Fractionation of Carlina acaulis L. Root Methanolic Extract as a Promising Path towards New Formulations against Bacillus cereus and Methicillin-Resistant Staphylococcus aureus.

The root of Carlina acaulis L. has been widely used in traditional medicine for its antimicrobial properties. In this study, the fractionation of methanol extract from the root was conducted. Four fractions (A, B, C, and D) were obtained and tested against a range of bacteria and fungi. The results showed promising antibacterial activity, especially against Bacillus cereus, where the minimal inhibitory concentration (MIC) was determined to be equal to 0.08 mg/mL and 0.16 mg/mL for heptane (fraction B) and ethyl acetate (fraction C), respectively. In the case of the methicillin-resistant Staphylococcus aureus (MRSA) ATCC 43300 strain, the same fractions yielded higher MIC values (2.5 and 5.0 mg/mL, respectively). This was accompanied by a lack of apparent cytotoxicity to normal human BJ foreskin fibroblasts, enterocytes derived from CaCo2 cells, and zebrafish embryos. Further analyses revealed the presence of bioactive chlorogenic acids in the fractionated extract, especially in the ethyl acetate fraction (C). These findings support the traditional use of the root from C. acaulis and pave the way for the development of new formulations for treating bacterial infections. This was further evaluated in a proof-of-concept experiment where fraction C was used in the ointment formulation, which maintained high antimicrobial activity against MRSA and displayed low toxicity towards cultured fibroblasts.

New cyclic arylguanidine scaffolds as a platform for development of antimicrobial and antiviral agents.

According to WHO, infectious diseases are still a significant threat to public health. The combine effects of antibiotic resistance, immunopressure, and mutations within the bacterial and viral genomes necessitates the search for new molecules exhibiting antimicrobial and antiviral activities. Such molecules often contain cyclic guanidine moiety. As part of this work, we investigated the selected antimicrobial and antiviral activity of compounds from the cyclic arylguanidine group. Molecules were designed using molecular modeling and obtained using microwave radiation (MW) and sonochemical ()))) methods, in accordance with the previously developed pathways. The obtained compounds were screened for the ability to inhibit the growth of Escherichia coli, Acinetobacter baumannii, Pseudomonas aeruginosa, Staphylococcus aureus, Candida albicans, and Cryptococcus neoformans. The capacity to block the entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into the host cell was probed using a bioluminescence immunoassay. The cytotoxicity and hemolytic properties of the most active molecules were also evaluated. The N-[2-(naphthalen-1-yl)ethyl]-5-phenyl-1,4,5,6-tetrahydro-1,3,5-triazin-2-amine 12j showed a high inhibition of Staphylococcus aureus and Cryptococcus neoformans (MIC ≤ 0.25 µg/mL), with no cytotoxic nor hemolytic effect (CC50, HC10 > 32 µm/mL). The CO-ADD platform identified many potentially useful molecules. A particularly rich population was examined in the database of the N.D. Zelinsky Institute of Organic Chemistry, in which 2517 active molecules (MIC ≤ 32 mg/mL) were found, of which about 10% are active at very low concentrations (MIC ≤ 1 mg/mL).

Carlina oxide inhibits the interaction of SARS-CoV-2 S glycoprotein with angiotensin-converting enzyme 2.

Carlina acaulis plant is a potential target for the industrial production of phytochemicals that display applicability in pharmacy and medicine. The dry roots of C. acaulis contain up to 2 % of essential oil, the main component (up to 99 %) of which is carlina oxide [2-(3-phenylprop-1-ynyl)furan]. This compound shows multidirectional biological activity, including antibacterial and antifungal properties. Here, we evaluated the capacity of carlina oxide to inhibit the interaction between SARS-CoV-2 and its human receptor in vitro and in silico. A bioluminescent immunoassay was used to study the interaction between the receptor binding domain (RBD) of viral spike protein and the human angiotensin-converting enzyme 2 (ACE2), which serves as a receptor for viral entry. A dose-effect relationship was demonstrated, and a concentration of carlina oxide causing half-maximal inhibition (IC50) of the RBD:ACE2 interaction was determined to be equal to 234.2 µg/mL. Molecular docking suggested the presence of carlina oxide binding sites within the RBD and at the interface between RBD and ACE2. Finally, this study expands the list of potential applications of C. acaulis as a crop species.

Zirconium-Based Metal-Organic Frameworks as Acriflavine Cargos in the Battle against Coronaviruses─A Theoretical and Experimental Approach.

In this study, we present a complementary approach for obtaining an effective drug, based on acriflavine (ACF) and zirconium-based metal-organic frameworks (MOFs), against SARS-CoV-2. The experimental results showed that acriflavine inhibits the interaction between viral receptor-binding domain (RBD) of spike protein and angiotensin converting enzyme-2 (ACE2) host receptor driving viral cell entry. The prepared ACF@MOF composites exhibited low (MOF-808 and UiO-66) and high (UiO-67 and NU-1000) ACF loadings. The drug release profiles from prepared composites showed different release kinetics depending on the local pore environment. The long-term ACF release with the effective antiviral ACF concentration was observed for all studied ACF@MOF composites. The density functional theory (DFT) calculations allowed us to determine that π-π stacking together with electrostatic interaction plays an important role in acriflavine adsorption and release from ACF@MOF composites. The molecular docking results have shown that acriflavine interacts with several possible binding sites within the RBD and binding site at the RBD/ACE2 interface. The cytotoxicity and ecotoxicity results have confirmed that the prepared ACF@MOF composites may be considered potentially safe for living organisms. The complementary experimental and theoretical results presented in this study have confirmed that the ACF@MOF composites may be considered a potential candidate for the COVID-19 treatment, which makes them good candidates for clinical trials.

