Spectroscopic Studies of Quinobenzothiazine Derivative in Terms of the In Vitro Interaction with Selected Human Plasma Proteins. Part 1

Plasma proteins play a fundamental role in living organisms. They participate in the transport of endogenous and exogenous substances, especially drugs. 5-alkyl-12(H)-quino[3,4-b][1,4]benzothiazinium salts, have been synthesized as potential anticancer substances used for cancer treatment. Most anticancer substances generate a toxic effect on the human body. In order to check the toxicity and therapeutic dosage of these chemicals, the study of ligand binding to plasma proteins is very relevant. The present work presents the first comparative analysis of the binding of one of the 5-alkyl-12(H)-quino[3,4-b][1,4]benzothiazinium derivatives (Salt1) with human serum albumin (HSA), α-1-acid glycoprotein (AGP) and human gamma globulin (HGG), assessed using fluorescence, UV-Vis and CD spectroscopy. In order to mimic in vivo ligand–protein binding, control normal serum (CNS) was used. Based on the obtained data, the Salt1 binding sites in the tertiary structure of all plasma proteins and control normal serum were identified. Both the association constants (K_a) and the number of binding site classes (n) were calculated using the Klotz method. The strongest complex formed was Salt1–AGP_complex (K_a = 7.35·10⁴ and 7.86·10⁴ mol·L⁻¹ at excitation wavelengths λ_ex of 275 and 295 nm, respectively). Lower values were obtained for Salt1–HSA_complex (K_a = 2.45·10⁴ and 2.71·10⁴ mol·L⁻¹) and Salt1–HGG_complex (K_a = 1.41·10⁴ and 1.33·10⁴ mol·L⁻¹) at excitation wavelengths λ_ex of 275 and 295 nm, respectively, which is a positive phenomenon and contributes to the prolonged action of the drug. Salt1 probably binds to the HSA molecule in Sudlow sites I and II; for the remaining plasma proteins studied, only one binding site was observed. Moreover, using circular dichroism (CD), fluorescence and UV-Vis spectroscopy, no effect on the secondary and tertiary structures of proteins in the absence or presence of Salt1 has been demonstrated. Despite the fact that the conducted studies are basic, from the scientific point of view they are novel and encourage further in vitro and in vivo investigations. As a next part of the study (Part 2), the second new synthetized quinobenzothiazine derivative (Salt2) will be analyzed and published.     

A New Pyrimidine Schiff Base with Selective Activities against Enterococcus faecalis and Gastric Adenocarcinoma

Enterococcus faecalis is known as a significant nosocomial pathogen due to its natural resistance to many antibacterial drugs. Moreover, it was found that E. faecalis infection causes inflammation, production of reactive oxygen species, and DNA damage to human gastric cancer cells, which can induce cancer. In this study, we synthesized and tested the biological activity of a new Schiff base, 5-[(4-ethoxyphenyl)imino]methyl-N-(4-fluorophenyl)-6-methyl-2-phenylpyrimidin-4-amine (3), and compared its properties with an analogous amine (2). In the biological investigation, 3 was found to have antibacterial activity against E. faecalis 29212 and far better anticancer properties, especially against gastric adenocarcinoma (human Caucasian gastric adenocarcinoma), than 2. In addition, both derivatives were non-toxic to normal cells. It is worth mentioning that 3 could potentially inhibit cancer cell growth by inducing cell apoptosis. The results suggest that the presence of the –C=N– bond in the molecule of 3 increases its activity, indicating that 5-iminomethylpyrimidine could be a potent core for further drug discovery research.     

Polymorphism and Conformational Equilibrium of Nitro-Acetophenone in Solid State and under Matrix Conditions

Conformational and polymorphic states in the nitro-derivative of o-hydroxy acetophenone have been studied by experimental and theoretical methods. The potential energy curves for the rotation of the nitro group and isomerization of the hydroxyl group have been calculated by density functional theory (DFT) to estimate the barriers of the conformational changes. Two polymorphic forms of the studied compound were obtained by the slow and fast evaporation of polar and non-polar solutions, respectively. Both of the polymorphs were investigated by Infrared-Red (IR) and Raman spectroscopy, Incoherent Inelastic Neutron Scattering (IINS), X-ray diffraction, nuclear quadrupole resonance spectroscopy (NQR), differential scanning calorimetry (DSC) and density functional theory (DFT) methods. In one of the polymorphs, the existence of a phase transition was shown. The position of the nitro group and its impact on the crystal cell of the studied compound were analyzed. The conformational equilibrium determined by the reorientation of the hydroxyl group was observed under argon matrix isolation. An analysis of vibrational spectra was achieved for the interpretation of conformational equilibrium. The infrared spectra were measured in a wide temperature range to reveal the spectral bands that were the most sensitive to the phase transition and conformational equilibrium. The results showed the interrelations between intramolecular processes and macroscopic phenomena in the studied compound.     

