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Rationale: Paper spray offers a rapid screening test without the need for sample preparation. The incomplete extraction of paper spray allows for further testing using more robust, selective and sensitive techniques such as liquid chromatography/mass spectrometry (LC/MS). Here we develop a two‐step process of paper spray followed by LC/MS to (1) rapidly screen a large number of samples and (2) confirm any disputed results. This demonstrates the applicability for testing medication adherence from a fingerprint. Methods: Following paper spray analysis, drugs of abuse samples were analysed using LC/MS. All analyses were completed using a Q Exactive™ Plus Orbitrap™ mass spectrometer. This two‐step procedure was applied to fingerprints collected from patients on a maintained dose of the antipsychotic drug quetiapine. Results: The extraction efficiency of paper spray for two drugs of abuse and metabolites was found to be between 15 and 35% (analyte dependent). For short acquisition times, the extraction efficiency was found to vary between replicates by less than 30%, enabling subsequent analysis by LC/MS. This two‐step process was then applied to fingerprints collected from two patients taking the antipsychotic drug quetiapine, which demonstrates how a negative screening result from paper spray can be resolved using LC/MS. Conclusions: We have shown for the first time the sequential analysis of the same sample using paper spray and LC/MS, as well as the detection of an antipsychotic drug from a fingerprint. We propose that this workflow may also be applied to any type of sample compatible with paper spray, and will be especially convenient where only one sample is available for analysis. [ABSTRACT FROM AUTHOR]

Complete heparin digestion with heparin lyase I and II results in a mixture of hexasaccharides and tetrasaccharides with 3‐O‐sulfo group‐containing glucosamine residues at their reducing ends. Because these tetrasaccharides are derived from antithrombin III‐binding sites of heparin, we examined whether this method could be applied to estimate the anticoagulant activity of heparin. Therefore, this paper presents a new low molecular weight heparin sample preparation method–chemical depolymerization. Qualitative analysis of the studied compounds and a comparison of their composition are an important contribution to the structural analysis of low molecular weight heparins, which has not been fully conducted so far. Qualitative on‐line liquid chromatography–mass spectrometric analysis of these resistant oligosaccharides is also described in this paper. [ABSTRACT FROM AUTHOR]

Highlights • A method for the analysis of typical HMOs was validated. • Neutral and acidic HMOs are concomitantly quantified in the same analysis. • The method was applied to analyze HMOs in real human milk samples. • Accurate and reliable quantification of HMOs was achieved. • The method allows the Lewis and Secretor phenotype of the mother to be determined. Abstract Human milk oligosaccharides (HMOs) are free glycans naturally present in human milk that act as prebiotics, prevent pathogen binding, modulate the immune system and support brain development in infants. The HMOs composition and concentrations vary significantly among different women mainly because of the direct influence of the Secretor and Lewis phenotypes on HMOs biosynthesis. Analytical methods that can identify the differences in the HMOs composition and concentrations are a fundamental tool in HMOs research. This paper describes a simple HMOs extraction and analysis for the simultaneous and absolute quantification of neutral and acidic HMOs by graphitized carbon liquid chromatography–electrospray ionization–mass spectrometry. This method was validated and applied to analyze HMOs in the human milk obtained from 10 women. This method allows accurate and reliable quantification of HMOs and can be used to determine differences in HMOs concentrations throughout lactation and among women with different Secretor and Lewis phenotypes. [ABSTRACT FROM AUTHOR]

This paper presents a multi-class target screening method for the detection of 105 veterinary drug residues from 11 classes in milk using ultra-high performance liquid chromatography electrospray ionization quadrupole Orbitrap mass spectrometry (UHPLC/ESI Q-Orbitrap). The method is based on a non-target approach of full mass scan and multiplexing data-independent acquisition (Full MS/mDIA). The veterinary drugs include endectocides, fluoroquinolones, ionophores, macrolides, nitroimidazole, NSAIDs, β-lactams, penicillins, phenicols, sulfonamides, and tetracyclines. Veterinary drug residues were extracted from milk using a salting-out and solid-phase extraction (SOSPE) procedure, which entailed the precipitation of milk proteins by an extraction buffer (oxalic acid and EDTA, pH 3) and acetonitrile, a salting-out acetonitrile/water phase separation using ammonium sulfate, and solid-phase extraction for clean-up using polymeric reversed-phase sorbent cartridges. The Q-Orbitrap Full MS/dd-MS2 (data-dependent acquisition) was used to acquire product-ion spectra of individual veterinary drugs to build a compound database and a mass spectral library, whereas its Full MS/mDIA was utilized to acquire sample data from milk for target screening of veterinary drugs fortified at 1.0 or 10.0 μg/kg. The in-spectrum mass correction or solvent background lock-mass correction was used to minimize mass error when building the compound database from experimental dd-MS2 accurate mass data. Retention time alignment and response threshold adjustment were used to eliminate or reduce false negatives and/or false positive rates. The validated method was capable of screening 58% and 96% of 105 veterinary drugs at 1.0 and 10.0 μg/kg, respectively, without manually evaluating every compound during data processing, which will reduce the workload in routine practice. [ABSTRACT FROM AUTHOR]

