Biomedical subjects

L Novotny

Publications and source records attributed to L Novotny.

At least 19 recordsLinked to original sources

PubMed2007

In vitro and in vivo antileukemic effect of novel dimers consisting of 5-fluorodeoxyuridine and arabinofuranosylcytosine.

Various amphiphilic heterodinucleoside phosphates containing 1-beta-D-arabinofuranosylcytosine (ara-C) and 5- fluorodeoxyuridine (5-FdUrd) have recently been synthesized in order to increase the efficacy of ara-C and 5-FdUrd. Employing growth inhibition and growth recovery assays, we evaluated the in vitro effects of four of these dimers (No. 2, 2A, 3, 10) in L1210 and P388D1 murine leukemia cells. Although ara-C and 5-FdUrd appeared equimolar in all dimers, their contribution to the cytotoxicity of these agents was different. Thus, the liberation of ara-C and 5-FdUrd from their dimeric origin and their subsequent metabolic activation had a different course. In another set of experiments, we examined the in vivo effects of these agents in mice. The dimer with the highest cytotoxicity in vitro exerted the lowest acute toxicity and yielded the lowest therapeutic effect in vivo. The obtained data indicate that dimers with slower liberation of ara-C and 5-FdUrd were less cytotoxic, but prolonged liberation of both antimetabolites protected them from inactivation and extended the time period of therapeutic action. Some of the dimers exceeded the synergistic effects yielded by simultaneous application of both ara-C and 5-FdUrd. The significantly higher therapeutic potential of these new antitumor agents indicates that further studies are warranted.

Animals↗

PubMed2006

Papillomatosis in a European bison.

Five European bison (Bison bonasus) from three European zoos were shipped to the Bukovské Vrchy Hills (Slovakia) in June 2004 and kept together in an acclimatization enclosure. The European bison were released into the wild in December 2004. At that time, papillomas were found at the medial canthus of the left eye of a 12-yr-old female bison. Cutaneous papillomatosis was confirmed histologically. Negative stain transmission electron microscopic examination revealed papillomavirus in the papillomas, and papillomavirus DNA also was detected using the polymerase chain reaction with FAP59 and FAP64 primers. The amplified 413 bp DNA sequence was identical to that of BAPV2 bovine papillomavirus. This paper is the first report of papillomatosis in European bison.

Animals↗

PubMed2006

Mutagenic and carcinogenic potential of menadione.

Menadione (2-methyl-l,4-naphthoquinone) or vitamin K3 is a lipid-soluble substance and promotes the hepatic biosynthesis of blood clotting factors. Carcinogenic potential of menadione was determined by a DC polarography method in strictly anhydrous N,N-dimethylformamide (DMF) in the presence of alpha-lipoic acid. Superoxide anion formation was measured after incubation of rat lung, liver and kidney microsomes with menadione. The genotoxic potential of menadione was investigated using the unscheduled DNA synthesis (UDS) and alkaline elution assays. The parameter of potential menadione carcinogenicity tg alpha was 0.0025 indicating no carcinogenic activity of menadione. Superoxide anion was generated in a concentration- and time-dependent manner when menadione was incubated with microsomes. In the mammalian cells (A 549) used for alkaline elution and UDS assays, menadione was cytotoxic at concentrations above 20 nmol/ml. The use of S9 mix (metabolic activation) fractions decreased the cytotoxicity of menadione. In the concentration range of above 20 nmol/ml menadione was genotoxic in the UDS test in absence of metabolic activation. In the presence of metabolic activation the menadione-induced DNA damage and repair was greatly reduced. Treatment of A 549 lung cells with 4-nitroquinoline-N-oxide (NQO) caused significant formation of DNA single-strand breaks both in the absence and presence of metabolic activation. Treatment of A 549 lung cells with menadione caused formation of DNA single-strand breaks in the absence of S9 mix. In the presence of metabolic activation menadione caused no significant formation of DNA strand breaks. Menadione-induced DNA repair in A 549 cells was concentration-, time-, and temperature- dependent. Measurement of unscheduled DNA (UDS) synthesis (repair) following treatment with NQO and menadione yielded strong UDS responses in the absence of S9 mix. Taken together the results of these studies suggest the mutagenic potential of NQO and menadione. These results indicate that menadione undergoes redox cycling with formation of reactive oxygen species which cause DNA damage and repair without having a carcinogenic potential.

