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PubMed1995

Thin-layer chromatographic procedures for lipid separation.

This review focuses on the thin-layer chromatographic (TLC) separation aspects of lipid analysis. Since the space limitations do not permit, the quantitative aspects of the analyses are not discussed at length although some indications of appropriate methodology and detection reagents will be indicated. Many separations carried out by TLC have the prerequisite of proper sample preparation. Using proper sample clean-up prior to TLC enables one to carry out precise separation as well as sensitive quantitation. Thus, the discussions are divided into the two main topics--sample preparation and TLC. Examples of applications are limited to those which illustrate the capabilities of the technique as well as practicability. Since there are a number of reviews in the literature, the discussions herein are focused mainly on reports after 1985.

Chromatography, Thin Layer↗
PubMed2003

Practical approaches to fast gas chromatography-mass spectrometry.

Fast gas chromatography-mass spectrometry (GC-MS) has the potential to be a powerful tool in routine analytical laboratories by increasing sample throughput and improving laboratory efficiency. However, this potential has rarely been met in practice because other laboratory operations and sample preparation typically limit sample throughput, not the GC-MS analysis. The intent of this article is to critically review current approaches to fast analysis using GC-MS and to discuss practical considerations in addressing their advantages and disadvantages to meet particular application needs. The practical ways to speed the analytical process in GC and MS individually and in combination are presented, and the trade-offs and compromises in terms of sensitivity and/or selectivity are discussed. Also, the five main current approaches to fast GC-MS are described, which involve the use of: (1) short, microbore capillary GC columns; (2) fast temperature programming; (3) low-pressure GC-MS; (4) supersonic molecular beam for MS at high GC carrier gas flow; and (5) pressure-tunable GC-GC. Aspects of the different fast GC-MS approaches can be combined in some cases, and different mass analyzers may be used depending on the analytical needs. Thus, the capabilities and costs of quadrupole, ion trap, time-of-flight, and magnetic sector instruments are discussed with emphasis placed on speed. Furthermore, applications of fast GC-MS that appear in the literature are compiled and reviewed. At this time, the future usefulness of fast GC-MS depends to some extent upon improvement of existing approaches and commercialization of interesting new techniques, but moreover, a greater emphasis is needed to streamline overall laboratory operations and sample preparation procedures if fast GC-MS is to become implemented in routine applications.

Gas Chromatography-Mass Spectrometry↗
PubMed2001

Isotope dilution gas chromatography/mass spectrometry method for the determination of methionine sulfoxide in protein.

We have developed a new technique for quantifying methionine sulfoxide (MetSO) in protein to assess levels of oxidative stress in physiological systems. In this procedure, samples are hydrolyzed with methanesulfonic acid (MSA) in order to avoid the conversion of MetSO to methionine (Met) that occurs during hydrolysis of protein in HCl. The hydrolysate is fractionated on a cation exchange column to remove the nonvolatile MSA from amino acids, and the amino acids are then derivatized as their trimethylsilyl esters for analysis by selected ion monitoring-gas chromatography/mass spectrometry. The limit of detection of the assay is 200 pmol of MetSO per analysis, and the interassay coefficient of variation is 5.8%. Compared to current methods, the SIM-GC/MS assay avoids the potential for conversion of Met to MetSO during sample preparation, requires less sample preparation time, has lower variability, and uses mass spectrometry for sensitive and specific analyte detection.

Collagen↗
PubMed2000

Characterization of synthetic polymers by MALDI-TOF/MS: investigation into new methods of sample target preparation and consequence on mass spectrum finger print

Characterization of synthetic polymers by Matrix assisted laser desorption (MALDI) is limited by the solubility of different oligomers in a suitable solvent, and the fingerprint of the mass spectrum is affected by the properties of solvents employed (eg., pH, secondary solvents, evaporation) during sample target preparation. If solvents are not used during sample target preparation, then solvent properties should not play an important role in determining the quality of the MALDI mass spectrum. We report here two solventless approaches for sample target preparation. It was observed that Poly(ethylene glycol) 6000 (PEG) showed the same molecular mass distribution in different modes of sample target preparation. Fluorinated polymer used in these studies was affected by sample target preparation protocol and by target surface. Pyrolysis of PEG oligomers was observed in all the methods of sample target preparation. The desorbed high mass neutral oligomers fragment to give small oligomers which are then cationized by the desolvation of the cationized matrix clusters. Moreover, the origin of the matrix clusters (i.e., formed in the condensed phase or in the gas phase) determines the relative intensities of PEG oligomers cationized by sodium or potassium.

Journal Article↗
PubMed2000

Molecular insights into neurodevelopmental and neurodegenerative diseases.

