Technical nonylphenol (tNP), used for industrial production of nonylphenol polyethoxylate surfactants, is a complex mixture of C₃₋₁₀-phenols. The major components, 4-nonylphenols, are weak endocrine disruptors whose estrogenicities vary according to the structure of the branched nonyl group. Thus, accurate risk assessment requires isomer-specific determination of 4-NPs. Comprehensive two-dimensional gas chromatography/time-of-flight mass spectrometry (GC × GC/ToFMS) was used to characterize tNP samples obtained from seven commercial suppliers. Under optimal chromatographic conditions, 153−204 alkylphenol peaks, 59−66 of which were identified as 4-NPs, were detected. The 4-NPs comprised ∼86−94% of tNP, with 2-NPs and decylphenols making up ∼2−9% and ∼2−5%, respectively. The tNP products were analyzed for eight synthetic 4-NP isomers, and results were compared with published data based on GC/MS analysis. Significant differences were found among the products and between two samples from a single supplier. The enhanced resolution of GC × GC coupled with fast mass spectral data acquisition by ToFMS facilitated identification of all major 4-NP isomers and a number of previously unrecognized components. Analysis of tNP altered by the bacterium, Sphingobium xenophagum Bayram, revealed several persistent 4-NPs whose structures and estrogenicities are presently unknown. The potential of this technology for isomer-specific determination of 4-NP isomers in environmental matrices is demonstrated using samples of wastewater-contaminated groundwater and municipal wastewater.