The civil union law was debated in Congress for over a decade until it was passed and signed into law by the president in April.
As it went into effect, couples began arriving at civil registry offices early to officially validate their unions.
Some wore matching traditional Chilean cowboy garb, others elegant tuxedos and dresses.
"It was beautiful. It was such a nice ceremony. It was all very emotional. Our families were here, everyone was shedding tears," Virginia Gomez told reporters after she registered her union with her partner, Roxana Ortiz.
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The couple had married in Spain but their union was not recognized in Chile.
"Now we can make decisions together like a couple. We're thrilled."
Civil union gives same-sex and unmarried couples many of the rights granted to married couples.
Partners can inherit each other's property, join one another's health plans and receive pension benefits.
They have been recognized in several South American countries, though only Argentina and Uruguay allow formal gay marriage.
Gay advocates in Chile are celebrating the right to same-sex civil unions as a step toward full rights.
Chile decriminalized gay sex in 1999 and it was one of the last countries in the world to legalize divorce, in 2004.
The brutal killing of a gay man in 2012 set off a national debate that prompted Congress to pass a hate crimes law.