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Effect of Digestion and Storage of Human Milk on Free Fatty Acid Concentration and Cytotoxicity

Penn, Alexander H.*; Altshuler, Angelina E.*; Small, James W.*; Taylor, Sharon F.; Dobkins, Karen R.; Schmid-Schönbein, Geert W.*

Journal of Pediatric Gastroenterology and Nutrition: September 2014 - Volume 59 - Issue 3 - p 365–373
doi: 10.1097/MPG.0000000000000441
Original Articles: Hepatology and Nutrition

Objectives: Fat is digested in the intestine into free fatty acids (FFAs), which are detergents and therefore toxic to cells at micromolar concentration. The mucosal barrier protects cells in the adult intestine, but this barrier may not be fully developed in premature infants. Lipase-digested infant formula, but not fresh human milk, has elevated FFAs and is cytotoxic to intestinal cells, and therefore could contribute to intestinal injury in necrotizing enterocolitis (NEC), but even infants exclusively fed breast milk may develop NEC. Our objective was to determine whether stored milk and milk from donor milk (DM) banks could also become cytotoxic, especially after digestion.

Methods: We exposed cultured rat intestinal epithelial cells or human neutrophils to DM and milk collected fresh and stored at 4°C or −20°C for up to 12 weeks and then treated for 2 hours (37°C) with 0.1 or 1 mg/mL pancreatic lipase and/or trypsin and chymotrypsin.

Results: DM and milk stored 3 days (at 4°C or −20°C) and then digested were cytotoxic. Storage at −20°C for 8 and 12 weeks resulted in an additional increase in cytotoxicity. Protease digestion decreased, but did not eliminate cell death.

Conclusions: Present storage practices may allow milk to become cytotoxic and contribute to intestinal damage in NEC.

*Departments of Bioengineering

Pediatrics, University of California, San Diego, La Jolla

Department of Psychology, University of California, San Diego, La Jolla.

Address correspondence and reprint requests to Alexander H. Penn, Department of Bioengineering, Institute of Engineering in Medicine, University of California, San Diego, La Jolla, CA 92093-0412 (e-mail:

Received 7 February, 2014

Accepted 12 May, 2014

The present study is supported by National Institutes of Health grants NS071580 and GM85072.

The authors report no conflicts of interest.

© 2014 by European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology,