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PubMed1995

New solid-phase extraction for an improved high-performance liquid chromatographic procedure for the quantitation of halofantrine and monodesbutylhalofantrine in blood or plasma.

A rapid, accurate, and sensitive high-performance liquid chromatographic (HPLC) method, with fluorimetric detection, for the simultaneous measurement of halofantrine and desbutylhalofantrine in human plasma or whole blood is described. Sample preparation involved protein precipitation, followed by an efficient solid-phase extraction on a C8 cartridge. Analytes were isolated from 1 ml of the biological fluids and recovered by a 2% acetic acid in ethyl acetate solution. Chromatographic separation was carried out on a LiChrospher 60 RP select B, C8 bonded phase (5 microns particle size, 25 cm x 4 mm I.D.) using a mobile phase of water-acetonitrile (35:65, v/v) containing triethylamine (1%) and adjusted to pH 4 with orthophosphoric acid. The total run time was 14 min. Relative standard deviations of the intra-and inter-assay precisions were less than 5.9%. Assumption of linearity was investigated by studying the y-residuals and by ANOVA (analysis of variance). Because of the wide range of calibration (0.1 to 2.0 microgram/ml) variances were non-homogeneous (Hartley's test) and the weighted regression line was computed in order to allow pharmacokinetic studies. Accuracy was tested using a t-statistic. Limits of decision, detection and quantification were realized from an analysis of the blanks. Application of the method to clinical specimens was demonstrated.

Antimalarials↗
PubMed1997

Sampling intercomparisons for aldehydes in simulated workplace air.

Thirty one laboratories of various EU Member States have participated in two interlaboratory comparisons in order to assess errors of personal sampling methods associated with both the sampling and the analytical steps. In contrast to conventional quality control schemes, this project particularly focuses attention on the sampling and identification step; it is executed by means of sampling exercises and has included discussions on potential sources of error. In a sampling exercise, participants come to a central facility and perform measurements on synthetic workplace air in a laboratory installation. Concentration levels of formaldehyde, acrolein, glutaraldehyde and acetaldehyde between 0.1 and 2 times the limit value for workplace air were prepared at various humidity levels and with acetone, occasionally, as interferent. Sampling times varied from 1-4 h. The related analytical work is performed at the analyst's own laboratory. The intention is for each participant to determine the observed value of the delivered standard atmosphere using the sampling method of his own choice. Trueness (bias), precision and relative overall uncertainty of each method-laboratory combination is calculated and verified towards compliance with EN 482, which outlines minimum performance criteria. The first challenge involved the precise gas phase generation of the selected analytes in high air flows (up to 300 1 min-1) and calculating the true value only by direct reference to primary standards. This was accomplished by modifying the capillary dosage injection technique so that reactive compounds, like low molecular mass aldehydes, could be dosed with the same accuracy and precision as unreactive solvents. A permeation tube with high emission rate was developed for formaldehyde. Up to ten different sampling techniques were evaluated. The measurement methods used by the majority of the participants were based on pumped sampling on silica cartridges (or tubes) and glass fiber filters, coated with 2,4-dinitrophenylhydrazine. It was observed that for formaldehyde, and in some cases for acetaldehyde and glutaraldehyde, the majority of the method-laboratory combinations complied with an overall uncertainty of 30%. The results for acrolein, however, indicated a systematic negative bias, often larger than minus 50% of the true value, caused by the decomposition of the acrolein DNPH derivative in the presence of excess acid and excess DNPH.

Air Pollutants, Occupational↗
PubMed2002

Electrochemical bioassay for the investigation of chlorpyrifos-methyl in vine samples.

This paper reports the optimization of an electrochemical bioassay for the determination of chlorpyrifos-methyl and its application to the analysis of grape and vine leaf samples treated with that pesticide. The analytical method was based on electrochemical determination of the extent of the inhibition exerted by the pesticide on acetylcholinesterase using the substrate acetylthiocholine. Two similar calibration plots were obtained, in the range of 1-300 ng/mL, respectively, for chlorpyrifos-methyl in pure standard form and in the commercial preparation Reldan, with comparable coefficients of variation (CV) in the range of 10% < CV < 20%. After an insecticide treatment, samples were analyzed to evaluate its persistence both in grapes and in vine leaves. Samples were evaluated using different extraction procedures: one based on solvent extraction of pesticide residue from grapes and the other based on aqueous extraction from vine leaves using phosphate buffer. The grape solvent extracts were analyzed using both gas chromatography and electrochemical bioassay, whereas the vine leaf buffer extracts were analyzed using the electrochemical bioassay. Quantitative analysis of chlorpyrifos-methyl determined in the two samples, with the electrochemical bioassay, showed a comparable decrease profile over the experimental period.

