Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Mass Spectrometry: Complex Analysis01:21

Mass Spectrometry: Complex Analysis

647
Mass spectrometry is an important technique for the identification of pure compounds. However, it has some limitations for the analysis of complex mixtures, often due to excessive fragmentation making the spectrum too complicated to decipher. Mass spectrometry can be combined with suitable separation methods in sequence, forming hyphenated methods, which are useful in the analysis of complex mixtures.
GC–MS is a powerful hyphenated method commonly used in forensics and environmental...
647
Peptide Identification Using Tandem Mass Spectrometry01:33

Peptide Identification Using Tandem Mass Spectrometry

6.2K
Tandem mass spectrometry, also known as MS/MS or MS2, is an analytical technique that employs two mass analyzers. Essentially it is a series of mass spectrometers that helps isolate a particular biomolecule and then helps study its chemical properties.
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...
6.2K

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Comparison of Mucosal and Faecal Microbiomes in Patients With Cirrhosis.

Gastroenterology research and practice·2026
Same author

Elevated Relative Levels of the C-3 Epimer of 25-Hydroxyvitamin D in Patients with Cirrhosis.

Nutrients·2026
Same author

Continuous Flow Paper Spray Ionization Mass Spectrometry for In-Depth Characterization of Anticancer Drugs in Tissues: Addressing Mass Spectral Complexity.

Journal of the American Society for Mass Spectrometry·2026
Same author

Tracking the Changes of DOM Composition, Transformation, and Cycling Mechanism Triggered by the Priming Effect: Insights from Incubation Experiments.

Environmental science & technology·2024
Same author

Measurement of Serum Free Vitamin D Concentrations: Importance, Challenges, and the Emerging Role of Mass Spectrometry.

Clinical chemistry·2024
Same author

Impact of Hydrothermal Fluids on Hydrocarbon Generation and Solid Bitumen Formation in the Kongdian Formation, Huanghua Depression, China.

Journal of the American Society for Mass Spectrometry·2024

Related Experiment Video

Updated: May 7, 2025

Workflow Based on the Combination of Isotopic Tracer Experiments to Investigate Microbial Metabolism of Multiple Nutrient Sources
12:47

Workflow Based on the Combination of Isotopic Tracer Experiments to Investigate Microbial Metabolism of Multiple Nutrient Sources

Published on: January 22, 2018

9.2K

Streamlined Vitamin D Metabolite Fingerprinting Analysis Using Isotope-Coded Multiplexing MS with Cost-Effective

Pascal Schorr1, Caroline S Stokes2, Dietrich A Volmer1

  • 1Department of Chemistry, Humboldt Universität zu Berlin, Brook-Taylor-Str. 2, Berlin 12489, Germany.

ACS Omega
|January 1, 2025
PubMed
Summary

This study introduces routine isotope-coded multiplex derivatization for vitamin D metabolites, enabling simultaneous analysis of two samples. The method improves separation of key vitamin D epimers on standard C-18 columns for clinical applications.

More Related Videos

Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization
12:11

Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization

Published on: February 27, 2020

6.7K
Automated Sample Multiplexing by using Combined Precursor Isotopic Labeling and Isobaric Tagging cPILOT
09:24

Automated Sample Multiplexing by using Combined Precursor Isotopic Labeling and Isobaric Tagging cPILOT

Published on: December 18, 2020

5.5K

Related Experiment Videos

Last Updated: May 7, 2025

Workflow Based on the Combination of Isotopic Tracer Experiments to Investigate Microbial Metabolism of Multiple Nutrient Sources
12:47

Workflow Based on the Combination of Isotopic Tracer Experiments to Investigate Microbial Metabolism of Multiple Nutrient Sources

Published on: January 22, 2018

9.2K
Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization
12:11

Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization

Published on: February 27, 2020

6.7K
Automated Sample Multiplexing by using Combined Precursor Isotopic Labeling and Isobaric Tagging cPILOT
09:24

Automated Sample Multiplexing by using Combined Precursor Isotopic Labeling and Isobaric Tagging cPILOT

Published on: December 18, 2020

5.5K

Area of Science:

  • Clinical Chemistry
  • Analytical Chemistry
  • Mass Spectrometry

Background:

  • Accurate analysis of vitamin D and its metabolites is crucial for clinical diagnostics.
  • Existing methods for vitamin D metabolite analysis face challenges in multiplexing and separating critical epimers.
  • Conventional separation techniques often require specialized columns (e.g., PFP) with limited long-term stability.

Purpose of the Study:

  • To develop the first routine isotope-coded multiplex derivatization method for vitamin D and its metabolites.
  • To enable simultaneous analysis of two samples in a single liquid chromatography-tandem mass spectrometry (LC-MS/MS) run.
  • To improve the chromatographic separation of 25-hydroxyvitamin D3 epimers on conventional C-18 columns.

Main Methods:

  • Extended a one-pot double derivatization reaction using commercial reagents.
  • Employed a Diels-Alder reaction with 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) followed by acetylation with acetic anhydride (or its d3-isotopologue).
  • Utilized C-18 column with methanol/water gradient elution for enhanced separation of vitamin D3 metabolites.

Main Results:

  • Established a routine isotope-coded multiplex derivatization enabling simultaneous measurement of two samples.
  • Achieved baseline separation of critical vitamin D3 metabolites (24,25(OH)2D3, 3α-25(OH)D3, 3β-25(OH)D3, and vitamin D3) on a C-18 column.
  • Demonstrated method applicability by testing on 40 serum samples from patients with chronic liver diseases (CLD).

Conclusions:

  • The developed duplex derivatization method offers a robust and routine approach for clinical vitamin D analysis.
  • This method overcomes limitations of specialized columns and enhances the separation of challenging epimers.
  • The isotope-coded multiplexing strategy facilitates higher throughput and efficiency in clinical laboratories.