Ceramide (Cer) and sphingomyelin (SM) are sphingolipids associated with conserved cellular processes; however, their involvement in the homeorhetic adaptation to lactation in cows is unknown. Our objectives were to characterize temporal responses in plasma sphingolipids during the periparturient period and determine whether these changes were related to adiposity and lipolysis. Multiparous Holstein cows were grouped by BCS at d −28 prepartum: lean (BCS 2.9 ± 0.13; n = 7) or overweight (OVER; BCS 4.0 ± 0.21; n = 7), fed a balanced diet. Blood samples were collected routinely from d −21 to 21. LC/MS was used to profile 37 Cer, monohexosylceramides (GlcCer), lactosylceramides (LacCer), and SM in plasma. Data were analyzed as repeated measures under a mixed model (fixed effects of BCS and day). Nonparametric correlations were analyzed. Plasma NEFA increased around calving and was higher in OVER from d −5 to 21 (P < 0.05). OVER lost more BCS and BW (P < 0.01). C16:0-Cer concentration decreased by 40% during transition and was higher in OVER prepartum (P < 0.05). C24:0-Cer increased 50% and was 33% higher in OVER at d 21 (P < 0.05). C22:0-Cer increased 49 and 66% at d 14 and 21, respectively, in OVER (P < 0.05). C24:0-GlcCer increased 52% during transition (P < 0.05) and tended to be higher in OVER at d 0 (P < 0.1). C16:0-GlcCer increased by 145% at d 4 (P < 0.01). C22:0-GlcCer increased over time and was higher in OVER at d 14 (P = 0.05). C24:1-GlcCer tended to increase by 100% in OVER during transition (P < 0.1). C16:0-LacCer increased postpartum and peaked at d 14 (P < 0.01). C18:1- and C24:1-LacCer levels increased during transition and were higher in OVER from d 4 to 21 (P < 0.01). Abundant C16:0-, C18:1- and C20:1-SM decreased over time and reached nadir around parturition (P < 0.01). C18:1- and C20:1-SM were higher in OVER pre- and postpartum (P < 0.05). C20:0-dihydro-SM decreased around calving (P < 0.01) and was higher in OVER at d −5 (P < 0.05). Cer, GlcCer and LacCer were positively correlated to NEFA and negatively to SM during transition (P < 0.001). Our results support a model of NEFA-induced ceramide synthesis during peripartum.