Variation of Human Milk Glucocorticoids over 24 hour Period

  • Shikha Pundir
  • Clare R. Wall
  • Cameron J. Mitchell
  • Eric B. Thorstensen
  • Ching T. Lai
  • Donna T. Geddes
  • David Cameron-SmithEmail author


Human milk (HM) contains a complex array of hormones, including members of the glucocorticoid family. The predominant glucocorticoids, cortisol and cortisone may influence the growth and behaviour of the breastfed infant. However, little is understood of the factors regulating the levels of these hormones within HM. The aim of the study was to examine HM cortisol and cortisone concentration, measured in samples collected at each feed during a 24 hour period. Twenty three exclusively breastfeeding mothers collected milk, prior to and after each breastfeeding session over 24 hour period at 3.2(1.60) months. HM cortisol and cortisone levels were measured using high pressure liquid chromatography mass spectroscopy. Cortisone was the predominant glucocorticoid (3.40 ng/ml), and cortisol was detected in all samples (1.62 ng/ml). A positive correlation was found between cortisone and cortisol (r = 0.61, y = 1.93 ± 0.24, p < 0.0001). Cortisol and cortisone concentrations were significantly higher in feeds in the morning (2.97 ng/ml and 4.88 ng/ml), compared to afternoon (1.20 ng/ml and 3.54 ng/ml), evening (0.69 ng/ml and 2.13 ng/ml) and night (1.59 and 3.27 ng/ml). No difference was found between glucocorticoids level of the milk expressed for collection either before or immediately after the breastfeed, or between milk collected from the left or right breast. This study shows that HM glucocorticoid concentrations exhibit a 24 hour pattern, with highest peak levels in the early morning, reflecting the circadian pattern as previously reported in plasma. Thus, HM glucocorticoid concentrations are likely to reflect those in the maternal circulation.


Glucocorticoids Human milk Cortisol Cortisone High-performance liquid chromatography 



Authors thank the research team and mothers for the kind donation of breast milk used in the current study.

S.P. carried out laboratory analysis and data interpretation and drafted manuscript. E.B.T developed the MS method and oversaw the laboratory work. C.R.W and C.J.M assisted with statistical analysis and contributed to the manuscript development. D.T.G and C.T.L provided samples and contributed to the manuscript development. D.C.S designed research question and supervised all aspects of the study. All authors approve the submission of this manuscript for peer review.

Compliance with Ethical Standards


This study was funded by the Liggins Institute, University of Auckland, Philanthropic trust, the Riddet Institute, Massey University.

Conflict of Interest

CRW is employed through, Faculty of Medical and Health Science, University of Auckland. CJM, EBT and DCS are employed through Liggins Institute, University of Auckland. CTL and DG receive a salary from an unrestricted research grant from Medela AG, administered by University of Western Australia.


