A binderless, bulk modified, renewable surface amperometric sensor for NADH and ethanol

Graphite particles are exfoliated and subsequently functionalized with toluidine blue. The resulting covalently modified graphite particles are restacked without any binder to form a surface-renewable, bulk-modified electrode. Electrocatalytic oxidation of NADH and its application in the amperometric biosensing of ethanol using alcohol dehydrogenase enzyme have been demonstrated with this material.

Mechanisms of azole resistance and carcinogenic acetaldehyde production in chronic mucosal candidosis

Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED, APS1) is an autoimmune disease caused by a loss-of function mutation in the autoregulator gene (AIRE). Patients with APECED suffer from chronic mucocutaneous candidosis (CMC) of the oral cavity and oesophagus often since early childhood. The patients are mainly colonized with Candida albicans and decades of exposure to antifungal agents have lead to the development of clinical and microbiological resistance in the treatment of CMC in the APECED patient population in Finland. A high incidence of oral squamous cell carcinoma is associated with oral CMC lesions in the APECED patients over the age of 25. The overall aim of this study was firstly, to investigate the effect of long-term azole exposure on the metabolism of oral C. albicans isolates from APECED patients with CMC and secondly, to analyse the specific molecular mechanisms that are responsible for these changes. The aim of the first study was to examine C. albicans strains from APECED patients and the level of cross-resistance to miconazole, the recommended topical compound for the treatment of oral candidosis. A total of 16% of the strains had decreased susceptibility to miconazole and all of these isolates had decreased susceptibility to fluconazole. Miconazole MICs also correlated with MICs to voriconazole and posaconazole. A significant positive correlation between the years of miconazole exposure and the MICs to azole antifungal agents was also found. These included azoles the patients had not been exposed to. The aim of our second study was to determine if the APECED patients are continuously colonized with the same C. albicans strains despite extensive antifungal treatment and to gain a deeper insight into the genetic changes leading to azole resistance. The strains were typed using MLST and our results confirmed that all patients were persistently colonized with the same or a genetically related strain despite antifungal treatment between isolations. No epidemic strains were found. mRNA expression was analysed by Northern blotting, protein level by western blotting, and TAC1 and ERG11 genes were sequenced. The main molecular mechanisms resulting in azole resistance were gain-of-function mutations in TAC1 leading to over expression of CDR1 and CDR2, genes linked to azole resistance. Several strains had also developed point mutations in ERG11, another gene linked to azole resistance. In the third study we used gas chromatography to test whether the level of carcinogenic acetaldehyde produced by C. albicans strains isolated from APECED patients were different from the levels produced by strains isolated from healthy controls and oral carcinoma patients. Acetaldehyde is a carcinogenic product of alcohol fermentation and metabolism in microbes associated with cancers of the upper digestive tract. In yeast, acetaldehyde is a by-product of the pyruvate bypass that converts pyruvate into acetyl-CoA during fermentation. Our results showed that strains isolated from APECED patients produced mutagenic levels of acetaldehyde in the presence of glucose (100mM, 18g/l) and the levels produced were significantly higher than those from strains isolated from controls and oral carcinoma patients. All strains in the study, however, were found to produce mutagenic levels of acetaldehyde in the presence of ethanol (11mM). The glucose and ethanol levels used in this study are equivalent to those found in food and beverages and our results highlight the role of dietary sugars and ethanol on carcinogenesis. The aims of our fourth study were to research the effect of growth conditions in the levels of acetaldehyde produced by C. albicans and to gain deeper insight into the role of different genes in the pyruvate-bypass in the production of high acetaldehyde levels. Acetaldehyde production in the presence of glucose increased by 17-fold under moderately hypoxic conditions compared to the levels produced under normoxic conditions. Under moderately hypoxic conditions acetaldehyde levels did not correlate with the expression of ADH1 and ADH2, genes catalyzing the oxidation of ethanol to acetaldehyde, or PDC11, the gene catalyzing the oxidation of pyruvate to acetaldehyde but correlated with the expression of down-stream genes ALD6 and ACS1. Our results highlight a problem where indiscriminate use of azoles may influence azole susceptibility and lead to the development of cross-resistance. Despite clinically successful treatment leading to relief of symptoms, colonization by C. albicans strains is persistent within APECED patients. Microevolution and point mutations that occur in strains may lead to the development of azole-resistant isolates and metabolic changes leading to increased production of carcinogenic acetaldehyde.

