Download Analytical data in support of the liver and peripheral blood

Original Research
European Journal of Forensic Sciences
www.ejfs.co.uk
DOI: 10.5455/ejfs.224357
Analytical data in support of the liver
and peripheral blood concentration
ratio as a marker of postmortem
redistribution
Iain M McIntyre
ABSTRACT
Forensic Toxicology
Laboratory Manager,
County of San Diego,
Department of the
Medical Examiner
Address for correspondence:
County of San Diego,
Department of the Medical
Examiner, 5570 Overland
Ave., Suite 101, San Diego,
CA 92123, U.S.A. Email: Iain.
[email protected]
Received: April 06, 2016
Accepted: May 18, 2016
Published: June 28, 2016
Objective: Postmortem redistribution (PMR) refers to the changes that can occur in drug concentrations
after death. Consequently, postmortem blood concentrations may not always reflect the antemortem drug
levels. A recent literature review has postulated a model describing drugs with a liver (L) to peripheral blood
(P) concentration ratio less than 5 L/kg as being prone to little or no PMR, while drugs with an L/P ratio greater
than 20-30 L/kg exhibit propensity for substantial PMR. Antidepressants including tricyclic antidepressants and
some selective serotonin re-uptake inhibitors, for example, were obviously distinguished from drugs confirmed
to be free from, or demonstrate little PMR. Methods: This current paper presents analytical L/P data from 867
postmortem cases yielding a ranking of 44 different drugs’ propensity for (and degree of) PMR. Results: The
results are recorded on a continuum of propensity for PMR ranging from the lowest to the greatest. Drugs
previously suspected to exhibit substantial PMR were noticeably differentiated from those thought not to
demonstrate such postmortem changes–the higher the L/P, the greater the propensity for PMR. Moreover, drugs
with intermediate propensity for PMR were discriminated from others with greater (or lesser) predisposition.
Conclusions: The resulting classification of drugs’ propensity for–and anticipated degree of–PMR, should
assist with a rational interpretation of postmortem drug concentrations for forensic experts.
KEY WORDS: Forensic sciences, forensic toxicology, autopsy, liver, peripheral blood, postmortem redistribution
INTRODUCTION
A potentially significant issue complicating interpretation of
postmortem drug concentrations results from the phenomenon
referred to as postmortem redistribution (PMR). Postmortem
drug concentrations in blood may not always reflect antemortem
drug concentrations due to the movement of the drugs after
death [1]. Accordingly, authors have argued a cautious approach
in interpreting postmortem blood concentrations [2]. The
mechanisms involved in PMR are both complicated and poorly
understood. However, postmortem drug concentrations in blood
have been thought to follow some commonly accepted trends
that may aid with interpretation. In general, the characteristics
of the drug itself have been used to estimate if a drug is subject
to PMR. Substantial changes in blood drug concentrations have
been predicted for basic, lipophilic drugs with a high volume of
distribution [3]. When PMR occurs, blood specimens drawn
from the central body cavity and heart generally exhibit higher
drug concentrations postmortem than specimens drawn from
peripheral areas, most commonly the femoral region. Diffusion
of drugs from organ tissues into the blood may explain the
observed phenomenon [1,4].
178
The collection, analysis, and comparison of antemortem blood
specimens would obviously be appropriate in assisting with the
interpretation of postmortem blood drug concentrations, but
relevant specimens are only rarely available. In a set of case
studies of six drugs, concentrations in the postmortem femoral
blood specimens exceeded the antemortem concentrations
in five of the drugs studied, suggesting that even peripheral
blood exhibited some redistribution [5]. The study reporting
principally cases of overdose did not, however, convey the
postmortem interval been death and autopsy. This interval (or
postmortem delay) has been suggested to influence PMR [6].
The potential for redistribution of other drugs in postmortem
peripheral blood has also been documented more recently [7].
In an early attempt to assess and demonstrate PMR, postmortem
blood specimens were collected from two areas of the body at
autopsy: A peripheral area and central area (often the heart/
cardiac) so that a comparison could be made. This approach
appeared to provide a partial answer to the difficulties associated
with the interpretation of postmortem drug concentrations.