Electrochemical properties of positive electrode in lead-acid battery modified by ammonium-based ionic liquids

The influence of selected types of ammonium ionic liquid (AIL) additives on corrosion and functional parameters of lead-acid battery positive electrode was examined. AILs with a bisulfate anion used in the experiments were classified as protic, aprotic, monomeric, and polymeric, based on the structure of their cation. Working electrodes consisted of a lead-calcium-tin alloy utilized in the industry for manufacturing current collectors of positive electrodes in lead-acid batteries (LABs). This alloy was used in the first part of the study for the evaluation of corrosion intensity and stability of electrolyte with AIL additives. In the second part, the grid made of the examined alloy was covered with positive active mass, the composition of which was modified by selected AIL. The selection of an appropriate substance was made based on parameters such as corrosion potential, corrosion current, polarization resistance, and hydrogen and oxygen evolution potentials. Techniques such as linear sweep voltammetry, corrosimetry, and electrochemical impedance spectroscopy were used. The conducted measurements revealed that polymeric AIL additive had an exceptionally positive influence on the inhibition of the corrosion process in LAB and electrochemical window of the electrolyte. The influence of this compound on the specific capacity and resistance of the active mass was also discussed.

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Synthesis of a cisplatin derivative from lithocholic acid

A new steroidal-platinum(II) hybrid compound was synthesized using a simple and efficient methodology. The synthesis was performed by a convergent approach with cross metathesis (CM) as a key step. An olefin derived from lithocholic acid and a vinyl substituted ethylenediamine derived from l-serine were used as chiral building blocks, which were combined in the CM step. The most important advantage of this method was the utilization of l-serine as a cheap, stereoisomerically pure substrate. A steroid with a diamino system in the side chain was subjected to reaction with potassium tetrachloroplatinate to obtain the target platinum(II) complex. Attempts to synthesize similar diamine systems using the asymmetric Strecker reaction were unsuccessful.     

Application of phosphonium deep eutectic solvents as extractants in ultrasound-assisted dispersive liquid–liquid microextraction for preconcentration of trace amounts of herbicides in drainage ditches waters

In this study, an efficient preconcentration method was presented that is based on dispersive liquid-liquid microextraction taking the advantage of newly synthesized phosphonium deep eutectic solvents used as extractants and ultrasound probe as a dispersing agent. The extracts obtained were analyzed by high-performance liquid chromatography. To optimize the five most important factors for the microextraction procedure a central composite design plan was used. Under optimal conditions (140 μl of extractant, 60 mg of NaCl, pH = 2.0, 120 s of extraction time with ultrasound probe as the dispersing agent, 16 min of centrifugation for phase separation), the proposed method allowed to achieve good precision with RSD between 3.2% and 9.7% at 1.0, 5.0 and 40.0 ng ml levels. The preconcentration factors were equal to 42, 39, and 41, and the limits of detection 0.128, 0.103, and 0.135 ng/ml for dicamba, 2-methyl-4-chlorophenoxyacetic acid, and 2-methyl-4-chlorophenoxypropionic acid, respectively. The proposed method was successfully applied for the determination of chlorophenoxy acid herbicides in water samples from drainage ditches with a good recovery in the range of 70%–93%.     

X-ray studies of three 3,6-bis(pyridin-2-yl)-1,2,4,5-tetrazine cocrystals: an unexpected molecular conformation stabilized by hydrogen bonds

The results of the X-ray structure analysis of three novel 3,6-bis(pyridin-2-yl)-1,2,4,5-tetrazine cocrystals are presented. These are 3,6-bis(pyridin-2-yl)-1,2,4,5-tetrazine–2,4,6-tribromophenol (1/2), C₁₂H₈N₆·2C₆H₃Br₃O, 3,6-bis(pyridin-2-yl)-1,2,4,5-tetrazine–isonicotinic acid N-oxide (1/2), C₁₂H₈N₆·2C₆H₅NO₃, and 3,6-bis(pyridin-2-yl)-1,2,4,5-tetrazine–4-nitrobenzenesulfonamide (1/1), C₁₂H₈N₆·C₆H₆N₂O₄S. Special attention is paid to a conformational analysis of the title tetrazine molecule in known crystal structures. Quantum chemistry methods are used to compare the energetic parameters of the investigated conformations. A structural analysis of the hydrogen and halogen bonds with acceptor aromatic tetrazine and pyrazine rings is conducted in order to elucidate factors responsible for conformational stability.     

