Simultaneous determination of procaine and para-aminobenzoic acid by LC-MS/MS method

A sensitive high performance liquid chromatography tandem mass spectrometry (LC-MS/MS) method has been developed for simultaneous determination of procaine and its metabolite p-aminobenzoic acid (PABA). N-Acetylprocainamide (NAPA) was used as an internal standard for procaine and PABA analysis. This assay method has also been validated in terms of linearity, lower limit of detection, lower limit of quantitation, accuracy and precision as per ICH guidelines. Chromatography was carried out on an XTerra MS C(18) column and mass spectrometric analysis was performed using a Quattro Micro mass spectrometer working with electro-spray ionization (ESI) source in the positive ion mode. Enhanced selectivity was achieved using multiple reaction monitoring (MRM) functions, m/z 237-->100, m/z 138-->120, and m/z 278-->205 for procaine, PABA and NAPA, respectively. Retention times for PABA, procaine and NAPA were 4.0, 4.7 and 5.8min, respectively. Linearity for each calibration curve was observed across a range from 100nM to 5000nM for PABA, and from 10nM to 5000nM for procaine. The intra- and inter-day relative standard deviations (RSD) were <5%.     

Enantioselective LC/MS method for the determination of an antimalarial agent Fenozan B07 in dog plasma

A chiral liquid chromatography/mass spectrometry (LC/MS) bioanalytical procedure has been developed for the analysis of the antimalaric agent Fenozan B07 in dog plasma. Normal-phase chromatography involving a phenylcarbamate derivative of cellulose coated on silica gel as the chiral stationary phase was used to resolve (-)-(S,S)-B07 from (+)-(R,R)-B07. The enantiomers were detected by a mass spectrometer equipped with an atmospheric pressure chemical ionization (APCI) interface operated in the negative ion mode. A mass spectrum, characterized by a base peak of m/z 285, was obtained for each enantiomer. The m/z 285 ion was very specific for the analysis of both enantiomers in the plasma. The selected ion monitoring analysis of the plasma samples was therefore performed at m/z 285 for quantitative purposes. The enantiomers were extracted from the plasma in a basic medium and purified by solid-phase extraction using a hydrophilic-lipophilic balanced sorbent. A lower limit of quantification of 2 ng/mL in plasma was achieved for both enantiomers. The quantitative procedure reported in this study was highly specific and sensitive, and was validated according to the FDA guidance on bioanalytical method validation.     

Determination of urinary phytoestrogens by HPLC-MS/MS: a comparison of atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI)

A comparison of the analytical performance of atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI) for the quantitative determination of six urinary phytoestrogens (daidzein, O-desmethylangolensin, equol, enterodiol, enterolactone and genistein) by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) is presented here. Both APCI and ESI were suitable for the analysis of these compounds; however, ESI did improve measurement imprecision and sensitivity in certain cases. Method imprecision (between-run coefficients of variation [CVs] from duplicate analysis of three quality control [QC] urine pools across 20 runs) was 5.6-12% for ESI, as opposed to 5.3-30% for APCI. At low concentrations (3-60 ng/mL, analyte dependent) imprecision was lower with ESI, whereas both techniques were generally commensurate at high concentrations (200-1000 ng/mL, analyte dependent). Method accuracy (spiked analyte recovery from the QC pools) was comparable between techniques: 86-114% for ESI; 95-105% for APCI. Limits of detection (LODs) were equivalent or better with ESI compared to APCI, with the most significant LOD improvement observed for equol (ESI: 0.3 ng/mL; APCI: 2.7 ng/mL). This translated into a substantial increase in equol detection frequency (% of sample results above LOD) within a random patient sample subset (98% for ESI, compared to 81% for APCI, n=378). Correlation (Pearson) and agreement (Deming regression, Bland-Altman bias) between ESI and APCI results in the patient subset was better in cases where imprecision and sensitivity was similar for both techniques (daidzein, enterolactone, genistein: r=0.993-0.998; slope=0.98-1.03; bias=-4.2 to -0.8%); correlation and/or agreement was poorer for analytes, where APCI imprecision and sensitivity were inferior (equol, O-desmethylangolensin, enterodiol). Baring significant factors arising from differences in ionization source design, these observations suggest that ESI is more appropriate for urinary biomonitoring of these compounds by LC-MS/MS.     