Animals↗

PubMed2005

Decreased levels of advanced glycation end-products in patients with Gilbert syndrome.

Oxidative stress plays an important role in the atherogenesis and bilirubin is one of the most potent antioxidant substances in human body. The aim of the present study was to investigate the impact of hyperbilirubinemia on plasma levels of advanced glycation end-products (AGEs), which contribute to atherogenesis. Two AGEs, pentosidine and Nepsilon-carboxymethyl lysine (CML), were determined in 23 subjects with Gilbert syndrome (GS) and in 21 age-matched healthy controls. Pentosidine was assessed by HPLC and CML was determined with ELISA. Logistic regression analysis was used for multiple adjustments of possible modifying factors. As expected, significantly higher serum bilirubin levels were found in GS subjects as compared to controls (28.9 +/- 9.6 vs 9.7 +/- 2.7 micromol/l, p<0.001). In contrast, serum levels of both AGEs were significantly lower in GS compared to normobilirubinemic controls (median; 25%-75% interquartile range (pentosidine: 1.12; 0.90-1.28 vs 1.31; 1.18-1.58 nmol/g protein, p<0.005; and CML: 6.70; 6.10-7.34 vs 7.33; 6.76-8.20 micromol/g protein, p = 0.01, respectively). Levels of both AGEs remained substantially lower even after adjustment for selected vascular risk and other modifying factors. In subjects with GS elevated serum bilirubin concentrations are associated with lower levels of AGEs. These results are in support of previous data on antioxidant properties of bilirubin.

Bilirubin↗

PubMed2004

Applications of field-enhanced near-field optical microscopy.

Metal nanostructures such as sharp tips can enhance emission yields through shape-induced local field enhancement. The enhancement originates from two mechanisms: surface plasmons and electrostatic lightening rod effects. We present fluorescence imaging using the strong local field created at the apex of a gold tip and demonstrate optical resolution of 25 nm. The enhancement effect gives also rise to photoemission from the tip itself. Measured spectra of the tip emission show a broad band continuum together with a second-harmonic peak. Both continuum and second-harmonic are confined at the apex of the tip. We find that, depending on the spectral position, the photoluminescence originates either from intraband or from interband transitions. The nonlinear response can be described by a single dipole oscillating at the second-harmonic frequency and oriented along the tip axis. These unique properties can be used to map focal fields distributions.

Journal Article↗

PubMed2003

Near-field second-harmonic generation induced by local field enhancement.

The field near a sharp metal tip can be strongly enhanced if irradiated with an optical field polarized along the tip axis. We demonstrate that the enhanced field gives rise to local second-harmonic (SH) generation at the tip surface thereby creating a highly confined photon source. A theoretical model for the excitation and emission of SH radiation at the tip is developed and it is found that this source can be represented by a single on-axis oscillating dipole. The model is experimentally verified by imaging the spatial field distribution of strongly focused laser modes.

Journal Article↗

PubMed2003

Plasmon-coupled tip-enhanced near-field optical microscopy.

Near the cut-off radius of a guided waveguide mode of a metal-coated glass fibre tip it is possible to couple radiation to surface plasmons propagating on the outside surface of the metal coating. These surface plasmons converge toward the apex of the tip and interfere constructively for particular polarization states of the initial waveguide mode. Calculations show that a radially polarized waveguide mode can create a strong field enhancement localized at the apex of the tip. The highly localized enhanced field forms a nanoscale optical near-field source.

Journal Article↗

PubMed2003

Near-field Raman spectroscopy using a sharp metal tip.

Near-field Raman spectroscopy with a spatial resolution of 20 nm is demonstrated by raster scanning a sharp metal tip over the sample surface. The method is used to image vibrational modes of single-walled carbon nanotubes. By combining optical and topographical signals rendered by the single-walled carbon nanotubes, we can separate near-field and far-field contributions and quantify the observed Raman enhancement factors.