Pd Supported on Titanium Nitride for Efficient Ethanol Oxidation

The excellent metal support interaction between palladium (Pd) and titanium nitride (TiN) is exploited in designing an efficient anode material. Pd-TN, that could be useful for direct ethanol fuel cell in alkaline media. The physicochemical and electrochemical characterization of the Pd-TiN/electrolyte interface reveals an efficient oxidation of ethanol coupled with excellent stability of the catalyst under electrochemical conditions. Characterization of the interface using in situ Fourier transform infrared spectroscopy (in situ FITR) shows the production CO2 at low overvoltages revealing an efficient cleaving of the C-C bond. The performance comparison of Pd supported on TiN (Pd-TiN) with that supported on carbon (Pd-C) clearly demonstrates the advantages of TiN support over carbon. A positive chemical shift of Pd (3d) binding energy confirms the existence of metal support interaction between pd and TiN, which in turn helps weaken the Pd-CO synergetic bonding interaction. The remarkable ability of TiN to accumulate -OH species on its surface coupled with the strong adhesion of Pd makes TiN an active support material for electrocatalysts.

Production of Carcinogenic Acetaldehyde by Oral Microbiome

Oral cancer is the seventh most common cancer worldwide and its incidence is increasing. The most important risk factors for oral cancer are chronic alcohol consumption and tobacco smoking, up to 80 % of oral carcinomas are estimated to be caused by alcohol and tobacco. They both trigger an increased level of salivary acetaldehyde, during and after consumption, which is believed to lead to carcinogenesis. Acetaldehyde has multiple mutagenic features and it has recently been classified as a Group 1 carcinogen for humans by the International Agency for Research on Cancer. Acetaldehyde is metabolized from ethanol by microbes of oral microbiota. Some oral microbes possess alcohol dehydrogenase enzyme (ADH) activity, which is the main enzyme in acetaldehyde production. Many microbes are also capable of acetaldehyde production via alcohol fermentation from glucose. However, metabolism of ethanol into acetaldehyde leads to production of high levels of this carcinogen. Acetaldehyde is found in saliva during and after alcohol consumption. In fact, rather low ethanol concentrations (2-20mM) derived from blood to saliva are enough for microbial acetaldehyde production. The high acetaldehyde levels in saliva after alcohol challenge are explained by the lack of oral microbiota and mucosa to detoxify acetaldehyde by metabolizing it into acetate and acetyl coenzymeA. The aim of this thesis project was to specify the role of oral microbes in the in vitro production of acetaldehyde in the presence of ethanol. In addition, it was sought to establish whether microbial metabolism could also produce acetaldehyde from glucose. Furthermore, the potential of xylitol to inhibit ethanol metabolism and acetaldehyde production was explored. Isolates of oral microbes were used in the first three studies. Acetaldehyde production was analyzed after ethanol, glucose and fructose incubation with gas chromatography measurement. In studies I and III, the ADH enzyme activity of some microbes was measured by fluorescence. The effect of xylitol was analyzed by incubating microbes with ethanol and xylitol. The fourth study was made ex vivo and microbial samples obtained from different patient groups were analyzed. This work has demonstrated that isolates of oral microbiota are able to produce acetaldehyde in the presence of clinically relevant ethanol and glucose concentrations. Significant differences were found between microbial species and isolates from different patient groups. In particular, the ability of candidal isolates from APECED patients to produce significantly more acetaldehyde in glucose incubation compared to healthy and cancer patient isolates is an interesting observation. Moreover, xylitol was found to reduce their acetaldehyde production significantly. Significant ADH enzyme activity was found in the analyzed high acetaldehyde producing streptococci and candida isolates. In addition, xylitol was found to reduce the ADH enzyme activity of C. albicans. Some results from the ex vivo study were controversial, since acetaldehyde production did not correlate as expected with the amount of microbes in the samples. Nevertheless, the samples isolated from patients did produce significant amounts of acetaldehyde with a clinically relevant ethanol concentration.

