# In situ isobaric lipid mapping by MALDI-ion mobility separation-mass spectrometry imaging

Tingting Fu, Janina Oetjen, Manuel Chapelle, Alexandre Verdu, Matthias Szesny, Arnaud Chaumot, Davide Degli-Esposti, Olivier Geffard, Yohann Clément, Arnaud Salvador, et al.

► To cite this version:

Tingting Fu, Janina Oetjen, Manuel Chapelle, Alexandre Verdu, Matthias Szesny, et al. In situ isobaric lipid mapping by MALDI-ion mobility separation-mass spectrometry imaging. 68th Conference on Mass Spectrometry and Allied Topics, May 2020, Online, United States. , 2020. hal-03996540

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Submitted on 20 Feb 2023

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<figure><img src="image_1.png" /></figure>

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# In situ isobaric/isomeric lipid mapping by MALDI-Ion Mobility Separation-MSI

Tingting Fu<fnref n="1" />, Janina Oetjen<fnref n="2" />, Manuel Chapelle<fnref n="2" />, Alexandre Verdu<fnref n="2" />, Matthias Szesny<fnref n="2" />, Arnaud Chaumot<fnref n="3" />, Davide Degli-Esposti<fnref n="3" />, Olivier Geffard<fnref n="3" />, Yohann Clément<fnref n="1" />, Arnaud Salvador<fnref n="1" />, Sophie Ayciriex<fnref n="1" />

## Introduction

The highly diverse chemical structures of lipids make their analysis directly from biological tissue sections extremely challenging. Here we report the in-situ mapping and identification of lipids in a freshwater crustacean *Gammarus fossarum* using MALDI mass spectrometry imaging (MSI) in combination with an additional separation dimension using ion mobility spectrometry (IMS) [1]. The high-resolution trapped ion mobility spectrometry (TIMS) allowed efficient separation of isobaric/isomeric lipids showing distinct spatial distributions. The structures of the lipids were further characterized by MS/MS analysis. It is demonstrated that MALDI MSI with mobility separation is a powerful tool for distinguishing and localizing isobaric/isomeric lipids.

## Methods

The workflow for MALDI-MSI sample preparation and data analysis is outlined in Figure 1. Fresh frozen female gammarid was sectioned at 12 µm and thaw mounted onto an ITO slide (Sigma-Aldrich). After drying, sections were sprayed with 10 mg/ml DHB in ACN/H₂O/TFA (70:30:0.1, v/v/v) using a TM sprayer (HTX Technologies, Chapel Hill, NC, USA). Tissues were imaged using the following parameters if not indicated otherwise: m/z range: 100-1000, 400 shots, 10 kHz laser frequency, pitch: 20 µm. For ion mobility separation, ions were separated and eluted in the second part of the dual TIMS device using a ramp time of 300 ms and a 1/K₀ range of 0.6-1.8. Imaging data processing was performed with SCiLS Lab version 2020a. Ion mobility data was visualized with TIMS data viewer. Assignment of the ions were achieved by interrogating open source databases including METLIN [2], Lipid Maps [3] and CCS Compendium [4].

<figure><img src="image_2.png" /><figcaption>Fig. 1: MALDI-MSI workflow</figcaption></figure>

## Results

<figure><img src="image_3.png" /><figcaption>Fig. 2: Ion mobility heat map (top) and average mass spectrum (bottom) generated from the tissue section of female *G. fossarum*.</figcaption></figure>

* Good sensitivity at high lateral resolution.

* Various ion species with different 1/K₀ values detected.

* Richer molecular information compared to non-TIMS Imaging

<figure><img src="image_4.png" /><figcaption>Muscle Oocytes HP</figcaption></figure>

<figure><img src="image_5.png" /><figcaption>Fig. 3: MALDI-IMS-MSI of female *G. fossarum* tissue section with 20 µm pixel size.</figcaption></figure>

A. Optical image of the tissue after MALDI Imaging analysis. HP: hepatopancreas; In: Intestine. VP: ventral pouch.

B. Ion mobility separation of two isobaric lipid species.

C. Ion image of the ion at m/z 782.561 with 1/K₀ = 1.426 ± 0.01(CCS = 290.7 Å<fnref n="2" />).

D. Ion image of ion at m/z 782.561 with 1/K₀ = 1.402 ± 0.01 (CCS = 285.7 Å<fnref n="2" />).

E. Overlay of C and D.

<figure><img src="image_6.png" /></figure>

<table><caption>Table 1: Most intense ions colocalizing to different organs. (Assignments by interrogating METLIN [2]).</caption><thead><tr><th>Organ</th><th>m/z</th><th>Tentative assignment</th></tr></thead><tbody><tr><td rowspan="4">Muscle</td><td>713.452</td><td>PC(32:0)+K+-N(CH<sub>3</sub>)<sub>3</sub></td></tr><tr><td>772.525</td><td>PC(32:0)+K+</td></tr><tr><td>834.679</td><td>GlcCer(d18:1/24:0)+Na+</td></tr><tr><td>883.678</td><td>TAG(53:8)+Na+</td></tr><tr><td rowspan="4">Oocyte</td><td>766.574</td><td>PC(O-16:0/20:5)+H<sup>+</sup>/PC(P-16:0/20:4)+H<sup>+</sup></td></tr><tr><td>792.590</td><td>PC(O-16:0/22:6)+H<sup>+</sup>/PC(P-16:0/22:5)+H<sup>+</sup></td></tr><tr><td>794.606</td><td>PC(O-16:0/22:5)+H<sup>+</sup>/PC(P-16:0/22:4)+H<sup>+</sup></td></tr><tr><td>806.569</td><td>PC(38:6)+H<sup>+</sup></td></tr><tr><td rowspan="3">Hepatopancreas</td><td>332.783</td><td rowspan="3">Not assigned</td></tr><tr><td>348.758</td></tr><tr><td>375.065</td></tr></tbody></table>

<figure><img src="image_7.png" /><figcaption>Fig. 5: Identification of isobaric lipid species at m/z 782.56.</figcaption></figure>

### In-situ MALDI MS/MS

<figure><img src="image_8.png" /></figure>

$$ \begin{gathered} \text{CCS} = 290.7 \AA^2 \\ \text{[M+Na]}^+ \\ \text{PC (16:0-18:1)} \end{gathered} $$

<figure><img src="image_9.png" /></figure>

### ESI PASEF MS/MS

<figure><img src="image_10.png" /></figure>

$$ \begin{gathered} \text{CCS} = 285.7 \AA^2 \\ \text{[M+H]}^+ \\ \text{PC (36:4)} \end{gathered} $$

<figure><img src="image_11.png" /></figure>

<figure><img src="image_12.png" /></figure>

## Conclusions

* MALDI TIMS Imaging leads to richer molecular information content.

* Isobaric and isomeric lipids were mapped to specific locations in a tiny crustacean sample.

* The combination of in-situ MALDI MS/MS and ESI-PASEF is a powerful tool to retrieve lipid identifications with different distributions.

## MS-Imaging

<footnote n="1">¹Université Claude Bernad Lyon 1, Institut des Sciences Analytiques, CNRS UMR 5280, Villeurbanne, France</footnote>

<footnote n="2">²Bruker Daltonik GmbH, Bremen, Germany</footnote>

<footnote n="3">³INRAE, UR RiverLy, Laboratoire d'écotoxicologie, Villeurbanne, France</footnote>