Browse
Submit Data
Databases
API
Help

Dataset Information

19 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Rapid quantification of insulin degludec by immunopurification combined with liquid chromatography high-resolution mass spectrometry.

ABSTRACT: Insulin degludec is an ultra-long-acting insulin analogue that is increasingly being used in diabetes due to its favourable efficacy and safety profile. Thus, there is an increasing demand for a reliable and specific analytical method to quantify insulin degludec for research, pharmaceutical industry and clinical applications. We developed and validated an automated, high-throughput method for quantification of insulin degludec in human blood samples across the expected clinical range combining immunopurification with high-resolution mass spectrometry. Validation was performed according to the requirements of the US Food and Drug Administration. The method satisfyingly met the following parameters: lower limit of quantification (120 pM), linearity, accuracy (error

SUBMITTER: Reverter-Branchat G

PROVIDER: S-EPMC7680744 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Rapid quantification of insulin degludec by immunopurification combined with liquid chromatography high-resolution mass spectrometry.

Reverter-Branchat Gemma G Groessl Michael M Nakas Christos T CT Prost Jean-Christophe JC Antwi Kwasi K Niederkofler Eric E EE Bally Lia L

Analytical and bioanalytical chemistry 20201002 30

Insulin degludec is an ultra-long-acting insulin analogue that is increasingly being used in diabetes due to its favourable efficacy and safety profile. Thus, there is an increasing demand for a reliable and specific analytical method to quantify insulin degludec for research, pharmaceutical industry and clinical applications. We developed and validated an automated, high-throughput method for quantification of insulin degludec in human blood samples across the expected clinical range combining ...[more]

PMID: 33006670

Similar Datasets

Simultaneous isotope dilution quantification and metabolic tracing of deoxyribonucleotides by liquid chromatography high resolution mass spectrometry.

Project description:Quantification of cellular deoxyribonucleoside mono- (dNMP), di- (dNDP), triphosphates (dNTPs) and related nucleoside metabolites are difficult due to their physiochemical properties and widely varying abundance. Involvement of dNTP metabolism in cellular processes including senescence and pathophysiological processes including cancer and viral infection make dNTP metabolism an important bioanalytical target. We modified a previously developed ion pairing reversed phase chromatography-mass spectrometry method for the simultaneous quantification and 13C isotope tracing of dNTP metabolites. dNMPs, dNDPs, and dNTPs were chromatographically resolved to avoid mis-annotation of in-source fragmentation. We used commercially available 13C15N-stable isotope labeled analogs as internal standards and show that this isotope dilution approach improves analytical figures of merit. At sufficiently high mass resolution achievable on an Orbitrap mass analyzer, stable isotope resolved metabolomics allows simultaneous isotope dilution quantification and 13C isotope tracing from major substrates including 13C-glucose. As a proof of principle, we quantified dNMP, dNDP and dNTP pools from multiple cell lines. We also identified isotopologue enrichment from glucose corresponding to ribose from the pentose-phosphate pathway in dNTP metabolites.

| S-EPMC7359880 | biostudies-literature

Quantification of Dolichyl Phosphates Using Phosphate Methylation and Reverse-Phase Liquid Chromatography-High Resolution Mass Spectrometry.

Project description:Dolichyl monophosphates (DolPs) are essential lipids in glycosylation pathways that are highly conserved across almost all domains of life. The availability of DolP is critical for all glycosylation processes, as these lipids serve as membrane-anchored building blocks used by various types of glycosyltransferases to generate complex post-translational modifications of proteins and lipids. The analysis of DolP species by reverse-phase liquid chromatography-mass spectrometry (RPLC-MS) remains a challenge due to their very low abundance and wide range of lipophilicities. Until now, a method for the simultaneous qualitative and quantitative assessment of DolP species from biological membranes has been lacking. Here, we describe a novel approach based on simple sample preparation, rapid and efficient trimethylsilyl diazomethane-dependent phosphate methylation, and RPLC-MS analysis for quantification of DolP species with different isoprene chain lengths. We used this workflow to selectively quantify DolP species from lipid extracts derived of Saccharomyces cerevisiae, HeLa, and human skin fibroblasts from steroid 5-α-reductase 3- congenital disorders of glycosylation (SRD5A3-CDG) patients and healthy controls. Integration of this workflow with global lipidomics analyses will be a powerful tool to expand our understanding of the role of DolPs in pathophysiological alterations of metabolic pathways downstream of HMG-CoA reductase, associated with CDGs, hypercholesterolemia, neurodegeneration, and cancer.

| S-EPMC9933046 | biostudies-literature

Quantification of protein phosphorylation by liquid chromatography-mass spectrometry.

