UPLC Tandem Mass Spectrometry Research Articles

Development and Validation of UPLC Tandem Mass Spectrometry Assay for Ceftibuten and Sulbactam in Human Plasma.

A sensitive and rapid ultra-performance liquid chromatography coupled with -tandem mass spectrometry (UPLC-MS/MS) method was developed and validated to determine ceftibuten (CTB) and sulbactam (SUL) in human plasma. An ACQUITY UPLC HSS T3 C18 (2.1 × 100 mm), 1.8 μm column with gradient elution of water (0.1% formic acid) and acetonitrile was used for separation at a flow rate of 0.2mL/min. This method involves a simple sample preparation with acetonitrile. The calibration curves of CTB and SUL in plasma showed good linearity over the concentration range of 0.50-25 μg/mL and with a correlation coefficient (r2) >0.99. This method was validated in terms of selectivity, linearity, precision, accuracy and stability. High precision was obtained with coefficients of variation <15%. Excellent recovery in the range of 90-104% was achieved for CTB and SUL was 86-110%. The method has the potential utility to support pharmacometric modeling in clinical practice and biopharmaceutic studies.

Robust and ultra-sensitive quantitation of plasma aldosterone by UPLC tandem mass spectrometry

Robust and ultra-sensitive quantitation of plasma aldosterone by UPLC tandem mass spectrometry

An advanced strategy for quality evaluation of Xinyi Biyan Pill by UPLC-DAD fingerprinting combined with multi-components UPLC-MS/MS analysis

Xinyi Biyan Pill (XBP) is a classical Chinese patent medicine and widely used to treat acute and chronic or allergic rhinitis in clinical practice. This study aimed to establish and validate a comprehensive strategy combining ultra-performance liquid chromatography with diode array detector (UPLC-DAD) fingerprinting and multi-component quantification for quality evaluation of XBP. In the fingerprint analysis, 32 peaks were selected as common peaks and used to evaluate the similarity of 12 batches of XBP. In addition, 141 compounds of XBP were identified or preliminarily characterized in both positive and negative ion modes by coupling with an advanced hybrid linear ion trap (LTQ)-Orbitrap mass spectrometer. Moreover, a rapid quantitative method based on UPLC tandem mass spectrometry (UPLC-MS/MS) has been optimized and validated for simultaneous determination of 10 chemical markers within 15 min, and applied to analyzing 12 batches of XBP. The proposed comprehensive strategy combining UPLC-DAD fingerprinting and multi-component UPLC-MS/MS quantification exhibited satisfactory results with high efficiency, accuracy and reliability, which can be used as a reference for overall quality consistency evaluation of Chinese herbal formulations.

B-166 Effect of Urine Adulteration on Quantifying both Delta 8- and Delta 9-carboxy tetrahydrocannabinol using a New UPLC-tandem Mass Spectrometry Method

Abstract Background The legality and use of marijuana is a controversial topic. With increased prevalence of usage, cannabis detection for legal, medical, and workforce drug testing is on the rise. LC-MS/MS quantification of delta 9-carboxy tetrahydrocannabinol (delta 9-THC-COOH) in urine is accepted as the standard to detect marijuana use. One challenge faced by testing laboratories has been identifying adulterated samples. Many products/reagents can be used to manipulate the urine to avoid positive drug testing results. In this study, we developed a dilute-and-shoot strategy to separate and quantify both the delta 8- and 9-THC-COOH in urine samples and tested the effects of adulteration using a combination of nitrite and/or acids which can be used to mask cannabinoid use. Methods Five authentic delta 8- and 9-THC-COOH positive specimens, and one spiked sample were treated with strong acids (1N HCL or 10% TCA), 500 µg/mL nitrite, or a combination. After hydrolysis with sodium hydroxide, the analytes were quantified using a 6495 Mass Spectrometer coupled to a StreamSelect LC system (Agilent). LC separation was performed with a CORTECS T3 column utilizing a shallow gradient. In addition, twelve urine specimens with varying concentrations of delta 8- and 9-THC-COOH were treated with nitrite and screened using a delta 9-THC-COOH immunoassay on the Roche Cobas C501 and analyzed by LC-MS/MS. Results Nitrite did not change the physical appearance or the pH of the urine samples. Strong acid treatment also did not alter the appearance but did drop the urine pH ≤2. Pre-treated specimens with 1N HCL, 10% TCA, or 500 µg/mL of nitrite did not affect the delta 8- and 9-THC-COOH concentrations up to 6 days (refrigerated); however, specimens treated with strong acid combined with 500 µg/mL of nitrite for 30 min made both analytes undetectable. Additional experiments confirmed that 0.5 and 1 mol/L of nitrite lowered the concentration of all analytes, including internal standard, to undetectable levels in 48 h. Twelve additional specimens were collected, 3 with delta 8-THC-COOH, 3 with delta 9-THC-COOH, and 6 with both. All 12 samples screened positive using the Roche immunoassay and were quantified using the LC-MS/MS method. After treatment with 0.5 mol/L of nitrite, 10 of the 12 samples were negative using the immunoassay, but 2 samples remained positive. However, all nitrite treated specimens were negative (&amp;lt;LLOQ) using the LC-MS/MS method and the internal standard was eliminated. Later experiments confirmed that the delta 8- and 9-THC-COOH could be eliminated with as little as 0.1 mol/L nitrite for 30 min. Conclusion The unique dilute-and-shoot method can separate and quantify delta 8- and 9-THC-COOH. In addition, high concentrations of nitrite were shown to degrade both analytes along with the isotopically labeled internal standard. As a result, labs can distinguish true negative delta 8- and 9-THC-COOH (no analyte peaks) from nitrite adulterated specimens since the adulterated samples have no peaks for both the analytes and internal standard. A limitation of the study is that only sodium nitrite was tested. Other adulterants, such as potassium nitrite, pyridinium chlorochromate, peroxide etc. require further investigation.