Journal Article↗

PubMed2003

Comparison of the carcinogenic potential of streptozotocin by polarography and alkaline elution.

The carcinogenic potential of streptozotocin (STZ) was evaluated by the polarographic determination of its reduction potential in the presence of alpha-lipoic acid and detection of DNA single-strand brakes by alkaline elution. After STZ electrochemical reduction in an anhydrous solvent, the half-wave potential (E1/2 ) was determined to be -1.340 V. The parameter of the carcinogenic potential (tg alpha) for STZ was 0.400. This is in good agreement with WHO data regarding STZ carcinogenicity. Additionally, it is in the good agreement with the tg alpha value determined for the positive control used, N-nitroso-N-methylurea (NMU), which was found to be 0.459. The 3 hours exposure of A549 human lung tumor cells to 250, 500, and 1000 nmol/ml STZ was followed by DNA single-strand brakes detection using the alkaline elution method. NMU, the positive control, was tested under identical experimental conditions at the same concentrations. Without metabolic activation, NMU induced a significant formation of DNA single-strand brakes only at 1000 nmol/ml. In the presence of the metabolic activation, NMU caused a significant, concentration-dependent formation of DNA single-strand brakes. In the absence of metabolic activation, STZ induced no significant formation of DNA single-strand brakes at any concentration used. In the presence of metabolic activation, STZ caused a significant, concentration-dependent formation of DNA single-strand brakes. The results of this study underline the crucial role of using a metabolic activation system when carcinogenic potential of drugs and chemicals is investigated in vitro studies. Results of polarographic experiments and alkaline elution correlate well with each other and they indicate that these methods are useful to early predict the carcinogenic potential of STZ and other xenobiotics.

Carcinogens↗

PubMed2003

Physico-chemical properties and spectrophotometric determination of biologically active 1-alkyl-2-(2-pyridyl)pyridinium bromides.

Physico-chemical properties of compounds prepared from 2,2'-bipyridine, 1-alkyl-2-(2-pyridyl)pyridinium bromides, were investigated by DC polarography and by GC-MS. Their ionization potentials were calculated. Additionally, the formation of associates with bromothymol blue and methyl orange during the spectrophotometric determination was measured. It was determined that 1-alkyl-2-(2-pyridyl)pyridinium ions are reduced by 2 one-electron steps in a DC polarography system. The reduction potentials are not related to the ionization potential values calculated for the substances investigated. The carcinogenic potential (tg alpha) of the parent compound 2,2'-bipyridine and of a series of 1-alkyl derivatives was very low indicating that the compounds are not carcinogenic. The MS fragmentation patterns indicate the low stability of the 1-alkyl substituents. It was shown that 2,2'-bipyridine is either fragmented to two pyridine ions or the--N=CH--fragments are removed. Additionally, spectrophotometric determinations of colored associates of 1-alkyl-2-(2-pyridyl)pyridinium bromides with bromothymol blue and methyl orange were investigated and the optimal condiditions for these determinations are reported.

2,2'-Dipyridyl↗

PubMed2003

The free radical scavenging activity of four flavonoids determined by the comet assay.

Flavonoids are reported to exhibit a wide variety of biological effects, including antioxidant and free radical scavenging activities. The aim of our study was to determine the cytotoxicity of flavonoids quercetin, rutin, apigenin and luteolin and their ability to protect DNA molecules against H2O2-induced damage. Cytotoxicity of studied flavonoids was tested in murine leukemia L1210 cells by the trypan blue exclusion technique. DNA strand breaks were determined using the alkaline single-cell gel electrophoresis (comet assay). Quercetin was found to possess the highest protective effect among the flavonoids studied (45%). The protective activity determined was lower for luteolin (40%). Protective effect of apigenin (600 microM/L) was only marginal (2%). However, at the higher concentration of apigenin (1200 microM/L), this flavonoid induced DNA single strand breaks. This indicates the ability of apigenin to serve as a pro-oxidant. Rutin had no protective effect on DNA single strand breaks induced by H2O2.

