Reforms under Pope Francis, by the Institute for Religious Works, has tried to make the church be a less a profit-at-all cost investment vehicle and more a service-oriented foundation.
The pontiff wants the bank to help religious orders and Vatican officials better-manage their money.
The Vatican said the resignation of Clemens Borsig and Carlo Salvatori from the bank's board "can be seen in light of legitimate reflections and opinions" about the management of such an unusual institution.
The bank has been going through a years-long process to comply with international anti-money laundering norms after a series of scandals.
Last year the Vatican's bank manager resigned and the institution had its accounts frozen by the state's top prosecutor amid allegations of embezzlement.
In the past few years the organisation has been through massive restructuring under Pope Francis as he attempted to stamp out corruption.
Prior to Francis' reorganistion there were allegations that the Vatican Bank was used by money launders forced Pope Benedict to start a process of reform at the organisation.
Since the election of Pope Francis the bank has been ordered to be more transparent.
