San Francisco also waived linebacker Keaton Kristick on Sunday, a week ahead of the season opener at home against the defending NFC West champion Seattle Seahawks.
San Diego had signed Tolzien as an undrafted free agent. During the preseason, he completed 25 of 40 passes for 302 yards with a touchdown and an interception.
Tolzien's quarterbacks coach/offensive coordinator at the University of Wisconsin was Paul Chryst, brother of 49ers QBs coach Geep Chryst. Tolzien completed 16 of 23 passes for 226 yards with a touchdown and an interception in San Diego's 20-17 loss to San Francisco on Thursday.
The Associated Press contributed to this report.