Discovery of anti-SARS-CoV-2 secondary metabolites from the heartwood of Pterocarpus santalinus using multi-informative molecular networking.

A pigment-depleted extract from the heartwood of Pterocarpus santalinus L. f. (PS-DE) showed promising anti-SARS-CoV-2 activity with an IC50 of 29.9μg/mL in Caco-2-F03 cells. To determine the potential active constituents within the extract prior to isolation, multi-informative molecular network (MN) was applied. Therefore, the extract was separated by high-performance counter-current chromatography (HPCCC) into 11 fractions which were subsequently tested for anti-SARS-CoV-2 activity and analysed by UPLC-tandem mass spectrometry (MS2). The resulting MN combines the bioactivity data of the fractions with the MS2 data. The MN analysis led to the targeted isolation of seven compounds including one pterocarpan (7) reported for the first time as constituent of P. santalinus and four so far undescribed natural products (NPs) that belong to the compound classes of arylpropanes (9), isoflavanones (10) coumestans (16) and 3-arylcoumarins (17), respectively. In total, 15 constituents from the heartwood of P. santalinus and one synthetic isoflavonoid that is structurally related to the natural metabolites were tested for anti-SARS-CoV-2 activity. Thereby, the two pterocarpans (-)-homopterocarpin (5) and (-)-medicarpin (2), the stilbene (E)-pterostilbene (1) and the isoflavonoid 7-O-methylgenistein (11) showed a distinct antiviral activity with IC50 values of 17.2, 33.4, 34.7, and 37.9µM, respectively, and no cytotoxic effects against Caco-2-F03 cells (CC50 > 100µM). In addition, a structure-activity relationship (SAR) was proposed indicating structural requirements of pterocarpans for anti-SARS-CoV-2 activity. The herein presented results support the implementation of multi-informative molecular networks as powerful tool for dereplication and targeted isolation of bioactive NPs.

Vitamin and Mineral Supplementation and Rate of Weight Gain during the First Trimester of Gestation in Beef Heifers Alters the Fetal Liver Amino Acid, Carbohydrate, and Energy Profile at Day 83 of Gestation.

