Screening of Plants for Antibacterial Properties: Growth Inhibition of Staphylococcus aureus by Artemisia Tridentata

Drug-resistant pathogenic and opportunistic bacteria are increasing in occurrence and prevalence, and pose a dangerous threat to human health. In the search for novel antibiotics with which to combat this threat, plants, specifically those used in traditional medicine with ascribed antibacterial properties, offer a promising and potentially vast source of such therapeutic compounds. The purpose of this study was therefore to screen chemical extracts created from various plant species for antibacterial properties versus pathogenic bacterial species. In the course of these antibacterial assays, we successfully identified a methanol extract derived from Artemisia tridentata tridentata plant material as capable of inhibiting the growth of the opportunistic pathogen Staphylococcus aureus. Three sub-fractions were created using hexane, ethyl acetate and water solvents. Each of these extracts displayed significant antibacterial activity versus a wild-type strain over a period of six hours, at concentrations as low as 62.5 µg/ml. The extracts also demonstrated an enhancement of antibiotic effects when combined with ampicillin, G418 sulfate or amikacin, for a period of up to twelve hours. Though the efficacy of the extracts was lessened when tested against an ampicillin-resistant strain, significant enhancement of the efficacy of this antibiotic was still observed. Gas chromatography-mass spectrometry analysis of these three extracts revealed the sesquiterpene lactone achillin as present in each. Column chromatography of the hexane extract resulted in a fraction retaining its antibacterial activity, and still containing this compound, further implicating it as responsible for the antibacterial activity of this plant. The results of serial dilution and plating of extract-treated samples, along with those of ethidium bromide assays and transmission electron microscopy analysis, indicated a bacteriostatic mechanism of action involving disruption of the bacterial membrane, which is in agreement with the literature on the antibacterial properties of this plant, and those of sesquiterpene lactones, respectively. We therefore conclude that achillin, likely produced as a secondary metabolite by Artemisia tridentata tridentata, possesses growth inhibitory properties versus Staphylococcus aureus, and should be isolated and studied further for the purposes of evaluating its potential use, either as a stand-alone antibiotic, or as an adjunctive therapeutic, in the treatment of drug-resistant bacterial pathogens

Characterisation of a functional rat hepatocyte spheroid model.

Many in vitro liver cell models, such as 2D systems, that are used to assess the hepatotoxic potential of xenobiotics suffer major limitations arising from a lack of preservation of physiological phenotype and metabolic competence. To circumvent some of these limitations there has been increased focus on producing more representative 3D models. Here we have used a novel approach to construct a size-controllable 3D hepatic spheroid model using freshly isolated primary rat hepatocytes (PRH) utilising the liquid-overlay technique whereby PRH spontaneously self-assemble in to 3D microtissues. This system produces viable spheroids with a compact in vivo-like structure for up to 21 days with sustained albumin production for the duration of the culture period. F-actin was seen throughout the spheroid body and P-glycoprotein (P-gp) and multidrug resistance-associated protein 2 (MRP2) transporters had polarised expression on the canalicular membrane of hepatocytes within the spheroids upon formation (day 3). The MRP2 transporter was able to functionally transport 5 μM 5-chloromethylfluorescein diacetate (CMFDA) substrates into these canalicular structures. These PRH spheroids display in vivo characteristics including direct cell-cell contacts, cellular polarisation, 3D cellular morphology, and formation of functional secondary structures throughout the spheroid. Such a well-characterised system could be readily exploited for pre-clinical and non-clinical repeat-dose investigations and could make a significant contribution to replace, reduce and refine the use of animals for applied research

Screening of Plants for Antibacterial Properties: Growth Inhibition of Staphylococcus aureus by Artemisia tridentata

Drug-resistant pathogenic and opportunistic bacteria are increasing in occurrence and prevalence, and pose a dangerous threat to human health. In the search for novel antibiotics with which to combat this threat, plants, specifically those used in traditional medicine with ascribed antibacterial properties, offer a promising and potentially vast source of such therapeutic compounds. The purpose of this study was therefore to screen chemical extracts created from various plant species for antibacterial properties versus pathogenic bacterial species. In the course of these antibacterial assays, we successfully identified a methanol extract derived from Artemisia tridentata tridentata plant material as capable of inhibiting the growth of the opportunistic pathogen Staphylococcus aureus. Three sub-fractions were created using hexane, ethyl acetate and water solvents. Each of these extracts displayed significant antibacterial activity versus a wild-type strain over a period of six hours, at concentrations as low as 62.5 µg/ml. The extracts also demonstrated an enhancement of antibiotic effects when combined with ampicillin, G418 sulfate or amikacin, for a period of up to twelve hours. Though the efficacy of the extracts was lessened when tested against an ampicillin-resistant strain, significant enhancement of the efficacy of this antibiotic was still observed. Gas chromatography-mass spectrometry analysis of these three extracts revealed the sesquiterpene lactone achillin as present in each. Column chromatography of the hexane extract resulted in a fraction retaining its antibacterial activity, and still containing this compound, further implicating it as responsible for the antibacterial activity of this plant. The results of serial dilution and plating of extract-treated samples, along with those of ethidium bromide assays and transmission electron microscopy analysis, indicated a bacteriostatic mechanism of action involving disruption of the bacterial membrane, which is in agreement with the literature on the antibacterial properties of this plant, and those of sesquiterpene lactones, respectively. We therefore conclude that achillin, likely produced as a secondary metabolite by Artemisia tridentata tridentata, possesses growth inhibitory properties versus Staphylococcus aureus, and should be isolated and studied further for the purposes of evaluating its potential use, either as a stand-alone antibiotic, or as an adjunctive therapeutic, in the treatment of drug-resistant bacterial pathogens

