Novel Cr(III)-imprinted poly(vinyl alcohol)/sodium alginate/AuNPs hydrogel membranes (Cr(III)-IIMs) were obtained and characterized and further applied as a sorbent for chromium speciation in waters. Cr(III)-IIMs were prepared via solution blending method using blends of poly(vinyl alcohol) and sodium alginate as film-forming materials, poly(ethylene glycol) as a porogen agent, sodium alginate stabilized gold nanoparticles (SA-AuNPs) as a crosslinking and mechanically stabilizing component, and Cr(III) ions as a template species. The physicochemical characteristics of pre-synthesized AuNPs and obtained hydrogel membranes Cr(III)-IIM were studied by UV-vis and FTIR spectroscopy, TEM and SEM observations, N2 adsorption–desorption measurements, and XRD analysis. The mechanism of the adsorption process toward Cr(III) was best described by pseudo-first-order kinetic and Langmuir models. Experiments performed showed that quantitative retention of Cr(III) is attained in 20 h at pH 6 and temperature 40 °C. Under the same conditions, the adsorption of Cr(VI) is below 5%. A simple and sensitive analytical procedure was developed for the speciation of Cr in an aquatic environment using dispersive solid phase extraction of Cr(III) by Cr(III)-IIM prior to selective Cr(VI) measurement by ETAAS in the supernatants. The detection limits and reproducibility achieved for the Cr speciation analysis fulfill the requirements for their monitoring in waters under the demand of the Water Framework Directive.