Studies of polyphenols in various natural products have shown their variability and often low amounts. Thus, we aimed to investigate the bioaccessibility of the main polyphenolic groups extracted from two poplar types of propolis collected in continental Croatia. The propolis complex, in the form of water-soluble powder of propolis extract with the carrier 2-hydroxypropyl-β-cyclodextrin (HP-β-CD), was obtained by applying ultrasonic-assisted extraction followed by lyophilization. The in vitro gastrointestinal digestion was conducted on the propolis complex extracts and a mixture of standards (pinocembrin, galangin, chrysin, caffeic acid, and caffeic acid phenylethyl ester). Fractions from the oral, gastric, and intestinal phase samples were analyzed spectrophotometrically by standard methods used to determine the content of total phenols (TPs), flavonols/flavones (TFFs), and flavanones/dihydroflavonols (TFDs). The small-intestinal phase was performed by two models: centrifugation and dialysis. The results of propolis-HP-β-CD extracts demonstrated high polyphenolic bioaccessibility during all three phases of digestion. The bioaccessibility of TPs, TFFs, and TFDs after the intestinal phase with centrifugation was from 0.02 to 8.86 times higher than in the dialysis phase, stressing that it is a better model than dialysis. Our findings show that the inclusion complex of propolis with HP-β-CD enables the good solubility of polyphenol groups.

Disruption of any stage of iron homeostasis, including uptake, utilization, efflux, and storage, can cause progressive damage to peripheral organs. The health hazards associated with occupational exposure to inhalation anesthetics (IA) in combination with chronic iron overload are not well documented. This study aimed to investigate changes in the concentration of essential metals in the peripheral organs of rats after iron overload in combination with IA. The aim was also to determine how iron overload in combination with IA affects tissue metal homeostasis, hepcidin–ferritin levels, and MMP levels according to physiological, functional, and tissue features. According to the obtained results, iron accumulation was most pronounced in the liver (19×), spleen (6.7×), lungs (3.1×), and kidneys (2.5×) compared to control. Iron accumulation is associated with elevated heavy metal levels and impaired essential metal concentrations due to oxidative stress (OS). Notably, the use of IA increases the iron overload toxicity, especially after Isoflurane exposure. The results show that the regulation of iron homeostasis is based on the interaction of hepcidin, ferritin, and other proteins regulated by inflammation, OS, free iron levels, erythropoiesis, and hypoxia. Long-term exposure to IA and iron leads to the development of numerous adaptation mechanisms in response to toxicity, OS, and inflammation. These adaptive mechanisms of iron regulation lead to the inhibition of MMP activity and reduction of oxidative stress, protecting the organism from possible damage.

The purpose of this investigation was to prepare a water-soluble complex by encapsulation of propolis extract with the carrier 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) and to examine the influence of in vitro simulated gastrointestinal digestion (GID) on the content of the main polyphenolic groups when administered as aqueous suspension. The research was conducted on two continental samples of propolis from Croatia and a mixture of selected polyphenol standards. By applying UAE followed by a lyophilization procedure, it was possible to obtain a powder with a high content of polyphenols. The content of total phenols (TPs), total flavonols/flavones (TFFs) and total flavanones/dihydroflavonols (TFDs) in the fractions of oral and gastric phase and small intestinal phase performed with two models: centrifugation and dialysis through a semi-permeable membrane, was determined by three UV/Vis spectrophotometric methods: Folin-Ciocalteu, aluminium complexation and 2,4-dinitrophenylhydrazine method. The obtained results for both propolis complexes showed stability in the polyphenol content and high bioaccessibility during all three phases of digestion. The bioaccessibility of TPs, TFFs and TFDs under in vitro conditions after dialysis through a semipermeable membrane was lower compared to the intestinal phase model with centrifugation.

Florfenicol (FF) is a broad-spectrum antibiotic used to treat gastrointestinal and respiratory infections in domestic animals. Considering FF’s rapid elimination via urine after drug treatment, its use increases concerns about environmental contamination. The objective of the study was to establish a sustainable chromatographic method for simple analysis of FF in pig urine to investigate the urinary excretion of FF after a single intramuscular administration of 20 mg FF/kg body weight. The urine sample was prepared using a centrifuge and regenerated cellulose filter, and the diluted sample was analyzed. The method was validated in terms of linearity, the limit of detection (0.005 µg/mL) and quantitation (0.016 µg/mL), repeatability and matrix effect (%RSD ranged up to 2.5), accuracy (varied between 98% and 102%), and stability. The concentration-time profile of pig urine samples collected within 48 h post-drug administration showed that 63% of FF’s dose was excreted. The developed method and previously published methods used to qualify FF in the urine of animal origin were evaluated by the National Environmental Method Index (NEMI), Green Analytical Procedure Index (GAPI) and Analytical GREENness Metric Approach (AGREE). The greenness profiles of published methods revealed problems with high solvents and energy consumption, while the established method was shown to be more environmentally friendly.

A study of florfenicol (FF) and its metabo- lite florfenicol amine (FFA) in pig cerebrospinal fluid was conducted following repeated intramuscular administration of the original (reference) and a generic veterinary medicinal product (VMP) under the same experimental conditions (20 mg FF/kg body weight, 48-hour interval). Both VMPs are solutions for injection containing FF as an active substance in the concentration of 300 mg/mL and have been authorized in Croatia for use in cattle and pigs. In this study, clinically healthy pigs were randomly divided into three groups. The first group was treated with the reference VMP, the second with the generic VMP, while the third served as the control group. Animals were sacrificed at 216, 288 and 384 hours after the first drug administration. Cerebrospinal fluid samples were analysed by the optimized and validated high-performance liquid chromatography-diode array detector method (HPLC-DAD). The solid-phase extraction (SPE) technique was chosen for sample preparation. The HPLC-DAD method provides good linearity over the concentration range of 0.05 to 5.00 μg/mL for FF and FFA. Limits of detection were 0.0023 μg/mL for FF and 0.0100 μg/mL for FFA. Extraction recoveries of FF were from 86.6% to 111.8%, and of FFA from 91.7% to 98.8%. The SPE-HPLC-DAD method has been demonstrated to be a selective, sensitive and suitable analytical method for the determination of FF and FFA in cerebrospinal fluid. The present study was based on a preliminary study that quantified FF in pig plasma at 216 hours after the first application of reference or generic VMP. However, FF and FFA were not detected in any of the cerebrospinal fluid samples during the experimental period. According to the nature of biological fluids, the SPE-HPLC-DAD method can be suitable for further pharmacokinetic studies of FF in pig plasma and serum after intramuscular administration of VMPs.