In 2014, incumbent State Representative Carl Zimmermann, a Democrat, ran for re-election in the 65th District, so Sprowls ran against him. He faced fellow attorney Debbie Ann Faulkner in the Republican primary. The Tampa Tribune endorsed Sprowls over Faulkner, which noted that, though "both candidates bring intelligence and passion to the race," Sprowls "is the more polished candidate and appears ready to serve on day one if elected." Sprowls ended up defeating Faulkner by a wide margin, winning 65% of the vote, and advanced to the general election, where he faced Zimmermann. Sprowls campaigned on his opposition to Medicaid expansion provided for under the Affordable Care Act, his support for charter schools, and his opposition to abortion. Ultimately, owing to the tendency of the district to vote for Republican candidates, Sprowls unseated Zimmermann, winning his first term in the legislature with 52% of the vote.

On September 17, 2019, Sprowls was chosen by his Republican colleagues to be the speaker of the Florida House for the 2020–2022 legislative term. The full House formally elected him speaker on November 17, 2020, after the 2020 elections.

During the 2020 elections, amid the COVID-19 pandemic, he participated in indoors Donald Trump campaign events where attendees and participants did not wear masks or socially distance.

In 2021, Sprowls praised Florida Governor Ron DeSantis for his handling of the COVID-19 pandemic.

After Florida voters voted in favor of a referendum to restore the voting rights of felons, Sprowls and other Republicans restricted this so that felons who had outstanding fines would not regain their voting rights. Sprowls argued that this restriction did not amount to a "poll tax", arguing that poll taxes had racial implications.