Application of the Alkaline-Digestion-HPLC Method to the Rapid Determination of Polyhydroxyalkanoate in Activated Sludge

Abstract

A new method for analyzing polyhydroxyalkanoate (PHA) in activated sludge based on alkaline digestion followed by high-performance liquid chromatography (HPLC) was evaluated. The effects of the concentration of NaOH, reaction time, and reaction temperature were examined using activated-sludge samples containing PHA and commercially purchased PHA. To determine PHA concentrations, 0.5 mL 2 N NaOH was added to 1 mL activated-sludge mixed liquor, and the resulting mixture was heated at 105°C in a drying oven for 1 h to convert 3-hydroxybutyrate units into 2-butenoate and 3-hydroxyvalerate units into 2-pentenoate. The mixture was then acidified with 0.5 mL of 2 N H₂SO₄, and the solids were removed and subsequently analyzed by HPLC. The present method gave results consistent with those obtained by the conventional method of methanolysis followed by gas chromatography. The present method can facilitate investigations of PHA in activated sludge by reducing the time and labor required for analysis and by eliminating the use of organic solvents.