| Summary: | Abstract Supplementation of the spent oyster substrate enhances its nutritional properties to produce a new mushroom cropping cycle. The study investigated the potential of a nano-fertilizer (Lithovit®-Amino25) with an admixture of 25% l-amino acids on Pleurotus ostreatus production, proteins, and amino acid contents. The product applied at spawning (t1), after the first harvest (t2), and at both timings (t3), in two doses: 3 g/kg (C1) or 5 g/kg (C2). Compared with control (C0t0), the first harvest was earlier by 2.3–3.3 days in C1t1 and C2t1. The biological yield of the second harvest was improved by 28.0% in C2t2. Superior results were in C1t3 where the number of crops increased to four, biological efficiency was optimized (117.3%) at the third harvest, and biological and economic yields increased by 36.7% and 36.4%, respectively. Lignin was the most degraded in C1t3, while residual cellulose, hemicellulose, neutral detergent fiber, and acid detergent fiber were higher in all treated substrates than in control. In C2t1, mushrooms were the richest in proteins, while in C1t1, they were the richest in the essential amino acids threonine, valine, isoleucine, leucine, and histidine. Lithovit®-Amino25 has a high potential for use in P. ostreatus production.
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