The Old Walnut Street Associate Presbyterian Church had its origins in the Scots Presbyterian Church, also called the First Associate Scots Church. In February of 1764 the first pastor, William Marshall began preaching at a hall in Videll's Alley. In 1770 the congregation purchased land on the south side of Spruce Street between 3rd and 4th and erected a building in 1771-72. John Adams attended the church once in 1777 and wrote a revealing letter about his experience among the congregation of Scots who were "not very genteel" and their imposing pastor, William Marshall, who was a "patriot," a supporter of the Revolutionary War. When the British occupied Philadelphia in 1777 Hessian troops took over the church building for use as a hospital.

The Reformed Presbyterian Church (Covenanters) and the Associate Presbyterian Church (seceders) merged in 1782 to form the Associate Reformed Presbytery of Philadelphia. At that point the First Associate Scots Church became the First Associate Reformed Church (Scots), but the leader of the Associate dissidents opposed to the 1782 merger was none other than William Marshall himself. He was forced out, and started a new Associate Presbyterian congregation.

In 1790, David Clark and William Young purchased for their Associate Presbyterian congregation a lot of ground on the north side of Walnut street, between Fourth and Fifth streets, being 36 feet 10 inches wide, by 124 feet deep — and shortly afterwards a brick meeting house was erected, with the residue of the lot being reserved for a burying ground. On the 2d day of August, 1802, John Cummings sold two lots of ground on the west side of 13th street, south of Spruce street, as an additional burying ground for the congregation.

Marshalls' congregation enjoyed a period of stability before undergoing a further schism in 1845. They left the Associate Church under the leadership of another fiery minister, Reverend Chauncy Webster, forming a tiny short-lived denomination called the Independent Associate Presbytery of Philadelphia with a total of three pastors.

The church and burial ground on Walnut Street were taken down in 1854 when they moved to a new church called The First United Presbyterian Church on the southwest corner of Broad and Lombard Streets.

In 1864 and 1869, burials at the second location at 13th and Walnut were removed to Mount Moriah Cemetery.