (AMP) and to evaluate the feasibility of these materials for being used as solid phase extraction sorbent for the selective preconcentration and determination of AMP in cow milk samples. MIPs were synthesized by bulk polymerization using methacrylic acid or methyl methacrylate as monomer and ethylene glycol dimethacrylate as cross-linker at different ratios. Characterization of the MIPs were carried out by Fourier-transform infrared spectrometry. The variables affecting the molecularly imprinted solid-phase extraction (MISPE) procedure were optimized. AMP recoveries were higher than 98%, and RSD less than 7%. A preconcentration factor of 20 was reached, which was sufficient to determine AMP at levels allowed by the EU (4µgkg-1) in cow milk. The selectivity of the AMP-MIP was evaluated in presence of other structurally related ß-lactam antibiotics (amoxicillin, oxacillin, penicillin G).

Synthesis and characterization of a molecularly imprinted polymer for the determination of spiramycin in sheep milk.

A series of molecularly imprinted polymers (MIPs) comprising reactionary sites which are complementary to macrolide antibiotic spiramycin (SPI) were synthetized by noncovalent bulk polymerization technique. MIPs were synthesized under different polymerization process and their recognition efficiency was evaluated in binding studies in comparison with non-imprinted polymers. The best MIP was morphologically characterized and equilibrium assays were carried out. The MIP was evaluated as a sorbent for extraction and preconcentration of SPI from aqueous and sheep milk samples, and an off-line MISPE method followed by high-performance liquid chromatography with UV diode-array detection was established. Good linearity were obtained for SPI in a range of 24-965µgkg-1 and the average recoveries at three spiked levels in milk samples were higher than 90% (RSD<5%). Limit of quantification was 24.1µgkg-1. Cross-reactivity studies from other macrolides with similar structure were tested. The optimum imprinted polymer showed a good selectivity and affinity for SPI, demonstrating the potential of the proposed MISPE for rapid, sensitive and effective sample pretreatment for selective determination of SPI in sheep milk samples.

Occurrence of erythromycin residues in sheep milk. Validation of an analytical method.

The paper describes a new and selective analytical sample treatment for quantitative extraction and preconcentration of erythromycin in presence of other macrolide antibiotics in sheep milk samples. The methodology is based on the use of a molecular imprinted polymer (MIP) employed as solid phase extraction sorbent (MISPE). The synthesized material by bulk polymerization using erythromycin (ERY) as template was evaluated as solid phase extraction sorbent, in a novel sample treatment technique that can be coupled to high-performance liquid chromatography with diode-array detector (HPLC-DAD). MIP selectivity was studied for other macrolide antibiotics with similar structures, such as tylosin (TYL), spiramycin (SPI), josamycin (JOS), roxithromycin (ROX) and ivermectin (IVER) getting recoveries for these interferents lower than 35%, for all cases except for ROX, which recoveries were around 85%. The variables affecting the molecularly imprinted solid-phase extraction (MISPE) procedure were optimized to select the best conditions of selectivity and sensitivity to determine ERY at concentration levels established by EU legislation in sheep milk. Under the selected experimental conditions, quantification limit was 24.1 µg kg(-1). Recoveries were higher than 98%, with RSDs between 0.7% and 2%. The proposed MISPE-HPLC method was validated and successfully applied to ERY analysis in sheep milk samples.

An HPLC-DAD method for the simultaneous determination of nine ß-lactam antibiotics in ewe milk.

The presence of ß-lactam residues in foodstuffs constitutes a potential risk to the human health and undesirable effects on consumers, and nowadays these antibiotic residues are also recognised as an emerging environmental problem. In addition, these are of great concern to prestigious Manchego cheese processors (Central Spain denomination of origin) because they reduce the curdling of milk and cause improper cheese ripening, which consequently lead to an important loss of monetary income. This work describes the development of a sensitive and reliable method using liquid chromatography with UV-diode array detection (LC-DAD) for simultaneous determination of the ß-lactam antibiotics, ampicillin (AMP), benzylpenicillin (PEG), cephalexin (CFX), cefazolin (CFL), cefoperazone (CFP), cloxacillin (CLO), dicloxacillin (DCL), oxacillin (OXA) and phenoxymethylpenicillin (PEV), in Manchega ewe milk. The column, mobile phase, temperature and flow rate were optimised to provide the best resolution of these analytes. The extraction method of the antibiotic residues involves the deproteinisation of the milk sample using acetonitrile and centrifugation followed by a solid-phase extraction (SPE) clean-up. The recoveries for the studied ß-lactams ranged from 79% to 96% with relative standard deviations between 0.5% and 4.9%. The limits of quantification (LOQs) for all these compounds were in the range of 3.4-8.6µgkg(-1), which are lower than the maximum residue limits (MRLs) established by the European Union for the studied ß-lactams in milk, making the method suitable for performing routine analyses. The proposed multi-residue LC-UV-diode array detection (LC-DAD) method is a powerful and popular alternative for the determination and confirmation of antibiotic residues in small milk industries and is the first one capable of determining nine ß-lactam antibiotics in samples of Manchega ewe milk.

