The couple announced the news on Twitter on January 9, but confirmed in an online statement that they will remain "cherished friends".
Mr Bezos, who was last year named as the richest man in modern history, has a fortune of around £107.3 billion and the divorce could see MacKenzie become the richest woman in the world.
Soon after the announcement was made, US tabloid the National Enquirer published a story about the Amazon boss' alleged affair with former TV presenter Lauren Sanchez.
On Friday, Mr Bezos made an extraordinary public statement in which he claimed he was the target of "extortion and blackmail" by Enquirer publisher American Media Inc over private photos.
Here's everything you need to know about the man behind one of the world's biggest companies.
What is Jeff Bezos' net worth?
But this week his fortune soared to more than $160 billion after his company surpassed Microsoft to become Wall Street's most valuable company.
According to Business Insider, Bezos is so wealthy that him spending $88,000 dollars is the equivalent to an average person spending $1.
How did Jeff Bezos make his money?
Bezos founded Amazon in 1994 when it began as an online bookshop before selling a variety of different products, and it ended-up taking on established retail giants such as Walmart.
The company has since continued to diversify and grow, through acquisitions such as Whole Foods and services such as Amazon Web Services.
Under Bezos, Amazon also launched the Kindle e-reader and revolutionized the way books are distributed and read. The company has also been a pioneer in cloud computing.
At the end of 2015, the e-commerce company announced 304 million active member accounts.
In 2018, Amazon's stock soared by 60% and had a market value of more than $890 billion.
Bezos is the largest shareholder with a 16% stake in the company. He also personally owns the Washington Post and Blue Origin, a space exploration company.
Who is his wife, Mackenzie Bezos?
Novelist Mackenzie Bezos married Jeff in 1993 before they both quit their jobs and relocated to Seattle to found Amazon.
Mackenzie, 48, grew up in San Francisco and attended Princeton University, before she met her husband when interviewing for a job at a New York hedge fund.
She was one of the first Amazon employees, doing the accounting for the company in the first year, with Bezos saying last year that her support was instrumental in founding the company.
He has said of his wife: "I think my wife is resourceful, smart, brainy and hot, but I had the good fortune of having seen her résumé before I met her, so I knew exactly what her SATs were."
The couple have four children and are the 25th largest landowners in the US.
Last year, they launched the Bezos Day One Fund, offering a two billion dollar fund to help homeless families and build pre-schools in low-income neighbourhoods.
Why are they divorcing?
According to the Twitter statement, the couple decided to divorce after a long period of "living exploration" and "trial separation".
Bezos wrote: "We want to make people aware of a development in our lives.
"As our family and close friends know, after a long period of loving exploration and trial separation, we have decided to divorce and continue our shared lives as friends.
"We feel incredibly lucky to have found each other and deeply grateful for every one of the years we have been married to each other. If we had known we would separate after 25 years, we would do it all again.
"We've had such a great life together as a married couple, and we also see wonderful futures ahead, and projects, and as individuals pursuing ventures and adventures. Though the labels might be different, we remain a family, and we remain cherished friends."
How will Jeff Bezos' divorce affect his net worth?
Mr Bezos is Amazon's main shareholder, owning around 16 percent of the the company's shares.
His divorce could be one of the costliest in history as MacKenzie could get in half of his fortune - making her the richest woman in the world.
According to the Bloomberg Billionaires Index, Jeff Bezos is worth $137 billion (roughly £107.3 billion) as of January 11, 2019.
However, he could see his net worth plummet following the proceedings of his divorce with MacKenzie.
In the early days, MacKenzie was reportedly involved in Amazon’s business dealings. According to a 1999 Jeff Bezos profile with she actually secured the company’s first freight contracts from inside a Barnes and Noble Starbucks - a favourite business meeting spot for the then-fledgling tech start up.
And according to TMZ, the couple did not get a prenup prior to their marriage. The TMZ sources, who claim to have direct knowledge of the situation, also added that the couple would be filing for divorce in Washington.
That means that MacKenzie is technically entitled to half of Bezos’ net worth, according to People. Since Washington is a community property state, “all assets and debts accrued during the marriage are considered owned equally by both spouses unless there was a prenuptial agreement stating otherwise”.