Impact on S. aureus and E. coli Membranes of Treatment with Chlorhexidine and Alcohol Solutions: Insights from Molecular Simulations and Nuclear Magnetic Resonance.

Membranes form the first line of defence of bacteria against potentially harmful molecules in the surrounding environment. Understanding the protective properties of these membranes represents an important step towards development of targeted anti-bacterial agents such as sanitizers. Use of propanol, isopropanol and chlorhexidine can significantly decrease the threat imposed by bacteria in the face of growing anti-bacterial resistance via mechanisms that include membrane disruption. Here we have employed molecular dynamics simulations and nuclear magnetic resonance to explore the impact of chlorhexidine and alcohol on the S. aureus cell membrane, as well as the E. coli inner and outer membranes. We identify how sanitizer components partition into these bacterial membranes, and show that chlorhexidine is instrumental in this process.

Effect of povidone-iodine and propanol-based mecetronium ethyl sulphate on antimicrobial resistance and virulence in Staphylococcus aureus.

BACKGROUND: Reports are available on cross-resistance between antibiotics and biocides. We evaluated the effect of povidone-iodine (PVP-I) and propanol-based mecetronium ethyl sulphate (PBM) on resistance development, antibiotics cross-resistance, and virulence in Staphylococcus aureus. METHODS: The minimum inhibitory concentration (MIC) of PVP-I and PBM were determined against S. aureus ATCC 25923 using the agar-dilution method. Staphylococcus aureus ATCC 25923 was subjected to subinhibitory concentrations of the tested biocides in ten consecutive passages followed by five passages in a biocide-free medium; MIC was determined after each passage and after the fifth passage in the biocide-free medium. The developed resistant mutant was tested for cross-resistance to different antibiotics using Kirby-Bauer disk diffusion method. Antibiotic susceptibility profiles as well as biocides' MIC were determined for 97 clinical S. aureus isolates. Isolates were categorized into susceptible and resistant to the tested biocides based on MIC distribution pattern. The virulence of the biocide-resistant mutant and the effect of subinhibitory concentrations of biocides on virulence (biofilm formation, hemolysin activity, and expression of virulence-related genes) were tested. RESULTS: PVP-I and PBM MIC were 5000 µg/mL and 664 µg/mL. No resistance developed to PVP-I but a 128-fold increase in PBM MIC was recorded, by repeated exposure. The developed PBM-resistant mutant acquired resistance to penicillin, cefoxitin, and ciprofloxacin. No clinical isolates were PVP-I-resistant while 48.5% were PBM-resistant. PBM-resistant isolates were more significantly detected among multidrug-resistant isolates. PVP-I subinhibitory concentrations (» and ½ of MIC) completely inhibited biofilm formation and significantly reduced hemolysin activity (7% and 0.28%, respectively). However, subinhibitory concentrations of PBM caused moderate reduction in biofilm activity and non-significant reduction in hemolysin activity. The ½ MIC of PVP-I significantly reduced the expression of hla, ebps, eno, fib, icaA, and icaD genes. The virulence of the biocide-resistant mutant was similar to that of parent strain. CONCLUSION: PVP-I is a highly recommended antiseptic for use in healthcare settings to control the evolution of high-risk clones. Exposure to PVP-I causes no resistance-development risk in S. aureus, with virulence inhibition by subinhibitory concentrations. Also, special protocols need to be followed during PBM use in hospitals to avoid the selection of resistant strains.

Volatiles of antagonistic soil yeasts inhibit growth and aflatoxin production of Aspergillus flavus.