Alterations in Kynurenine and NAD+ Salvage Pathways during the Successful Treatment of Inflammatory Bowel Disease Suggest HCAR3 and NNMT as Potential Drug Targets.

A meta-analysis of publicly available transcriptomic datasets was performed to identify metabolic pathways profoundly implicated in the progression and treatment of inflammatory bowel disease (IBD). The analysis revealed that genes involved in tryptophan (Trp) metabolism are upregulated in Crohn's disease (CD) and ulcerative colitis (UC) and return to baseline after successful treatment with infliximab. Microarray and mRNAseq profiles from multiple experiments confirmed that enzymes responsible for Trp degradation via the kynurenine pathway (IDO1, KYNU, IL4I1, KMO, and TDO2), receptor of Trp metabolites (HCAR3), and enzymes catalyzing NAD+ turnover (NAMPT, NNMT, PARP9, CD38) were synchronously coregulated in IBD, but not in intestinal malignancies. The modeling of Trp metabolite fluxes in IBD indicated that changes in gene expression shifted intestinal Trp metabolism from the synthesis of 5-hydroxytryptamine (5HT, serotonin) towards the kynurenine pathway. Based on pathway modeling, this manifested in a decline in mucosal Trp and elevated kynurenine (Kyn) levels, and fueled the production of downstream metabolites, including quinolinate, a substrate for de novo NAD+ synthesis. Interestingly, IBD-dependent alterations in Trp metabolites were normalized in infliximab responders, but not in non-responders. Transcriptomic reconstruction of the NAD+ pathway revealed an increased salvage biosynthesis and utilization of NAD+ in IBD, which normalized in patients successfully treated with infliximab. Treatment-related changes in NAD+ levels correlated with shifts in nicotinamide N-methyltransferase (NNMT) expression. This enzyme helps to maintain a high level of NAD+-dependent proinflammatory signaling by removing excess inhibitory nicotinamide (Nam) from the system. Our analysis highlights the prevalent deregulation of kynurenine and NAD+ biosynthetic pathways in IBD and gives new impetus for conducting an in-depth examination of uncovered phenomena in clinical studies.

Kynurenine emerges from the shadows - Current knowledge on its fate and function.

Kynurenine (KYN), a main metabolite of tryptophan in mammals, is a direct precursor of kynurenic acid, anthranilic acid and 3-hydroxykynurenine (3-HK). Under physiological conditions, KYN is produced endogenously mainly in the liver by tryptophan 2,3-dioxygenase (TDO). Tumorigenesis and inflammatory conditions increase the activity of another KYN synthetizing enzyme, indoleamine 2,3-dioxygenase (IDO). However, knowledge about the exogenous sources and the fate of KYN in mammals is still limited. While most papers deal with the contribution of KYN to pathologies of the central nervous system, its role in the periphery has almost been ignored. KYN is a ligand for the aryl hydrocarbon receptor (AhR). As a receptor for KYN and its downstream metabolites, AhR is involved in several physiological and pathological conditions, including inflammation and carcinogenesis. Recent studies have shown that KYN suppresses immune response and is strongly involved in the process of carcinogenesis and tumour metastasis. Thus, inhibition of activity of the enzymes responsible for KYN synthesis, TDO, IDO or genetic manipulation leading to reduction of KYN synthesis, could be considered as innovative strategies for improving the efficacy of immunotherapy. Surprisingly, however, genetic or pharmacological approaches for reducing tryptophan catabolism to KYN do not necessarily result in decrease of KYN level in the main circulation. This review aims to summarize the current knowledge of KYN fate and function and to emphasize its importance for vital physiological and pathological processes.

Toxicity of Carlina Oxide-A Natural Polyacetylene from the Carlina acaulis Roots-In Vitro and In Vivo Study.

There are several reports indicating that the roots of the Carlina acaulis L. used to be commonly applied as a treatment measure in skin diseases and as an antiparasitic agent, starting from antiquity to the 19th century; however, nowadays, it has lost its importance. Currently, numerous studies are being conducted assessing the possibility of reintroducing C. acaulis-derived extracts to phytotherapy. Determining the safety profile of the main constituents of the plant material is crucial for achieving this goal. Here, we aimed to determine the toxicity profile of carlina oxide, one of the most abundant components of the C. acaulis root extract. We obtained the carlina oxide by distillation of C. acaulis roots in the Deryng apparatus. The purity of the standard was evaluated using GC-MS, and the identity was confirmed by IR, Raman, and NMR spectroscopy. In vitro cytotoxicity was assessed using a panel of human cell lines of skin origin, including BJ normal fibroblasts and UACC-903, UACC-647, and C32 melanoma cells. This was accompanied by an in vivo zebrafish acute toxicity test (ZFET). In vitro studies showed a toxic effect of carlina oxide, as demonstrated by an induction of apoptosis and necrosis in both normal and melanoma cells. Decreased expression of AKT kinase and extracellular signal-regulated kinase 1/2 (ERK1/2) was noted in the UACC-647 melanoma cell line. It was also observed that carlina oxide modified the expression of programmed cell death-ligand 1 (PD-L1) in tested cell lines. Carlina oxide exhibited high in vivo toxicity, with LC50 = 10.13 µg/mL upon the 96 h of exposure in the ZFET test. Here, we demonstrate that carlina oxide displays toxic effects to cells in culture and to living organisms. The data indicate that C. acaulis-based extracts considered for therapeutic use should be completely deprived of carlina oxide.