Prosecutors said Dr Virginia Soares de Souza and her medical team administered muscle relaxing drugs to patients, then reduced their oxygen supply, causing them to die of asphyxia at the Evangelical Hospital in the southern city of Curitiba.
De Souza, a 56-year-old widow, was arrested last month and charged with seven counts of aggravated first degree murder. Three other doctors, three nurses and a physiotherapist who worked under De Souza have also been charged with murder.
Prosecutors for the state of Parana said wiretaps of De Souza's phone conversations revealed that her motive was to free up hospital beds for other patients.
"I want to clear the intensive care unit. It's making me itch," she said in one recording released to Brazilian media. "Unfortunately, our mission is to be go-betweens on the springboard to the next life," she added in the same phone call.
De Souza's lawyer, Elias Mattar Assad, said investigators had misunderstood how an intensive care unit works and she would prove her innocence.
More cases are expected to emerge as investigators comb through 1,700 medical records of patients who died in the last seven years at the hospital, where De Souza headed the intensive care unit.
"We already have more than 20 cases established, and there are nearly 300 more that we are looking into," the chief investigator assigned by Brazil's Health Ministry, Dr. Mario Lobato, said on Globo TV's Fantastico program on Sunday.
If prosecutors prove that De Souza killed 300 patients, this could be one of the world's worst serial killings, rivalling the notorious case of Harold Shipman, the British doctor who was found to have killed at least 215 patients.
Lobato said the deaths he reviewed occurred under similar circumstances: a muscle relaxant such as Pancuronium (trademark Pavulon) was administered, increasing the patients' dependence on artificial respiration; then the oxygen supply was reduced, causing death by asphyxia.
Some of the patients were conscious moments before they died, he said.
Prosecutors said De Souza felt "all powerful" running the intensive care unit homicide, to the point where she "had the power to decree the moment when a victim would die".
In some cases, De Souza was absent from the hospital and gave instructions to end the life of a patient by telephone to members of her medical team, according to documents detailing the charges.
Last week, a Curitiba judge ordered the release of De Souza and her medical team. Prosecutors sought on Monday to have her returned to custody because she was the leader of the team and witnesses had reported being intimidated.
Parana state prosecutors asked police on Wednesday to investigate whether more hospital employees, including former managers, were involved in the case.
President Dilma Rousseff's government will announce steps on Thursday to reorganise the hospital, a spokesman for the Health Ministry said.