Last season's Johnstone's Paint Trophy finalists Chesterfield bowed out in the first round this time in a 2-0 defeat at Scunthorpe.
The home side took control with first-half goals from Sean McAllister and former Spireite Gary McSheffrey.
McAllister netted in the 15th minute with a left-foot shot on the turn from inside the box after McSheffrey had threaded the ball through.
Ten minutes later McSheffrey made it 2-0 after a mis-kick by substitute goalkeeper Aaron Chapman left him with an open goal.
Chapman then made amends with a full stretch save from a Paddy Madden shot to prevent his side falling further behind.
Twice more before the interval Madden had a chance to make it 3-0 but the striker could not find his finishing touch.
For an hour Chesterfield offered little threat in front of goal until midfielder Ollie Banks tried his luck from long range with a shot that brought the best out of home keeper Bobby Olejnik.
The visitors finished the game with 10 men when centre-back Jack Broadhead was sent off in the 84th minute for a second yellow card offence as Scunthorpe eased comfortably into round two.