Liquid Chromatography-ion Trap Mass Spectrometry Research Articles

Linking biogenic hydrocarbons to biogenic aerosol in the Borneo rainforest

Abstract. Emissions of biogenic volatile organic compounds are though to contribute significantly to secondary organic aerosol formation in the tropics, but understanding these transformation processes has proved difficult, due to the complexity of the chemistry involved and very low concentrations. Aerosols from above a Southeast Asian tropical rainforest in Borneo were characterised using liquid chromatography–ion trap mass spectrometry, high-resolution aerosol mass spectrometry and Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) techniques. Oxygenated compounds were identified in ambient organic aerosol that could be directly traced back to isoprene, monoterpenes and sesquiterpene emissions, by combining field data on chemical structures with mass spectral data generated from synthetically produced products created in a simulation chamber. Eighteen oxygenated species of biogenic origin were identified in the rainforest aerosol from the precursors isoprene, α-pinene, limonene, α-terpinene and β-caryophyllene. The observations provide the unambiguous field detection of monoterpene and sesquiterpene oxidation products in SOA above a pristine tropical rainforest. The presence of 2-methyl tetrol organosulfates and an associated sulfated dimer provides direct evidence that isoprene in the presence of sulfate aerosol can make a contribution to biogenic organic aerosol above tropical forests. High-resolution mass spectrometry indicates that sulfur can also be incorporated into oxidation products arising from monoterpene precursors in tropical aerosol.

The in vivo and in vitro metabolism and the detectability in urine of 3’,4’‐methylenedioxy‐alpha‐pyrrolidinobutyrophenone (MDPBP), a new pyrrolidinophenone‐type designer drug, studied by GC‐MS and LC‐MSn

3',4'-Methylenedioxy-alpha-pyrrolidinobutyrophenone (MDPBP), a designer drug of the pyrrolidinophenone-type, was first seized in Germany in 2009. It was also identified in 'legal high' samples investigated in the UK. Therefore, the aim of the presented work was to identify its in vivo and in vitro phase I and II metabolites using gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-ion trap mass spectrometry (LC-MS(n) ). Furthermore, detectability of MDPBP in rat and human urine using standard urine screening approaches (SUSA) by GC-MS and LC-MS(n) was studied. The metabolites were isolated either directly or after enzymatic cleavage of conjugates by solid-phase extraction (C18, HCX). The metabolites were then analyzed and structures proposed after GC-MS (phase I) and LC-MS(n) (phase II). Based on these identified metabolites, the following main metabolic steps could be proposed: demethylenation followed by methylation of one hydroxy group, aromatic and side chain hydroxylation, oxidation of the pyrrolidine ring to the corresponding lactam as well as ring opening to the corresponding carboxylic acid. Furthermore, in rat urine after a typical user's dose as well as in human urine, mainly the metabolites could be detected using the authors' SUSA by GC-MS and LC-MS(n) . Thus, it should be possible to monitor an application of MDPBP assuming similar toxicokinetics in humans. Finally, CYP2C19 and CYP2D6 could be identified as the isoenzymes mainly responsible for demethylenation.

A preliminary study of arecoline and guvacoline presence in the saliva of a "betel-quid" chewer using liquid-chromatography ion trap mass spectrometry.

It has been reported that the parasympathomimetic alkaloid arecoline and the nootropic agent guvacoline have been detected in areca nut (Areca catechu L.) during extraction using a basic medium. Here, we have studied the detection of arecoline and guvacoline in vivo in saliva of a "betel-quid" chewer using liquid-chromatography ion trap mass spectrometry. In this paper, we provide evidence that guvacoline is absent in the neutral aqueous extract of betel nut, but is present in abundance in the aqueous extract with added time (pH 11.9). In an in vivo experiment, we demonstrated that guvacoline is present in the salivary extracts in the mouth with time (pH 9.5) and without lime (pH5.3).

