Carolina Fernández-Ramos

New method for the determination of carbamate and pyrethroid insecticides in water samples using on-line SPE fused core column chromatography.

A new HPLC column-switching method using large volume sample injection and fused-core columns for on-line solid phase extraction have been developed for the determination of the following carbamates and pyrethroids: aldicarb, carbaryl, pirimicarb, carbofuran, kadethrin, flumethrin, fenpropathrin, fenoxycarb, tau-fluvalinate and fenvalerate, in surface water samples. Sudan I was used as internal standard. The proposed method was performed using 100 µl sample injection followed by an on-line solid phase extraction procedure and finally the compounds were identified and quantified by liquid chromatography with ultraviolet detection. The separation was carried out on C-18 reversed phase column based on fused-core particle technology. The influence of the injected sample volume, the variables affecting to SPE process and the conditions for the separation on an analytical column, were studied and optimized. The limits of detection ranged from 5.5 to 8.9 µg L(-1), and limits of quantification from 18.4 to 29.7 µg L(-1), while inter- and intra-day variability was under 15%. This new analytical procedure was satisfactorily applied for the determination of these organic pollutants in surface water samples located in Czech Republic. Concentration levels were found for some of these pollutants up to 26.11 µg L(-1) in the river Elbe and up to 34.53 µg L(-1) in the closed lakes samples.

Phytomonas spp: superoxide dismutase in plant trypanosomes

In 1909, Donovan [1] proposed the creation of a new genus, Phytomonas, to differentiate plant from animal trypanosomatids. Phytomonas spp. live in the latex, sap, sieve tubes, and fruit of many plant species [2]. At first, considerable controversy surrounded the pathogenicity of phytomonads in plants, as initially these parasitic protozoans were not thought to be particularly harmful. However, evidence from a number of plants of great economic significance, such as coffee, coconut, and palm, has shown that Phytomonas infections can have devastating economic consequences [3]. The defence mechanisms of trypanosomatids against the toxic products of O2 reduction – e.g. superoxide anion, hydrogen peroxide or hydroxyl radicals – are not completely understood. Trypanosomatids are protected from the damaging effects of reactive oxygen intermediates by scavengers such as trypanothione and specific enzymes such as superoxide dismutase (SOD, EC 1.15.1.1) and trypanothione peroxidase. Catalase, involved in the elimination of hydrogen peroxide, is absent from most trypanosomatids, although it has been detected in Crithidia luciliae [4] and in Phytomonas spp. [5]. Cyanide-insensitive SOD activity has been reported in Trypanosoma cruzi [6], as well as in other trypanosomatids such as Crithidia fasciculata [7] and Trypanosoma brucei [4]. The SOD activities of Leishmania tropica and T. cruzi are also cyanide-insensitive but peroxide-sensitive [7]. SOD of C. fasciculata is located in the cytosol and exists in three forms, which may represent three distinct isozymes. Comparisons of the amino-acid sequence of this SOD with those of SODs from other sources suggest that the crithidial enzyme is closely related to bacterial FeSOD of the alga Euglena gracilis. SOD activity has also been detected in L. dono ani and Phytomonas spp., although its nature has not been determined [8,9]. Most of the information on the biochemistry of plant flagellates has come from experiments in which the flagellates were used as instruments in biochemical research rather than from studies investigating the biochemistry of the plant flagellates themselves. Probably for this reason, these studies are scattered through a variety of subjects, and, despite the amount of information gathered in recent years, our knowledge of the biochemistry of Phytomonas remains fragmentary [3]. In this sense, the detoxifying mechanisms of oxygen radicals in plant trypanosomatids are unknown. From previous studies [10], we know that plant flagellates have SOD activity, but we have yet to identify the role of this enzyme in the destruction of the superoxide radicals, and there is no information available on the enzyme itself. In the present work, we confirm and quantify the SOD activity in three trypanosomes isolated from different plants: from phloem of the Coco nucifera (Hartrot disease) [11], from latex vessels of Euphorbia characias [12], from the fruits of Lycopersicon esculenAbbre iations: Cu/ZnSOD, copper/zinc-containing superoxide dismutase; FeSOD, iron-containing superoxide dismutase; ME, malic enzyme; MnSOD, manganese-containing superoxide dismutase; NBT, nitro-blue tetrazolium salt; PK, pyruvate kinase; PFK, 6-phospho fructose kinase; SOD, superoxide dismutase. * Corresponding author. Tel.: +34-958-242369; fax: +34-958243174. E-mail address: msanchem@goliat.ugr.es (M. Sanchez-Moreno).