Chlortetracycline is (CTC) is a tetracycline antibiotic which is being in the white-tailed deer industry to improve production and animal health. In this paper, we present a method for determining chlortetracycline residues in edible white-tailed deer tissues, using liquid chromatography with heated electrospray ionization and mass spectrometry detection. The procedure involved extraction with EDTA-McIlvaine buffer at pH 4.0, followed by solid-phase extraction cleanup using a hydrophilic-lipophilic balance (HLB) cartridge. The liquid chromatography analysis was performed with heated electrospray ionization and mass spectrometry detection. The limit of quantification for the method was 2.7 ng/g and limit of detection was 0.8 ng/g. The recovery values were >78.5% for muscle, 65.1% for kidney, 63.1% for liver. Mean tissue residue concentration of chlortetracycline and it's epimer, 4-epi chlortetracycline (4-epi-CTC) at 10-day withdrawal period for kidney, liver, muscle was 122.8, 44.7 and 26.7 ng/g, respectively. Chlortetracycline tissue residue concentration at 45-day withdrawal period for kidney, liver, muscle was 19.2, 28.9 and 10.7 ng/g, respectively. Mean tissue concentration of CTC was less than the established maximum residual limit (MRL) values for bovine tissues. We have validated and successfully applied this method in the qualitative and quantification of chlortetracycline in white-tailed deer tissue samples. [ABSTRACT FROM AUTHOR]

Buxus papillosa is one of the most extensively studied species of the genus Buxus known to possess steroidal alkaloids which can be used for assessing the various pharmacological activities of this plant. This paper describes the liquid chromatography–electrospray ionization triple quadrupole mass spectrometry (LC–ESI-QQQ-MS) method for the quantification of six steroidal alkaloids as chemical markers in the extracts of leaves, roots and stems of B. papillosa . Quantitative MS/MS analysis was carried out by optimization of the most sensitive transition for each analyte. This has yielded detection and quantification limits of 0.486–8.08 ng/mL and 1.473–24.268 ng/mL, respectively for all analytes. Linearity of response was also achieved and the regression coefficient found to be >0.99 for all analyzed compounds. The newly developed MRM (Multiple Reaction Monitoring) method showed excellent sensitivity for the quantification of steroidal alkaloids within 15 min run time. This paper describes the application of LC–QQQ-MS technique for steroidal alkaloids analysis in plant samples. [ABSTRACT FROM AUTHOR]

Phytohormones are signaling and regulating metabolites involved in numerous plant processes, including growth, development, and responses to stress. Currently, the focus is on the analysis of multiple phytohormones in order to characterize crosstalk and hormone signaling networks. In this paper, representative phytohormones of the major classes are simultaneously determined in rice tissues by a generic solid-liquid extraction, followed by liquid chromatography and electrospray ionization high-resolution tandem mass spectrometry using a Q-Exactive™ instrument. After a thorough optimization of the sample preparation, the analytical method was fully validated toward the ultra-trace quantification of six a priori selected plant hormones using three scan modes of the quadrupole-Orbitrap instrument: full-scan high-resolution mass spectrometry, targeted single ion monitoring (t-SIM), and t-SIM followed by data-dependent tandem mass spectrometry. Overall, a similar quantitative performance was noticed for the different scan modes. The analytical method was successfully applied to measure basal phytohormone levels in six different rice accessions, comprising Oryza sativa ssp. japonica, indica, and Oryza glaberrima. Hormone concentrations were higher in shoots than in roots or at least similar. Except for a lower level of salicylic acid in shoots of O. glaberrima versus O. sativa, no other differences in hormone levels could be noticed that were dependent of the (sub)species assignment of the analyzed accessions. Making use of the benefits of full-scan high-resolution mass spectrometry, a first post-run suspect screening was performed, suggesting - based on accurate mass measurements and isotopic patterns - the possible presence of about 50 additional plant hormones in the rice tissues.ᅟ [ABSTRACT FROM AUTHOR]

