The Mercedes driver has a 53-point lead with seven rounds remaining but hopes of dominating his rivals at the Marina Bay Street Circuit were dampened by the resurgent Red Bulls and competitive Ferraris.
While Hamilton’s team mate and closest rival Nico Rosberg led a Mercedes one-two in the first session, both men were outpaced in the second as Russia’s Daniil Kvyat put Red Bull on top for the first time this season.
Kvyat lapped the 23-turn floodlit circuit in one minute, 46.142 to edge out Kimi Raikkonen’s Ferrari by just 0.039 seconds.
Red Bull’s Australian Daniel Ricciardo was third fastest after lapping in 1:46.256.
Hamilton was fourth, 0.337 seconds off the pace, while Rosberg languished in seventh place behind Ferrari’s Sebastian Vettel and Sergio Perez’s Force India.
Red Bull, who with the exception of a double podium appearance in Hungary have endured a miserable season with their underperforming Renault engine, looked to have found real pace around the streets of Singapore.
Ferrari also looked up for the fight in the muggy Southeast Asian conditions with very little between the top three teams in terms of lap times over the longer, race simulation runs with heavy fuel loads.
Rosberg complained over the radio that he had problems with his drinks bottle but the German generally enjoyed a trouble-free first 90-minute session to lap more than a second faster than a year ago.
Hamilton, however, endured an erratic evening. He went off the track three times as he pushed his Mercedes to the limit at the start of his quest to emulate boyhood hero Senna’s haul of race wins in the same number of starts (161).
Fortunately for the double champion, the mistakes occurred on parts of the track with run off areas with Vettel also venturing off in the first session after running wide at the quick Turn Five but stopping short of the barriers.
American rookie Alexander Rossi crashed as he prepared for his first weekend as a race driver for tailenders Manor Marussia after flying in at short notice to replace Spaniard Roberto Merhi for five of the remaining seven grands prix.
He completed 18 laps at the back of the field before losing control and slamming into the barriers after Turn 18, his broken Manor stranded and unable to continue as the session was halted with three minutes remaining.
He managed 20 minutes of action in his repaired Manor in the second session but only after team mate Will Stevens had crashed out early on.
(Editing by Amlan Chakraborty/Alan Baldwin)