An on-line SPE–HPLC method for effective sample preconcentration and determination of fenoxycarb and cis, trans-permethrin in surface waters

A new on-line SPE-HPLC method using fused-core columns for on-line solid phase extraction and large volume sample injection for increasing the sensitivity of detection was developed for the determination of insecticides fenoxycarb and cis-, trans-permethrin in surface waters. The separation was carried out on fused-core column Phenyl-Hexyl (100×4.6 mm), particle size 2.7 µm with mobile phase acetonitrile:water in gradient mode at flow rate 1.0 mL min(-1), column temperature 45°C. Large volume sample injection (1500 µL) to the extraction dimension using short precolumn Ascentis Express RP C-18 (5×4.6 mm); fused-core particle size 2.7 µm allowed effective sample preconcentration and efficient ballast sample matrix removal. The washing mobile phase consisting of a mixture of acetonitrile:water; 30:70, (v/v) was pumped at flow rate of 0.5 mL min(-1) through the extraction precolumn to the waste. Time of the valve switch for transferring the preconcentrated sample zone from the extraction to the separation column was set at 3rd min. Elution of preconcentrated insecticides from the extraction precolumn and separation on the analytical column was performed in gradient mode. Linear gradient elution started from 40% of acetonitrile at time of valve switch from SPE column (3rd min) to 95% of acetonitrile at 7th min. Synthetic dye sudan I was chosen as an internal standard. UV detection at wavelength 225 nm was used and the method reached the limits of detection (LOD) at ng mL(-1) levels for both insecticides. The method showing on-line sample pretreatment and preconcentration with highly sensitive determination of insecticides was applied for monitoring of fenoxycarb and both permethrin isomers in different surface water samples in Czech Republic. The time of whole analysis including on-line extraction, interferences removal, chromatography separation and system equilibration was less than 8 min.

Determination of alcohol sulfates in wastewater treatment plant influents and effluents by gas chromatography-mass spectrometry.

In the present paper, we developed an accurate method for the analysis of alcohol sulfates (AS) in wastewater samples from wastewater treatment plant (WWTP) influents and effluents. Although many methodologies have been published in the literature concerning the study of anionic surfactants in environmental samples, at present, the number of analytical methodologies that focus in the determination of AS by gas chromatography in the different environmental compartments is limited. The reason for this is that gas chromatography-mass spectrometry (GC-MS) technique requires a previous hydrolysis reaction followed by derivatization reactions. In the present work, we proposed a new procedure in which the hydrolysis and derivatization reactions take place in one single step and AS are directly converted to trimethylsilyl derivatives. The main factors affecting solid-phase extraction (SPE), hydrolysis/derivatization and GC-MS procedures were accurately optimised. Quantification of the target compounds was performed by using GC-MS in selected ion monitoring (SIM) mode. The limits of detection (LOD) obtained ranged from 0.2 to 0.3 μg L(-1), and limits of quantification (LOQ) from 0.5 to 1.0 μg L(-1), while inter- and intra-day variability was under 5%. A recovery assay was also carried out. Recovery rates for homologues in spiked samples ranged from 96 to 103%. The proposed method was successfully applied for the determination of anionic surfactants in wastewater samples from one WWTP located in Granada (Spain). Concentration levels for the homologues up to 39.4 μg L(-1) in influent and up to 8.1 μg L(-1) in effluent wastewater samples.

