Acid-detergent insoluble nitrogen (ADIN) is thought to represent nitrogen that is not degraded during ruminal and post-ruminal digestion of forages by cattle. Forage ADIN can be determined following direct acid detergent extraction or following sequential extraction with neutral and acid detergents. In this study, we evaluated how direct verses sequential detergent extraction influences the ADIN content of legume silages differing in polyphenol composition and degree of conditioning at harvest. In 2002 and 2003, cuttings of polyphenol-free alfalfa, low to high tannin birdsfoot trefoil, and red clover with o-diphenols and polyphenol oxidase were conventionally conditioned or macerated and then ensiled in minisilos for 90 d. ADIN, as a proportion of total N, was determined by combustion after direct and sequential detergent extraction by the ANKOM filter bag system, without the addition of sulphite or amylase. In conventionally conditioned silages, ADIN concentrations by direct and sequential methods averaged 40 and 34 g/kg for alfalfa, 43 and 34 g/kg for low tannin trefoil, 46 and 37 g/kg for moderate tannin trefoil, 48 and 40 g/kg for high tannin trefoil, and 41 and 31 g/kg for red clover. In macerated silages, ADIN concentrations by direct and sequential methods averaged 42 and 36 g/kg for alfalfa, 43 and 39 g/kg for low tannin trefoil, 35 and 43 g/kg for moderate tannin trefoil, 44 and 52 g/kg for high tannin trefoil, and 50 and 37 g/kg for red clover. Thus, analysis method, forage type, and degree of conditioning at harvest differentially influenced ADIN concentrations. The potential impact of these shifts in ADIN concentrations on milk production and nitrogen utilization by dairy cattle will be evaluated using nutrition and whole-farm models.