The mini-sub was deployed late on Tuesday, but was forced to resurface due to a technical issue, officials said. It has since been redeployed.
The data downloaded from the mini-sub on Wednesday had "no significant detections", Australian officials add.
Flight MH370 went missing on 8 March with 239 people on board.
Air traffic controllers lost contact with it over the South China Sea while it was en route from Kuala Lumpur to Beijing.
Based on satellite data, officials believe it ended its flight thousands of kilometres off course, in seas west of the Australian city of Perth.
However, so far not a single piece of debris from the jet has been found.
'No detections'On Monday, the US Navy's Bluefin-21 robotic submarine was sent on its first mission to search the sea floor for wreckage after signals believed to be consistent with "black box" flight recorders were detected.
Other ships and aircraft are scouring the search area for debris But the drone exceeded its operating limit of 4,500m (15,000ft) and built-in safety features returned it to the surface. "No objects of interest were found," Australia's Joint Agency Co-ordination Centre (JACC) said.
The mini-sub was again deployed on Tuesday from the Australian Defence Vessel Ocean Shield.
It was forced to resurface on Wednesday morning to rectify an unspecified technical issue and redeployed, JACC said.
"Initial analysis of the data downloaded this morning indicates no significant detections," it said.
The 5m-long Bluefin-21 can create a sonar map of the sea floor.
The US Navy has estimated that it could take the unmanned submarine from six weeks to two months to scan the search zone.
Continue reading the main story 8 March: Malaysia Airlines Kuala Lumpur-Beijing flight carrying 239 people disappearsPlane's transponder, which communicates with ground radar, was switched off as it left Malaysian airspaceSatellite 'pings' indicate plane was still flying seven hours after satellite contact was lost24 March: Based on new calculations, Malaysian PM says "beyond reasonable doubt" that plane crashed in southern Indian Ocean with no survivorsSome 11 military planes and three civilian planes were also flying out of Perth on Wednesday to scour the Indian Ocean for floating debris.Isolated showers are forecast in the search area with sea swells of up to 2m.
In another development, officials are investigating an oil slick about 5.5km (3.4 miles) from the area where the last underwater signals were detected.
An oil sample has been sent back to Perth for analysis, a process that will take several days, said Air Chief Marshal Angus Houston, who heads the JACC.
Australian officials have said they are confident they are searching in the right area for the missing plane.
But ACM Houston warned on Monday that the search of the sea floor could be a long, painstaking process that might not yield results.
Officials so far have no idea why the plane diverted so far from its intended flight path. Investigators are looking at theories including hijacking, mechanical failure, sabotage and pilot action.