Ethanol-dependent oxygen consumption and acetaldehyde formation during vanadyl oxidation by H2O2

Sequential addition of vanadyl sulfate to a phosphate-buffered solution of H2O2 released oxygen only after the second batch of vanadyl. Ethanol added to such reaction mixtures progressively decreased oxygen release and increased oxygen consumption during oxidation of vanadyl by H2O2. Inclusion of ethanol after any of the three batches of vanadyl resulted in varying amounts of oxygen consumption, a property also shared by other alcohols (methanol, propanol and octanol). On increasing the concentration of ethanol, vanadyl sulfate or H2O2, both oxygen consumption and acetaldehyde formation increased progressively. Formation of acetaldehyde decreased with increase in the ratio of vanadyl:H2O2 above 2:1 and was undetectable with ethanol at 0.1 mM. The reaction mixture which was acidic in the absence of phosphate buffer (pH 7.0), released oxygen immediately after the first addition of vanadyl and also in presence of ethanol soon after initial rapid consumption of oxygen, with no accompanying acetaldehyde formation. The results underscore the importance of some vanadium complexes formed during vanadyl oxidation in the accompanying oxygen-transfer reactions.

Renewable surface electrodes based on dopamine functionalized exfoliated graphite: NADH oxidation and ethanol biosensing

Exfoliated graphite (EG) was modified by covalently attaching dopamine (DA) (3,4-dihydroxyphenethylamine) through amide linkages, using -COOH groups introduced on the EG surface. The modified material was characterized by FT-IR spectroscopy, Xray photoelectron spectroscopy and electrochemical techniques. Composites of DA modified EG dispersed in organically modified silicates were prepared by a sol-get process. Electrodes were fabricated by casting the composites in glass tubes. The sol-gel based electrodes were found to be active for the electrocatalytic oxidation of NADH and biosensing of ethanol in presence of NAD(+) and alcohol dehydrogenase enzyme. The modified composite electrodes were found to be stable for several months. The surface of the electrode could be renewed just by mechanically polishing the electrode using emery sheets. The modified EG was also pressed and restacked in the form of a pellet and the use of this material as a binderless bulk-modified electrode was also demonstrated. The performance of sol-gel derived composite EG electrodes with binderless bulk-modified EG electrodes was compared. (C) 2002 Elsevier Science B.V. All rights reserved.

Ethanol sensing using CuO/MWNT thin film

Copper (II) oxide (CuO)/multiwall carbon nanotube (MWNT) thin film based ethanol-sensors were fabricated by dispersing CVD-prepared MWNTs in varying concentration over DC magnetron sputtered-CuO films. The responses of these sensors as a function of MWNT concentrations and temperatures were measured, and compared. The sensing response was the maximum at an operating temperature near 400 degrees C for all the samples irrespective of the MWNTs dispersed over them. At optimum operating temperature (T(opt)) of 407 +/- 1 degrees C, the response is linear for 100-700 ppm range and tends to saturate at higher concentrations. In comparison with bare CuO sample, the response of CuO/MWNT sensing films increased up to 50% in the linear range. The response improvement for 2500 ppm of ethanol was up to 90% compared to bare CuO sample. In addition, the sensing response time also reduced to around 23% for lowest ethanol concentration at T(opt). However, a decrease in the sensor response was observed on films with very high concentrations of MWNTs. (C) 2011 Elsevier B.V. All rights reserved.

