Ketone Bodies

In many forensic cases alcohol abusers have been found dead and the cause of death cannot be ascertained. In order to examine the possible role of ketoacidosis as the cause of death the concentrations of ketone bodies (acetone, acetoacetate, d-^-hy-droxybutyrate) have to be determined in postmortem blood specimens. The phenomenon of ketoacidosis is often seen as typical in periods of abstinence with low intake of food. It is due to the accumulation of d-^-hydroxybutyrate and acetoacetic acid. The accumulation is probably the result of various factors such as volume depletion and starvation, which have a lipolytic effect.

A routine procedure is a coupled enzymatic head-space GC method (Figure 3). This procedure is based

Figure 2 Total ion chromatograms ((A) selected ion monitoring and (B) full scan mode) of a standard solution of 28 substances relevant in congener analysis in concentrations of 2 mg L~1 (methanol 10 mg L~1, acetaldehyde 0.5 mg L~1). 1, Acetaldehyde; 2, methanol; 3, ethanol; 4, propionaldehyde; 5, acetone; 6, propanol-2; 7, methyl acetate; 8, t-butanol (internal standard); 9, i-butyraldehyde; 10, propanol-1; 11, n-butyraldehyde; 12, methyl ethyl ketone; 13, ethyl acetate; 14, butanol-2; 15, i-butanol; 16, i-valeraldehyde; 17, 2-methylbutyraldehyde; 18, butanol-1; 19, n-valeraldehyde;20,1,1-diethoxyethane;21, 3-hydroxybutanone-2;22, 3-methylbutanol-1; 23, 2-methylbutanol-1; 24, i-butyl acetate; 25, pentanol-1; 26, butyl acetate; 27, ethyl lactate; 28, hexanol-1. GC parameter: HP 5890 II GC with HP MSD 5972, equipped with a DB 624 column (60 m x0.32 mm, df = 1.8 ^m); helium flow 1 mL min~1; injector 150°C; detector 200°C; oven initially 30°C for 8 min, 3°C min~1 up to 180°C. (Reproduced with permission from Roemhild W (1998) Congener analysis by means of 'headspace'- GC/MS. Blutalkohol35: 10.)

Figure 2 Total ion chromatograms ((A) selected ion monitoring and (B) full scan mode) of a standard solution of 28 substances relevant in congener analysis in concentrations of 2 mg L~1 (methanol 10 mg L~1, acetaldehyde 0.5 mg L~1). 1, Acetaldehyde; 2, methanol; 3, ethanol; 4, propionaldehyde; 5, acetone; 6, propanol-2; 7, methyl acetate; 8, t-butanol (internal standard); 9, i-butyraldehyde; 10, propanol-1; 11, n-butyraldehyde; 12, methyl ethyl ketone; 13, ethyl acetate; 14, butanol-2; 15, i-butanol; 16, i-valeraldehyde; 17, 2-methylbutyraldehyde; 18, butanol-1; 19, n-valeraldehyde;20,1,1-diethoxyethane;21, 3-hydroxybutanone-2;22, 3-methylbutanol-1; 23, 2-methylbutanol-1; 24, i-butyl acetate; 25, pentanol-1; 26, butyl acetate; 27, ethyl lactate; 28, hexanol-1. GC parameter: HP 5890 II GC with HP MSD 5972, equipped with a DB 624 column (60 m x0.32 mm, df = 1.8 ^m); helium flow 1 mL min~1; injector 150°C; detector 200°C; oven initially 30°C for 8 min, 3°C min~1 up to 180°C. (Reproduced with permission from Roemhild W (1998) Congener analysis by means of 'headspace'- GC/MS. Blutalkohol35: 10.)

on enzymatic dehydrogenation of d-^-hydroxy-butyrate into acetoacetate and subsequent decar-boxylation of this compound into acetone. Three portions are taken from each sample. One portion is heated to 60°C in a headspace sampler, which gives the free acetone. Acetoacetate is converted into

Figure 3 Schematic presentation of a standard procedure for determination of ketone bodies in blood specimens. Three portions are taken from each sample to determine free acetone and the sums of acetone # acetoacetate and acetone # acetoacetate # D-^-hydroxybutyrate.

acetone by decarboxylation at 100°C in the second portion. This part gives the combined amount of acetone and acetoacetate. In the third portion, d-^-hydroxybutyrate is first enzymatically dehydrogenized into acetoacetate by d-^-hydroxybutyrate dehydrogenase and then decarboxylated into acetone. Quantification of acetone then yields the molar equivalent of the total ketone bodies. Omission of the enzymatic stage of the analysis allows quantification of the molar equivalent of acetone and acetoacetate present, and the substraction of this value from total ketone quantitation allows calculation of the d-^-hydroxybutyrate concentration.

The reported ketone body concentrations vary a lot. It was held that if the ketone body concentration of the blood exceeds 531 |imol L"1 and if there is no other plausible cause of death in a group of alcohol abusers, the term ketoalcoholic death should be used. In another study it was pointed out that very hight levels, above 10mmolL_1, are indicative of profound alcoholic ketoacidosis.

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