Determination of morphine in postmortem rabbit bone marrow and comparison with blood morphine concentrations

doi:10.1016/j.forsciint.2004.12.017

Forensic Science International

Volume 156, Issues 2–3, 27 January 2006, Pages 91-94

https://doi.org/10.1016/j.forsciint.2004.12.017 Get rights and content

Abstract

The aim of this study is to predict how long after time of death a buried body could be analyzed for opiates in soft tissues and to show the accessibility and suitability of bone marrow as a useful toxicological specimen from buried bodies. Morphine solutions were injected in nine albino rabbits. Doses ranged from 0.3 to 1.1 mg/kg with 0.1 mg/kg increments. One hour after the injections, the rabbits were sacrificed. Blood, urine and bone marrow samples were collected for analysis. After the whole bodies were buried, femur bone marrow specimens were collected on the seventh and fourteenth days. CEDIA^® was used to monitor morphine contents of the collected samples. All experimental cases showed that the increase in the given morphine doses correlated with the increase in blood and bone marrow morphine concentrations. High morphine concentrations were detected in urine samples, but there was no correlation between the urine and blood or urine and bone marrow morphine concentrations. Statistically meaningful increases in bone marrow morphine concentrations were found parallel to increase of blood morphine concentrations. Seventh and fourteenth day postmortem morphine concentrations also followed this correlation. Morphine concentrations in bone marrow at 7 and 14 day postmortem decreased consistently when compared with bone marrow morphine concentrations collected immediately after death. We conclude that in sudden death when other specimens are unavailable due to degradation, bone marrow can be a most useful specimen. Further experimental research in this area is required to validate bone marrow as an alternative tissue.

Introduction

Toxic substances and toxicology have been important for many years [1]. The number of intoxications around the world is not known. For example in 1993, over 1.7 million exposures to potential poisons were reported by poison centers participating in the American Association of Poison Control Centers Toxic Exposure Surveillance System (AAPCC-TESS). Intoxication rate is estimated as 0.04% per year in Turkey [2]. Since the majority of drugs and poisons do not produce characteristic pathological lesions, their presence in the body can be demonstrated only by sensitive chemical methods of analysis. Collection and preservation of appropriate specimens is a critical component in all types of forensic examination. Moreover in toxicological analyses, geographical variations in drug use and ability of laboratories must be considered [3].

Traditionally, forensic toxicology and drug analyses have been centered on the use of blood, body fluids and selected organs such as liver, kidney and brain. On the other hand non-parenchymal tissues such as bone and bone marrow may account for 14% of an average body mass. This clearly represents a significant potential drug depot, which is not usually considered in toxicological investigations [4]. Bone marrow, due to high degree of vascularity and lipid matrix, acts as a repository for drugs. Additionally it is encased in bone and reduces the possibility for contamination even after trauma [5], [6].

Frequently forensic toxicologists are faced with situations in which contamination and decomposition make the collection of blood samples impossible for suitable analytical purposes. Because bone marrow is protected by bony tissue and because this tissue is highly vascularized it may be useful as an alternative specimen when contamination of blood specimens is suspected or not available.

Studies have been performed primarily on rabbits, showing that a linear relationship exist between bone marrow and postmortem blood concentration of a given drug for up to 24 h for many substances including tricyclic antidepressants, barbiturates, benzodiazepines and ethyl alcohol. Although putrefaction is delayed in bone marrow, usually this tissue is not considered as an alternative specimen in postmortem toxicology unless others are unavailable [7].

The aim of this study was to predict how long after time of death a buried body could be analyzed for opiates and to show the accessibility and suitability of bone marrow as a valid specimen for toxicological analysis.

Section snippets

Materials and methods

Morphine solutions were injected in nine albino rabbits into the marginal ear vein. Doses ranged from 0.3 to 1.1 mg/kg with 0.1 mg/kg increments. One hour after the injections, the rabbits were sacrificed and their blood, urine and bone marrow samples were collected for morphine content analysis. After the whole bodies with all extremities were buried on the seventh and fourteenth days, the bodies were excavated. Bone marrow specimens were collected and analysed for morphine content.

The analysis

Results and discussion

The morphine dose and the morphine concentrations in blood, urine, and bone marrow in the test rabbits are shown in Table 1.

Postmortem median morphine bone marrow concentrations decreased 21% during the first 7 days and 38% during the second 7 days (Fig. 1). At 14 days the bone marrows were still suitable for toxicological analysis. These experiments showed about 51% loss in bone marrow morphine concentrations at the end of second week. This could be caused by back diffusion from marrow to

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Chromatography was performed on a fused silica capillary column and analytes were determined in the selected-ion-monitoring (SIM) mode. The assay was validated in the range 0.3–1 ng/mg (depending on the drug) to 150 ng/mg, the mean absolute recoveries ranging from 66% to 110%, the matrix effect from 62% to 121% and process efficiency from 61% to 89% depending on the analyte. The intra- and inter-assay accuracy values were always better than 20%. The validated method was then successfully applied to real bone samples from forensic cases in which toxicological analysis for these drugs in blood was positive. Drugs were detected in bone in 12 of the 15 blood positive results. The approximate concentration range was 3–5 ng/g for 6-monoacetylmorphine, 3–7 ng/g for morphine, 14–28 ng/g for methadone and 6 ng/g and 11 ng/g for tramadol and benzoylecgonine.
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^☆: This work was presented at the AAFS 56th Annual Meeting February 16–21, 2004. DALLAS and supported by Research Fund of Istanbul University, project no: UDP222.

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Determination of morphine in postmortem rabbit bone marrow and comparison with blood morphine concentrations☆

Abstract

Introduction

Section snippets

Materials and methods

Results and discussion

Forensic Sci. Int.

Forensic Sci. Int.

Forensic Sci. Int.

Forensic Sci. Int.

Forensic Sci. Int.