This study was aimed at determining the effective ingredients of yeast culture (YC) for animal breeding. First, the contents of YCs obtained from various fermentation times were detected using gas-chromatography. A total of 85 compounds were identified. Next, 336 Arbor Acres (AA) broilers were randomly divided into seven experimental groups and fed a basal diet, diets supplemented with YCs obtained at various fermentation times, or SZ1 (a commercial YC product). A significant increase in body weight gain (BWG) and a significant decrease in feed conversion ratio (FCR) of AA broiler chicks were observed with YC supplementation. Additionally, most of blood and immunological indices were improved with YC supplementation. According to the production performance and the results of multivariate analysis, glycine, fructose, inositol, galactose, and sucrose were found as the potential effective compounds of YC and were involved in metabolic pathways including glycine, serine, and threonine metabolism. Supplementation with diets based on combinations of effective compounds improved weight gain, feed efficiency, serum immunoglobulin A, and immunoglobulin G, but decreased blood urea concentration. These findings suggest YCs as effective and harmless feed additives with improved nutritional properties for broiler chicks.