Surreal illustration of metabolite analysis with isotopes and chromatography.

Unlock Metabolite Secrets: How HILIC-NMR and Isotope Tagging Revolutionize Analysis

Rhea Morgan in Science & Nature August 2025 • 4 min read.

"A cutting-edge approach combines advanced chromatography and isotopic labeling to identify crucial metabolites, even at low concentrations, transforming our understanding of health and disease."

The ability to accurately measure small molecules in biological samples is key to understanding the body and disease. Scientists are constantly looking for better ways to identify and measure these metabolites, which can act as indicators of health and disease. This is where metabolomics comes in, offering a detailed snapshot of the body's chemical processes.

Nuclear Magnetic Resonance (NMR) spectroscopy has long been a staple tool in metabolomics. It's known for being accurate and non-destructive. However, NMR struggles with sensitivity, making it hard to detect all the metabolites present, especially those at very low concentrations. This limitation can make it difficult to get a complete picture of what's happening in a biological sample.

To overcome these limitations, researchers have combined NMR with other techniques, such as chromatography. This combination helps to simplify complex samples before NMR analysis. One such approach involves Hydrophilic Interaction Chromatography (HILIC), which is particularly good at separating polar metabolites. This article explores a new approach that combines HILIC with isotope tagging to boost the detection and identification of specific metabolites, opening new doors for biological research.

HILIC-NMR and Isotope Tagging: A Powerful Combination

Surreal illustration of metabolite analysis with isotopes and chromatography.

The study highlights a method that combines two-dimensional Hydrophilic Interaction Chromatography (HILIC) with isotope-tagged NMR to identify carboxyl-containing metabolites in human urine. This approach leverages the separation capabilities of HILIC and the enhanced detection offered by isotope tagging. By tagging metabolites with 15N-ethanolamine, researchers can selectively target and identify carboxyl groups, even at low concentrations.

The process involves several key steps:

Isotope Tagging: Metabolites are labeled with 15N-ethanolamine, which selectively binds to carboxyl groups.
HILIC Separation: The labeled sample is then separated using two-dimensional HILIC to isolate compounds based on their polarity.
NMR Analysis: The separated fractions are analyzed using 2D HSQC NMR to identify the tagged metabolites.

This combination allows for the identification of metabolites that might otherwise be missed due to low concentrations or overlapping signals. The two-dimensional HILIC separation further simplifies the sample, reducing the complexity of the NMR spectra and making it easier to identify individual compounds.

New Avenues in Metabolite Research

The research demonstrates the effectiveness of combining HILIC separation with isotope tagging for identifying metabolites in complex biological samples. This method not only improves the detection of low-concentration metabolites but also enhances the accuracy of metabolite identification.

By improving metabolite identification and quantification, this approach opens up new avenues for research in various fields, including biomarker discovery and personalized medicine. Identifying unique metabolic signatures can help in the early diagnosis of diseases and the development of targeted therapies.

While this study focuses on urine samples, the same approach could be applied to other biological fluids and tissue samples. As technology advances, combining HILIC-NMR with micro-coil NMR probes could further improve the sensitivity and detection limits, allowing for even more detailed metabolic profiling.

About this Article -

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This article is based on research published under:

DOI-LINK: 10.3390/metabo3030575, Alternate LINK

Title: Combining Hydrophilic Interaction Chromatography (Hilic) And Isotope Tagging For Off-Line Lc-Nmr Applications In Metabolite Analysis

Subject: Molecular Biology

Journal: Metabolites

Publisher: MDPI AG

Authors: Emmanuel Appiah-Amponsah, Kwadwo Owusu-Sarfo, G.A. Gowda, Tao Ye, Daniel Raftery

Published: 2013-07-18

Everything You Need To Know

What is metabolomics and why is it important?

Metabolomics is the comprehensive study of small molecules, known as metabolites, within a biological sample. These metabolites offer a snapshot of the chemical processes within the body and can serve as indicators of health and disease. Analyzing these molecules provides critical insights into the inner workings of biological systems.

What is the role of Nuclear Magnetic Resonance (NMR) in the analysis?

Nuclear Magnetic Resonance (NMR) spectroscopy is a non-destructive analytical technique used in metabolomics to identify and quantify metabolites. It is highly accurate but can lack sensitivity, making it challenging to detect metabolites present at low concentrations. This limitation can obscure a comprehensive understanding of the metabolic landscape.

How does Hydrophilic Interaction Chromatography (HILIC) contribute to this process?

Hydrophilic Interaction Chromatography (HILIC) is a chromatographic technique used to separate polar metabolites based on their affinity for a hydrophilic stationary phase. Combining HILIC with other techniques, like NMR, simplifies complex samples and enhances the detection of specific metabolites. In the context of the study, two-dimensional HILIC further separates the sample, reducing the complexity of the NMR spectra.

What is the purpose of isotope tagging?

Isotope tagging is a technique used to enhance the detection of specific metabolites. In this approach, metabolites are labeled with an isotope, such as 15N-ethanolamine, which selectively binds to carboxyl groups. This tagging improves the sensitivity of detection, especially for low-concentration metabolites, and allows for easier identification through techniques like NMR. This method is instrumental in targeting and identifying specific types of metabolites.

How do HILIC-NMR and Isotope Tagging work together to advance research?

The combination of Hydrophilic Interaction Chromatography (HILIC) and Isotope Tagging with Nuclear Magnetic Resonance (NMR) represents a powerful analytical method. This approach enhances the detection and identification of metabolites, particularly carboxyl-containing ones, which can be challenging to detect at low concentrations. The two-dimensional HILIC separation further simplifies the sample, improving the accuracy of metabolite identification in biological samples like human urine. This method opens new avenues for biological research by providing a more complete picture of the metabolic landscape.