Pyrimidine Schiff Bases: Synthesis, Structural Characterization and Recent Studies on Biological Activities.

Recently, 5-[(4-ethoxyphenyl)imino]methyl-N-(4-fluorophenyl)-6-methyl-2-phenylpyrimidin-4-amine has been synthesized, characterized, and evaluated for its antibacterial activity against Enterococcus faecalis in combination with antineoplastic activity against gastric adenocarcinoma. In this study, new 5-iminomethylpyrimidine compounds were synthesized which differ in the substituent(s) of the aromatic ring attached to the imine group. The structures of newly obtained pyrimidine Schiff bases were established by spectroscopy techniques (ESI-MS, FTIR and 1H NMR). To extend the current knowledge about the features responsible for the biological activity of the new 5-iminomethylpyrimidine derivatives, low-temperature single-crystal X-ray analyses were carried out. For all studied crystals, intramolecular N-H∙∙∙N hydrogen bonds and intermolecular C-H∙∙∙F interactions were observed and seemed to play an essential role in the formation of the structures. Simultaneously, their biological properties based on their cytotoxic features were compared with the activities of the Schiff base (III) published previously. Moreover, computational investigations, such as ADME prediction analysis and molecular docking, were also performed on the most active new Schiff base (compound 4b). These results were compared with the highest active compound III.

In silico and in vitro screening for carcinogenic potential of angiotensin-converting enzyme inhibitors and their degradation impurities.

According to some clinical observations, the use of angiotensin-converting enzyme inhibitors (ACEI) may be associated with an increased risk of cancer. The aim of the present study was to screen for the potential carcinogenicity, mutagenicity and genotoxicity of these drugs using in silico methodology. Delapril, enalapril, imidapril, lisinopril, moexipril, perindopril, ramipril, trandolapril, spirapril were thereby analyzed. In parallel, the corresponding degradation impurities, the diketopiperazine (DKP) derivatives, were also investigated. (Q)SAR computer software (VEGA-GUI and Lazar), available in the public domain, was employed. The obtained predictions suggested that none of the compounds tested (from the group of ACE-Is and DKPs) was mutagenic. Moreover, none of the ACE-Is was carcinogenic. The reliability of these predictions was high to moderate. In contrast, in the DKP group, ramipril-DKP and trandolapril-DKP were found to be potentially carcinogenic, but the reliability of this prediction was low. As for the genotoxicity screening, all compounds tested (ACE-I and DKP) were predicted to be active and genotoxic, with moexipril, ramipril, spirapril, and all DKP derivatives within the highest risk group. They were prioritized for experimental verification studies to confirm or exclude their toxic activity. On the other hand, the lowest risk of carcinogenicity was assigned to imidapril and its DKP. Then, a follow-up in vitro micronucleus assay for ramipril was performed. It showed that this drug was genotoxic via aneugenic activity, but only at concentrations exceeding real-life levels. At concentrations found in human blood after standard dose, ramipril was not genotoxic in vitro. Therefore, ramipril was considered safe for human use with a standard dosing regimen. The other compounds of concern (spirapril, moexipril and all DKP derivatives) should be subjected to analogous in vitro studies. We also concluded that the adopted in silico software was applicable for ACE-I toxicity prediction.

Zn(II) Induces Fibril Formation and Antifungal Activity in Shepherin I, An Antimicrobial Peptide from Capsella bursa-pastoris.

Shepherin I is a glycine- and histidine-rich antimicrobial peptide from the root of a shepherd's purse, whose antimicrobial activity was suggested to be enhanced by the presence of Zn(II) ions. We describe Zn(II) and Cu(II) complexes of this peptide, aiming to understand the correlation between their metal binding mode, structure, morphology, and biological activity. We observe a logical sequence of phenomena, each of which is the result of the previous one: (i) Zn(II) coordinates to shepherin I, (ii) causes a structural change, which, in turn, (iii) results in fibril formation. Eventually, this chain of structural changes has a (iv) biological consequence: The shepherin I-Zn(II) fibrils are highly antifungal. What is of particular interest, both fibril formation and strong anticandidal activity are only observed for the shepherin I-Zn(II) complex, linking its structural rearrangement that occurs after metal binding with its morphology and biological activity.

Semenogelins Armed in Zn(II) and Cu(II): May Bioinorganic Chemistry Help Nature to Cope with Enterococcus faecalis?