Enantioselective extraction of (+)-(S)-citalopram and its main metabolites using a tailor-made stir bar chiral imprinted polymer for their LC-ESI-MS/MS quantitation in urine samples

This paper reports the application of a chiral imprinted polymer (CIP)-coated stir bar for the selective extraction of (+)-(S)-citalopram (SCIT) and its main metabolites, (+)-(S)-desmethylcitalopram (SDCIT) and (+)-(S)-didesmethylcitalopram (SDDCIT), from urine samples. The developed device has been demonstrated to be capable of selectively extracting the three target analytes from urine samples without saturating the imprinted sites. A CIP-coated stir bar sorptive extraction procedure (CIP-SBSE) is proposed for the isolation of SCIT, SDCIT and SDDCIT followed by their subsequent analysis using liquid chromatography ion trap mass spectrometry (LC-ITMS). Deuterated SCIT-d6 was used as an internal standard. The method was validated using a standard procedure, which revealed that a quantification of 5ngmL−1 was obtained in urine samples and that the accuracy and precision were within the established values while no matrix effect was observed.

Steady turnover of amyloid fibril proteins in gastric mucosa after liver transplantation in familial amyloid polyneuropathy

Introduction: Our previous study demonstrated marked regression of amyloid deposits in abdominal fat tissues of familial amyloid polyneuropathy (FAP) patients treated with liver transplantation (LT). To determine whether similar changes in deposited amyloid can also occur in other organs, we examined gastric mucosal amyloid before and after LT in FAP patients with ATTRV30M.Methods: We histopathologically and biochemically investigated gastric mucosal amyloid before and after LT in six FAP patients with ATTRV30M.The amounts of amyloid deposits in biopsied gastric mucosa were determined by microscopy, and the proportion of wild-type transthyretin (TTR) in extracted amyloid fibril proteins was assayed by liquid chromatography-ion trap mass spectrometry. Similar examinations were also performed in 21 untreated FAP patients and 13 transplanted patients with ATTRV30M.Results: The amount of deposited amyloid was not markedly different before and after LT in six patients. However, the composition ratios of wild-type TTR in gastric mucosal amyloid increased markedly from 20.0% ± 11.4% before LT to 43.2% ± 13.8% after LT. In addition, the ratio of wild-type TTR in all transplanted patients was significantly higher than that in untransplanted patients (72.7% ± 25.5%, 23.8% ± 14.3%, respectively).Conclusions: Our results showed that the components of amyloid fibril proteins in gastric mucosa of transplanted FAP patients are different from those in untreated patients: a significant portion of preexisting ATTRV30M-derived amyloid seemed to be replaced by wild-type TTR-derived amyloid postoperatively, indicating that continuous turnover of amyloid deposits can occur in all organs in transplanted FAP patients. It was also confirmed that wild-type TTR plays an important role in the pathogenesis of postoperative amyloid deposition in transplanted FAP patients.

Characterization and quantification of 10‐hydroxycamptothecine in Camptotheca acuminate and its medicinal preparation by liquid chromatography–ion trap mass spectrometry

A rapid and sensitive method for the identification and quantification of 10-hydroxycamptothecine (HCPT) in Camptotheca acuminata Decne is described. The HCPT standard solution was directly infused into the ion trap mass spectrometers (IT/MS) for collecting the MS(n) spectra. The electrospray ionization (ESI) mass spectral fragmentation pathway of HCPT was proposed and the ESI-MS(n) fragmentation behavior of HCPT was deduced in detail. The major fragment ions of HCPT were confirmed by MS(n) in both negative ion and positive ion mode. The possible main cleavage pathway of fragment ions was studied. Quantification of HCPT was assigned in negative-ion mode at a product ion at m/z 363 → 319 by LC-MS. The LC-MS method was validated for linearity, sensitivity, accuracy and precision, and then used to determine the content of the HCPT. Lastly, the LC-MS method was successfully applied to determine HCPT in real samples of Camptotheca acuminate Decne and its medicinal preparation in the first time.