Monitoring phospholipids for assessment of matrix effects in a liquid chromatography-tandem mass spectrometry method for hydrocodone and pseudoephedrine in human plasma

Matrix effects resulting in ion suppression or enhancement have been shown to be a source of variability and inaccuracy in bioanalytical mass spectrometry. Glycerophosphocholines may cause significant matrix ionization effects during quantitative LC/MS/MS analysis and are known to fragment to form characteristic ions (m/z 184) in electrospray mass spectrometry. This ion was used to monitor ion suppression effects in the determination of hydrocodone and pseudoephedrine in human plasma as a means to track and avoid these effects. The m/z 184 ion fragment was detected in both plasma extracts and solutions of phosphatidylcholine. Post-column infusion studies showed that the ion suppression for both drugs and internal standards correlated with the elution of phospholipids. HPLC conditions were adjusted to chromatographically resolve the peaks of interest from the phospholipids. Upon repeated injection, the elution time of the phospholipids decreased while elution of the analyte peaks remained unchanged. This resulted in co-elution and significantly affected peak shape and internal standard response for the analytes. It was decided to use the phospholipid fragment to monitor this matrix effect in validation samples. The resulting method demonstrated intra-day and inter-day precision within 4.5 and 5.6% for hydrocodone and pseudoephedrine, respectively, and accuracy within 8.9 and 8.7% for hydrocodone, and pseudoephedrine, respectively. There was no statistically significant difference in the internal standard response for the determination with and without monitoring the phospholipid fragment ion. We found that monitoring the phospholipid fragment was useful in method development to avoid the matrix effects, and in routine analysis to provide a practical way to ensure the avoidance of matrix effects in each individual sample.     

HPLC/MS analysis of fusarium mycotoxins, fumonisins and deoxynivalenol

Fusarium fungi are widely found in agricultural products, worldwide and can produce a great variety of mycotoxins. Fumonisins, produced by F. moniliforme, and deoxynivalenol, produced by F. graminearum, are two such mycotoxins that have received considerable attention as food safety concerns by regulatory agencies. High Performance Liquid Chromatography/Mass Spectrometry (HPLC/MS) was found to be a convenient analytical method to detect and quantify the naturally occurring fumonisin homologs and deoxynivalenol in extracts from grains and food products. The fumonisins are detected primarily as protonated molecules in the positive ion electrospray ionization (ESI) mode as they elute from a C-18 reverse phase column during a methanol water gradient containing acetic acid to facilitate chromatography. Deoxynivalenol can be detected as positive or negative ions in the atmospheric pressure chemical ionization (APCI) mode or in the negative ion ESI mode. One nanogram amounts of fumonisins or deoxynivalenol injected into the HPLC system are easily detected with signal to noise allowing detection limits of 1 microg g(-1) or better to easily be achieved with minimal clean-up of grain extracts.     

Electrospray ionization/mass spectrometry compatible reversed-phase separation of phospholipids: piperidine as a post column modifier for negative ion detection

An electrospray ionization (ESI) compatible separation of phospholipids (PL), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), and phosphatidylcholine (PC), was performed on a C18 column by reversed phase High Performance Liquid Chromatography (HPLC) with minimal ESI suppression. The mobile phase, used isocratically, consisted of methanol and water. ESI was used to efficiently transfer the ions present in solution to the gas phase for mass spectrometric (MS) detection. Formation of negative ions was reinforced by incorporating piperidine post column. Limits of detection (LOD) and limits of quantitation (LOQ) were experimentally determined to be 20 and 60 fmol/microl, respectively, when acquiring data in the selected ion monitoring (SIM) mode monitoring three ions with a single quadrupole MS. When acquiring data from m/z 110-900 in the scanning mode, the LOD and LOQ were experimentally determined to be 1 pmol/microl and 3 pmol/microl. When acquiring product ion spectra for m/z 747, the LOD and LOQ were experimentally determined to be 446 attomol/microl and 1.3 fmol/microl, respectively.     

A fast LC-APCI/MS method for analyzing benzodiazepines in whole blood using monolithic support

A simple and fast procedure was developed for the simultaneous determination of eight benzodiazepines (BZDs) in whole blood using liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS). Sample pretreatment was carried out using a simple liquid-liquid extraction (LLE) with n-butylchloride, and chromatographic separation was performed using a monolithic silica column to speed up the analytical process. APCI and electrospray ionization (ESI) were compared. Whereas both ionization techniques appeared suitable for BZDs, APCI was found to be slightly more sensitive, especially for the determination of frequently low-dosed compounds. The method was validated according to the guidelines of the "Société Fran?aise des Sciences et Techniques Pharmaceutiques" (SFSTP) in the concentration range of 2.5-500 microg/L. The limit of quantification (LOQ) was 2.5 microg/L for all the compounds. Validation data including linearity, precision, and trueness were obtained, allowing subtherapeutic quantification of frequently low-dosed BZDs. The high selectivity of the mass spectrometer, along with the properties of the monolithic support, allowed unequivocal analysis of the eight compounds in less than 5 min. To demonstrate the potential of the method, it was used for the analysis of benzodiazepines in postmortem blood samples.     

A new strategy for ionization enhancement by derivatization for mass spectrometry

Liquid chromatography-mass spectrometry (LC-MS) using atmospheric pressure ionization is drastically different from hitherto available analytical methods used to detect polar analytes. The electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) sources of MS have contributed to the advancement of LC-MS and LC-MS/MS techniques for the analysis of biological samples. However, one major obstacle is the weak ionization of some analytes in the ESI and APCI techniques. In this review, we introduce high-sensitivity methods using several derivatization reagents for ionization enhancement. We also present an overview of chemical derivatization methods that have been applied to small molecules, such as amino acids and steroids, in biological samples.     

A comparison of nLC-ESI-MS/MS and nLC-MALDI-MS/MS for GeLC-based protein identification and iTRAQ-based shotgun quantitative proteomics.

The use of nLC-ESI-MS/MS in shotgun proteomics experiments and GeLC-MS/MS analysis is well accepted and routinely available in most proteomics laboratories. However, the same cannot be said for nLC-MALDI MS/MS, which has yet to experience such widespread acceptance, despite the fact that the MALDI technology offers several critical advantages over ESI. As an illustration, in an analysis of moderately complex sample of E. coli proteins, the use MALDI in addition to ESI in GeLC-MS/MS resulted in a 16% average increase in protein identifications, while with more complex samples the number of additional protein identifications increased by an average of 45%. The size of the unique peptides identified by MALDI was, on average, 25% larger than the unique peptides identified by ESI, and they were found to be slightly more hydrophilic. The insensitivity of MALDI to the presence of ionization suppression agents was shown to be a significant advantage, suggesting it be used as a complement to ESI when ion suppression is a possibility. Furthermore, the higher resolution of the TOF/TOF instrument improved the sensitivity, accuracy, and precision of the data over that obtained using only ESI-based iTRAQ experiments using a linear ion trap. Nevertheless, accurate data can be generated with either instrument. These results demonstrate that coupling nanoLC with both ESI and MALDI ionization interfaces improves proteome coverage, reduces the deleterious effects of ionization suppression agents, and improves quantitation, particularly in complex samples.     

Determination of 17 illicit drugs in oral fluid using isotope dilution ultra-high performance liquid chromatography/tandem mass spectrometry with three atmospheric pressure ionizations

The collection of oral fluid for drug testing is easy and non-invasive. This study developed a drug testing method using ultra-high performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS) in selected-reaction monitoring (SRM) mode. We tested the method on the analysis of four opiates and their metabolites, five amphetamines, flunitrazepam and its two metabolites, and cocaine and its four metabolites in oral fluid. 100-μL samples of oral fluid were diluted with twice the amount of water then spiked with isotope-labeled internal standards. After the samples had undergone high-speed centrifugation for 20 min, we analyzed the supernatant. The recovery of the sample preparation ranged from 81 to 108%. We compared the performance of electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI) and atmospheric pressure photoionization (APPI). The ion suppression of most analytes on ESI (28-78%) was lower than that of APCI and APPI. A post-column flow split (5:1) did not reduce the matrix effect on ESI. Direct APPI performed better than dopant-assisted APPI using toluene. ESI, APCI and APPI limits of quantitation mostly ranged from 0.11 to 1.9 ng/mL, 0.02 to 2.2 ng/mL and 0.02 to 2.1 ng/mL, respectively, but were much higher on amphetamine and ecgonine methyl ester (about 2.7-4.7 ng/mL, 8.7-14 ng/mL, and 10-19 ng/mL, respectively). Most of the bias percentages (accuracy) and relative standard deviations (precision) on spiked samples were below 15%. This method greatly simplifies the process of sample preparation and shortens the chromatographic time to only 7.5 min per run and is able to detect analytes at sub-ppb levels.     