Animals↗

PubMed2002

Stability of the new antileukemic 4-pyranone derivative, BTMP, using HPLC and LC-MS analyses.

The stability of the new antileukemic kojic acid derivative, 5-benzyloxy-2-thiocyanatomethyl-4-pyranone (BTMP) was investigated. The degradation of BTMP was studied using specific and reproducible HPLC and LC-MS methods. Accelerated stability studies of BTMP were conducted in 0.1 M hydrochloric acid solution, physiological phosphate buffer solution (pH 7.5) and basic phosphate buffer solution (pH 9.0) at 30, 40 and 60 degrees C, respectively. The degradation of BTMP was found to follow pseudo-first order kinetics. In basic solution (pH 9.0) BTMP underwent rapid hydrolysis at a degradation rate constant (0.183-0.638 h-1) and degradation half-life (3.67-1.06 h) depending on the temperature setting. On the other hand, BTMP was significantly stable in 0.1 M hydrochloric acid solution (kdeg: 0.0017-0.0052 h-1; degradation half-life t1/2: 408.6-135.7 h), whereas in physiological phosphate buffer solution (pH 7.5), BTMP was only moderately stable (kdeg: 0.006-0.231 h-1; degradation half-life: 117.7-3.0 h). Arrhenius plots were constructed to predict the degradation kinetic parameters of BTMP at 25 degrees C and 4 degrees C. LC-MS analyses confirmed the degradation of BTMP in basic solutions and indicated at least two degradation products; namely 5-benzyloxypyran-2-ol-4-one (m/z 217.8) and 2-thiocyanatomethylpyran-5-ol-4-one (m/z 181.6).

Antineoplastic Agents↗

PubMed2001

Longitudinal field modes probed by single molecules.

We demonstrate that a strong longitudinal, nonpropagating field is generated at the focus of a radially polarized beam mode. This field is localized in space and its energy density exceeds the energy density of the transverse field by more than a factor of 2. Single molecules with fixed absorption dipole moments are used to probe the longitudinal field. Vice versa, it is demonstrated that orientations of single molecules are efficiently mapped out in three dimensions by using a radially polarized beam as the excitation source. We also show that there is no momentum or energy transport associated with the longitudinal field.

Journal Article↗

PubMed2001

Liquid chromatographic-mass spectrometric determination of celecoxib in plasma using single-ion monitoring and its use in clinical pharmacokinetics.

Celecoxib is a cyclooxygenase-2 specific inhibitor, that has been recently and intensively prescribed as an anti-inflammatory drug in rheumatic osteoarthritis. A robust, highly reliable and reproducible liquid chromatographic-mass spectrometric assay is developed for the determination of celecoxib in human plasma using sulindac as an internal standard. The run cycle-time is <4 min. The assay method involved extraction of the analytes from plasma samples at pH 5 with ethyl acetate and evaporation of the organic layer. The reconstituted solution of the residue was injected onto a Shim Pack GLC-CN, C18 column and chromatographed with a mobile phase comprised of acetonitrile-1% acetic acid solution (4:1) at a flow-rate of 1 ml/min. The mass spectrometer (LCQ Finnigan Mat) was programmed in the positive single-ion monitoring mode to permit the detection and quantitation of the molecular ions of celecoxib and sulindac at m/z 382 and 357, respectively. The peak area ratio of celecoxib/sulindac and concentration are linear (r2>0.994) over the concentration range 50-1000 ng/ml with a lowest detection limit of 20 ng/ml of celecoxib. Within- and between-day precision are within 1.58-4.0% relative standard deviation and the accuracy is 99.4-107.3% deviation of the nominal concentrations. The relative recoveries of celecoxib from human plasma ranged from 102.4 to 103.3% indicating the suitability of the method for the extraction of celecoxib and I.S. from plasma samples. The validated LC-MS method has been utilized to establish various pharmacokinetic parameters of celecoxib following a single oral dose administration of celecoxib capsules in two selected volunteers.

Celecoxib↗

PubMed2001

Determination of diclofenac sodium, flufenamic acid, indomethacin and ketoprofen by LC-APCI-MS.

