Identifying TB Markers Using UHPLC–IMS-HRMS and Non-Invasive Skin Sampling

A team of scientists from the University of Pretoria in South Africa, developed a non-invasive approach utilizing ultrahigh-performance liquid chromatography–ion mobility spectrometry-high-resolution mass spectrometry (UHPLC–IMS-HRMS) for the identification of biological markers for tuberculosis (TB) diagnosis using human skin (1). Their findings were published in the Journal of Chromatography Open.

Pattern of human skin © PixieMe - stock.adobe.com

Tuberculosis is one of the world's deadliest infectious diseases, claiming millions of lives annually, despite being preventable and curable with appropriate treatment. According to the World Health Organization (WHO), TB caused 1.23 million deaths in 2023 alone, making it the deadliest infectious disease in the world, after several years when it was replaced by COVID-19 (2). Individuals with weakened immune systems face a significantly higher risk of developing TB-related illness. For example, someone living with HIV is approximately 13 times more likely to develop active TB (3). Early and accurate diagnosis is crucial to combating TB, yet current diagnostic methods remain expensive, invasive, and time-consuming. As TB continues to pose a major global health challenge, more accessible, cost-effective, and non-invasive diagnostic techniques are increasingly required.

One promising area of research involves metabolomics, the study of small molecules known as metabolites found in biological samples. Metabolomics offers a potential avenue for non-invasive TB diagnostics by analyzing biomarkers present in various bodily fluids, including blood, urine, breath, and skin. Recent advancements in analytical techniques, particularly HRMS combined with ion mobility spectrometry (IMS), have allowed researchers to identify and validate metabolites with a higher degree of confidence (4).

In this current study, a novel wearable polydimethylsiloxane (PDMS) sampler, designed for the passive collection of metabolites from the skin surface, was employed. The sampler works by concentrating chemical compounds emitted through the skin, which are then desorbed into solvent within a liquid chromatography (LC) vial for further analysis. This process eliminates the need for complex extraction techniques and allows for the quick and efficient analysis of metabolites directly from the skin. HRMS allows for the precise identification of metabolites by measuring their mass-to-charge ratio, while IMS provides data on the collision cross section (CCS) of molecules. This allows the researchers to align product and precursor ions, improving the accuracy of biomarker identification.

By employing this non-invasive method, the researchers were able to identify 14 potential biomarkers that could distinguish active TB patients from healthy controls. Among these biomarkers, para-aminobenzoic acid (PABA) and 2,6-diethylaniline emerged as promising candidates. PABA demonstrated a specificity of 1.0, sensitivity of 0.9, and an area under the curve (AUC) of 0.961. To the best of the authors’ knowledge, 2,6-diethylaniline has never been reported as a potential TB marker (1).

The method provided limits of detection ranging from 6 ng for PABA to 172 ng for phenylalanine. The method’s calibration range of 10–800 ng and a high R² value (≥0.99) indicate robust reproducibility and reliability. These findings highlight the potential of this non-invasive skin sampling method to be used in a diagnostic setting, offering an effective and accurate alternative to current invasive TB tests.

Skin surface sampling offers several advantages over traditional diagnostic approaches, such as breath and sputum testing, which require more complex and often invasive procedures. Additionally, analyzing skin metabolites avoids some of the challenges associated with breath-based testing, which can be influenced by environmental factors and may raise concerns in infectious settings.

Traditional metabolomic studies have primarily focused on volatile organic compounds (VOCs) found in breath and urine, but this study demonstrated that skin surface compounds could provide just as much diagnostic potential by highlighting that skin semi-volatile and non-volatile compounds can effectively distinguish between individuals who are TB-positive and those who are TB-negative (1).

While these initial results are promising, the team has stressed the need for further research, particularly with larger patient cohorts, to confirm the reliability of these biomarkers. The findings suggest that skin sampling, coupled with the appropriate analytical tools, could be integrated into routine TB diagnostic protocols in the future.

References

(1) Wooding, M.; van Pletzen, K.; Naudé, Y. Identifying Skin Surface Chemicals as Potential Tuberculosis Diagnostic Biomarkers Using Ultra Performance Liquid Chromatography-High Resolution Mass Spectrometry. J. Chromatogr. Open 2025, 7, 100204. DOI: 10.1016/j.jcoa.2025.100204

(2) WHO, Global Tuberculosis Report 2024, https://iris.who.int/bitstream/handle/10665/379339/9789240101531-eng.pdf?sequence=1 (accessed 2025-04-17).

(3) 10 Facts on Tuberculosis; WHO website. https://www.who.int/news-room/facts-in-pictures/detail/tuberculosis(accessed 2025-04-17).

(4) Wooding, M.; Dodgen, T.; Rohwer, E. R.; Naudé, Y. Advancing the Analytical Toolkit in the Investigation of Vector Mosquito Host Biting Site Selection. J. Mass Spectrom. 2023, 59, e4992. DOI: 10.1002/jms.4992