BACKGROUND This study aimed to evaluate the effect of l‐tryptophan (l‐TRP) used in the cultivation of Lolium perenne on the yield, and chemical and microbiological quality of its herbage and silage. l‐Tryptophan was applied in doses of 5, 0.5, 0.05 kg ha−1. The experiment was conducted with a control group (C) and a comparative control group (C+) with higher nitrogen fertilization. RESULTS: The dose of 5 kg ha−1 had a significant effect on herbage yield, which increased by 15% compared to group C and approximated the value achieved in group C+. The treatment with l‐TRP caused a significant increase in water‐soluble carbohydrate (WSC) content only in Tr5 (165 g kg−1 DM), which was reflected in a more beneficial course of fermentation, lower pH (4.59), and a higher sum of fermentation acids, including lactic acid (94.7 g kg−1 DM). Ryegrass treatment with a high l‐TRP dose effectively reduced the loads of Clostridium spp. and fungi, and increased the count of Bacillus spp. The l‐TRP significantly reduced N‐NH3 content in Tr05 (98.6 g kg−1 TN) compared with C+ (123 g kg−1 TN) and butyric acid content in Tr05 (from 1.35 g kg−1 DM in the C to 0.38 g kg−1 DM). CONCLUSION: The most effective dose turned out to be a dose of 5 kg ha−1, which allowed a higher yield and a better fermentation course to be achieved. This work presents the feasibility of using l‐TRP to optimize nutrient consumption by Lolium perenne and ultimately to affect the quality of its silage as a feedstuff. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]