Performance of zwitterionic hydrophilic interaction LC for the determination of iodinated X‐ray contrast agents

This study compares the separation performance of a group of iodinated X‐ray contrast media on four different columns. The first three were two stationary phases (SPs) modified with C₁₈ and a polar‐embedded SP (polar amide group bonded to an alkyl chain), all of which worked under RP‐LC mode. The fourth was a zwitterionic sulphoalkylbetaine SP, working under the hydrophilic interaction LC (HILIC) mode. After the optimisation of the different parameters, the zwitterionic column displayed the best separation, which also overcomes the problems encountered when these analytes were separated under RP‐LC. Moreover, when HILIC is coupled to MS/MS, sensitivity is enhanced. However, when sewage samples were analysed by SPE followed by the optimal HILIC–MS/MS, the sensitivity of the method was affected due to the high matrix effect, which had to be solved by dilution of the extract. Finally, the method was preliminarily validated with sewage and the figures of merit were comparable to those of the SPE–RP‐LC–MS/MS.     

Identification of phototransformation products of sildenafil (Viagra) and its N‐demethylated human metabolite under simulated sunlight

Recent publications on pharmaceutical monitoring are increasingly covering the field of illicit drugs and lately the forensic evaluation of designing illegal analogs of lifestyle drugs like the phosphodiesterase type 5 (PDE‐5) inhibitors Viagra (sildenafil), Levitra (vardenafil) and Cialis (tadalafil). Recently, the presence of all three erectile dysfunction treatment drugs has been reported in wastewaters at very low concentrations. In the environment, contaminants undergo various physical or chemical processes classified into abiotic (photolysis, hydrolysis) and biotic (biodegradation) reactions. Thus, changes in the chemical structure lead to the formation of new transformation products, which may persist in the environment or be further degraded. This study describes the photolysis of sildenafil (SDF) and its human metabolite N‐demethylsildenafil (DM‐SDF) under simulated solar radiation (Xenon lamp). Following chromatographic separation of the irradiated samples, eight photoproducts in the SDF samples and six photoproducts for DM‐SDF were detected and characterized. The combination of ultra performance liquid chromatography‐electrospray ionization‐quadrupole time‐of‐flight‐mass spectrometry (UPLC‐ESI‐QToF‐MS), liquid chromatography‐atmospheric pressure chemical ionization‐triple quadrupole mass spectrometry (LC‐APCI‐QqQ‐MS) and hydrogen/deuterium‐exchange experiments allowed to propose plausible chemical structures for the photoproducts, taking into account the characteristic fragmentation patterns and the accurate mass measurements. These mass spectral data provided sound evidence for the susceptibility of the piperazine ring toward photodegradation. A gradual breakdown of this heterocyclic structure gave rise to a series of products, which in part were identical for SDF and DM‐SDF. The sulfonic acid, as the formal product of sulfonamide hydrolysis, was identified as key intermediate in the photolysis pathway. In both drug/metabolite molecules, phototransformation processes taking place beyond the sulfonamide group were deemed to be of minor relevance. Copyright © 2012 John Wiley & Sons, Ltd.     

ESI‐MS Studies on Alcoholysis Mechanism of Electron‐deficient Cyclopropane Derivatives

Electrospray ionization mass spectrometry (ESI‐MS) was utilized to perform monitoring of the intermediates in the reaction of 1,2,3‐trisubstituted electron‐deficient cyclopropane derivatives, cis‐1‐thien‐2′‐oyl‐2‐(p‐subustituted phenyl‐6,6‐dimethyl)‐5,7‐dioxaspiro[2.5]‐4,8‐octadiones, with methanol. Key intermediates, either cationic or protonated forms of neutral species, were intercepted and characterized by ESI‐MS and its tandem version (ESI‐MS/MS). Therefore, the mechanism of the ring‐opening process for electron‐deficient cyclopropane derivatives was fully confirmed by the intermediates monitored.     

