Although stratum corneum (SC) hydration has been primarily of concern to the cosmetic industry, it serves an important biosensor function. In murine models, not only deiminated products of filaggrin-derived amino acids ("NMF") but also endogenous glycerol from circulation into the epidermis via aquaporin 3 channel and from triglyceride turnover in sebaceous glands (SG) are important determinants. We assessed here whether endogenous glycerol could also be linked to SC hydration in humans. SG-enriched sites are more hydrated than SG-impoverished sites, and SC hydration correlates with both sebum production and SC glycerol content, but the correlation is more significant for SC glycerol content than for sebum content. Moreover, gender-related differences in sebum content are not associated with altered SC hydration. SC hydration is also linked to SC glycerol content in SG-impoverished sites, suggesting a role for non-SG-derived (? from circulation) glycerol in SC hydration. Finally, short-term water immersion produces a parallel decline in SC hydration and SC glycerol content, with glycerol levels returning to normal over several hours. These results suggest that endogenous glycerol of both circulatory and SG origin comprises an H2O-extractable pool that influences SC hydration in humans. These results also provide a rationale for the development of glycerol-containing therapeutic moisturizers.