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PubMed2004

Determination of residual styrene monomer in polystyrene granules by gas chromatography-mass spectrometry.

Polystyrene is widely used in formulations intended for children use. The main problem with this plastic is the residual styrene, which can migrate from the product, and therefore, be in contact with children. The acute toxicity of styrene is well known, raising the need of an efficient and fast method of analysis for this compound. Several extraction methods have been evaluated and compared for the determination of residual styrene monomer in polystyrene granules used in toys: supercritical fluid extraction (SFE), microwave-assisted extraction (MAE), Soxhlet extraction, headspace emission and dissolution-precipitation. The analyte was subsequently detected by gas chromatography (GC) with MS detection. The results indicated that the most efficient method was dissolution-precipitation giving even higher extraction efficiency than SFE. For validating the method, PS samples spiked with known quantities of styrene at three concentration levels were prepared to calculate the extraction recovery. The founded validation data proved the suitability of the proposed method.

Gas Chromatography-Mass Spectrometry↗
PubMed2006

Application of a semi-automated 96-well format solid-phase extraction, column-switching, fluorescence detection protocol for the determination of alendronate in human urine samples obtained from a bioequivalence study.

In the current study, a semi-automated, 96-well format, solid-phase extraction (SPE), analytical column-switching method for alendronate determination in human urine is developed, validated and applied to a bioequivalence study. The current protocol was a substantial improvement of an existing classical method. A robotic liquid handling system was employed to simplify and reduce the time of sample preparation procedure. Automated SPE was carried out using a 96-well cartridge plate and a vacuum control system. Urine samples were determined by applying a column-switching protocol with fluorescence detection. Analysis time, due to the column-switching procedure, was about half of the conventional LC approach (11.5 min instead of 21 min). The method application required the determination of alendronate in urine samples obtained from 96 healthy volunteers as part of a bioequivalence study of two 70 mg alendronate sodium tablets. All major pharmacokinetic parameters of the bioequivalence study were estimated and reported.

Alendronate↗
PubMed2003

Detection of fumonisin B1: comparison of flow-injection liposome immunoanalysis with high-performance liquid chromatography.

Fumonisins are secondary metabolites of the fungus Fusarium moniliforme, a common mycotoxin in corn, which are known to cause cancer in a number of experimental animals and have been linked to human esophageal cancer in China and South Africa. A high-performance liquid chromatographic (HPLC) method is currently the most widely used method for the quantitative determination of fumonisins. This method utilizes precolumn derivatization with o-phthaldialdehyde, isocratic elution, and fluorescence detection. In this study, the HPLC method was chosen as the reference method to evaluate the reproducibility and accuracy of FILIA (flow-injection liposome immunoanalysis) for the detection of the fumonisin B1 (FmB1). Studies indicate that a recovery of 86-90% could be obtained when commercial yellow cornmeal spiked with FmB1 was extracted in 75% methanol, which correlated favorably (correlation coefficient, r(2)=0.945) with the result of 80-92% obtained using the flow-injection liposome immunoanalysis (FILIA) system. The data suggest that the FILIA method is comparable to HPLC for the detection of fumonisins in corn, animal feeds, and human foods. Important features of FILIA as compared to HPLC are, most importantly, lower detection limit (ca. 25 x lower), and also less complex and faster sample preparation and therefore increased analytical throughput. In addition, 24 human corn-based foods and 6 animal feeds were examined for the presence of FmB1 using HPLC and FILIA.

Chromatography, High Pressure Liquid↗
PubMed1990

Novel assay method for mitoxantrone in plasma, and its application in cancer patients.

Mitoxantrone is an anthracene derivative that acts as a cytostatic in a variety of cancers. A quantitative analytical method has been established for the determination of mitoxantrone in plasma. The method employed C18 reversed-phase ion-pair chromatography with an isocratic mobile phase of 50.0% methanol in 10 mM phosphate buffer (pH 3.0) plus 0.09% 1-pentanesulphonic acid and ultraviolet detection. Sample preparation consisted of two extraction steps using same organic solvent system at different pH to remove plasma impurities efficiently. Potential adsorption of mitoxantrone onto glassware was considered. Silanization of all glassware with 5% dichlorodimethylsilane in chloroform increased the extraction recovery in plasma from 50 to 85% with high reproducibility. Mitoxantrone was unstable in human plasma. To maintain plasma sample integrity, each millilitre of sample should be fortified with 0.1 ml of 5% vitamin C (in citrate buffer) and kept frozen until analysis. Using this new method, the calibration curve of mitoxantrone in plasma in the range of interest (1-500 ng/ml) showed good linearity (r = 0.996) and precision (both between-day and within-day coefficients of variation less than 10%). The lower detection limit of this assay method was 1 ng. The application of this method allowed us to study the stability of mitoxantrone in plasma, and the pharmacokinetics of mitoxantrone in nasopharyngeal carcinoma patients receiving 12 mg/m2. The study revealed a prolonged terminal phase half-life for mitoxantrone.

