Tendulkar, 36, smashed 19 fours and four sixes for his 45th one-day century before India were bowled out for 347 in 49.4 overs while chasing Australia´s imposing 350-4 in 50 overs.
Tendulkar shared a 137-run partnership with Suresh Raina (59) to take India close in the day-night encounter.
But his dismissal in the 48th over proved the turning point of the game and handed Australia a 3-2 lead in the seven-match series.
Debutant fast bowler Clint McKay picked three wickets, including that of Tendulkar, while Shane Watson took 3-47 to cap a fine all-round performance, having scored 93 earlier.
Australia were also indebted to Shaun Marsh, who struck a career-best 112 and shared 145 runs for the opening wicket with Watson after the world champions won the toss and elected to bat at the Rajiv Gandhi stadium.
India began their chase in rollicking fashion with openers Virender Sehwag (38) and Tendulkar putting on 66 runs in 53 balls but lost their way in the middle with three quick dismissals.
Gautam Gambhir, Yuvraj Singh and skipper Dhoni all failed to reach double-figures as they tried to go for the big shots, India slipping to a shaky 162-4 by the 24th over.
Raina kept his cool, reaching his 12th one-day half-century with a six before a Watson double strike tilted the scale back in Australia´s favor.
Earlier, Marsh clobbered eight fours and two sixes in his run-a-ball knock for his maiden one-day hundred, reached with a clipped two to deep square leg off paceman Ashish Nehra (1-79).
Watson too thrived in batsmen-friendly conditions, hitting nine fours and three sixes in his 89-ball effort before being caught at deep mid-wicket as he tried to slog sweep off-spinner Harbhajan Singh (1-44).
Cameron White then hit a sparkling 57 off 33 balls to complete a run-feast by the injury-hit Aussies, missing nine of their regular one-day players to various niggles.
India´s bowlers were left with little answer to the Aussie onslaught, as reflected in the dismal figures of fast bowlers Nehra and Munaf Patel (0-73). Praveen Kumar picked two wickets but at the cost of 68 runs in nine overs.
The visitors were off to a flying start with Watson and Marsh laying the platform for the huge total with their aggressive stroke-making.
Marsh, dropped on 29 by Dhoni and on 52 by Sehwag, was finally dismissed by Nehra who had him caught by Gambhir at long-on.
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