Characterization of Spanish honeys with protected designation of origin "Miel de Granada" according to their mineral content.

Honey attributes such as geographical origin or specified botanical sources often command a premium price due to their organoleptic or pharmacoactive properties. "Miel de Granada" is a highly quality product with protected designation of origin (PDO) which includes six monofloral honeys and two multifloral honeys. Our objective was the characterization of "Miel de Granada" according to their metal content. Metal content was specific enough and allowed discrimination from honeys of different botanical and geographical origins and confirmed the authenticity of PDO labelling as Granada product with the determination of only five elements (K, Na, Ca, Mg and Zn). Chemometric techniques as cluster analysis and ANOVA were used to classify honeys according to their botanical and geographical origin in the metal data. Metal content marks the differences in honey samples and can be used as a tool to assess the quality of honeys. ANOVA showed significant differences among rosemary honeys from different geographical areas despite the botanical factor weight. Our research contributes to the groundwork studies to determine the geographical origin of Spanish honeys.

Development of a molecularly imprinted polymer-matrix solid-phase dispersion method for selective determination of ß-estradiol as anabolic growth promoter in goat milk.

A simple, fast, and sensitive method for determination of 17 ß-estradiol (E2) in goat milk samples has been developed by combining selective molecularly imprinted matrix solid-phase dispersion (MIP-MSPD) and liquid chromatography with diode-array detection (DAD). The molecularly imprinted polymer was synthesized by use of 17ß-estradiol as template molecule, methacrylic acid as functional monomer, ethylene glycol dimethacrylate as crosslinker monomer, azobisisobutyronitrile as initiator, and acetonitrile as porogen, and was used as selective solid support for matrix solid-phase dispersion. The selected dispersant had high affinity for E2 in the goat milk matrix and the extract obtained was sufficiently clean for direct injection for HPLC analysis without any interferences from the matrix. The proposed MIP-MSPD method was validated for linearity, precision, accuracy, decision limit (CCα) and detection capability (CCß), in accordance with European Commission Decision 2002/657/EC criteria. Linearity ranged from 0.3-10 µg g(-1) (correlation coefficient r(2) > 0.999). Mean recovery of E2 from goat milk samples at different spiked levels was between 89.5 and 92.2%, with RSD values within 1.3-2%. CCα and CCß values were 0.36 and 0.39 µg g(-1), respectively. The developed MIP-MSPD method was successfully applied to direct determination of E2 in goat milk samples.

Spectrophotometric simultaneous determination of nitrite, nitrate and ammonium in soils by flow injection analysis.

A new rapid flow injection procedure for the simultaneous determination of nitrate, nitrite and ammonium in single flow injection analysis system is proposed. The procedure combines on-line reduction of nitrate to nitrite and oxidation of ammonium to nitrite with spectrophotometric detection of nitrite by using the Griess-Ilosvay reaction. The formed azo dye was measured at 543 nm. The influence of reagent concentration and manifold parameters were studied. Nitrite, nitrate and ammonium can be determined within the range of 0.02-1.60 microgmL(-1), 0.02-1.60 microgmL(-1) and 0.05-1.40 microgmL(-1), respectively. R.S.D. values (n=10) were 2.66; 1.41 and 3.58 for nitrate, nitrite and ammonium, respectively. This procedure allows the determination and speciation of inorganic nitrogen species in soils with a single injection in a simple way, and high sampling rate (18 h(-1)). Detection limits of 0.013, 0.046 and 0.047 microgmL(-1)were achieved for nitrate, nitrite and ammonium, respectively. In comparison with others methods, the proposed one is more simple, it uses as single chromogenic reagent less injection volume (250 mL in stead of 350 mL) and it has a higher sampling rate.