Qualitative and quantitative analysis of the major constituents in Acorus tatarinowii Schott by HPLC/ESI‐QTOF‐MS/MS

Acorus tatarinowii Schott (ATS) is a well‐known traditional Chinese medicine (TCM) for the treatment of epilepsy, amnesia and insomnia. In this study, a methodology utilizing high‐performance liquid chromatography (HPLC) coupled with electrospray ionization quadrupole time‐of‐flight mass spectrometry (ESI‐QTOF‐MS/MS) was established for the separation and structural identification of the major chemical constituents in ATS for the first time. Overall, 46 major constituents including flavonoid glycosides, phenylpropane derivatives, amides and lignans were identified or tentatively characterized. Seven major constituents, including four phenylpropane derivatives and three lignans, were further quantified as marker substances, which showed good linearity within the test ranges. These results indicated that the developed quantitative method was linear, sensitive, and precise for quality control of ATS. Copyright © 2014 John Wiley & Sons, Ltd.     

Structural analysis of glycosphingolipid analogues obtained by the saccharide primer method using CE‐ESI‐MS

A glycosphingolipid analogue (12‐azidododecyl β‐lactoside) as a saccharide primer has been shown to be useful for the synthesis of oligosaccharide libraries by mammalian cells. In the present study, CE‐ESI‐MS was employed to elucidate the structure of glycosphingolipid analogues derived from 12‐azidododecyl β‐lactoside (Lac‐C12N3) by mammalian cells. MDCK cells and COLO201 cells were cultured with Lac‐C12N3, and the glycosylated products secreted into the medium were collected and separated into acidic and neutral products by column chromatography. The acidic products could be directly analyzed by CE‐ESI‐MS, while the neutral products were converted to anionic derivatives via a reaction with propiolic acid. With this method, it was possible to analyze both acidic and neutral products glycosylated by MDCK cells and COLO201 cells at high sensitivity.     

Analysis of the chemical components of Qixianqingming granules and their metabolites in rats by UPLC‐ESI‐Q‐TOF‐MS

Qixianqingming granules (QXQM) comprise a traditional Chinese medicine (TCM) formula that was developed based on the combination of TCM theory and clinical practice. This formula has been proven to effectively treat asthma. In this study, an analytical procedure using ultraperformance liquid chromatography, coupled with electrospray ionization quadrupole time‐of‐flight mass spectrometry, was established for the rapid separation and sensitive identification of the chemical components in QXQM and its metabolites in serum of rats. Seventy‐two compounds were systematically identified in QXQM, including flavonoids, terpenoids, anthraquinones, phenylethanoid glycosides, stilbenes, alkaloids, and organic acids. Thirteen prototype compounds and 29 metabolites were detected in the serum of rats. The results provided fundamental information for further studying the mechanisms and clinical application of QXQM.     

Chemical UPLC‐ESI‐MS/MS profiling of aconitum alkaloids and their metabolites in rat plasma and urine after oral administration of Aconitum carmichaelii Debx. Root extract

In this paper, an ultra high performance liquid chromatography tandem mass spectrometric (UPLC‐ESI‐MS/MS) method in positive ion mode was established to systematically identify and to compare the major aconitum alkaloids and their metabolites in rat plasma and urine after oral administration of Fuzi extract. A total twenty‐nine components including twenty‐five C19‐diterpenoid alkaloids and four C20‐diterpenoid alkaloids were identified in Fuzi extract. Thirteen of the parent components and five metabolites were detected in rat plasma and sixteen parent compounds and six metabolites in urine. These parent components found in rat plasma and urine were mainly C19‐diterpenoid alkaloids. All of the metabolites in vivo were demethylated metabolites (phase I metabolites), which suggested that demethylation was the major metabolic pathway of aconitum alkaloids in vivo. A comparison of the parent components in rat plasma and urine revealed that 3‐deoxyacontine was found in plasma but not in urine, while kalacolidine, senbusine and 16‐β‐hydroxycardiopetaline existed in urine but not in plasma, which indicated that most alkaloids components were disposed and excreted in prototype form. This research provides some important information for further metabolic investigations of Fuzi in vivo.     

