Medela, Inc., P.O. Box 660, 1101 Corporate Drive, McHenry, IL 60051-0660Phone: (800) 435-8316 or (815) 363-1166 Fax: (815) 363-1246 Email: customer.service@medela.com

Medela is a registered trademark of Medela, Inc.1547637 A 0111 © 2011 Medela, Inc. Printed in the USA.

www.medela.com

2011 Innovating Practice Through Research & Evidence

Collection & Storage of Human Milk

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Research Overview:The Storage of Breastmilk

The benefi ts of breastfeeding and the negatives of formula feeding are becoming increasingly apparent through research based evidence. Importantly, this evidence is being communicated to pregnant women, mothers and healthcare professionals. As a result breastfeeding rates in the US have recorded modest increases as reported by the CDC in its latest ‘Breastfeeding Report Card – United States 2010’. Currently, 75% of mothers start breastfeeding, however, research indicates many stop shortly thereafter with exclusive breastfeeding rates at three months at only 33%.

There are many barriers to increased breastfeeding duration in the US, however, separation of the mother from her baby, for whatever reason, stands out as one of the more signifi cant obstacles to overcome. At times of separation, expressed breastmilk is the preferred feeding option for babies (WHO, 2003). Indeed, the act of expressing breastmilk provides distinct benefi ts for both mother and baby. With regard to the mother, the regular and adequate removal of milk from the breast (in the absence of the infant) supports the ‘supply and demand’ relationship between milk synthesis and removal (Daly et al, 1993) allowing for the continuation of lactation (Win et al, 2006) so that breastfeeding may last as long as the mother and baby mutually wish. For the baby, the known benefi ts of breastmilk over formula are well documented and new research continues to unearth more benefi ts at a rapid pace.

For the mother expressing and collecting her breastmilk there are extra challenges. Regardless of the setting (home or hospital) her milk will need to be collected and stored prior to being fed to her baby. This raises the question of optimal storage conditions for breastmilk. While there has been a signifi cant amount of research attempting to optimize storage conditions for cow’s milk, much less work has been done for breastmilk. With respect to milk storage two main questions continually arise, those of bacterial contamination and degradation of milk components. In this edition we have sourced three noteworthy, peer-reviewed research publications all addressing these issues and have summarized the key points for each one.

• ‘Effects of storage on the physiochemical and antibacterial properties of human milk’, by Michael O. Ogundele (2002).

• ‘Refrigerator storage of expressed human milk in the neonatal intensive care unit’, by Richard Schanler (2010).

• Academy of Breastfeeding Medicine, Clinical protocol #8: Human milk storage information for home use for full-term infants (2010).

In reviewing these three articles we cover the biochemistry and breastmilk storage scenarios in both hospital and home.

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References: CDC Breastfeeding Report Card – United States 2010.

World Health Organization. (2003). Global strategy for infant and young child feeding <http://whqlibdoc. who.int/publications/2003/9241562218.pdf> . Geneva, Switzerland: World Health Organization <http://en.wikipedia.org/wiki/World_Health_Organization> and UNICEF <http://en.wikipedia.org/wiki/UNICEF> . ISBN <http://en.wikipedia.org/wiki/International_Standard_Book_ Number> 9241562218 <http://en.wikipedia.org/wiki/Special:BookSources/9241562218> . http://whqlibdoc.who.int/publications/2003/9241562218.pdf. Retrieved 2011-01-28.

Daly SE, Kent JC, Owens RA, Hartmann PE. Frequency and degree of milk removal and the short-term control of human milk synthesis. Exp Physiol. 1996;81;861-875.

Win NN, Binns CW, Zhao Y, Scott JA, Oddy WH. Breastfeeding duration in mothers who express milk: a cohort study. Int Breastfeed J. 2006;1:28.

Effects of Storage on the Physiochemical and Antibacterial Properties of Human Milk. Ogundele. Michael O. British Journal of Biomedical Science, 2002, 59(4): 205-211.

Refrigerator Storage of Expressed Human Milk in the Neonatal Intensive Care Unit. Slutzah M, Codipilly C N, Potak D, Clark RM, and Schanler RJ. The Journal of Pediatrics, 2010, 156(1): 26-28.

ABM Clinical Protocol #8: Human Milk Storage Information for Home Use for Full-Term Infants.

Academy of Breastfeeding Medicine Protocol Committee. Breastfeeding Medicine, 2010, 5(3): 127-130.

