Leftover Milk in the Bottle After a Feeding: Can It Be Kept for More Than 2 Hours?

Source: Zychlinsky Scharff A. et al., medRxiv preprint, February 2026 (not yet peer-reviewed)

Background and Rationale for the Study

Current recommendations from the WHO, the CDC, and the Academy of Breastfeeding Medicine (ABM) are particularly stringent: any milk remaining in the bottle after a feeding must be discarded within 2 hours for expressed breast milk, and immediately for infant formula. These recommendations are widely followed, yet their actual microbiological evidence base is surprisingly thin.

The study by Zychlinsky Scharff et al. (2026) is a preprint published on medRxiv in February 2026. It combines a cross-sectional survey of 1,056 parents with a prospective microbiological cohort study involving 44 bottle-fed infants (27 receiving infant formula, 17 receiving expressed breast milk), recruited at Hannover Medical School in Germany.

⚠️ Important caveat: this is a preprint, meaning it has not yet undergone peer review. The findings are preliminary and do not constitute a clinical recommendation. Lactation consultants should contextualise these data carefully when discussing them with families.

Methodology

Parental Survey

An online questionnaire was administered between June and October 2023 to 1,056 parents (99% mothers) of children under 36 months of age. Participants were asked to rate the financial, emotional, logistical, and time-related burden associated with each feeding method (direct breastfeeding, expressed breast milk via bottle, infant formula via bottle) on a Likert scale from 0 to 10.

Microbiological Study

For each infant, an oral cavity swab and two milk samples (before and after the feeding) were collected under sterile technique. Samples were then incubated at 4°C and at 20°C for 4, 8, and 24 hours. Bacterial quantification (CFU/ml) and species identification by MALDI-TOF mass spectrometry were performed at each time point. Statistical analyses used Mood's median test with Holm–Bonferroni correction for multiple comparisons.

Main Findings

Parental Burden and Milk-Discarding Practices

• 46% of parents discard leftover milk at least once a day.

• 84% would prefer to discard less if they were confident it was safe to do so.

• 67% discard milk immediately after the feeding, even at room temperature.

• Only 7% re-offer milk kept at room temperature for more than 2 hours.

• Expressed breast milk generates a higher emotional and logistical burden than direct breastfeeding; infant formula generates the greatest financial burden.

Bacterial Growth in Leftover Milk

In breast milk, the median bacterial load before the feeding was 4,200 CFU/ml (range: 300–350,000). After the feeding, it rose to 24,600 CFU/ml, reflecting transfer of the infant's oral flora. At 4 and 8 hours, no statistically significant increase was observed, whether the bottle was stored at 4°C (p = 0.82 at 4 h; p = 0.64 at 8 h) or at 20°C. Significant bacterial growth only appeared at 24 hours at 20°C (p < 0.001).

In infant formula, the baseline bacterial load before the feeding was expectedly lower (median: 0 CFU/ml). After the feeding, it rose to 11,700 CFU/ml. Again, no significant difference was observed at 4 h and 8 h, regardless of storage temperature. Significant growth also occurred only at 24 h at 20°C (p < 0.001).

The bacterial species identified in the milk after feeding were predominantly Staphylococcus epidermidis and Streptococcus mitis, commensals of breast milk and oral flora, with no pathogenic potential in this context for a healthy term infant.

Important Limitations to Be Aware Of

This study has several limitations:

• Modest sample size (n = 44) for the microbiological study, with high inter-individual variability.

• Population from a high-income country (Germany): findings are not directly applicable to resource-limited settings.

• Controlled laboratory conditions (fixed 20°C) do not reflect real-life domestic temperature fluctuations, particularly in summer or warmer environments.

• MALDI-TOF methodology does not detect viruses or certain fastidious pathogens.

• Participants, aware of the study's topic, likely adhered to hygiene practices more rigorously than in real-life conditions.

• Premature, immunocompromised, or hospitalised infants are explicitly excluded from the study's scope: for these populations, current recommendations to discard milk immediately remain fully valid.

Implications for Lactation Consultant Practice

This study provides the first in vivo microbiological data on milk remaining in the bottle after a feeding in infants, covering both breast milk and infant formula. Its results suggest that current recommendations (discard immediately or within 2 hours) are likely more conservative than necessary for healthy term infants in a home setting.

For lactation consultants supporting bottle-feeding families, these data can inform reflection on the unnecessary burden of wasting expressed breast milk, milk obtained at considerable physical and emotional cost. They invite a more nuanced approach to infant feeding safety, without anticipating official recommendations that have not yet been revised.

In practice: this data does not replace current institutional recommendations (WHO, CDC, ABM, French Society of Paediatrics). It provides a foundation for future research and, potentially, for a revision of clinical guidelines. In the meantime, healthcare professionals should inform families of the limitations of these data and of individual risk factors (prematurity, immunosuppression, local health context).