Development and application of a mini-QuEChERS method for the determination of pesticide residues in anuran adipose tissues.

The expansion of monocultures to regions close to conservation areas has put biodiversity at risk, mainly due to the intense use of pesticides. Anurans are highly susceptible to pesticides and may be a biological marker in the contamination of an area. However, methods for determining pesticides in anurans are incipient. In this work, a miniaturized QuEChERS method was developed for the extraction of atrazine, chlorpyrifos, α- and ß-endosulfan, α-, ß-, θ- and ζ-cypermethrin in anuran adipose tissues. The method was optimized for the tissue sample size scale according to sample mass availability. Extracting solvent and adsorbents for the clean-up step was evaluated, achieving recoveries next to 100% with acetonitrile and without a clean-up step. The mini-QuEChERS method, using 500 mg of adipose tissue, 50 mg of NaCl and 200 mg of MgSO4, 100 µL of ultrapure water, and 1.50 mL of acetonitrile with no purification step, followed by high-performance liquid chromatography analysis and photodiode array detection was validated following the European Community guidelines. The methodology showed a moderate matrix effect for some pesticides, which was corrected using the matrix-matched calibration. The limits of quantification for the pesticide residues in adipose tissues ranged from 10 to 75 µg kg-1. Pesticide recoveries ranged from 74% to 115%, and repeatability and within-lab reproducibility showed relative standard deviations < 11%. The mini-QuEChERS method was applied to extract pesticide residues from the adipose tissues of two species of anurans: Leptodactylus macrosternum and Scinax x-signatus. 25% of samples were positive, detecting endosulfan and chlorpyriphos, confirmed by liquid chromatography coupled to tandem mass spectrometry. The mini-QuEChERS was a simple, economical, and eco-friendly method for extracting pesticide residues in anuran adipose tissue samples.

The exposure in ovo of embryos belonging to Amazonian turtle species Podocnemis expansa (Testudines) to commercial glyphosate and fipronil formulations impairs their growth and changes their skeletal development.

Pesticides are widely used in agricultural production; moreover, they can have direct and indirect effect on both flora and fauna. Aquatic organisms, among other animals, including reptiles, are mainly susceptible to contamination effects. Accordingly, the aim of the present study is to test the hypothesis that the incubation of Podocnemis expansa eggs in substrate added with glyphosate and fipronil formulations changes their viability, interferes with their growth and induces bone alterations. Eggs collected in natural environment were artificially incubated in sand moistened with water added with glyphosate Atar 48, at concentrations of 65 or 6500 µg/L (groups G1 and G2, respectively), and with fipronil Regent 800 WG at 4 or 400 µg/L (groups F1 and F2, respectively) or, yet, with the combination of 65 µg/L glyphosate and 4 µg/L fipronil, or with 6500 µg/L glyphosate and 400 µg/L fipronil (groups GF1 and GF2, respectively). The level of exposure to the herein assessed pesticides was quantified at the end of the incubation period; it was done by dosing its concentration in eggshells. Eggs exposed to the tested pesticides did not have their viability affected by it; however, all embryos exposed to the tested pesticides showed lowered body mass at hatch, as well as impaired development. In addition, bone malformation in the scleral ossicular ring was observed in individuals in groups F1, F2 and GF1. Pesticides accumulated in eggshells at concentrations related to exposure level. Thus, the recorded results have evidenced some remarkably relevant, and previously unknown, impacts associated with the exposure of a species listed as lower risk/conservation dependent, which spends most of its life in the water, to two widely used pesticides, at a very sensitive stage of its life, namely: egg incubation on land.

Disposable pipette extraction using a selective sorbent for carbendazim residues in orange juice.