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Research Overview:The Storage of Breastmilk

Mothers often refer to breastmilk as ‘liquid gold,’ therefore for mothers who express breastmilk, storage is of a key concern -- regardless of their environment (home or hospital). In the next few pages please fi nd summaries of two key research articles as well as the clinical protocols for milk storage from the Academy of Breastfeeding Medicine (ABM). These data provide an up-to-date view of the evidence supporting the development of optimal breastmilk storage conditions and provide you with key talking points.

Key points to note are:

• Bactericidal activity in refrigerated human milk declines in the fi rst 2-3 days of storage; however, the sequestration of bacteria seems to increase at this time. Both activities provide protection to the breastmilk fed infant.

• Human milk has unique properties specifi cally tailored to human infants.

• Human milk stored in a NICU refrigerator for 96 hours maintains its overall integrity and is safe for infant consumption.

• Fresh milk refrigerated for 4 days compares favorably to frozen or donor milk.

• The ABM guidelines are for full-term infants, not preterm or hospitalized infants.

• Cleanliness of milk expression, storage and handling are emphasized as it affects the numbers and types of bacteria in expressed milk. Milk that has higher bacteria counts will have a shorter shelf life.

• The ABM is very specifi c about this protocol serving as a guideline, not “an exclusive course of treatment” or standard of care.

• The CDC cites the ABM guidelines for human milk storage on their website.

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Notes

Research SummaryPrepared by: Jean Rhodes PhD, CNM, IBCLC

Effects of Storage on the Physiochemical andAntibacterial Properties of Human Milk. Ogundele. Michael O. British Journal of Biomedical Science, 2002, 59(4): 205-211.

Study Background

• This highly technical study examines the effects of different storage temperatures on bactericidal activities, bacteria sequestration activities and the pH of human milk.

• Human milk samples, obtained from 13 healthy lactating women, were grouped by postpartum period of lactation: colostrum, transitional milk and mature milk.

• Some milk samples were exposed to bacteria (e.coli) to test the degree to which human milk can destroy bacteria.

• The authors studied this bactericidal activity of human milk after exposure to refrigeration and freezing.

• Of particular interest, the authors also studied the milk fat globule membrane’s (MFGM) ability to sequester bacteria, preventing them from infecting their host. “Electron microscopic studies of the outer later of human MFGM shows the presence of numerous thin fi laments…oriented…into the aqueous phase of milk.” These glycoprotein stands trap bacteria, preventing them from adhering to the mucosal lining of the GI system.

Study Results

• Milk became more acidic over time due to an increase in free fatty acids rather than an increase in bacterial growth.

• Bactericidal properties were diminished but survived in samples frozen for up to 12 weeks.

• Milk bacterial sequestration - or the ability of milk fat globule membrane to adhere to suspended bacteria - was greatly enhanced during the fi rst three to seven days in refrigerated milk when compared to fresh milk. Bacterial sequestration was evident to a lesser degree in human milk frozen for four weeks.

Conclusion:

This study suggests enhanced bacteria sequestration activity compensates for the loss of bactericidal activity in human milk during the fi rst week of refrigeration. The decline in pH of stored human milk is not due to excessive contamination or deterioration of its protective abilities, but rather, the rise in free fatty acids from lipolysis.

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Notes

Research SummaryPrepared by Jean Rhodes, PhD, CNM, IBCLC

Refrigerator Storage of Expressed Human Milk in the Neonatal Intensive Care Unit. Slutzah M, Codipilly C N, Potak D, Clark RM, and Schanler RJ. The Journal of Pediatrics, 2010, 156(1): 26-28.

Study Background

• The goal of this study was to provide recommendations for refrigerator storage of fresh human milk in the NICU setting.

• This is a descriptive, comparative, longitudinal study of 36 samples of fresh, un-fortifi ed human milk stored in NICU refrigerators at 4° C for 96 hours.

• Milk samples were tested at 24, 48, 72 and 96 hours after collection. Variables analyzed were: pH, white blood cells (WBC) and osmolality.

• Additional samples were frozen and analyzed later for bacteria, fat, protein, and several factors known to or which may inhibit the growth of pathogens - secretory IgA, lactoferrin and free fatty acids.

• Results were compared to 5 samples of thawed pasteurized human donor milk (HDM) that were refrigerated for 96 hours and tested for bacteria, pH and free fatty acid concentrations.

Study Results

• Study results suggest human milk stored in a NICU refrigerator for 96 hours maintains its overall integrity and is safe for infant consumption.

• Small changes after 96 hours included:

- Gram-positive bacteria declined and free fatty acids increased (both refl ect anti-microbial aspects of human milk).

- Milk pH declined.

