Identification of flunixin glucuronide and depletion of flunixin and its marker residue in bovine milk

Residues of flunixin [and its marker residue 5‐hydroxyflunixin (5OHFLU)] were determined in milk from cows that intravenously received therapeutic doses of the drug. The samples were collected during each milking (every 12 h) for six consecutive days, and concentrations of flunixin and its metabolites were determined by the method with and without enzymatic hydrolysis (beta‐glucuronidase). The highest flunixin concentration in milk was observed 12 h after dosing (2.4 ± 1.42 μg/kg, mean ± SD). Flunixin concentrations in the samples determined with enzymatic hydrolysis were significantly higher (P < 0.05), which suggests the transfer of flunixin glucuronide to the milk. Additionally, unambiguous identification of flunixin glucuronide in the bovine milk was performed with linear ion‐trap mass spectrometry. The 5OHFLU concentrations analyzed without enzymatic hydrolysis (22.3 ± 16.04 μg/kg) were similar to this obtained with enzymatic hydrolysis. Flunixin and 5OHFLU concentrations dropped below the limits of detection at 48 h after last dosing.     

In Vitro Studies on Atrazine Effects on Human Intestinal Cells

Considering the importance of the oral route for human exposure to atrazine, we have investigated the possible effect of this herbicide on the human intestinal cells and the integrity of the epithelial barrier, using Caco-2 cells as the intestinal model in vitro. We evaluated possibile cytotoxic and genotoxic effects of atrazine in concentrations ranging from 1 to 250 μM on the Caco-2 cells at different stages of growth after short- and long-term exposure. Results from the tetrazolium blue (MTT) test and the Trypan blue exclusion assay showed that atrazine cytotoxicity was dose- and time-dependent. Obtained data indicated that atrazine at high concentrations (50 and 250 μM) was able to induce effects on Caco-2 proliferation and viability. Moreover, it was found that the long-term exposure to atrazine at the non-cytotoxic dose caused inhibition of the intestinal cell maturation and decreased the transepithelial electrical resistance, the indicator of the epithelial barrier integrity. Studies on the atrazine genotoxicity determined using the single cell microelectrophoresis assay indicated that atrazine did not induce DNA damages in the Caco-2 cells at concentrations of up to 50 μM, whereas enhancement in the DNA damage was observed at 250 μM. Altogether, our results indicate that atrazine at expected human oral exposure concentrations is not able to induce effects on the Caco-2 cell proliferation and viability, but may suppress the intestinal cell differentiation and reduce the cell monolayer integrity. We suggest that chronic exposure on low levels of atrazine may lead to alteration in the expression of the morphological and functional features of the Caco-2 cells related to the transport and barrier function of small intestinal enterocytes. In consequence, this may lead to alterations in the intestinal absorption process.     

Thin-layer chromatography of tocopherols of oil seeds

Summary The thin- layer chromatography technique on Silicagel G/250 and with chloroform as solvent, has been adapted for separation and estimation of tocopherols in linseed, rapeseed and soybean. The isolation of an additional tocopherol with Rf value between and tocopherol has been achieved in all investigated seeds. Moreover, linseed has been found to contain only traces of tocopherol, the main amount, till now determined as tocopherol has been localised in the new fraction.The absorption maximum of this fraction has been found to be at 293 mµ. It gives positive reaction with Emmerie-Engel and Further-Meyer reagent and is more susceptible to oxidation than other tocopherols found in the benzene solution of unsaponified matter of oil of investigated seeds. It is supposed that the new tocopherol is identical with 5.7,8-tocotrienol, i.e. with zetatocopherol. It has been stated, that this fraction is separable only, when the chromatography cell is full saturated with solvent vapors and the adsorbent is long time activated at 130°C. The content of this tocopherol in investigated oil seeds has been determined.

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Zusammenfassung Die DC-Technik auf Kieselgel G (250 µ) mit Chloroform als Lösungsmittel wurde zur Trennung und zur quantitativen Analyse der Tocopherole in Samen von Raps, Lein und Soja angewandt. Bei allen untersuchten Samen erhielt man die Trennung eines zusätlichen Tocopherols mit einem Rf Zwischenwert (zwischen den alpha- und den gamma-Tocopherolen). Ferner wurde festgestellt, da Leinsamen nur Spuren von alpha-Tocopherol enthält, daß der Hauptanteil, der bis jetzt als alpha-Tocopherol bestimmt werden konnte, sich in dieser neuen Fraktion befindet. Die maximale Absorbierung dieser Fraktion liegt bei 293 mµ. Sie gibt eine positive Reaktion mit dem Reagenz Emmerie-Engel und Further-Meyer und ist leichter oxidierbar als die anderen Tocopherole, die man in der Benzollösung des Unverseifbaren der untersuchten Samenöle fand. Es ist zu vermuten, daß das neue Tocopherol identisch ist mit 5,7,8-Tocotrienol, das heißt mit dem zeta-Tocopherol. Es wurde gezeigt, daß diese Fraktion nur getrennt werden kann, wenn die chromatographische Zelle mit Lösungsmitteldämpfen gesättigt ist und der Adsorptionsstoff einer längeren Aktivierung bei 130°C unterzogen wurde. Der Gehalt der Ölsamen an diesem Tocopherol wurde untersucht.