Prouty and Anderson [6] first detailed information about blood
drug concentrations attained from the different sites for over
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McIntyre: L/P ratio as a marker of postmortem redistribution
50 drugs. Then, Dalpe-Scott et al. [8] presented a tabular list
of the drug concentrations from both cardiac and peripheral
blood samples expressed as a ratio of cardiac to peripheral
blood (C/P) for over 100 drugs. The C/P ratio became the
accepted benchmark with the accepted guideline that ratios
>1.0 were associated with redistribution, and high ratios
indicated potential for substantial PMR [8]. This guideline was
repeated in a review a few years later where many C/P ratios
were republished [9].
Despite some apparent value, limitations of the C/P model,
however, have been documented. While drug properties such
as the volume of distribution, protein binding, and pKa were
thought to contribute to PMR, a relationship between C/P
and drug properties has not been established [10]. In addition,
there has been little agreement as to what ratio actually defines
a compound as one that is prone to substantial or minimal
PMR [11]. Furthermore, reports of a C/P ratio >1.0 have
been published for salicylate [5] and carisoprodol [11] which
are not prone to redistribution. Arteriovenous differences,
anatomic variability within individuals, and statistical chance
may result in a C/P ratio >1.0 in drugs that do not redistribute.
In addition, resuscitation attempts may result in a C/P ratio
<1.0 [12]. Inaccurate ratios may also be obtained as an artifact
of sampling on depletion of the cardiac blood volume by the
collection of blood from connected blood vessels, or in cases of
an acute overdose where the drug has not undergone complete
absorption and/or distribution. Consequently, the traditional
C/P ratios can be inconclusive and even misleading with respect
to the interpretation of PMR [13].
Alternately, the liver to peripheral blood (L/P) ratio has been
proposed as a marker for PMR. Ratios <5 L/kg were advocated
to indicate little to no propensity toward PMR, and ratios
exceeding 20-30 L/kg indicative of a propensity for substantial
PMR [11]. A number of reports and a literature review
elaborating on, and supporting, this model have now been
published by this author [13-18].
The current paper describing analytical postmortem data
from 867 cases is presented in support of the L/P ratio theory.
Results were then applied to create a ranking 44 different drug’s
propensity for (and degree of) PMR.
METHODS
Autopsy and Postmortem Specimen Collection
In all cases, a full autopsy was conducted at the Medical
Examiner’s Department, San Diego County. Autopsies were
performed within 48 h of the recorded time of death. In cases
in which the decedent was found dead, the time of death was
recorded as the time found. This was not the exact time of death,
which may have occurred several hours earlier. No attempt was
made to establish the exact time of death for these cases. No
cases showed obvious signs of decomposition; cases were not
included if decomposition was noted by the medical examiner
investigator or observed during the autopsy procedure.
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Autopsies on the 867 cases examined in this investigation were
performed over several years (2007-2015) by a staff of 10 board
certified forensic pathologists. Although individual pathologists
had slightly different approaches to details within the autopsy
procedure, the general technique and specimen collection were
consistent. The autopsy was started with a usual Y incision to
allow viewing of the chest and abdominal organs. Following an
initial inspection, each organ was removed for a more detailed
examination. During the examination of the liver, sections of
the right lobe of the liver (approximately 100 g) were collected
and stored in an opaque plastic 4 ounce container without
preservative. There was minimal chance of contamination from
gastric contents or other sources with this section and collection
technique. Upon removal of the intestines, the common iliac
vein was visualized and punctured or cut and the peripheral
blood specimens (generally 10-20 ml) collected and stored
in standard glass tubes containing sodium fluoride (100 mg)
and potassium oxalate (20 mg). Using this technique (visual
identification of the iliac vein in the pelvis), the pathologist
was able to ensure collection of blood returning from the leg.
However, as the upper section of iliac vein was not usually
clamped, there was potential for a small volume of blood to
accumulate from more proximal regions in some cases. Despite
this relatively minor exception, there was minimal opportunity
for substantial contamination of the blood, especially from other
sources. All samples were stored at 4°C until analyzed.
