Werder Bremen, playing their first official game under new coach Robin Dutt, were beaten 3-1 in extra time by third-division side Saarbrucken.
After a poor string of results in pre-season, following on from their worst Bundesliga season in 14 years, the alarm bells will already be starting to ring in northern Germany with Sebastian Prodl's goal not enough to prevent them from losing in the first round for the third successive year, and for the third time to third-division opposition.
Bundesliga newcomers Eintracht Braunschweig went down 2-1 to 2. Bundesliga side Arminia Bielefeld while Borussia Monchengladbach were beaten in a penalty shoot-out by Darmstadt.
After a goalless 120 minutes, Luuk de Jong and Branimir Hrgota missed from the spot to give Lucien Favre's side a losing start to the season.
Nurnberg became the fourth Bundesliga side to fall at the first hurdle today when they were beaten on penalties by second-division Sandhausen.
Hanno Balitsch and Marvin Plattenhardt both missed for Michael Wiesinger's men, who were held to a 1-1 draw after 120 minutes.
Hannover had no such concerns against Victoria Hamburg with Artur Sobiech and Szabolcs Huszti scoring in a 2-0 win while Greuther Furth saw off Pfeddersheim by the same scoreline.
Hertha Berlin left it late to beat Neumunster and successfully come through their first test of the season to reach the second round of the DFB-Pokal.
With the game tied at 2-2 after 90 minutes, extra time was needed to find a winner. Just as it looked like the game would be determined by a penalty shoot-out, Hertha got the chance to snatch it from the spot.
Sami Allagui stepped up and, with the final kick of the game, sent Jos Luhukay's men through.
They were joined in the last 32 by Hamburg and Stuttgart, who made easier work of their first-round ties.
Artjoms Rudnevs grabbed a brace as Hamburg beat SV Schott Jena 4-0 while Vedad Ibisevic scored both goals in Stuttgart's 2-0 win over Berliner FC Dynamo.
Eintracht Frankfurt beat FV Illertissen by the same scoreline in Augsburg, and they were joined in the second-round draw by city rivals FSV Frankfurt, who needed extra time to beat Optik Rathenow 3-1.
Preussen Munster caused a minor stir by knocking out 2. Bundesliga side St. Pauli while fourth-division SC Wiedenbruck upset Fortuna Dusseldorf with a 1-0 home win.
Information from the Press Association was used in this report.