A Study of the Spatial Distribution of Gossypol and Other Terpenoids in Cotton Leaves and Ovules

Significance of the topic

Plant terpenoids are crucial for defense and signaling; imaging their in situ distribution at cellular resolution informs biosynthetic and functional studies.

Objectives and study overview

This study employed Shimadzu’s iMScope QT to map the spatial distribution of gossypol and related terpenoids in cotton leaves and ovules,
evaluating tissue-specific accumulation and comparing glandular profiles between organs.

Methodology and instrumentation

The experimental workflow included:

• Sample preparation: Leaf and ovule sections (10 µm) were obtained from gelatin-embedded, frozen samples using a cryomicrotome at –20 °C.
• Matrix application: α-Cyano-4-hydroxycinnamic acid (CHCA) was sublimated onto ITO slides (0.7 µm coating), followed by methanol vapor recrystallization.
• Instrumentation: iMScope QT combining optical microscopy, an AP-MALDI source, and a Q-TOF analyzer enabled integrated high-resolution imaging.
• Data acquisition: Negative-ion mode (m/z 150–650) with 5×5 µm pixel pitch, 5 µm laser diameter, 700 shots per pixel at 2000 Hz.
• Data analysis: Mass images were processed in IMAGEREVEAL MS; regions of interest (ROIs) within five glands per tissue were quantified.

Key results and discussion

• High-resolution MS images (5 µm) revealed that gossypol, hemigossypol, and hemigossypolone localize predominantly to lysigenous glands.
• Gossypol and hemigossypol levels were substantially higher in ovule glands (average gossypol intensity ~148 856) than in leaf glands (~23 654), consistent with prior reports.
• Conversely, hemigossypolone accumulated more in leaf glands (average ~13 471) than in ovule glands (~145).
• Gland size correlated with compound intensity, and detailed ROI images showed heterogeneous versus uniform distributions in leaf versus ovule glands, respectively.

Benefits and practical applications of the method

• Combines morphological observation with chemical imaging for precise localization of metabolites at cellular resolution.
• Enables comparative analysis of compound distribution across tissues, aiding functional metabolomics and QA/QC in plant sciences.
• High sensitivity and spatial resolution facilitate studies of low-abundance metabolites in microscopic structures.

Future trends and potential applications

• Integration with multimodal imaging could further link metabolite patterns to gene expression and protein localization.
• Application to other plant species and stress conditions may uncover dynamic changes in terpenoid biosynthesis.
• Advances in matrix application and ionization techniques could enhance resolution and sensitivity for broader metabolite classes.

Conclusion

iMScope QT provides a robust platform for high-resolution, in situ metabolite imaging, elucidating the spatial distribution of terpenoids in cotton tissues and guiding future studies on their biosynthesis and physiological roles.

Reference

• Mol Plant. 2023, 16(12):1990–2003