High mass and spatial resolution MS imaging of soft fruit sample cross-sections using desorption electrospray ionization (DESI)

Importance of the topic

Desorption electrospray ionization (DESI) mass spectrometry imaging (MSI) enables ambient, high-spatial-resolution chemical mapping of biological tissues without extensive sample preparation. Applying DESI-MSI to soft fruits (high water content tissues such as strawberries and raspberries) addresses a practical analytical challenge: preserving spatial metabolite localization while extracting a chemically representative set of small molecules for high mass‑resolution detection. Reliable workflows for imaging soft fruits are valuable for food quality control, plant physiology studies, breeding, and metabolomics-driven phenotyping.

Study objectives and overview

This work evaluated a simple direct-transfer approach to prepare cross-sections of fresh strawberries and raspberries for DESI-MSI and compared three receiving surfaces (plain glass slides, nitrocellulose membrane on glass, and Hamamatsu Poropore/Poropare plates). The goals were to determine which substrate best preserves spatial distribution of metabolites, which provides the broadest metabolite coverage, and how mass‑resolution capability aids in tentative assignment of detected species.

Methodology

Fresh fruit (strawberries and raspberries) were cut through the centre to expose cross-sections and transferred directly to the receiving surface by a short transfer (approximately 10 seconds). After drying, samples were imaged by DESI-MSI in negative ion mode. Key acquisition parameters included a 50 µm pixel size and a spray solvent of 95:5 methanol:water. Sucrose (m/z 341.1089) was used as an internal lockmass for high mass accuracy. Imaging emphasized high mass resolution to permit tentative metabolite annotation based on exact mass.

Used instrumentation

• DESI source: Waters DESI XS ambient ionization source.
• Mass spectrometer: Waters Xevo MRT (P10) operated in negative ion mode.
• Spray solvent: 95:5 MeOH:H2O.
• Imaging parameters: 50 µm pixel size; sucrose m/z 341.1089 as lockmass.
• Receiving substrates tested: plain glass microscope slides, nitrocellulose membrane (0.2 µm pore) mounted on glass, and Hamamatsu Poropore transfer plates.

Main results and discussion

• Substrate-dependent delocalization and sensitivity: Glass slides produced strong signal intensities but showed pronounced delocalization of metabolites—spatial features were blurred during the liquid-mediated transfer. This delocalization was evident both visually and from MSI distributions.
• Poropore (Hamamatsu) plates best preserved spatial detail: Imaging from Poropore plates exhibited minimal metabolite delocalization, allowing clear resolution of internal tissue structures and fine chemical heterogeneity. However, overall signal intensities for detected metabolites were lower on Poropore compared to glass.
• Nitrocellulose membrane increased metabolite transfer breadth but suffered mechanical fragility: Nitrocellulose transfers enabled observation of complementary metabolites not prominent on the other surfaces and appeared to extract a broader set of analytes. The trade-off was increased delocalization relative to Poropore plates and practical issues with substrate integrity—membranes were damaged after a single DESI run.
• High mass resolution enabled tentative metabolite annotations: Using accurate mass, several fruit metabolites were tentatively assigned, including citric acid (m/z ~191.0196), asparagine (m/z ~131.0457–131.0459), sucrose (m/z 341.1089), malic acid (m/z ~133.0138), glutamic acid (m/z ~146.0454), and long‑chain fatty acids such as palmitic (m/z ~255.2329) and stearic acid (m/z ~283.2643). Spatial overlays of selected ions illustrated differences in tissue localization (e.g., sugars and organic acids in distinct regions).

Benefits and practical applications of the method

• Enables MSI of previously challenging, high-water-content plant tissues without complex cryo-embedding or sectioning protocols by using a rapid direct-transfer approach.
• Poropore plates provide a practical compromise when the priority is preserving spatial localization, useful for mapping metabolites at tissue-level resolution in studies of ripening, nutrient distribution, or infection responses.
• Nitrocellulose transfers can be exploited when maximum metabolite coverage is desired (e.g., exploratory metabolomics), but users should account for increased delocalization and substrate fragility.
• High mass resolution imaging supports tentative compound identification directly from images, improving confidence in chemical mapping even in complex plant matrices.

Future trends and potential uses

• Optimization of transfer protocols: Systematic variation of transfer time, pressure, and drying conditions could reduce delocalization while maximizing signal intensity.
• Substrate engineering: Development of more robust membranes or porous plates that combine the extraction efficiency of nitrocellulose with the localization preservation of Poropore surfaces.
• Integration with MS/MS and ion mobility: Adding on‑line fragmentation and orthogonal separation will allow definitive identifications and disentangling of isobaric species detected in situ.
• Quantitative workflows: Calibration strategies and internal standards compatible with DESI imaging would move applications from qualitative mapping to quantitative spatial metabolomics for food quality and regulatory testing.
• Broader applications: Plant breeding, provenance and authenticity studies, assessment of post‑harvest changes and stress responses, and spatially resolved pesticide/metabolite residue analysis.

Conclusion

A direct-transfer DESI-MSI workflow permits imaging of soft fruit cross-sections with high mass resolution. Among the evaluated receiving surfaces, Hamamatsu Poropore plates best preserved metabolite localization at the spatial scale used (50 µm), while nitrocellulose membranes provided broader metabolite extraction but were physically fragile and caused greater delocalization. Glass produced stronger signals but substantial delocalization. High mass accuracy enabled plausible assignments of key fruit metabolites, demonstrating the method's utility for spatial metabolomics in soft, high‑water-content plant tissues. Choice of substrate therefore depends on whether localization fidelity or metabolite breadth is the primary goal.

References

• Rao W., Scanlan C., Towers M., Ballantyne J., Molyneux G. High mass and spatial resolution MS imaging of soft fruit sample cross-sections using desorption electrospray ionization (DESI). Waters Corporation technical poster/report. 2026.