- WBCs declined but were higher than WBC levels in frozen or pasteurized milk.

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Study Results (continued)

- Total protein declined.

- When compared to HDM, refrigerated fresh milk had a higher pH (more normal pH) and lower free fatty acid concentrations but at levels deemed safe for infant feeding.

Study limitations

The authors did not study the changes or evaluate the safety of fresh refrigerated milk beyond 4 days.

• Clinical trial data is available regarding feeding mothers’ milk stored for 96 hours in the refrigerator to NICU infants.

Notes

Research SummaryPrepared by: Jean Rhodes, PhD, CNM, IBCLC

ABM Clinical Protocol #8: Human Milk Storage Information for Home Use for Full-Term Infants. Academy of Breastfeeding Medicine Protocol Committee. Breastfeeding Medicine, 2010, 5(3): 127-130.

Background

• The Academy of Breastfeeding Medicine (ABM) recognizes the importance of human milk storage and handling guidelines to the overall success of breastfeeding.

• These guidelines are a revision of the 2004 Clinical Protocol on human milk storage. They are an excellent synthesis of the issues related to human milk storage and handling, both in terms of what is known and what information is lacking. These guidelines are for term infants, not preterm or hospitalized.

Preparation for Human Milk Storage

• The protocol stresses the importance of hand and pump cleaning prior to milk expression and the use of clean storage containers.

• The authors recommend avoiding milk containers with bisphenol A (BPA) for the storage of Human Milk. • The authors discuss many of the variables involved in milk storage at different temperatures including cleanliness of collection, room temperature variation and the effects of temperature on specifi c milk components.

• Recommendations include avoid adding warm milk to previously cooled or frozen milk, and store milk in small increments to avoid wastage.

Using Stored Human Milk

• Infants may drink thawed milk at a variety of temperatures from cool to warm, depending on infant preference.

• Milk that has been brought to room temperature should be used within several hours.

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Using Stored Human Milk (continued)

• Milk that has been used for a feeding, i.e., infant’s mouth has had contact with the milk, should be used within 1-2 hours after the feeding.

• Milk from a mother with a breast infection is acceptable for use. • However, milk that is “stringy, foul or purulent” should be discarded.

• The authors do not have enough data to make recommendations about refreezing thawed milk.

ABM Milk Storage Guidelines

Location of storage Temperature Maximum recommended storage duration

Room temperature 16-29°C (60-85°F)

• 3-4 hours optimal

• 6-8 hours acceptable under very clean

conditions

Refrigerator ≤ 4°C (39°F)• 72 hours optimal

• 5-8 days under very clean conditions

Freezer < - 4°C (24°F)• 6 months optimal

• 12 months acceptable

Notes

Additional Human Milk Collection and Storage Resources

1. ABM. ABM clinical protocol #8: human milk storage information for home use for full-term infants (original protocol March 2004; revision #1 March 2010).

Breastfeed Med. Jun 2010;5(3):127-130.

2. Ajusi JD, Onyango FE, Mutanda LN, Wamola. Bacteriology of unheated expressed breast milk stored at room temperature. East Afr Med J. Jun 1989;66(6): 381-387.

3. Ankrah NA, Appiah-Opong R, Dzokoto C. Human breastmilk storage and the glutathione content. J Trop Pediatr. Apr 2000;46(2):111-113.

4. Bank MR, Kirksey A, West K, Giacoia G. Effect of storage time and temperature on folacin and vitamin C levels in term and preterm human milk. Am J Clin Nutr. Feb 1985;41(2):235-242.

5. Berkow SE, Freed LM, Hamosh M, et al. Lipases and lipids in human milk: effect of freeze-thawing and storage. Pediatr Res. Dec 1984;18(12):1257-1262.

6. Biagioli F. Returning to work while breastfeeding. Am Fam Physician. Dec 1 2003;68(11):2201-2208.

7. Boo NY, Nordiah AJ, Alfi zah H, Nor-Rohaini AH, Lim VK. Contamination of breast milk obtained by manual expression and breast pumps in mothers of very low birthweight infants. J Hosp Infect. Dec 2001;49(4):274-281.

8. Buss IH, McGill F, Darlow BA, Winterbourn CC. Vitamin C is reduced in human milk after storage. Acta Paediatr. Jul 2001;90(7):813-815.

9. CDC. Proper handling and storage of human milk. http://www.cdc.gov/breastfeeding/recommendations/ handling_breastmilk.htm. 2010.