Magnetic resonance spectroscopy (MRS) is a non-invasive physical technique that is routinely used to determine the quantity and structure of organic molecules in solution. Technical advances that have expanded the usefulness of this technique include: (1) high resolution MRS to identify and quantify individual molecules present in complex mixtures of tissue extracts; (2) in vivo MRS techniques to non-invasively monitor metabolites in humans; (3) structure determination of proteins of moderate size; and (4) improved structure characterization of solids and liquid crystals, such as the detection of phase changes in membranes. The focus of this review is on the first two technical advances mentioned above. The strengths of MRS as a research tool to investigate molecular alterations in disease states include ease of sample preparation, minimum sample manipulation, avoidance of the preparation of derivatives, and the ability to analyze an unfractionated sample. The strengths of MRS in the clinic are its ability to measure neuronal metabolite levels non-invasively in humans and its potential for disease diagnosis, monitoring disease progression, and assessing the efficacy of experimental therapies.

Aging↗
PubMed1990

Development of a laboratory robotic system for automated bioanalytical methods--I. The determination of theophylline in human plasma: a comparison between the robotized and manual method.

The quality of bioanalytical methods is often determined by the quality of sample preparation. Using a robot for sample treatment may give better results than manual sample preparation, since the robot lacks human behaviour and incidental errors that are part of it. The use of a laboratory robot has the additional advantage of giving each sample the same analytical history, resulting in better reproducibility. An automated method has been developed for the analysis of drugs in plasma using a laboratory robot. Theophylline was used as a probe drug. Sample preparation was automated with a Zymate II robot, followed by separation and quantitation on an HPLC-system. The robotic method showed a good correlation with the manual method, while sample throughput was doubled.

Chemistry, Clinical↗
PubMed1999

[Preparation technique of S2OF10 gas standard sample and determination method of the trace S2OF10 in SF6].

In this paper a series of methods and techniques for the S2OF10 standard sample preparation and quantitative determination are presented. They are, the preparation of S2OF10 by preparative chromatography with adsorption/thermal desorption, the standard sample of S2OF10 gas prepared by exponential dilution and the gas chromatography/flame photometric detector(GC/FPD) determination of trace S2OF10 from an SF6 sample with quantitative calibration factor. Especially, the S2OF10 gas from a used SF6 sample was directly separated and concentrated through a U-adsorbent-tube packed with 300 mg of Porasil A in a cold trap (-63 degrees C) with liquid-nitrogen and chloroform. Then it was purified by preparative-GC and to be injected into a preparative system of standard gas sample. In the meantime, the S2OF10 gas obtained was confirmed by the methods of GC/FPD, infrared spectrophotometer(IR) and gas chromatography/mass spectrometer(GC/MS) separately. The sub-ppm(by volume) level of the S2OF10 and SF6 mixture samples were prepared by use of the exponential dilution system. The GC/FPD experimental results showed that the detection linear range of S2OF10 gas concentration was 0.80 x 10(-6)-2.60 x 10(-4) (volume fraction) and the quantitative calibration factor of the S2OF10 was 0.197 based on SF6. The determination errors of quantitative calibration factor were 1.8%-20% and S2OF10 recovery of the adsorption/thermal desorption was 98.2% (n = 9) and its relative standard deviation was 6.2%. In addition, the results also showed that it is a simple and rapid method with good linearity and reproducibility.

English Abstract↗
PubMed1996

Characterization of specific noncovalent protein complexes by UV matrix-assisted laser desorption ionization mass spectrometry.

While electrospray (ESI) mass spectrometry has already established its potential for the characterization of non-covalent protein complexes, matrix-assisted laser desorption/ionization mass spectrometry (MALDI/MS) seemed not to be applicable hitherto because of limitations in matrix chemistry and sample preparation. In this work, a sample preparation method has been developed in which 6-aza-2-thiothymine (ATT) was used as a matrix without any addition of organic cosolvents, and proteins were dissolved in aqueous buffers such as ammonium hydrogencarbonate, ammonium citrate and ammonium acetate. Under these conditions, the intact non-covalent protein complexes, RNAse S, the non-covalent complex of S-protein and S-peptide and specific dimers of coiled-coil leucine zipper polypeptides were observed by UV-MALDI/MS. The specificity of complex formation was ascertained by admixture of non-specific peptides which did not yield detectable aggregate ions. In addition, on-target tryptic digestion of cytochrome c and leucine zipper peptides was carried out after MALDI/MS molecular mass determination in the presence of the ATT matrix. Mass spectrometric analyses of these tryptic digests yielded spectra that showed complete digestion of the proteins. These results indicate that proteins maintained intact tertiary structures necessary for the formation of specific non-covalent complexes, and that trypsin retained its functional enzymatic structure and full biological activity with the present sample preparation method.

