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PubMed1994

A spectrophotometric assay for the detection of 2-aminophenols in biological samples.

A sensitive and efficient spectrophotometric assay is described for the determination of 2-aminophenols in biological samples. Using ferric ions as a metal catalyst, 2-aminophenol dimerizes in an acidic aqueous environment to 2-hydroxyisophenoxazin-3-one, an intensively colored dye. The newly developed assay is suitable for the detection of 2-aminophenols in the micromolar range. The paper demonstrates that this chemical assay is also applicable for the determination of 2-aminophenols substituted with a halogen at the ortho, meta, or para position of the aromatic ring with respect to the amino moiety, lambda max, and the molar extinction coefficient varying with the substituent pattern. Results obtained for fluorinated 2-aminophenol detection in biological samples by either this new method or 19F NMR are similar. This observation corroborates that the newly developed assay is suitable for detection of 2-aminophenols in biological samples.

Aminophenols↗
PubMed2005

Simultaneous determination of arsenic and selenium in biological samples by HG-AFS.

A new method is proposed for simultaneous determination of traces of arsenic (As) and selenium (Se) in biological samples by hydride-generation double-channel non-dispersive atomic-fluorescence spectrometry (HG-AFS) from tartaric acid media. The effects of analytical conditions on fluorescence signal intensity were investigated and optimized. Interferences from coexisting ions were evaluated. Under optimum conditions linear response ranges above 20 microg L(-1) for As and 32 microg L(-1) for Se were obtained with detection limits of 0.13 and 0.12 microg L(-1), respectively. The precision for eleven-fold determination of As at the 4 microg L(-1) level and of Se at the 8 microg L(-1) level were 2.7 and 1.9% (RSD), respectively. Recoveries of 92.5-95.5% for As and 101.2-108.4% for Se were obtained for four biological samples and two certified biological reference materials. The proposed method has the advantages of simple operation, high sensitivity, and high efficiency; it was successfully used for simultaneous determination of As and Se in biological samples.

Animals↗
PubMed2005

Screening and identification of antioxidants in biological samples using high-performance liquid chromatography-mass spectrometry and its application on Salacca edulis Reinw.

In this study, a new approach was developed for screening and identifying antioxidants in biological samples. The approach was based on significant decreases of the intensities of ion peaks obtained from high-performance liquid chromatography (HPLC) coupled with mass spectrometry (MS) upon reaction with 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radicals. HPLC-MS/MS was further applied to elucidate structures of antioxidant peaks characterized in a spiking test. The new approach could also be used to monitor the reactivity of antioxidants in biological sample with free radicals. The approach was successfully applied to the identification of antioxidants in salak (Salacca edulis Reinw), a tropical fruit that is reported to be a very good source of natural antioxidants, but it was still not clear which compounds were responsible for its antioxidant property. The antioxidants in salak were identified to be chlorogenic acid, (-)-epicatechin, and singly linked proanthocyanidins that mainly existed as dimers through hexamers of catechin or epicatechin. In salak, chlorogenic acid was identified to be an antioxidant of the slow reaction type as it reacted with free radicals much more slowly than either (-)-epicatechin or proanthocyanidins. The new approach was proved to be useful for the characterization and identification of antioxidants in biological samples as a mass detector combined with an HPLC separation system not only serves as an ideal tool to monitor free radical active components but also provides their possible chemical structures in a biological sample.

Antioxidants↗
PubMed1994

Solid-phase extraction technique for the analysis of biological samples.

This article reviews the literature on solid-phase extraction published in the last 10 years. Emphasis has been placed on dealing with samples of biological origin. The sections consist of introduction, history and development, types of columns, selection of a suitable column, types of samples, advantages and applications.

Animals↗
PubMed2000

Direct determination of lead in biological samples by electrothermal vaporization inductively coupled plasma mass spectrometry (ETV-ICP-MS) after furnace-fusion in the sample cuvette-tungsten boat furnace.