10. Chantry CJ, Israel-Ballard K, Moldoveanu Z, et al. Effect of fl ash-heat treatment on immunoglobulins in breast milk. J Acquir Immune Defi c Syndr. Jul 1 2009;51(3):264-267.

11. Clark RM, Hundrieser KH, Ross S, Brown PB. Effect of temperature and length of storage on serum-stimulated and serum-independent lipolytic activities in human milk. J Pediatr Gastroenterol Nutr. Sep 1984;3(4):567-570.

12. Czank C, Prime DK, Hartmann B, Simmer K, Hartmann PE. Retention of the immunological proteins of pasteurized human milk in relation to pasteurizer design and practice. Pediatr Res. Oct 2009;66(4):374-379.

13. Czank C, Simmer K, Hartmann PE. Simultaneous pasteurization and homogenization of human milk by combining heat and ultrasound: effect on milk quality. J Dairy Res. May 2010;77(2):183-189.

14. Davanzo R, Travan L, Demarini S. Storage of human milk: accepting certain uncertainties. J Hum Lact. Aug 2010;26(3):233-234.

15. Eidelman AI, Szilagyi G. Patterns of bacterial colonization of human milk. Obstet Gynecol. May 1979;53(5):550-552.

16. Eteng MU, Ebong PE, Eyong EU, Ettarh RR. Storage beyond three hours at ambient temperature alters the biochemical and nutritional qualities of breast milk. Afr J Reprod Health. Aug 2001;5(2): 130-134.

17. Evans TJ, Ryley HC, Neale LM, Dodge JA, Lewarne VM. Effect of storage and heat on antimicrobial proteins in human milk. Arch Dis Child. Mar 1978;53(3):239-241. 18. Friend BA, Shahani KM, Long CA, Vaughn LA. The effect of processing and storage on key enzymes, B vitamins, and lipids of mature human milk. I. Evaluation of fresh samples and effects of freezing and frozen storage. Pediatr Res. Jan 1983;17(1):61-64.

19. Groh-Wargo S, Sapsford A. Enteral nutrition support of the preterm infant in the neonatal intensive care unit. Nutr Clin Pract. Jun-Jul 2009;24(3):363-376.

20. Hamosh M, Ellis LA, Pollock DR, Henderson TR, Hamosh P. Breastfeeding and the working mother: effect of time and temperature of short-term storage on proteolysis, lipolysis, and bacterial growth in milk. Pediatrics. Apr 1996;97(4):492-498. 21. Hamosh M, Henderson TR, Ellis LA, Mao JI, Hamosh P. Digestive enzymes in human milk: stability at suboptimal storage temperatures. J Pediatr Gastroenterol Nutr. Jan 1997;24(1):38-43.

22. Hanna N, Ahmed K, Anwar M, Petrova A, Hiatt M, Hegyi T. Effect of storage on breast milk antioxidant activity. Arch Dis Child Fetal Neonatal Ed. Nov 2004;89(6):F518-520.

23. Hernandez J, Lemons P, Lemons J, Todd J. Effect of storage processes on the bacterial growth-inhibiting activity of human breast milk. Pediatrics. Apr 1979;63(4):597-601.

24. Igumbor EO, Mukura RD, Makandiramba B, Chihota V. Storage of breast milk: effect of temperature and storage duration on microbial growth. Cent Afr J Med. Sep 2000;46(9):247-251.

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25. Jocson MA, Mason EO, Schanler RJ. The effects of nutrient fortifi cation and varying storage conditions on host defense properties of human milk. Pediatrics. Aug 1997;100(2 Pt 1):240-243.

26. Jones F, Tully MR. Best practice for expressing, storing and handling human milk in hospitals, homes and child care settings. Raleigh, NC: HumanMilk Banking Association of North America, Inc.;2006. 27. Larson E, Zuill R, Zier V, Berg B. Storage of human breast milk. Infect Control. Mar 1984;5(3):127-130. 28. Lavine M, Clark RM. Changing patterns of free fatty acids in breast milk during storage. J Pediatr Gastroenterol Nutr. Sep-Oct 1987;6(5):769-774.

29. Lawrence RA. Milk banking: the infl uence of storage procedures and subsequent processing on immunologic components of human milk. Adv Nutr Res. 2001; 10:389-404.

30. Liebhaber M, Lewiston NJ, Asquith MT, Sunshine P. Comparison of bacterial contamination with two methods of human milk collection. J Pediatr. Feb 1978;92(2): 236-237.

31. Marin ML, Arroyo R, Jimenez E, Gomez A, Fernandez L, Rodriguez JM. Cold storage of human milk: effect on its bacterial composition. J Pediatr Gastroenterol Nutr. Sep 2009;49(3):343-348.

