Uncovering the small-molecule landscape of fetal bovine serum: Integrated untargeted metabolomics and lipidomics identify compositional programs in three commercial grade products

Fetal bovine serum (FBS) is foundational to mammalian cell culture, yet the small-molecule composition of commercial grades—and its functional implications remain poorly defined. We performed integrated, untargeted LC–MS metabolomics and lipidomics on Low-, Medium-, and High-grade FBS (Thermo Fisher Scientific), and PCA showed clear grade separation with tight within-grade replication. Most features formed a conserved core (metabolites: 2414 ≈80%; lipids: 6558 ≈89%), indicating primarily quantitative rather than qualitative differences. Differential abundance revealed two coordinated programs: a storage/raft program in High-grade marked by higher TG/DG/CE and SM/Cer/HexCer together with aromatic/secondary-metabolite chemistry, and a signaling/membrane-remodeling program in Medium-grade enriched for PI/PLD pathways, lysophospholipids, Cer/HexCer, and vitamin/cofactor and bile-axis metabolites; Low-grade retained the core but lacked dense signaling hubs. DIABLO circos maps and cross-omics correlation networks confirmed strong lipid–metabolite covariance with inverse coupling between neutral-lipid storage and PI/sphingolipid signaling modules. Endocrine-linked molecules (steroidogenic lipids, bile-like species, and phytohormone-like metabolites) showed consistent grade biases with clear relevance to receptor signaling, redox balance, and stress responses. Together, these compositional data suggest that the three FBS products analyzed differ by coordinated small-molecule programs that rebalance membrane order, endocrine tone, and micronutrient capacity, providing a compositional basis for hypotheses about grade-associated differences in cellular behavior that warrant functional validation. We propose candidate sentinel lipids and metabolites as practical QC markers to augment lot qualification and guide the rational formulation of serum-reduced or defined media.