Tyrosol-gold nanoparticle functionalized acacia gum-PVA nanofibers for mitigation of Candida biofilm.

Increasing incidences of fungal infections and prevailing antifungal resistance in healthcare settings has given rise to an antifungal crisis on a global scale. The members of the genus Candida, owing to their ability to acquire sessile growth, are primarily associated with superficial to invasive fungal infections, including the implant-associated infections. The present study introduces a novel approach to combat the sessile/biofilm growth of Candida by fabricating nanofibers using a nanoencapsulation approach. This technique involves the synthesis of tyrosol (TYS) functionalized chitosan gold nanocomposite, which is then encapsulated into PVA/AG polymeric matrix using electrospinning. The FESEM, FTIR analysis of prepared TYS-AuNP@PVA/AG NF suggested the successful encapsulation of TYS into the nanofibers. Further, the sustained and long-term stability of TYS in the medium was confirmed by drug release and storage stability studies. The prepared nanomats can absorb the fluid, as evidenced by the swelling index of the nanofibers. The growth and biofilm inhibition, as well as the disintegration studies against Candida, showed 60-70 % biofilm disintegration when 10 mg of TYS-AuNP@PVA/AG NF was used, hence confirming its biological effectiveness. Subsequently, the nanofibers considerably reduced the hydrophobicity index and ergosterol content of the treated cells. Considering the challenges associated with the inhibition/disruption of fungal biofilm, the fabricated nanofibers prove their effectiveness against Candida biofilm. Therefore, nanocomposite-loaded nanofibers have emerged as potential materials that can control fungal colonization and could also promote healing.

Cholesterol vs Ergosterol: Influence on the Dynamic and Structural Properties of the Cobalt-Based Metallosomal Bilayer Membrane.

Sterol derivatives are a crucial part of liposomes, as their concentration and nature can induce significant alternations in their characteristic features. For natural liposomal-based (phospholipid-based) studies, the bulk literature is already present depicting the role of the concentration or nature of different sterol derivatives in modulation of membrane properties. However, the studies aiming at evaluating the effect of sterol derivatives on synthetic liposomal assemblies are limited to cholesterol (Chl), and a comparative effect with other sterol derivatives, such as ergosterol (Erg), has never been studied. To fill this research gap, through this work, we intend to provide insights into the concentration-dependent effect of two sterol derivatives (Chl and Erg) on a synthetic liposomal assembly (i.e., metallosomes) prepared via thin film hydration route using a double-tailed metallosurfactant fabricated by modifying cetylpyridinium chloride with cobalt (Co) (i.e., Co:CPC II). The morphological evaluations with cryogenic-transmission electron microscopy (cryo-TEM), atomic force microscopy (AFM), and field emission-scanning electron microscopy (FE-SEM) indicated that metallosomes retained their spherical morphology irrespective of the nature and concentration of sterol derivatives. However, the size, ζ-potential, and lamellar width values were significantly modified with the incorporation of sterol derivatives in a concentration-dependent manner. In-depth studies affirmed that the extent of modulation of the bilayer in terms of hydrophobicity, fluidity, and rigidity was more severe with Chl than Erg. Such differences in the membrane properties lead to their contrasting behavior in the delivery of the broad-spectrum active compound "curcumin". From entrapment to in vitro behavior, the metallosomes demonstrated dissimilar behavior as even though Erg-modified metallosomes (at higher concentrations of Erg) exhibited low entrapment efficiency, they still could easily release >80% of the entrapped drug. In vitro studies conducted with Staphylococcus aureus bacterial cultures further revealed an interesting pattern of activity as the incorporation of Chl reduced the toxicity of the self-assembly, whereas their Erg-modified counterparts yielded slightly augmented toxicity toward these bacterial cells. Furthermore, Chl- and Erg-modified assemblies also exhibited contrasting behavior in their interaction studies with bacterial DNA.

Druggable Sterol Metabolizing Enzymes in Infectious Diseases: Cell Targets to Therapeutic Leads.