Project description:The identification and quantification of specific phosphorylation sites within a protein by mass spectrometry has proved challenging when measured from peptides after protein digestion because each peptide has a unique ionization efficiency that alters with modification, such as phosphorylation, and because phosphorylation can alter cleavage by trypsin, shifting peptide distribution. In addition, some phosphorylated peptides generated by tryptic digest are small and hydrophilic and, thus, are not retained well on commonly used C18 columns. We have developed a novel C-terminal peptide (2)H-labeling derivatization strategy and a mass balance approach to quantify phosphorylation. We illustrate the application of our method using electrospray ionization liquid chromatography-mass spectrometry by quantifying phosphorylation of troponin I with protein kinase A and protein kinase C. The method also improves the retention and elution of hydrophilic peptides. The method defines phosphorylation without having to measure the phosphorylated peptides directly or being affected by variable miscleavage. Measurement of phosphorylation is shown to be linear (relative standard error <5%) with a detection limit of <10%.

| S-EPMC3050605 | biostudies-literature

3,4-Methylenedioxypyrovalerone (MDPV) and metabolites quantification in human and rat plasma by liquid chromatography-high resolution mass spectrometry.

Project description:Synthetic cathinones are recreational drugs that mimic the effects of illicit stimulants like cocaine, amphetamine or Ecstasy. Among the available synthetic cathinones in the United States, 3,4-methylenedioxypyrovalerone (MDPV) is commonly abused and associated with dangerous side effects. MDPV is a dopamine transporter blocker 10-fold more potent than cocaine as a locomotor stimulant in rats. Previous in vitro and in vivo studies examining MDPV metabolism reported 3,4-dihydroxypyrovalerone (3,4-catechol-PV) and 4-hydroxy-3-methoxypyrovalerone (4-OH-3-MeO-PV) as the two primary metabolites. We developed and validated a liquid chromatography-high resolution mass spectrometry method to quantify MDPV and its primary metabolites in 100 μL human and rat plasma. Plasma hydrolysis was followed by protein precipitation before analysis. Limits of detection were 0.1 μg L(-1), with linear ranges from 0.25 to 1000 μg L(-1). Process efficiency, matrix effect, total imprecision (%CV) and accuracy (%target) were 36-93%, from -8 to 12%, 2.1 to 7.3% and 86 to 109%, respectively. MDPV and metabolites were stable at room temperature for 24 h, 4 °C for 72 h and after 3 freeze-thaw cycles with less than 10% variability. Human-rat plasma cross validation demonstrated that rat plasma could be accurately quantified against a human plasma calibration curve. As proof of this method, rat plasma specimens were analyzed after intraperitoneal and subcutaneous dosing with MDPV (0.5 mg kg(-1)). MDPV, 3,4-catechol-PV and 4-OH-3-MeO-PV concentrations ranged from not detected to 107.5 μg L(-1) prior to and up to 8h after dosing. This method provides a simultaneous quantification of MDPV and two metabolites in plasma with good selectivity and sensitivity.

| S-EPMC4150738 | biostudies-literature

Accurate and reliable quantification of 25-hydroxy-vitamin D species by liquid chromatography high-resolution tandem mass spectrometry.

