The application of 1H nuclear magnetic resonance (NMR) spectroscopy to characterize and quantitate paraquat in urine is described. Characterization was performed taking advantage of two NMR spectroscopy parameters: chemical shifts and coupling patterns. Without any pretreatment of the biological samples, herbicide was detected by its aromatic doublets at 8.49 and 9.02 ppm. Quantitation of the xenobiotic was realized by relative integration of the dipyridyl protons to an internal standard. After a validation step using control urine samples, quantitation was performed in urine obtained from two poisoned patients. On admission, mean paraquat concentrations were 985 (patient 1) and 500 (patient 2) micromol/L. Results are compared and found to be in good agreement, using a second-derivative spectroscopy method.