The aim of this paper is to present the development and validation of a high-resolution full scan (HR-FS) electrospray ionization (ESI) liquid chromatography coupled to quadrupole Orbitrap mass spectrometer (LC/Q/Orbitrap MS) platform for the screening of prohibited substances in human urine according to World Antidoping Agency (WADA) requirements. The method was also validated for quantitative analysis of six endogenous steroids (epitestosterone, testosterone, 5α-dihydrotestosterone, dehydroepiandrosterone, androsterone and etiocholanolone) in their intact sulfates form. The sample preparation comprised a combination of a hydrolyzed urine liquid–liquid extraction and the dilute & shoot addition of original urine in the extracted aliquot. The HR-FS MS acquisition mode with Polarity Switching was applied in combination of the Quadrupole-Orbitrap mass filter. The HR-FS acquisition of analytical signal, for known and unknown small molecules, allows the inclusion of all analytes detectable with LC–MS for antidoping investigations to identify the use of known or novel prohibited substances and metabolites after electronic data files’ reprocessing. The method has been validated to be fit-for-purpose for the antidoping analysis. [ABSTRACT FROM AUTHOR]

This paper described a reliable and simple method for the selective determination of MQCA in animal tissues using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). A highly targeted immunoaffinity column was used for sample purification after enzymatic hydrolysis. The purified extracts were analyzed by reversed-phase HPLC–MS/MS in positive ESI and multiple reaction monitoring mode. The calibration curves showed good linearity with correlation coefficient (r 2 ) larger than 0.995. The average recoveries at the spiked levels of 0.5, 2.0 and 20 μg kg − 1 were 90.2% to 103.5% with intra-day and inter-day relatives standard deviations (RSD, n = 6) ranging from 1.8% to 6.7% and 3.5% to 7.6% respectively. The limit of quantification (LOQ) was 0.5 μg kg − 1 , which can fulfil the maximum residue level (MRL) of 4.0 μg kg − 1 stipulated by the Agricultural Minister of China and the requirement of the confirmatory criteria according to the European Commission Decision 2002/657/EC. The method is sensitive, accurate, convenient and rapid, and has been successfully applied in real samples. [ABSTRACT FROM AUTHOR]

In recent years, an increasing number of research papers revealed that the compositional and volumetric alterations in the extracellular matrix are the consequences of aging and may be related to Alzheimer's disease (AD). In this study, we aimed to demonstrate the alterations in hippocampal extracellular fluid proteins in vivo using the 5XFAD mouse model. Samples were obtained from hippocampi of 5XFAD mice (n = 6) and their non-transgenic littermates by intracerebral push-pull perfusion technique at 3 months of age, representing the pre-pathological stage of the AD. Proteins in the hippocampal perfusates were analyzed by Ultra Performance Liquid Chromatography-Electrospray Ionization Quadrupole Time-of-Flight Mass Spectrometry (UPLC-ESI-qTOF-MS/MS). 178 proteins were identified and 19 proteins of them were found to be statistically significantly altered (p≤0.05, fold change ≥40%, unique peptide count ≥3) in the hippocampal CA1 extracellular fluid of the 5XFAD mouse model. Ingenuity pathway analysis of the protein expression results identified IL6 as an upstream regulator. The upregulation of IL6 was validated by immunohistochemical staining of the hippocampus and cortex of the 5XFAD mice prior to Aβ plaque formation. Furthermore, the iron level in the hippocampus was measured by inductively coupled plasma-mass spectrometry as IL6 is mentioned in several studies to take part in iron homeostasis and inflammation and found to be increased in 5XFAD mice hippocampus. Alterations in extracellular matrix proteins in addition to increasing amount of hippocampal IL6 and iron in the early stages of AD may reveal inflammation-mediated iron dyshomeostasis in the early stages of neurodegeneration. [ABSTRACT FROM AUTHOR]