Liquid Chromatography-Mass Spectrometry-Based Chemotaxonomic Classification of Aspergillus spp. and Evaluation of the Biological Activity of Its Unique Metabolite, Neosartorin

This work aimed to classify Aspergillus (8 species, 28 strains) by using a secondary metabolite profile-based chemotaxonomic classification technique. Secondary metabolites were analyzed by liquid chromatography ion-trap mass spectrometry (LC-IT-MS) and multivariate statistical analysis. Most strains were generally well separated from each section. A. lentulus was discriminated from the other seven species (A. fumigatus, A. fennelliae, A. niger, A. kawachii, A. flavus, A. oryzae, and A. sojae) with partial least-squares discriminate analysis (PLS-DA) with five discriminate metabolites, including 4,6-dihydroxymellein, fumigatin, 5,8-dihydroxy-9- octadecenoic acid, cyclopiazonic acid, and neosartorin. Among them, neosartorin was identified as an A. lentulus-specific compound that showed anticancer activity, as well as antibacterial effects on Staphylococcus epidermidis. This study showed that metabolite-based chemotaxonomic classification is an effective tool for the classification of Aspergillus spp. with species-specific activity.

Ultra-high-performance liquid chromatography–ion trap mass spectrometry characterisation of milk polar lipids from dairy cows fed different diets

Milk polar lipids are an important class of biologically active species for human health and for improving the physical functionality of food ingredients. Milk polar lipids from 144 multiparous Holstein–Friesian dairy cows fed different diets were analysed using ultra-high-performance liquid chromatography–ion trap mass spectrometry (UHPLC–MSn). A complex profile of polar lipids, consisting of 7 species of phosphatidylinositol (PI), 12 species of phosphatidylethanolamine (PE), 18 species of phosphatidylcholine (PC) and 13 species of sphingomyelin (SM) were identified from the molecular ions and sequential MSn fragmentation. Qualitative assessment of the data suggested that different cow diets influenced the relative amounts of a small number of species in the milk samples, e.g. PE 14:0/18:1, PE 18:0/18:1, PC 15:0/18:1, PC 18:0/18:1, SM d18:1/14:0, SM d18:1/15:0, SM d18:1/22:0 and SM d18:1/23:0.

Analysis of impurities in vertilmicin sulfate by liquid chromatography ion-trap mass spectrometry

The characterization of impurities present in vertilmicin by liquid chromatography (LC) coupled with mass spectrometry (MS) is described. A reversed phase (RP)-LC method using a C18 column resistant to basic pH with an alkaline (pH 11) aqueous mobile phase was developed and coupled to MS with an electrospray ionization (ESI) source in the positive ion mode which provides MSn capability. In total, 18 impurities were detected in a commercial sample. Eleven impurities described in this work were newly identified.

Identification and quantification of active alkaloids in Catharanthus roseus by liquid chromatography–ion trap mass spectrometry

Catharanthus roseus is an important dicotyledonous medicinal plant that produces anticancer compounds. The active alkaloids vinblastine, vindoline, ajmalicine, catharanthine, and vinleurosine were identified by direct-injection ion trap-mass spectrometry (IT-MS) for collecting MS1–2 spectra. The determinations of five alkaloids were accomplished by liquid chromatography (LC) with UV and MS detections. The analytes provided good signals corresponding to the protonated molecular ions [M+H]+ and product ions. The precursor ions and product ions for quantification of vinblastine, vindoline, ajmalicine, catharanthine, and vinleurosine were m/z 825→807, 457→397, 353→144, 337→144 and 809→748 by LC–IT-MS, respectively. Two methods were used to evaluate a number of validation characteristics (repeatability, LOD, calibration range, and recovery). MS provided a high selectivity and sensitivity for determination of five alkaloids in positive mode. After optimisation of the methods, separation, identification and quantification of the five components in C. roseus were comprehensively accomplished by HPLC with UV and MS detection.