A rapid fluorimetric screening method for the 1,4-benzodiazepines: determination of their metabolite oxazepam in urine.

Oxazepam is the major metabolite screened in urine samples for the evidence of the use of benzodiazepine drugs. The methods currently used, however, are laborious and time consuming. This paper proposes an oxazepam detection method based on its hydrolysis and cyclization--a reaction catalysed by cerium (IV) in an ortho-phosphoric acid-containing medium--to form 2-chloro-9(10H)-acridinone, a strongly fluorescent molecule. The variables involved in the hydrolysis and cyclization stages were optimised. Oxazepam was detectable in the 5-900 ng mL(-1) range, with a detection limit of 4.15 ng mL(-1) for k=3. The method was successfully used for the determination of oxazepam in urine samples collected at different times after the oral administration of Valium and Tranxilium.

Removal of carbaryl, linuron, and permethrin by Lupinus angustifolius under hydroponic conditions.

The metabolism of organic pollutants by plants normally requires contaminant direct uptake by cells. Factors affecting this uptake and the later distribution of chemicals within the plant include the physicochemical properties of the compounds (concentration, structure, solubility, log k(ow), diffusion rate) and the biochemical characteristics of the plant. This paper reports the tolerance, uptake, and effects of the pesticides carbaryl, linuron, and permethrin on Lupinus angustifolius germination and growth as well as contaminant intraplant distribution and possible degradation. Lupine plants were grown in hydroponic culture containing either 1 or 5 mg of the individual pesticides, or combinations of these (1, 5, or 10 mg of each), in 100 mL nutrient and water solutions. Analysis of the remaining solutions 8 days post-germination showed the water solutions to have higher remaining pesticide concentrations than nutrient solutions. Furthermore, in the presence of pesticides, germination was more frequent in the water solutions. After 16 days of growth, the plants were harvested, and their tissues were microwaved digested and analyzed by reversed-phase liquid chromatography. Although only minor quantities of each pesticide were detected in plant tissues, their amount in the roots was higher than in the stems. No accumulation was noted in the cotyledons, and only 2% of linuron was detected in the leaves. Mass recovery at the end of the experiment showed that 57, 53, and 55% of carbaryl, linuron, and permethrin, respectively, were degraded and/or bound in an irreversible manner to plant material. The results suggest that L. angustifolius could be useful for the cleaning/remediation of pesticide-contaminated water.

Liquid chromatography-UV diode-array detection method for multi-residue determination of macrolide antibiotics in sheep's milk.

A rapid, simple and sensitive liquid chromatography-UV diode-array detection method was developed for the simultaneous determination of seven macrolides (erythromycin, oleandomycin, roxithromycin, josamycin, spiramycin, tylosin and ivermectin) in sheep's milk. The column, mobile phase, temperature and flow rate were optimised to provide the best resolution of these analytes. The extraction of the antibiotic residues involves the treatment of protein-free samples with a combination of concentrated sodium hydroxide and ethyl acetate. Necessary defatting is achieved by alkaline hydrolysis. The recovery of each antibiotic was between 55% and 77%, with relative standard deviations ranging from 1% to 6.5%. The limit of quantification was 72.4 microg/kg for ivermectin, 48.3 microg/kg for roxithromycin, and 24.1 microg/kg for erythromycin, oleandomycin, spiramycin, josamycin and tylosin. The procedure was successfully used in the multi-residue determination of these macrolides at levels below the maximum concentrations legally allowed in milk samples.

Simultaneous determination of maneb and its main metabolites in tomatoes by liquid chromatography using diode array ultraviolet absorbance detection.

Liquid chromatography (LC) with diode array ultraviolet absorbance (DAD UV) detection is used for the simultaneous determination of the fungicide maneb and its main metabolites (ethylenethiourea--ETU, ethylenebis (isothiocyanate) sulfide--EBIS, and ethyleneurea--EU) in tomatoes. The identity of EBIS, one of the main UV degradation products of maneb, was verified by both DAD UV detection and mass spectrometry. The analytes were extracted three times with 3 mL of 1:1:1 acetonitrile-dichloromethane-chloroform by 2 min of mechanical shaking and separated on a C-18 column by gradient elution with an acetonitrile-methanol-aqueous 100mM sodium dodecylsulfate (SDS) mixture. The quantification limits of 0.45, 0.04, and 0.35 mg kg(-1) obtained for maneb, ETU, and EU, respectively, show that the proposed method is suitable for their determination in tomatoes.

