The study describes practices relating to syringe acquisition and disposal by Syringe Exchange Programme (SEP) participants. A cross-sectional multi-city study enrolled 857 injection drug users (IDUs) from six SEPs in different Brazilian regions, and assessed self-reported acquisition and disposal behaviours. Seven hundred and nine males (82.9%) and 146 females (17.1%) were recruited through outreach and interviewed, most from the streets or their neighbourhoods (54.1%). The average age was 28.5 years; 76.4% reported injecting cocaine in the past 6 months. Sources for acquiring new syringes differed significantly between time of injection drug use debut and the 6 months prior to interview. Fifty-three percent of IDUs reported acquiring their syringes in pharmacies when they initiated injection drug use, whereas most reported acquiring new syringes in the 6 months before interview from several simultaneous sources: 69% through SEPs; 58% through pharmacies; 36% from friends and/or sexual partners; and 17% from other health services. Across SEPs, acquisition and disposal varied widely. Most interviewees discarded their syringes on the streets, in open fields, or in the garbage or sewage. Restrictions on syringe availability and unsafe practices may be functioning as barriers to the public health recommendation of one-time use of sterile syringes for IDUs and discouraging community support to SEPs. Further increase in access to legal, inexpensive and timely sterile syringes, as well as counselling about the merits of one-time use and safer disposal must be reinforced as part of efforts to minimise high-risk behaviours and curb the spread of blood-borne infections.