| Summary: | This study investigates the potential of utilizing three food wastes: cheese whey (CW), beet molasses (BM), and corn steep liquor (CSL) as alternative nutrient sources for the cultivation of the diatom <i>Phaeodactylum tricornutum</i>, a promising source of polyunsaturated eicosapentaenoic acid (EPA) and the carotenoid fucoxanthin. The CW media tested did not significantly impact the growth rate of <i>P. tricornutum</i>; however, CW hydrolysate significantly enhances cell growth. BM in cultivation medium enhances biomass production and fucoxanthin yield. The optimization of the new food waste medium was conducted through the application of a response surface methodology (RSM) using hydrolyzed CW, BM, and CSL as factors. The results showed a significant positive impact of these factors (<i>p</i> < 0.005), with an optimized biomass yield of 2.35 g L<sup>−1</sup> and a fucoxanthin yield of 3.64 mg L<sup>−1</sup> using a medium composed of 33 mL L<sup>−1</sup> of CW, 2.3 g L<sup>−1</sup> of BM, and 2.24 g L<sup>−1</sup> of CSL. The experimental results reported in this study showed that some food by-products from a biorefinery perspective could be utilized for the efficient production of fucoxanthin and other high-added-value products such as eicosapentaenoic acid (EPA).
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