Diurnal rhythms in melatonin and cortisol in very preterm human milk

Human milk (HM) contains bioactive compounds and hormones vital for infant neuro- and circadian-development. While healthy term-born infants receive time-of-day cues naturally through breastfeeding, preterm infants in the Neonatal Intensive Care Unit (NICU) are typically fed with pooled expressed milk via tube feeding, which lacks time-of-day specificity. Despite growing interest in chrononutrition and circadian entrainment in the NICU, little is known about diurnal hormonal rhythms in preterm HM and their potential role in circadian development.

We investigated diurnal rhythmicity of cortisol and melatonin in human milk from mothers of very preterm infants (born before 30 weeks gestational age) across different lactational stages. Milk series consisted of 13 to 17 (median 15) samples, collected over three consecutive days. Cortisol and melatonin concentrations were measured using enzyme-linked immunosorbent assays (ELISA) and rhythmicity was assessed using non-linear cosine curve fitting.

In total, 25 milk series were included. Melatonin exhibited a clear diurnal rhythm in all 25 HM series, while cortisol showed rhythmicity in 18 out of 25 HM series. The mean peak time for melatonin occurred at 03:58 (AM) ± 1.46 h; for cortisol, the mean peak time occurred at 08:29 (AM) ± 1.86 h. There was no significant association between lactational stage and peak time or amplitude for either hormone. In a preliminary subgroup analysis, milk from mothers of female infants showed a significantly earlier cortisol peak compared to mothers of male infants (p < 0.01).

Despite the immaturity of the lactational system after very preterm birth, consistent diurnal variations in cortisol and melatonin were present across all lactational stages. These findings demonstrate that maternal hormonal rhythmicity is preserved in human milk from the earliest stages of preterm lactation, providing a biological rationale for further research into whether time-of-day matched feeding strategies may provide biologically relevant temporal signals that warrant investigation regarding their potential role in circadian entrainment in very preterm infants.