Tim Farron says there will be no pacts with other parties after two senior Lib Dems were recorded discussing support for a Labour candidate.
Ex-business secretary Sir Vince Cable said he would find it "difficult to vote against" a Labour candidate whose views were "very close" to his own.
Lib Dem Richmond Park candidate Sarah Olney suggested the use of "paper candidates" or "not campaigning".
But Mr Farron said: "Let me be clear: no pact, no deal, no coalition."
In a recording from a question-and-answer session obtained by the BBC, Sir Vince, who is the party's candidate in the Twickenham seat he lost in 2015, said he had a good relationship with people from other parties, adding: "I particularly met a lot of people on the Labour side whose views are very close to mine."
He gave the example of a conversation with Rupa Huq, who is seeking re-election for Labour in Ealing Central and Acton.
"We talked for a couple of hours, and it was very clear that on almost every issue our views were almost identical," he said.
"And so I would find it difficult to vote against somebody like that, and I hope that our people around the country are discriminating and think and act in a constructive way."
Also in the recording, Ms Olney warned that the Conservatives were trying to make a so-called "progressive alliance" appear to be "a negative thing".
She added: "We do have to be very conscious about how some of our efforts are viewed, and being tactical isn't just about standing down or voting for the right candidate, it also can be about paper candidates, but not campaigning...
"Certainly I know that I've had a lot of Ealing Lib Dems coming down here, and I'm sure they've been over in Twickenham as well, because yes - we may be standing candidates there, but we know that we want Rupa to win in Ealing, and so on and so forth."
Labour gained Ealing Central and Acton from the Tories in 2015 by 274 votes, with the Lib Dems adrift in third.
The Green Party is not fielding a candidate, as part of its call for an anti-Tory alliances in some seats.
The constituency's Lib Dem candidate, Jon Ball, claimed only two results were possible in the seat - a Conservative gain or a victory for him. Labour did not have a chance of holding the seat, he said.
Asked by ITV News if he was embarrassed by the recorded comments, Mr Farron said that "on the ground", it was right that politicians of all political parties "actually treat each other like human beings, not members of different isolated tribes".
But, he said, "it's also important that we as a party are very clear in our message there will be no deal, no pacts, no coalitions.
"It's about the Lib Dems being the only serious and clear alternative to a Tory landslide."
But the Conservatives, who have repeatedly warned of a "coalition of chaos" despite Labour and the Lib Dems ruling out a coalition, seized on the recorded remarks.
Work and Pensions Secretary Damien Green claimed the recording showed the Lib Dems were "scheming" to make a coalition happen.
Labour, meanwhile, has excluded three activists who appeared in a BBC report backing a rival party's candidate who is standing against Health Secretary Jeremy Hunt.
The trio had told BBC Radio 4's Today programme they supported a "progressive alliance" in favour of the National Health Action Party candidate in South West Surrey.