The newly conceived electrothermal vaporization (ETV) system using a tungsten boat furnace (TBF) sample cuvette was designed for the direct analysis of solid samples with detection by inductively coupled plasma mass spectrometry (ICP-MS). Into this small sample cuvette, a solid mixture of the biological samples and diammonium hydrogenphosphate powder as a fusion flux was placed and situated on a TBF. Tetramethylammonium hydroxide solution was added to the mixture. After the on-furnace digestion had been completed, the analyte in the cuvette was vaporized and introduced into the ICP mass spectrometer. The solid samples were analyzed by using a calibration curve prepared from the aqueous standard solutions. The detection limit was estimated to be 5.1 pg of lead, which corresponds to 10.2 ng g(-1) of lead in solid samples when a prepared sample amount of 1.0 mg was applied. The relative standard deviation for 8 replicate measurements obtained with 100 pg of lead was calculated to be 6.5%. The analytical results for various biological samples are described.

Animals↗
PubMed1996

Detection of Toxoplasma gondii in feline and canine biological samples by use of the polymerase chain reaction.

OBJECTIVE: To develop a polymerase chain reaction (PCR) technique to identify Toxoplasma gondii DNA in biological samples from cats and dogs. DESIGN: To artificially create samples that would mimic those acquired in a clinical setting from animals with naturally acquired toxoplasmosis. Using these samples, a PCR test to identify T gondii DNA was developed. SAMPLE POPULATION: Feline and canine aqueous humor, CSF, serum, and blood samples. PROCEDURE: Tachyzoites of several strains of T gondii grown in cell culture were added to feline and canine aqueous humor, CSF, serum, and blood samples. Protocols for identifying T gondii DNA by use of the PCR were developed. RESULTS: The DNA from as few as 10 tachyzoites of T gondii could be identified in feline and canine aqueous humor, CSF, and serum samples. One hundred tachyzoites could be identified in blood samples. CONCLUSIONS: Toxoplasma gondii can be identified in feline and canine biological samples by use of the PCR. CLINICAL RELEVANCE: Correlation of clinical disease to T gondii serum antibodies provides only a presumptive diagnosis of toxoplasmosis. Use of PCR to detect T gondii DNA in biological samples from cats and dogs may provide a sensitive tool for the antemortem diagnosis of toxoplasmosis and may be most beneficial when used in conjunction with serum antibody titers.

Animals↗
PubMed1995

Development of an electronically tunable L-band resonator for EPR spectroscopy and imaging of biological samples.

While interest in the EPR spectroscopy and imaging of biological samples triggered the development of numerous resonator and microwave-bridge designs operating at high RF and L-band frequencies, a number of important technical problems remain. Two of these problems are the occurrence of sample-induced drifts in resonance frequency of the resonator and the intrinsic limitations in sensitivity of low-frequency EPR measurements. In an effort to address these problems, an approach was developed for obtaining fixed-frequency EPR measurements, utilizing a low-phase-noise fixed-frequency oscillator as a microwave source and an electronically tunable ceramic re-entrant L-band resonator, which was locked to the oscillator via automatic frequency control (AFC). The tunable microwave resonator described in this paper employs a voltage-controlled piezoelectric actuator for adjustment of the resonance frequency via variations of the equivalent capacitance of the resonator. Design considerations and data, as well as schematic drawings and test results, are presented. The reported approach provides true fixed-frequency operation under a wide range of sample and environment conditions and over prolonged periods of time. The use of this tunable resonator design in conjunction with an ultra-low-noise fixed-frequency oscillator should enable enhanced sensitivity in EPR measurements of large lossy biological samples.

Algorithms↗
PubMed2005

Concentration and characteristics of depleted uranium in water, air and biological samples collected in Serbia and Montenegro.

