Substance use can be a barrier to stable housing for homeless persons with mental disorders. We examined DSM-IV symptoms among homeless adults (N = 497), comparing those who reported daily substance use (DSU) with non-daily substance users. Multivariable linear regression modeling was used to test the independent association between DSU and symptoms using the Colorado Symptom Index total score. DSU was independently associated with higher symptoms (beta = 3.67, 95 % CI 1.55–5.77) adjusting for homelessness history, age, gender, ethnicity, education, marital status, and mental disorder sub-type (adjusted R 2 = 0.24). We observed a higher prevalence of DSU in our sample than has been previously reported in a Housing First intervention. DSU was also independently associated with more DSM-IV symptomatology. We have an opportunity to observe this cohort longitudinally and examine if there are changes in substance use based on treatment assignment and commensurate changes in housing stability, community integration, health status, and quality of life.