## 1 Introduction  
As the supernatant by-product left after coagulation of casein in the dairy industry, the whey protein (WP) fraction represents approximately 15-20% of all protein in bovine milk and has been discovered to be an excellent source of food-grade protein [1]. The peptides released during enzymatic hydrolysis also enable WP to present many in vivo biological functions, such as benefits on the nervous, endocrine, immune, cardiovascular, and digestive systems [2]. As an enriched source of essential amino acids and other nutrients [3], WP is now commonly applied in infant formula (IF), which has been ubiquitously used as a substitute for breastfeeding. Due to the high prevalence of allergies to cows' milk in infants, whey protein hydrolysate has been commercially developed as a major kind of hypoallerggenic dairy product [4]. Moreover, \"predigested\" IFs seem to induce a better feeding tolerance for preterm infants, which was indicated by higher gastrointestinal transport rates, more rapid gastric emptying, less gastro-esophageal reflux, and more rapid establishment of full enteral feeding [5]. More recently, an IF with partially hydrolyzed WP was proved to support adequate growth for healthy term infants [6]. WP hydrolysate was also found to serve as a probiotic growth and metabolism enhancer for Wistar rats, which were fed with a standard diet [7]. However, few studies have been conducted to evaluate the potential effect of hydrolyzed WP on the infant gut microbiome.  
The microbiota in human guts was affected by copious factors such as delivery pattern, diet, genetics, antibiotics, and prebiotics [8]. During the period of approximately three to five years after birth, the gut microbiota of individuals could have swift changes in microbial configuration till the relatively steady stage in adulthood. Those early-life occurrences even have a long-term impact on the brain and behavior [9]. The introduction of IF early in the postnatal period also shows interference in the colonization and proliferation of the neonatal intestinal microbiota, even given in small amounts during breastfeeding [10]. To properly modulate the microbiome, many attempts have focused on the application of prebiotics, which preferentially stimulates the growth and metabolic activity of probiotics. In general, prebiotics are chiefly thought to be various types of carbohydrates that are resistant to the digestion of the host [11]. However, some fragments from bovine and human breast milk were also reported to generate beneficial modulation of the gut microbiota [12]. WP-derived peptides could not only present bacteriostasis against pathogenic strains of _E. coli_, _Bacillus subtilis_, and _Staphylococcus aureus_, but also show growth-promoting effects on _Bifidobacterium longum_ ATCC 15,707 [4]. Additionally, WP-derived peptides have also been associated with the improvement of oxidative stress and the maintenance of the intestinal barrier, such as a commercial WP-derived product named Hilmar(tm) 8350 [13].  
In the current study, we sought to evaluate the potential modulation of the Hilmar(tm) whey protein hydrolysate (WPH) on the infant gut microbiome, using infant fecal inoculum and the cultivation of five probiotic strains. As a valuable reference for further nutritional supplement applications, information on the components of amino acids and peptides in WPH were obtained using HPLC-MS/MS.