Proteolytic degradation of semenogelins, the most abundant proteins from human semen, results in the formation of 26- and 29-amino acid peptides (SgIIA and SgI-29, respectively), which share a common 15 amino acid fragment (Sg-15). All three ligands are effective Zn(II) and Cu(II) binders; in solution, a variety of differently metalated species exist in equilibrium, with the [NH2, 3Nim] donor set prevailing at physiological pH in the case of both metals. For the first time, the Cu(II)-induced antimicrobial activity of Sg-15 against Enterococcus faecalis is shown. In the case of the two native semenogelin fragment metal complexes, the strong local positive charge in the metal-bound HH motif correlates well with their antimicrobial activity. A careful analysis of semenogelins' metal coordination behavior reveals two facts: (i) The histamine-like Cu(II) binding mode of SgI-29 strongly increases the stability of such a complex below pH 6 (with respect to the non-histamine-like binding of SgIIA), while in the case of the SgI-29 Zn(II)-histamine-like species, the stability enhancement is less pronounced. (ii) The HH sequence is a more tempting site for Cu(II) ions than the HXH one.

Novel Antimicrobial Peptides from Saline Environments Active against E. faecalis and S. aureus: Identification, Characterisation and Potential Usage.

Microorganisms inhabiting saline environments have been known for decades as producers of many valuable bioproducts. These substances include antimicrobial peptides (AMPs), the most recognizable of which are halocins produced by halophilic Archaea. As agents with a different modes of action from that of most conventionally used antibiotics, usually associated with an increase in the permeability of the cell membrane as a result of a formation of channels and pores, AMPs are a currently promising object of research focused on the investigation of antibiotics with non-standard modes of action. The aim of this study was to investigate antimicrobial activity against multidrug-resistant human pathogens of three peptides, which were synthetised based on sequences identified in metagenomes from saline environments. The investigations were performed against Enterococcus faecalis, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Escherichia coli and Candida albicans. Subsequently, the cytotoxicity and haemolytic properties of the tested peptides were verified. An in silico analysis of the interaction of the tested peptides with molecular targets for reference antibiotics was also carried out in order to verify whether or not they can act in a similar way. The P1 peptide manifested the growth inhibition of E. faecalis at a MIC50 of 32 µg/mL and the P3 peptide at a MIC50 of 32 µg/mL was shown to inhibit the growth of both E. faecalis and S. aureus. Furthermore, the P1 and P3 peptides were shown to have no cytotoxic or haemolytic activity against human cells.

Chemical "Butterfly Effect" Explaining the Coordination Chemistry and Antimicrobial Properties of Clavanin Complexes.

Can a minor difference in the nonmetal binding sequence of antimicrobial clavanins explain the drastic change in the coordination environment and antimicrobial efficiency? This study answers the question with a definite "yes", showing the details of the bioinorganic chemistry of Zn(II) and Cu(II) complexes with clavanins, histidine-rich, antimicrobial peptides from hemocytes of the tunicate Styela clava.

Zn-Enhanced Asp-Rich Antimicrobial Peptides: N-Terminal Coordination by Zn(II) and Cu(II), Which Distinguishes Cu(II) Binding to Different Peptides.

The antimicrobial activity of surfactant-associated anionic peptides (SAAPs), which are isolated from the ovine pulmonary surfactant and are selective against the ovine pathogen Mannheimia haemolytica, is strongly enhanced in the presence of Zn(II) ions. Both calorimetry and ITC measurements show that the unique Asp-only peptide SAAP3 (DDDDDDD) and its analogs SAAP2 (GDDDDDD) and SAAP6 (GADDDDD) have a similar micromolar affinity for Zn(II), which binds to the N-terminal amine and Asp carboxylates in a net entropically-driven process. All three peptides also bind Cu(II) with a net entropically-driven process but with higher affinity than they bind Zn(II) and coordination that involves the N-terminal amine and deprotonated amides as the pH increases. The parent SAAP3 binds Cu(II) with the highest affinity; however, as shown with potentiometry and absorption, CD and EPR spectroscopy, Asp residues in the first and/or second positions distinguish Cu(II) binding to SAAP3 and SAAP2 from their binding to SAAP6, decreasing the Cu(II) Lewis acidity and suppressing its square planar amide coordination by two pH units. We also show that these metal ions do not stabilize a membrane disrupting ability nor do they induce the antimicrobial activity of these peptides against a panel of human pathogens.

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.

Impact of metal coordination and pH on the antimicrobial activity of histatin 5 and the products of its hydrolysis.