  1. 1.
    Ballard O, Morrow AL. Human milk composition: nutrients and bioactive factors. Pediatr Clin N Am. 2013;60(1):49–74.CrossRefGoogle Scholar
  2. 2.
    Hinde K, Skibiel AL, Foster AB, Del Rosso L, Mendoza SP, Capitanio JP. Cortisol in mother’s milk across lactation reflects maternal life history and predicts infant temperament. Behav Ecol. 2015;26(1):269–81.CrossRefPubMedGoogle Scholar
  3. 3.
    Neville MC, Allen JC, Archer PC, Casey CE, Seacat J, Keller RP, et al. Studies in human lactation: milk volume and nutrient composition during weaning and lactogenesis. Am J Clin Nutr. 1991;54(1):81–92.PubMedGoogle Scholar
  4. 4.
    German JB, Freeman SL, Lebrilla CB, Mills DA. Human milk oligosaccharides: evolution, structures and bioselectivity as substrates for intestinal bacteria. Nestle Nutr Workshop Ser Pediatr Program. 2008;62:205–22.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Chan S, Debono M. Replication of cortisol circadian rhythm: new advances in hydrocortisone replacement therapy. Ther Adv Endocrinol Metab. 2010;1(3):129–38.CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Grosvenor CE, Picciano MF, Baumrucker CR. Hormones and growth factors in milk. Endocr Rev. 1993;14(6):710–28.CrossRefPubMedGoogle Scholar
  7. 7.
    Grey KR, Davis EP, Sandman CA, Glynn LM. Human milk cortisol is associated with infant temperament. Psychoneuroendocrinology. 2013;38(7):1178–85.CrossRefPubMedGoogle Scholar
  8. 8.
    Tucker HA. Hormones, mammary growth, and lactation: a 41-year perspective. J Dairy Sci. 2000;83(4):874–84.CrossRefPubMedGoogle Scholar
  9. 9.
    Brisken C, O’Malley B. Hormone action in the mammary gland. Cold Spring Harb Perspect Biol. 2010;2(12):3178.CrossRefGoogle Scholar
  10. 10.
    Khani S, Tayek JA. Cortisol increases gluconeogenesis in humans: its role in the metabolic syndrome. Clin Sci. 2001;101:739–47.CrossRefPubMedGoogle Scholar
  11. 11.
    Catalani A, Casolini P, Cigliana G, Scaccianoce S, Consoli C, Cinque C, et al. Maternal corticosterone influences behavior, stress response and corticosteroid receptors in the female rat. Pharmacol Biochem Behav. 2002;73(1):105–14.CrossRefPubMedGoogle Scholar
  12. 12.
    Hahn-Holbrook J, Le TB, Chung A, Davis EP, Glynn LM. Cortisol in human milk predicts child BMI. Obesity. 2016;24(12):2471–4.CrossRefPubMedGoogle Scholar
  13. 13.
    Sullivan EC, Hinde K, Mendoza SP, Capitanio JP. Cortisol concentrations in the milk of rhesus monkey mothers are associated with confident temperament in sons, but not daughters. Dev Psychobiol. 2011;53(1):96–104.CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Hart S, Boylan LM, Border B, Carroll SR, McGunegle D, Lampe RM. Breast milk levels of cortisol and secretory immunoglobulin a (SIgA) differ with maternal mood and infant neuro-behavioral functioning. Infant Behav Dev. 2004;27(1):101–6.CrossRefGoogle Scholar
  15. 15.
    Cubero J, Valero V, Sánchez J, Rivero M, Parvez H, Rodríguez AB, et al. The circadian rhythm of tryptophan in breast milk affects the rhythms of 6-sulfatoxymelatonin and sleep in newborn. Neuro Endocrinol Lett. 2005;26(6):657–61.PubMedGoogle Scholar
  16. 16.
    Mitoulas LR, Kent JC, Cox DB, Owens RA, Sherriff JL, Hartmann PE. Variation in fat, lactose and protein in human milk over 24 h and throughout the first year of lactation. Br J Nutr. 2002;88(1):29–37.CrossRefPubMedGoogle Scholar
  17. 17.
    Khan S, Hepworth AR, Prime DK, Lai CT, Trengove NJ, Hartmann PE. Variation in fat, lactose, and protein composition in breast milk over 24 hours: associations with infant feeding patterns. J Hum Lact. 2013;29(1):81–9.CrossRefPubMedGoogle Scholar
  18. 18.