Acetylcholinesterase↗
PubMed2004

Chemical screening by mass spectrometry to identify inhibitors of anthrax lethal factor.

Mass spectrometry (MS) analysis is applicable to a broad range of biological analytes and has the important advantage that it does not require analytes to be labeled. A drawback of MS methods, however, is the need for chromatographic steps to prepare the analyte, precluding MS from being used in chemical screening and rapid analysis. Here, we report that surfaces that are chemically tailored for characterization by matrix-assisted laser-desorption ionization time-of-flight MS eliminate the need for sample processing and make this technique adaptable to parallel screening experiments. The tailored substrates are based on self-assembled monolayers that present ligands that interact with target proteins and enzymes. We apply this method to screen a chemical library against protease activity of anthrax lethal factor, and report a compound that inhibits lethal factor activity with a K(i) of 1.1 microM and blocks the cleavage of MEK1 in 293 cells.

Antigens, Bacterial↗
PubMed2005

Analytical performance and workflow evaluation of the Roche E170 modular immunoassay analyzer in a pediatric setting.

OBJECTIVES: To evaluate the analytical performance of the Roche E170 modular analytics immunoassay analyzer and assess its impact on workflow efficiency and ability to consolidate workstations in a pediatric setting. DESIGN AND METHODS: Analytical performance of eleven common immunoassays was assessed. Total imprecision was determined using Roche PreciControl Universal controls, Bio-Rad Lyphochek Immuno Plus, Anemia controls, and a human serum pool. Method comparison was performed with approximately 100 patient specimens. High dose hook effect, sample carryover, and results comparison between the two measuring channels were evaluated. For the workflow study, the time required for sample and reagent handling, instrument preparation, and hands-on time were assessed. RESULTS: Correlation coefficients with existing methods ranged from 0.941 to 0.999. Biases of -19% to 70% were observed. Total imprecision ranged from 1.1 to 7.6%. No sample carryovers were encountered. Results from both measuring channels were comparable. CONCLUSION: E170 is suitable for use in a pediatric setting. The analytical performance is acceptable and gives equivalent results to our existing systems. The precision is comparable and acceptable. Some improvement in efficiency, workflow, cost saving, and consolidation of workstations is possible. Significant workflow improvements can only be realized when integrated with the chemistry modules.

Autoanalysis↗
PubMed2005

Rapid measurement of plasma acylcarnitines by liquid chromatography-tandem mass spectrometry without derivatization.

BACKGROUND: Tandem mass spectrometry (MS/MS) is being increasingly used to identify and measure acylcarnitines in blood and urine of children suspected of having fatty oxidation disorders and other inborn errors of metabolism. Rapid MS/MS analysis requires simple and efficient sample preparation. We developed a LC-MS/MS method for the online extraction of acylcarnitines in plasma without derivatization that requires only precipitation of proteins by acetonitrile followed by centrifugation, thus increasing efficiency. METHODS: An API-3000 tandem mass spectrometer (SCIEX, Toronto, Canada) equipped with electrospray ionization (ESI), TurboIon Spray source, three Shimadzu LC10AD micropumps and autosampler (Shimadzu Scientific Instruments, Columbia, MD) was used to perform the analysis. Within-day and between-day imprecision was evaluated for 10 analytes in the MRM mode using 3 levels of controls. Accuracy was determined by comparing the method with another MS/MS procedure and by recovery experiments. Sensitivity and specificity were evaluated by identifying patient samples under a wide variety of clinical conditions. RESULTS: Within-day CVs was <10% for all analytes tested and between-day CVs ranged from 4.4% to 14.2%. The method was linear in the range between 1.0 and 100 micromol/l for C2 and 0.1 and 10 micromol/l for the other acylcarnitines. The results of the comparison study yielded r values ranging between 0.948 and 0.999. Recovery ranged from 84% to 112%. The method correctly identified patients with a variety of fatty acid oxidation disorders and organic acidemias. CONCLUSIONS: Our method is a simple procedure for the analysis of acylcarnitines in plasma with minimal sample preparation. It is thus ideal in a routine clinical setting where efficient processing of clinical samples is necessary to reduce turnaround time under conditions of high-throughput.

