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PubMed2006

The acid tolerance response of Bacillus cereus ATCC14579 is dependent on culture pH, growth rate and intracellular pH.

The food pathogen Bacillus cereus is likely to encounter acidic environments (i) in food when organic acids are added for preservation purposes, and (ii) during the stomachal transit of aliments. In order to characterise the acid stress response of B. cereus ATCC14579, cells were grown in chemostat at different pH values (pH(o) from 9.0 to 5.5) and different growth rates (micro from 0.1 to 0.8 h(-1)), and were submitted to acid shock at pH 4.0. Cells grown at low pH(o) were adapted to acid media and induced a significant acid tolerance response (ATR). The ATR induced was modulated by both pH(o) and micro, and the micro effect was more marked at pH(o) 5.5. Intracellular pH (pH(i)) was affected by both pH(o) and micro. At a pH(o) above 6, the pH(i) decreased with the decrease of pH(o) and the increase of micro. At pH(o) 5.5, pH(i) was higher compared to pH(o) 6.0, suggesting that mechanisms of pH(i) homeostasis were induced. The acid survival of B. cereus required protein neo-synthesis and the capacity of cells to maintain their pH(i) and DeltapH (pH(i) - pH(o)). Haemolysin BL and non-haemolytic enterotoxin production were both influenced by pH(o) and micro.

Acids↗

PubMed2007

Ordered adsorption of coagulation factor XII on negatively charged polymer surfaces probed by sum frequency generation vibrational spectroscopy.

Electrostatic interactions between negatively charged polymer surfaces and factor XII (FXII), a blood coagulation factor, were investigated by sum frequency generation (SFG) vibrational spectroscopy, supplemented by several analytical techniques including attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), quartz crystal microbalance (QCM), zeta-potential measurement, and chromogenic assay. A series of sulfonated polystyrenes (sPS) with different sulfonation levels were synthesized as model surfaces with different surface charge densities. SFG spectra collected from FXII adsorbed onto PS and sPS surfaces with different surface charge densities showed remarkable differences in spectral features and especially in spectral intensity. Chromogenic assay experiments showed that highly charged sPS surfaces induced FXII autoactivation. ATR-FTIR and QCM results indicated that adsorption amounts on the PS and sPS surfaces were similar even though the surface charge densities were different. No significant conformational change was observed from FXII adsorbed onto surfaces studied. Using theoretical calculations, the possible contribution from the third-order nonlinear optical effect induced by the surface electric field was evaluated, and it was found to be unable to yield the SFG signal enhancement observed. Therefore it was concluded that the adsorbed FXII orientation and ordering were the main reasons for the remarkable SFG amide I signal increase on sPS surfaces. These investigations indicate that negatively charged surfaces facilitate or induce FXII autoactivation on the molecular level by imposing specific orientation and ordering on the adsorbed protein molecules.

Adsorption↗

PubMed2001

Quantitative analysis of active compounds in pharmaceutical preparations by use of attenuated total-reflection Fourier transform mid-infrared spectrophotometry and the internal standard method.

A new method is presented for the quantitative analysis of compounds in pharmaceutical preparations Fourier transform (FT) mid-infrared (MIR) spectroscopy with an attenuated total reflection (ATR) module. Reduction of the quantity of overlapping absorption bands, by interaction of the compound of interest with an appropriate solvent, and the employment of an internal standard (IS), makes MIR suitable for quantitative analysis. Vigabatrin, as active compound in vigabatrin 100-mg capsules, was used as a model compound for the development of the method. Vigabatrin was extracted from the capsule content with water after addition of a sodium thiosulfate IS solution. The extract was concentrated by volume reduction and applied to the FTMIR-ATR module. Concentrations of unknown samples were calculated from the ratio of the vigabatrin band area (1321-1610 cm(-1)) and the IS band area (883-1215 cm(-1)) using a calibration standard. The ratio of the area of the vigabatrin peak to that of the IS was linear with the concentration in the range of interest (90-110 mg, in twofold; n=2). The accuracy of the method in this range was 99.7-100.5% (n=5) with a variability of 0.4-1.3% (n=5). The comparison of the presented method with an HPLC assay showed similar results; the analysis of five vigabatrin 100-mg capsules resulted in a mean concentration of 102 mg with a variation of 2% with both methods.