During the Balkan conflicts, in 1995 and 1999, depleted uranium (DU) rounds were employed and were left in the battlefield. Health concern is related to the risk arising from contamination of the environment with DU penetrators and dust. In order to evaluate the impact of DU on the environment and population in Serbia and Montenegro, radiological surveys of DU in water, air and biological samples were carried out over the period 27 October-5 November 2001. The uranium isotopic concentrations in biological samples collected in Serbia and Montenegro, mainly lichens and barks, were found to be in the range of 0.67-704 Bqkg(-1) for (238)U, 0.48-93.9 Bqkg(-1) for (234)U and 0.02-12.2 Bqkg(-1) for (235)U, showing uranium levels to be higher than in the samples collected at the control sites. Moreover, (236)U was detectable in some of the samples. The isotopic ratios of (234)U/(238)U showed DU to be detectable in many biological samples at all examined sites, especially in Montenegro, indicating widespread ground-surface DU contamination, albeit at very low level. The uranium isotopic concentrations in air obtained from the air filter samples collected in Serbia and Montenegro were found to be in the range of 1.99-42.1 microBqm(-3) for (238)U, 0.96-38.0 microBqm(-3) for (234)U, and 0.05-1.83 microBqm(-3) for (235)U, being in the typical range of natural uranium values. Thus said, most of the air samples are DU positive, this fact agreeing well with the widespread DU contamination detected in the biological samples. The uranium concentrations in water samples collected in Serbia and Montenegro were found to be in the range of 0.40-21.9 mBql(-1) for (238)U, 0.27-28.1 mBql(-1) for (234)U, and 0.01-0.88 mBql(-1) for (235)U, these values being much lower than those in mineral water found in central Italy and below the WHO guideline for drinking water. From a radiotoxicological point of view, at this moment there is no significant radiological risk related to these investigated sites in terms of possible DU contamination of water, air and/or plants.

Air Pollutants, Radioactive↗
PubMed1999

An integrated microfabricated device for dual microdialysis and on-line ESI-ion trap mass spectrometry for analysis of complex biological samples.

A microfabricated dual-microdialysis device in a single integrated microfabricated platform was constructed using laser micromachining techniques for the rapid fractionation and cleanup of complex biological samples. On-line dual microdialysis and ESI-MS of biological samples was demonstrated using an ion trap mass spectrometer. The mass spectra obtained demonstrated the efficiency of dual microdialysis for removing both high-molecular-weight and low-molecular-weight species that interfere with effective ESI-MS analysis of target biopolymers. Signal-to-noise ratios were also greatly improved compared to direct sample infusion. In addition to its compactness, negligible dead volume, and robustness, the device can be used at a flow rate of only 200 nL/min, an order of magnitude lower than that obtained previously. This reduced sample consumption and improved sensitivity with ESI-MS. The results suggest the potential for integration of such microfabricated devices with other sample manipulations for the rapid ESI-MS analysis of complex biological samples.

Escherichia coli↗
PubMed1977

Nickel gas chromatographic columns: an alternative to glass for biological samples.

Nickel tubing may be substituted for glass in the fabrication of gas chromatographic columns for use with samples of biological interest. Comparisons of separations of mixtures of steroids, narcotic alkaloids, phenothiazines, and amphetamines on stainless stell, glass, and nickel packed columns showed little or no observable sample decomposition on glass or nickel as contrasted to complete loss of certain compounds on stainless steel. The nickel columns are easily prepared, durable, economical, and not subject to breakage.

Alkaloids↗
PubMed2002

Generalizing molecular results arising from incomplete biological samples: expected bias and unexpected findings.