The objective of this study was to evaluate the effects of feeding heifers a vitamin and mineral supplement and targeting divergent rates of weight gain during early gestation on the fetal liver amino acid, carbohydrate, and energy profile at d 83 of gestation. Seventy-two crossbred Angus heifers were randomly assigned in a 2 × 2 factorial arrangement to one of four treatments comprising the main effects of vitamin and mineral supplementation (VTM or NOVTM) and feeding to achieve different rates of weight gain (low gain [LG] 0.28 kg/day vs. moderate gain [MG] 0.79 kg/day). Thirty-five gestating heifers with female fetuses were ovariohysterectomized on d 83 of gestation and fetal liver was collected and analyzed by reverse phase UPLC-tandem mass spectrometry with positive and negative ion mode electrospray ionization, as well as by hydrophilic interaction liquid chromatography UPLC-MS/MS with negative ion mode ESI for compounds of known identity. The Glycine, Serine, and Threonine metabolism pathway and the Leucine, Isoleucine, and Valine metabolism pathway had a greater total metabolite abundance in the liver of the NOVTM-LG group and least in the VTM-LG group (p < 0.01). Finally, both the TCA Cycle and Oxidative Phosphorylation pathways within the Energy Metabolism superpathway were differentially affected by the main effect of VTM, where the TCA cycle metabolites were greater (p = 0.04) in the NOVTM fetal livers and the Oxidative Phosphorylation biochemicals were greater (p = 0.02) in the fetal livers of the VTM supplemented heifers. These data demonstrate that the majority of metabolites that are affected by rate of weight gain or vitamin/mineral supplementation are decreased in heifers on a greater rate of weight gain or vitamin/mineral supplementation.

Integration of multiplatform metabolomics and multivariate analysis for geographical origin discrimination of Panax ginseng

As the health food industry grows, the market for ginseng also expands globally. Korean ginseng (Panax ginseng C. A. Meyer) is cultivated in East Asian countries such as Korea, China, and Japan. The metabolic profile of plants can vary depending on the cultivation environment. As such, in this study, we aimed to compare the differences in the metabolic profiles of P. ginseng cultivated in Korea, China, and Japan, and to construct a library of these metabolite data. Using ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS), we profiled 62 types of secondary metabolites, ginsenosides, using optimized analysis conditions to separate peaks with a high-resolution for about 30 min. In addition, we selected ginsenosides showing differences between their origins were selected among the possible origins in the S-line plot of orthogonal partial least squares discriminant analysis (OPLS-DA), which we quantitatively analyzed using UPLC-tandem mass spectrometry (UPLC-MS/MS). The contents of ginsenosides Ra1, Ra2, Ra3, and Rk1 were high in Korean P. ginseng; the contents of ginsenosides Rb1, Rb2, and Rc were high in Japanese P. ginseng; and the contents of the ginsenoside Ro was high in Chinese P. ginseng. We also analyzed the primary metabolite contents using nuclear magnetic resonance (NMR) spectroscopy and gas chromatography time-of-flight mass spectrometry (GC-TOF/MS). Japanese P. ginseng showed a high sucrose content and Korean P. ginseng showed high contents of most amino acids and organic acids. In the PLS-DA results of multivariate statistical analysis using the data obtained from each analysis instrument, we observed a clear clustering among the three origins. Although a genetically identical species, the metabolic profile substantially differs depending on the cultivation environment. Because ginsenoside, having many biological activities, showed origin-dependent origins, when P. ginseng is used for medicinal purposes, its content by origin should be considered. After disclosing the profiling results of these metabolites, we expect that they will be used in future ginseng research.

Establishing an untargeted-to-MRM liquid chromatography–mass spectrometry method for discriminating reconstituted milk from ultra-high temperature milk

The counterfeit use of reconstituted milk to simulate fresh milk in some countries is largely driven by profiteering; hence, establishing a robust market-surveillance method is an important objective. In this study, an untargeted metabolomics approach that uses ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC–Q-TOF-MS) was established to discover biomarkers that can be used to classify UHT and reconstituted milk. Furthermore, 36 positive- and 24 negative-ionization mode features were selected as candidate markers to establish a UPLC–tandem mass spectrometry (UPLC–MS/MS) multiple reaction monitoring (MRM) method. The support vector machine (SVM) method was used to process the responses of the selected potential markers, and 100% classifiability was observed. The marker metabolites could be divided into three categories by hierarchical clustering analysis: peptides, lipids, and nucleic acids. The results suggest that the untargeted-to-MRM metabolomics method is an effective tool for distinguishing between UHT and reconstituted milk.

