The minister holds portfolio responsibility for gambling laws and the Australian Communications and Media Authority.
On Thursday, Nine newspapers reported Sportsbet paid $8960 for a place at a dinner held to support Ms Rowland's 2022 federal election campaign.
Three days before the election, Sportsbet also donated $10,000 to her campaign.
While the betting giant disclosed the $10,000 donation to the Australian Electoral Commission, Labor's NSW branch was not required to declare the payments because they were under the $14,500 mandatory disclosure threshold.
In Question Time, independent MP Zoe Daniel asked the minister to confirm the reports and explain how they were not a conflict of interest.
Ms Rowland, who did not directly address the claims, said every member of parliament had to comply with the rules of disclosure.
"I have and will continue to comply with the disclosure requirements of the AEC, the register of members' interests and the ministerial code of conduct," she said.
Independent MP Andrew Wilkie said it was "unfathomable" for Ms Rowland to think it was acceptable to take donations from an online gambling company.
He noted that at the time of the donations Ms Rowland was Labor's communications spokeswoman and was expected to become communications minister if the party won the election.
"Michelle Rowland reportedly made shocking errors of judgment when she accepted a number of big donations from Sportsbet before the most recent federal election," Mr Wilkie said.
"(She) is now deeply conflicted and it is patently untenable for her to remain in her position."
Mr Wilkie urged Prime Minister Anthony Albanese to sack the minister and return the funds to Sportsbet.
Greens senator Larissa Waters said the minister had not broken any laws, but that showed how weak Australia's donation rules were.
"It is clearly untenable for the minister to remain as gambling regulator when she has such a cosy relationship with betting agencies," she said.
"These donations are an insult to voters and gambling reform advocates across Australia."