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Résumé La technique de la CCM sur Silicagel G (250 g), avec du chloroforme comme solvant a été adaptée pour la séparation et l'analyse quantitative des tocopherols dans les graines de lin, de colza et de soja. La séparation d'un tocopherol supplémentaire avec une valeur de Rf intermédiaire entre celles des alpha et des gamma tocopherols a été obtenue dans toutes les graines analysées. De plus il a été établi que les graines de lin ne contenaient des alphatocopherols qu'à l'état de traces, la majeure partie de la teneur, déterminée à ce jour comme des alpha-tocopherols, se trouvant dans cette nouvelle fraction.L'absorption maximum de cette fraction a été obtenue à 193 mµ. Elle donne une réaction positive au réactif de Emmerie-Engel et Further-Meyer, et est plus facilement oxydable que les autres tocopherols trouvés dans la solution benzenique de l'insaponifiable des huiles des graines étudieées. On suppose que ce nouveau tocopherol est identique au 5.7,8-tocotrienol, c'est-à-dire, avec le zetatocopherol. Il a été montré que cette fraction n'est séparable que lorsque la cellule chromatographique est saturée de vapeurs du solvant et que l'adsorbant a subi une longue activation à 130°C. La teneur des graines oléagineuses en ce tocopherol a été étudiée.

     

Unmanned aerial vehicle canopy reflectance data detects potassium deficiency and green peach aphid susceptibility in canola

There is growing evidence that potassium deficiency in crop plants increases their susceptibility to herbivorous arthropods. The ability to remotely detect potassium deficiency in plants would be advantageous in targeting arthropod sampling and spatially optimizing potassium fertilizer to reduce yield loss due to the arthropod infestations. Four potassium fertilizer regimes were established in field plots of canola, with soil and plant nutrient concentrations tested on three occasions: 69 (seedling), 96 (stem elongation), and 113 (early flowering) days after sowing (DAS). On these dates, unmanned aerial vehicle (UAV) multi-spectral images of each plot were acquired at 15 and 120 m above ground achieving spatial (pixel) resolutions of 8.1 and 65 mm, respectively. At 69 and 96 DAS, field plants were transported to a laboratory with controlled lighting and imaged with a 240-band (390–890 nm) hyperspectral camera. At 113 DAS, all plots had become naturally infested with green peach aphids (Hemiptera: Aphididae), and intensive aphid counts were conducted. Potassium deficiency caused significant: (1) increase in concentrations of nitrogen in youngest mature leaves, (2) increase in green peach aphid density, (3) decrease in vegetation cover, (4) decrease in normalized difference vegetation indices (NDVI) and decrease in canola seed yield. UAV imagery with 65 mm spatial resolution showed higher classification accuracy (72–100 %) than airborne imagery with 8 mm resolution (69–94 %), and bench top hyperspectral imagery acquired from field plants in laboratory conditions (78–88 %). When non-leaf pixels were removed from the UAV data, classification accuracies increased for 8 mm and 65 mm resolution images acquired 96 and 113 DAS. The study supports findings that UAV-acquired imagery has potential to identify regions containing nutrient deficiency and likely increased arthropod performance.     

Biological Denitrification of High Nitrate Processing Wastewaters from Explosives Production Plant

Wastewater samples originating from an explosives production plant (3,000 mg N l⁻¹ nitrate, 4.8 mg l⁻¹ nitroglycerin, 1.9 mg l⁻¹ nitroglycol and 1,200 mg l⁻¹ chemical oxygen demand) were subjected to biological purification. An attempt to completely remove nitrate and to decrease the chemical oxygen demand was carried out under anaerobic conditions. A soil isolated microbial consortium capable of biodegrading various organic compounds and reduce nitrate to atmospheric nitrogen under anaerobic conditions was used. Complete removal of nitrates with simultaneous elimination of nitroglycerin and ethylene glycol dinitrate (nitroglycol) was achieved as a result of the conducted research. Specific nitrate reduction rate was estimated at 12.3 mg N g⁻¹ VSS h⁻¹. Toxicity of wastewater samples during the denitrification process was studied by measuring the activity of dehydrogenases in the activated sludge. Mutagenicity was determined by employing the Ames test. The maximum mutagenic activity did not exceed 0.5. The obtained results suggest that the studied wastewater samples did not exhibit mutagenic properties.