Despite the extensive archaeological surveys carried out in the last decades, little attention has been paid to one of the longest water supply systems of ancient times – the Byzantine water infrastructure which fed Constantinople from the mid-late fourth century ad. This work uses modern satellite terrain data and Global Positioning System (GPS) data to assess this system and provide an improved description of its route, total length and gradient profile. Forty-four validated GPS Control Points were correlated with ASTER GDEM V2 digital satellite data and archaeological information in a Geographic Information System (GIS) environment. We concluded that the total length of the water supply system was 426 km, and possibly even 565 km if the fifth-century aqueduct continued in parallel all the way to Constantinople rather than merging with the fourth-century aqueduct. The gradient of the channels varied across their length, being steepest near the spring sources, with a gradient mostly in the region of 5 m/km, and flattest at around 0.4 m/km in the most downstream section nearest the City. This reconstruction of the gradient profile provides valuable insight into the physical characteristics of the system, allowing future study of its hydraulic function.