Minimizing Aspergillus flavus growth is an effective strategy to mitigate aflatoxin contamination in food and agricultural products. In the present investigation, we attempted to utilize soil-associated yeasts from the Western and Eastern Ghats of India against A. flavus to reduce aflatoxin contamination. Forty-five yeast isolates were screened against A. flavus using overlay and dual plate assays. Among them, 12 isolates effectively inhibited the growth of A. flavus. The 18S rDNA gene sequence analysis identified the twelve antagonistic isolates as belonging to Saccharomyces cerevisiae, Suhomyces xylopsoci, Pichia kudriavzevii, and Candida tropicalis. From the isolated yeasts, S. cerevisiae strains were selected for further evaluation based on the potential antagonistic activity. Volatiles of S. cerevisiae effectively suppressed the mycelial growth of A. flavus (P < 0.05) up to 92.1 % at 7 DAI. Scanning electron microscopic images of the fungus exposed to volatiles showed hyphal deformity and mycelial damage. Aflatoxin B1 (AFB1) production was drastically reduced up to 99.0 % in the volatile-exposed fungus compared to the control. The yeast strain YKK1 showed consistent Aspergillus flavus growth inhibition (80.7 %) and AFB1 production (98.1 %) for 14 days. Gas chromatography-mass spectrophotometry analysis of the yeast volatiles revealed the presence of antimicrobial compounds, including 1-pentanol, 1-propanol, ethyl hexanol, ethanol, 2-methyl-1-butanol, ethyl acetate, dimethyl trisulfide, p-xylene, styrene, and 1,4-pentadiene. The evaluated compounds of yeast volatiles, including ethyl acetate, hexanal, 1-propanol, 1-heptanol, 1-butanol, and benzothiazole, inhibited the fungal growth and AFB1 production of Aspergillus flavus when applied as pure chemicals. Benzothiazole at 5 mM was responsible for a high level of growth inhibition (23.6 %) and reduction of AFB1 synthesis (93.5 %). Hence, volatile compounds produced by soil yeast strains could be a potential biocontrol mechanism against aflatoxin contamination.

Olfactory Detection Thresholds for Primary Aliphatic Alcohols in Mice.

Probing the neural mechanisms that underlie each sensory system requires the presentation of perceptually appropriate stimulus concentrations. This is particularly relevant in the olfactory system as additional odorant receptors typically respond with increasing stimulus concentrations. Thus, perceptual measures of olfactory sensitivity provide an important guide for functional experiments. This study focuses on aliphatic alcohols because they are commonly used to survey neural activity in a variety of olfactory regions, probe the behavioral limits of odor discrimination, and assess odor-structure activity relationships in mice. However, despite their frequent use, a systematic study of the relative sensitivity of these odorants in mice is not available. Thus, we assayed the ability of C57BL/6J mice to detect a homologous series of primary aliphatic alcohols (1-propanol to 1-heptanol) using a head-fixed Go/No-Go operant conditioning assay combined with highly reproducible stimulus delivery. To aid in the accessibility of our data, we report the animal's threshold to each odorant according to the 1) ideal gas condition, 2) nonideal gas condition (factoring in the activity of the odorant in the solvent), and 3) the liquid dilution of the odorant in the olfactometer. Of the odorants tested, mice were most sensitive to 1-hexanol and least sensitive to 1-butanol. These updated measures of murine sensitivity will hopefully guide experimenters in choosing appropriate stimulus concentrations for experiments using these odorants.

Investigating the interaction of porcine pancreatic elastase and propanol: A spectroscopy and molecular simulation study.

The response of porcine pancreatic elastase (PPE) to propanol was examined by various techniques including UV-vis spectrophotometry, spectrofluorometry and circular dichroism, as well as molecular docking and molecular simulation. These techniques were used to investigate the structural changes and elastase activity in the presence of propanol. This work was performed at three temperatures of 303, 313 and 323 K, with the pH value of 8.5 (Tris buffer). The results of the UV-vis spectrophotometry indicated the transfer of tryptophan to an environment with low hydrophobicity. Fluorescence measurements also revealed the quenching of fluorescence intensity was induced by propanol, and dynamic quenching was the proposed quenching mechanism. Kinetic studies also suggested the inhibitory effect (noncompetitive) of propanol on elastase. Further, Circular Dichroism (CD) spectra showed that propanol caused slight alterations in the secondary structures of PPE (0.3% increase for the α-helix and 0.5% decrease for the ß-sheet). Addition of propanol decreased the Tm (Melting Temperature) parameter from 332.8 K to 330.1 K.