32. Martinez-Costa C, Silvestre MD, Lopez MC, Plaza A, Miranda M, Guijarro R. Effects of refrigeration on the bactericidal activity of human milk: a preliminary study. J Pediatr Gastroenterol Nutr. Aug 2007;45(2): 275-277.

33. Molinari C, Casadio Y, Arthur P, Hartmann P. The effect of storage at 25° C on proteins in human milk.  International Dairy Journal.  Volume 21, Issue 4, April 2011, Pages 286-293.

34. Nwankwo MU, Offor E, Okolo AA, Omene JA. Bacterial growth in expressed breast-milk. Ann Trop Paediatr. Jun 1988;8(2):92-95.

35. Ogundele MO. Techniques for the storage of human breast milk: implications for anti-microbial functions and safety of stored milk. Eur J Pediatr. Nov 2000;159(11): 793-797.

36. Ogundele MO. Effects of storage on the physicochemical and antibacterial properties of human milk. Br J Biomed Sci. 2002;59(4):205-211.

37. Omarsdottir S, Casper C, Akerman A, Polberger S, Vanpee M. Breastmilk handling routines for preterm infants in Sweden: a national cross-sectional study. Breastfeed Med. Sep 2008;3(3):165-170.

38. Pardou A, Serruys E, Mascart-Lemone F, Dramaix M, Vis HL. Human milk banking: infl uence of storage processes and of bacterial contamination on some milk constituents. Biol Neonate. 1994;65(5):302-309.

39. Paxson CL, Jr., Cress CC. Survival of human milk leukocytes. J Pediatr. Jan 1979;94(1):61-64. 40. Permanyer M, Castellote C, Ramirez-Santana C, et al. Maintenance of breast milk Immunoglobulin A after high-pressure processing. J Dairy Sci. Mar 2010;93(3): 877-883.

41. Pittard WB, 3rd, Anderson DM, Cerutti ER, Boxerbaum B. Bacteriostatic qualities of human milk. J Pediatr. Aug 1985;107(2):240-243.

42. Pittard WB, 3rd, Bill K. Human milk banking. Effect of refrigeration on cellular components. Clin Pediatr (Phila). Jan 1981;20(1):31-33.

43 Rechtman DJ, Lee ML, Berg H. Effect of environmental conditions on unpasteurized donor human milk. Breastfeed Med. Spring 2006;1(1):24-26.

44. Santiago MS, Codipilly CN, Potak DC, Schanler RJ. Effect of human milk fortifi ers on bacterial growth in human milk. J Perinatol. Oct 2005;25(10):647-649.

45. Silprasert A, Dejsarai W, Keawvichit R, Amatayakul K. Effect of storage on the creamatocrit and total energy content of human milk. Hum Nutr Clin Nutr. Jan 1987;41(1):31-36.

46. Silvestre D, Ferrer E, Gaya J, et al. Available lysine content in human milk: stability during manipulation prior to ingestion. Biofactors. 2006;26(1):71-79.

47. Silvestre D, Lopez MC, March L, Plaza A, Martinez-Costa C. Bactericidal activity of human milk: stability during storage. Br J Biomed Sci. 2006;63(2):59-62.

48. Slutzah M, Codipilly CN, Potak D, Clark RM, Schanler RJ. Refrigerator storage of expressed human milk in the neonatal intensive care unit. J Pediatr. Jan 2010;156(1):26-28.

49. Sosa R, Barness L. Bacterial growth in refrigerated human milk. Am J Dis Child. Jan 1987;141(1):111-112.

50. Tacken KJ, Vogelsang A, van Lingen RA, Slootstra J, Dikkeschei BD, van Zoeren-Grobben D. Loss of triglycerides and carotenoids in human milk after processing. Arch Dis Child Fetal Neonatal Ed. Nov 2009;94(6):F447-450.

51. Williamson MT, Murti PK. Effects of storage, time, temperature, and composition of containers on biologic components of human milk. J Hum Lact. Mar 1996;12(1):31-35.

Compiled March 2011

Medela, Inc., P.O. Box 660, 1101 Corporate Drive, McHenry, IL 60051-0660Phone: (800) 435-8316 or (815) 363-1166 Fax: (815) 363-1246 Email: customer.service@medela.com

Medela is a registered trademark of Medela, Inc.1547637 A 0111 © 2011 Medela, Inc. Printed in the USA.

www.medela.com

2011 Innovating Practice Through Research & Evidence

Collection & Storage of Human Milk