Anticonvulsants↗

PubMed2007

Analysis of urinary calculi using an infrared microspectroscopic surface reflectance imaging technique.

This investigation highlights the use of infrared microspectroscopy for the morphological analysis of urinary stones. The research presented here has utilized the reflectance mode of an infrared microscope for use in creating chemically specific maps of cross-sectioned renal calculi surfaces, precisely showing the placement of renal stone components in a calculus sample. The method has been applied to renal stones of both single and multiple components consisting primarily of hydroxyapatite, calcium oxalate monohydrate and calcium oxalate dihydrate. Factors discussed include the photometric accuracy of the spectra obtained, a comparison of the surface reflectance method with existing methods such as diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) and attenuated total internal reflection (ATR) analysis, and the influence of specular reflectance between polished and unpolished sample spectra. Full spectral maps of cross-sectioned renal stones provided positive localization of components using qualitatively accurate spectra similar in appearance to DRIFTS spectra. Unlike ATR and DRIFTS spectra, surface reflectance spectra lack photometric accuracy and are therefore not quantifiable; at present, however, spectra are suitable for qualitative analysis. It was found that specular reflectance increases minimally with a highly polished stone cross-section surface, though qualitative data is not affected. Surface reflectance imaging of sections of renal stones is useful for determining the identity of stone components while simultaneously providing precise locations of mineral components within the stone using presently available instruments.

Animals↗

PubMed2005

Interfacial interaction of tartaric acid with hydroxyapatite and enamel.

The purpose of this study was to investigate the interfacial interaction of tartaric acid with hydroxyapatite and enamel. Hydroxyapatite particles were mixed with 15% (w/v) alcohol-aqueous (1:1) solution of tartaric acid for 72 h. After the mixture, the filtrate was separated from the solid. The filtrate, the solid, hydroxyapatite, and tartaric acid were dried for FTIR and XRD analysis. Enamel disks etched with tartaric acid for 60 s were analyzed by attenuated total reflectance (ATR). Enamel disks etched with 15% tartaric acid for 15, 30, 60, 120 s were analyzed by SEM. FTIR spectra showed carboxylate in the solid and in subtraction spectra. ATR spectra revealed carboxylate on the enamel surfaces. XRD data indicated the formation of calcium tartrate and calcium hydrogen phosphate after the reaction of tartaric acid with hydroxyapatite. SEM observations revealed typical decalcification of the periphery of the enamel rods due tartaric acid etching. It is concluded that tartaric acid can decalcify and chemisorb onto HA simultaneously, hence, tartaric acid could be used as etchant and ingredient of self-etching primers in adhesive dentistry.

Acid Etching, Dental↗

PubMed2005

Characterization of trypsin immobilized on the functionable alkylthiolate self-assembled monolayers: a preliminary application for trypsin digestion chip on protein identification using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Self-assembled monolayers (SAMs) on coinage metal provide versatile modeling systems for studies of interfacial electron transfer, biological interactions, molecular recognition and other interfacial phenomena. Recently the bonding of enzyme to SAMs of alkanethiols onto Au electrode surfaces was exploited to produce a bio-sensing system. In this work, the attachment of trypsin to a SAMs surface of 11-mercaptoundecanoic acid was achieved using water soluble N-ethyl-N '-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide as coupling agent. The thickness of SAMs was determined by optical ellipsometer; contact angles of the modified Au surfaces were measured in air using a goniometer. The Second Harmony Generation data displays the last few percents of the alkylthiol molecules adsorbed and produced the complete monolayer by inducing the transition from a high number of gauche defects to an all-trans conformation. Using X-ray Photoelectron Spectroscopy (XPS) and Fourier-Transformed Infrared Reflection-Absorption and Attenuated Total Reflection Spectroscopes (FTIR-RAS and ATR), we examined the chemical structures of samples with different treatments. By matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), we demonstrated the digestion of bovine serum albumin (BSA) on the trypsin-immobilized SAMs surface. Experimental results have revealed that the XPS C1s core levels at 286.3 and 286.5 eV (Amine bond), 288.1 eV (Amide bond) and 289.3 eV (Carboxylic acid) illustrate the immobilization of trypsin. These data were also in good agreement with FTIR-ATR spectra for the peaks valued at 1659.4 cm(-1) (Amide I) and 1546.6 cm(-1) (Amide II). Using MALDI-TOF MS observations, analytical results have demonstrated the BSA digestion of the immobilized trypsin on the functionalized SAMs surface. For such surfaces, BSA was digested on the trypsin-immobilized SAMs surface, which shows the enzyme digestion ability of the immobilized trypsin. The terminal groups of the SAMs structure can be further functionalized with biomolecules or antibodies to develop surface-base diagnostics, biosensors, or biomaterials.