In this work, a selective sorbent for carbendazim was prepared by the thermal immobilization of the poly(glycidoxypropylmethyl-co-dimethylsiloxane), PGDMS, on silica. The lab-made Si(PGDMS) sorbent was physicochemically and morphologically characterized and was used into the pipette tip for the disposable pipette extraction (DPX) of carbendazim residues of orange juice. The DPX method was optimized from a central composite design and validated according to the SANTE/11813/2017 document recommended by the European Union. The proposed method presented recoveries between 93 and 110% with RSD <16% and the limit of quantification below the MRL for carbendazim in citrus. The whole sample preparation process was carried out at less than 3 min, with good accuracy and precision without the need of any cleanup step or electrical equipment, allowing its portability for the extraction of carbendazim residues in orange juice.

Preparation and Characterization of Cellulose Triacetate as Support for Lecitase Ultra Immobilization.

The use of polymers as supports for enzyme immobilization is a strategy that enables to remove the enzymes from a chemical reaction and improve their efficiency in catalytic processes. In this work, cellulose triacetate (CTA) was used for physical adsorption of phospholipase Lecitase ultra (LU). CTA is more hydrophobic than cellulose, shows good performance in the lipases immobilization being a good candidate for immobilization of phospholipases. We investigated the immobilization of LU in CTA, the stability of the immobilized enzyme (CTA-LU) and the performance of CTA-LU using soybean oil as a substrate. LU was efficiently immobilized in CTA reaching 97.1% in 60 min of contact with an enzymatic activity of 975.8 U·g-1. The CTA-LU system presents good thermal stability, being superior of the free enzyme and increase of the catalytic activity in the whole range of pH values. The difference observed for immobilized enzyme compared to free one occurs because of the interaction between the enzyme and the polymer, which stabilizes the enzyme. The CTA-LU system was used in the transesterification of soybean oil with methanol, with the production of fatty acid methyl esters. The results showed that CTA-LU is a promising system for enzymatic reactions.

Análise de risco de contaminação de águas subterrâneas por resíduos de agrotóxicos no município de Campo Novo do Parecis (MT), Brasil / Risk analysis of groundwater contamination by pesticide residues in Campo Novo do Parecis (MT), Brazil

RESUMO Considerando a possibilidade de que os agrotóxicos utilizados nas principais culturas, como soja, algodão e milho, podem gerar resíduos que contaminam o meio ambiente, faz-se necessária uma análise preliminar sobre a possibilidade de contaminação das águas subterrâneas por resíduos de agrotóxicos utilizados nas principais regiões agrícolas do país. Uma dessas áreas agrícolas se localiza na cidade de Campo Novo do Parecis, Mato Grosso, região de estudo deste trabalho. Os principais agrotóxicos aplicados nas diferentes culturas da região foram avaliados quanto às suas propriedades físico-químicas, seus potenciais de contaminação de acordo com critérios da Agência de Proteção Ambiental dos Estados Unidos e pelo índice de vulnerabilidade de águas subterrâneas (GUS). As informações obtidas foram empregadas para avaliar o risco de contaminação das águas subterrâneas. De acordo com os resultados encontrados, a região apresenta risco real de contaminação ambiental por resíduos de agrotóxicos, uma vez que 45,6% dos agrotóxicos comumente empregados na agricultura local são classificados como extremamente tóxicos ou altamente tóxicos. Além disso, vários desses ingredientes aditivos, cerca de 22%, já foram detectados em diferentes compartimentos ambientais de outras regiões do estado de Mato Grosso.

ABSTRACT Considering the possibility that pesticides used in main crops, such as soybeans, cotton and corn, generate toxic residues that contaminate the environment, a preliminary analysis of the possibility of groundwater contamination is necessary in the main agricultural regions in Brazil. One of these agricultural areas is located in Campo Novo do Parecis, Mato Grosso, the study area of this work. The main pesticides used in different cultures of this region were evaluated considering their physicochemical properties, their contamination potentials according to criteria of the Environmental Protection Agency of the United States and the groundwater ubiquity score index (GUS). The information obtained was used to assess the risk of groundwater contamination. According to the results, the region presents a real risk of environmental contamination by pesticide residues, since 45.6% of the pesticides commonly used in local agriculture are classified as extremely toxic or highly toxic. In addition, several of these pesticides, about 22%, have been detected in different environmental compartments from other regions of the state of Mato Grosso, Brazil.