## 4 Conclusions  
This study reported the application of LDPI-MS (Laser Desorption Ionization Mass Spectrometry) for in situ detection of CBZ pesticide residues in carrot tubers. The method offered several advantages, such as simplified sample preparation, minimal matrix interference, and rapid and convenient analysis, having a particular advantage in detecting both endogenous and exogenous compounds, particularly small molecules. The LOD of the present method reached 0.019 ppm. The LDPI-MS imaging results demonstrated viable visual differentiation of the levels of CBZ residues using distinct colors, effectively providing in situ molecular information on pesticide absorption within the tissues. The distribution of CBZ in carrot tubers was closely related to the tissue structure and morphology of carrot itself. Most CBZ molecules could not penetrate the epidermis of carrot, and mainly accumulated on the outer surface of the epidermis after washing. Since the leaves of the carrot were removed in the experiment, the transpiration of the carrot was inhibited, so the entry of CBZ is likely related to the permeability of the epidermis. With prolonged culturing time, we found that small amounts of CBZ molecules were able to enter and accumulate in the central xylem of the cross section, while relatively small amounts of CBZ were present in the cambium between the epidermis and the xylem. This difference in distribution may be related to differences in hydrophobicity among different tissues in carrot tubers. Furthermore, by analyzing images at different stages of slicing, dynamic information regarding the distribution of drug molecules within tissues was obtained, supporting theoretical investigations of exogenous drug metabolism. Clearly, LDPI-MS can be considered an effective and promising method, providing significant insight into plant metabolomics and pesticide residue analysis. Compared with traditional MSI, LDPI-MSI greatly improves the sensitivity of the analysis through postionization technology and can detect more molecules that are not ionized under conventional conditions. It is particularly suitable for applications that require high sensitivity and rapid spatial imaging of small-molecule compounds, such as determining the distribution of functional molecules in cell tissues and the distribution of drugs in organisms. Despite multiple advantages offered by this technique, laboratory equipment used in our experiments currently exhibits a lower imaging resolution, compared to existing commercial devices. Additionally, the analysis of experimental data has yet to be refined to the level of absolute quantification. However, the integration of novel machine-learning techniques has the potential to overcome these challenges. Addressing these limitations would further enhance the application spectrum of LDPI-MS and contribute to its advancement in the fields of plant metabolomics and pesticide residue analysis. The following supporting information can be downloaded at: [[https://www.mdpi.com/article/10.3390/app14135431/s1](https://www.mdpi.com/article/10.3390/app14135431/s1)]([https://www.mdpi.com/article/10.3390/app14135431/s1](https://www.mdpi.com/article/10.3390/app14135431/s1)), Figure S1: The schematic diagram of ion lens of time-of-flight mass spectrometer in this experiment; Figure S2: The time-of-flight mass spectrometer in this experiment; Figure S3: Spatial distribution of carbendazim dominate ion at m/z 160. The MSI data of this image and Figure 7b were obtained at the same time; Figure S4: Comparison of carrot sample LDPI-MS with or without carbendazim residue; Figure S5: Mass spectrum of Carbendazim by ESI-MS and carbendazim possible fragmentation pathways in LDPI-MS; Table S1: Effect of delay T between two laser beams and distance Z between the sample surface and ionization laser on mass spectrometry resolution; Table S2: Effect of delay T between two laser beams and distance Z between the sample surface and ionization laser on mass spectrometry peak relative intensity. Experiment, T.W.