Biosensing Techniques↗

PubMed2004

Photochemical production of conjugated linoleic acid from soybean oil.

Conjugated linoleic acid (CLA), an anticarcinogenic compound with numerous other health benefits, is present mainly in dairy and beef lipids. The main CLA isomer present in dairy and beef lipids is cis 9, trans 11 CLA at a 0.5% concentration. The typical minimum human dietary intake of CLA is 10 times less than the 3 g/d suggested requirement that has been extrapolated from animal and cell-line studies. The objectives of this study were to produce CLA isomers from soybean oil by photoisomerization of soybean oil linoleic acid and to study the oxidation status of the oil. Refined, bleached, and deodorized soybean oil with added iodine concentrations of 0, 0.1, 0.25, and 0.5% was exposed to a 100-W mercury lamp for 0 to 120 h. An SP-2560 fused-silica capillary GC column with FID was used to analyze the esterified CLA isomers in the photoisomerized oil. The CLA content of the individual isomers was optimized by response surface methodology. Attenuated total reflectance (ATR)-FTIR spectra in the 3400 to 3600 cm(-1) range and 1H NMR spectra in the 8 to 12 ppm range of the photoisomerized soybean oil were obtained to follow hydroperoxide formation. The largest amount of cis 9, trans 11 CLA isomer in soybean oil was 0.6%, obtained with 0.25% iodine and 84 h of photoisomerization. Lipid hydroperoxide peaks in the ATR-FTIR spectra and aldehyde peaks in the 1H NMR spectra were not observed in the photoisomerized soybean oil, and the spectra were similar to that of fresh soybean oil. This study shows that CLA isomers can be produced simply and inexpensively from soybean oil by photoisomerization.

Hydrogen Peroxide↗

PubMed1988

A technique for monitoring mammalian cell growth and inhibition in situ via Fourier transform infrared spectroscopy.

A single culture of Chinese hamster ovary cells was grown on germanium attenuated total reflectance (ATR) crystals and continuously monitored in situ via ATR/Fourier transform infrared (FT-IR) spectroscopy for approximately 60 h. The cells were seeded into a specially designed flow cell which controlled physiological conditions, flow rate, and addition of growth medium or metabolic inhibitors. Infrared spectra were taken at 20-min intervals until a confluent monolayer was formed. Several strong bands are evident in the spectra which can be generally ascribed to molecular features of cellular components. Cell growth kinetics were measured as a function of infrared band intensity over time and exhibited the normal lag phase, logarithmic growth, and stationary phase on reaching confluence. Spectra of growing cells, normalized to the area under the spectral region 1800-1000 cm-1, were subtracted from reference spectra of confluent cells at 60 h. Difference spectra showed that the largest differences were observed between confluent cells and cells in early growth stages. Differences may reflect cell morphological changes, biochemical activity, and degree of ATR crystal exposure to the bulk medium. ATR/FT-IR spectroscopy of living Chinese hamster ovary cells was also used in a toxicological study to monitor the effects of hydroxyurea, an inhibitor of DNA synthesis. Delayed growth was observed in the cell growth curve of the hydroxyurea-treated cells during the course of treatment as compared to the control culture.

Animals↗

PubMed1994

The effect of topical anesthetics on skin sensation and soleus motoneuron reflex excitability.

