Save the Children says its research suggests 150,000 women could lose up to £68 a week when the new universal credit takes effect next year.
The report also claims second earners will be affected.
But the Department for Work and Pensions said 600,000 lone parents would be better off under a system that “incentivised work and made work pay”.
Save the Children says single mothers on low incomes would be forced to make ends meet by either working longer hours or by getting into debt.
The charity’s report - Ending Child Poverty - claims the changes would make it less attractive for parents to come off benefits and into work because of poor childcare support.
It also suggests that couples where both work part-time in low paid jobs would be hit by the changes
The charity is urging the Chancellor, George Osborne, to take action in next week’s budget to head off the problems and ensure that single mothers keep more of their income before losing benefits.
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The truth is 600,000 lone parents will be better off under a system which will incentivise work”
Department for Work and Pensions Chief executive of Save the Children Justin Forsyth said: “Universal credit will help some families, but mums working hard to stay above the breadline are its big blind spot.
“It’s incredibly hard bringing up three kids on £370 a week - losing almost a fifth of that will push many families over the edge.
“The government must make sure mums who want to work keep more of their incomes and get more support with childcare.
“Otherwise we’ll see fewer women in the workplace and more children growing up in poverty.”
‘Better-off’ By the time universal credit is fully implemented, the government expects 900,000 people to be lifted out of poverty.
A spokeswoman for the DWP said the charity’s claims were based on hypothetical examples and it was wrong to assert that lone parents would lose out under universal credit.
“The truth is 600,000 lone parents will be better off under a system which will incentivise work and make work pay”, said the spokeswoman.
“This is in stark contrast to the broken system this government inherited which only rewards lone parents who work 16 hours or more.
“Under universal credit 80,000 more families, including lone parents, will be able to claim childcare support - no matter how few hours they work,” she added.
The government did however admit that payments to some new claimants would be lower under the new system.
Shadow employment minister Stephen Timms said the government must work harder to get universal credit right.
“The best way to get children out of poverty is to get more parents in work,” he said.
“But as this report shows, their current plans will lock in a parents’ penalty, chip away at the incentives for thousands to work and push 150,000 working parents deeper into poverty.”
Published by BBC News - 13 March 2012 Last updated at 12:22 See: http://www.bbc.co.uk/news/education-17351634