Toxicological Analyses
In general, the toxicological screening regimen consisted of the
analysis of postmortem blood for alcohol and simple volatile
compounds (GC-FID headspace), drugs of abuse by ELISA (at
a minimum: Cocaine metabolite, opiates, methamphetamine,
benzodiazepines, cannabinoids, fentanyl) (Immunalysis Inc.,
CA), and an alkaline drug screen by gas chromatography-mass
spectrometry (GC-MS) following solid phase extraction.
An acid/neutral drug screen with high-performance liquid
chromatographic (HPLC)-photodiode array detection following
specimen precipitation with acetonitrile was performed as
required (often dictated by medications found at the scene).
Positive results were confirmed and quantified by subsequent
and specific techniques. Most of the drugs studied were
quantified by an alkaline liquid-liquid extraction followed
by GC-nitrogen-phosphorus detector detection which has
been previously described [17]. Drugs determined by other
procedures were carisoprodol and meprobamate by GCMS [11]; amphetamine and methamphetamine determined
by GC-MS [18]; fentanyl by GC-MS [19]; lamotrigine and
quetiapine, which were determined by the HPLC method [20];
acetaminophen by HPLC [21].
RESULTS AND DISCUSSION
Table 1 presents the mean, standard deviation, and median L/P
ratio data for 867 cases where 44 drugs were examined. The
L/P values represent data determined from autopsied cases at
the Department of the Medical Examiner, San Diego County.
Some drugs investigated clearly exhibited a significant L/P
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McIntyre: L/P ratio as a marker of postmortem redistribution
ratio variability. This was particularly evident for a number of
drugs previously considered to display substantial potential
for PMR. Amitriptyline, for example, revealed a mean value
of 30.4 L/kg with a standard deviation of 39.0 (for 57 cases).
This substantial variability exemplifies the interpretative
complexity of postmortem forensic toxicology. There are
several potential explanations including individual variability,
recent compared to long-term drug use, and possibly overdose
cases – with potentially an inadequate time for complete
distribution of the drug before sudden death. This concept
has been described previously [8]. Consequently, the median
value of 16.4 L/kg was projected to be a more representative
value of the intrinsic L/P ratio for both this drug (amitriptyline)
and most of the other drugs as well (the median is the most
resistant statistic and having a breakdown point of 50%: So long
as no more than half the data are contaminated, the median
Table 1: L/P blood ratio data: Alphabetical listing for 44 drugs
Drug
Acetaminophen
Amiodarone
Amitriptyline
Amlodipine
Amphetamine
Bupropion
Carisoprodol
Chlorpheniramine
Citalopram
Clomipramine
Clozapine
Cyclobenzaprine
Desipramine
Dextromethorphan
Diltiazem
Diphenhydramine
Doxepin
Doxylamine
Fentanyl
Fluoxetine
Gabapentin
Guaifenesin
Hydrocodone
Hydroxyzine
Imipramine
Lamotrigine
Meprobamate
Methadone
Methamphetamine
Metoprolol
Mirtazapine
Naproxen
Olanzapine
Paroxetine
Promethazine
Propoxyphene
Propranolol
Quetiapine
Sertraline
Tramadol
Trazodone
Venlafaxine
Zolpidem
Ethanol
L/P: Liver/peripheral
180
Number
of cases
L/P (L/kg)
(mean)
SD
L/P (L/kg)
(median)
15
5
57
9
15
11
11
3
37
5
12
13
6
4
8
54
20
3
16
48
28
5
38
10
6
3
8
94
18
6
5
20
20
19
9
35
6
65
9
36
19
42
14
1.2
61.5
30.4
45.5
8.0
1.3
2.8
9.5
10.1
45.4
7.4
27.5
43.8
8.7
23.8
10.0
22.1
2.8
6.9
42.2
0.67
1.4
3.4
13.8
30.8
8.0
1.2
5.5
5.7
4.0
14.3
1.1
14.0
33.3
14.0
14.2
13.5
18.0
97.0
2.5
3.5
4.3
6.2
0.91
0.41
42.4
39.0
34.6
4.0
0.6
1.5
1.9
7.9
27.9
2.4
27.4
14.3
5.5
24.2
8.9
14.0
0.4
4.5
30.1
0.26
1.3
1.7
6.2
15.3
2.5
0.7
3.0
2.3
1.7
10.4
0.65
9.6
21.2
16.1
12.1
10.9
21.6
40.0
1.4
2.0
3.3
9.6
1.1
41.9
16.4
27.8
7.0
1.0
2.0
9.5
7.5
61.0
6.7
19.6
44.6
8.6
17.2
6.7
19.4
2.9
5.9
30.0
0.65
0.90
3.0
12.3
28.8
8.5
0.90
4.8
6.2
3.5
12.0
1.0
12.0
29.2
9.1
9.0
10.9
11.2
76.0
2.3
2.8
3.3
2.4
will not give an arbitrarily large result. The median can be used
generally as a measure when a distribution is skewed, or when
one requires reduced importance to be attached to outliers.