1904-P: Metabolomic Fingerprints after Metabolic Surgery: The STAMPEDE Trial

Introduction: Metabolic surgery (MS) results in durable resolution of hyperglycemia in ∼50% of patients with type 2 diabetes (T2D). Roux-en-Y gastric bypass (RYGB) and sleeve gastrectomy (SG) account for more than 90% of all metabolic surgeries, and both have been demonstrated to have greater efficacy when compared to intensive medical and lifestyle therapy (IMT) to improve glycemic control. However, a subset of patients following MS exhibit inadequate weight loss, poor glycemic control, or both. Thus, the purpose of this study was to identify novel biomarkers of biochemical resolution of T2D following IMT and MS. Methods: Plasma samples from ninety patients (49.9±7.6 years, 57.7% female) randomly assigned to receive either IMT (n=30), RYGB (n=30) or SG (n=30) were retrospectively analyzed and quantified by untargeted metabolomic analysis using UPLC-tandem mass spectrometry at baseline (PRE) and 24 months (POST) of treatment. Results: We identified 384 metabolites altered by RYGB, SG and IMT treatment. 2-hydroxybutyrate was significantly decreased following all 3 interventions with a greater decrease in SG and RYGB at POST (p≤0.05). Pyruvate was significant lower in the IMT and RYGB group (p≤0.05), but not in SG at POST (vs. PRE). Markers of fatty acid metabolism, including acetylcarnitine and 3-hydroxybutyrate (BHBA), were also decreased following the SG intervention (p≤0.05). Several primary and secondary bile acid metabolites were significantly increased after RYGB compared to SG. Conclusion: We identified a novel metabolomic fingerprint characterizing the longer-term adaptations to IMT, RYGB, and SG. Notably, the metabolomic profile of both RYGB and SG procedures were distinct, suggesting that the roughly equivalent weight loss is achieved by divergent mechanisms. Disclosure W.S. Dantas: None. C.L. Axelrod: None. A. Hari: None. S. Kashyap: Other Relationship; Self; GI Dynamics Inc. P. Schauer: Advisory Panel; Self; GI Dynamics Inc. Consultant; Self; Ethicon US, LLC, Medtronic. Research Support; Self; Ethicon US, LLC, Medtronic. J.P. Kirwan: None. Funding Ethicon

Utilization of UPLC-tandem mass spectrometry for quantitation of colouring matters in dietary formulation

A simple and sensitive ultra-performance liquid chromatography with Tandem Mass Spectrometric detection (UPLC-MS/MS) method was developed and validated for the simultaneous quantitation of two food colouring matters, Curcumin (CUR) and Riboflavin (RIB). Chromatographic separation was done on Hypersil gold 50×2.1 mm (1.9 μm) column, with gradient programing of mobile phase starting with aqueous 0.1% formic acid and increasing the percentage of 0.1% formic acid in acetonitrile. The utilization of multiple reaction monitoring (MRM) improved the selectivity of detection and decreased the matrix effect. The method was linear in the range of 5-500 ng/mL for CUR and RIB. Intra- and inter-day reproducibility were within the accepted criteria. The method was successfully applied for the determination the laboratory prepared mixtures of the two selected colouring matters. The use of mass spectrometry enhanced the selectivity and sensitivity of detection which allows the robust use of the method in routine quality control tests of the two colouring matters (CUR and RIB).

Development of an UPLC-MS/MS method coupled with in-source CID for quantitative analysis of PEG-PLA copolymer and its application to a pharmacokinetic study in rats

Poly(ethylene glycol)-block-poly(lactic acid) (PEG-PLA) is a biocompatible and amphiphilic block copolymer composed of a hydrophilic PEG block and a hydrophobic PLA block, which can self-assemble into micelles in water. It is one of the most commonly used biodegradable polymers for drug encapsulation, drug solubilization and drug delivery. Due to the complexity and heterogeneity of PEG-PLA, the precise analysis of this polymer is a great challenge. This study reports an application of an UPLC tandem mass spectrometry coupled with in-source collision induced dissociation (CID) technique for the analysis of a model compound mPEG2000-PDLLA2500-COOH, which could be dissociated in source and generate a series of fragment ions corresponding to its subunits. These surrogate ions including PLA-specific and PEG-specific fragment ions could be further broken into specific product ions in collision cell. Finally, the ion transition at m/z 505.0 → 217.0 was selected for the quantitation of mPEG2000-PDLLA2500-COOH. This assay achieved a lower limit of quantitation (LLOQ) of 0.05 μg/mL with only 30 μL rat plasma. The linear range is 0.05 to 5 μg/mL. Intraday and interday accuracy and precision were within ±12.1%. The method was successfully applied to the pharmacokinetic study of mPEG2000-PDLLA2500-COOH in rats. The results revealed that LC-MS/MS coupled with in-source CID is a sensitive and specific strategy for analysis of PEG-PLA. This method can be potentially extended to the analysis of other pharmaceutical polymer excipients.