Using 1-propanol to significantly enhance the production of valuable odd-chain fatty acids by Rhodococcus opacus PD630.

Odd-chain fatty acids (OCFAs) have been reported to possess pharmacological activity and have been used in the manufacture of agricultural and industrial chemicals. We here provided a new method to increase the OCFAs content in oil produced by Rhodococcus opacus PD630 through addition of 1-propanol to the fermentation media. The OCFAs in oil of R. opacus PD630 are primarily pentadecanoic acid (C15:0), heptadecanoic acid (C17:0) and heptadecenoic acid (C17:1). After adding 0.5-1.5% (v/v) 1-propanol, the production of oil increased from 1.27 g/L to 1.31-1.61 g/L, and the OCFAs content in oil increased by 46.7-55.1%. Metabolic intermediates determination and transcriptome analysis revealed that R. opacus assimilated 1-propanol through methylmalonyl-CoA pathway. When the nitrogen source was limited, propionyl-CoA was converted to propionyl-acyl carrier protein (ACP) which could be used as primer during the elongation of fatty acid synthesis. Then OCFAs were produced when odd number of propionyl-ACP was incorporated in the cycles of fatty acid synthesis.

Bactericidal Activity of Ready-To-Use Alcohol-Based Commercial Wipes According to EN 16615 Carrier Standard.

BACKGROUND: The effectiveness of ready-to-use disinfectant wipes was previously assessed in standardized suspension tests, which were inadequate because they ignored that the wipes are rubbed against a surface. Thus, we assessed the effectiveness of commercially available disinfectant wipes impregnated with an alcoholic solution according to the 16615 standard, which includes a test with mechanical action. METHODS: According to the EN 16615 standard, under clean conditions, four squares (5cm x 5 cm), placed next to one another, were marked on a test surface. Enterococcus hirae, Pseudomonas aeruginosa, and Staphylococcus aureus were inoculated on the leftmost square, and a wipe impregnated with an alcoholic solution was placed to the left of that square. Then, the wipe was pressed with a 2.5 kg weight and moved to the right and back to the left. After contact times of 1, 5, 10, or 15 minutes, we measured the reduction in bacterial load. RESULTS: Alcohol-based ready-to-use commercial wipes did not show sufficient bactericidal activity at the contact times of 1, 5, 10 and 15 minutes. Wipes containing propan-1-ol and a mixture of propan-1-ol and propan-2-ol were active against Pseudomonas aeruginosa at the contact times of 1 minute and 15 minutes. None of the examined wipes were active against Enterococcushirae or Staphylococcusaureus. CONCLUSION: Bactericidal parameters of ready-to-use disinfectant wipes should be determined in surface tests, in addition to suspension tests, because suspension tests do not simulate the conditions under which disinfectant wipes are used in practice.

High Environmental Stability of Hepatitis B Virus and Inactivation Requirements for Chemical Biocides.

Hepatitis B virus (HBV) infection is considered a major public health problem worldwide, and a significant number of reports on nosocomial and occupational outbreaks have been reported. This systematic investigation of HBV stability and susceptibility to different antiseptics revealed that HBV infectivity was very stable, with a half-life of >22 days at 37°C. At 4°C, infectivity was barely reduced for up to 9 months. Different alcohols and commercially available hand antiseptics had a virucidal effect against HBV. We propose that very strict compliance with established hygienic guidelines should be mandatory to avoid and prevent HBV infections.

Plant phenyl-propanoids-conjugated silver nanoparticles from edible plant Suaeda maritima (L.) dumort. Inhibit proliferation of K562-human myeloid leukemia cells.