The degradation kinetics, pathways and disinfection by-products (DBPs) formation of iopamidol by chlorine and chloramines were investigated in this paper. The chlorination kinetics can be well described by a second-order model. The apparent second-order rate constants of iopamidol chlorination significantly increased with solution pH. The rate constants of iopamidol with HOCl and OCl − were calculated as (1.66 ± 0.09) × 10 −3 M −1 s −1 and (0.45± 0.02) M −1 s −1 , respectively. However, the chloramination of iopamidol fitted well with third-order kinetics and the maximum of the apparent rate constant occurred at pH 7. It was inferred that the free chlorine (i.e., HOCl and OCl − ) can react with iopamidol while the combined chlorine species (i.e., NH 2 Cl and NHCl 2 ) were not reactive with iopamidol. The main intermediates during chlorination or chloramination of iopamidol were identified using ultra performance liquid chromatography - electrospray ionization-mass spectrometry (UPLC-ESI-MS), and the destruction pathways including stepwise deiodination, hydroxylation as well as chlorination were then proposed. The regular and iodinated DBPs formed during chlorination and chloramination of iopamidol were measured. It was found that iodine conversion from iopamidol to toxic iodinated DBPs distinctly increased during chloramination. The results also indicated that although chloramines were much less reactive than chlorine toward iopamidol, they led to the formation of much more toxic iodinated DBPs, especially CHI 3 . [ABSTRACT FROM AUTHOR]

In this paper, the effect of isotope-labeled analogs on the liquid chromatography-isotope dilution mass spectrometry (LC-IDMS) measurement was evaluated based on the comparison research of electrospray ionization responses (ESI) and matrix effect of melamine, C-melamine, C+N-melamine, and N-melamine. The isotope-labeled melamines had similar ionization efficiency with melamine in the electrospray ionization source, but the intensity of corresponding quantitative fragment ions had distinctive differences. Based on the density functional theory at the B3LYP/6-311+G** level, this phenomenon was explained very well. The rare cleavage pathways of melamine, which just could be exactly identified by N-labeled melamines, resulted in the difference of quantitative fragment ions between N-labeled melamines and melamine. The interaction of ESI response between melamine and isotope-labeled melamines was investigated using MRM monitor mode. N-labeled melamine had significant ion inter-suppression effect on melamine, while C-labeled melamine had little influence on melamine. Finally, the influence of different isotope-labeled melamines on the LC-IDMS result was evaluated using the IDMS correction factor ( θ). Taking the determination of melamine in milk powder as an example, the matrix effects of different isotope-labeled melamines and melamine had notable difference and the impact of this difference on the measurement results depended on the concentrations of analyte and matrix solution. It was worth noting that N-melamine exhibited significant ion suppression to melamine in matrix solution. The deviation of the results from IDMS method might reach 59% using N-melamine as internal standard in special matrix solution. [ABSTRACT FROM AUTHOR]

Purpose: Currently, mass spectrometry has become an effective method for the qualitative analysis of plant-derived samples. Precursor and product ions can be obtained by tandem mass spectrometry, supplying rich information for determining the structural formulas of compounds. In this work, we review the optimization of qualitative methods using liquid chromatography-electrospray ionization-quadrupoletime of flight (LC-ESI-Q-TOF), coupled with ultraviolet and infrared spectroscopy, nuclear magnetic resonance, and mass spectrometry. This paper provides a systemic reference for researchers engaged in the qualitative analysis of plant-derived samples using LC-ESI-Q-TOF. [ABSTRACT FROM AUTHOR]

Electrospray ionization (ESI) tandem mass spectrometry coupled with liquid chromatography is a routine technique for identifying and quantifying compounds in complex mixtures. The identification step can be aided by matching acquired tandem mass spectra (MS2) against reference library spectra as is routine for electron ionization (El) spectra from gas chromatography/mass spectrometry (GC/MS). However, unlike the latter spectra, ESI MS2 spectra are likely to originate from various precursor ions for a given target molecule and may be acquired at varying energies and resolutions and have characteristic noise signatures, requiring processing methods very different from El to obtain complete and high quality reference spectra for individual analytes. This paper presents procedures developed for creating a tandem mass spectral library that addresses these factors. Library building begins by acquiring MS2 spectra for all major MS1 peaks in an infusion run, followed by assigning MS2 spectra to clusters and creating a consensus spectrum for each. Intensity-based constraints for cluster membership were developed, as well as peak testing to recognize and eliminate suspect peaks and reduce noise. Consensus spectra were then examined by a human evaluator using a number of criteria, including a fraction of annotated peaks and consistency of spectra for a given ion at different energies. These methods have been developed and used to build a library from >9000 compounds, yielding 230,000 spectra. [ABSTRACT FROM AUTHOR]