Sterol biosynthesis via the mevalonate-isoprenoid pathway produces ergosterol (24ß-methyl cholesta-5,7-dienol) necessary for growth in a wide-range of eukaryotic pathogenic organisms in eukaryotes, including the fungi, trypanosomes and amoebae, while their animal hosts synthesize a structurally less complicated product-cholesterol (cholest-5-enol). Because phyla-specific differences in sterol metabolizing enzyme architecture governs the binding and reaction properties of substrates and inhibitors while the order of sterol metabolizing enzymes involved in steroidogenesis determine the positioning of crucial chokepoint enzymes in the biosynthetic pathway, the selectivity and effectiveness of rationally designed ergosterol biosynthesis inhibitors toward ergosterol-dependent infectious diseases varies greatly. Recent research has revealed an evolving toolbox of mechanistically distinct tight-binding inhibitors against two crucial methylation-demethylation biocatalysts-the C24 sterol methyl transferase (absent from humans) and the C14-sterol demethylase (present generally in humans and their eukaryotic pathogens). Importantly for rational drug design and development, the activities of these enzymes can be selectively blocked in ergosterol biosynthesis causing loss of ergosterol and cell killing without harm to the host organism. Here, we examine recent advances in our understanding of sterol biosynthesis and the reaction differences in catalysis for sterol methylation-demethylation enzymes across kingdoms. In addition, the novelties and nuances of structure-guided or mechanism-based approaches based on crystallographic mappings and substrate specificities of the relevant enzyme are contrasted to conventional phenotypic screening of small molecules as an approach to develop new and more effective pharmacological leads.

Nonstereotypical Distribution and Effect of Ergosterol in Lipid Membranes.

Ergosterol, found in fungi and some protist membranes, is understudied compared with cholesterol from animal membranes. Generally, ergosterol is assumed to modulate membranes in the same manner as cholesterol, based on their similar chemical structures. Here we reveal some fundamental structural and dynamical differences between them. Neutron diffraction shows that ergosterol is embedded in the lipid bilayer much shallower than cholesterol. Ergosterol does not change the membrane thickness as much as cholesterol does, indicating little condensation effect. Neutron spin echo shows that ergosterol can rigidify and soften membranes at different concentrations. The lateral lipid diffusion measured by quasielastic neutron scattering indicates that ergosterol promotes a jump diffusion of the lipid, whereas cholesterol keeps the same continuous lateral diffusion as the pure lipid membrane. Our results point to quite distinct interactions of ergosterol with membranes compared with cholesterol. These insights provide a basic understanding of membranes containing ergosterol with implications for phenomena such as lipid rafts and drug interactions.

Comparative study on the anti-tumor effect of steroids derived from different organisms in H22 tumor-bearing mice and analysis of their mechanisms.

This study aimed to comparatively investigate the anti-tumor mechanisms of steroids including ergosterol, ß-sitosterol, cholesterol, and fucosterol. The model of H22 tumor-bearing mice was constructed based on histopathological data and biochemical parameters, while serums were subjected to metabolomics analysis to study the potential anti-tumor mechanisms. The results indicated that the four steroids exhibited different degrees of anti-tumor effects on H22 mice. The tumor inhibition rates were 63.25% for ergosterol, 56.41% for ß-sitosterol, 61.54% for cholesterol, and 72.65% for fucosterol. Metabolomic analyses revealed that 87, 71, and 129 differential metabolites were identified in ergosterol, cholesterol, and fucosterol treatment groups, respectively. The fucosterol treatment group had the highest number of differential metabolites. At the same time, it mainly inhibited purine and amino acid metabolism to exert anti-tumor effects. Ergosterol enhanced immunity and affected pyruvate metabolism, and cholesterol inhibited purine metabolism. The chemical structure difference among ergosterol, cholesterol, and fucosterol is mainly at the number and position of sterol double bonds and the number and length of side chain carbons. Therefore, there is a structure-activity relationship between the structure of steroid compounds and their efficacy. This study provides a key foundation for the exploitation of the anti-tumor effects of steroids derived from different organisms.

How Does the Antibiotic Amphotericin B Enter Membranes and What Does It Do There?

Amphotericin B is a popular antifungal antibiotic, but the exact way it works is still a matter of debate. Here, we used monolayers composed of phosphatidylcholine with ergosterol as a model of fungal lipid membranes to study drug incorporation from the aqueous phase and analyze the molecular reorganization of membranes underlying the biological activity of the antibiotic. The results show that the internalization of antibiotic molecules into membranes occurs only in the presence of ergosterol in the lipid phase. Comparison of images of solid-supported monolayers obtained by atomic force microscopy and lifetime imaging fluorescence microscopy shows the formation of intramembrane clusters of various sizes in the lipid phase, consisting mainly of antibiotic dimers and relatively large membrane pores (∼15 nm in diameter). The results reveal multiple modes of action of amphotericin B, acting simultaneously, each of which adversely affects the structural properties of the lipid membranes and their physiological functionality.

Water-in-oil Pickering emulsion using ergosterol as an emulsifier solely.