Method for rapidly discovering active components in Yupingfeng granules by UPLC‐ESI‐Q‐TOF‐MS

Yupingfeng granules (YPFG) were isolated from a traditional Chinese medicine (TCM) formulation composed of three herbs (Astragali Radix, Atractylodis Macrocephalae Rhizoma, and Saposhnikoviae Radix). This formulation is used in TCM to tonify qi, and it can help strengthen exterior and reduce sweating. Nevertheless, the active components of YPFG remain unclear. In this study, the chemical constituents of YPFG were systematically characterized by ultra‐performance liquid chromatography coupled with electrospray ionization/ quadrupole time‐of‐flight mass spectrometry (UPLC‐ESI‐Q‐TOF‐MS). Fifty‐eight compounds, namely, 20 flavonoids, 19 saponins, nine organic acids, four volatile coumarins, three lactones, one alkaloid, and two other components, were identified. In addition, the constituents of YPFG with the potential for in vivo bioactivities following oral administration were investigated in Sprague–Dawley rats. Thirteen compounds, namely, 11 flavonoid‐related and 2 saponin‐related components, were detected in rat plasma. After enriching flavonoids and saponins in YPFG by extraction, the extracts and YPFG were administrated to immunosuppressed rats, respectively. Plasma samples were analyzed by UPLC‐ESI‐Q‐TOF‐MS, and principal component analysis (PCA) confirmed that the extracts had similar effects to YPFG. This method could discover active ingredients in YPFG quickly and provide a scientific basis for quality control and mechanism research.     

Characterization of trace embedded impurities in thin multilayer structures using synchrotron X‐ray standing waves

X‐ray standing wave (XSW) field generated under Bragg reflection condition in a periodic Mo/Si multilayer structure has been used to determine the concentration and location of various trace element contaminants embedded in different layers of that multilayer structure. We have used intense synchrotron X rays for XSW analysis. It is observed that various trace element impurities such as Cr, Fe, Ni and W get embedded unintentionally in the multilayer structure during the deposition process. Consequences of such impurity incorporation on the optical properties of the multilayer structure are discussed in hard and soft X‐ray regions. Present measurements are important in order to optimize the deposition methods on one hand and to better correlate the measured optical properties of a multilayer structure with theoretical models on the other. Copyright © 2010 John Wiley & Sons, Ltd.     

A quantitative analysis of the polyamine in lung cancer patient fingernails by LC‐ESI‐MS/MS

A quantitative analysis of polyamines in lung cancer patient fingernails by the combination of 4‐(N,N‐dimethylaminosulfonyl)‐7‐fluoro‐2,1,3‐benzoxadiazole derivatives and liquid chromatography–electrospray ionization tandem mass spectrometry is described. The reaction of the reagent with eight kinds of polyamines, that is, N¹‐acetylputrescine (N¹‐actPUT), N⁸‐acetylspermidine, N¹‐acetylspermine, 1,3‐diaminopropane, putrescine (PUT), cadaverine, spermidine and spermine (SPM) effectively occurs at 60 °C for 30 min. The detection limits (signal‐to‐noise ratio 5) were 5–100 fmol. A good linearity was achieved from the calibration curves, which was obtained by plotting the peak area ratios of the analytes relative to the internal standard (IS), that is, 1,6‐diaminohexane, vs the injected amounts of polyamines (r² > 0.996), and the intra‐day and inter‐day assay precisions were <9.84%. Furthermore, the recoveries (%) of the polyamines spiked in the human fingernails were 89.14–110.64. The present method was applied to human fingernail samples from 17 lung cancer patients and 39 healthy volunteers. The polyamine concentration was different based on the gender, that is, the N¹‐actPUT and PUT contents were 3.10 times and 2.56 times higher in healthy men than in women, respectively. Additionally, in the lung cancer patient group, as compared with the healthy volunteers, the concentrations of SPM had a statistically significant (p < 0.05) correlation. Therefore, because the proposed method provides a good mass accuracy and the trace detection of the polyamines in human fingernails, this analytical technique could be a noninvasive technique to assist in the diagnosis and assessment of disease activity in lung cancer patients. Copyright © 2013 John Wiley & Sons, Ltd.     