Fluorescein-core piperazine as a fluorescent labeling reagent for highly sensitive determination of jasmonic acid by high-performance liquid chromatography

In the present work, a pre-column fluorescent derivatization reagent, fluorescein-core piperazine (FCP) was prepared for labeling jasmonic acid (JA). Based on the enhanced fluorescence after derivatization, a simple and sensitive method was developed for determination of JA by high performance liquid chromatography with fluorescence detection (HPLC/FD). Several key derivatization conditions were examined. The results indicated that FCP labeled JA could be well separated from FCP in the sample within 13 min on a C18 reversed-phase column using a mobile phase of methanol–water (55 : 45, v/v) with fluorescence detection at excitation and emission wavelength of 507 nm and 526 nm, respectively. The detection limit obtained was 0.4 nM (signal to noise ratio of 3), which was lower than most of the existing methods. The method is reproducible and yields single peak for JA regardless of isomer. The specificity of the proposed method was confirmed by high-performance liquid chromatography-ion trap mass spectrometry (HPLC/IT-MS). Finally, it has been demonstrated that the proposed HPLC/FD method can be applied to the assay of JA in plant samples.

Generic detection of basic taxoids in wood of European Yew (Taxus baccata) by liquid chromatography–ion trap mass spectrometry

The occurrence of the cardiotoxin taxine (comprising taxine B and several other basic taxoids) in leaves of Taxus baccata L. (European yew) is well known and has led to public concerns about the safety of eating or drinking from utensils crafted from the wood of this poisonous species. The occurrence of basic taxoids in the heartwood of T. baccata had not been examined in detail, although the bark is known to contain 2'β-deacetoxyaustrospicatine. Initial examination of heartwood extracts for 2'β-deacetoxyaustrospicatine by liquid chromatography-mass spectrometry (LC-MS) revealed the presence of this basic taxoid at about 0.0007% dry weight, using a standard isolated from bark. Analyses for taxine B, however, proved negative at the extract concentration analysed. Observing other basic taxoids within the heartwood extracts was facilitated by developing generic LC-MS methods that utilised a fragment arising from the N-containing acyl group of basic taxoids as a reporter ion. Of the various MS strategies available on a hybrid ion trap-orbitrap instrument that allowed observation of this reporter ion, combining all-ion collisions with high resolution ion filtering by the orbitrap was most effective, both in terms of the number of basic taxoids detected and sensitivity. Numerous basic taxoids, in addition to 2'β-deacetoxyaustrospicatine, were revealed by this method in heartwood extracts of T. baccata. Red wine readily extracted the basic taxoids from heartwood while coffee extracted them less efficiently. Contamination with basic taxoids could also be detected in soft cheese that had been spread onto wood. The generic LC-MS method for detecting basic taxoids complements specific methods for detecting taxine B when investigating yew poisoning cases in which the analysis of complex extracts may be required or taxine B has not been detected.

Impurity profiling of micronomicin sulfate injection by liquid chromatography–ion trap mass spectrometry

The characterization of impurities present in micronomicin sulfate injection by liquid chromatography (LC) coupled with mass spectrometry (MS) is described. A reversed phase (RP)-LC method using a C18 column resistant to an alkaline (pH 11) aqueous mobile phase was developed and coupled to MS with an electrospray ionization (ESI) source in the positive ion mode which provides MSn capability. A total of thirty six impurities were detected in commercial samples: five impurities were identified by comparison of their fragmentation patterns with those of available related substances, eleven of them were identified in accordance with relevant literature, while the other twenty impurities were newly identified using the MS/MS spectra of the available related reference substances as interpretative templates combined with knowledge of the nature of functional group fragmentation behaviors. This work was applied to evaluate the quality of micronomicin sulfate injection from different manufacturers.

Identification and quantification of atractylenolide I and atractylenolide III in Rhizoma Atractylodes Macrocephala by liquid chromatography–ion trap mass spectrometry

