Related Experiment Video

Updated: Mar 26, 2026

A Method for Measuring Metabolism in Sorted Subpopulations of Complex Cell Communities Using Stable Isotope Tracing

A Method for Measuring Metabolism in Sorted Subpopulations of Complex Cell Communities Using Stable Isotope Tracing

Published on: February 4, 2017

3D TOCSY-HSQC NMR for Metabolic Flux Analysis Using Non-Uniform Sampling

P N Reardon¹, C L Marean-Reardon^1,2, M A Bukovec³

¹Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory , 3335 Innovation Boulevard, Richland, Washington 99352, United States.

Analytical Chemistry

|February 6, 2016

Abstract:

(13)C-Metabolic Flux Analysis ((13)C-MFA) is rapidly being recognized as the authoritative method for determining fluxes through metabolic networks. Site-specific (13)C enrichment information obtained using NMR spectroscopy is a valuable input for (13)C-MFA experiments. Chemical shift overlaps in the 1D or 2D NMR experiments typically used for (13)C-MFA frequently hinder assignment and quantitation of site-specific (13)C enrichment. Here we propose the use of a 3D TOCSY-HSQC experiment for (13)C-MFA. We employ Non-Uniform Sampling (NUS) to reduce the acquisition time of the experiment to a few hours, making it practical for use in (13)C-MFA experiments. Our data show that the NUS experiment is linear and quantitative. Identification of metabolites in complex mixtures, such as a biomass hydrolysate, is simplified by virtue of the (13)C chemical shift obtained in the experiment. In addition, the experiment reports (13)C-labeling information that reveals the position specific labeling of subsets of isotopomers. The information provided by this technique will enable more accurate estimation of metabolic fluxes in large metabolic networks.

More Related Videos

PTR-ToF-MS Coupled with an Automated Sampling System and Tailored Data Analysis for Food Studies: Bioprocess Monitoring, Screening and Nose-space Analysis

PTR-ToF-MS Coupled with an Automated Sampling System and Tailored Data Analysis for Food Studies: Bioprocess Monitoring, Screening and Nose-space Analysis

Published on: May 11, 2017

A Strategy for Sensitive, Large Scale Quantitative Metabolomics

A Strategy for Sensitive, Large Scale Quantitative Metabolomics

Published on: May 27, 2014

Related Concept Videos

2D NMR: Heteronuclear Single-Quantum Correlation Spectroscopy (HSQC)

2D NMR: Heteronuclear Single-Quantum Correlation Spectroscopy (HSQC)

Heteronuclear single-quantum correlation spectroscopy (HSQC) is a 2D NMR technique that reveals one-bond correlations between hydrogen and a heteronucleus. The HSQC experiment is similar to the heteronuclear correlation experiment (HETCOR) but is more sensitive. In the HSQC spectrum, the proton chemical shift is plotted on the horizontal F2 axis, while the 13C chemical shift is plotted on the vertical F1 axis. The corresponding proton and 13C spectra are also shown. The HSQC contour plot does...

2D NMR: Homonuclear Correlation Spectroscopy (COSY)

2D NMR: Homonuclear Correlation Spectroscopy (COSY)

Homonuclear correlation spectroscopy, or COSY, is a 2-dimensional NMR technique that provides information about coupled protons. Typically, the geminal and vicinal coupling are observed. For example, consider the COSY spectrum of ethyl acetate, where its 1D proton NMR spectrum is plotted along the vertical and horizontal axes with their corresponding chemical shift scale. Three spots on the diagonal corresponding to the three peaks in the 1D proton spectrum are called diagonal peaks. The COSY...

2D NMR: Overview of Homonuclear Correlation Techniques

2D NMR: Overview of Homonuclear Correlation Techniques

Homonuclear correlation spectroscopy (COSY) is a powerful technique used in Nuclear Magnetic Resonance (NMR) spectroscopy to study the correlations between nuclei of the same type within a molecule. It provides information about scalar couplings between adjacent nuclei, which helps determine connectivity and structural information. There are several COSY variants, each with its unique strengths and experimental parameters.
COSY90 is the standard two-dimensional (2D) COSY experiment that...

2D NMR: Overview of Heteronuclear Correlation Techniques

2D NMR: Overview of Heteronuclear Correlation Techniques

Heteronuclear correlation spectroscopy is an analytical technique that investigates the coupling between different types of nuclei, often a proton and an X-nucleus, such as carbon-13 or nitrogen-15. This method is commonly used in nuclear magnetic resonance (NMR) spectroscopy to gain insights into complex chemical compounds' structural and compositional aspects. A typical heteronuclear correlation spectrum displays X-nucleus chemical shifts on one axis and a proton spectrum on the other...

Noncompartmental Analysis: Mean Transit, Absorption and Dissolution Time

Noncompartmental Analysis: Mean Transit, Absorption and Dissolution Time

When drugs are administered extravascularly, a comprehensive evaluation through noncompartmental analysis becomes imperative. This analytical approach considers various parameters that play a crucial role in understanding the pharmacokinetics of these drugs.
One of the key parameters is the mean transit time (MTT), which refers to the total duration required for drug molecules to transit through the body. MTT is determined by calculating the ratio of the area under the moment curve to the area...

Two-Dimensional (2D) NMR: Overview

Two-Dimensional (2D) NMR: Overview

The 1D NMR spectrum of large and complex molecules like natural products has complicated splitting patterns and overlapping signals, which can be easily interpreted using 2-dimensional (2D) NMR. Unlike 1D NMR, 2D NMR has two frequency axes that provide the coupling information between the nucleus A and nucleus B in a molecule. The process from which 2D spectra are obtained has four steps.
The first step is the preparation period, during which nucleus A is excited with a radiofrequency pulse....

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

The weak interdomain coupling observed in the 70 kDa subunit of human replication protein A is unaffected by ssDNA binding.

Nucleic acids research·2001

DNA-XPA interactions: a (31)P NMR and molecular modeling study of dCCAATAACC association with the minimal DNA-binding domain (M98-F219) of the nucleotide excision repair protein XPA.

Nucleic acids research·2001

Human nucleotide excision repair protein XPA: NMR spectroscopic studies of an XPA fragment containing the ERCC1-binding region and the minimal DNA-binding domain (M59-F219).

Mutation research·2001

A comprehensive approach for accurate measurement of proton-proton coupling constants in the sugar ring of DNA.

Journal of magnetic resonance (San Diego, Calif. : 1997)·2000

Interactions of human nucleotide excision repair protein XPA with DNA and RPA70 Delta C327: chemical shift mapping and 15N NMR relaxation studies.

Biochemistry·1999

Human replication protein A: global fold of the N-terminal RPA-70 domain reveals a basic cleft and flexible C-terminal linker.

Journal of biomolecular NMR·1999

See all related articles