• Certain metabolites of tacrolimus could contribute to its toxicity. • Few papers describe LC-MS methods allowing for the quantification of 3 main metabolites. • A LC-MS method for the quantification of 3 main metabolites was developed. • Metabolic ratios were determined in a solid organ transplant population. • It's a useful tool for further studies on the role of tacrolimus metabolites in graft survival. Tacrolimus is the cornerstone of immunosuppressive therapy in solid organ transplantation and its blood concentrations are routinely monitored. Tacrolimus is extensively metabolized into metabolites that are supposed to be nephrotoxic. Yet, few analytical methods have been described to simultaneously quantify tacrolimus and its main metabolites. We developed and validated a simple liquid chromatography-mass spectrometry method for the quantification of tacrolimus and its three desmethylated metabolites, 13-O, 15-O, and 31-O-desmethylated tacrolimus (M−I, M−III, and M−II respectively) in human whole blood. Protein precipitation of 50 µL of whole blood with 100 µL methanol and zinc sulfate was used as a single-extraction procedure. Tacrolimus and its metabolites were quantified using electrospray ionization-triple quadrupole mass spectrometry in combination with selected reaction monitoring detection in the positive ionization mode. The method was validated following FDA recommendations. This method was precise (intra- and inter-assay coefficients of variation: 2.88–7.81% and 3.96–12.10% for low and high levels of internal quality controls, respectively) and accurate (intra- and inter-assay biases: −1.67–10.30%, and −0.77–-9.36%, respectively). In adult kidney transplant patients who were treated with tacrolimus prolonged release formulation, the median (10th-90th percentiles) trough concentrations (n = 16) of tacrolimus, M−I, and M−III were 5.85 (3.37–7.09), 0.100 (0.037–0.168), 0.051 (0.03–0.104), respectively. M−II was measured in only 2 trough samples. The metabolic ratios M−I/tacrolimus and M−III/tacrolimus were 0.017 (0.009–0.027) and 0.009 (0.006–0.015) when measured on trough concentration and 0.022 (0.011–0.037) and 0.008 (0.006–0.015) when measured on area under the curves 0–24 h. This method is a suitable and easy-to-perform tool for future pharmacokinetic-pharmacodynamics studies investigating the importance of tacrolimus and its metabolites blood exposure for solid organ graft survival. [ABSTRACT FROM AUTHOR]

Dalbergia odorifera, a traditional Chinese medicine, has been used to treat cardio- and cerebrovascular diseases in China for thousands of years. Flavonoids are major active compounds in D. odorifera. In this paper, a rapid and sensitive ultra-high performance liquid chromatography-triple quadrupole mass spectrometry method was developed and validated for simultaneous determination of 17 flavonoids in D. odorifera. Quantification was performed by multiple reaction monitoring using electrospray ionization in negative ion mode. Under the optimum conditions, calibration curves for the 17 analytes displayed good linearity (r2 > 0.9980). The intra- and inter-day precisions (relative standard deviations) were lower than 5.0%. The limit of quantitation ranged from 0.256 to 18.840 ng/mL. The mean recovery range at three spiked concentrations was 94.18–101.97%. The validated approach was successfully applied to 18 samples of D. odorifera. Large variation was observed for the contents of the 17 analytes. Sativanone and 3′-O-methylviolanone were the dominant compounds. The fragmentation behaviors of six flavonoids were investigated using UPLC with quadrupole time-of-flight tandem mass spectrometry. In negative ion electrospray ionization mass spectrometry, all the flavonoids yielded prominent [M − H]− ions. Fragments for losses of CH3, CO, and CO2 were observed in the mass spectra. Formononetin, liquiritigenin, isoliquiritigenin, sativanone, and alpinetin underwent retro-Diels–Alder fragmentations. The proposed method will be helpful for quality control of D. odorifera. [ABSTRACT FROM AUTHOR]