Quantitative screening and prioritization based on UPLC-IM-Q-TOF-MS as an alternative water sample monitoring strategy.

Suspect and non-target screening based on the use of high-resolution mass spectrometry (HRMS) has become more common in water analysis over the past years. However, this only yields lists of features or suspects without quantitative information. To expand the use of HRMS data to a quantitative screening, we have developed and validated a simple and fast method for more than 140 micropollutants using ultra high-performance liquid chromatography coupled to traveling wave ion mobility quadrupole time-of-flight mass spectrometry (UPLC-IM-Q-TOF-MS). Positive findings from suspect and non-target screening can be prioritized and identified by reference standards. The quantitative screening is then performed by additional measurement of calibration standards. This is carried out by means of direct injection and external calibration, without consideration of matrix effects. For all substances, limits of quantification (LOQs) of less or equal than 100ng/L are achieved. The calibration is carried out in a range of 100 to 1000ng/L and the results are reported as concentration ranges, in which the concentration of the analyte in the sample is to be expected. All substances were evaluated using quadratic regressions. For the verification of the accuracy, different water matrices (drinking water, groundwater, and surface water) were spiked with five concentration levels (50ng/L, 300ng/L, 500ng/L, 800ng/L, and 2000ng/L) and indicate that for the drinking water and groundwater sample, 97% correct assignments were found, whereas for the surface water sample, 88% correct assignments were achieved. A comparative study of water samples of various matrices was accomplished using the quantitative screening analysis method and validated target methods by means of three UPLC tandem mass spectrometry (MS/MS) methods and two gas chromatography (GC) coupled to MS and MS/MS methods. A total of 510 data could be compared, which showed a good match of both approaches in more than 80% of the results. As an alternative strategy for the monitoring of water samples by UPLC-IM-Q-TOF-MS, this method provides quantitative information about target components, besides tentatively or identified substances from suspect or non-target screening. Depending on the resulting concentration range and reporting requirements, validated target methods can be further used for the previously detected targets. Graphical abstract.

108: Molecular similarities and dissimilarities between hellp syndrome and preeclampsia (PE) using shotgun and targeted proteomics

Although clinical management of HELLP syndrome is similar to PE, hypertension and proteinuria do not always occur with HELPP. Thus, it was suggested that HELLP and PE could be separate clinical entities. Based on the ability of the urine misfolded proteins to retain Congo red (congophilia) it was suggested PE is a protein conformational disorder similar to Alzheimer’s disease. Molecular diagnostics are becoming more often part of modern clinical medicine. Here we used state of the art shotgun proteomics and targeted multiple reaction monitoring (MRM) mass spectrometry to discover and validate molecular signatures in crude urine samples of pregnant women with PE and HELLP that potentially differentiate between these two clinical conditions. We analyzed urine samples from six groups of patients classified phenotypically as: non-hypertensive healthy controls (CRL, n=9, 30±1 wks), women with idiopathic proteinuria (IP, n=15, 28±2 wks), PE mild features (mPE, n=16, 33±1 wks), superimposed PE (spPE, n=16, 31±1 wks), PE severe features (sPE, n=16, 32±1 wks) and HELLP (n=16, 29±1 wks). In the discovery phase, urine proteins were enriched by CR precipitation and subjected to UPLC-tandem mass spectrometry. Linear discriminant analysis (LDA) was used to cluster clinical groups based on molecular signatures. We choose a non-parsimonious set of peptides that distinguished among clinical phenotypes and were suitable for targeted validation using MRM in crude urine samples. 1,747 molecular IDs were identified in CR precipitates. These merged into 690 protein groups that generated good separation among clinical phenotypes with a false discovery rate of 1.12% (Fig. A). 21 key peptides were prioritized through pairwise comparison among groups (Wilkes Lambda MANOVA=0.006, p-value of 3.6 x 10-14) (Fig. B). Based on a full MS/MS spectral library, 98 MRM transitions were generated. CLIC1 (oxidant sensor activated by misfolded proteins), ABHD14B (lipid metabolizing-enzyme) and β2-microglobulin were among the parent proteins to distinguish sPE from HELLP. Principal component analysis demonstrated that targeted proteome from raw urine represented CR precipitate well and closely correlated with clinical criteria. Based on molecular signatures, HELLP syndrome appears to be a different clinical entity from sPE.