Acylation↗
PubMed1997

Solid-phase extraction of phospholipids from hemoglobin solutions using Empore styrene-divinylbenzene disks.

Styrene-divinylbenzene Empore disks were investigated for the extraction of phospholipids from red blood cells or aqueous solutions of hemoglobin as a means to reduce the time and solvent use required in sample preparation. Red blood cells are the source for hemoglobin used in the preparation of a hemoglobin-based oxygen carrier which is being developed to replace blood in transfusion therapy. Phospholipids are a major component of the membrane of red blood cells, and are toxic when administered directly into the vasculature. Sensitive analytical methods are required to detect phospholipids to ensure that concentrations in purified hemoglobin are well below toxic levels. This requires isolation from large volumes of purified hemoglobin solutions. The method described utilizes Empore disks to extract phospholipids from 30 ml of stroma free Hb preparations. Phosphatidylethanolamine, phosphatidylinositol, phosphatidylcholine and sphingomyelin were recovered with an average of 92% yield. The recovery of phosphatidylserine was 65%. The use of solvent and time required for sample preparation were reduced by an average of 80% relative to liquid-liquid extraction. The capacity of the 47-mm disk for the total of five phospholipids exceeds 0.3 mg. The method has been used for quantitation of phospholipids in red blood cells and stroma free hemoglobin solutions.

Chromatography, High Pressure Liquid↗
PubMed2003

Multi-element trace determinations in pure alkaline earth fluoride powders by high-resolution ICP-MS using wet-chemical sample preparation and laser ablation.

Four alternative analytical procedures for the determination of ten important trace impurities (Mg, Cr, Fe, Cu, Zn, Sr, Zr, Cd, Ba, and Pb) in pure alkaline earth fluoride powders were applied using high-resolution inductively coupled plasma mass spectrometry (ICP-MS). Two procedures are based on a wet-chemical microwave digestion with boric acid and quantification by the standard addition technique and isotope dilution mass spectrometry (IDMS), respectively. In addition, analyses are also performed by laser ablation as a direct solid sampling technique applying matrix-matched external calibration as well as isotope dilution of the powdered sample. For most elements good agreement between the different methods is found. Detection limits for laser ablation vary between 0.05 ng g(-1) for Zr and 20 ng g(-1) for Mg. They are about one to two orders of magnitude lower than those of the wet-chemical procedures, which is mainly due to the high dilution factor during the sample preparation step. Advantages and restrictions of the different analytical procedures are discussed with respect to their routine applicability. Due to its relatively high accuracy, low detection limits, and time-efficiency LA-ICP-IDMS is the preferred choice if no standard reference materials are available.

Journal Article↗
PubMed1996

Microchip-based Devices for Molecular Diagnosis of Genetic Diseases.

Microchips, constructed with a variety of microfabrication technologies (photolithography, micropatterning, microjet printing, light-directed chemical synthesis, laser stereochemical etching, and microcontact printing) are being applied to molecular biology. The new microchip-based analytical devices promise to solve the analytical problems faced by many molecular biologists (eg, contamination, low throughput, and high cost). They may revolutionize molecular biology and its application in clinical medicine, forensic science, and environmental monitoring. A typical biochemical analysis involves three main steps: (1) sample preparation, (2) biochemical reaction, and (3) detection (either separation or hybridization may be involved) accompanied by data acquisition and interpretation. The construction of a miniturized analyzer will therefore necessarily entail the miniaturization and integration of all three of these processes. The literature related to the miniaturization of these three processes indicates that the greatest emphasis so far is on the investigation and development of methods for the detection of nucleic acid, followed by the optimization of a biochemical reaction, such as the polymerase chain reaction. The first step involving sample preparation has received little attention. In this review the state of the art of, microchip-based, miniaturized analytical processes (eg, sample preparation, biochemical reaction, and detection of products) are outlined and the applications of microchip-based devices in the molecular diagnosis of genetic diseases are discussed.

Journal Article↗
PubMed2005

Determination of tannin in green tea infusion by flow-injection analysis based on quenching the fluorescence of 3-aminophthalate.

A flow-injection analytical system was developed to determine tannin content in green tea infusions. The flow-injection system is based on measuring the quenching effect of tannin on the fluorescence of 3-aminophthalate. Fluorophore was obtained by auto-oxidation of luminol during solution preparation. System performance was satisfactory for routine analysis (sample throughput >20 h(-1); linear dynamic range for tannic acid, 0.005-0.3 mg/mL; linear dynamic range for green tea tannin, 0.02-1.0 mg/mL; CV < 3%). The flow-injection method is immune from interference by coexisting ascorbate in green tea infusion. Analytical results were verified by the ferrous tartrate method, the Japanese official analytical method.

