Plzen raised eyebrows all over Europe as they cantered to a 3-0 win in Naples last week and completed a professional performance over two legs with an impressive victory on home soil on Thursday night to set up a meeting with Fenerbahce in the next round.
Second-half goals by Jan Kovarik and Stanislav Tecl proved the difference as sorry Napoli fell well short in their bid to become the first team in this competition to overturn a three-goal first-leg deficit.
Napoli coach Walter Mazzarri clearly thought the game was up before kick-off as he kept the competition's top scorer Edinson Cavani on the bench until the start of the second half, and with hindsight that is where he should perhaps have stayed as Plzen put the game to bed within minutes of his introduction.
Napoli's exit at least gives them the chance to focus their attention on domestic matters with the Naples club sitting second in Serie A, four points behind Juventus.
Plzen claimed their shock first-leg win at the Stadio San Paulo thanks to goals from Vladimir Darida, Frantisek Rajtoral and Tecl, and were in determined mood not to let that advantage slip on Thursday night.
The Czech league leaders exited the tournament at this stage last season after a 4-2 extra-time defeat by Schalke but a repeat was never on the cards as Napoli were reduced to optimistic efforts from long range in a forgettable first half.
Mazzarri was not in the mood to go down without a fight, however, and duly introduced Gokhan Inler and Cavani at the interval in a final throw of the dice.
Uruguay international Cavani has caught the eye with seven Europa League goals this season but his introduction failed to have the desired effect as Napoli fell further behind in the 50th minute.
Rajtoral teed up the chance with a cross from the right wing that Kovarik turned home low into the left corner from near the penalty spot.
And Napoli's misery was compounded with 16 minutes remaining when Daniel Kolar laid the ball off for Tecl to fire home a thumping effort from long range.