Andy Williams' rasping 25-yard shot saw Chesterfield relegated from League One after just one season.
The Spirites took the lead when Jordan Bowery turned the ball in off Mark Randall's cross past a passive defence.
Yeovil levelled when Dominic Blizzard passed into the net from deep inside the box after Andy Williams' assist.
Danny Whitaker's deflected shot put Chesterfield back in front before Keiran Agard slotted home and Williams struck from long range.
VIEWS FROM THE DRESSING ROOM
Chesterfield assistant manager Tommy Wright tells BBC Radio Sheffield:
"We're obviously disappointed, gutted. When we got the 2-1 lead I thought we were going to win the game.
"Two poor goals on our part have cost us dearly. I thought second half we were the better side, we created a lot of chances.
"But when you let people turn and shoot at goal, you're going to pay the consequences."