Post mortem concentrations of endogenous gamma hydroxybutyric acid (GHB) and in vitro formation in stored blood and urine samples

Gamma-hydroxybutyrate (GHB) is a central nervous system depressant, primarily used as a recreational drug of abuse with numerous names. It has also been involved in various instances of drug-facilitated sexual assault due to its potential incapacitating effects. The first aim of this paper is to measure the post-mortem concentration of endogenous GHB in whole blood and urine samples of 30 GHB free-users, who have been divided according to the post-mortem interval (PMI) in three groups (first group: 24-36h; second group: 37-72h; third group: 73-192h), trying to evaluate the role of PMI in affecting post mortem levels. Second, the Authors have evaluated the new formation of GHB in vitro in blood and urine samples of the three groups, which have been stored at -20°C, 4°C and 20°C over a period of one month. The concentrations were measured by GC-MS after liquid-liquid extraction according to the method validated and published by Elliot (For. Sci. Int., 2003). For urine samples, GHB concentrations were creatinine-normalized. In the first group the GHB mean concentration measured after autopsy was: 2.14mg/L (range 0.54-3.21mg/L) in blood and 3.90mg/g (range 0.60-4.81mg/g) in urine; in the second group it was: 5.13mg/L (range 1.11-9.60mg/L) in blood and 3.93mg/g (range 0.91-7.25mg/g) in urine; in the third group it was: 11.8mg/L (range 3.95-24.12mg/L) in blood and 9.83mg/g (range 3.67-21.90mg/g) in urine. The results obtained in blood and urine samples showed a statistically significant difference among groups (p<0.001) in the first analysis performed immediately after autopsy. Throughout the period of investigation up to 4 weeks, the comparison of storage temperatures within each group showed in blood and urine samples a mean difference at 20°C compared to -20°C not statistically significant at the 10% level. These findings allow us to affirm that the PMI strongly affects the post mortem production of GHB in blood and urine samples. Regarding the new formation of GHB in vitro both in blood and urine samples of the three groups, which have been stored at -20°C, 4°C and 20°C over a period of one month, although there was no significant increases of GHB levels throughout the period of investigation, the lowest increases were found both in blood and urine at -20°C, therefore we recommend the latter as optimal storage temperature.