Enhancing natural products extraction:using metabolomics for evaluating analytical methods

www.helmholtz-hips.de Abstract Microbial natural products are historically known for their role in antibiotic discovery and potential for other drugs development [1]. The success of isolating new microbial natural products depends not only on enhancing biotechnological cultivation techniques, but also on extraction and analytical methods [2]. From this perspective, alternative methods are being explored both to maximize the extraction efficiency of secondary metabolites and to achieve environmentally friendly processes. To assess if a greener technique can substitute or complement traditional approaches, we employ here a metabolomics-based approach for evaluating alternative methodologies from the natural products perspective. In this context, techniques that aim to diminish the quantity of solvent use, and therefore being eco-friendlier, were chosen for the current study. This poster outlines two experimental setups in which metabolomics is used to evaluate extraction and analytical analysis of bacterial crude extracts, respectively. To address the extraction process, a microreactor is being tested for facilitating the extraction of endometabolites by disrupting cells without the need of solvent use. For the analytical step, a supercritical fluid chromatography method –UltraPerformance Convergence Chromatography (UPC²) –coupled to mass spectrometry (MS) is compared to a conventional UltraHigh Performance Liquid Chromatography (UHPLC) coupled to mass spectrometry-system for analysing myxobacterial crude extracts. Acknowledgements This poster is part of the project GreenX3 - funded by the European Union Horizon Europe MSCA Doctoral Network program under EC Grant Agreement 101120061 and by UK Research and Innovation (UKRI) under the UK government’s Horizon Europe funding guarantee [grant number EP/Y032039/1]. Enhancing natural products extraction: using metabolomics for Elisa de Lannoy Guerra1,Simon Cremers2, Felix Penth3, Daniela Becher3, Ronald Garcia2, Alexander Voltz2, Sebastian Walesch2, Chantal Bader2, Daniel Krug1,2, Rolf Müller1,2 1Universität des Saarlandes, Department of Pharmacy, Campus E8.1, 66123 Saarbrücken, Germany 2Department of Microbial Natural Products, Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS), Helmholtz Centre for Infection Research (HZI) and Department of Pharmacy at Saarland University, Campus E8.1, 66123 Saarbrücken, Germany ³ MyBiotech GmbH, Industriestraße 1b, 66802 Überherrn, Germany References [1] Hutchings MI, Truman AW, Wilkinson B. Antibiotics: past, present and future. Curr Opin Microbiol. 2019;51:72–80. doi:10.1016/j.mib.2019.10.008 [2] Bader CD, Neuber M, Panter F, Krug D, Müller R. Supercritical fluid extraction enhances discovery of secondary metabolites from myxobacteria. Anal Chem. 2020;92(23):15403–11.doi:10.1021/acs.analchem.0c02995 [3] Workflow created with Biorender.com [4]MyBiotech GmbH. Microjet Reactor (MJR) [Image]. Available at: https://mybiotech.de/mybiotech/ (Accessed: 17 April 2025). [5] Baumstümmler, B., Türeli, A. E., Penth, F., Müller, D. & Krumbholz, R.,2020.EP 3 938 088 B1. European Patent Office MICROBIAL NATURAL PRODUCTS ANALYSIS WORKFLOW EXTRACTION EVALUATION Traditional extraction Adding an extraction facilitator step previously + Sample after MJR treatment Sample before MJR treatment MicroJet Reactor (MJR) [4,5] VS + [3] CRUDE EXTRACT ANALYSIS METABOLOMICS DATA-ANALYSIS OUTCOME 1: Analysis of chromatogram + m/z spectrogram 2. Identification of compounds (annotation) Crude extracts 3. Comparison of number of features 4. Group different features/ Statistical tools for visualization Understanding when a greener method can substitute or enhance extraction and identification of natural products Understanding which type of natural products supercritical fluid chromatography techniques can highlight To facilitate the discovery of a new microbial natural product Ultimate goal: To find a new molecular structure with the potetial to the development of novel antibiotic Cultivation Harvest Extraction Crude Extract Analysis Fractionation Isolation Structural Elucidation evaluating analytical methods Microscope image at 40x magnification LC-MS Chromatogram of crude extract samples with and without previous MJR treatment overlapped. In red: sample undergone MJR process, in gray: sample without MJR process. OUTCOME Understanding when greener methods can enhance extraction and identification of natural products Understanding which type of natural products supercritical fluid chromatography can highlight To facilitate the discovery of a new microbial natural product To find a new molecular structure with the potential to the development of novel antibiotic Ultimate goal