Despite the fact many of the drugs did not demonstrate such
large variability; the median value was applied to evaluate all
drugs for purposes of consistency. Data from these 44 drugs
corroborated earlier published reports supporting a relationship
between the L/P ratio and susceptibility for drugs to exhibit
potential for PMR [11,13,22]. Drugs known to be susceptible
to significant PMR, such as the tricyclic antidepressants
and some selective serotonin re-uptake inhibitors, were
clearly distinguished from ethanol, gabapentin, carisoprodol,
zolpidem, and other compounds not thought to exhibit
substantial PMR. Several of these data have been independently
published: Carisoprodol/meprobamate [11], sertraline [14],
fentanyl [16], hydroxyzine [17], and methamphetamine/
amphetamine [18,22]. The data for ethanol were taken from
the previous publication [23].
Nevertheless, the resulting L/P ratio (and consequent
interpretation of potential for PMR) may not be upheld in
all casework; especially, on occasion of substantially longer
postmortem delay (>48 h), particularly for those compounds
displaying extensive PMR potential. In such cases, and
certainly in the event of decomposition, the possibility
of considerable physical and chemical changes may cause
additional and inconsistent drug redistribution, thereby
increasing interpretative complexity. Furthermore, in cases
of overdose, where the incomplete distribution of a drug
can result in variable concentrations throughout the body’s
organs and tissues, the current approach may not always be
suitable. Such cases, therefore, should be interpreted even
more cautiously.
Collection procedures employed for postmortem blood and liver
tissue samplings are also important matters for consideration.
Consistency in the collection technique of postmortem blood
is critical. Concentrations of many drugs have been shown
to have substantial site-dependence [24]. Concentrations
attained from drug analyses performed on heart (or central)
blood, pericardial blood, chest blood, and perhaps subclavian
blood, where drug concentrations can be erroneously elevated,
may not be applicable to this particular model. Likewise, the
site/location of collection of the liver sample is important.
Liver concentrations may differ if collected near the lower
left quadrant where contamination from gastric content can
occur [25]. Nonetheless, if sample collections were consistent
and contamination prevented (or at least marginalized)
assumptions and calculations analogs to those made in this
paper can be practicable.
The results were then recorded on a continuum of propensity for
PMR ranging from the lowest to the greatest [Table 2]. Again,
as recognized earlier, drugs previously suspected to exhibit
substantial PMR were noticeably differentiated from those
thought not to demonstrate such postmortem changes – The
higher the L/P, the greater the propensity for PMR. Moreover,
drugs with intermediate propensity for PMR were discriminated
from others with greater (or lesser) predisposition. Citalopram
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McIntyre: L/P ratio as a marker of postmortem redistribution
Table 2: L/P ratio: Listed in increasing propensity for PMR
Drug
Gabapentin
Guaifenesin
Meprobamate
Ethanol
Naproxen
Bupropion
Acetaminophen
Carisoprodol
Tramadol
Zolpidem
Trazodone
Doxylamine
Hydrocodone
Venlafaxine
Metoprolol
Methadone
Fentanyl
Methamphetamine
Clozapine
Diphenhydramine
Amphetamine
Citalopram
Lamotrigine
Dextromethorphan
Propoxyphene
Promethazine
Chlorpheniramine
Propranolol
Quetiapine
Mirtazapine
Olanzapine
Hydroxyzine
Amitriptyline
Diltiazem
Doxepin
Cyclobenzaprine
Amlodipine
Imipramine
Paroxetine
Fluoxetine
Amiodarone
Desipramine
Clomipramine
Sertraline
L/P (L/kg)
0.65
0.90
0.90
0.91
1.0
1.0
1.1
2.0
2.3
2.4
2.8
2.9
3.0
3.3
3.5
4.8
5.9
6.2
6.7
6.7
7.0
7.5
8.5
8.6
9.0
9.1
9.5
10.9
11.2
12.0
12.0
12.3
16.4
17.2
19.4
19.6
27.8
28.8
29.2
30.0
41.9
44.6
61.0
76.0
PMR: Postmortem redistribution, L/P: Liver/peripheral
(L/P = 7.5 L/kg), for example, can be identified as an example
of a drug which possesses propensity for PMR somewhat greater
than that of venlafaxine (L/P = 3.3 mg/L), but substantially
less than that of the traditional tricyclic antidepressants and
some of the other SSRIs such as fluoxetine (L/P = 30.0).
