Preterm infants can inoculate virulent cytomegalovirus (CMV) through their mothers' raw breast milk. Complete virus inactivation is achieved only by heat treatment, but the effect on growth factors has never been assessed systematically. Insulin-likegrowth-factor-1-, IGF-2-, insulin-like-growth-factor-binding-protein-2-, and IGFBP-3-concentrations were measured, before and after heating, in 51 breast-milk-samples from 28 mothers, and epidermalgrowth-factor-concentrations in a subgroup of 35 samples from 22 mothers. Two heating methods were applied: Short-term (5 s) pasteurisation at 62, 65, and 72°C, and long-term Holder-Pasteurisation (30 min) at 63°C. IGF-1, IGF-2, IGFBP-2, and IGFBP-3 were measured by RIA, and EGF by ELISA. Heating for 30 min decreased significantly IGF-1 by 39.4%, IGF-2 by 9.9%, IGFBP-2 by 19.1%, and IGFBP-3 by 7.0%. In contrast, IGF-1, IGF-2, IGFBP-2, and IGFBP-3 were not altered significantly when using a short heating duration of 5 s, irrespective of the level of temperature, except for IGF-2 at 62°C for 5 s (p ϭ 0.041) and IGFBP-2 at 72°C for 5 s (p ϭ 0.025). Neither long-nor short-time heating methods changed the concentration of EGF. Only short heating methods (5 s, 62-72°C) can preserve, almost completely, the concentrations of IGFs in human milk, whereas Holder-Pasteurization does not. (Pediatr Res 65: 458-461, 2009) P reterm infants are at risk of acquiring Cytomegalovirus (CMV) infection in their first weeks of life through their mothers' raw breast milk, because CMV-IgG-positive mothers reactivate the virus during lactation and excrete it into their breast milk (1,2). For total inactivation of CMV in breast milk, heating procedures are necessary. Freeze-thawing may reduce the virus count but does not eliminate virulent CMV completely (3-5). There is growing evidence that, in addition to conventional Holder pasteurization (63°C, 30 min), short-term heat inactivation methods are effective (1,3; Müller, D ͓Establishing of a new procedure for the short-time heat pasteurization for inactivating of human Cytomegalievirus in mother's milk͔. Doctoral thesis 2006 at The Medical Faculty of the Eberhard-Karls-University of Tuebingen). However, the effect of the duration of heating and the temperature level achieved on bioactive substances like insulin-like growth factors (IGF), insulin-like growth factor binding proteins (IGFBP), and epidermal growth factor (EGF) in human milk has not yet been assessed systematically. We, therefore, evaluated the concentrations of growth hormones in human breast milk before and after processing with different heat inactivation parameters. METHODSMilk samples. IGF-1, IGF-2, IGFBP-2, and IGFBP-3 were analyzed in 51 freshly expressed breast milk specimens from 28 mothers; 21 of preterm and 7 of term infants. The expressed milk specimens, originating from lactation day 5-46, were stored at Ϫ20°C soon after collection until use. For processing, they were thawed, aliquoted, and labeled as either control or treated. The latter were heated according to fou...