Determination of 5-aminoimidazole-4-carboxamide in human plasma by ion-pair extraction and LC-MS/MS

A liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was established for the determination of 5-aminoimidazole-4-carboxamide (AICA) in human plasma. The method included a solvent extraction of AICA as an ion pair with 1-pentanesulfonate ion and a separation on a Hypersil ODS2 column with the mobile phase of methanol-water (68:32, v/v). Determination was performed using an electrospray ionization source in positive ion mode (ESI(+)). Multiple reaction monitoring (MRM) was utilized for the detection monitoring m/z at 127-->110 for AICA, and 172-->128 for IS. The calibration curve was linear within a range from 20 to 2000 ng/mL and the limit of quantity for AICA in plasma was 20 ng/mL. RSD of intra-assay and inter-assay were no more than 5.90% and 5.65%.     

A sample extraction and chromatographic strategy for increasing LC/MS detection coverage of the erythrocyte metabolome

Reproducible and comprehensive sample extraction and detection of metabolites with a broad range of physico-chemical properties from biological matrices can be a highly challenging process. A single LC/MS separation method was developed for a 2.1mmx100mm, 1.8mum ZORBAX SB-Aq column that was used to separate human erythrocyte metabolites extracted under sample extraction solvent conditions where the pH was neutral or had been adjusted to either, pH 2, 6 or 9. Internal standards were included and evaluated for tracking sample extraction efficiency. Through the combination of electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) techniques in both positive (+) and negative (-) ion modes, a total of 2370 features (compounds and associated compound related components: isotopes, adducts and dimers) were detected across all pHs. Broader coverage of the detected metabolome was achieved by observing that (1) performing extractions at pH 2 and 9, leads to a combined 92% increase in detected features over pH 7 alone; and (2) including APCI in the analysis results in a 34% increase in detected features, across all pHs, than the total number detected by ESI. A significant dependency of extraction solvent pH on the recovery of heme and other compounds was observed in erythrocytes and underscores the need for a comprehensive sample extraction strategy and LC/MS analysis in metabolomics profiling experiments.     

Systematic and comprehensive strategy for reducing matrix effects in LC/MS/MS analyses

A systematic, comprehensive strategy that optimizes sample preparation and chromatography to minimize matrix effects in bioanalytical LC/MS/MS assays was developed. Comparisons were made among several sample preparation methods, including protein precipitation (PPT), liquid-liquid extraction (LLE), pure cation exchange solid-phase extraction (SPE), reversed-phase SPE and mixed-mode SPE. The influence of mobile phase pH and gradient duration on the selectivity and sensitivity for both matrix components and basic analytes was investigated. Matrix effects and overall sensitivity and resolution between UPLC technology and HPLC were compared. The amount of specific matrix components, or class of matrix components, was measured in the sample preparation extracts by LC/MS/MS with electrospray ionization (ESI) using both precursor ion scanning mode and multiple reaction monitoring (MRM). PPT is the least effective sample preparation technique, often resulting in significant matrix effects due to the presence of many residual matrix components. Reversed-phase and pure cation exchange SPE methods resulted in cleaner extracts and reduced matrix effects compared to PPT. The cleanest extracts, however, were produced with polymeric mixed-mode SPE (both reversed-phase and ion exchange retention mechanisms). These mixed-mode sorbents dramatically reduced the levels of residual matrix components from biological samples, leading to significant reduction in matrix effects. LLE also provided clean final extracts. However, analyte recovery, particularly for polar analytes, was very low. Mobile phase pH was manipulated to alter the retention of basic compounds relative to phospholipids, whose retention tends to be relatively independent of pH. In addition to the expected resolution, speed and sensitivity benefits of UPLC technology, a paired t-test demonstrated a statistically significant improvement with respect to matrix effects when this technology was chosen over traditional HPLC. The combination of polymeric mixed-mode SPE, the appropriate mobile phase pH and UPLC technology provides significant advantages for reducing matrix effects resulting from plasma matrix components and in improving the ruggedness and sensitivity of bioanalytical methods.     

