Corn steep liquor (CSL), an aqueous stream from the corn industry, is used as a nutritional supplement in animal feed and biotechnological processes like the production of Penicillin. CSL was also identified as a source of biosurfactants, patent WO 2020/234501, and Gramicidin S. This raises the possibility of evaluating the residual supernatant and biomass from CSL, obtained after biosurfactant and Gramicidin S extraction, as potential nutritional supplements for culture media. Hence CSL was subjected to cell-bound biosurfactant and Gramicidin S extraction with phosphate buffered saline (PBS) or ethanol, respectively. Subsequently, fermentations with Lactobacillus pentosus were ran at 31ºC for 30 h, using culture media based on extracted CSL containing a mixture of liquid and solid residual fractions and compared with commercial CSL. L. pentosus was chosen due to its high nutritional requirements, and the colony-forming units (CFU)/ mL of L. pentosus on the different CSL-based media were considered as dependent variable. The results revealed that CSL subjected to extraction with either PBS or ethanol promoted similar growth of L. pentosus. Therefore, it can be concluded that extracted CSL can be used as a nutritional supplement in fermentative processes, increasing the added value of this secondary waste stream.