Prosecutors shut down a multibillion-dollar Internet gambling ring and charged 27 with involvement in a criminal enterprise, including a baseball scout for the Washington Nationals and a professional poker player.

Charges were brought against the top echelon of individuals associated with Playwithal.com, a sports gambling site run by the high-stakes poker player James Giordano, Police Commissioner Raymond Kelly and District Attorney Richard Brown of Queens said yesterday. Also named in the indictment was Major League Baseball scout, Frank Falzarano, whom prosecutors named as one of the enterprise’s highest bet processors.

Prosecutors allege the enterprise employed more than 2,000 people, booking more than $3.3 billion in wagers during a 28-month police investigation. In addition to charges filed against 27 individuals, prosecutors indicted three corporations and filed for $500 million in asset forfeiture, one of the largest cases ever filed, they said. The proceeds would go to federal, state, and city municipalities.

The arrests are the first since President Bush last month signed the Unlawful Internet Gambling Enforcement Act, which prohibits Americans from using credit cards to place online bets. Included in yesterday’s indictment were bookies accused of accepting cash wagers in exchange for access to Playwithal.com, where gamblers placed bets, prosecutors said.

“This is the largest illegal gambling operation that this department has ever encountered,” Mr. Kelly said. He said search warrants executed during the past 24 hours resulted in the seizure of more than $1.5 million in cash, casino chips, art, and sports collectibles, including a football signed by the 1969 Super Bowl champion Jets. More arrests and seizures are expected.

Officials said they launched the investigation in 2005, after they were tipped off to the scheme while investigating Queens pharmacists who allegedly worked with Bonanno crime family drug dealers. Further inquiry uncovered a multi-tiered network of bookies, money collectors, and distributors headed by Mr. Giordano.

Police also learned that the enterprise employed two technology companies named in the indictment — Prolexic Technologies Inc. and Digital Solutions — to protect their system.

In describing the scope of the enterprise, officials said it operated in Queens and elsewhere in the city, as well as across the country and overseas. The network’s top distributor, Donald Clarke, processed $652,000 in bets in one week, they said. Low-level bookies collected between $8,000 and $20,000 weekly.

“The operation was massive,” Mr. Brown said. The scheme relied on Internet, e-mail, and multiple toll-free numbers that allowed “constant and immediate access to betters around the country and overseas,” he said.

The attorney for at least one defendant downplayed the indictment and called the arrests a targeted campaign. “I’ll reserve comment on this case until all law enforcement are finished patting each other on the back about the arrests of more Italian people for gambling during football season,” the attorney, Vincent Romano, said. Mr. Romano’s client, Rocco Renelle, pleaded not guilty on Tuesday.

However, prosecutors said they hoped the arrests would diminish the online gambling industry. The American Gaming Association estimated online gambling is a $12 billion industry, with more than 2,000 online gambling sites worldwide. About 8 million Americans gambled online in 2005, according to the market research firm Christiansen Capital Advisors. “It’s been said of Internet gambling that once you click your mouse, you can lose your house,” Mr. Brown said.

Given other recent crackdowns, the head of at least one online gaming company said his company came up with a legal alternative to comply with regulatory oversight. “Since I am a legal alternative, I approve of a thoughtfully regulated and strictly enforced gaming regulatory alternative,” the CEO of GameLogic, Steven Kane, said. “It’s good for the consumer, it’s good for the industry.”