최재원 (국립창원대학교)

임재민 (국립창원대학교)

최재원 (국립창원대학교)

박정훈 (국립창원대학교)

우현주 (경남생명항노화 핵심지원센터)

임재민 (국립창원대학교)

Mass spectrometry (MS)-based glycomics is essential for the precise relative quantification of in vivo N-glycan profiles. While the Metabolic Isotope Labeling of Polysaccharides with Isotopic Glucose (MILPIG) strategy provides a robust framework for such quantification, its application in animal models, Caenorhabditis elegans (C. elegans), has been constrained by challenges in achieving consistent isotope incorporation, leading to potential quantification errors. In our previous study, we demonstrated the feasibility of MILPIG in C. elegans by utilizing a strictly controlled holidic diet and optimizing the isotope-labeled glucose (1-¹³C₁) concentration to 20 mg/mL.

Building upon this foundation, the current study refines this platform to ensure highly reproducible and accurate relative quantification. To enhance the robustness of the quantitative data, we implemented a time-course tracking strategy, analyzing glycan profiles at multiple developmental intervals to effectively control for metabolic variations. By monitoring these changes, we successfully stabilized the isotope cluster distribution and enhanced the monoisotopic peak intensity, effectively resolving the quantification errors caused by labeling inconsistencies. Our findings confirm that this refined MILPIG strategy establishes a superior, high-resolution platform for the precise relative quantification of glycan dynamics, providing a reliable framework for future glycomics studies in complex animal models.

Acknowledgements: This work was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education (RS-2024-00463300), and by the Korea Basic Science Institute (National Research Facilities and Equipment Center) grant funded by the Ministry of Education (2023R1A6C101B022).