A simple chemical method for the oxidation of beta-hydroxybutyrate; application of the method for the estimation of ketone bodies.

A simple one-step method employing potassium persulphate as an oxidising agent in presence of catalyst, Ag2+, for the oxidation of beta-hydroxybutyrate (beta-OHB) to acetoacetate (AcAc) has been developed and standardized. Under the condition of assay, beta-OHB (0.079-0.395 microM) was quantitatively transformed to AcAc. The reaction linearity was observed from 0.079 to 0.634 microM. Optimum conditions were: pH, 6.2; temp., 40 degrees C; persulphate saturation, 40% and catalyst, 1.82 mM. Under the experimental condition, no reversal of inhibition caused by chloride (22.96 mM) was observed at Ag+ concentrations (9.09 and 18.18 mM), while higher conc. of Ag+ (27.27 mM) caused significant reversal of inhibition (about 60%). The maximum reversal of inhibition was achieved at Ag+ (36.36 mM). The level of ketone bodies, when estimated by the present method, was greatly enhanced during starvation period and about 2- and 12-fold higher level of ketone bodies was observed (compared to control) in rats fasted for 24 and 48 hr respectively.

Spectral evidence for the oxidation of beta-hydroxybutyrate to acetoacetate: diminution of spectra by glucose and chloride.

Chemical oxidation of beta-hydroxybutyrate (beta-OHB) to acetoacetate (AcAc) has been carried out by a simple and new method employing potassium persulphate as an oxidising agent. Under the conditions of assay, beta-OHB (0.079-0.395 microM) was instantaneously oxidised to AcAc and the authenticity of the oxidised product was proved by absorption spectroscopy. A common absorption maxima at about 446 nm was observed in all the spectra recorded for the product (AcAc-complex) obtained after the oxidation of beta-OHB (0.079-0.395 microM) to AcAc followed by coupling with diazotized p-nitroaniline. This absorption maxima was almost equal to that obtained for AcAc-complex using AcAc as reference standard. It implies that AcAc formed by the chemical oxidation of beta-OHB is identically similar to the AcAc used as reference standard for the study. This fact was further strengthened when absorption spectra, recorded either individually or in combination (mixed-type), exhibited a single peak with a common absorption maxima at about 446 nm. Absorption spectra was found to be partially diminished by glucose (1.77 microM) and chloride (17.1 microM), while almost complete diminution of absorption spectra was observed at higher concentration of glucose (8.88 microM) and chloride (51.3 microM).