But the international Red Cross, which is due to coordinate the operation, said it had no information on what the trucks were carrying or where they were going.
That could raise fears in Ukraine and the West, where leaders have voiced concerns that Russia could use the initiative as a pretext for sending troops into rebel-held territory.
Russian television and news agencies reported Tuesday that 2,000 tons of aid was en route to Ukraine, where fighting between pro-Russian separatists and government forces has claimed more than 1,300 lives since April, according to a UN report.
A Russian Orthodox Priest sprinkled holy water on the trucks, some of which bore a red cross, before their departure. They could take up to a day to arrive at their destination.
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Andre Loersch, a spokesman for the International Committee of the Red Cross mission in Ukraine, said that while the organization had reached a general agreement about delivery of humanitarian aid to the region, he had "no information about the content" of the trucks and did not know where they were headed.
It was unclear whether the trucks would cross the border into the rebel-controlled territory of Luhansk or the government-controlled territory in Kharkiv. At least 100 kilometers (60 miles) of the border is currently in rebel hands."
Yesterday, the ICRC issued a statement saying that it was ready to facilitate the aid operation with the involvement of all sides concerned.
However, Laurent Corbaz, the ICRC's head of operations for Europe and central Asia, said that "practical details of this operation need to be clarified" before the deliveries can move ahead.