The 29-year-old Jamaican clocked 9.81 seconds, beating rival Justin Gatlin of the United States by eight hundredths of a second at the Olympic Stadium on Sunday. Canada's Andre de Grasse took bronze in 9.91, reports Xinhua.
It was the seventh Olympic gold medal of Bolt's career as he confirmed his status as one of the greatest sprinters in history.
The result kept intact Bolt's hopes of being the first man to achieve an unprecedented triple-triple: three successive Olympic gold medals in the 100m, 200m and 4x100m relay.
Bolt enjoyed overwhelming support from the capacity crowd while his long-time rival Gatlin was roundly jeered.
Bolt beat Gatlin by one-hundredth of a second at the World Championships in Beijing last year.
The 34-year-old American won the 100m gold medal at the Athens 2004 Games but later served a four-year ban for doping.
Britain became the biggest winner at Rio on Sunday as their athletes pocketed five gold medals to overtake China on the medals table, only behind the United States.
Britain's gold haul was hightlighted by Max Whitlock's two gymnastics golds in one day. the 23-year-old talent won both the men's floor exercise and pommel horse titles.
Whitlock first stood out of a strong squad including all-around winner Kohei Uchimura and world champion Kenzo Shirai for the floor title and less than two hours later, Whitolck edged experienced teammate Louis Smith in the pommel horse final.
"It's just an incredible feeling," said Whitlock, who had already got an all-around bronze medal on Wednesday.
"All the gymnasts out there know how much work goes into it. You get one minute to show what you've been working on for the last however many years."
In tennis, Andy Murray became the first player to win two gold medals in the men's singles event as he scraped past Argentine Juan Martin Del Potro 7-5, 4-6, 6-2, 7-5 to retain his London 2012 Olympic Games title.
The newly crowned Rio Games champion hugged Del Potro after the four-hour match, both men with tears in the eyes.
Britain's Justin Rose won the men's golf gold medal on the sport's return to the Olympic Games, outlasting Sweden's Henrik Stenson on the final day to secure the two-shot victory.
"That felt better than anything I've ever won," said Rose. "It was the best tournament I've ever done."
Jason Kenny defeated Callum Skinner in an all-Britain final to defend his cycling individual sprint gold and win his second gold of the Rio Games.
Kenny and Skinner combined with Philip Hindes to win the team sprint Thursday night.
China bagged two gold medals with lifter Meng Suping claiming the women's over 75kg category and diver Shi Tingmao winning the women's 3m springboard title.
The diving gold was also Shi's second after she won the women's 3m synchronized springboard with Wu Minxia days ago.
"Today I can finally relax," said Shi, who finished with 406.5 points to beat her compatriot He Zi by 18.15 points.
"I will probably just go back to the Olympic Village and sleep all day long and then cheer for the other athletes from the Chinese team," Shi added.
China has won four of the five diving gold medals decided so far at the Rio Games.
Elsewhere, South Africa's Wayde van Niekerk shocked the Olympic Stadium by smashing the world record as he clinched the gold medal in the men's 400m.
The 24-year-old finished in 43.03 seconds, beating the previous record held by Michael Johnson of the US by 15 hundredths of a second.
Johnson's mark had been one of the longest standing records in athletics, having stood since August 1999.
In the morning session of the athletics competitions, Jemima Sumgong claimed Kenya's first gold medal of the Rio Olympics by winning the women's marathon.
After Sunday's finals, the US still tops medals table on 26 golds, while Britain and China are level on 15 gold medals with Britain placing second due to more silvers.
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