A sensitive, selective and accurate high-performance liquid chromatography-mass spectrometry (LC-MS) assay for the determination of selected non-steroidal anti-inflammatory drugs (NSAIDs), namely diclofenac sodium (DIC), flufenamic acid (FLU), indomethacin (IND) and ketoprofen (KET), either individually or in mixtures, was developed. The examined drugs were injected onto Shim-pack GLC-CN column and were eluted with a mobile phase consisting of acetonitrile and 20 mM ammonium acetate solution (5:1 v/v)/pH 7.4 at a flow rate l ml min(-1). The mass spectrometer, operated in the single ion monitoring mode, was programmed to admit the negative ions [M-H] at m/z 295.9 (DIC), 280.1 (FLU), 355.8 (IND) and 252.9 (KET), respectively. The calibration curves were linear (r > or = 0.9993) over the concentration range 50-300 ng ml(-1) (FLU, DIC) and 100-500 ng ml(-1) (KET, IND) with detection limits of 0.5-4.0 ng. The mean predicted concentrations for the analytes were in the range -5.9 and 5.2% of the nominal concentrations. Within-day and between-day precision were in the range of 0.8-9.1% of the R.S.D. Mean recovery percentages of the individual compounds from laboratory-made mixtures and pharmaceutical formulations were (99.5-101.5%) and (100.6-102.2%), respectively.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2001

Antitumor activity of benzamide riboside and its combination with cisplatin and staurosporine.

Benzamide riboside (BR), a new synthetic nucleoside analogue, has demonstrated a potent cytotoxic activity in murine leukemia in vitro. The purpose of the present investigation was to examine the antitumor activity of BR in mice bearing leukemia L1210. The results revealed that BR possesses a potent antitumor activity in vivo. It increases life-span of mice with leukemia. Synergistic cytotoxicity of BR with select DNA damaging agents, cisplatin (cis-Pt) and staurosporine (STP) was examined in MTT chemosensitivity assay, FACS analyses and apoptotic DNA fragmentation on L1210 cells in culture. A simultaneous treatment of leukemia L1210 cells with the combination of BR and STP resulted in synergistic cytotoxicity that correlated with increased apoptotic activity in those cells. On the other hand, treatment of L1210 cells with combination of BR and cis-Pt resulted in antagonistic cytotoxic effect. Finally, to elucidate the synergistic effect of BR and STP in inducing apoptosis, the attention was directed to the activation of cell death processes through various cell cycle signals. This is the first report describing in vivo antitumor activity of BR and its utilization in combination chemotherapy.

Animals↗

PubMed2001

Probing confined fields with single molecules and vice versa.

Single dye molecules are used as local probes to map the spatial distribution of the squared electric field components in the focus of a high numerical aperture lens. Simulated field distributions are quantitatively verified by experimentally obtained fluorescence excitation maps. We show that annular illumination can be used to engineer the field distribution in the focus at a dielectric/air interface such that electric field components in all directions acquire comparable magnitudes. The 3D orientation of molecular absorption dipoles can be determined by comparing measured to simulated image patterns. The presence of longitudinal electric field components in a focus is of particular interest in tip-enhanced scanning near-field optical microscopy.

Journal Article↗

PubMed2001

Near-field effects in single molecule emission.

We present the first experimental proof of the influence of a nearby nano-sized metal object on the angular photon emission by a single molecule. A novel angular sensitive detection scheme is implemented in an existing near-field scanning optical microscope (NSOM). The positioning accuracy ( approximately 1 nm) of the NSOM allows a systematic investigation of the intensity ratio between two different half-spaces as a function of the position of the metal-glass interfaces of the probe with respect to the single emitter. The observed effects are shown to be particularly strong for molecules that are excited mainly below the rims of the aperture. An excellent agreement is found between experiments and numerical simulations for these molecules. The observed angular redistribution of the angular emission of a single molecule could explain the alteration of the emission polarization observed for certain molecules in earlier experiments (Veerman et al. (1999) J. Microsc. 194, 477-482).

Journal Article↗