A novel method for quantitation of acylglycines in human dried blood spots by UPLC-tandem mass spectrometry

BackgroundSeveral acylcarnitines used as primary markers on dried blood filter papers (DBS) for newborn screening lack specificity and contribute to a higher false positive rate. The analysis of urine acylglycines is useful in the diagnosis of inborn errors of metabolism (IEM) including medium chain acyl-CoA dehydrogenase deficiency (MCADD), isovaleric acidemia, and beta-ketothiolase deficiency (BKTD). Currently, no method for analyzing acylglycines from DBS has been published. MethodsAcylglycines were extracted from two 3.2 mm DBS punches and butylated using Butanol-HCl. Ultra Performance Liquid Chromatography (UPLC-MS/MS) with run time of 10 min permits resolution and quantitation of 15 acylglycines; including several isobaric. Method development was completed. Reference intervals (n = 573) were established for four birth weight groups. Furthermore, samples from patients with a confirmed IEM (n = 11), and false positive screens (n = 78) were analyzed to validate the interpretation obtained from the newly established reference intervals. ResultsCalibration curves were linear from 0.005 to 25.0 μM. Ion suppression was evaluated as minimal (2 to 10%). Samples from known patients were used to validate the reference intervals. For C5OH-related disorders, tiglylglycine (TG), TG/acetylglycine (AG) ratio, 3methylcrotonylglycine (3MCG) and 3MCG/AG ratio increased specificity. Propionylglycine (PG) and PG/acetylglycine ratio were two discriminatory markers in the investigation of C3-related disorders. Hexanoylglycine (HG), octanoylglycine (OG), suberylglycine (SG), and the ratios HG/AG, OC/AG and SG/AG were excellent markers of MCADD deficiency. ConclusionThis method shows potential application as a second tier screen in order to reduce the false positive rate for a number of IEM targeted by newborn screening.

Exemestane potency is unchanged by common nonsynonymous polymorphisms in CYP19A1: results of a novel anti-aromatase activity assay examining exemestane and its derivatives.

Exemestane (EXE) treats estrogen receptor positive (ER+) breast cancer in postmenopausal women by inhibiting the estrogen‐synthesizing cytochrome P450 CYP19A1. Variability in the severity and incidence of side effects as well as overall drug efficacy may be partially explained by genetic factors, including nonsynonymous variation in CYP19A1, also known as aromatase. The present study identified phase I EXE metabolites in human liver microsomes (HLM) and investigated mechanisms that may alter the extent of systemic estrogen deprivation in EXE‐treated women with breast cancer, including whether functional polymorphisms in aromatase cause differential inhibition by EXE and whether EXE metabolites possess anti‐aromatase activity. The potency of EXE and ten of its derivatives was measured with HEK293‐overexpressed wild type aromatase (CYP19A1*1) using a rapid novel UPLC tandem mass spectrometry method. Of the ten compounds assayed, five were poor inhibitors (IC 50 ˃ 50 μmol/L) of wild type aromatase while five others, including the major metabolite, 17β‐dihydroexemestane (17β‐DHE), exhibited moderate potency, with IC 50 values ranging between 1.2 and 7.1 μmol/L. The anti‐aromatase activity of EXE was also tested with two common allozymes, aromataseThr201Met (CYP19A1*3) and aromataseArg264Cys (CYP19A1*4). Differential inhibition of variant aromatase is unlikely to account for variable clinical outcomes as EXE‐mediated inhibition of aromataseThr201Met (IC 50 = 0.86 ± 0.12 μmol/L) and aromataseArg264Cys (IC 50 = 1.7 ± 0.65 μmol/L) did not significantly differ from wild type (IC 50 = 0.92 ± 0.17 μmol/L). Although less potent than the parent drug, these results suggest that active metabolites may contribute to the therapeutic mechanism of EXE.

Determination and dissipation of mesotrione and its metabolites in rice using UPLC and triple-quadrupole tandem mass spectrometry.