Adsorption↗
PubMed2004

Molecule by molecule direct and quantitative counting of antibody-protein complexes in solution.

We have used two-color fluorescence coincidence detection to directly count individual protein-antibody complexes of protein G or herpes simplex virus labeled with one or more red- and blue-excited antibodies. This allowed quantitative measurement of the concentration of the protein-antibody complexes over 3 orders of magnitude down to the femtomolar level. Single molecule measurements in diluted serum are also possible. The sample preparation is simple, takes place in solution, and requires no separation. Both the antibody affinity and complex dissociation rate are important in determining the sensitivity of the method. At present, the sensitivity limit of 50 fM is determined by the encounter rate of the labeled analyte with the probe volume. This method can be used to detect and quantitate proteins and to measure the stoichiometry, equilibrium constant, and dissociation rate of protein-protein complexes at low concentrations.

Animals↗
PubMed2005

Long-range magnetic ordering in iron jarosites prepared by redox-based hydrothermal methods.

The iron jarosites, plumbojarosite, Pb0.5Fe3(OH)6(SO4)2, argentojarosite, AgFe3(OH)6(SO4)2, and thallium jarosite, TlFe3(OH)6(SO4)2, along with the selenate-capped jarosite analogues of potassium, KFe3(OH)6(SeO4)2, and rubidium, RbFe3(OH)6(SeO4)2, have been prepared in their analytically pure forms by employing redox-based hydrothermal methods. The crystal structures of these materials have been determined, and all are found to be essentially isostructrual including Pb0.5Fe3(OH)6(SO4)2, which is distinct from the structure reported for naturally mined samples. All iron jarosites show long-range order (LRO), signified by a sharp transition temperature, T(N), which falls in the narrow temperature range of 61.4 +/- 5 K. The mechanism responsible for this ordering has been established by examining magnetostructural correlations for the jarosites possessing various interlayer cation and capping groups. We show that all magnetic properties of jarosites, including LRO, find their origin in the basic magnetic unit, the intralayer Fe3(mu-OH)3 triangle. Field-dependent magnetization experiments are consistent with the antiferromagnetic stacking of an out of plane moment developed from spin canting within Fe3(mu-OH)3 triangles. Together with the previously reported AFe3(OH)6(SO4)2 (A = Na+, K+, Rb+ and NH4+) jarosites, these compounds provide a framework for probing magnetic ordering in a spin frustrated lattice of the largest series of isoelectronic and isostructural kagomé systems yet discovered.

Ferric Compounds↗
PubMed2000

Determination of 4-nonylphenol and 4-octylphenol in human blood samples by high-performance liquid chromatography with multi-electrode electrochemical coulometric-array detection.

Alkylphenols can affect human health because they disrupt the endocrine system. In this study, an analytical method for determining trace amounts of 4-nonylphenol (NP) and 4-octylphenol (OP) in human blood samples was developed. Reversed-phase HPLC with multi-electrode electrochemical coulometric-array detection was used for the determination of NP and OP in plasma and serum samples prepared with a solid-phase extraction method. The separation was achieved using an isocratic mobile phase of 0.7% phosphoric acid-acetonitrile with a C18 reversed phase column. The detection limits of NP and OP were 1.0 and 0.5 ng ml-1, respectively. The recoveries of NP and OP added to human plasma samples were above 70.0% with a relative standard deviation of less than 15.5%. The method was found to be applicable to the determination of NP and OP in various human blood samples such as serum and plasma.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

Problems of official methods and new techniques for analysis of foods and feeds for vitamin A.

The official methods for measuring vitamin A and carotene in foods and feeds have evolved over several decades. New procedures are needed to permit the use of modern analytical techniques such as liquid chromatography (LC), and to expand the application of official methods to a wide variety of foods. Recent improvements in methodology are reviewed. Emphasis is often placed on the separations achieved by LC, but all factors that affect quantitation, such as sampling, preparation of standards, and protection of the vitamin against oxidation, must be considered in the development of new methods. Many approaches to analysis are possible because of the versatility of LC but not all of them can be used in official methods. Methods for milk, margarine, and feeds can be prepared now for collaborative assays. Although straightforward methods can be developed for the measurement of beta-carotene in certain foods, a full examination of provitamins will probably continue to require a flexible approach and special expertise.