Ascorbic Acid↗
PubMedDate not supplied

Precision and accuracy of asbestos fiber counting by phase contrast microscopy.

A new method was developed to prepare permanent asbestos slides with relocatable fields of view by imprinting these fields directly on the cleared wedges of filters containing asbestos fibers. The image quality of the fibers is comparable with that of the fibers prepared by the acetone/triacetin and the dimethyl formamide/Euparal method. The slides are suitable for evaluating the intercounter precision and accuracy of fiber counts by phase contrast microscopy. Seventeen chrysotile and 16 amosite slides, prepared from American Industrial Hygiene Association/National Institute for Occupational Safety and Health Proficiency Analytical Testing program samples, were evaluated by 58 analysts of 38 government and private laboratories in Canada. By asking the analysts to examine the same fields of view of the slides, the present study found that when examined at 400x, an average of 59.2 fiber counting errors were made for every 100 chrysotile fibers reported and 24.4 errors for 100 amosite fibers reported. The chrysotile fibers were underestimated by 25.0%, but there was no bias in counting amosite fibers. Remedial steps have been proposed to control the major source of errors, which is the subjective ability of the analyst to observe and size fibers. The slides may also be used to harmonize various phase contrast optical microscopy methods and proficiency testing programs.

Asbestos, Amosite↗
PubMed2004

Separation and determination of two sesquiterpene lactones in Radix inulae and Liuwei Anxian San by microemulsion electrokinetic chromatography.

A novel microemulsion electrokinetic chromatography (MEEKC) method for separating and determining two sesquoterpene lactones, alantolactone (AL) and isoalantolactone (IAL), in Radix inulae and Liuwei Anxian San has been developed. The effects of several important factors such as internal organic phases, concentration of microemulsion, concentration of acetonitrile, injection time and running voltage were systematically investigated to determine the optimum conditions. The optimum microemulsion system was composed of n-hexane (0.32% w/w), SDS (1.24% w/w), 1-butanol (2.64% w/w), acetonitrile (10% w/w) and 10 mm sodium tetraborate buffer (85.80% w/w, pH 9.2). The applied voltage was 20 kV. The analytes were detected at 214 nm. Regression equations revealed linear relationships (correlation coefficients 0.9950 for AL and 0.9946 for IAL) between the peak area of each analyte and the concentration. The limits of detection (defined as a signal-to-noise ratio of about 3) were approximately 0.45 microg/mL for AL and 0.56 microg/mL for IAL. The levels of the analytes were successfully determined with recoveries ranging from 98.2 to 104.3%. Furthermore, a simple and effective extraction method, with methanol in an ultrasonic water bath for 60 min, was used for sample preparing. Also, MEEKC was compared with micellar electrokinetic chromatography (MEKC) and shown better separation results.

Acetonitriles↗
PubMed2004

Capillary electrophoresis analysis of fosfomycin in biological fluids for clinical pharmacokinetic studies.

A feasible capillary zone electrophoresis (CZE) method with indirect UV and contactless conductivity detection was developed for the determination of fosfomycin, an antibiotic, in human plasma and microdialysis samples. Samples were collected from test persons during a clinical trial. The background electrolytes used consisted of 25 mM benzoic acid and 0.5 mM hexadecyltrimethylammonium bromide, adjusted with tris(hydroxymethyl)aminomethane solution to pH 6.95 for plasma, and to pH 8.05 for microdialysis samples. CZE separations of the anionic analyte were carried out with reversed electroosmotic flow directed towards the anode. The limit of detection was between 0.6 and 2 microg/mL, depending on the matrix and the detection method. No sample preparation was needed for microdialysis samples; for plasma samples, proteins were precipitated with methanol (1+2, v+v), and the supernatant was analyzed. The yield determined with spiked samples was about 100%, the reproducibility of the entire method, expressed by the RSD% of three independent determinations of fosfomycin in triplicate after spiking Ringer's solutions and plasma samples, respectively, was better than 8%. The method is thus well-suited for clinical studies for the determination of the antibiotic in biological fluids.

Benzoic Acid↗
PubMed2002

Speciation of mercury, tin, and lead compounds by gas chromatography with microwave-induced plasma and atomic-emission detection (GC-MIP-AED).