Cellular Mechanistic Considerations on Cytotoxic Mode of Action of Phosphino Ru(II) and Ir(III) Complexes

Two piano-stool ruthenium(II) complexes Ru(η⁶-p-cymene)Cl₂PPh₂CH₂OH ( RuPOH ) and Ru(η⁶-p-cymene)Cl₂P(p-OCH₃Ph)₂CH₂OH ( RuMPOH ) and two half-sandwich iridium(III) complexes Ir(η ⁵-Cp*)Cl₂PPh₂CH₂OH ( IrPOH ) and Ir(η ⁵-Cp*)Cl₂P(p-OCH₃Ph)₂CH₂OH ( IrMPOH ) have been studied in terms of potential anticancer activity on previously selected cell line (human lung adenocarcinoma). Based on experimental results obtained in monoculture in vitro model mechanistic considerations on the possible cellular modes of action have been carried out. ICP-MS analysis revealed the higher cellular uptake for less hydrophobic Ir(III) complexes in comparison to the corresponding Ru(II) compounds. Cytometric analysis showed a predominance of apoptosis over the other types of cell death for all complexes. The apoptotic pathway was confirmed by a decrease in mitochondrial membrane potential and the activation of caspases-3/9 for both Ru(II) and Ir(III) complexes. It was concluded that in the case of Ru(II) complexes the intense ROS generation is mainly responsible for the resulting cytotoxicity. The corresponding Ir(III) complexes trigger simultaneously at least three different cytotoxic pathways i. e., depletion of mitochondrial potential, activation of caspases-dependent apoptosis, and ROS-associated oxidation. Thus, it can be assumed that the final accumulation of toxic effects over time via parallel activation of different pathways results in the highest cytotoxicity in vitro exhibited by Ir(III) complexes when compared with Ru(II) complexes.     

Effect of Elicitation with (+)-Usnic Acid on Accumulation of Phenolic Acids and Flavonoids in Agitated Microshoots of Eryngium alpinum L.

The present work was aimed at studying the potential of elicitation on the accumulation of phenolic compounds in in vitro shoot cultures of Eryngium alpinum L., a protected plant from the Apiaceae family. The study examined the influence of (+)-usnic acid on the biomass growth as well as on the biosynthesis of the desired flavonoids and phenolic acids in the cultured microshoots. The phenolic compound content was determined by HPLC-DAD. The flavonoid of the highest concentration was isoquercetin, and the phenolic acids of the highest amount were rosmarinic acid, caffeic acid and 3,4-dihydroxyphenylacetic acid, both in the non-elicited and elicited biomass. Isoquercetin accumulation was efficiently increased by a longer elicitation with a lower concentration of lichenic compound (107.17 ± 4.67 mg/100 g DW) or a shorter elicitation with a higher concentration of acid (127.54 ± 11.34 and 108.37 ± 12.1 mg/100 g DW). Rosmarinic acid production generally remained high in all elicited and non-elicited microshoots. The highest content of this acid was recorded at 24 h of elicitation with 3.125 µM usnic acid (512.69 ± 4.89 mg/100 g DW). The process of elicitation with (+)-usnic acid, a well-known lichenic compound with allelopathic nature, may therefore be an effective technique of enhancing phenolic compound accumulation in alpine eryngo microshoot biomass.     

Oclacitinib,a Janus Kinase Inhibitor,Reduces the Frequency of IL-4- and IL-10-, but Not IFN-γ-, Producing Murine CD4+ and CD8+ T Cells and Counteracts the Induction of Type 1 Regulatory T Cells

The purpose of the present study was to broaden the knowledge and understanding of the effects of oclacitinib (OCL), a Janus kinase inhibitor, on T cells in the context of both the immune mechanisms underlying anti-inflammatory and anti-allergic properties of the drug and its safety. The results indicate that beneficial effects of OCL in the treatment of skin allergic diseases may be partially mediated by the inhibition of IL-4 production in CD4⁺ and CD8⁺ T cells. To a certain extent, the antiproliferative effect of OCL on CD8⁺ T cells may also contribute to its therapeutic effect. The study found that OCL does not affect the proliferation of CD4⁺ T cells or the number of IFN-γ- and IL-17-producing CD4⁺ and CD8⁺ T cells. Moreover, OCL was found to counteract the induction of type 1 regulatory T (Tr1) cells and to act as a strong inhibitor of IL-10 production in both CD4⁺ and CD8⁺ T cells. Thus, these results indicate that beneficial effects of OCL in the treatment of skin allergic diseases are not mediated through: (a) the abolishment of IFN-γ and IL-17-production in CD4⁺ and CD8⁺ T cells; (b) generation of Tr1 cells; (c) inhibition of CD4⁺ T cell proliferation; (d) induction of IL-10 production in CD4⁺ T cells. The results of this study strongly suggest that, with respect to the evaluated parameters, OCL exerts a suppressive effect on Th2- but not Th1-mediated immunity.