Glyphosate [(N-phosphonomethyl) Glycine], one of the most worldwide commercialized herbicides, has provoked many debates about its carcinogenic effects. Here, a smartphone-based surface plasmon resonance (SPR) sensor is proposed for glyphosate detection using different pH and concentrations. CuO nanoparticles have been added to glyphosate samples, diluted in ultra-pure water solutions, to enhance its detection. An increase of sensitivity was observed in acidic solutions reaching a dilution of 10−8 (v/v), which is equivalent to 5·10−7 ppm. This novel smartphonebased SPR device for glyphosate detection besides presenting very high sensitivity, it has also favorable features such as easy handling, portability, and real-time analysis.