Because of their high toxicity and widespread distribution, the reliable selective quantification of alkyl and aryl species containing mercury, tin, or lead has been one of the goals of speciation analysis in recent years. Since becoming commercially available, GC-MIP-AED has been one of the most-used tools in this work. In this paper, the value and limitations of GC-MIP-AED for the speciation of Hg, Sn, and Pb compounds in environmental samples are reviewed and compared with the analytical characteristics of other hyphenated GC-based techniques. Because quantification of Hg, Sn, and Pb species by GC techniques normally requires complex sample preparation involving several steps, the effect of sample-preparation methods on the accuracy and precision of the results is discussed. Finally, we describe the current status of a rapid, low-cost GC-MIP-PED system specifically designed for routine quantification of Hg, Sn, and Pb species in environmental control laboratories.

Journal Article↗
PubMed2002

A highly sensitive spectrophotometric determination of platinum(IV) using leuco xylene cyanol FF.

A new, simple, highly sensitive and rapid spectrophotometric method has been described for the determination of platinum(IV). The method is based on the oxidation of leuco xylene cyanol FF (LXCFF) to its blue form of xylene cyanol FF by platinum(IV) in sulfuric acid medium (pH 1.0-2.5), the formed dye shows an absorption maximum at 620 nm in acetate buffer medium (pH 3.0-4.5). The method obeys Beer's law over a concentration range of 0.3 to 2.6 micro g mL(-1) platinum, having molar absorptivity and Sandell's sensitivity of 5.1x10(4) L mol(-)(1) cm(-1) and 0.0038 micro g cm(-2), respectively. The optimum reaction conditions and other analytical parameters have been evaluated. The developed method has been successfully applied to the determination of platinum in pharmaceutical preparations, soil, natural water, plant material, platinum-containing catalyst, and synthetic alloy samples.

Journal Article↗
PubMed1992

Measurement of dothiepin and its major metabolites in plasma by high-performance liquid chromatography.

This paper describes a reversed-phase high-performance liquid chromatographic method which will simultaneously measure dothiepin and its three major metabolites (northiaden, northiaden-S-oxide and dothiepin-S-oxide) in plasma using trimipramine as internal standard. Sample preparation involved a basic extraction using diethyl ether followed by an acid back-extraction. The method we report is linear over the range 50-1000 ng/ml (r = 0.999), for all analytes. Total imprecision is less than 11% (coefficient of variation) and accuracy is greater than 94% (n = 20). Recovery of analytes varied considerably from 51.7% for northiaden-S-oxide to 90.2% for dothiepin-S-oxide.

Chromatography, High Pressure Liquid↗
PubMed2003

Closed-loop stripping analysis of synthetic musk compounds from fish tissues with measurement by gas chromatography-mass spectrometry with selected-ion monitoring.

Synthetic musk compounds have been found in surface water, fish tissues, and human breast milk. Current techniques for separating these compounds from fish tissues require tedious sample clean-up procedures. A simple method for the determination of synthetic musk compounds in fish tissues has been developed. Closed-loop stripping of saponified fish tissues in a 1-1 Wheaton purge-and-trap vessel is used to strip compounds with high vapor pressures such as synthetic musks from the matrix onto a solid sorbent (Abselut Nexus). This technique is useful for screening biological tissues that contain lipids for musk compounds. Analytes are desorbed from the sorbent trap sequentially with polar and nonpolar solvents, concentrated, and directly analyzed by high resolution gas chromatography coupled to a mass spectrometer operating in the selected ion monitoring mode. In this paper, we analyzed two homogenized samples of whole fish tissues with spiked synthetic musk compounds using closed-loop stripping analysis and pressurized liquid extraction (PLE). The analytes were not recovered quantitatively but the extraction yield was sufficiently reproducible for at least semi-quantitative purposes (screening). The method was less expensive to implement and required significantly less sample preparation than the PLE technique.

Animals↗
PubMed1999

On-line combination of aqueous-sample preparation and capillary gas chromatography.

Methods currently in use to combine the preparation of aqueous samples on-line with capillary gas chromatography (GC) comprise heartcut-orientated reversed-phase liquid chromatography-GC and analyte-isolation-orientated analyte extraction-GC. These approaches either use techniques in which water is directly introduced onto the GC column, or an indirect approach in which water is eliminated, i.e., by solid-phase extraction, solid-phase microextraction or liquid-liquid extraction, prior to introduction of the analytes onto the GC column. The latter type of approach is much more successful and user-friendly, and many applications have been reported.

Chromatography, Gas↗