BACKGROUND Computational research plays an important role in predicting the chemical and physical properties of biologically active compounds important in future structural modifications to improve or modify biological activity. OBJECTIVE This research focuses on quantum chemical and spectroscopic investigations properties of synthesized 4-hydroxycoumarin derivatives. METHODS Quantum chemical calculations were obtained using B3LYP, HF, and M06-2x level methods with the 6-31++G (d,p) basis set. Afterward, IR, 1H, 13C, UV-Visible experimentally parameters were compared with the results obtained using the B3LYP/6-31+G*(d) basis set of the molecules to be able to characterize the structures. RESULTS Based on the quantum chemical calculations compound with acetamido group on the phenyl ring is the most reactive, and compound with nitro substituent is the least reactive and the the strongest electrophile among tested compounds. With the exception of compounds with dimethylamino group, all other compounds have a pronounced tautomer between between OH and C = O group. The calculated and experimental values are in agreement with each other. CONCLUSION The molecular structure in the ground state of six 3-cinnamoyl 4-hydroxycoumarin derivatives was optimized using density functional theory. The observed and computed values were compared and it can be concluded that the theoretical results were in good linear agreement with the experimental data.

BACKGROUND Preclinical drug testing requires in vitro and in vivo assessments that are vital for studying drug pharmacokinetics and toxicity. Distinct factors that play an important role in drug screening, such as hydrophobicity, solubility of the substance and serum protein binding can be challenging by inducing result inconsistencies. Hence, establishing accurate methods to quantify drug concentrations in cell cultures becomes pivotal for reliable and reproducible results important for in vivo dosing predictions. OBJECTIVE This research focuses on developing an optimized analytical approach via high-pressure liquid chromatography (HPLC) to determine thymoquinone (TQ) levels in monolayer cell cultures. METHODS The method's validation adheres to the International Council for Harmonisation (ICH) guideline M10, ensuring its acceptance and applicability. Using an HPLC system with a Diode Array Detector (DAD), the study fine-tuned various parameters to achieve an efficient separation of TQ. Validation covered specificity, sensitivity, matrix effects, linearity, precision, and accuracy, alongside assessing TQ stability in RPMI-1640 medium. RESULTS The HPLC method exhibited remarkable TQ specificity, free from interfering peaks at the analyte retention. Sensitivity analysis at the lower limit of quantification (LLOQ) revealed 5.68% %CV and 98.37% % mean accuracy. Matrix effect evaluation showcased accuracy within 85-115%. Linearity spanned in the concentration range of 2-10 μM with a correlation coefficient (r2) of 0.9993. Precision and accuracy were aligned with acceptance criteria. The proposed method was found to be greener in terms of usage of persistent, bioaccumulative, and toxic chemicals and solvents, corrosive samples, and waste production. CONCLUSION The developed HPLC-DAD method emerges as specific, accurate, sensitive, and reliable for TQ determination in cell cultures. It ensures robust TQ quantification, enhancing precise in vitro assessments and dependable dosing predictions for in vivo studies. Further research is advocated to investigate TQ's stability across diverse environmental conditions.

Solvent and substitution effects on the UV/Vis spectroscopic and fluorescence behaviour of seven synthesized 3-substituted 4-hydroxycoumarin derivatives were tested. The tested compounds were dissolved in ethyl acetate, acetonitrile, and dimethyl sulfoxide. Absorption and emission spectra were recorded in the range of 200–800 nm. All tested 4-hydroxycoumarin derivatives showed good absorption in a wide range of 200–550 nm, depending on the properties of the substituents on the benzene ring of the cinnamoyl moiety and the type of solvent. In comparison to the unsubstituted analogue, compounds with an electron-donating group exhibited bathochromically shifted UV/Vis absorption and emission spectra. The highest fluorescence quantum yield was observed for compounds with dimethylamino and acetamido groups as substituents at the benzene ring. Considering that both substitution and solvent affect the absorption and emission spectra of the tested compounds, it can be concluded that judiciously selecting these parameters can improve their absorption and fluorescence properties, making them suitable for various analytical uses.

Skin sensitization is a crucial endpoint in the safety assessment of chemicals, with the Direct Peptide Reactivity Assay (DPRA) emerging as a valuable in chemico method for evaluating a substance's sensitization potential. This review delves into the principles, applicability, and limitations of the DPRA within the context of the Adverse Outcome Pathway (AOP) framework for skin sensitization. We examine the DPRA'srole in addressing the molecular initiating event of skin sensitization, its integration into Integrated Approaches to Testing and Assessment (IATA), and its performance in predicting sensitizers. The review also highlights the challenges in testing certain categories of chemicals and the importance of considering the DPRA's results alongside other complementary methods. By providing a comprehensive overview of the DPRA, this review aims to inform researchers, regulators, and clinicians about its utility and limitations in the context of skin sensitization testing.

The main MoR discussion led to further suggestions on KE terminology, including ensuring coherence to directionality in terms of the KE descriptions (e.g., specifying increase, decrease, altered, no direction, etc.) and clarifying differences in ROS and reactive oxygen and nitrogen species (RONS), and enzymatic and non-enzymatic events. The consortium highlighted the importance of the role of ROS as a KE and an associative event in the AOP framework. Additionally, participants highlighted modification to macromolecules from the resultant RONS generation (e.g., lipid peroxidation) as a relevant endpoint to include in the KE. The possibility of grouping ROS-related KEs in the AOP framework needs to be discussed further.

Aflatoxins M1 and M2 are the hydroxylated metabolites of aflatoxins B1 and B2 and can be found in milk or milk products. The aim of this study was to obtain the exposure and health risk assessment of this toxin in raw milk using a selection of existing test results of different periods in a different region of Bosnia and Herzegovina. For AFM1 intake assessment, the data available from the report of Pre-Accession Assistance To Agriculture And Rural Development Sector Analyses in Bosnia and Herzegovina was used. For the concentration of the aflatoxin M1 in raw milk, the data of the previous studies and unpublished data from the Laboratory of Toxicological Chemistry Faculty of the Pharmacy University of Sarajevo were used. Risk characterization was performed by calculating the risk of hepatocellular carcinoma (HCC) incidence per year for four exposure scenario. The results indicated that the average estimated daily intake of aflatoxin M1 was 0.049 ng/kg body weight (bw) per day. The risk of hepatocellular carcinoma (cases/year/100 000 individuals), depending on the HBsAg prevalence, was relatively low (average 0.0000640.000074 cases/year/100 000). It is concluded that in recent years, the carcinogenic risk of aflatoxin M1 has been low and not critical in Bosnia and Herzegovina.