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This study evaluated the variation of the concentration of mimosine in Leucaena forage with season, the time of defoliation and dehydration. Fresh samples were collected randomly from Leucaena regrowth and sorted into four types of samples (primordial tips, young leaves, mature leaves and edible twigs). The samples were divided into two portions and handled as fresh or air-dried. Mimosine was extracted from the fresh and air-dried samples with dilute hydrochloric acid. Chemical analysis was done by indirect spectrophotometric methods. A General Linear Model procedure was used to determine the impact of the variables on the concentration of mimosine. The results showed that air-dried samples in both seasons contained significantly (P < 0.05) more mimosine than the corresponding fresh samples by an average of 34%. Fresh autumn samples collected at 12:00 contained more mimosine (15.43%) than samples collected at 15:00 (9.09%) and 9:00 (4.48%) in comparison with corresponding summer samples. Similarly, air-dried autumn samples collected at 12:00 contained more mimosine (47.15%) than samples collected at 9:00 (43.57%) and 15:00 (40.12%) in comparison with the corresponding summer samples. Dehydration of samples elevated the concentration of mimosine by 39.09%, 31.72% and 31.03% for samples collected at 9:00, 12:00 and 15:00, respectively. It was concluded that dehydration of samples had the most significant impact on the concentration of mimosine followed by the time of defoliation and the season.