Application of high-performance liquid chromatography-mass spectrometry to detection of diuretics in human urine

A rapid, sensitive and reliable high-performance liquid chromatographic-mass spectrometric method for the detection of 25 diuretics in human urine has been developed. Atmosphere pressure chemical ionization (APCI) and electrospray ionization (ESI) modes were evaluated. A 2-ml volume of urine was extracted under basic conditions and separated on an Agilent Zorbax SB-C(18) column (150 x 2.1 mm, 5 microm). The mobile phase consisted of formic ammonium-formic acid buffer (pH 3.5) and acetonitrile. The effects of capillary temperature, sheath gas pressure and compositions of mobile phase on the sensitivity were studied. The recoveries of most of the diuretics were 75-95%. In the full scan mode, the limits of detection of the 25 diuretics were 0.25-25 ng/ml for APCI and 0.6-250 ng/ml for ESI. Under the optimal conditions, 14 diuretics from authentic urine samples were detected successfully by LC-APCI-MS. To obtain more fragmentation information on the chemical structure for positive confirmation, tandem mass analysis was also investigated.     

Quantitative determination of endogenous sorbitol and fructose in human erythrocytes by atmospheric-pressure chemical ionization LC tandem mass spectrometry

Evaluation of different extraction methods for quantification of endogenous sorbitol and fructose in human red blood cells (RBCs) and matrix effects in ESI and APCI showed that protein-precipitation followed by mixed-mode solid-phase extraction was more effective extraction method and APCI more effective ionization method. Then the LC/APCI-MS/MS method was fully validated and successfully applied to analysis of clinical RBC samples. The concentrations of endogenous sorbitol and fructose were determined using calibration curves employing sorbitiol-13C6 and fructose-13C6 as surrogate analytes. The method has provided excellent intra- and inter-assay precision and accuracy with a linear range of 50.0-10,000 ng/mL (correlation coefficient >0.999) for sorbitol-13C6 and 250-50000 ng/mL (correlation coefficient >0.999) for fructose-13C6 in human RBCs.     

Simultaneous determination of five flavonoids from Scutellaria Barbata extract in rat plasma by LC-MS/MS and its application to pharmacokinetic study

A new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been developed and validated for the determination of five flavonoids including scutellarin, naringenin, apigenin, luteoline and wogonin in rat plasma using sulfamethalazole as internal standard (IS). Plasma samples were pretreated with liquid-liquid extraction procedure and acid hydrolysis method was used for converting conjugated flavonoids to their respective free forms. The chromatographic separation was performed on a C(18) column with a linear gradient elution using a mobile phase consisted of 0.01% acetic acid and methanol. The detection was accomplished by multiple-reaction monitoring (MRM) scanning with electrospray ionization (ESI) source operating in the negative ionization mode. The optimized mass transition ion-pairs (m/z) monitored for scutellarin, naringenin, apigenin, luteoline, wogonin and IS were 461.1/285.1, 271.0/119.0, 269.0/117.0, 285.0/132.9, 283.0/268.0 and 252.0/155.9, respectively. The method was linear for all analytes over investigated ranges with all correlation coefficients greater than 0.9915. The lower limit of quantitation (LLOQ) of scutellarin was 9.15 ng/mL and other compounds were all less than 2.0 ng/mL. The proposed method showed appropriate accuracy and repeatability and was suitable for pharmacokinetic studies of the five flavonoids after oral administration of Scutellaria Barbata extract.     

Evaluation of electrospray ionization and atmospheric pressure chemical ionization for simultaneous detection of estrone and its metabolites using high-performance liquid chromatography/tandem mass spectrometry

The levels of estrogens and/or their metabolites play important roles in carcinogenesis, reproductive function, and sexual development during perinatal and adolescence periods. The main purpose of this report was to investigate the applicability of high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) with electrospray ionization (ESI) and/or atmospheric pressure chemical ionization (APCI) for simultaneous detection of estrone (E1) and its six metabolites. Both positive and negative ionization modes in ESI and APCI were used to evaluate the signal responses of seven target analytes. Among the seven target analytes, five analytes, E1, 16alpha-hydroxyestrone, 2-methoxyestrone, 4-methoxyestrone, and 2-hydroxyestrone-3-methyl, produced signals with the best signal-to-noise (S/N) ratios in positive APCI-MS/MS mode, while the other two analytes, 2-hydroxyestrone and 4-hydroxyestrone, yielded the best S/N ratios in negative ESI-MS/MS mode. Based on the results of the evaluation, HPLC-APCI-MS/MS with switching between positive and negative modes was recommended for simultaneous detection of E1 and its six metabolites. The proposed analytical scheme was successfully applied in the analysis of cell culture medium of Human liver carcinoma cells treated with varying amounts of 2,3,7,8-tetrachlorodibenzo-p-dioxin.     