This work focuses on the relationship between the coordination chemistry and antimicrobial activity of Zn(II) and Cu(II) complexes of histatin 5 and the products of its hydrolysis: its N-terminal fragment (histatin 5-8) and C-terminal fragment (histatin 8). Cu(II) coordinates in an albumin-like binding mode and Zn(II) binds to up to 3 His imidazoles. The antimicrobial activity of histatins and their metal complexes (i) strongly depends on pH - they are more active at pH 5.4 than at 7.4; (ii) the complexes and ligands alone are more effective in eradicating Gram-positive bacteria than the Gram-negative ones, and (iii) Zn(II) coordination is able to change the structure of the N-terminal region of histatin 5 (histatin 5-8) and moderately increase all of the studied histatins' antimicrobial potency.

Pluronic F-127 Enhances the Antifungal Activity of Fluconazole against Resistant Candida Strains.

Candida strains as the most frequent causes of infections, along with their increased drug resistance, pose significant clinical and financial challenges to the healthcare system. Some polymeric excipients were reported to interfere with the multidrug resistance mechanism. Bearing in mind that there are a limited number of marketed products with fluconazole (FLU) for the topical route of administration, Pluronic F-127 (PLX)/FLU formulations were investigated in this work. The aims of this study were to investigate (i) whether PLX-based formulations can increase the susceptibility of resistant Candida strains to FLU, (ii) whether there is a correlation between block polymer concentration and the antifungal efficacy of the FLU-loaded PLX formulations, and (iii) what the potential mode of action of PLX assisting FLU is. The yeast growth inhibition upon incubation with PLX formulations loaded with FLU was statistically significant. The highest efficacy of the azole agent was observed in the presence of 5.0 and 10.0% w/v of PLX. The upregulation of the CDR1/CDR2 genes was detected in the investigated Candida strains, indicating that the efflux of the drug from the fungal cell was the main mechanism of the resistance.

Bioinorganic chemistry of shepherin II complexes helps to fight Candida albicans?

The fungal cell wall and cell membrane are an important target for antifungal therapies, and a needle-like cell wall or membrane disruption may be an entirely novel antifungal mode of action. In this work, we show how the coordination of Zn(II) triggers the antifungal properties of shepherin II, a glycine- and histidine-rich antimicrobial peptide from the root of Capsella bursa-pastoris. We analyze Cu(II) and Zn(II) complexes of this peptide using experimental and theoretical methods, such as: mass spectrometry, potentiometry, UV-Vis and CD spectroscopies, AFM imaging, biological activity tests and DFT calculations in order to understand the correlation between their metal binding mode, structure, morphology and biological activity. We observe that Zn(II) coordinates to Shep II and causes a structural change, resulting in fibril formation, what has a pronounced biological consequence - a strong anticandidal activity. This phenomenon was observed neither for the peptide itself, nor for its copper(II) complex. The Zn(II) - shepherin II complex can be considered as a starting point for further anticandidal drug discovery, which is extremely important in the era of increasing antifungal drug resistance.

Zn(II) binding to pramlintide results in a structural kink, fibril formation and antifungal activity.

The antimicrobial properties of amylin, a 37-amino acid peptide hormone, co-secreted with insulin from the pancreas, are far less known than its antidiabetic function. We provide insight into the bioinorganic chemistry of amylin analogues, showing that the coordination of zinc(II) enhances the antifungal properties of pramlintide, a non-fibrillating therapeutic analogue of amylin. Zinc binds to the N-terminal amino group and His18 imidazole, inducing a kink in the peptide structure, which, in turn, triggers a fibrillization process of the complex, resulting in an amyloid structure most likely responsible for the disruption of the fungal cell.

Activity of erythrocyte antioxidant enzymes in healthy women depends on age, BMI, physical activity, and diet.

INTRODUCTION: Antioxidant enzymes protect the human body against the harmful effects of oxidative stress. The activity of antioxidant enzymes changes with age and depends on dietary nutrients such as fats and vitamins, which can have a significant impact on minimizing or exacerbating oxidative stress. AIM: To examine the effect of age, BMI, diet, physical activity, and smoking status on the activity of erythrocyte antioxidant enzymes catalase, glutathione reductase, glutathione peroxidase glutathione S-transferase, superoxide dismutase, and glutathione concentrations in healthy women. MATERIAL AND METHODS: This study included 98 healthy women aged between 20 and 65 years. All women underwent anthropometric tests: body weight, height, hip, and waist circumference. Antioxidant activity in erythrocytes was measured by spectrophotometric methods. RESULTS: Catalase activity increased significantly with age (p < 0.001), while superoxide dismutase activities and glutathione decreased with age (p = 0.008, p = 0.023, respectively). Women with a lower BMI (emaciation) had higher superoxide dismutase activity than those in the first degree of obesity (p = 0.009). CONCLUSIONS: (1) Increased catalase activity with age may signify a large amount of hydrogen peroxide resulting from malfunctioning antioxidant systems in old age. (2) A decline in superoxide dismutase activity with age may indicate inactivation of this enzyme, inappropriate SOD function in the presence of excessive amounts of hydrogen peroxide, and glycation of superoxide dismutase molecules. (3) A negative correlation between superoxide dismutase activity and the BMI index may indicate a decreased enzymatic activity in obese people.