Interlaboratory study comparing analyses of simulated angle‐resolved X‐ray photoelectron spectroscopy data

An interlaboratory study has been conducted to determine the following: (i) the similarities and differences of film thicknesses and composition profiles obtained from analyses of simulated angle‐resolved X‐ray photoelectron spectroscopy (ARXPS) data by different analysts using different algorithms for data analysis, and (ii) the effects of two assumptions commonly made in data‐analysis algorithms for ARXPS on derived film thicknesses and composition profiles. The analyzed data were generated by the National Institute of Standards and Technology Database for the Simulation of Electron Spectra for Surface Analysis, (SESSA) which provides a simple way to study the influence of the aforementioned effects on compositional depth profile reconstruction. Sets of simulated ARXPS data were produced for thin films of SiO₂, SiON, HfO₂, and HfON of varying thicknesses on a Si substrate. For some HfON films, the N concentration varied with depth. Eleven groups participated in the round robin study. The majority (eight) employed a commercial ARXPS instrument in which the angular distribution is measured for a fixed sample geometry, in contrast to conventional ARXPS in which the sample is tilted for angular variation. The average deviations between the reported average depth, film thickness, and amount of material typically varied between 20% and 30% but were considerably larger, between 30% and 80%, for some cases. The average errors were generally larger for simulations that included elastic scattering and the finite analyzer‐acceptance angle (realistic conditions) than those for simulations that neglected elastic scattering and the finite analyzer‐acceptance angle (simplified conditions). The retrieved N depth profiles were quantitatively different from the true depth profiles and showed substantial variability among the group of members who used the same instrument and analysis software. Copyright © 2014 John Wiley & Sons, Ltd.     

Simultaneous determination of ten flavonoids of crude and wine‐processed Radix Scutellariae aqueous extracts in rat plasma by UPLC‐ESI‐MS/MS and its application to a comparative pharmacokinetic study

Radix Scutellariae (RS) is a herbal medicine with various pharmacological activities to treat inflammation, respiratory and gastrointestinal infections, etc. In this study, a rapid, sensitive and selective UPLC‐ESI‐MS/MS method was developed for simultaneous determination of 10 flavonoids – scutellarin, scutellarein, chrysin, wogonin, baicalein, apigenin, wogonoside, oroxylin A‐7‐O‐glucuronide, oroxylin A and baicalin – from RS aqueous extracts in rat plasma with propyl paraben as internal standard (IS). Chromatographic separation was achieved on a C₁₈ column using gradient elution with the mobile phase consisting of methanol and water (containing 0.1% formic acid) at a flow rate of 0.2 mL/min. The detection was performed in multiple reaction monitoring mode using electrospray ionization in negative mode. The validated method showed good linearity over a wide concentration range (r >0.9935). The intra‐ and interday assay variabilities were <9.5% and <12.4% for all analytes, respectively. The extraction recovery ranged from 71.2 to 89.7% for each analyte and IS. This method was successfully applied to pharmacokinetic comparision after oral administration of crude and wine‐processed RS aqueous extracts. There were significant differences in some pharmacokinetic parameters of most analytes between crude and wine‐processed RS. This suggested that wine‐processing exerted effects absorption of most flavonoids. Copyright © 2014 John Wiley & Sons, Ltd.     