The present study portrays the isolation of four phenylpropanoids - ferulic acid (FA), sinapic acid (SA), caffeic acid (CA), and chlorogenic acid (CHA) from the water extract of Suaeda maritima (L.) Dumort, a phytochemically less explored Indian medicinal plant. Further, synthesis and characterization of silver and gold nanoparticles using the isolated phenylpropanoids were done. The silver nanoparticles synthesized from S. maritima water extract along with silver nano-conjugated forms of the isolated compounds exhibited appreciable anti-leukemic activity against K562 cells (human myeloid leukemia). Especially, the ferulic and CA-conjugated silver nanoparticles showed significant (P < .01) activity against leukemia.

Antimicrobial evaluation of selected naturally occurring oxyprenylated secondary metabolites.

This study tested the antimicrobial activity of eight selected naturally occurring oxyprenylated secondary metabolites against Staphylococcus aureus ATCC 29213, S. epidermidis ATCC 35984, Escherichia coli ATCC 8739, Pseudomonas aeruginosa ATCC 9027 and Candida albicans ATCC 10231. Results showed a moderate antimicrobial activity. The most active compounds were 3-(4-geranyloxyphenyl)-1-ethanol (4) and 3-(4-isopentenyloxyphenyl)-1-propanol (5) that were tested on mature and in-formation biofilms of all micro-organisms, moreover the cytotoxic activity was evaluated. Except for S. epidermidis, both compounds reduced significantly (p < 0.05) the microbial biofilm formation at 1/2 MIC and 1/4 MIC, in particular, compounds 4 and 5 at each concentration, inhibited E. coli biofilm formation to a greater extent, the biofilm formation was never more than 44% in respect to the control, moreover both compounds showed a low cytotoxic effect. Oxyprenylated derivatives may be of great interest for the development of novel antimicrobial therapeutic strategies and the synthesis of semi-synthetic analogues with anti-biofilm efficacy.

Controlling the feed rate of propanol to optimize erythromycin fermentation by on-line capacitance and oxygen uptake rate measurement.

The aim of the present study was to optimize the feeding proportion of glucose and propanol for erythromycin biosynthesis by real-time monitoring and exploring its limited ratio by the on-line multi-frequency permittivity measurement. It was found that the capacitance values were sensitive to the variation of biomass concentration and microbial morphology as well as the true state of cell growth. It was most favorable to both cell growth and secondary metabolism to keep the ratio of glucose to propanol at 4.3 (g/g). The specific growth rate calculated by the capacitance measurement correctly and accurately reflected the cell physiological state. An appropriate feed rate of propanol was crucial for cell growth and secondary metabolism, as well as to improve the quality of erythromycin-A. In addition, the erythromycin production titer (10,950 U/mL) was further enhanced by 4 % when the propanol feed was regulated by step-down strategy based on both OUR (oxygen uptake rate) and the on-line monitoring capacitance.

Use of three-carbon chain compounds as biosynthesis precursors to enhance tacrolimus production in Streptomyces tsukubaensis.

Tacrolimus, a 23-membered polyketide macrolide, is isolated as a metabolite from the whole fermentation broth of Streptomyces species. This potent immunosuppressive calcineurin inhibitor has been widely used in various fields of medicine, including transplantology, dermatology and pharmacotherapy of autoimmune diseases. The current study was focused on optimisation of tacrolimus biosynthesis by Streptomyces tsukubaensis through the use of culture media supplements. Enrichment of the fermentation medium with propionic acid, propylene glycol or propanol resulted in a 5.5-, 3.5- and 1.8-fold, respectively, increase in tacrolimus production. The optimal concentration of the precursors was 0.25% for both propanol and propionic acid and 0.75% for propylene glycol. The mode of action of each media supplement tested was unique. For instance, propionic acid acted as a tacrolimus biosynthesis precursor while propylene glycol induced mycelial growth of S. tsukubaensis. Results from the current study clearly demonstrate that a set of novel culture medium supplements considerably increased tacrolimus production. Application of such promoters of tacrolimus biosynthesis may lead to a substantial improvement in the production of tacrolimus by S. tsukubaensis in industrial fermentation processes.

Synthesis of new N-acryl-1-amino-2-phenylethanol and N-acyl-1-amino-3-aryloxypropanols and evaluation of their antihyperlipidemic, LDL-oxidation and antioxidant activity.