Synthesis, Crystal Structure, and Biological Evaluation of Novel 5-Hydroxymethylpyrimidines.

Pyrimidine displays a wide array of bioactivities, and thence, it is still considered a potent unit of new drug research. Its derivative, 5-hydroxymethylpyrimidine, can be found as a scaffold of nontypical nitrogen bases in DNA and as a core of some natural bioactive compounds. In this study, we obtained a series of 5-hydroxymethylpyrimidines that vary in the 4-position by the reduction of proper esters. All compounds were characterized by spectroscopic analysis, and single-crystal X-ray diffraction was performed for some of them. Biological investigations estimated cytotoxic properties against normal (RPTEC) and cancer (HeLa, HepaRG, Caco-2, AGS, A172) cell lines. It was found that the derivatives with an aliphatic amino group at the 4-position are generally less toxic to normal cells than those with a benzylsulfanyl group. Moreover, compounds with bulky constituents exhibit better anticancer properties, though at a moderate level. The specific compounds were chosen due to their most promising IC50 concentration for in silico study. Furthermore, antimicrobial activity tests were performed against six strains of bacteria and one fungus. They demonstrated that only derivatives with at least three carbon chain amino groups at the 4-position have weak antibacterial properties, and only the derivative with 4-benzylsulfanyl constituent exhibits any antifungal action.

Zn(II)-alloferon complexes - Similar sequence, different coordination modes, no antibacterial activity.

Often, in the search for a highly defined scientific phenomenon, a different one becomes apparent. This was also the case of this work, in the scope of which we planned to search for metal-enhanced, novel antibacterial/antifungal compounds. Instead, we denied the existence of such and revealed the details of the bioinorganic chemistry of Zn(II)-alloferon complexes. Zinc(II) complexes of alloferon 1 and 2, ligands with a sequential difference of one amino acid only, show a substantially different coordination pattern at physiological pH. In the case of Zn(II)-alloferon 1 species, a histamine-like binding mode is observed (N-terminal amine and imidazole of His-1) and the coordination sphere is completed with the imidazole nitrogens of His-6 and His-9; His-12 is not involved in binding. In the case of Zn(II)-alloferon 2, the N-terminal amine and all the three imidazoles present in the sequence participate in the coordination, however, with the chemical shift of His-5 being less affected than those of other imidazoles. The histamine-like binding in Zn(II)-alloferon 1 complex strongly enhances its thermodynamic stability in comparison to the His-1 lacking alloferon 2 analogue. Despite previous reports on the antibacterial and antifungal activity of alloferon 1, no such activity was detected, neither for alloferon 1 and 2 nor for their Zn(II) complexes.

Bioinorganic chemistry of calcitermin - the picklock of its antimicrobial activity.

Calcitermin, an antimicrobial peptide from the fluid of the human airways, is a well-conserved, 15 amino acid C-terminal cleavage fragment of calgranulin C (VAIALKAAHYHTHKE), which is active under acidic pH conditions (pH 5.4). In an attempt to understand the impact of the coordination of Zn(ii) and Cu(ii) on the biological activity of calcitermin, we mutated each of the histidines with an alanine and studied the thermodynamics, binding mode and antimicrobial activity of wild type calcitermin and its H9A, H11A and H13A mutants and their Zn(ii) and Cu(ii) complexes. Both metals strongly enhance the antimicrobial activity of calcitermin-like peptides, although the link between the minimal inhibitory concentration (MIC) values and the stability, charge or structure of the complexes is not so obvious. As expected, the increase in the number of histidines makes the coordination of both metals more effective. There is no preferred Cu(ii) binding site in calcitermin: the stabilities of the Cu(ii)-H9A and Cu(ii)-H13A complexes are almost identical, while the Cu(ii)-H11A complex (in which two histidines are separated by three amino acids and only one His residue is involved in binding) is less stable. On the other hand, the higher stability of the Zn(ii)-H13A complex with respect to those formed by H9A and H11A suggests a pivotal role of His9 and His11 in Zn(ii) complexation. Impressive MIC breakpoints were obtained, similar and lower than those for commonly used antimicrobial agents that treat Candida albicans (Zn(ii) and Cu(ii) complexes of WT calcitermin and H9A, as well as H9A alone), Enterococcus faecalis (H11A, H13A and their metal complexes) and Staphylococcus aureus (H13A and its complexes).