Foreign Affairs Minister Marise Payne says there are about 1400 Australians in Ukraine and they should use commercial means to leave the country if safe to do so.
"This is a cautious and prudent step because the security situation is unpredictable," Senator Payne told the ABC on Tuesday.
"It is about protecting Australians on the ground."
The Department of Foreign Affairs and Trade on Monday night raised its advice to "Do Not Travel".
"Australians in Ukraine should leave now by commercial means, where safe to do so, noting that flight availability could change or be suspended at short notice," it said in a statement.
"Australians who decide to remain in Ukraine should review their personal security plans, be prepared to shelter in place if required, maintain heightened security awareness and register with DFAT."
Senator Payne said the decision by DFAT was sparked by consultations with "like-minded counterparts in Kyiv and on the subject of Ukraine".
"But importantly this decision doesn't change one iota our steadfast support for Ukraine sovereignty and territorial integrity."
DFAT's advice follows similar moves by the United States and the United Kingdom.
But Ukrainian diplomat in Australia Volodymyr Shalkivskyi urged people not to panic, saying pulling people out of the country "sends the wrong message".
"We would like to avoid panic and rumours that the invasion is inevitable," the Ukrainan embassy's Chargé d'Affaires told the ABC.
"The official responsibility of Australian and Ukrainian officials is to facilitate and assist in every possible way. But in this situation we would like to avoid panic and different kinds of rumours that the invasion is inevitable."
Tensions in Ukraine have been increasing for months after the Kremlin massed about 100,000 troops near Ukraine's borders, a build-up the West says is preparation for a war to prevent Ukraine joining NATO.
The Kremlin has repeatedly denied planning to invade.
Senator Payne again ruled out military assistance but said Australia will consider providing cyber security support.
"There has been a significant cyber attack already on Ukraine, understood to have come from potentially Russian sources," she said.
"Australia has been working closely with partners in the international community, including with Ukraine, on cyber resilience and cybersecurity, and if Australia can assist in that regard we will."
Further sanctions are also under review.