The effect that topical anesthetics has on cutaneous sensation and on soleus motoneuron reflex excitability was studied in a healthy population. Data were obtained from 16 volunteers (7 men and 9 women) between 20 and 47 years of age. The experiment consisted or recording H-reflexes and Achilles tendon reflexes (ATR) before and after administration of a 20% benzocaine spray or a placebo to the skin of the posterior calf. In addition, a sensory evaluation was conducted to determine objectively the actual quality of anesthesia obtained. Subjects served as their own controls, randomly receiving both the active medication and a placebo. The study was conducted in a single blind fashion with only the experimenter being aware of what medication (active/placebo) was being applied during a given experiment. H-reflex amplitudes increased significantly following the application of benzocaine and the placebo spray compared to control (p < .01). This increase was similar whether the active drug or placebo was administered. No significant difference was shown between active benzocaine and the placebo application. The ATR was not significantly affected by any of the treatment techniques used in this study, although some reduction in reflex amplitude was noted. No change in skin sensibility could be attributed to the topical anesthetic in any subject in this study. It was concluded that benzocaine spray did not have an effect on skin sensation and that the increased motoneuron reflex excitability was more likely in response to a stimulatory action upon cutaneous receptors from the spray technique. It is further suggested that topical anesthetics applied to intact skin may not permeate cutaneous tissue in large enough quantities to have a desensitizing effect.

Achilles Tendon↗

PubMed1986

Orientation of gramicidin D incorporated into phospholipid multibilayers: a Fourier transform infrared-attenuated total reflection spectroscopic study.

Polarized Fourier transform infrared (FTIR)-attenuated total reflection (ATR) spectroscopy was applied to study the orientation of the linear pentadecapeptide antibiotic gramicidin D incorporated into phospholipid multibilayers, which were cast on a germanium ATR plate from chloroform solution. In DMPC and DPPC multibilayers, the CH2 stretching bands of lipid hydrocarbon chains were slightly shifted to the higher frequency side and bandwidth was increased in the presence of gramicidin. However, in DPPE multibilayers, frequencies and bandwidths of these bands were unaltered. In each case, gramicidin produced little effect on the orientation of lipid hydrocarbon chains, suggesting that gramicidin penetrates into lipid layers without noticeable perturbations. Upon incubation of cast films in contact with water above the gel-liquid-crystalline transition temperature (Tc) of lipids, the reorientation of gramicidin in lipid multibilayers occurred, the degree thereof depending upon the fluidity of the lipid hydrocarbon chains and the amount of surrounding water. In DMPC multibilayers, the helix axis of gramicidin was oriented almost parallel to the lipid hydrocarbon chains after incubation. In DPPC multibilayers, on the other hand, the helix axis of gramicidin was tilted on average about 15 degrees from the lipid hydrocarbon chains after incubation. However, in DPPE multibilayers, which are known to have the most rigid bilayer structures, the reorientation of gramicidin could not be seen.

Gramicidin↗

PubMed1985

Subfornical organ neurons with efferent projections to the hypothalamic paraventricular nucleus: an electrophysiological study in the rat.

Seventeen neurons in the subfornical organ (SFO) were antidromically activated by electrical stimulation of the paraventricular nucleus (PVN) in the rat. The activity of all identified SFO neurons was excited by microiontophoretically (MIPh) applied angiotensin II (AII) and the effect of AII was blocked by MIPh-applied saralasin (Sar), an AII antagonist, but not by atropine (Atr), a muscarinic antagonist. In these identified SFO neurons, 9 were also excited and 8 were not affected by MIPh-applied acetylcholine (ACh) and the effect of ACh was attenuated by not only MIPh-applied Atr but also by Sar. These results suggest that there are specific AII- and both AII- and ACh-sensitive types of SFO neurons with efferent projections to the PVN.

Acetylcholine↗

PubMed1982

Topical anesthesia: modulation of the monosynaptic reflexes by desensitization of the skin.

