Livestock production plays a leading role in agricultural land-use change. Producing biogas from livestock waste and subsequently using the biogas effluent as fertilizer for crops is a promising option to solve environmental problems resulting from expanding livestock production. However, it is difficult to promptly and accurately measure the nitrogen (N) concentration of effluent for farmers in developing countries, making precise N management difficult. The objectives of the current study were (1) to evaluate the feasibility of variable-timing, fixed-rate application of cattle biogas effluent using a leaf color chart (LCC) for rice (Oryza sativa L.) and (2) to determine the optimum LCC threshold for grain yield. We conducted two microcosm experiments in the Mekong Delta of Vietnam in 2018 using eight treatments of N-fertilizer application. In the Zero treatment, we applied no N. In the Estd treatment, we split-applied N as effluent (E) at fixed rate and timing as the standard method. In E2.75, E3.00, E3.25, E3.50, and E3.75, we applied effluent whenever the LCC value went below 2.75, 3.00, 3.25, 3.50, and 3.75, respectively. In U3.25, we applied N as urea (U) whenever the LCC value fell below 3.25. The total effluent-N application rate ranged from 90 to 210 kg N ha−1 season−1 . Rice growth was normal but there was a substantial yield gap between the two microcosm experiments due to the seasonal difference in solar radiation. Rice yield tended to increase with increasing LCC threshold. There was a positive linear relationship between LCC and chlorophyll content (SPAD) values (R2 = 0.73–0.79). Grain yield was well explained (R2 = 0.70–0.89) by the seasonal mean LCC or SPAD value. Plant total N uptake increased with increasing LCC threshold, but the three calculated indices of N use efficiency (NUE) – apparent N recovery, agronomic NUE, and internal NUE – were not always improved with a higher LCC threshold. Our results showed that the tested variable-timing, fixed-rate strategy for the application of cattle biogas effluent was feasible and the optimum LCC threshold for grain production was 3.75 under the current microcosm conditions