Amino Acid Sequence↗
PubMed2006

Ultrasound in analytical chemistry.

Ultrasound is a type of energy which can help analytical chemists in almost all their laboratory tasks, from cleaning to detection. A generic view of the different steps which can be assisted by ultrasound is given here. These steps include preliminary operations usually not considered in most analytical methods (e.g. cleaning, degassing, and atomization), sample preparation being the main area of application. In sample preparation ultrasound is used to assist solid-sample treatment (e.g. digestion, leaching, slurry formation) and liquid-sample preparation (e.g. liquid-liquid extraction, emulsification, homogenization) or to promote heterogeneous sample treatment (e.g. filtration, aggregation, dissolution of solids, crystallization, precipitation, defoaming, degassing). Detection techniques based on use of ultrasonic radiation, the principles on which they are based, responses, and the quantities measured are also discussed.

Analytic Sample Preparation Methods↗
PubMed2004

[GC-MS analysis of methamphetamine and amphetamine in hair of Thai drug addicts].

UNLABELLED: Psycho-stimulant dependence in young individuals has become a serious problem in Thailand, where consumption of the so-called YaBa methamphetamine tablets has become a fashionable trend. Due to its easy availability in the form of a tablet, young individuals abuse methamphetamine. Methamphetamine tablets are known to be potently addictive and its difficulty in cessation of drug use and to be abstinent from the drug. We herein report the results obtained from GC-MS analysis of methamphetamine and amphetamine in 33 samples of urine and hair from patients with psycho-stimulant dependence. These samples were collected from patients registered at the outpatient clinic in the Department of Psychiatry, Chiang Mai University Hospital and were sent to Nippon Medical School, Department of Legal Medicine (Tokyo, Japan) for further analysis by Dr. Werawan Ruangyttikarn, Department of Forensic Medicine, Chiang Mai University. Sample preparation: Hairs samples were cropped near the hair root. After washing, they were cut into 1.2-cm sections and extracted with methanol/5N HCl (2:1) for an hour and then, solid-phase extraction was conducted using Bond-Elut Certify. Following extraction, GC-MS analysis was performed. Urine samples were subjected to GC-MS analysis after preparation with Bond Elut Certify. RESULTS AND DISCUSSION: In 6 samples, both urine and hair samples analyzed were negative for detection of the stimulant drugs. In those cases individuals might stop taking drug for about 5 months. In 18 samples, urine samples were negative whereas hair samples were positive. These results suggest that individuals might stop using drugs for a few days before they went to the hospital but they abuse drugs continuously. In 9 samples, both urine and hair samples were positive. These results show that individuals always abuse drugs. In order to treat drug dependence effectively it is necessary to obtain the patient history of drug use and to evaluate and determine short-term and long-term drug use with urinalysis and hair analysis, respectively. Our present data revealed that useful information concerned with the long term drug abuse can be obtained from hair analysis, and that this method of analysis is applicable not only to forensic cases but also for evaluating clinical cases.

Adolescent↗
PubMed1999

Coupling of biological sample handling and capillary electrophoresis.

The analysis of biological samples (e.g., blood, urine, saliva, tissue homogenates) by capillary electrophoresis (CE) requires efficient sample preparation (i.e., concentration and clean-up) procedures to remove interfering solutes (endogenous/exogenous and/or low-/high-molecular-mass), (in)organic salts and particulate matter. The sample preparation modules can be coupled with CE either off-line (manual), at-line (robotic interface), on-line (coupling via a transfer line) or in-line (complete integration between sample preparation and separation system). Sample preparation systems reported in the literature are based on chromatographic, electrophoretic or membrane-based procedures. The combination of automated sample preparation and CE is especially useful if complex samples have to be analyzed and helps to improve both selectivity and sensitivity. In this review, the different modes of solid-phase (micro-) extraction will be discussed and an overview of the potential of chromatographic, electrophoretic (e.g., isotachophoresis, sample stacking) and membrane-based procedures will be given.

Electrophoresis, Capillary↗
PubMed2004

Matrix effect in LC-ESI-MS and LC-APCI-MS with off-line and on-line extraction procedures.