Project description:In general, mass spectrometric quantification of small molecules in routine laboratory testing utilizes liquid chromatography coupled to low mass resolution triple-quadrupole mass spectrometers (QQQs). Here we introduce high-resolution tandem mass spectrometry (quadrupole-Orbitrap) for the quantification of 25-hydroxy-vitamin D [25(OH)D], a marker of the vitamin D status, because the specificity of 25(OH)D immunoassays is still questionable and mass spectrometric quantification is becoming increasingly important. Liquid chromatography coupled to high-resolution tandem mass spectrometry (LC-MS/HR-MS) was used to quantify 25-hydroxy-cholecalciferol [25(OH)D3], 25-hydroxy-ergocalciferol [25(OH)D2], and their C3-epimers 3-epi-25(OH)D3 and 3-epi-25(OH)D2. The method has a run time of 5 min and was validated according to the US Food and Drug Administration and the European Medicines Agency guidelines. High mass resolution was advantageously applied to separate a quasi-isobaric interference of the internal standard D6-25(OH)D2 with 3-epi-25(OH)D3. All analytes showed an imprecision of below 10% coefficient of variation (CV), trueness between 90% and 110%, and limits of quantification below 10 nM. Concentrations measured by LC-MS/HR-MS are in good agreement with those of the National Institute of Standards and Technology reference methods using LC-MS/MS (QQQ). In conclusion, quantification of 25(OH)D by LC-MS/HR-MS is applicable for routine testing and also holds promise for highly specific quantification of other small molecules.

| S-EPMC4442879 | biostudies-literature

Ultra-High-Performance Liquid Chromatography-Electrospray Ionization-Mass Spectrometry for High-Neuroanatomical Resolution Quantification of Brain Estradiol Concentrations.

Project description:Ventromedial hypothalamic nucleus (VMN) control of glucostasis is estradiol (E-2)-dependent. E-2 regulation of VMN reactivity to hypoglycemia may involve changes in signal volume due to altered aromatase expression. Here, high-resolution micropunch dissection tools for isolation of segmental VMN tissue were used with Design of Experiments-refined uHPLC-electrospray ionization-mass spectrometry (LC-ESI-MS) methodology to investigate the premise that effects of acute and/or recurring hypoglycemia on VMN E-2 content are sex-dimorphic. Relationships among multiple independent mass spectrometric operational variables were assessed by Central Composite Design (CCD) to amplify E-2 chromatogram area. Combinations of spectrometric temperature and gas pressure variable combinations were screened by Akaike Information Criterion correction modeling. A Fibonacci Sequence design using CCD minimum and maximal variable limits produced a small-run model that replicated maximal response from CCD. E-2 chromatographic response was further enhanced by optimization of solid phase extraction and instrument source and collision-induced dissociation voltages. In male rats, acute and chronic hypoglycemia respectively elevated or diminished E-2 concentrations relative to baseline in both rostral and caudal VMN. However, females exhibited regional variability in tissue E-2 profiles during acute (increased, rostral VMN; no change, caudal VMN) and recurring (no change, rostral VMN; increased, caudal VMN) hypoglycemia. Outcomes demonstrate requisite LC-ESI-MS sensitivity for E-2 quantification in small-volume brain tissue samples acquired with high-neuroanatomical specificity. Current methodology will facilitate efforts to investigate physiological consequences of VMN rostro-caudal segment-specific acclimation of E-2 profiles to recurring hypoglycemia, including effects on gluco-regulatory function, in each sex.

| S-EPMC7581564 | biostudies-literature

Quantification of regional DNA methylation by liquid chromatography/tandem mass spectrometry.

Project description:Promoter hypermethylation-associated tumor suppressor gene (TSG) silencing has been explored as a therapeutic target for hypomethylating agents. Promoter methylation change may serve as a pharmacodynamic endpoint for evaluation of the efficacy of these agents and predict the patient's clinical response. Here a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay has been developed for quantitative regional DNA methylation analysis using the molar ratio of 5-methyl-2'-deoxycytidine (5mdC) to 2'-deoxycytidine (2dC) in the enzymatic hydrolysate of fully methylated bisulfite-converted polymerase chain reaction (PCR) amplicons as the methylation indicator. The assay can differentiate 5% of promoter methylation level with an intraday precision ranging from 3 to 16% using two TSGs: HIN-1 and RASSF1A. This method was applied to characterize decitabine-induced promoter DNA methylation changes of these two TSGs in a breast cancer MCF-7 cell line. Promoter methylation of these TSGs was found to decrease in a dose-dependent manner. Correspondingly, the expression of these TSGs was enhanced. The sensitivity and reproducibility of the method make it a valuable tool for specific gene methylation analysis that could aid characterization of hypomethylating activity on specific genes by hypomethylating agents in a clinical setting.

| S-EPMC3939067 | biostudies-literature

Automated annotation and quantification of glycans using liquid chromatography-mass spectrometry.

