Published: Journal of Analytical Toxicology, ISSN 0146-4760, Volume 24, Number 5, July/August, pp.383-384

Here is where the title stuff goes

Elevated GHB in Citrate-Buffered Blood

To the Editor:

With the recent decision to make gamma-hydroxybutyrate (GHB) a federally controlled Schedule I substance (1), it is anticipated that more laboratories will include GHB screens in their routine analytical schemes. Although it is important to recognize that recreational abuse of GHB poses a serious problem, GHB has been implicated in a number of drug-facilitated sexual assault cases as well.

Recently, our laboratory discovered an important finding in the analysis of whole blood for the presence of GHB. We were asked to verify the findings of another laboratory that found no GHB in a blood sample of a sexual assault victim. Our results identified GHB at a concentration of 14 µg/mL. Conversation with the other laboratory revealed their analysis had been on a whole-blood specimen stored in a tube containing the anticoagulant EDTA (lavender top). The blood we analyzed was stored in a yellow-top tube containing a different anticoagulant-citrate buffer (trisodium citrate, citric acid, and dextrose). To resolve the differences, the other laboratory agreed to reanalyze both specimens.

Upon completion of the reanalysis, we were informed that the blood specimen from the EDTA tube was still negative for GHB, but GHB was detected in the citrate-buffered blood. After confirming that both blood samples had originated from the same victim at approximately the same time, we agreed to do further testing that may explain the differences in the GHB findings between the two blood specimens.

During the course of this investigation, we received another case of suspected drug-facilitated sexual assault. As in the above case, blood samples from the victim were collected in both lavender-top and yellow-top specimen tubes. Analyses of these specimens for GHB resulted in similar findings-GHB was not detected in the blood from the EDTA tube, yet was identified in the blood from the citrate-containing tube (31 µg/mL).

Given that the specimens from both cases had been stored for months prior to the GHB analyses, it was felt that GHB may either be artificially produced in citrate-buffered blood specimens or destroyed in EDTA blood specimens during extended storage.

In an effort to explore the idea that GHB may be artificially produced in blood stored in citric acid, we analyzed 10 citrate-buffered whole blood specimens from random sexual assault cases. None of these cases was suspected of being GHB-related assaults. The results of the analyses are shown in Table I. These specimens had been stored at –20°C for periods of 6–36 months before analysis.

These results support the notion that GHB may be artificially elevated in citrate-buffered blood. It is a disturbing finding because these levels may be considered indicative of GHB ingestion by the victim. Although it has been documented that GHB is artificially elevated in postmortem blood specimens (2), it was our belief that this did not occur with antemortem blood.

Several experiments were conducted in our lab to examine GHB levels in various blood tubes. We have not been able to determine the source of the elevated GHB in the citrate-buffered specimens, nor have we been able to produce it in any significant quantities under laboratory-controlled storage conditions. Additionally, we have not uncovered any evidence that GHB loss occurs in blood specimens stored in EDTA tubes. However, given the discrepancies between the citrate-buffered and EDTA-stored specimens, our laboratory has taken the position that we will not report positive GHB results from citrate-buffered whole blood unless a urine specimen from the individual was also collected. “Positive” GHB findings in these blood samples will only be reported when the results of a paired urine specimen support such a finding.

This emphasizes just one of many concerns facing the toxicologist in their investigation into claims of drug-facilitated sexual assault. Many rape evidence collection kits that are in use today contain tubes with citrate buffer. Few contain the preferred grey-top tube (sodium fluoride/potassium oxalate) for blood specimens, and even fewer contain collection cups for urine specimens (4). When these kits are used to collect specimens from victims of drug-facilitated sexual assault, one should be wary of the anticoagulant or preservative used in the specimen tube before reporting GHB-positive results.

Marc A. LeBeau, Madeline A. Montgomery, Rebecca A. Jufer, and Mark L. Miller
FBI Laboratory
Washington, D.C. 20535

References

1. Fed. Regist. 65: 13235–13238 (2000).
2. E. Fieler, D. Coleman, and R. Baselt. Hydroxybutyrate concentrations in pre- and postmortem blood and urine. Clin. Chem. 44: 692 (1998).
3. B. Stevens, D. Coleman, and R.C. Baselt. In vitro stability of endogenous gamma-hydroxybutyrate in postmortem blood. J. Forensic Sci. 44: 231 (1999).
4. M. LeBeau, W. Andollo, W.L. Hearn, R. Baselt, E. Cone, B. Finkle, D. Fraser, A. Jenkins, J. Mayer, A. Negrusz, A. Poklis, H.C. Walls, L. Raymon, M. Robertson, and J. Saady. Recommendations for toxicological investigations of drug-facilitated sexual assaults. J. Forensic Sci. 44: 227–230 (1999).

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