Animal Feed↗
PubMed2005

Quantification of phenolic metabolites of environmental chemicals in human urine using gas chromatography-tandem mass spectrometry and isotope dilution quantification.

We have developed a method to measure 12 urinary phenolic metabolites of pesticides or related chemicals. The target chemicals for our method are 2-isopropoxyphenol; 2,4-dichlorophenol; 2,5-dichlorophenol; carbofuranphenol; 2,4,5-trichlorophenol; 2,4,6-trichlorophenol; 3,5,6-trichloro-2-pyridinol; para-nitrophenol, ortho-phenylphenol, pentachlorophenol, 1-naphthol and 2-naphthol. The sample preparation involves enzyme hydrolysis, isolation of the target chemicals using solid phase extraction cartridges, a phase-transfer catalyzed derivatization, cleanup using sorbent-immobilized liquid/liquid extraction cartridges, and concentration of the sample. Derivatized samples are analyzed by capillary gas chromatography-tandem mass spectroscopy using isotope dilution calibration for quantification. The limits of detection are in the mid ng/L range and the average coefficient of variation was below 15% for most of the analytes. Using our method, we measured concentrations of the target chemicals in urine samples from the general population.

Calibration↗
PubMed2001

Signal amplification using "spot-on-a-chip" technology for the identification of proteins via MALDI-TOF MS.

The presented "spot-on-a-chip" technology enables easy enrichment of samples in the low nanomolar (1-5 nM) range and provides a fast and reliable automated sample preparation method for performing matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis with high sensitivity and throughput. Through microdispensing, which allows accurate deposition of 60-pL droplets, dilute samples were enriched by making multiple droplet depositions in nanovials. The sample was confined to a defined spot area (300 x 300 microm), and multiple depositions increase the surface density of analyte in the nanovial, thereby providing detection of low attomole levels. The impact of the nanovial geometry with respect to the MALDI-TOF MS resolution for peptides deposited in the microfabricated silicon vials was investigated and the optimal geometry and size were determined. The spot-on-a-chip technology, that is, the combination of microdispensing, micromachined silicon nanovials and on-spot enrichment provides a signal amplification of at least 10-50 times as compared to an ordinary sample preparation. The linearity of the enrichment effect is shown by the analysis of a peptide mixture at the 5 nM level. The signal amplification provided by the spot-on-a-chip enrichment is demonstrated by the analysis of relevant biological samples, interleukin-8 from a spiked cell supernatant, and by successful protein identification of an excised spot from a high-sensitivity silver-stained two-dimensional electrophoresis gel separation.

Adrenocorticotropic Hormone↗
PubMedDate not supplied

Determination of coenzyme Q10 in over-the-counter dietary supplements by high-performance liquid chromatography with coulometric detection.

A rapid and sensitive method is described for the determination of coenzyme Q10 (Q10) in over-the-counter dietary supplements by automated high-performance liquid chromatography (HPLC) with coulometric detection. Sample solutions of powder-filled capsules, oil-based softgels, and tablets were prepared by serial dilution with 1-propanol. After dilution, a known volume of sample solution containing Q10 and the internal standard, coenzyme Q9 (Q9), was directly injected into the HPLC system. Most of electrochemically active compounds in the injection were oxidized at the precolumn conditioning cell and postcolumn guard cell. Q9 and Q10 were monitored at an analytical cell that contained 2 coulometric electrodes, where Q9 and Q10 were reduced to the corresponding ubiquinol-9 and -10 and then oxidized to produce currents. This method produced a linear detector response for peak height measurements over the concentration range of 0.05-8 microg/mL (r > 0.999). The lower limit of detection was 5 ng/mL (signal-to-noise ratio, > or =3). The mean recovery was 98.9 +/- 0.6%; coefficients of variation for intra- and interday precisions were 1.8-4.0%. The proposed method was successfully applied to the determination of Q10 in marketed products.

1-Propanol↗
PubMed2005

Analytical method development and validation of mianserin hydrochloride and its metabolite in human plasma by LC-MS.