Biomedical properties of sesquiterpene lactones, with a focus on dehydroleucodine

Dehydroleucodine, a sesquiterpene lactone, belongs to the terpenoid class of secondary metabolites. Dehydroleucodine and other Artemisia-derivedphytochemicals evolved numerous biodefenses that were first co-opted for human pharmacological use by traditional cultures in the Middle East, Asia, Europeand the Americas. Later, these phytochemicals were modified through the use of medicinal chemical techniques to increase their potency. All sesquiterpenelactones contain an α-methylene-γ-lactone group, which confers thiol reactivity, which is responsible, in part, for their therapeutic effects. A wide range oftherapeutic uses of sequiterpene lactones has been found, including anti-adipogenic, cytoprotective, anti-microbial, anti-viral, anti-fungal, anti-malarial and,anti-migraine effects. Dehydroleucodine significantly inhibits differentiation of murine preadipocytes and also significantly decreases the accumulation oflipid content by a dramatic down regulation of adipogenic-specific transcriptional factors PPARγ and C-EBPα. Dehydroleucodine also inhibits secretion ofmatrix metalloprotease-2 (MMP-2), which is a known protease involved in migration and invasion of B16 cells. In addition to these anti-adipogenic and anticancereffects, dehydroleucodine effectively neutralizes several bacterial species, including Bacillus cereus, Staphylococcus aureus, Escherichia coli,Klebsiella pneumoniae, Helicobacter pylori, methicillin resistant Staphylococcus aueus (MRSA) and S. epidermis (MRSE). The compound also inhibits thegrowth and secretion of several toxins of Pseudomonas aeruginosa, possesses gastro-protective qualities and possesses anti-parasitic properties againstTrypanosoma cruzi, responsible for Chagas disease. Other sesquiterpene lactones, such as parthenolide, costunolide, and helanin, also possess significanttherapeutic utility.Fil: Abood, Steven. Florida International University; Estados UnidosFil: Mustafi, Sushmita. Florida International University; Estados UnidosFil: Veisaga, Maria-Luisa. Florida International University; Estados UnidosFil: Lopez, Luis Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos. Universidad Nacional de Cuyo. Facultad de Cienicas Médicas. Instituto de Histología y Embriología de Mendoza Dr. Mario H. Burgos; ArgentinaFil: Barbieri, Manuel Alejandro. Fairchild Tropical Botanic Garden; Estados Unido

Genetic determinants of variable metabolism have little impact on the clinical use of leading antipsychotics in the CATIE study

PURPOSE: To evaluate systematically in real clinical settings whether functional genetic variations in drug metabolizing enzymes influence optimized doses, efficacy, and safety of antipsychotic medications. METHODS: DNA was collected from 750 patients with chronic schizophrenia treated with five antipsychotic drugs (olanzapine, quetiapine, risperidone, ziprasidone and perphenazine) as part of the Clinical Antipsychotic Trials of Intervention Effectiveness (CATIE) study. Doses for each of the medicines were optimized to 1, 2, 3, or 4x units in identically-appearing capsules in a double blind design. We analyzed 25 known functional genetic variants in the major and minor metabolizing enzymes for each medication. These variants were tested for association with optimized dose and other relevant clinical outcomes. RESULTS: None of the tested variants showed a nominally significant main effect in association with any of the tested phenotypes in European-Americans, African-Americans or all patients. Even after accounting for potential covariates no genetic variant was found to be associated with dosing, efficacy, overall tolerability, or tardive dyskinesia. CONCLUSION: There are no strong associations between common functional genetic variants in drug metabolizing enzymes and dosing, safety or efficacy of leading antipsychotics, strongly suggesting merely modest effects on the use of these medicines in most patients in typical clinical settings