Structural Transformations, Composition Anomalies and a Dramatic Collapse of Linear Polymer Chains in Dilute Ethanol-Water Mixtures

Water-ethanol mixtures exhibit many interesting anomalies, such as negative excess partial molar volume of ethanol, excess sound absorption coefficient at low concentrations, and positive deviation from Raoult's law for vapor pressure, to mention a few. These anomalies have been attributed to different, often contradictory origins, but a quantitative understanding is still lacking. We show by computer simulation and theoretical analyses that these anomalies arise from the sudden emergence of a bicontinuous phase that occurs at a relatively low ethanol concentration of x(eth) approximate to 0.06-0.10 (that amounts to a volume fraction of 0.17-0.26, which is a significant range!). The bicontinuous phase is formed by aggregation of ethanol molecules, resulting in a weak phase transition whose nature is elucidated. We find that the microheterogeneous structure of the mixture gives rise to a pronounced nonmonotonic composition dependence of local compressibility and nonmonotonic dependence in the peak value of the radial distribution function of ethyl groups. A multidimensional free energy surface of pair association is shown to provide a molecular explanation of the known negative excess partial volume of ethanol in terms of parallel orientation and hence better packing of the ethyl groups in the mixture due to hydrophobic interactions. The energy distribution of the ethanol molecules indicates additional energy decay channels that explain the excess sound attenuation coefficient in aqueous alcohol mixtures. We studied the dependence of the solvation of a linear polymer chain on the composition of the water-ethanol solvent. We find that there is a sudden collapse of the polymer at x(eth) approximate to 0.05-a phenomenon which we attribute to the formation of the microheterogeneous structures in the binary mixture at low ethanol concentrations. Together with recent single molecule pulling experiments, these results provide new insight into the behavior of polymer chain and foreign solutes, such as enzymes, in aqueous binary mixtures.

Influence of ethanol/water mixture on the undrained shear strength of pure clays

The mechanical properties of clays are highly dependent not only on the stress/strain ratio to which the material is subjected but also on the chemistry of the pore fluids which in turn affects the intergranular or the effective stresses. Atterberg limits and vane shear tests were performed with different pore fluids in order to observe how the fine-grained material mechanically responded. The diffuse double layer theory has been used to interpret the data of vane shear tests in order to explain the variation of geotechnical responses with the different clays. Van der Waals forces and double layer forces were obtained and capillary forces calculated. The results show that while for kaolinite and illite the chemistry of the pore fluids has no influence on the water content and hence on the mechanical behaviour of the material, Na-smectite shows a strong correlation between the dielectric constant of the pore fluids and an increase in undrained shear strength. The data obtained extends an understanding of the influence of the dielectric constant (epsilon) of the pore fluids on the geotechnical properties of fine-grained materials.

Spray characterization of gasoline-ethanol blends from a multi-hole port fuel injector

This work reports the measured spray structure and droplet size distributions of ethanol-gasoline blends for a low-pressure, multi-hole, port fuel injector (PFI). This study presents previously unavailable data for this class of injectors which are widely used in automotive applications. Specifically, gasoline, ethanol, and gasoline-ethanol blends containing 10%, 20% and 50% ethanol were studied using laser backlight imaging, and particle/droplet image analysis (PDIA) techniques. The fuel mass injected, spray structure and tip penetrations, droplet size distributions, and Sauter mean diameter were determined for the blends, at two different injection pressures. Results indicate that the gasoline and ethanol sprays have similar characteristics in terms of spray progression and droplet sizes in spite of the large difference in viscosity. It appears that the complex mode of atomization utilized in these injectors involving interaction of multiple fuel jets is fairly insensitive to the fuel viscosity over a range of values. This result has interesting ramifications for existing gasoline fuel systems which need to handle blends and even pure ethanol, which is one of the renewable fuels of the future. (C) 2012 Elsevier Ltd. All rights reserved.