Identification of ilaprazole metabolites in human urine by HPLC‐ESI‐MS/MS and HPLC‐NMR experiments

Ilaprazole is a new proton pump inhibitor designed for the treatment of gastric ulcers, and limited data is available on the metabolism of the drug. In this article, the structural elucidation of urinary metabolites of ilaprazole in human was described by HPLC‐ESI‐MS/MS and stopped‐flow HPLC‐NMR experiments. Urinary samples were precipitated by sodium carbonate solution, and then extracted by liquid–liquid extraction after adding ammonium acetate buffer solution. The enriched sample was separated using a C₁₈ reversed‐phase column with the mobile phase composed of acetonitrile and 0.05 mol/L ammonium acetate buffer solution in a gradient solution, and then directly coupled to ESI‐MS/MS detection in an on‐line mode or ¹H‐NMR (500 MHz) spectroscopic detection in a stopped‐flow mode. As a result, four sulfide metabolites, ilaprazole sulfide (M1), 12‐hydroxy‐ilaprazole sulfide (M2), 11,12‐dihydroxy‐ilaprazole sulfide (M3) and ilaprazole sulfide A (M4), were identified by comparing their MS/MS and NMR data with those of the parent drug and available standard compounds. The main biotransformation reactions of ilaprazole were reduction and the aromatic hydroxylation of the parent drug and its relative metabolites. The result testified that HPLC‐ESI‐MS/MS and HPLC‐NMR could be widely applied in detection and identification of novel metabolites. Copyright © 2010 John Wiley & Sons, Ltd.     

Simultaneous determination of eight components in Angelica sinensis based on UHPLC‐ESI‐MS/MS method for quality evaluation

A method employing ultra‐high performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) for determination of eight components including ferulic acid, senkyunolide A, butylphthalide, ligustilide, butylidenephalide, senkyunolide I, senkyunolide H and levistolide A in Angelica sinensis was established. The separation was carried out using a Waters ACQUITY UHPLC BEH C₁₈ column with gradient elution with 0.1% formic acid aqueous and acetonitrile at a flow rate of 0.4 mL/min. Good linearity was attained with R² of 0.9983–0.9998 in wide concentration ranges. The method had limit of detection (LOD) and quantitation (LOQ) in the range of 0.42–6.98 ng/mL and 1.39–23.28 ng/mL, respectively. Intra‐ and inter‐day precisions varied with relative standard deviations (RSDs) from 0.33% to 0.88% and 0.37% to 1.04%, respectively. Moreover, the average recoveries were in a satisfactory range of 92.7%–102.1% with RSDs of less than 3.60%. Finally, the method was successfully applied to analyze 19 batches of A. sinensis samples grown in Min County, Gansu province, China, as well as that collected in other regions. The findings indicated that the established method is reliable and may thus be applied as a powerful tool for qualitative and quantitative analysis of components in A. sinensis, which has its implications in quality control of A. sinensis.     

Study of the retention behavior of iodinated X‐ray contrast agents in hydrophilic interaction liquid chromatography,comparing bare silica and zwitterionic stationary phases

Iodinated X‐ray contrast media are the most widely used pharmaceuticals for intravascular administration in X‐ray diagnostic procedures. The increasing concern of the fate of these compounds into the environment has led to the development of analytical methods to determine them. However, these methods present problems due to the polar character of these analytes. In this paper, hydrophilic interaction LC is presented as an alternative technique. The retention of iodinated X‐ray contrast media was studied in two bare silica phases with different particle designs (i.e. porous and Fused Core?) and a zwitterionic sulfoalkylbetaine phase. The effect of the most important parameters of the mobile phase was studied for each stationary phase. It was observed that optimal mobile phase conditions included buffers with a high buffering capacity. Additionally, the retention mechanisms involved were studied in order to provide some insight into the possible occurring interactions. The contributions of partition and adsorption and the effect of the temperature on the retention of analytes were evaluated on all of the stationary phases.     