Mianserin is a tetracyclic antidepressant drug and administered as racemate of R (-) and S (+) mianserin hydrochloride in a dose of 30-90 mg/day in divided doses. Liquid chromatography-mass spectroscopy (LC-MS) is a tool, which is widely used for determination of drug and their metabolites in biological fluids because of its high sensitivity and precision. Here we describe a liquid chromatography mass spectroscopy method for simultaneous determination of mianserin and its metabolite, N-desmethylmianserin, from human plasma using a liquid-liquid extraction with hexane:isoamylalcohol (98:2) and back extraction with 0.005 M formic acid solution. This method is specific and linear over the concentration range of 1.00-60.00 ng/ml for mianserin and 0.50-14.00 ng/ml for N-desmethylmianserin in human plasma. The lowest limits of quantification (LLQ) is 1.00 ng/ml for mianserin and 0.50 ng/ml for N-desmethylmianserin. Intraday and interday precision (%C.V.) is <10% for both mianserin and N-desmethylmianserin. The accuracy ranges from 94.44 to 112.33% for mianserin and 91.85-100.13% for N-desmethylmianserin. The stability studies showed that mianserin and N-desmethylmianserin in human plasma are stable during short-term period for sample preparation and analysis. The method was used to assay mianserin and its metabolite, N-desmethylmianserin, in human plasma samples obtained from subjects who had been given an oral tablet of 30 mg of mianserin.

Calibration↗
PubMed2001

Fully automated on-line determination of olanzapine in serum for routine therapeutic drug monitoring.

A method is presented for unattended fully automated extraction and on-line determination of the atypical antipsychotic drug olanzapine in serum. An ASPEC automatic sample-preparing apparatus with Isolute cyanopropyl-bonded silicagel cartridges was used for solid-phase extraction of the drugs from serum. The adsorbed drugs were eluted with methanol and an aliquot injected into a high-performance liquid chromatograph (HPLC) apparatus. Trifluoperazine was used as internal standard, and the analytes were separated on an unmodified silicagel column using methanol-ammonium acetate buffer pH 9.9 (85:15) as mobile phase. Ultraviolet detection at 257 nm was used for quantitation. Within the therapeutic range for the serum concentration of olanzapine, the interday variations for the quantitative determinations were <8%. Comparisons between concentrations measured using liquid-liquid extraction and the present on-line extraction method showed good agreement. Other drugs often used in combination with olanzapine did not interfere with the quantitative determinations. The method has been in routine use for more than 1 year for therapeutic drug monitoring.

Antipsychotic Agents↗
PubMed2006

A rapid and sensitive method for determination of sorafenib in human plasma using a liquid chromatography/tandem mass spectrometry assay.

A rapid, sensitive and specific method was developed and validated using LC/MS/MS for determination of sorafenib in human plasma. Sample preparation involved a single protein precipitation step by the addition of 0.1 mL of plasma with 0.5 mL acetonitrile. Analysis of the compounds of interest including the internal standard ([(2)H(3)(15)N] sorafenib) was achieved on a Waters X-Terra C(18) (150 mm x 2.1mm i.d., 3.5 microm) analytical column using a mobile phase consisting of acetonitrile/10 mM ammonium acetate (65:35, v/v) containing 0.1% formic acid and isocratic flow at 0.2 mL/min for 6 min. The analytes were monitored by tandem mass spectrometry with electrospray positive ionization. Linear calibration curves were generated over the range of 7.3-7260 ng/mL for the human plasma samples with values for the coefficient of determination of >0.96. The values for both within day and between day precision and accuracy were well within the generally accepted criteria for analytical methods (<15%).

Area Under Curve↗
PubMed2005

On-line cation-exchange preconcentration and capillary electrophoresis coupled by tee joint interface.

An on-line preconcentration method based on ion exchange solid phase extraction was developed for the determination of cationic analytes in capillary electrophoresis (CE). The preconcentration-separation system consisted of a preconcentration capillary bonded with carboxyl cation-exchange stationary phase, a separation capillary for zone electrophoresis and a tee joint interface of the capillaries. Two capillaries were connected closely inside a 0.3 mm i.d. polytetrafluoroethylene tube with a side opening and fixed together by the interface. The preparations of the preconcentration capillaries and interface were described in detail in this paper. The on-line preconcentration and separation procedure of the analysis system included washing and conditioning the capillaries, loading analytes, filling with buffer solution, eluting analytes and separating by capillary zone electrophoresis (CZE). Several analysis parameters, including sample loading flow rate and time, eluting solution and volume, inner diameter and length of preconcentration capillary etc., were investigated. The proposed method enhanced the detection sensitivity of CE-UV about 5000 times for propranolol and metoprolol compared with normally electrokinetic injection. The detection limits of propranolol and metoprolol were 0.02 and 0.1 microg/L with the proposed method respectively, whereas those were 0.1 and 0.5 mg/L with conventional electrokinetic injection. The experiment results demonstrate that the proposed technique can increase the preconcentration factor evidently.