Optimization of a matrix solid-phase dispersion method with subsequent clean-up for the determination of ethylene bisdithiocarbamate residues in almond samples.

A matrix solid-phase dispersion (MSPD) method with subsequent clean-up has been developed to isolate the ethylene bisdithiocarbamate (EBDC) main metabolites (ethylenethiourea, ETU, and ethylenebis [isothiocyanate] sulphide, EBIS) in almond samples. The optimized experimental set-up configuration involved 0.2 g of almond sample, washed sand as MSPD support and NaOH as defatting agent. A subsequent purification step on alumina using acetonitrile as extraction solvent was enough to remove all interferent matrix components, including the fatty material, and provide clean extracts. Quantitative analysis was performed by reversed phase liquid chromatography (RPLC) with diode-array ultraviolet absorbance (DAD UV) detector. Analytes recoveries were between 76 and 85% with relative standard deviations ranging from 3 to 12%. The low limits of quantification of 0.05 and 0.07 mg kg(-1) achieved for ETU and EBIS, respectively, make the method useful for the determination of EBDC residues on almond samples.

Evaluation of pesticide uptake by Lupinus seeds.

Pesticide uptake by seeds depends on the properties of the chemical, such as structure, stability, log k(ow) and diffusion rate, type of water, pH, temperature, content of organic matter and composition, and on seed characteristics such as permeability of the seed coat. The efficiency with which Lupinus angustifolius seeds retain different herbicides (simazine, atrazine, isoproturon, linuron,) and insecticides (carbaryl, fenamiphos, permethrin) was evaluated using both a batch and a continuous system. Factors which affect pesticide uptake by seeds, such as flow rate, seed biomass, pesticide concentration, contact time, pH, seed saturation and also the speed of the retention process for 17 days, were tested. L. angustifolius showed a high retention capacity for the above mentioned pesticides. The extraction of pesticides from seeds using different organic solvents, such as methanol, acetonitrile, ethyl acetate and n-hexane was evaluated and no pesticide residues were detected in any of the solvents tested. This could be attributed to the capacity of the seed to degrade the pesticides. From the results obtained, L. angustifolius seems to be a promising seed to be applied for phytoremediation of industrial effluents or contaminated water.

Trace elements and electrolytes in human resting mixed saliva after exercise.

OBJECTIVES: Exercise is known to cause changes in the concentration of salivary components such as amylase, Na, and Cl. The aim of this investigation was to evaluate the effect of physical exercise on the levels of trace elements and electrolytes in whole (mixed) saliva. METHODS: Forty subjects performed a maximal exercise test on a cycle ergometer. Samples of saliva were obtained before and immediately after the exercise test. Sample concentrations of Fe, Mg, Sc, Cr, Mn, Co, Cu, Zn, Se, Sr, Ag, Sb, Cs, and Hg were determined by inductively coupled plasma mass spectrometry and concentrations of Ca and Na by atomic absorption spectrometry. RESULTS: After exercise, Mg and Na levels showed a significant increase (p < 0.05) while Mn levels fell (p < 0.05). Zn/Cu molar ratios were unaffected by exercise. CONCLUSIONS: Intense physical exercise induced changes in the concentrations of only three (Na, Mg, and Mn) of the 16 elements analysed in the saliva samples. Further research is needed to assess the clinical implications of these findings.

An on-line immunoassay method for theophylline using a protein A immunoreactor.

A heterogeneous fluorescence immunoassay for theophylline has been automated using a flow injection analysis system containing a protein A solid phase reactor to separate antibody-bound and unbound fluorescein-theophylline. For each sample the antibody-protein A reaction takes place at near neutral pH, and the complexes are eluted at acid pH. The antibody-binding capacity of the reaction greatly exceeds the antibody level in each sample incubation mixture, and a single reactor can be repeatedly cycled between neutral and acid pHs. Experimental variations such as reactor size, flow rate, pH values, and reactant concentrations have been studied. Theophylline could readily be determined at the micrograms ml-1 level with on-line incubation with antibodies.