The detention of observers from the Organization for Security and Co-operation in Europe's special monitoring mission in Ukraine "cannot be tolerated", Burkhalter said in a statement.
He described their detention as "acts of sabotage of international efforts to assist Ukraine in overcoming the crisis".
Burkhalter called for their "immediate and unconditional release".
The Vienna-based Organisation for Security and Co-operation in Europe said on Tuesday that it had lost contact with a team of four observers, monitoring the situation in restive eastern Ukraine.
The team — a Dane, an Estonian, a Turk and a Swiss national — were apparently detained at a checkpoint shortly before contact was lost.
Ukraine's foreign ministry has, meanwhile, accused rebels of holding them.
On Wednesday, an OSCE spokesperson confirmed there still hadn't been any contact with them.
A second group of 11 observers was detained in the same region of Donetsk on Wednesday but the OSCE later said it had managed to re-establish contact with them.
"We can't confirm their status until we have them back with the rest of the mission, safe and sound," an OSCE spokesperson said in Vienna.
Concern about the status of the observers arose as reports emerged that pro-Russian rebels shot down a Ukrainian helicopter later Wednesday, killing 12 soldiers, including a general.
The attack dealt Kiev its biggest setback since president-elect Petro Poroshenko's vow to crush the seven-week eastern insurgency quickly.