Rhizoma Atractylodes Macrocephala (RAM) is an important traditional Chinese medicinal herb that is used for treatment of dyspepsia and anorexia. The active ingredients, atractylenolide I (AO-I) and atractylenolide III (AO-III), were identified by direct-injection ion trap-mass spectrometry (IT-MS) for collecting MS(n) spectra. The major fragment ions of AO-I and AO-III were confirmed by MS(n) both in negative ion mode and in positive ion mode. The possible main cleavage pathway of fragment ions was studied. The determinations of AO-I and AO-III were accomplished by liquid chromatography (LC) with UV and MS. The analytes provided good signals corresponding to the protonated molecular ions [M + H](+) and product ions. The precursor ions and product ions for quantification of AO-III and AO-I were m/z 249 → 231 and m/z 233 → 215, respectively, using selected ion monitoring by LC-IT-MS. Two methods were evaluated for a number of validation characteristics (repeatability, limit of detection, calibration range, and recovery). MS provides a high selectivity and sensitivity for determination of AO-III and AO-I in positive mode. After optimization of the methods, separation, identification and quantification of the two components in RAM were comprehensively tested by HPLC with UV and MS.

System Response of Metabolic Networks in Chlamydomonas reinhardtii to Total Available Ammonium

Drastic alterations in macronutrients are known to cause large changes in biochemistry and gene expression in the photosynthetic alga Chlamydomonas reinhardtii. However, metabolomic and proteomic responses to subtle reductions in macronutrients have not yet been studied. When ammonium levels were reduced by 25-100% compared with control cultures, ammonium uptake and growth rates were not affected at 25% or 50% nitrogen-reduction for 28 h. However, primary metabolism and enzyme expression showed remarkable changes at acute conditions (4 h and 10 h after ammonium reduction) compared with chronic conditions (18 h and 28 h time points). Responses of 145 identified metabolites were quantified using gas chromatography-time of flight mass spectrometry; 495 proteins (including 187 enzymes) were monitored using liquid chromatography-ion trap mass spectrometry with label-free spectral counting. Stress response and carbon assimilation processes (Calvin cycle, acetate uptake and chlorophyll biosynthesis) were altered first, in addition to increase in enzyme contents for lipid biosynthesis and accumulation of short chain free fatty acids. Nitrogen/carbon balance metabolism was found changed only under chronic conditions, for example in the citric acid cycle and amino acid metabolism. Metabolism in Chlamydomonas readily responds to total available media nitrogen with temporal increases in short-chain free fatty acids and turnover of internal proteins, long before nitrogen resources are depleted.

Impurity profiling of etimicin sulfate by liquid chromatography ion-trap mass spectrometry

A reversed phase (RP)-LC method using a C(18) column resistant to basic pH and an alkaline (pH 10) aqueous mobile phase was developed and coupled to MS with an electrospray ionization (ESI) source in the positive ion mode which provides MS(n) capability. In total, 26 impurities were detected in a commercial sample. The structures of the impurities were proposed based on comparison of their fragmentation patterns with those of the available reference substances, the synthetic route and literature data. Starting material and its residual impurities, intermediates, synthetic by-products and degradation products were the main sources of those impurities. 14 impurities described in this work were newly identified.

Application of Matrix Solid-Phase Dispersion and Liquid Chromatography–Ion Trap Mass Spectrometry for the Analysis of Pesticide Residues in Fruits

This study presents an application of rapid and sensitive multiresidue method for the analysis of acephate, acetamipride, atrazine, carbendazim, carbaryl, carbofuran, dimethoate, imidacloprid, linuron, malathion, monocrotophos, monuron, propazine, simazine, and tebufenozide in fruits. The method involves an extraction procedure based on matrix solid-phase dispersion using diatomaceous earth as a dispersant and dichloromethane as the eluent. The target pesticides were determined using liquid chromatography–ion trap mass spectrometry. Quantification of the analytes was carried out using the most sensitive ion transition. Ion trap parameters, like activation q and time, were found to have a prominent influence on method sensitivity for some pesticides and they were optimized accordingly. The confirmation of residues detected in real samples was performed by repeated injection and acquiring additional ion transitions besides the ones used for quantification. The method was validated for accuracy, linearity, reproducibility, and sensitivity. Mean values for recoveries were in the range of 70–120 % for all tested matrices. Repeatability of the method, expressed as the relative standard deviation, was in general lower than 20 %. The applicability of the method to routine analysis was tested in real fruit samples with good performance.

