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Purpose Limited options are available to treat critically ill patients with acute liver failure (ALF) and acute-on-chronic liver failure (AoCLF), therefore we set up an in vitro study in order to test the bilirubin adsorption capacity of the Lixelle S-35 cartridge by direct hemoperfusion (DHP).Methods Mock DHP was performed for 120 min using hyperbilirubinic human plasma and blood obtained from a plasmapheresis and exchange transfusion, respectively. The total bilirubin (TBIL) and direct bilirubin (DBIL) baseline concentrations were 17.57 ± 0.53, 12.57 ± 0.23 mg/dl for plasma and 23.10 ± 0.47, 15.37 ± 0.24 mg/dl for blood. Plasma and blood were separately circulated in a closed circuit simulating DHP using an adsorption column (Lixelle S-35) at flow rate of 100 ml/min. TBIL and DBIL levels were measured at 10, 30, 60, and 120 min from arterial and venous ports and assessed with the Jendrassik-Grof method. All tests were performed in triplicate.Results The total removal subsequent to DHP (120 min) was seen as TBIL in plasma 55.60%, TBIL in blood 62.16%, DBIL plasma 58.87%, DBIL in blood 64.41%, respectively. The estimated mass adsorption of TBIL in plasma 958.20 ± 5.72 mg, TBIL in blood 1233.60 ± 10.22 mg, DBIL in plasma 680.70 ± 10.68, DBIL in blood 818.10 ± 4.68, respectively.Conclusions The bilirubin adsorption rates after DHP were very promising for both hyperbilirubinic plasma and blood. Although further in vitro investigations are required, including comparisons with other techniques, these findings have shown that the Lexille S-35 should represent an option for the management of hyperbilirubinemia in ALF or AoCLF.