Skin desensitization by topical anesthesia was studied for its effect on the motoneuron excitability of the soleus muscle. Skin areas overlying calf, tibial, quadriceps and hamstrings muscles and skin dermatomes (L2, L3, L4, L5, S1 and S2) were studied separately. Motoneuron excitabilities were measured by the H-reflex and Achilles tendon reflex (for alpha and gamma motoneurons). It was shown that anesthesia applied to all skin areas and dermatomes, except those overlying the antagonist muscles, resulted in significant facilitation of the soleus H-reflex. In these cases, the ATR showed either slight inhibition or no significant changes. Anesthesia to the skin overlying the anterior tibial antagonistic muscle produced varied and inconsistent modifications in the amplitude of the H-reflex. In these cases the ATR was either slightly facilitated or showed no significant changes. These results appear to indicate the existence of on-going excitatory and inhibitory effects from the skin on the alpha and gamma motoneuron pool probably via segmental and suprasegmental levels. A possible clinical application of these results to modulate the motoneuron pool excitability is proposed.

Administration, Topical↗

PubMed1983

Bronchodilator characteristics of nebulized metaproterenol sulfate, isoetharine, and atropine in chronic asthma.

A randomized double-blind comparison of aerosolized META, ISO, ATR, and PI solutions delivered by motorized mist nebulizer was conducted in subjects with known reversible airway obstruction. Vital signs and spirometric parameters were monitored sequentially during each of four 6 hr test periods. Abstention from all medication was required for at least 12 hr before each test period. Baseline percent predicted FEV1 and FEF25-75 among subjects for all test periods were statistically similar. Compared to the other agents tested META treatment (15 mg) resulted in significantly improved global response and higher mean percent increases in FEV1 and FEF25-75 over the entire test period as well as at individual measurement points up to 4 hr for FEV1 and 5 hr for FEF25-75. Peak FEV1 and FEF25-75 values with META treatment were noted at 30 min and persisted above baseline during the entire 6 hr. ATR (2 mg) treatment resulted in later onset of bronchodilatory effect and, as with ISO (125 micrograms), more rapid diminution in measured response. Administration of saline alone resulted in a net 9% increase in FEV1 and 11% in FEF25-75 over the entire test period. Clinically insignificant tremor was noted in 41% of META-treated subjects. Delivery of META by a motorized mist nebulizer provides safe, effective bronchodilation and META is superior to the other agents tested in terms of onset of action, peak reversal of airway obstruction, and duration of effect.

Adolescent↗

PubMed1984

Surface analysis of biomedical polymers by attenuated total reflectance-fourier transform infra-red.

An infra- red approach has been developed to characterize hydrophilic biomaterials, particularly contact lenses. Water was utilized as the optical coupling agent between the sample and the ATR element. This method enables the study of hydrophilic polymers in their natural aqueous environment and simultaneously solves the optical contact problem that usually arises when the ATR technique is used. Some studies were made utilizing this approach: dry and hydrated samples were structurally compared; also, a depth profiling study and a surface comparison were made on soft contact lenses fabricated by different industrial processes. Finally, the three dimensional orientation of one hydrated structure was characterized.

Biocompatible Materials↗

PubMed1984

Characterization of protein adsorption on soft contact lenses. I. Conformational changes of adsorbed human serum albumin.

Adsorption of human serum albumin on three different soft contact lens surfaces (lathe cut and spin cast crosslinked PHEMA and spin cast PHEMA/MAA) was studied. Using ATR--FTIR spectroscopy the spectra of the adsorbed protein were obtained at different times of adsorption. Structural changes were detected, initially characterized by an increase in hydrogen bonding and after that by involvement of the protein hydrophobic side chain residues. At long adsorption times, the protein becomes denatured, its alpha-helix content is drastically reduced and the amounts of random coil and beta-sheet conformations are increased. ATR-FTIR and circular dichroism studies of albumin solutions reveal similar conformational changes to those experienced by the adsorbed protein. Differences in the adsorption behaviour for the hydrogel surface, indicate the importance of the hydrophilicity, surface regularity and the chemical composition of the contact lens surfaces as the controlling parameters in the protein adsorption phenomena.

Adsorption↗

PubMed1986

Surface modification of soft contact lenses: silanization, wettability and lysozyme adsorption studies.

