Limbaugh, who said on Monday that "it's up to me and Fox News" to kill immigration reform efforts, gave Rubio a respectful hearing but said he feared the senator was being exploited to advance Democratic goals.
Rubio defended his efforts as a shield against accusations that Republicans don't care about immigrants in case negotiations fail.
"If they want to go further than [the Senate agreement], I think they have a problem because they cant argue we haven't tried to do our part to come up with something that's reasonable," he said.
Responding to challenges from Limbaugh that Obama would demand reforms with fewer border security measures, Rubio emphasized his willingness to walk away from a bill if he didn't get what he wanted on that front. In particular, he said including enforcement measures as a "trigger" for undocumented immigrants to seek permanent residency was key.
"Unless there's real enforcement triggers we are not going to have a bill that moves on the opportunity to apply for a green card," Rubio said. He added: "I'm not going to be part of a bidding war to see who can put the most lenient path forward" if Obama demands a smoother path to citizenship.
As for the political benefit to Republicans of an agreement, Rubio said he believed "given a fair chance, I can convince most Americans including Americans of Hispanic descent that limited government and free enterprise is better for them and better for their upward mobility than big government is - because that's the reason they came here."
Limbaugh questioned the political utility of a deal, saying he had reviewed "scholarly research" indicating that immigrants no longer came to America to better their lot through hard work but instead because they believed "government is the source of prosperity." In short, they were not viable Republicans.
"It's not about conservative principles and so forth, not the way it used to be," he said, referring to immigration patterns. Rubio responded that he felt he could sell Hispanics on a small government philosophy.
In addition to his fear that immigrants may lack a basic sense of industry and responsibility, Rush added that he worried Republicans would be falsely attacked as intolerant.
"My concern is the president wants people to believe something that isn't true ... and that is that you guys are not being truthful in what you say, that you really don't want an improved life for Hispanics, that you really are still racist," he said.
Rubio repeated his argument that preempting Obama protected Republicans from those charges.
"I think outlining our principles is important because it takes away the ability of the other side to mischaracterize what we are for and what we are against," he said.