As a crucial component of the fungal cell membranes, ergosterol has been demonstrated to possess surface activity attributed to its hydrophobic region and polar group. However, further investigation is required to explore its emulsification behavior upon migration to the oil-water interface. Therefore, this study was conducted to analyze the interface properties of ergosterol as a stabilizer for water in oil (W/O) emulsion. Moreover, the emulsion prepared under the optimal conditions was utilized to load the water-soluble bioactive substance with the chlorogenic acid as the model molecules. Our results showed that the contact angle of ergosterol was 117.017°, and its dynamic interfacial tension was obviously lower than that of a pure water-oil system. When the ratio of water to oil was 4: 6, and the content of ergosterol was 3.5 % (ergosterol/oil phase, w/w), the W/O emulsion had smaller particle size (438 nm), higher apparent viscosity, and better stability. Meanwhile, the stability of loaded chlorogenic acid was improved under unfavorable conditions (pH 1.2, 90 °C, ultraviolet irradiation, and oxidation), which were 73.87 %, 59.53 %, 62.53 %, and 69.73 %, respectively. Additionally, the bioaccessibility of chlorogenic acid (38.75 %) and ergosterol (33.69 %), and the scavenging rates of the emulsion on DPPH radicals (81.00 %) and hydroxyl radicals (82.30 %) were also enhanced. Therefore, a novel W/O Pickering emulsion was prepared in this work using ergosterol as an emulsifier solely, which has great potential for application in oil-based food and nutraceutical formulations.

Ten ring-B aromatized ergosterols from Aspergillus spectabilis.

Spectasterols F-O (1-10), ten interesting ergosterols with an aromatized B ring, were obtained from Aspergillus spectabilis. Their structures and absolute configurations were determined using a combination of high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), nuclear magnetic resonance (NMR) spectroscopy, single-crystal X-ray diffraction analyses, and electronic circular dichroism (ECD) calculations. Structurally, these aromatic ergosterols feature versatile side chains. Notably, compound aromatic ergosterols featured versatile side chains, and compound 4 is an unusual C23 ergosterol characterized by a shorter side chain due to oxidative cleavage between C-23 and C-24. All compounds were evaluated for their neuroprotective activities, with compound 8 showing a dose-dependent ability to reduce apoptosis and protect mitochondrial function in glutamate-induced SH-SY5Y cells.

Psathrosterols C-E, highly conjugated ergosterol derivatives from Psathyrella rogueiana with anti-inflammatory and immunosuppressive activity.

Three new ergosterols featuring with a highly conjugated ring system, psathrosterols C-E (1-3), have been isolated from the fungus Psathyrella rogueiana. The structures with the absolute configurations were elucidated by means of spectroscopic methods and single crystal X-ray diffraction. Compounds 1-3 exhibit inhibitory activity against NO production with IC50 values ranging from 8.4 to 21.8 µM. Compound 1 inhibits the LPS-induced proliferation of B lymphocyte cells with an IC50 value of 12.3 µM.

Minor ergosteroids and a 19-nor labdane-type diterpenoid with anti-inflammatory effects from Ganoderma lucidum.

A chemical investigation on the fruiting bodies of Ganoderma lucidum led to the isolation and identification of five undescribed ergosteroids including two des-D-steroids (3 and 4) and one rare 6/6/7/5-fused carbon skeletal ergosterol (5) along with one 19-nor labdane-type diterpenoid (6). Their structures including their absolute configurations, were assigned by spectroscopic methods, ECD calculations, and X-ray diffraction analysis. In addition, the anti-inflammatory activities of all the isolates were evaluated. The results indicated that compound 1 can significantly down-regulate the protein expression of iNOS and COX-2 at 20 µM in LPS- stimulated RAW264.7 cells.

Ergosterols with rare peroxide, oxetane ring moiety, and a lactone ring from Aspergillus spectabilis and their immunosuppressive activities.

Ten ergostane-type steroids, including seven undescribed ones named spectasteroids A-G, were obtained from Aspergillus spectabilis. Their structures and absolute configurations were determined based on HRESIMS, NMR, ECD calculations, and single-crystal X-ray diffraction analyses. Structurally, spectasteroid A was a unique example of aromatic ergostane-type steroid that featured a rare peroxide ring moiety; spectasteroid B contained a rare oxetane ring system formed between C-9 and C-14; and spectasteroid C was an unusual 3,4-seco-ergostane steroid with an extra lactone ring between C-3 and C-9. Spectasteroids F and G specifically showed inhibitory effects against concanavalin A-induced T lymphocyte proliferation and lipopolysaccharide-induced B lymphocyte proliferation, with IC50 values ranging from 2.33 to 4.22 µM. Spectasteroid F also showed excellent antimultidrug resistance activity, which remarkable enhanced the inhibitory activity of PTX on the colony formation of SW620/Ad300 cells.