Sorption and desorption of alcohol sulfate surfactants in an agricultural soil

Alcohol sulfates are one of the most important types of commercial anionic surfactants and may pose serious environmental problems. The present study examines the sorption behavior of alcohol sulfates in an agricultural soil using a batch reactor and column experiments. Kinetics and equilibrium isotherms of the sorption and desorption processes of the following alcohol sulfate homologues have been determined: AS-C12, AS-C14, AS-C16 , and AS-C18. Sorption and desorption occurred rapidly, and equilibrium was achieved in approximately 1 h. The sorbed mass percentages were 35.1% AS-C12 , 67.2% AS-C14 , and 100% for both AS-C16 and AS-C18. A pseudo-first-order kinetic model provided an adequate fit of data. Sorption and desorption equilibrium data were fitted into 1) a linear model, with r(2) values in the ranges 0.712 to 0.988 and 0.736 to 0.983, respectively, and 2) a Freundlich model, with r(2) values in the ranges 0.956 to 0.991 and 0.891 to 0.981, respectively. Continuous-flow experiments in soil columns were carried out to obtain the breakthrough curves for each compound. The present study provides basic theoretical concepts and key parameters for developing mathematical models that simulate the migration of alcohol sulfate into agricultural soils.

Determination of alcohol sulfates and alcohol ethoxysulfates in marine and river sediments using liquid chromatography-tandem mass spectrometry

A novel and successful method has been developed for the identification and quantification of alcohol sulfates (AS) homologues and alcohol ethoxysulfates (AES) ethoxymers in marine and river sediment samples. The method involves the extraction of 5.00 g of dry sample with methanol using pressurized liquid extraction (PLE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). 2-Octylbenzene sulfonic acid sodium salt (2ØC8-LAS) was used as internal standard. The analytical methods were applied to marine sediments collected from the coast of Almeria (South-east Spain) and river sediments collected from the Monachil river (Granada, South-east Spain). For AS homologues, the found limits of detection were 0.04-0.08 μg g(-1) for marine and river sediments. For AES ethoxymers, the found limits of detection were 0.03-0.09 μg g(-1) and 0.06-0.22 μg g(-1) for marine and river sediments, respectively. The highest concentrations of AS and AES were found in river sediment samples. Significant differences were also observed between the behavior of short-chain compounds (C12) and long-chain compounds (C14 to C18). The influence of the physic-chemical properties of water on the occurrence of these compounds was also evaluated, and differences between long- and short-chain compounds were also observed. Additionally, principal components analyses were carried out in order to study the relationship between variables and to evaluate the sources of data variability and behavior patterns. Finally, important conclusions were drawn regarding the environmental behavior of AS and AES.

A new procedure of determination of alcohol sulfates and alcohol ethoxysulfates in agricultural soils

The number of analytical methodologies that focus in the determination of alcohol sulfates (AS) and alcohol ethoxysulfates (AES) in terrestrial environment is very limited. In the present work, a new methodology to improve the extraction and determination of AS and AES in agricultural soil samples has been developed. Prior to instrumental analysis, an extraction procedure using pressurized liquid extraction with methanol (PLE) was carried out in order to obtain the highest recoveries and improve sensitivity. The most influential variables affecting the PLE procedure were optimized. Then, the separation and quantification of analytes were performed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). The limits of detection (LOD) ranged from 0.03 to 0.08μgg(-1) for AS homologues and in the case of AES ethoxymers from 0.03 to 0.09μgg(-1) for AES-C12Ex and from 0.03 to 0.08μgg(-1) for AES-C14Ex. Matrix-matched calibration was used. Trueness was evaluated by using a spike recovery assay with spiked blank samples, and the recoveries ranged from 98.3% to 101.0% for AS and from 99.9% to 100.1% for AES. The method was satisfactorily applied in a field study designed to evaluate the environmental behavior of these compounds in agricultural soil.

Biochemical characterization of a trypanosomatid isolated from the plant Amaranthus retroflexus

A protozoan flagelate has recently been isolated from Amaranthus retroflexus. This plant grows near economically important crops in southeastern Spain, which are known to be parasitized by Phytomonas spp. The present study focuses on the characterization of the energy metabolism of this new isolate. These flagellates utilize glucose efficiently as their primary energy source, although they are unable to completely degrade it. They excrete ethanol, acetate, glycine, and succinate in lower amount, as well as ammonium. The presence of glycosomes was indicated by the early enzymes of the glycolytic pathway, one enzyme of the glycerol pathway (glycerol kinase), and malate dehydrogenase. No evidence of a fully functional citric-acid cycle was found. In the absence of catalase activity, these flagellates showed significant superoxide dismutase activity located in the glycosomal and cytosolic fractions. These trypanosomes, despite being morphologically and metabolically similar to other Phytomonas isolated from the same area, showed significant differences, suggesting that they are phylogenetically different species.