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Seminal plasma (SP) mirrors the local pathophysiology of the male reproductive system and represents a non-invasive fluid for the study of infertility. Matrix-Assisted Laser Desorption/Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF-MS) provides a high-throughput platform to rapidly extrapolate the diagnostic profiles of information-rich patterns. In this study, dispersive solid phase extraction (d-SPE) combined with MALDI-TOF-MS was applied for the first time to the human SP, with the aim of revealing a diagnostic signature for male infertility. Commercially available octadecyl (C18)-, octyl (C8)-bonded silica sorbents and hexagonal mesoporous silica (HMS) were tested and the robustness of MALDI-TOF peptide profiling was evaluated. Best performances were obtained for C18-bonded silica with the highest detection of peaks and the lowest variation of spectral features. To assess the diagnostic potential of the method, C18-bonded silica d-SPE and MALDI-TOF-MS were used to generate enriched endogenous peptide profiles of SP from 15 fertile and 15 non-fertile donors. Principal component analysis (PCA) successfully separated fertile from non-fertile men into two different clusters. An array of seven semenogelin-derived peptides was found to distinguish the two groups, with high statistical significance. These findings, while providing a rapid and convenient route to selectively enrich native components of SP peptidome, strongly reinforce the prominent role of semenogelins in male infertility.

Seminal plasma (SP) mirrors the local pathophysiology of the male reproductive system and represents a non-invasive fluid for the study of infertility. Matrix-Assisted Laser Desorption/Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF-MS) provides a high-throughput platform to rapidly extrapolate the diagnostic profiles of information-rich patterns. In this study, dispersive solid phase extraction ( d -SPE) combined with MALDI-TOF-MS was applied for the first time to the human SP, with the aim of revealing a diagnostic signature for male infertility. Commercially available octadecyl (C 18 )-, octyl (C 8 )-bonded silica sorbents and hexagonal mesoporous silica (HMS) were tested and the robustness of MALDI-TOF peptide profiling was evaluated. Best performances were obtained for C 18 -bonded silica with the highest detection of peaks and the lowest variation of spectral features. To assess the diagnostic potential of the method, C 18 -bonded silica d -SPE and MALDI-TOF-MS were used to generate enriched endogenous peptide profiles of SP from 15 fertile and 15 non-fertile donors. Principal component analysis (PCA) successfully separated fertile from non-fertile men into two different clusters. An array of seven semenogelin-derived peptides was found to distinguish the two groups, with high statistical significance. These findings, while providing a rapid and convenient route to selectively enrich native components of SP peptidome, strongly reinforce the prominent role of semenogelins in male infertility.