PURPOSE: In molecular epidemiology, obtaining biological samples for all subjects targeted for study is frequently hampered by ethical, clinical, and logistic factors. The extent to which the incompleteness of biological samples could cause bias is rarely analyzed in depth. Here we report some expected bias and some unexpected findings during a study on mutations in the K-ras gene in exocrine pancreatic cancer (EPC). METHODS: In this case-case study, all patients registered with EPC between 1980 and 1990 at two general hospitals were retrospectively identified from the hospital tumor registries. Their clinical records were abstracted and paraffin-embedded samples retrieved from pathology records. DNA was amplified, and mutations in codon 12 of the K-ras gene were detected using the artificial RFLP technique. RESULTS: Results on the mutations (RM) were obtained for 51 of the 149 cases of EPC (34.2%). There were no significant differences on the availability of RM by age, gender, and tumor stage at diagnosis, but RM were over five times more likely to be available from one of the hospitals. Subjects with RM were more likely to have received a treatment with curative intent (OR = 11.56, 95% CL: 2.88-46.36). The existence of RM was positively associated with the availability of information on alcohol use and family history of cancer. Subjects with RM tended to belong to higher occupational groups and to smoke less than subjects without RM. Unexpectedly--given that in EPC K-ras mutations have consistently been found unrelated to age, gender, tumor stage, and other clinical factors-, cases with a K-ras mutation were more likely than wild-type cases to have information on tobacco and alcohol use (OR = 3.29, p = .21), medical history (OR = 4.46, p = .41), and family history of cancer (OR = 4.80, p = .01). The relationship between completeness of clinical records and K-ras mutations among cases with RM could not be accounted by age, gender, and occupational group. CONCLUSIONS: Simple tests of age and gender distributions among subjects with and without available clinical information and molecular results may not rule out selection and information bias. Studies using biologic specimens are even more in need than classic studies to explain clearly the process followed to include and exclude subjects. Additional caution is needed when generalizing molecular results arising from incomplete biological specimens.

Aged↗
PubMed1975

The simultaneous determination by selected ion monitoring of the levels of homovanillic, isohomovanillic, 3,4-dihydroxyphenylacetic and 3-methoxy-4-hydroxymandelic acids in single biological samples.

Methods for the determination and quantitation of homovanillic, isohomovanillic, 3,4-dihydroxyphenylacetic and 3-methoxy-4-hydroxymandelic acids in single samples of biological material by gas chromatography, gas chromatographic mass spectrometry and selected ion monitoring during gas chromatographic mass spectrometry are described. Examples of the levels of these four acid metabolites of catecholamines in serum and CSF of untreated dog, cat and human, in urine of parkinsonian subjects treated with varying doses of L-DOPA and in CSF of dogs which had received 1 g of L-DOPA per day for 7 weeks are presented.

3,4-Dihydroxyphenylacetic Acid↗
PubMed2002

Spectral variation of the infrared absorption coefficient in pulsed photothermal profiling of biological samples.

Pulsed photothermal radiometry can be used for non-invasive depth profiling of optically scattering samples, including biological tissues such as human skin. Computational reconstruction of the laser-induced temperature profile from recorded radiometric signals is sensitive to the value of the tissue absorption coefficient in the infrared detection band (muIR). While assumed constant in reported reconstruction algorithms, muIR of human skin varies by two orders of magnitude in the commonly used 3-5 microm detection band. We analyse the problem of selecting the effective absorption coefficient value to be used with such algorithms. In a numerical simulation of photothermal profiling we demonstrate that results can be markedly impaired, unless the reconstruction algorithm is augmented by accounting for spectral variation muIR(lambda). Alternatively, narrowing the detection band to 4.5-5 microm reduces the spectral variation muIR(lambda) to a level that permits the use of the simpler, unaugmented algorithm. Implementation of the latter approach for depth profiling of port wine stain birthmarks in vivo is presented and discussed.

Algorithms↗
PubMed1995

Enzyme immunoassay for quantification of tenascin in biologic samples.

An enzyme immunoassay was developed for quantification of tenascin in biologic samples. An enzyme conjugate prepared by coupling peroxidase to a well-characterized, affinity-purified monoclonal antibody EB2 to human tenascin was used as principal reagent. The assay comprises 96-well microtitration strip plates with immobilized monoclonal antibody DB7 to human tenascin. By using a novel monoclonal antibody suppressing human-anti-mouse-factor, MAK33, in the sample buffer, the specificity of the test could be improved. The method has a minimum detectable sensitivity of 1.5 ng tenascin and permits determination of tenascin in various biologic samples. The coefficients of variation within run and between run ranged from 0.9% to 5.0%. The average tenascin concentration in normal plasma was 0.97 mg/L (n = 200) and in serum 0.73 mg/L (n = 200). The tenascin concentrations were also determined in samples of urine, bile, amniotic fluid, seminal fluid, cerebrospinal fluid, bronchoalveolar lavage (BAL) fluid, and pleural fluid showing general applicability of the assay. The method permits the determination of tenascin in samples of different body fluids from various diseases, including cancer, showing increased amounts of the protein at the tissue level.

