Objectives To examine different timeframes for calculating acute to chronic workload ratio (ACWR) and whether this variable is associated with intrinsic injury risk in elite Australian football players. Design Prospective cohort study. Methods Internal (session rating of perceived exertion: sRPE) and external (GPS distance and sprint distance) workload and injury data were collected from 70 players from one AFL club over 4 seasons. Various acute (1–2 weeks) and chronic (3–8 weeks) timeframes were used to calculate ACWRs: these and chronic load categories were then analysed to determine the injury risk in the subsequent month. Poisson regression with robust errors within a generalised estimating equation were utilised to determine incidence rate ratios (IRR). Results Altering acute and/or chronic timeframes did not improve the ability to detect high injury risk conditions above the commonly used 1:4 week ACWR. Twenty-seven ACWR/chronic load combinations were found to be “high risk conditions” (IRR > 1, p < 0.05) for injury within 7 days. Most (93%) of these conditions occurred when chronic load was low or very low and ACWR was either low (<0.6) or high (>1.5). Once a high injury risk condition was entered, the elevated risk persisted for up to 28 days. Conclusions Injury risk was greatest when chronic load was low and ACWR was either low or high. This heightened risk remained for up to 4 weeks. There was no improvement in the ability to identify high injury risk situations by altering acute or chronic time periods from 1:4 weeks.