Role of 70 °C heating on immunostimulatory effects of probiotic bifidobacteria in infant formula

Bifidobacteria, dominant members of the infant gut microbiota, are essential for immune system development and are commonly added to infant formula as probiotics. However, the preparation of powdered infant formulas involves dissolving the powder in hot water (>70 °C) to inhibit the growth of harmful bacteria, which can also affect probiotic viability. In this study, we assessed the cytokine production capacity of probiotic bifidobacteria exposed to 70 °C for 1 min, replicating the heating conditions for infant formula preparation. We subjected Bifidobacterium longum subsp. longum BB536, B. breve M-16V, and B. longum subsp. infantis M-63 to 70 °C for 1 min. Next, we exposed these strains to dendritic cells in vitro to observe the immunostimulatory activity. We found that heat treatment resulted in reduced cytokine production compared to that of unheated bacterial cells. The reduction in the immunostimulatory effect was less significant in bacterial cells heated at 70 °C for 1 min than in those heated at 90 °C for 15 min-the latter representing the heating condition used to completely kill bacteria during manufacturing. We also discovered that certain heat shock proteins, such as IbpA, which are highly expressed in B. longum BB536, retained their cytokine production capacity, even at 70 °C, but lost this capacity at 90 °C. These findings indicate that incorporating live bifidobacteria into infant formula as immunostimulatory ingredients is beneficial, even after the 70 °C preparation step that renders most cells non-viable.