The aim of this study was to investigate oxidative stress, biotransformation and bioaccumulation of polycyclic aromatic hydrocarbons (PAHs) in tropical estuarine food webs including fish (Chrysicthys nigrodigitatus), fiddler crab (Uca tangeri), blue crab (Calinectis amnicola), prawn (Macrobrachium vollenhovenii), periwinkle (Tympanotonus fuscatus) and sediment samples at three sites, Adiabo (control site), Obutong and Nsidung representing different degrees of anthropogenic contamination along Cross River Estuary, Nigeria. Hepatic oxidative stress and biotransformation enzyme activities glutathione peroxidase (Gpx), glutathione reductase (Gr), glutathione S-transferase (Gst), uridine diphosphate glucuronosyltranferease (Udpgt), 7-ethoxy-, methoxy-, pentoxy-, and benzyloxyresorufin O-deethylase (EROD, MROD, PROD and BROD) and PAHs levels were determined. Data demonstrated species- and site-specific mediated toxicological effects in oxidative stress, biotransformation responses, and PAHs bioaccumulation in biota and sediments from contaminated sites (Obutong and Nsidung), compared to control (Adiabo). The EROD, MROD, BROD, PROD activities and GPx, Gr, Gst, Udpgt exhibited significant increase in biota collected from contaminated sites at Obutong and Nsidung compared with control Adiabo. These biomarker response observations paralleled PAHs accumulation at Obutong and Nsidung suggesting PAHs exposure induced oxidative and biotransformation biomarker responses. Principal component analysis (PCA) produced significant associations between variables indicating sites were major factors determining contaminants uptake and biomarker responses in biota (fish, crabs, prawn and periwinkle). Data demonstrated site and species-specific occurrence and concentrations of PAHs in sediment and tropical estuarine food webs with corresponding biotransformation and oxidative stress responses on resident biota. Concentrations of PAHs detected in these tropical food webs indicate serious human food safety and environmental health concerns.