Optimizing Ergothioneine Biosynthesis and Antioxidant Activity in <i>Agaricus</i> spp. Through Amino Acid Supplementation and Yeast–Peptone Mixtures

With increasing demand for antioxidant-rich foods, research has focused on cost-effective methods to produce natural antioxidants. Mushrooms, especially <i>Agaricus</i> species, are rich in bioactive compounds like ergothioneine, a potent antioxidant. Ergothioneine has been shown to offer significant health benefits, such as protecting against oxidative stress, cardiovascular diseases, and premature aging. This study explores the effects of amino acid supplementation (methionine, cysteine, and histidine) and yeast–peptone mixtures on ergothioneine production, antioxidant activity, total phenolic content, and growth rate in various <i>Agaricus</i> species; this was conducted through two distinct experiments within a completely randomized design. In the first experiment, 13 treatment combinations were tested, involving varying concentrations of individual amino acids (methionine, cysteine, and histidine) at 0.5, 1, and 2 mM, as well as their combined concentrations (0.5 + 0.5 + 0.5, 1 + 1 + 1, and 2 + 2 + 2 mM), compared to a control (no amino acids). The second experiment tested yeast extract and peptone mixtures at seven concentrations: control (no supplementation), yeast (2 and 4 g/L), peptone (2 and 4 g/L), and combinations of yeast and peptone (2 + 2 and 4 + 4 g/L). Results revealed that supplementation with amino acids at 1 + 1 + 1 mM significantly enhanced ergothioneine content and antioxidant activity, though it resulted in decreased growth rates. In contrast, lower concentrations of amino acids (0.5 + 0.5 + 0.5 mM) increased ergothioneine production, with minimal impact on growth. Yeast and peptone supplementation at 2 + 2 g/L yielded the highest ergothioneine content, antioxidant activity, and growth rates across all <i>Agaricus</i> species tested. The most effective combination for maximizing ergothioneine production, antioxidant activity, and growth was found to be 0.5 mM of methionine, cysteine, and histidine, combined with 2 g/L of yeast extract and 2 g/L of peptone. <i>Agaricus bitorquis</i> (Quél.) Sacc. emerged as a promising candidate for ergothioneine production due to its genetic potential and metabolic efficiency. However, the strong responsiveness of <i>Agaricus bisporus</i> (white) to optimized culture conditions offers a viable alternative to <i>A. bitorquis</i>, which may require more complex and costly cultivation strategies.