    Kent JC, Mitoulas LR, Cregan MD, Ramsay DT, Doherty DA, Hartmann PE. Volume and frequency of breastfeedings and fat content of breast milk throughout the day. Pediatrics. 2006;117(3):387–95.CrossRefGoogle Scholar
  19. 19.
    Katzer D, Pauli L, Mueller A, Reutter H, Reinsberg J, Fimmers R, et al. Melatonin concentrations and antioxidative capacity of human breast milk according to gestational age and the time of day. J Hum Lact. 2016;32(4):NP105–110.Google Scholar
  20. 20.
    Van der voorn B, de Waard M, van Goudoever JB, Rotteveel J, Heijboer AC, Finken MJ. Breast-milk cortisol and cortisone concentrations follow the diurnal rhythm of maternal hypothalamus-pituitary-adrenal axis activity. J Nutr. 2016;146(11):2174–9.CrossRefPubMedGoogle Scholar
  21. 21.
    Copinschi G, Challet E. Endocrine rhythms, the sleep-wake cycle, and biological clocks. In: Jameson JL, De Groot LJ, de Kretser DM, Giudice LC, Grossman AB, Melmed S, Potts JT, Weir GC WBS, editors. Endocrinology: adult and pediatric. 7th ed. Philadelphia: Elsevier; 2016. p. 147–173.Google Scholar
  22. 22.
    Dickmeis T. Glucocorticoids and the circadian clock. J Endocrinol. 2009;200(1):3–22.CrossRefPubMedGoogle Scholar
  23. 23.
    Peckett AJ, Wright DC, Riddell MC. The effects of glucocorticoids on adipose tissue lipid metabolism. Metabolism. 2011;60:1500–10.CrossRefPubMedGoogle Scholar
  24. 24.
    Hausman Kedem M, Mandel D, Domani KA, Mimouni FB, Shay V, Marom R, et al. The effect of advanced maternal age upon human milk fat content. Breastfeed Med. 2013;8(1):116–9.CrossRefPubMedGoogle Scholar
  25. 25.
    Barrett KE, Ganong WF. Ganong’s review of medical physiology. 15th ed. New York: McGraw-Hill Medical; 2012.Google Scholar
  26. 26.
    Verkerk GA, Phipps AM, Matthews LR. Milk cortisol concentrations as an indicator of stress in lactating dairy cows. Proc N Z Soc Anim Prod. 1996;56:77–9.Google Scholar
  27. 27.
    Engstrom JL, Meier PP, Jegier B, Motykowski JE, Zuleger JL. Comparison of milk output from the right and left breasts during simultaneous pumping in mothers of very low birthweight infants. Breastfeed Med. 2007;2(2):83–91.CrossRefPubMedGoogle Scholar
  28. 28.
    Cannon AM, Kakulas F, Hepworth AR, Lai CT, Hartmann PE, Geddes DT. The effects of leptin on breastfeeding behaviour. Int J Environ Res Public Health. 2015;12(10):12340–55.CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Heinrichs M, Neumann I, Ehlert U. Lactation and stress: protective effects of breast-feeding in humans. Stress. 2002;5(3):195–203.CrossRefPubMedGoogle Scholar
  30. 30.
    Nissen E, Uvnäs-Moberg K, Svensson K, Stock S, Widström AM, Winberg J. Different patterns of oxytocin, prolactin but not cortisol release during breastfeeding in women delivered by caesarean section or by the vaginal route. Early Hum Dev. 1996;45(1–2):103–18.CrossRefPubMedGoogle Scholar
  31. 31.
    Amico JA, Johnston JM, Vagnucci AH. Suckling-induced attenuation of plasma cortisol concentrations in postpartum lactating women. Endocr Res. 1994;20(1):79–87.CrossRefPubMedGoogle Scholar
  32. 32.
    Ramsay DT, Hartmann PE. Milk removal from the breast. Breastfeed Rev. 2005;13(1):5–7.PubMedGoogle Scholar
  33. 33.
    Khan S, Prime DK, Hepworth AR, Lai CT, Trengove NJ, Hartmann PE. Investigation of short-term variations in term breast milk composition during repeated breast expression sessions. J Hum Lact. 2013;29(2):196–204.CrossRefPubMedGoogle Scholar
  34. 34.
    Ettyang GA, van Marken Lichtenbelt WD, Esamai F, Saris WHM, Westerterp KR. Assessment of body composition and breast milk volume in lactating mothers in pastoral communities in Pokot, Kenya, using deuterium oxide. Ann Nutr Metab. 2005;49(2):110–7.CrossRefPubMedGoogle Scholar
  35. 35.
    Miller EM, Aiello MO, Fujita M, Hinde K, Milligan L, Quinn EA. Field and laboratory methods in human milk research. Am J Hum Biol. 2013;25(1):1–11.CrossRefPubMedGoogle Scholar