As a part of our drug discovery program, we identified an alkaloidal amide i.e. Aegeline (V) isolated from the leaves of Aegle marmelos as a dual acting agent (antihyperlipidemic and antihyperglycemic). In continuation of this program, we synthesized new N-acyl-1-amino-2-alcohols (N-acrylated-1-amino-2-phenylethanol and N-acylated-1-amino-3-aryloxypropanols) via Ritter reaction and screened for their in-vivo antihyperlipdemic activity in Triton induced hyperlipidemia model, LDL-oxidation and antioxidant activity. Compounds 3, 11 and 13 showed good antihyperlipidemic activity, LDL-oxidation as well as antioxidant activity and comparable activity with marketed antidyslipidemic drug.

Effects of atropine and propranolol on lung inflammation in experimental envenomation: comparison of two buthidae venoms

Background: Previous works had shown that scorpion venom induced neurotransmitter elevation and an inflammatory response associated with various anatomo-pathological modifications. The most dangerous scorpions species in Algeria responsible for these effects are Androctonus australis hector (Aah) and Androctonus amoreuxi (Aam). Results: Comparison of the physiopathological effects induced by the two venoms showed differences in the kinetic of cytokine release and in lung injury. The lung edema was only observed in response to Aah venom and it was correlated with cell infiltration. In order to better understand the involved mechanism in inflammatory response, we used two antagonists, atropine (non-selective muscarinic antagonist) and propranolol (ß adrenergic antagonist), which lead to a decrease of cell infiltration but has no effect on edema forming. Conclusion: These results suggest another pathway in the development of lung injury following envenomation with Aam or Aah venom.(AU)

Polyproline fold-In imparting kinetic stability to an alkaline serine endopeptidase.

Polyproline II (PPII) fold, an unusual structural element was detected in the serine protease from Nocardiopsis sp. NCIM 5124 (NprotI) based on far UV circular dichroism spectrum, structural transitions of the enzyme in presence of GdnHCl and a distinct isodichroic point in chemical and thermal denaturation. The functional activity and conformational transitions of the enzyme were studied under various denaturing conditions. Enzymatic activity of NprotI was stable in the vicinity of GdnHCl upto 6.0M concentration, organic solvents viz. methanol, ethanol, propanol (all 90% v/v), acetonitrile (75% v/v) and proteases such as trypsin, chymotrypsin and proteinase K (NprotI:protease 10:1). NprotI seems to be a kinetically stable protease with a high energy barrier between folded and unfolded states. Also, an enhancement in the activity of the enzyme was observed in 1M GdnHCl upto 8h, in organic solvents (75% v/v) for 72h and in presence of proteolytic enzymes. The polyproline fold remained unaltered or became more prominent under the above mentioned conditions. However, it diminished gradually during thermal denaturation above 60°C. Thermal transition studies by differential scanning calorimetry (DSC) showed scan rate dependence as well as irreversibility of denaturation, the properties characteristic of kinetically stable proteins. This is the first report of PPII helix being the global conformation of a non structural protein, an alkaline serine protease, from a microbial source, imparting kinetic stability to the protein.

Substituted phenoxypropyl-(R)-2-methylpyrrolidine aminomethyl ketones as histamine-3 receptor inverse agonists.

Optimization of a series of aminomethyl ketone diamine H(3)R antagonists to reduce the brain exposure by lowering the pKa, led to molecules with improved pharmacokinetic properties. Compounds 9, 19, and 25 had high affinity for human H(3)R and demonstrated in vivo H(3)R functional activity in the rat dipsogenia model. Compound 9 displayed modest wake-promoting activity in the rat EEG/EMG model.

Effect of rat brain tissue extracts on osteoblast proliferation and differentiation.