High throughput quantification of cholesterol and cholesteryl ester by electrospray ionization tandem mass spectrometry (ESI-MS/MS)

Analysis of free cholesterol (FC) is not well suited for electrospray ionization (ESI); however, cholesteryl ester (CE) form ammonium adducts in positive ion mode and generate a fragment ion of m/z 369 upon collision-induced fragmentation. In order to allow parallel analysis of FC and CE using ESI tandem mass spectrometry (ESI-MS/MS), we developed an acetyl chloride derivatization method to convert FC to cholesteryl acetate (CE 2:0). Derivatization conditions were chosen to provide a quantitative conversion of FC to CE 2:0 without transesterification of naturally occurring CE species. FC and CE were analyzed by direct flow injection analysis using a fragment of m/z 369 in a combination of selected reaction monitoring (SRM) and precursor ion scan for FC and CE, respectively. Quantification was achieved using deuterated D(7)-FC and CE 17:0/CE 22:0 as internal standards as well as calibration lines generated by addition of FC and naturally occurring CE species to the respective sample matrix. The developed assay showed a precision and detection limit sufficient for routine analysis. A run time of 1.3 min and automated data analysis allow high throughput analysis. Loading of human skin fibroblast and monocyte derived macrophages with stable isotope labeled FC showed a potential application of this method in metabolism studies. Together with existing mass spectrometry methodologies for lipid analysis, the present methodology will provide a useful tool for clinical and biochemical studies and expands the lipid spectrum that can be analyzed from one lipid sample on a single instrumental platform.     

Application of electrospray ionization MS/MS and matrix-assisted laser desorption/ionization-time of flight mass spectrometry to structural analysis of the glycosyl-phosphatidylinositol-anchored protein.

We applied the improved sensitivity and soft ionization characteristics of electrospray Ionization (ESI)-MS/MS and matrix-assisted laser desorption/ionization(MALDI)-time of flight (TOF) mass spectrometry (MS) to analysis of the GPI-anchored C-terminal peptide derived from 5'-nucleotidase. ESI-MS/MS analysis was applied to the core structure (MW, 2,743). In the collision-induced dissociation (CID) spectrum, single-charged ions such as m/z 162 (glucosamine), 286 (mannose-phosphate-ethanolamine), and 447 ([mannose-phosphate-ethanolamine]-glucosamine) were clearly detected as characteristic fragment ions of the GPI-anchored peptide. On MALDI-TOF-MS analysis, heterogeneous peaks of GPI-anchored peptides were detected as single-charged ions in the positive mode. Product ions were obtained by post-source decay (PSD) of m/z 2,905 using curved field reflectron of TOF-MS. Most of the expected product ions derived from the GPI-anchored peptide, containing the core structure and an additional mannose side chain, were successively obtained. Thus, ESI-MS/MS and MALDI-TOF-PSD-MS proved to be effective and sensitive methods for analyzing the GPI-anchored peptide structure with less than 10 pmol of sample. These characteristic fragments or fragmentation patterns seem to be very useful for identification of GPI-anchored C-terminal peptides derived from any kind of GPI-anchored protein.     

Chiral liquid chromatography-tandem mass spectrometric methods for stereoisomeric pharmaceutical determinations

The characterization of the drug metabolism and pharmacokinetic (DMPK) profiles of stereoisomers is a fundamental aspect of the drug discovery and development processes. Therefore, chiral drug bioassays are very important to pharmaceutical and biomedical researchers. The recent developments in chiral liquid chromatography coupled to atmospheric pressure ionization tandem mass spectrometry (LC-API-MS/MS) for the analysis of pharmaceuticals are reviewed. Various ionization techniques including electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI) and atmospheric photoionization (APPI) interfaced with chiral liquid chromatographic methods are described in terms of their ionization efficiencies, matrix effects and limitations. Examples were selected to demonstrate the applicability of these methods for enantioselective bioanalysis.