  36. 36.
    Gong S, Miao YL, Jiao GZ, Sun MJ, Li H, Lin J, et al. Dynamics and correlation of serum cortisol and corticosterone under different physiological or stressful conditions in mice. PLoS One. 2015;10(2):117503.Google Scholar
  37. 37.
    Morita H, Isomura Y, Mune T, Daido H, Takami R, Yamakita N, et al. Plasma cortisol and cortisone concentrations in normal subjects and patients with adrenocortical disorders. Metabolism. 2004;53(1):89–94.CrossRefPubMedGoogle Scholar
  38. 38.
    Chen DC, Nommsen-Rivers L, Dewey KG, Lönnerdal B. Stress during labor and delivery and early lactation performance. Am J Clin Nutr. 1998;68(4):335–44.PubMedGoogle Scholar
  39. 39.
    Chida D, Miyoshi K, Sato T, Yoda T, Kikusui T, Iwakura Y. The role of glucocorticoids in pregnancy, parturition, lactation, and nurturing in melanocortin receptor 2-deficient mice. Endocrinology. 2011;152(4):1652–60.CrossRefPubMedGoogle Scholar
  40. 40.
    Powe CE, Knott CD, Conklin-Brittain N. Infant sex predicts breast milk energy content. Am J Hum Biol. 2010;22(1):50–4.CrossRefPubMedGoogle Scholar
  41. 41.
    Hinde K. Lactational programming of infant behavioral phenotype. In: Clancy KBH, Hinde K, Rutherford JN, editors. Building babies. New York: Springer New York; 2013. p. 187–207.CrossRefGoogle Scholar
  42. 42.
    Van Cauter E, Leproult R, Kupfer DJ. Effects of gender and age on the levels and circadian rhythmicity of plasma cortisol. J Clin Endocrinol Metab. 1996;81(7):2468–73.PubMedGoogle Scholar
  43. 43.
    Kajantie E, Phillips DIW. The effects of sex and hormonal status on the physiological response to acute psychosocial stress. Psychoneuroendocrinology. 2006;31(2):151–78.CrossRefPubMedGoogle Scholar
  44. 44.
    Van der Voorn B, Martens F, Peppelman NS, Rotteveel J, Blankenstein MA, Finken MJJ, et al. Determination of cortisol and cortisone in human mother’s milk. Clin Chim Acta. 2015;444(0):154–5.CrossRefPubMedGoogle Scholar
  45. 45.
    Ju Bae Y, Gaudl A, Jaeger S, Stadelmann S, Hiemisch A, Kiess W, et al. Immunoassay or LC-MS/MS for the measurement of salivary cortisol in children? Clin Chem Lab Med. 2016;54(5):811–22.PubMedGoogle Scholar
  46. 46.
    Matsui F, Koh E, Yamamoto K, Sugimoto K, Sin H-S, Maeda Y, et al. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for simultaneous measurement of salivary testosterone and cortisol in healthy men for utilization in the diagnosis of late-onset hypogonadism in males. Endocr J. 2009;56(9):1083–93.CrossRefPubMedGoogle Scholar
  47. 47.
    Guo T, Chan M, Soldin SJ. Steroid profiles using liquid chromatography-tandem mass spectrometry with atmospheric pressure photoionization source. Arch Pathol Lab Med. 2004;128(4):469–75.PubMedGoogle Scholar
  48. 48.
    Carrozza C, Lapolla R, Gervasoni J, Rota CA, Locantore P, Pontecorvi A, et al. Assessment of salivary free cortisol levels by liquid chromatography with tandemass spectrometry (LC-MS/MS) in patients treated with mitotane. Hormones. 2012;11(3):344–9.CrossRefPubMedGoogle Scholar
  49. 49.
    Panda S, Hogenesch JB, Kay SA. Circadian rhythms from flies to human. Nature. 2002;417(6886):329–35.CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Shikha Pundir
    • 1
  • Clare R. Wall
    • 2
  • Cameron J. Mitchell
    • 1
  • Eric B. Thorstensen
    • 1
  • Ching T. Lai
    • 3
  • Donna T. Geddes
    • 3
  • David Cameron-Smith
    • 1
    Email author
  1. 1.Liggins InstituteThe University of AucklandAucklandNew Zealand
  2. 2.Faculty of Medical and Health ScienceThe University of AucklandAucklandNew Zealand
  3. 3.School of Molecular SciencesThe University of Western AustraliaPerthAustralia

Personalised recommendations