Abstract Measurement of polymers in water and wastewater is a major challenge, and there is currently no quick and simple method that can achieve this. This study presents a method for quantification of polyacrylamide concentration using an in-line UV–vis spectrophotometer that can generate real-time data. Absorbance spectra of seven polymers were established in the wavelength range of 191.5–750 nm and the highest absorbance was recorded at 191.5 nm for all polymers. UV absorbance of the polymers gradually decreased between 191.5–240 nm and a strong linear relationship (R2 > 0.97) between concentration and absorbance held at 191.5, 200 and 210 nm. Detection limits were established in distilled water and in four other samples collected from water and wastewater treatment plants. Polymer chemistry and presence of organic and inorganic impurities impacted the detection limits. In distilled water samples, the lowest detection limit measured was 0.05 mg/L. In samples collected from water and wastewater treatment plants, detection limits varied between 0.07 and 1.35 mg/L depending on the polymer type and water quality. Polymers are known to be toxic to aquatic ecosystems and they are suspected carcinogens, and the presented method could be useful in monitoring the polymer concentrations and minimizing its excessive use.