Ammonium Chloride↗
PubMed1997

An isotope-dilution gas chromatography-mass spectrometry method for trace analysis of xylene metabolites in tissues.

A gas chromatography-mass spectrometry (GC-MS) method using isotope dilution was developed to measure trace levels of xylene metabolites in brain tissues. The primary metabolites of xylene are dimethylphenol (DMP), methylbenzyl alcohol (MBA), toluic acid (TA), and methylhippuric acid (MHA). The internal standard was a mixture of deuterated DMP-d3, TA-d7, and MHA-d7. DMP-d3 was commercially available and was used as the internal standard for both DMP and MBA. TA-d7 and MHA-d7 were biosynthesized by administering xylene-d10 to rats and collecting their urine. Based on the noise peaks in 10 blank samples, the on-column limits of quantitation (mean +10 SD of noise peaks) were approximately 305, 1220, 545, and 386 pg for DMP, MBA, TA, and MHA, respectively. Analyte detection and recovery tests from brain tissues of control rats were conducted by spiking the tissues with 32 nmol/g of each analyte, together with the deuterated metabolites. The tissues were homogenized, extracted with ethyl acetate, and derivatized by trimethylsilylation. One microliter of the sample was injected into the GC-MS. The recoveries of the analytes were 104 +/- 8%, 80 +/- 9%, 93 +/- 10%, and 92 +/- 11% (mean +/- SD, n = 7) for DMP, MBA, TA, and MHA, respectively. The tissue preparation efficiency, which was indicated by absolute recoveries of internal standards, was approximately 33% for DMP, MBA, and TA and approximately 80% for MHA. No metabolites were detected in untreated control tissues. This simple and sensitive method to simultaneously detect major xylene metabolites in brain tissues could also be used for the analysis of blood and urine samples from workers to monitor p-xylene exposure.

Animals↗
PubMed2005

Separation of proteins with a molecular mass difference of 2 kDa utilizing preparative double-inverted gradient polyacrylamide gel electrophoresis under nonreducing conditions: application to the isolation of 24 kDa human growth hormone.

A method for separating proteins with a molecular mass difference of 2 kDa using SDS-PAGE under nonreducing conditions is presented. A sample mixture containing several human growth hormone (hGH) isoforms was initially separated on a weak anion-exchange column. Fractions rich in 24 kDa hGH as determined by analytical SDS-PAGE were pooled and further separated by cation-exchange chromatography. The fractions pooled from the cation-exchange chromatography contained two hGH isoforms with a 2 kDa molecular mass difference according to SDS-PAGE analysis, 22 and 24 kDa hGH. The 22 and 24 kDa hGH were separated using continuous-elution preparative double-inverted gradient PAGE (PDG-PAGE) under nonreducing conditions. The preparative electrophoresis gel was composed of three stacked tubular polyacrylamide matrices, a 4% stacking gel, a 13-18% linear gradient gel, and a 15-10% linear inverted gradient gel. Fractions containing purified 24 kDa hGH were pooled and Western blot analysis displayed immunoreactivity to antihGH antibodies. PDG-PAGE provides researchers with an electrophoretic technique to preparatively purify proteins under nonreducing conditions with molecular mass differences of 2 kDa.

Chromatography↗
PubMed1999

Chromatographic separation of carotenoids.

The carotenoids are extremely reactive and consequently unstable due to their long system of conjugated double bonds. Several precautions, such as protection against light and oxygen, use of low temperature and antioxidants, analysis in the shortest possible time, should be taken during isolation and chromatography. The food samples, preferably fresh, are homogenized and immediately extracted with a suitable organic solvent. Saponification has been employed in order to hydrolyze the carotenoid esters, remove fatty material and destroy chlorophyll. This optional step facilitates subsequent carotenoid separation, identification and quantification. The separation of carotenoids is usually carried out by column chromatography, thin layer chromatography and high performance liquid chromatography, in analytical or preparative scale, on many stationary phases such as silica-gel, alumina, MgO, Ca(OH)2 and reversed-phase material (C18 and C30). The choice of the most suitable chromatographic method depends on the amount of sample, carotenoid composition, resolution, speed and purity required. Examples of carotenoid separation in different stationary phases will be shown and discussed.

Carotenoids↗