In this study, an efficient and fast method for the determination of the herbicide mesotrione and its two metabolites [4-methylsulfonyl-2-nitrobenzoic acid (MNBA) and 2-amino-4-methylsulfonylbenzoic acid (AMBA)] in rice, rice plants, rice hulls, water, and soil was developed, using ultra performance liquid chromatography coupled with tandem quadrupole (Xevo TQ-S) mass spectrometry (UPLC-Xevo TQ-S-MS/MS). The recoveries of the compounds were 68.6%-108.4% at different levels. The intra-day relative standard deviation (RSD) ranged from 2.0% to 14.4%, and the inter-day RSD, from 1.9% to 11.4%. The limits of quantification (LOQ) were ranged from 0.146 to 4.545μgkg-1for mesotrione, MNBA and AMBA. Dissipation and residual assays with the commercial formulation were conducted at three locations. The dissipation rates of the compounds were fast in rice plants, soil, and water, and residues at harvest were not detected in any of the three localities.

UPLC-Tandem Mass Spectrometry Method for Simultaneous Determination of Fluoxetine, Risperidone, and Its Active Metabolite 9-Hydroxyrisperidone in Plasma: Application to Pharmacokinetics Study in Rats.

Risperidone (RIS) is used as an antipsychotic drug alone or with other drugs, like fluoxetine (FLX). A simple method was developed and validated for the determination of both RIS and its metabolite 9-hydroxyrisperidone (9-OH-RIS), FLX, and olanzapine (OLA) as an internal standard in rat's plasma using UPLC-MS/MS. FLX, RIS, 9-OH-RIS, and OLA were purified using acetonitrile as a protein precipitating agent. Separation was performed on an ACQUITY™ “UPLC BEH™” C18 column (50 mm × 2.1 mm i.d., 1.7 μm; Waters Corp., USA). The ranges of the calibration curves were 1.0–1000.0 ng/mL for FLX and 0.2–1000.0 ng/mL for RIS and 9-OH-RIS. Linearity, recovery, precision, and stability were within the acceptable range. This method is rapid, fast, and precise for the determination of RIS and FLX in plasma and is applicable in pharmacokinetic studies.

Pressurized hot water extraction and chemometric fingerprinting of flavonoids from Bidens pilosa by UPLC-tandem mass spectrometry

ABSTRACTThe need for greener extraction procedures that are quick and efficient has prompted the evolution of pressurized hot water extraction (PHWE). Here, the extraction of flavonoids from Bidens pilosa was demonstrated using PHWE at 50°C, 100°C and 150°C. The extracts were analyzed on UPLC-qTOF-MS/MS and 28 flavonoids of different classes were identified. Further analysis of the data using principal component analysis revealed differential distribution patterns of the identified molecules. In overall, the extraction yield increased proportionately with increasing temperature. It can thus be deduced that PHWE is an excellent extraction method of flavonoids from plant tissues. Again, this study reiterates B. pilosa as a rich source of flavonoids.

Urine Purine Metabolite Determination by UPLC-Tandem Mass Spectrometry.

Inborn errors of purine metabolism, either deficiencies of synthesis or catabolism pathways, lead to a wide spectrum of clinical presentations: urolithiasis (adenine phosphoribosyltransferase), primary immune deficiency (adenosine deaminase deficiency and purine nucleoside phosphorylase deficiency), severe intellectual disability, and other neurological symptoms (Lesch-Nyhan disease, adenylosuccinase deficiency, and molybdenum cofactor deficiency). A rapid quantitative purine assay was developed using UPLC-MS/MS to determine purine nucleoside and base concentrations in urine. Taking advantages of ultra performance liquid chromatography, we achieved satisfactory analyte separation and recovery with a polar T3 column in a short run time with no requirement of time-consuming sample preparation or derivatization. This targeted assay is intended for diagnosis and management of purine diseases, newborn screening follow-up of SCID, and evaluation of autism spectrum disorders.

Identification of hydroxylcinnamoyl tartaric acid esters in Bidens pilosa by UPLC-tandem mass spectrometry

Bidens pilosa is a medicinal plant used for the treatment of several physiological illnesses. In South Africa, as the case may be for other African countries, this plant is equally consumed as a vegetable. In the current study, pressurized hot water extraction (PHWE) technology was employed for the extraction of polyphenolic compounds from leaves of B. pilosa under two different temperature conditions (100 and 150°C). Accordingly, extraction of these compounds was made possible at 150°C and analysis of these extracts using UPLC-qTOF-MS/MS revealed the presence of several hydoxylcinnamoyl tartaric acids. Here, different isomers of coutaric-, caftaric-, fertaric-, chicoric acid and caftaric acid glycosides were detected. The contribution of mass spectrometry fragmentation towards the characterization of these molecules is also presented. To the best of our knowledge, this is the first report of these molecules in B. pilosa.