Small extracellular vesicles (sEVs) are gaining recognition as potential biomarkers for diseases, including cancer, due to their involvement in key pathophysiological processes. However, the glycosylation of EVs and the specific roles of their glycans remain poorly understood. While several methods exist for isolating sEVs from complex biological samples, achieving sufficient purity and quantity for mass spectrometry-based glycoproteomic analysis remains a significant challenge. In this study, we compared two commonly used isolation methods, ultracentrifugation (UC) and immunoaffinity capture (MagCapture kit), across different starting volumes of human serum (200 µL and 500 µL) to evaluate their performance for downstream glycoproteomic analysis. While prior studies have examined protein content across isolation methods, our work uniquely investigates how isolation technique and sample volume affect glycoproteomic yield and quality. We show that UC, particularly at higher sample volumes, enables deeper glycoproteomic coverage, whereas MagCapture is advantageous when serum availability is limited. Notably, we report for the first time site-specific glycan microheterogeneity on sEV glycoproteins derived from human serum, including multiple glycoforms at the same glycosylation site. These findings highlight the complexity and biological relevance of glycosylation in sEV proteins and offer practical guidance for optimizing isolation protocols based on specific omics applications.

Glyphosate-based herbicides (GBHs) have gained extensive popularity in recent decades. For many years, glyphosate has been regarded as harmless or minimally toxic to mammals due to the absence of its primary target, the shikimic acid pathway in humans. Nonetheless, mounting evidence suggests that glyphosate may cause adverse health effects in humans via other mechanisms. In this study, we described the metabolomic changes in the serum of experimental rats exposed to chronic GBH using the highly sensitive LC-MS/MS technique. We investigated the possible relationship between chronic exposure to GBH and neurological disorders. Our findings suggest that chronic exposure to GBH can alter spatial learning memory and the expression of some important metabolites that are linked to neurophysiological disorders in young rats, with the female rats showing higher susceptibility compared to the males. This indicates that female rats are more likely to show early symptoms of the disorder on exposure to chronic GBH compared to male rats. We observed that four important metabolites (paraxanthine, epinephrine, L-(+)-arginine, and D-arginine) showed significant changes and involvement in neurological changes as suggested by ingenuity pathway analysis. In conclusion, our results indicate that chronic exposure to GBH can increase the risk of developing neurological disorders.

Glyphosate, the active ingredient in many herbicides, has been widely used in agriculture since the 1970s. Despite initial beliefs in its safety for humans and animals due to the absence of the shikimate pathway, recent studies have raised concerns about its potential health effects. This study aimed to identify glycomic changes indicative of glyphosate-induced toxicity. Specifically, the study focused on profiling N-glycosylation, a protein post-translational modification increasingly recognized for its involvement in various disorders, including neurological conditions. A comprehensive analysis of rat serum N-glycomics following chronic exposure to glyphosate-based herbicides (GBH) was conducted using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The results revealed significant changes in the N-glycan profile, particularly in sialylated and sialofucosylated N-glycans. The analysis of N-glycans across gender subgroups provided insights into gender-specific responses to GBH exposure, with the male rats exhibiting a higher susceptibility to these N-glycan changes compared to females. The validation of significantly altered N-glycans using parallel reaction monitoring (PRM) confirmed their expression patterns. This study provides novel insights into the impact of chronic GBH exposure on serum N-glycan composition, with implications for assessing glyphosate toxicity and its potential neurological implications.

Clinical studies have linked glyphosate exposure to substantial morbidity, with acute kidney injury occurring in some cases. Although the toxic effects of glyphosate-based herbicides (GBHs) have been reported in several studies, their molecular impact on renal function remains poorly understood. Given the kidney’s critical role in excretion, it is particularly susceptible to damage from xenobiotic exposure. In this study, we aim to identify N-glycomics and proteomics change in the kidney following chronic GBH exposure, to better understand the mechanisms behind glyphosate-induced kidney damage. Kidney tissues from female and male rats were analyzed using liquid chromatography–tandem mass spectrometry. The results revealed notable changes in the N-glycan composition, particularly in the fucosylated and sialofucosylated N-glycan types. The proteomic analysis revealed the activation of immune signaling and inflammatory pathways, including neutrophil degranulation, integrin signaling, and MHC class I antigen presentation. Transcription regulators, such as IL-6, STAT3, and NFE2L2, were upregulated, indicating a coordinated inflammatory and oxidative stress response. Sex-specific differences were apparent, with female rats exhibiting more pronounced alterations in both the N-glycan and protein expression profiles, suggesting a higher susceptibility to GBH-induced nephrotoxicity. These findings provide new evidence that chronic GBH exposure may trigger immune activation, inflammation, and potentially carcinogenic processes in the kidney.

Mass spectrometry (MS) is a powerful analytical technique that is widely used to characterize a variety of analytes across diverse fields. In the area of biomass conversion, which is essential for producing sustainable materials and energy, the role of MS is pivotal. Biomass conversion processes, such as solvent liquefaction and pyrolysis, generate a wide range of industrially valuable materials including bio-based polymers, fuels, and activated carbons. However, the inherent complexity and heterogeneity of biomass and its transformation products pose significant analytical challenges. Advanced MS techniques, such as GC-MS, LC-MS, ICP-MS and MALDI-MS, are essential for a comprehensive analysis, providing detailed insights into the compositions, impurities, and potential inhibitors that influence process optimization and product quality. This review systematically explores recent advancements in MS-based methods for the analysis of biomass-derived products. We discuss fundamental innovations in biomass conversion processes and highlight the applications of various MS techniques in assessing the chemical complexity of these materials. The insights provided by MS techniques not only facilitate process optimization but also support the sustainable production of high-value materials from biomass, aligning with ongoing efforts to enhance environmental sustainability and resource efficiency.

This study applies a comprehensive MS-based glycomics workflow, supported by advanced bioinformatics, to resolve complex glycan structures for biomedical applications. Figure created in BioRender; glycan structures generated using GlycoWorkbench 2.0.