In vitro metabolism of corydaline in human liver microsomes and hepatocytes using liquid chromatography‐ion trap mass spectrometry

Corydaline is a pharmacologically active isoquinoline alkaloid isolated from Corydalis tubers. It exhibits the antiacetylcholinesterase, antiallergic, antinociceptive, and gastric emptying activities. The purposes of this study were to establish in vitro metabolic pathways of corydaline in human liver microsomes and hepatocytes by identification of their metabolites using liquid chromatography-ion trap mass spectrometry. Human liver microsomal incubation of corydaline in the presence of an NADPH-generating system resulted in the formation of nine metabolites, namely, four O-desmethylcorydaline [M1 (yuanhunine), M2 (9-O-desmethylcorydaline), M3 (isocorybulbine), and M4 (corybulbine)], three di-O-desmethylcorydaline [M5 (9,10-di-O-desmethylcorydaline), M6 (2,10-di-O-desmethylcorydaline), and M7 (3,10-di-O-desmethylcorydaline)], M8 (hydroxyyuanhunine), and M9 (hydroxycorydaline). Incubation of corydaline in human hepatocytes produced four metabolites including M1, M5, M6, and M9. O-Demethylation and hydroxylation were the major metabolic pathways for the metabolism of corydaline in human liver microsomes and hepatocytes.

The simultaneous determination of vitamins A, E and β-carotene in bovine milk by high performance liquid chromatography–ion trap mass spectrometry (HPLC–MSn)

Bovine milk is considered to be a good dietary source of vitamins A, E and β-carotene. This paper describes a robust method for the simultaneous determination of these compounds using high performance liquid chromatography–ion trap mass spectrometry (HPLC–MSn) as the determinative step. The compounds were separated with a 5 μm Polaris 2.1 × 150 mm C18-A column and an aqueous-methanol mobile phase and the levels determined using the MS2 fragments (vitamin A, m/z 213 and m/z 199, vitamin E m/z 165 and β-carotene m/z 413). The HPLC–MSn was operated in the positive ion APCI mode. The method was validated using repeatability studies, duplicate analyses, recovery experiments, proficiency study data and comparison with previously validated high performance liquid chromatographic ultraviolet/visible (HPLC–UV/Vis) and fluorescence (HPLC–Fl) procedures used in our laboratory. Expanded measurement uncertainties were similar for HPLC–MSn, HPLC–UV/Vis and HPLC–Fl, e.g. vitamin A 45 μg/100 ml, HPLC–MSn ±8%, HPLC–UV ±9%; vitamin E 150 μg/100 ml, HPLC–MSn ±11%, HPLC–Fl ±9%; β-carotene 12 μg/100 ml, HPLC–MSn ±18%, HPLC–Vis ±21%.

Determination of Fluoroquinolones in Grass Carp by High Performance Liquid Chromatography-Ion Trap Mass Spectrometry Using Mixed-templates Imprinted Polymer Extraction

The molecularly imprinted polymer(MIP) has been prepared using levofloxacin and ciprofloxacin as the mixed-templates,α-methacrylic acid(MAA) as functional monomers and trimethylolpropane trimethylacrylate(TRIM) as crosslinker.A method has been established for simultaneous determination of residues of ten fluoroquinolones in grass carp by high performance liquid chromatography-ion trap mass spectrometry(HPLC-ITMS) combining with molecularly imprinted polymer solid phase extraction(MISPE).The samples were extracted with 2% acetic acid in acetonitrile and defatted with n-hexane,then purified by the self-made molecularly imprinted cartridge.The preparation of samples and the separation conditions had been optimized,HPLC was performed with a gradient elution program while acetonitrile and water(containing 0.05% formic acid) were used as mobile phase.The identification and quantification were done with ion trap mass spectrometry.The recove-ries ranged from 80.6% to 104.6% with a relative standard deviations of 8.6%.The limits of detection(LOD) were 0.11-0.25 μg/kg and the limits of quantification(LOQ) were 0.35-0.84 μg/kg.This method is suitable for validation and multi-residue analysis of fluoroquolones in aquatic products.