The Prime Minister was speaking in the Commons before a debate on the bill to trigger Article 50, the process of leaving the European Union.
MPs are expected to debate possible amendments to the bill into the early hours of Tuesday morning.
Rebels from the Conservative Party have threatened to vote with Labour and the SNP on changes to the EU (Notification Of Withdrawal) Bill.
"The House has spoken and now is not the time to obstruct democratically expressed wishes of the British people," Mrs May told MPs.
She also spoke about discussions with leaders of the other 27 EU states in Malta at the weekend.
She said EU leaders welcomed the UK's clarity of objectives set out for the negotiations and the ambition to build a new partnership in the interest of both sides.
Tories who wanted to remain in the EU want assurances that Parliament will get a vote on the "endgame" if negotiations with other member states collapse without a deal.
Mrs May has already pledged to give Parliament a vote on any agreement she reaches in Brussels.
Former ministers Anna Soubry and Dominic Grieve were reported to be among the potential rebels, alongside veteran pro-European Ken Clarke, the only Tory to oppose the Bill in last week's second reading vote.
Asked earlier whether Mrs May was minded to accept any of the tabled amendments to the bill, the Prime Minister's official spokeswoman said: "We've been very clear, we think this should be a straightforward bill about giving the Government the power to deliver on the decision of the British people.
"Parliament has already legislated to put the decision in the hands of the British people and we are not going to allow there to be attempts to remain inside the EU or to rejoin it through the back door.
"The Government has a very clear approach, it has a mandate from the British people for the UK to leave the EU and to carry out that decision."
Labour leader Jeremy Corbyn is also expected to face rebellion again after ordering his MPs to back the passage of the bill. Forty-seven defied the leadership last week.