Matrix effect on mass spectrometry response was investigated with commercially available electrospray ionisation (ESI) and atmospheric pressure chemical ionisation (APCI) sources coupled with a single quadruple mass spectrometer. A post-column infusion system was used to observe the MS signal alterations of methadone, selected as the model compound, in plasma. For this purpose, two sample preparation procedures were tested: (1) conventional off-line sample preparations with liquid-liquid extraction (LLE), solid-phase extraction (SPE) and protein precipitation (PP) with perchloric acid (PA) and acetonitrile (ACN) and (2) on-line SPE with two different extraction columns packed with a large particle support (LPS) and with restricted access material (RAM), respectively. Whatever the sample preparation procedures, APCI source appeared to be less liable to matrix effect than ESI source. Among the different off-line sample preparations, LLE was the most efficient extraction procedure. With other techniques, MS signal was affected not only by endogenous material but also by procedure introduced compounds. Moreover, on-line SPE-LC-MS configuration exhibited matrix effects, which depend on the API source and the extraction support.

Chromatography, Liquid↗
PubMed2003

A device for automated direct sampling and quantitation from solid-phase sorbent extraction cards by electrospray tandem mass spectrometry.

A new solid-phase extraction (SPE) device in the 96-well format (SPE Card) has been employed for automated off-line sample preparation of low-volume urine samples. On-line automated analyte elution via SPE and direct quantitation by micro ion spray mass spectrometry is reported. This sample preparation device has the format of a microtiter plate and is molded in a plastic frame which houses 96 separate sandwiched 3M Empore sorbents (0.5-mm-thickness, 8-microm particles) covered on both sides by a microfiber support material. Ninety-six discrete SPE zones, each 7 mm in diameter, are imbedded into the sheet in the conventional 9-mm pitch (spacing) of a 96-well microtiter plate. In this study one-quarter of an SPE Card (24 individual zones) was used merely as a convenience. After automated off-line interference elution of applied human urine from 24 samples, a section of SPE Card is mounted vertically on a computer-controlled X, Y, Z positioner in front of a micro ion spray direct sampling tube equipped with a beveled tip. The beveled tip of this needle robotically penetrates each SPE elution zone (sorbent disk) or stationary phase in a serial fashion. The eluted analytes are sequentially transferred directly to a microelectrosprayer to obtain tandem mass spectrometric (MS/MS) analysis. This strategy precludes any HPLC separation and the associated method development. The quantitative determination of Ritalin (methylphenidate) from fortified human urine samples is demonstrated. A trideuterated internal standard of methylphenidate was used to obtain ion current response ratios between the parent drug and the internal standard. Human control urine samples fortified from 6.6 to 3300 ng/mL (normal therapeutic levels have been determined in other studies to be between 50 and 100 ng/mL urine) were analyzed and a linear calibration curve was obtained with a correlation coefficient of 0.9999, where the precision of the quality control (QC) samples ranged from 9.6% at the 24 ng/mL QC level to 1.2% at the 3000 ng/mL QC level, and the accuracy for the four levels of QC samples ranged from 98.1% to 100.3%. The QC samples were prepared at four concentrations which included 24, 240, 1200, and 3000 ng/mL, respectively. The run time per sample in this work was 1.5 min not including the sample preparation time.

Automation↗
PubMed2001

[Determination of Fe, Al, Ca, Ti, P and Cu in silicon metal by XRF].

This article puts forward a sample preparation method that first dissolve the sample by acid and evaporate the solution then melt the remains by flux. This method resolves the problem that silicon metal can't easy to prepare sample by fusion, and it can determine several elements in the sample simultaneously. This method overcomes the limit of standard sample and removes the matrix effects. The range of determination is wide and the results is accurate. The condition of preparing sample was confirmed by test.

Aluminum↗
PubMed1977

Apparent metabolism of allopurinol by blood--a preliminary report.

Allopurinol is converted to a presently unidentified metabolite by whole blood, both in samples prepared in vitro as well as samples from subjects administered the drug. The apparent plasma concentration of allopurinol, as determined by HPLC, is dependent upon the length of time between sample preparation or sample withdrawl and the centrifugation and removal of plasma. Oxypurinol does not undergo the same conversion, nor does it affect the loss of allopurinol.

Allopurinol↗
PubMed2000

Automated extraction of 11-nor-delta9-tetrahydrocannabinol carboxylic acid from urine samples using the ASPEC XL solid-phase extraction system.

