Continuing his rant to his 497,000 followers, the Arrow actor went on: "It's absolutely ridiculous and every customer behind me thought it was ridiculous also.
"That gentleman just needed a bit of help to get him up, to make his day a little better, and Target, you were trying to deny that, so I'm a little p***ed off with you right now."
Barrowman tagged the official Target account in his post, who replied swiftly to apologize and tell him they were going to investigate.
They have since released a statement. It read: "We're very sorry for the experience you had at the LA Target store.
"We absolutely do not have a policy against this type of purchase, and are addressing it immediately with the team members involved.
"Thank you for bringing this to our attention."
The homeless population in Los Angeles has surged more than 75 per cent in the past six years, as California grapples with a homelessness crisis.
Official estimates of the city's homeless population run about 60,000, but outreach workers say the true number is even higher.
In Barrowman's initial tweet, he captioned the video: "I am disgusted by what I was told. Arrest me for trying to help someone. The stupidity behind that rule. I want an explanation. My fans and followers please message #Target regarding this. JB."
And indeed, his fan-base chimed in, praising the star for taking on the retail chain.
"Well done John, what a disgrace, where is compassion in this world, looks like common sense is not on targets agenda [sic]," one person posted.
Another replied: "That is awful!!!!! You have every right to purchase whatever you want for whomever you want! Thank you for being a kind human being!!! [sic]"
Another sympathised with the homeless man Barrowman helped out: "This is why I love you, John. Your heart is huge. From someone who's been there, thank you for helping that man. [sic]"
Target has stores across the US at 1,828 locations [as of 2017] and invested a massive $7billion ($9.6B) in its shops, a revamped digital strategy and private label pushing since 2015.
In 2017, Target returned $591million ($813.4M) to shareholders in the fourth quarter through dividends and share repurchases, bringing the total to $2.4 billion ($3.3M) for full-year.