Portable accumulation chambers (PAC) enable short term spot measurements of gaseous emissions including methane (CH4), carbon dioxide (CO2) and oxygen (O2) consumption from small ruminants. To date the differences in morning and evening gaseous measurements in the PAC have not been investigated. The objectives of this study were to investigate: 1) the optimal measurement time in the PAC, 2) the appropriate method of accounting for the animal’s size when calculating the animal’s gaseous output, and 3) the intra-day variability of gaseous measurements. A total of 12 ewe lambs (c. 10 to 11 months of age) were randomly selected each day from a cohort of 48 animals over nine consecutive days. Methane emissions from the 12 lambs were measured in 12 PAC during two measurement runs daily, AM (8 to 10 h) and PM (14 to 16 h). Animals were removed from Perennial ryegrass silage for at least 1 hour prior to measurements in the PAC and animals were assigned randomly to each of the 12 chambers. Methane (ppm) concentration, O2 and CO2 percentage were measured at 5 time points (T1 = 0.0 min, T2 = 12.5 min, T3 = 25.0 min, T4 = 37.5 min and T5 = 50.0 min from entry of the first animal into the first chamber) using an Eagle 2 monitor. The correlation between time points T5-T1 (i.e. 50 min minus 0 min after entry of the animal to the chamber) and T4-T1 was 0.95, 0.92 and 0.77 for CH4, O2 and CO2, respectively (P<0.01). The correlation between CH4 and CO2 output and O2 consumption, calculated with live-weight and with body volume was 0.99 (P<0.001). The correlation between the PAC measurement recorded on the same animal in the AM and PM measurement runs was 0.73. Factors associated with CH4 production included: day and time of measurement, the live-weight of the animal and the hourly relative humidity. Results from this study suggest that the optimal time for measuring an animal’s gaseous output in the PAC is 50 min, that live-weight should be used in the calculation of gaseous output from an animal and that the measurement of an animal’s gaseous emissions in either the AM or PM does not impact on the ranking of animals when gaseous emissions are measured using the feeding and measurement protocol outlined in the present study.