The analysis of 11-nor-delta9-tetrahydrocannabinol-carboxylic acid (THCCOOH, the major metabolite of cannabis) in urine with gas chromatography and mass spectrometry (GC-MS) and solid-phase extraction (SPE) sample preparation is well documented. Automated SPE sample preparation of THCCOOH in urine, although potentially advantageous, is to our knowledge poorly investigated. The objective of the present study was to develop and validate an automated SPE sample-preparation step using ASPEC XL suited for GC-MS confirmation analysis of THCCOOH in urine drug control. The recoveries showed that it was not possible to transfer the protocol for the manual SPE procedure with the vacuum manifold to the ASPEC XL without loss of recovery. Making the sample more lipophilic by adding 1 mL 2-propanol after hydrolysis to the urine sample in order to overcome the problem of surface adsorption of THCCOOH led to an extraction efficiency (77%) comparable to that reached with the vacuum manifold (84%). The reproducibility of the automated SPE procedure was better (coefficient of variation 5%) than that of the manual procedure (coefficient of variation 12%). The limit of detection was 1 ng/mL, and the limit of quantitation was 4 ng/mL. Precision at the 12.5-ng/mL level was as follows: mean, 12.4 and coefficient of variation, 3.0%. Potential carryover was evaluated, but a carryover effect could not be detected. It was concluded that the proposed method is suited for GC-MS confirmation urinalysis of THCCOOH for prisons and detoxification centers.

Dronabinol↗
PubMedDate not supplied

Determination of two oxy-pyrimidine metabolites of diazinon in urine by gas chromatography/mass selective detection and liquid chromatography/electrospray ionization/mass spectrometry/mass spectrometry.

An analytical method was developed for the determination in urine of 2 metabolites of diazinon: 6-methyl-2-(1-methylethyl)-4(1H)-pyrimidinone (G-27550) and 2-(1-hydroxy-1-methylethyl)-6-methyl-4(1H)-pyrimidinone (GS-31144). Two of the urine sample preparation procedures presented rely on gas chromatography/mass selective detection (GC/MSD) in the selected ion monitoring mode for determination of G-27550. For fast sample preparation and a limit of quantitation (LOQ) of 1.0 ppb, urine samples were purified by using ENV+ solid-phase extraction (SPE) columns. For analyte confirmation at an LOQ of 0.50 ppb, classical liquid/liquid partitioning was used before further purification in a silica SPE column. An SPE sample preparation procedure and liquid chromatography/electrospray ionization/mass spectrometry/mass spectrometry (LC/ESI/MS/MS) were used for both G-27550 and GS-31144. The limit of detection was 0.01 ng for G-27550 with GC/MSD, and 0.016 ng when LC/ESI/MS/MS was used for both G-27550 and GS-31144. The LOQ was 0.50 ppb for G-27550 when GC/MSD and the partitioning/SPE sample preparation procedure were used, and 1.0 ppb for the SPE only sample preparation procedure. The LOQ was 1.0 ppb for both analytes when LC/ESI/MS/MS was used.

Cholinesterase Inhibitors↗
PubMed1997

Glass wool column filtration, an advantageous way of preparing semen samples for intracytoplasmic sperm injection: an auto-controlled randomized study.

The characteristics of the glass wool filtration technique for the preparation of semen samples in an intracytoplasmic sperm injection programme (ICSI) were compared with a two-layer Percoll density gradient procedure. Half of each of 25 semen samples were prepared by each technique. The oocytes from each patient were randomly injected, half with spermatozoa prepared by glass wool filtration and half with Percoll-separated spermatozoa from the same semen sample. The percentage of recovered motile spermatozoa, the total motile sperm count, the percentage of morphologically normal forms, the fertilization rate, the cleavage rate and embryo quality obtained with both preparations were analysed. The 95% confidence intervals obtained through the Altman-Bland analysis showed that the percentage of motile spermatozoa recovered was 3.5-9.9% higher with the glass wool filtration technique, and this was highly correlated (r = 0.92, P = 0.0001) to the method. Similarly, the total number of spermatozoa available for ICSI was 0.18 x 10(6)-2.44 x 10(6) higher with the glass wool column and was highly correlated to the method (r = 0.956, P = 0.0001). Also, the percentage of normal forms was 1.25-3.31% higher after glass wool filtration but was poorly correlated to the method (r = 0.47, P = 0.017). Out of 100 metaphase II oocytes injected with glass wool-extracted spermatozoa, 77 fertilized and 72 cleaved. Out of 97 metaphase II oocytes injected with Percoll-selected spermatozoa, 71 fertilized and 69 cleaved. These results were not statistically different. The mean +/- SEM embryo quality score for the glass wool group (2.90 +/- 0.27) was the same as that for the Percoll group (2.80 +/- 0.24). No fertilization failures occurred and 11 patients (44% per oocyte retrieval) became pregnant. It was concluded that glass wool filtration for semen preparation in an ICSI programme offers higher sperm recovery and sperm morphology of superior quality than with the classic two-layer Percoll gradient method, without affecting the fertilization rate and embryo quality.

Cell Separation↗