Wildland firefighters conducting prescribed burns are exposed to a complex mixture of pollutants, requiring an integrated measure of exposure. We used urinary mutagenicity to assess if systemic exposure to mutagens is higher in firefighters after working at prescribed burns versus after non-burn work days. Other biomarkers of exposure and oxidative stress markers were also measured. Using a repeated measures study design, we collected urine before, immediately after, and the morning after a work shift on prescribed burn and non-burn work days from 12 healthy subjects, and analyzed for malondialdehyde (MDA), 8-isoprostane, 1-hydroxypyrene (OH-pyrene), and mutagenicity in YG1041 +S9. Particulate matter (PM) and carbon monoxide (CO) were measured by personal monitoring. Light-absorbing carbon (LAC) of PM was measured as a surrogate for black carbon exposure. Linear mixed-effect models were used to assess cross-work shift changes in urinary biomarkers. No significant differences occurred in creatinine-adjusted urinary mutagenicity across the work shift between burn days and non-burn days. Firefighters lighting fires had a non-significant, 1.6-fold increase in urinary mutagenicity for burn versus non-burn day exposures. Positive associations were found between cross-work shift changes in creatinine-adjusted urinary mutagenicity and MDA ( = 0.0010), OH-pyrene ( = 0.0001), and mass absorption efficiency which is the LAC/PM ratio ( = 0.2245), respectively. No significant effect of day type or work task on cross-work shift changes in MDA or 8-isoprostane was observed. Urinary mutagenicity may serve as a suitable measure of occupational smoke exposures among wildland firefighters, especially among those lighting fires for prescribed burns.