Optimization of feeding strategy for the ergosterol production by yeasts saccharomyces cerevisiae / Optimización de la estrategia de alimentación para la producción de ergosterol por levaduras saccharomyces cerevisiae

Objective of this study was to optimize ergosterol production by yeast strain Saccharomyces cerevisiae with the use of computer controlled feeding of cultivation medium. Baker´s yeasts strain of Saccharomyces cerevisiae originally modified and selected as mutant D7 was further applied in an industrial scale and also in this investigation. Composition of cultivation medium was optimized with the use of a modified Rosenbrock´s method with regard to following components: glucose, yeast extract, ammonium sulphate, potassium dihydrogen phosphate, magnesium sulphate and calcium chloride. Cultivation of yeast culture was performed in 7 L laboratory bioreactor with a working volume of 5 L equipped with a control unit and linked to a computer, with dissolved oxygen tension measurement, oxygen and carbon dioxide analyzers. BIOGENES prototype software was created from the commercial control system Genesis for Windows 3.0 (GFW), from Iconics and CLIPS 6.04 for the PC-Windows platform. From various factors affecting sterol biosynthesis a specific growth rate was chosen. Feed rate was controlled according to mathematical model. In this case it dealt with a design of optimal profile of specific growth rate with consequent calculation of carbon dioxide profile. Sterol concentration in the dry biomass increased from 1.0 % up to 3 %.

El objetivo de este estudio fue optimizar la producción de ergosterol por una cepa de levadura Saccharomyces cerevisiae, controlando la alimentación de medio de cultivo por computadora. La cepa de levadura panadera Saccharomyces cerevisiae originalmente modificada y seleccionada como mutante D7 fue posteriormente utilizada a escala industrial y también para esta investigación. La composición del medio de cultivo fue optimizada usando el método modificado de Rosenbrock respecto a los siguientes componentes: glucosa, extracto de levadura, sulfato de amonio, fosfato dihidrógeno de potasio, sulfato de magnesio y cloruro de calcio. El cultivo de las células de levadura se llevó a cabo en un biorreactor de laboratorio de 7L con un volumen de trabajo de 5L, equipado con una unidad de control conectada a una computadora, con medición de la tensión de oxígeno disuelto y analizadores de oxígeno y dióxido de carbono. Un software prototipo BIOGENES fue creado a partir del sistema de control comercial Genesis para Windows 3.0 (GFW), de Iconics y CLIPS 6.04 para la plataforma de PC-Windows. A partir de varios factores que afectan la biosíntesis de esterol se escogió una tasa específica de crecimiento. La tasa de alimentación se controló mediante un modelo matemático. En este caso, se trató con un diseño de perfil óptimo de tasa de crecimiento específico con un consecuente cálculo del perfil de dióxido de carbono. La concentración de esterol en la biomasa seca se incrementó desde 1,0% hasta 3%.

Ergosterol biosynthesis and drug development for Chagas disease

This article presents an overview of the currently available drugs nifurtimox (NFX) and benznidazole (BZN) used against Trypanosoma cruzi, the aetiological agent of Chagas disease; herein we discuss their limitations along with potential alternatives with a focus on ergosterol biosynthesis inhibitors (EBI). These compounds are currently the most advanced candidates for new anti-T. cruzi agents given that they block de novo production of 24-alkyl-sterols, which are essential for parasite survival and cannot be replaced by a host's own cholesterol. Among these compounds, new triazole derivatives that inhibit the parasite's C14± sterol demethylase are the most promising, as they have been shown to have curative activity in murine models of acute and chronic Chagas disease and are active against NFX and BZN-resistant T. cruzi strains; among this class of compounds, posaconazole (Schering-Plough Research Institute) and ravuconazole (Eisai Company) are poised for clinical trials in Chagas disease patients in the short term. Other T. cruzi-specific EBI, with in vitro and in vivo potency, include squalene synthase, lanosterol synthase and squalene epoxidase-inhibitors as well as compounds with dual mechanisms of action (ergosterol biosynthesis inhibition and free radical generation), but they are less advanced in their development process. The main putative advantages of EBI over currently available therapies include their higher potency and selectivity in both acute and chronic infections, activity against NFX and BZN-resistant T. cruzi strains, and much better tolerability and safety profiles. Limitations may include complexity and cost of manufacture of the new compounds. As for any new drug, such compounds will require extensive clinical testing before being introduced for clinical use, and the complexity of such studies, particularly in chronic patients, will be compounded by the current limitations in the verification of true parasitological...