The surface reactions of poly(2-hydroxyethylmethacrylate) (PHEMA) and the copolymer poly(HEMA-methacrylic acid) (PHEMA/MAA) with methyltrimethoxysilane, ethyltrimethoxysilane and phenyltrimethoxysilane have been characterized by attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy. A model compound, hydroxyethyl isobutyrate was synthesized and subsequently reacted with phenyltrimethoxysilane. Its FTIR spectrum was compared with the ATR-FTIR spectra mentioned above. Protein adsorption experiments showed that silanized PHEMA/MAA soft contact lenses adsorbed less lysozyme than the untreated lenses.

Adsorption↗

PubMed1994

In vitro exposure of a novel polyesterurethane graft to enzymes: a study of the biostability of the Vascugraft arterial prosthesis.

The biostability of the Vascugraft arterial prosthesis, a porous synthetic graft made by a novel spinning process from a unique poly(ester urethane) polymer, has been studied by means of an in vitro enzyme incubation technique. Samples of the Vascugraft were exposed to buffered solutions of collagenase and pancreatin, as well as the buffer solutions alone, for periods of up to 100 days at 37 +/- 1 degrees C. On removal and after cleaning, a number of different analytic methods, including X-ray photoelectron spectroscopy for chemical analysis (ESCA), attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), differential scanning calorimetry (DSC), size exclusion chromatography (SEC), scanning electron microscopy (SEM), interference microscopy, moisture content and contact angle measurements, were used to examine the changes in chemical structure and surface morphology of the samples. During incubation in both enzymes the molecular weight of the polyurethane appeared to decrease in the presence of enzyme but increase in the presence of buffer. Further microphase separation in the polyurethane material developed during incubation in buffer solutions. Such changes in microstructure were associated with increased surface hydrophilicity, increased moisture content and a significant improvement in the extent of order and preferred orientation of the hard segment domains within the fibres. In the sampling depth of about 5 nm, both enzymes decreased the carbonate group content at the surface of the prosthesis to as little as 40% of their original values. The results from ATR-FTIR and DSC demonstrated that this phenomenon was limited primarily to the soft segment phase. While the Vascugraft prosthesis did exhibit some limited chemical modifications on exposure to concentrated enzyme solutions, nevertheless such changes were confined to the surface layer of the polyurethane microfibres. The importance and significance of those results will be more adequately determined by in vivo investigation.

Blood Vessel Prosthesis↗

PubMed1996

Plasma-induced grafted polymerization of acrylic acid and subsequent grafting of collagen onto polymer film as biomaterials.

Polyacrylic acid (pAA) was introduced onto Ar-plasma treatment silicone rubber (SR) membrane surfaces by plasma-induced grafted polymerization. Collagen (type III) was also linked with the carboxylic group of pAA grafted onto the SR surface via a carbodiimine agent to obtain a secondary structure of SR. The SR surface properties were characterized by ATR-FTIR, ESCA, contact angle, and SEM. The biocompatibility of the SR surface was evaluated by a culture of cornea epithelial (CE) cells. Subsequently, 75-450 micrograms cm-2 of pAA were obtained on the SR surfaces under different reactive conditions; 3-12 micrograms cm-2 of collagen were linked on modified surfaces of SR. Moreover, ATR-FTIR and ESCA were utilized to confirm the proceedings of these reactions. The hydrophility of the modified SR was measured by a contact angle meter. The values of contact angle for SR grafted with pAA were approximately 45-50 degrees; a 50-55 degrees contact angle on pAA-g-SR to be further linked with collagen was subsequently obtained. Moreover, the influence of surface properties toward migration, growth and attachment of CE cells on the modified surfaces was also examined. Here, untreated SR was used as a control. Experimental results indicated that the number of CE cells attached onto the controlled SR was negligible. The attachment of cells onto pAA-grafted surfaces was clearly observed and peusopoda occurred; however, cell growth was depressed. This depression may have been caused by the acid environment of the pAA-grafted membrane. Nevertheless, both cell attachment and growth onto collagen-linked surfaces were significant. In addition, the morphology of the cells attached onto this surface was considered normal for primary cells. Collagen introduced on the SR surface was not denatured, i.e the natural properties of collagen were maintained. The results obtained in this study will hopefully lead to the successful development of modified SR for clinical applications.

Acrylic Resins↗