A bioassay, based on the response of wheat roots, was used to quantify the concentrations of bioavailable S-metolachlor when applied at different timings. The application timings of S-metolachlor (0.48 kg/ha) were 20 days before crop sowing (DBS), at crop sowing (AS) applied either post-sowing pre-emergence (PSPE) or incorporated by sowing (IBS). The upper 0 to 5 cm soil layer was sampled from all treatments at 0, 8, 14, 23 and 33 days after crop sowing (DAS). The concentration of bioavailable S-metolachlor was similar between the application timings AS (IBS) and AS (PSPE) at both 0 DAS (94 to 96%) and 8 DAS (86 to 89%). After this period, herbicide bioavailability was significantly greater in the AS (IBS) than AS (PSPE). The bioavailabity of S-metolachlor was always greater for the herbicide applied AS (IBS) than applied at 20 DBS. The bioavailability of the herbicide applied at 20 DBS was 55% of the original applied herbicide at seeding. On the last sampling time (33 DAS) the bioavailability of S-metolachlor was 45, 27 and 28% of the original amount applied for application timings of AS (IBS), AS (PSPE) and 20 DBS, respectively. The implications of this information for weed management strategies are discussed.