Deputies said the mare was probably in "extreme distress and pain" when she died.
The Maricopa County Sheriff's Office have arrested New River residents Laurel George and John Henkle, both 54.
The pair were booked into the local jail on two counts of felony animal cruelty, two of misdemeanor animal cruelty and a class six felony for cruelty to an animal that causes protracted suffering.
A malnourished 15-year-old horse was also found on the pair's property and has been taken to a no-kill animal-care facility operated through the sheriff's office and run by inmates.
Deputies allege the 26-year-old Arabian mare was killed in early March.
An examination of the dead horse revealed that the mare died through a homemade injection of pool acid and sodium bicarbonate.
Police said in a statement they believed the method resulted in "extreme distress and pain" to the horse.
The surviving horse is now being housed at Sheriff Joe Arpaio's no-kill animal shelter, called MASH, which was created to house and care for animals that have been abused or neglected by their caretakers and rescued by the Animal Cruelty Investigative Unit.
Its aim, pending the outcome of court cases, is to have the animals rehomed into loving, permanent homes.
The shelter is operated in a former jail no longer suitable for housing inmates. It is run by inmates.
The pool acid injection was mixed with sodium bicarbonate, deputies said.