New data for the isotopic composition of carbon in carbonate sediments deposited between 2.6 and 1.6 Ga indicate that the value of δ13C in these sediments underwent a very large positive excursion between 2.22 and 2.06 Ga. A reassessment of the earlier δ13C data for carbonate sediments shows that this excursion was probably worldwide, and that it was preceded and followed by several hundred million years during which the δ13C of carbonate sediments differed little from that of modern carbonates. The large δ13C excursion between 2.22 and 2.06 Ga was probably related to an abnormally high rate of organic carbon deposition, which generated an abnormally high rate of O2 production. We estimate that the total excess O2 produced during the excursion was between 12 and 22 times the present atmospheric O2 inventory. The δ13C data therefore suggest that the O2 content of the atmosphere increased very significantly between 2.22 and 2.06 Ga. This inference is supported strongly by several other lines of evidence.