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Citrus flavanones may improve oxidative stress and insulin resistance induced by western diets. However, there is a paucity of studies investigating the change in protein expression levels. This study evaluated the protection and the mechanisms of action of citrus flavanone metabolites, hesperetin 7-glucuronide (H7G) and 3-(4'-hydroxyphenyl) propanoic acid (PA), on pancreatic ß-cell function under oxidative stress induced by cholesterol using the global proteomics approach. Cholesterol induced changes in the global proteomic profile in the pancreatic ß-cell line Min6. On the other hand, proteomics analysis identified 254 proteins differentially expressed with H7G and 352 with PA treatments, most of them were opposite to the changes induced by cholesterol. Bioinformatics analysis showed that these proteins are implicated in cell functions like cell signaling (insulin signaling, p30MAPK signaling, and others), metabolism (glucokinase and glutathione metabolisms), and inflammation pathways (TNF-α and NF-κB pathways). Also, the results of molecular docking suggest that H7G and PA could bind to putative transcription factors (PPAR-γ, STAT-3, CREB1, NF-κB, NFYA) and cell signaling proteins (IKK, RAS, Pi3K, ERK), which results in changes in protein expression observed. Altogether, these data suggest that the treatment with H7G and PA protects pancreatic ß-cells against stress induced by cholesterol through multi-proteomic mechanisms of action.

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Cholesterol is one of the triggers of oxidative stress in the pancreatic-ß cell, generating high levels of reactive oxygen species, which leads to impairment of insulin synthesis and secretion. Bioactive compounds, such as citrus flavanones, which possess anti-inflammatory and antioxidant activities, could reduce oxidative stress in ß-cells and improve their function. We describe for the first time the protective effects of the phase-II flavanone metabolites [naringenin 7-O-glucuronide, hesperetin 3'-O-glucuronide, and hesperetin 7-O-glucuronide], and two flavanones-catabolites derived from gut microbiota metabolism [hippuric acid and 3-(4-hydroxyphenyl)propionic acid], on pancreatic ß-cell line MIN6 under oxidative stress, at physiologically relevant concentration. Cholesterol reduced cell viability in a dose and time-dependent manner, with an improvement in the presence of the metabolites. Moreover, flavanone metabolites attenuated oxidative stress by reducing levels of lipid peroxides, superoxide anions, and hydrogen peroxide. In response to the reduction of reactive oxygen species, a decrease in superoxide dismutase and glutathione peroxidase activities was observed; these activities were elevated by cholesterol. Moreover, all the flavanone metabolites improved mitochondrial function and insulin secretion, and reduced apoptosis. Flavanone metabolites were found uptake by ß-cells, and therefore could be responsible for the observed protective effects. These results demonstrated that circulating phase-II hesperetin and naringenin metabolites, and also phenolics derived from gut microbiota, protect pancreatic-ß cells against oxidative stress, leading to an improvement in ß-cell function and could be the bioactive molecules derived from the citrus consumption.

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Herein, an organic-inorganic hybrid molecularly imprinted polymer (MIP) was successfully synthesized with albendazole sulfoxide (ABZSO) as a template and 3-(trimethoxysilyl)propyl methacrylate, a bifunctional group compound, as a single cross-linking agent. In this study, a simple method using HPLC-DAD was developed for the determination of ABZSO enantiomers in human urine using pipette tip-based molecularly imprinted polymer solid phase extraction (PT-MIP-SPE). Enantioseparation with satisfactory retention times (5.17 and 7.09 min), acceptable theoretical plates (N = 4,535 and 5,091) and strong resolution (Rs = 5.45) was performed with an Agilent® Eclipse Plus C18 (100 mm × 4.6 mm, 3.5 µm) coupled with a Chiralpak® IA column (100 mm × 4.6 mm, 3 µm), a mixture with ethanol:water (50:50, v/v) as the mobile phase, temperature at 40°C, flow rate at 0.9 mL min-1 and λ = 230 nm. Thereafter, certain parameters affecting the PT-MIP-SPE were investigated in detail and the better conditions were: 300 µL of water as washing solvent, 500 µL of ethanol:acetic acid (9:1, v/v) as eluting solvent, 20 mg of MIP, 500 µL of human urine at pH 9 and no addition of NaCl. Recoveries/relative standard deviation (RSD%) for (R)-(+)-ABZSO and (S)-(-)-ABZSO were 78.2 ± 0.2% and 69.7 ± 1.7%, respectively.

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This study aimed to show that the physicochemical proprieties obtained by Fourier transform infrared spectroscopy (FTIR), thermogravimetry (TG), and scanning electronic microscopy (SEM) can be useful tools for evaluating the quality of active pharmaceutical ingredients (APIs) and pharmaceutical products. In addition, a simple, sensitive, and efficient method employing HPLC-DAD was developed for simultaneous determination of lidocaine (LID), ciprofloxacin (CFX) and enrofloxacin (EFX) in raw materials and in veterinary pharmaceutical formulations. Compounds were separated using a Gemini C18 (250 mm × 4.6 mm, 5 µm) Phenomenex® column, at a temperature of 25 °C, with a mobile phase containing 10 mM of phosphoric acid (pH 3.29): acetonitrile (85.7:14.3, v/v) and a flow rate of 1.5 mL/min. Physicochemical characterization by TG, FTIR, and SEM of raw materials of LID, CFX, and EFX provided information useful for the evaluation, differentiation, and qualification of raw materials. Finally, the HPLC method was proved to be useful for evaluation of raw material and finished products, besides satisfying the need for an analytical method that allows simultaneous determination of EFX, CFX, and LID, which can also be extended to other matrices and applications.

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A simple method using HPLC-DAD was developed for the determination of fluoroquinolones in human urine including ciprofloxacin (CIPRO), enrofloxacino (ENRO), marbofloxacino (MARBO) and norfloxacin (NOR). In addition, it was studied the extraction of fluoroquinolones in human urine samples using pipette tip-based molecularly imprinted polymers solid phase extraction (PT-MIPs-SPE). With the goal of finding the best procedure for extraction of four fluoroquinolones in human urine, several parameters that are likely to affect the efficiency of extraction during sample preparation, including the washing solvent, type and volume of eluent, amount of material, the volume of the sample, pH and the ionic strength were systematically optimized. Chromatographic separations of fluoroquinolones were hit within 10min using a Synergi(®) C18 (250×4.6mm, 4µm) column and mobile phase consisting of water (10mM of phosphoric acid, the pH adjusted at 3.29 with triethylamine) : acetonitrile (85.7: 14.3, v/v) at a flow rate of 1.5mLmin(-1). Detection was performed at 290nm. The average extraction recoveries/standard deviation relative to ENRO, CIPRO, NOR and MARBO were 96.40±5.51%, 42.47±4.81%, 41.82±7.99% and 87.49±4.70, respectively. The method was liner from 39 to 1260ngmL(-1) for each fluoroquinolone with correlation coefficient of 0.9904, 0.9910, 0.9914 and 0.9919, to ENRO, CIPRO, NOR and MARBO, respectively. The assays of within-day and between-day precision and accuracy for all analytes were studied at three concentration levels and were lower than 15%. The method was successfully employed in a preliminary cumulative urinary excretion study after administration of CIPRO to a healthy volunteer.

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