Project description:UnlabelledAs a common post-translational modification, protein glycosylation plays an important role in many biological processes, and it is known to be associated with human diseases. Mass spectrometry (MS)-based glycomic profiling techniques have been developed to measure the abundances of glycans in complex biological samples and applied to the discovery of putative glycan biomarkers. To automate the annotation of glycomic profiles in the liquid chromatography-MS (LC-MS) data, we present here a user-friendly software tool, MultiGlycan, implemented in C# on Windows systems. We tested MultiGlycan by using several glycomic profiling datasets acquired using LC-MS under different preparations and show that MultiGlycan executes fast and generates robust and reliable results.AvailabilityMultiGlycan can be freely downloaded at http://darwin.informatics.indiana.edu/MultiGlycan/.Supplementary informationSupplementary data are available at Bioinformatics online.

| S-EPMC4542666 | biostudies-literature

Liquid Chromatography Tandem Mass Spectrometry Quantification of 13C-Labeling in Sugars.

Project description:Subcellular compartmentation has been challenging in plant 13C-metabolic flux analysis. Indeed, plant cells are highly compartmented: they contain vacuoles and plastids in addition to the regular organelles found in other eukaryotes. The distinction of reactions between compartments is possible when metabolites are synthesized in a particular compartment or by a unique pathway. Sucrose is an example of such a metabolite: it is specifically produced in the cytosol from glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P). Therefore, determining the 13C-labeling in the fructosyl and glucosyl moieties of sucrose directly informs about the labeling of cytosolic F6P and G6P, respectively. To date, the most commonly used method to monitor sucrose labeling is by nuclear magnetic resonance, which requires substantial amounts of biological sample. This study describes a new methodology that accurately measures the labeling in free sugars using liquid chromatography tandem mass spectrometry (LC-MS/MS). For this purpose, maize embryos were pulsed with [U-13C]-fructose, intracellular sugars were extracted, and their time-course labeling was analyzed by LC-MS/MS. Additionally, extracts were enzymatically treated with hexokinase to remove the soluble hexoses, and then invertase to cleave sucrose into fructose and glucose. Finally, the labeling in the glucosyl and fructosyl moieties of sucrose was determined by LC-MS/MS.

| S-EPMC7022953 | biostudies-literature

Combined liquid chromatography-tandem mass spectrometry analysis of progesterone metabolites.

Project description:Progesterone has a number of important functions throughout the human body. While the roles of progesterone are well known, the possible actions and implications of progesterone metabolites in different tissues remain to be determined. There is a growing body of evidence that these metabolites are not inactive, but can have significant biological effects, as anesthetics, anxiolytics and anticonvulsants. Furthermore, they can facilitate synthesis of myelin components in the peripheral nervous system, have effects on human pregnancy and onset of labour, and have a neuroprotective role. For a better understanding of the functions of progesterone metabolites, improved analytical methods are essential. We have developed a combined liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for detection and quantification of progesterone and 16 progesterone metabolites that has femtomolar sensitivity and good reproducibility in a single chromatographic run. MS/MS analyses were performed in positive mode and under constant electrospray ionization conditions. To increase the sensitivity, all of the transitions were recorded using the Scheduled MRM algorithm. This LC-MS/MS method requires small sample volumes and minimal sample preparation, and there is no need for derivatization. Here, we show the application of this method for evaluation of progesterone metabolism in the HES endometrial cell line. In HES cells, the metabolism of progesterone proceeds mainly to (20S)-20-hydroxy-pregn-4-ene-3-one, (20S)-20-hydroxy-5?-pregnane-3-one and (20S)-5?-pregnane-3?,20-diol. The investigation of possible biological effects of these metabolites on the endometrium is currently undergoing.

| S-EPMC4332660 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data