Furthermore, this ranking of drugs’ potential for PMR addressed
the apparent incongruity observed in the interpretation of some
postmortem drug concentrations. Sertraline, for example, a
compound with a large volume of distribution (20-50 L/kg)
has been reported with a relatively insignificant C/P ratio of
<1.3 [14], and consequently, inconsistent with a drug exhibiting
a substantial degree of PMR. However, postmortem blood
concentrations (considered therapeutic to 1.0 or 1.5 mg/L) are
commonly reported to be greater than expected in consideration
of clinical investigations - studies report plasma sertraline
concentrations averaging up to 0.20 mg/L, even with chronic
high dosing of 300 mg daily [26]. This new analysis (displaying
Eur J Forensic Sci ● 2016 ● Vol 3 ● Issue 4
L/P = 76.0 L/kg) confirmed the substantial potential for
sertraline PMR and presented a more characteristic explanation
of observed postmortem blood concentrations. This conclusion
is also more consistent with expectation resulting from
consideration of the particularly large volume of distribution.
In addition, the list (showing drug ranking of potential for
PMR) resolved interpretive confusion concerning other drugs
such as metoprolol. Reports of inconsistent C/P ratio data have
made assessment of potential PMR difficult – One report listed
the C/P ratio data as l.0 or less [8] and another as 3.8 [6]. The
current model confirmed the minimal potential for metoprolol
PMR, with an L/P = 3.5 L/kg.
One of the principal goals of these investigations was to
attempt to provide a cataloging of drugs’ propensity for and,
subsequently, their potential extent of PMR. Until now, most
efforts in interpretation have simply described PMR by an
aphorism ranging from “the drug has not been found to exhibit
PMR” to “the drug is subject to PMR.” Such descriptions
have never been particularly useful in the interpretation of
postmortem drug concentrations, especially in relation to
deducing what the drug concentration may have been at the
time of death. Development of the L/P ratio model and the
resulting estimation of propensity for PMR will hopefully
provide a more definitive, and ultimately, a more authoritative
numerical interpretation of PMR for many drugs and poisons.
Although the approach described in this paper was capable
of providing an accurate explanation and interpretation for
data collected at this Medical Examiner’s Office, it can be
anticipated (or even expected) that this will not be possible in
all casework. As discussed earlier, the site of specimen collection
and collection technique, excessive postmortem delay, cases
of overdose, or contamination may cause susceptibility to
significant changes in drug concentrations, producing additional
interpretation difficulties, and rendering the current model
unsuitable, or ineffective. Nevertheless, a technique to estimate
potential for PMR, together with a reference list for 44 of some
of the most commonly encountered drugs in postmortem
forensic toxicology, is now attainable. The resulting classification
of drugs’ propensity for – and anticipated degree of PMR,
should assist with a rational interpretation of postmortem drug
concentrations for forensic experts.
ACKNOWLEDGMENTS
The author would like to thank the San Diego County Chief
Medical Examiner, Dr. Glenn Wagner, for making available
case details described in this manuscript and Christina Meyer
Escott, for collating much of the drug data.
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Source of Support: Nil, Conflict of Interest: None declared.
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