The speciation of selenium in selenium-enriched kelp (Laminaria japonica)  was studied by using chromatographic separation coupled with mass spectrometry detection (HPLC-ICP-MS). The kelp was cultivated hydroponically for 60 hours in seawater containing selenite (200 mg/dm³ Na₂SeO₃) and N and P (150 mg/ dm³ NaNO₃ + 25 mg/ dm³ NaH₂PO₄) nutrient substance. Biotransformation of selenium in the kelp was studied by measuring the concentration of selenium in biological samples. The Selenium-enriched kelp samples were divided into upper, middle and lower parts, and the Selenium values in different parts (dry weight) were 2.26mg/g, 2.29mg/g and 1.89mg/g respectively. The different parts were extracted with water and methanol (90:10), then shook automatically for 24h, selenium speciation was carried out by high performance liquid chromatography coupled with inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The result was that different organic Se was transformed from inorganic Se in different parts of kelp samples. The species of Se in upper, middle and lower parts in kelp samples were found as three, five and six respectively. Some organic Se was unknown because lack of standards, and further study was needed to identify their structure by high performance liquid chromatography coupled with electro-spray ionization time-of-flight mass spectrometry (HPLC-ESI-TOF-MS).