PURPOSE: The reason for enhanced fracture healing in traumatic brain injury patients is not clearly understood. It is possible that factors inherent in the brain passing through the blood-brain barrier to the peripheral circulation, or a disruption of central nervous system (CNS) control of the sympathetic nervous system (SNS), stimulates the process of fracture healing. METHODS: In this study, we assessed proliferation [using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay] and differentiation [using alkaline phosphatase (ALP)] in rat osteoblasts incubated with gray matter or other tissue extracts with and without the addition of an α- or ß-adrenergic receptor blocker (phentolamine or propranolol). RESULTS: Gray matter extract from normal brain caused a dose-dependent increase in osteoblast proliferation and differentiation. Serum from normal rats enhanced differentiation but not proliferation. Alpha-receptor blockade had no effect on proliferation or differentiation. Beta-receptor blockade caused a partial, but statistically significant, decrease in gray matter stimulation of osteoblast differentiation. CONCLUSION: The results of this study indicate that gray matter extract from normal brain increases osteoblast proliferation and differentiation and that ß receptors may be involved in differentiation under these conditions.

Inactivation and survival of hepatitis C virus on inanimate surfaces.

BACKGROUND: Hepatitis C virus (HCV) cross-contamination from inanimate surfaces or objects has been implicated in transmission of HCV in health-care settings and among injection drug users. We established HCV-based carrier and drug transmission assays that simulate practical conditions to study inactivation and survival of HCV on inanimate surfaces. METHODS: Studies were performed with authentic cell culture derived viruses. HCV was dried on steel discs and biocides were tested for their virucidal efficacy against HCV. Infectivity was determined by a limiting dilution assay. HCV stability was analyzed in a carrier assay for several days or in a drug transmission assay using a spoon as cooker. RESULTS: HCV can be dried and recovered efficiently in the carrier assay. The most effective alcohol to inactivate the virus was 1-propanol, and commercially available disinfectants reduced infectivity of HCV to undetectable levels. Viral infectivity on inanimate surfaces was detectable in the presence of serum for up to 5 days, and temperatures of about 65-70°C were required to eliminate infectivity in the drug transmission assay. CONCLUSIONS: These findings are important for assessment of HCV transmission risks and should facilitate the definition of stringent public health interventions to prevent HCV infections.

Age related alterations of adrenoreceptor activity in erythrocyte membrane.

The aim of the study was the investigation of age-related functional alterations of adrenoreceptors and the effect of agonist and antagonist drugs on age related adrenoreceptor activity in erythrocyte membrane. The impact of isopropanol and propanol on functional activity ß- adrenergic receptors in red blood cell membrane were studied in 50 practically healthy men--volunteers. (I group--75-89 years old, II group--22-30 years old). The EPR signals S1 and S2 were registered in red blood cell membrane samples after incubation with isopropanol and propanol respectively. It was found that decreasing sensitivity (functional activity) of red blood cells membrane adrenoreceptors comes with aging (S1old

Ethanol exposure modulates hepatic S-adenosylmethionine and S-adenosylhomocysteine levels in the isolated perfused rat liver through changes in the redox state of the NADH/NAD(+) system.

Methionine metabolism is disrupted in patients with alcoholic liver disease, resulting in altered hepatic concentrations of S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), and other metabolites. The present study tested the hypothesis that reductive stress mediates the effects of ethanol on liver methionine metabolism. Isolated rat livers were perfused with ethanol or propanol to induce a reductive stress by increasing the NADH/NAD(+) ratio, and the concentrations of SAM and SAH in the liver tissue were determined by high-performance liquid chromatography. The increase in the NADH/NAD(+) ratio induced by ethanol or propanol was associated with a marked decrease in SAM and an increase in SAH liver content. 4-Methylpyrazole, an inhibitor the NAD(+)-dependent enzyme alcohol dehydrogenase, blocked the increase in the NADH/NAD(+) ratio and prevented the alterations in SAM and SAH. Similarly, co-infusion of pyruvate, which is metabolized by the NADH-dependent enzyme lactate dehydrogenase, restored the NADH/NAD(+) ratio and normalized SAM and SAH levels. The data establish an initial link between the effects of ethanol on the NADH/NAD(+) redox couple and the effects of ethanol on methionine metabolism in the liver.