A comprehensive method for simultaneously detecting species of arsenic and selenium including arsenite (As(III)), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), arsenate (As(V)), selenocystine (SeCys), selenomethionine (SeMet) and selenate (Se(IV)) was developed with high performance liquid chromatography-hydride generation-double channel atomic fluorescence spectrometry (HPLC-HG-AFS). An anion-exchange column (PRP-X100) with eluent of 10 mmol/L NH4H2PO4 containing 2.5% (v/v) methanol was employed to separate these species within 12 min. The detection limits of As(III), DMA, MMA, As(V), SeCys, SeMet and Se(IV) were 1, 3, 2, 3, 4, 18 and 3 microg/L (200 microL of injection), respectively. The relative standard deviations in five independent determinations varied from 1.9% to 6.1% for arsenic and selenium species at the concentration levels of 100 and 300 microg/L. The proposed method was applied to analyze the selenium yeast tablet and human urine samples. The recoveries from spiked selenium yeast tablet and urine samples ranged from 88% to 105% and from 83% to 108%, respectively. The results showed that this method can be used for determining arsenic and selenium species in urinary metabolites and drug samples in daily analysis conveniently.