Generation of poly-β-hydroxybutyrate from externally provided acetate in rice root

During the investigation of the metabolism of ¹⁴C-acetate or ¹⁴C-succinate in rice seedlings, an unknown organic acid (X) with a high specific radioactivity was detected in 10,000 × g 30 min precipitate-fraction of rice roots. The X was hardly extracted by 0.1 N-H₂SO₄ boiling, but was extracted by 0.5 N-KOH boiling. The X was co-chromatographed with several known organic acids, and the radioactive peak of the X matched β-hydroxybutyric acid (β-hydroxybutyrate). The radioactive X and β-hydroxybutyrate were then heated with concentrated H₂SO₄. The radioactivity and the titration value were completely converted to crotonic acid. Thus, it was concluded that the X was β-hydroxybutyrate, and the original form of this acid was presumed to be poly-β-hydroxybutyrate (PHB). Then rice root incubated with 2-¹⁴C-acetate was extracted with hot-ethanol, ethanol/ether, and hot-chloroform. Approximately 10% of the radioactivity absorbed was detected in the chloroform fraction. The chloroform fraction was co-precipitated with authentic PHB by the addition of acetone/ether, and almost all the radioactivity was co-precipitated with the PHB. The radioactive co-precipitate was then heated with 0.5 N-NaOH, and chromatographed. The radioactivity of β-hydroxybutyrate plus crotonic acid almost matched that of the co-precipitate before alkaline-hydrolysis. Hence the radioactive co-precipitate was confirmed to be PHB. In wheat and radish seedlings, 2-¹⁴C-acetate was also assimilated into PHB. It is concluded that externally provided acetate was rapidly converted to PHB in higher plants.