A new specific phosphine modified polymer microsphere (PPMs) was designed and used as the core adsorbent to comprehensively enrich both inorganic and organic arsenic species in environmental waters by on line solid-phase extraction (SPE). Avoiding any redox reagent, all the four arsenic species have been quantitatively retained on the home made mini-column with large number of positively charged adsorption groups, and then eluted rapidly (within seconds) with a mixed solution of ammonium nitrate and ammonium dihydrogen phosphate. The trace separation and determination of As(III), DMA, MMA and As(V) species have been simultaneously achieved by high performance liquid chromatography hyphenated to inductively coupled plasma spectrometry (HPLC–ICP-MS) technique. This work has developed a versatile tri-n-butylphosphine functionalized polymer microsphere for an efficient and reliable on-line of simultaneous preconcentration and detection of inorganic and organic arsenic speciation. Under the optimized experimental conditions, the enrichment factors obtained for As(III) and DMA with 25mL sample solution were 28, while for MMA and As(V) reached 30. The low detection limits of 1.2ngL−1, 0.96ngL−1, 0.82ngL−1 and 0.91ngL−1, with the relative standard deviations (RSDs) of 3.9%, 5.6%, 3.2% and 4.5% were obtained for As(III), DMA, MMA and As(V), respectively. The developed method was validated by analyzing Certified Reference Materials GSBZ 07-3171-2014, promising for routine monitoring of arsenic species in lake water, river water and seawater samples.