The reserves went down to a 4-2 defeat at home to Walsall on Wednesday night.Chesterfield came back from 2-1 down to draw level following a goal from Kristian Ramsey-Dickson, but two late goals gave the Saddlers victory.
After going behind early on, the Spireites equalised when Armand Gnanduillet played the ball across the face of goal from the right-hand side and it was turned into the net by an opposition player.
Gnanduillet failed to re-appear in the second-half with Alex Hewitt taking his place.
The visitors regained the lead with a free header from a corner and Ramsey-Dickson equalised ten minutes after the break, lobbing the keeper after a ball played over the top was flicked on.
A second substitution after 70 minutes saw Cormac Noel replace Ashley Burbeary.
Chesterfield's hopes of completing a successful comeback were ended when a couple of good individual goals towards the end of the game gave Walsall victory.
Coach John Dungworth said: "Our defending could have been better but they were a good, footballing team. They were probably as good a team as we've played this season."
Chesterfield: Barrington, Wafula, Waddle, Donegan, Clay, Whittaker, Burbeary (Noel), Edge, Ramsey-Dickson, Gnanduillet (Hewitt), Compton. Unused substitutes: Mason, Slack, Mathers.