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Context: The Australian dairy industry is characterised by pasture-based feeding systems, where grazed pasture is commonly supplemented with cereal grain to increase milk production of dairy cows. Perennial ryegrass (PRG; Lolium perenne L.) is the most commonly used pasture species, with a wide range of cultivars available. However, the nutritive characteristics and the degradation of nutrients in individual PRG cultivars grown in different environments in Australia are unknown, and significant variation between cultivars may impact on supplementary nutrient requirements. Aims: The objective of this experiment was to quantify the extent of ruminal degradation of nutrients in three contrasting PRG cultivars (Bealey NEA2, Trojan NEA2 and Victorian SE) harvested during early spring and summer from Gippsland, northern Victoria and south-west Victoria in Victoria, Australia. Methods: Degradation parameters were determined by in situ incubation in the rumens of non-lactating, rumen-fistulated Holstein–Friesian cows for 72 h using a nylon bag technique. Key results: During both seasons, Bealey NEA2 had the greatest effective degradability of crude protein and neutral detergent fibre compared with Trojan NEA2 and Victorian SE. Cultivars harvested during early spring had greater effective degradability of crude protein and neutral detergent fibre, and a greater amount of rumen degradable protein compared with cultivars harvested during summer. Cultivars harvested from Gippsland had greater crude protein and neutral detergent fibre effective degradability, as well as a greater amount of rumen degradable protein, whereas cultivars harvested from northern Victoria had a greater amount of undegraded dietary protein. Conclusions: These results demonstrate that the ruminal degradation of PRG cultivars declines with increasing maturity, and that there are differences between cultivars and regions; however, differences between seasons are of a greater magnitude and are more important to consider when formulating optimal supplementary grain rations. Implications: This research has positive implications for farmers who will be able to understand the degradation of nutrients in PRG, and the amounts of rumen degradable protein and undegraded dietary protein available from pasture during each season. Farmers will be able to use this information to formulate supplementary grain rations that complement the nutrients from pasture and optimise milk production. Dairy production in south-eastern Australia is based on pasture-based feeding systems, where perennial ryegrass is the most common pasture species grown. This research provides novel information on the degradation characteristics of perennial ryegrass cultivars grown on farms in Australia, with degradability decreasing as the pasture matures. This will help farmers to formulate supplementary grain rations to match nutrient supply from pasture. [ABSTRACT FROM AUTHOR]