The circadian timing system (CTS) synchronizes internal physiology with regularly occurring cues such as light-dark cycle. To understand the role of circadian rhythms in regulation of lactogenesis we exposed cows to chronic phase shifts in light-dark cycle (PS) to disrupt the CTS during the dry period. Milk production efficiency increased in PS exposed cattle with no detrimental effects on blood NEFA or BHB levels. Our objective was to measure effect of PS exposure on melatonin (MEL) and serotonin (5HT) levels and circadian rhythms in cows to determine if alterations were associated with changes in production efficiency. Briefly, 5 wks before expected calving (BEC) multiparous cows were moved to tie-stalls and divided into 2 treatments: control (C, n = 16; 16 h light: 8 h dark) or phase-shifted (PS, n = 16; exposed 6 h light-dark phase shifts every 3 d). All cows were exposed to control lighting after calving. Blood samples were taken at 0600 on d 35 BEC, 21 BEC, 2 before calving (BC), and d 0, 2, 9, 15 and 22 postpartum (PP). A subset of cows (n = 6/group) were sampled every 4 h over 48 h at 3 time points: d 23 BEC, 9 BEC, and 5 PP. LC-MS/MS was used to measure MEL and 5HT. Body temperature was recorded every 30 min vaginally using calibrated iButton data loggers at d 23 BEC and d 9 BEC. PS (P < 0.05) shifted phase and attenuated circadian rhythms of core body temperature. The overall effect of PS was to increase MEL and decrease 5HT (P < 0.05) at 0600 sampling. MEL was > 1.5-fold higher on d 15 PP and 22 PP in PS than C; whereas 5HT was 1.3-fold lower (P < 0.05). Cosine fit analysis found MEL circadian rhythms were significantly affected by treatment with C cows exhibiting strong rhythms across 3 time points (r² ≥ 0.72; P < 0.01) and PS cows only fitting 24 h rhythm at d 23 BEC (r² ≥ 0.72; P < 0.01). Circadian rhythms of blood 5HT was only evident at d 9 BEC in C cows (r² = 0.68; P < 0.01), area under the curve of 5HT (170 ± 29 and 235 ± 29 mg/ml; P = 0.13) and MEL (1.62 ± 0.15 and 1.94 ± 0.14 ng/mL; P = 0.14) was increased from d 9 BEC to 5 PP, respectively across the treatments. These data led to hypothesis that the attenuation of circadian rhythms facilitates lactogenesis to increase milk production efficiency.