The Doctor Who and Torchwood actor won the first poll from PetNet after four weeks of voting, beating competition from Alesha Dixon and Paul O'Grady, among others.
"I'm delighted to receive this award," Barrowman said. "My dogs play such an important role in my life and I am proud to promote responsible ownership.
"I will be donating my prize money to Dogs Trust. As their patron, I understand how incredibly hard they work to find new homes for stray and abandoned dogs."
One voter said: "Not only does John look after his own three adorable dogs he also helps dogs at Dogs Trust and does a lot for dogs more than anyone will know. If anyone deserves to win this John does."
Meanwhile, '60s model Celia Hammond won the PetNet award for top 'Celebrity Cat Owner', beating McFly's Tom Fletcher.
"I am honoured to receive this award but am even more delighted that our wonderful supporters came out in force to ensure the Trust benefited from this donation," Hammond said.
"This will enable us to help even more animals that we rescue from the streets and help low income pet owners have their pets treated."
One of her fans said: "Celia should win this hands down. I remember her when she was a super model in the '60s and since then she's dedicated her life to saving and caring for cats and dogs and other animals."
Head of digital at PetNet Lisa Robb said: "We are delighted that John Barrowman and Celia Hammond have won our first PetNet Celebrity Pet Owner Awards.
"From an original list of 82 nominees, 12 celebrities were shortlisted by our expert panel of judges and the overwhelming support of their fans has resulted in two very deserving winners.
"We are delighted that Dogs Trust and the Celia Hammond Animal Trust will receive £2,000 each plus a photoshoot thanks to Venture Photography."