MLB said Barrera, a 25-year-old who made his major league debut last September, tested positive for the performance-enhancing substance Dehydrochlormethyltestosterone (DHCMT) -- the chemical compound used in a drug that fueled Olympic athletes in the former East Germany decades ago.
Berrera's suspension is another element in a turbulent opening to Washington's defense of its 2019 World Series championship.
Hours before the Nationals lost their rain-shortened opener of the pandemic-delayed season to the New York Yankees on Thursday night, star slugger Juan Soto was placed on the COVID-19 injured list because he tested positive for the coronavirus.
Barrera said Saturday that he appealed his suspension but it was upheld by an arbitrator.
"From the initial failed drug test to losing the appeal, this all has come as a complete surprise as I have never, and will never knowingly use a banned substance," Barrera said in a statement released by the players' association.
"I have worked too hard and given too much to this sport to disrespect or cheat the game that I love. I would never do anything to dishonor my family, teammates, and coaches or do anything to let down the many young ball players that look up to me."
Barrera was on Washington's 40-man roster and part of its 60-man player pool.
He appeared in two games in 2019 and was not a member of the postseason roster as the Nationals went on to win the franchise's first title.
Barrera was selected by the Nationals in the sixth round of the 2016 amateur draft out of the University of Texas.
He is the seventh player suspended this year under the major league drug program, one shy of last year's total.
Washington was scheduled to host the Yankees in the second game of their season-opening three-game series at Nationals Park on Saturday night.