Sustrans, which developed and co-ordinates the National Cycle Network, said its use had risen in Scotland by seven per cent from 2012 to 2013.
It forms part of an all-over increase of seven per cent across the UK, as walkers and cyclists made 748 million journeys on the network.
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The figures come as operators of Glasgow's new hire cycles have reported an encouraging start to use of their bikes, which are already being used more frequently than London's famous Boris bikes.
John Lauder, national director of Sustrans Scotland, said: "Our report provides clear evidence of the value of the National Cycle Network in Scotland, and indeed across the whole of the UK.
"It is great to see so many people enjoying the many benefits of cycling and walking more often, from the health benefits of being more active to enjoying the convenience of knowing how long their journeys will take."
The National Cycle Network (NCN) is a 2,000-mile signed, mapped and promoted network of routes throughout Scotland. It stretches 14,700 miles around the UK and aims to provide a range of active travel options from everyday commuting to longer routes.
The report, which details the usage and benefits of the National Cycle Network in 2013, also said data indicated that new sections of the route in Scotland and Community Links projects are attracting more users than in other parts of the UK. Community links are links of three miles or less which Sustrans Scotland also operates.
Previously, a report by Sustrans Scotland had indicated that cycle routes, including St Mary's Industrial Estate, Dumfries, Goose Green in Musselburgh and Inveralmond Park Link had seen a significant increase in their usage, as did routes at the Golf Course Link in Troon, Uddingston Grammar School, and Bells Burn Path, Linlithgow.
Mr Lauder added: "This demonstrates that investment by the Scottish Government is paying dividends in meeting the demand for cycling and walking.
"Sustrans Scotland has worked, and continues to work, effectively with a large number of partners, including local authorities, to deliver a wide range of projects."
Sustrans also said the increase in 2013 delivered an overall benefit of £1 billion to the UK economy.
This includes £803 million in health benefits derived from physical activity, potential fuel savings for individuals valued at almost £215m and £25m worth of potential carbon saving.
The charity said more than one-third of users of the cycle network could have driven but chose not to, helping to ease congestion by saving 157 million car journeys.
It also said that older people accounted for around 10 per cent of journeys made on the network last year, with 55 per cent of pedestrians aged 65 or over and 57 per cent of cyclists in this age group feeling that the network helped them meet new people.
More than 285 million of the journeys last year were made by women, with 75 per cent saying the network helped them increase levels of physical activity.
Of the trips taken by women, 25 per cent were commuting journeys and 42 per cent were for recreation. The research indicated that women aged 45-54 are using the walking and cycling routes the most.
Sustrans chief executive Malcolm Shepherd said: "Official data shows levels of cycling and walking to be in long-term decline, but year on year we are seeing increases in the number of people taking to the National Cycle Network, both by bike and foot.
"The rise in the number of people using the network shows that there is a demand for safe, convenient and welcoming walking and cycling routes."