Abstract Polyethylene glycols (PEGs) are non-ionic, water-soluble synthetic polymers which have been widely used for many applications. Since they are of very low toxicity and are readily excreted in urine, PEGs in the molecular weight range 400–6000 have been used extensively in the study of intestinal physiology in man. A high-performance liquid chromatographic (HPLC) method has been developed for the determination of PEG 600 in human urine, which includes a pre-column derivatisation step. The dibenzoate derivatives of PEG 600 can be quantitatively prepared, and this, coupled with ultraviolet detection at 230 nm, has greatly improved the limit of detection for the determination of PEGs by HPLC. A suitable extraction procedure has also been developed which enabled PEG levels in urine to be monitored with much greater sensitivity than any previously reported method.