Antibodies, Monoclonal↗
PubMed1995

Enzymatic determination of carbon (14C)-labeled glycerol in biological samples.

A method for determination of glycerol-specific-radioactivity in biological samples is presented. It is based on the following steps: (a) enzymatic conversion of glycerol to dihydroxyacetone-phosphate, (b) quantitative trapping of dihydroxyacetone-phosphate in SPE amino (NH2) columns, (c) eluation with HCl 0.5 N of dihydroxyacetone-phosphate followed by radioactivity counting and (d) estimation of the radioactivity thus trapped compared with that of enzymatically untreated aliquots of the same samples. No interferences from other 14C-labeled materials tested such as D-glucose, L-alanine, L-glutamine and D-beta-hydroxybutyrate were observed. This inexpensive and high-speed method can be applied in routine multiple estimations of glycerol-specific-radioactivity in biological samples in tracer metabolic studies.

Animals↗
PubMed1985

Enzymatic lactate-specific radioactivity determination in biological samples.

A method for the measurement of specific lactate radioactivity in biological samples is presented. It is based on the following steps: (a) enzymatic conversion of lactate to pyruvate, (b) pyruvate conversion to 2,4-dinitrophenylhydrazone, (c) concentration-separation of the latter in reusable Amberlite XAD-7 polymeric adsorbent columns, and finally (d) estimation of the radioactivity thus retained compared with that of enzymatically untreated aliquots of the same samples. Specificity was ensured by the use of lactate dehydrogenase as specific recognizing agent for lactic acid. No interference from glucose, lactate, or amino acids was observed. The method presented is simple and can be applied in routine multiple estimations of lactic acid radioactivity in conjunction with the enzymatic measurement of lactate in biological samples in tracer metabolic studies.

Animals↗
PubMed1994

Quantitative analysis of 2-oxoglutarate in biological samples using liquid chromatography with electrochemical detection.

This paper describes a technique for quantitative analysis of 2-oxoglutarate (alpha-ketoglutarate) in biological samples using liquid chromatography with electrochemical detection (LC-EC). This method utilizes a simplified, efficient sample preparation designed to select for 2-oxoglutarate and similar compounds by derivatization with phenylhydrazine. The response was linear over the range from 62.5 to 1000 ng/ml. The least quantifiable concentration was 62.5 ng/ml and the least detectable concentration was 25 ng/ml. To test the ability of the assay to measure 2-oxoglutarate in biological samples, this method was used to quantitate the 2-oxoglutarate content in chick osteoblast cultures and to determine the ability of the assay to accurately measure a standard addition of 500 ng/ml 2-oxoglutarate when added to a sample of the forementioned groups. The 2-oxoglutarate content of these cells was 6.67 +/- 1.20 ng/micrograms DNA or 105 +/- 18 ng/cell layer (mean +/- 95% confidence interval) and the assay accurately measured the standard addition. This method was also used to quantitate 2-oxoglutarate content in whole embryonic chick calvariae containing 6.40 +/- 0.95 ng/mg dry bone weight or 37.5 +/- 5.5 ng/bone. This assay provides significantly lower detection limits than the currently available procedures and is suitable for determination of 2-oxoglutarate in biological samples where very low amounts of 2-oxoglutarate are found. This method is the first application of LC-EC for quantitating 2-oxoglutarate.

Animals↗
PubMed1988

Sensitive determination of D-lactic acid in biological samples by high-performance liquid chromatography.

D-Lactate in biological samples was converted into the hydrazone of pyruvate in the presence of D-lactate dehydrogenase, an NADH-reoxidation system using diaphorase, DL-6,8-thioctamide and hydrazine. The hydrazone was converted into 2-methylquinoxanol by o-phenylenediamine in hydrochloric acid, and then the quinoxanol was determined by high-performance liquid chromatography with fluorescence detection. The calibration curve of D-lactate was linear up to at least 60 nmol/ml, and the determination limit was 600 fmol. Using this method, D-lactate was determined in biological samples.

Adult↗