Screening anti‐allergic components of Astragali Radix using LAD2 cell membrane chromatography coupled online with UHPLC‐ESI‐MS/MS method

Huangqi (Astragali Radix), a traditional Chinese herb, is widely used in clinical therapy in China. In addition, an anti‐allergic effect of constituents in Huangqi has been reported in the scientific literature. In the present study, cell membrane chromatography coupled online with UHPLC‐ESI‐MS/MS method was developed to screen, analyze and identify the anti‐allergic components of Huangqi. The Laboratory of Allergic Disease 2 (LAD2) cell was used to establish cell membrane chromatography, which was combined with UHPLC‐ESI‐MS/MS. The coupled system was then used to screen anti‐allergic components from Huangqi. Effects of active components were verified by histamine release assay. A component retained on the LAD2 cell membrane chromatography was identified as formononetin. Bioactivity of formononetin was investigated by histamine release assay in LAD2 cells, and it was found that formononetin could inhibit histamine release in a dose‐dependent manner from 1 to 100 μm . The LAD2 cell membrane chromatography online with UHPLC‐ESI‐MS/MS method is an effective technique for screening the anti‐allergic components of Huangqi.     

Chemical composition differentiation of Shen‐Shuai‐Ning granule between combined decoction and separated decoction using HPLC‐DAD‐ESI‐QTOF‐MS

Shen‐Shuai‐Ning (SSN) granule, a traditional Chinese medicine formula, is widely used in clinical practice for treating chronic renal failure. However, its detailed chemical profile is unknown. Here, HPLC‐ESI‐QTOF‐MS was employed for the systematic chemical analysis of SSN. A total of 52 compounds were identified and the characteristic ions of the compounds were described. Furthermore, chemical consistency between the combined decoction and the separated decoction of SSN was evaluated using HPLC‐DAD. A chemical comparison between two preparations of SSN granule (combined decoction and separated decoction of Coptides Rhizoma) indicated a significant difference in the content of many compounds, including salvianolic acid A, salvianolic acid B, berberine, palmatine and epiberberine. As a result, separated decoction of Coptides Rhizoma would lead to a significantly decrease in depsides in Salviae Miltiorrhizae Radix et Rhizoma and an increase in alkaloids in Coptidis Rhizoma.     

Fragmentation pathways of metopimazine and its metabolite using ESI‐MSn,HR‐MS and H/D exchange

Metopimazine (MPZ) is a phenothiazine derivative used to prevent emesis during chemotherapy where few structural analysis of the aforementioned compound have been described in the literature. Thus, this work reports, for the first time, the detailed study of fragmentation pathways of MPZ and its metabolite (AMPZ) using electrospray ionization (EI) with multistage mass spectrometry (ESI‐MSⁿ) in positive‐ion mode. The structures of 21 product ions were identified and their accurate masses were determined using high resolution mass spectrometry (HRMS) experiments. Characteristic product ions of these two phenothiazine derivatives are more particularly displayed along with differences between their relative abundances and their structures checked by H/D exchange experiments. Copyright © 2010 John Wiley & Sons, Ltd.     

Determination of spiramycin and neospiramycin antibiotic residues in raw milk using LC/ESI‐MS/MS and solid‐phase extraction

A simple confirmatory method for the determination of spiramycin and its metabolite neospiramycin in raw milk using LC ESI MS/MS is presented. Macrolide residues in raw milk were extracted by ACN, and sample extracts were further cleaned up and concentrated using SPE cartridges. Both spiramycin and neospiramycin were protonated in electrospray positive ion mode to form singly and/or doubly charged pseudomolecular ions. Data acquisition was achieved using multiple reaction monitoring, i.e., two transitions, for quantification and confirmation. Matrix‐matched standard calibration curves were utilized to achieve the best accuracy for the method. The method performance was evaluated according to both a conventional validation procedure and a designed experimental result. The measurement uncertainty arising from accuracy and precision was estimated. The method accuracy, expressed as a percentage of an overall recovery, was from 82.1 to 108.8%, and its intermediate precision was less than 20%. LC/ESI‐MS/MS method LODs (S/N ? 3:1) of spiramycin and neospiramycin were less than 1.0 μg/kg.