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Journal of Analytical Toxicology, Vol. 34, September 2010
Uptake and Distribution of the Abused Inhalant
1,1-Difluoroethane in the Rat
Joseph Avella1,2,*, Naveen Kunaparaju1, Sunil Kumar1, Michael Lehrer3, S. William Zito1, and Michael Barletta1
1Department
of Pharmaceutical Sciences, St. John’s University, Jamaica, New York; 2Department of Health Services,
Office of the Chief Medical Examiner, Nassau County, New York; and 3Department of Health Services,
Division of Medical-Legal and Forensic Investigations, Suffolk County, New York
Introduction
Abstract
1,1-Difluoroethane (DFE) is a halogenated hydrocarbon used as a
propellant in products designed for dusting electronic equipment
and air brush painting. When abused, inhaled DFE produces
intoxication and loss of muscular coordination. To investigate DFE
toxicokinetics, groups (n = 3) of Sprague-Dawley rats were exposed
to 30 s of 20 L/min DFE. The experimental model was designed to
mimic exposure during abuse, a protocol which has not been
conducted. Tissue collection (blood, brain, heart, liver, and kidney)
occurred at 0, 10, 20, 30, 45, 60, 120, 240, 480, and 900 s.
Average peak DFE levels were blood 352, brain 519, heart 338,
liver 187, and kidney 364 mg/L or mg/kg. The total percent uptake
of the administered dose was 4.0%. Uptake into individual
compartments was 2.72, 0.38, 0.15, 0.41, and 0.32% for blood,
brain, heart, liver, and kidney, respectively. All animals showed
signs of intoxication within 20 s manifested as lethargy, prostration
and loss of righting reflex. Marked intoxication continued for
about 4 min when DFE averaged 21 mg/L in blood and 17 mg/kg in
brain. Between 4 and 8 min, animals continued to show signs of
sedation as evidenced by reduced aggression and excitement
during handling. No discernable intoxication was evident after 8
min and blood and brain levels had fallen to 10 and 6 mg/L or kg,
respectively. Plots of concentration (log) versus time were
consistent with a two compartment model. Initial distribution was
rapid with average half life (t½) during the α phase of 9 s for blood,
18 s for brain and 27 s in cardiac tissue. During β slope elimination
average t½ was 86 s in blood, 110 s in brain and 168 s in heart.
Late elimination half lives were longer with blood γ = 240 s,
brain γ = 340 s, and heart γ = 231 s. Following acute exposure the
Vd = 0.06 L, β = 0.48 min–1, AUC = 409.8 mg · min L–1, and CL
from blood was 0.03 L min–1. The calculated toxicokinetic data
may underestimate these parameters if DFE is abused chronically
due to continued uptake into lowly perfused tissues with repeated
dosing.
* Author to whom correspondence should be addressed: Joseph Avella, Ph.D., FTS-ABFT,
Chief Toxicologist, Nassau County Medical Examiner’s Office, Bldg. R, 2251 Hempstead
Tpke, East Meadow, NY 11554. Email: [email protected].
Products containing 1,1-difluoroethane (DFE) have been
cited with increasing frequency as sources of inhalant exposure
in reports from government agencies (1) and organizations
involved in inhalant abuse (2). According to a 2004 SAMSHA
survey, 23 million U.S. residents reported that they had abused
inhalants and of these, 10% had done so within the last year
(3). Among inhalant abusers, the abuse of duster products
rose from 12.6 to 25.4% during the period 2002–2005 (4).
When combined these surveys suggest that abuse of duster
products doubled between 2002 and 2005, rising from nearly
2.9 million to 5.8 million US residents.
Inhalation of DFE is reported to produce rapid onset of intoxication and impairment. In addition, abuse of inhalants like
DFE has also been reported to cause sudden death (5–8).
Deaths have therefore occurred due to DFE’s ability to impair
cognitive function, which sometimes leads to accidental death,
and also due to DFE’s inherent cardio-toxicity (9–11). In addition, drivers under the influence of inhaled DFE are regularly
involved in motor vehicle crashes that have resulted not only
in property loss but have also had fatal outcome to passengers,
pedestrians and other motorists.
DFE has both central nervous system (CNS) and cardiac effects that were until recently thought to be adequately described by the Meyer-Overton theory, which said that the lipid
solubility of volatile agents resulted in alterations to cell membrane integrity allowing unregulated flow of ions (12–14).
However that theory of volatile chemistry has been largely replaced with the concept that, like other abused drugs, there are
specific protein targets for volatile agents and that inhalant induced effects on receptor mediated electrochemistry occur at
concentrations that have no effect on membrane integrity
(15–17).
A primary target for volatile agents such as toluene and
1,1,1-trichloroethane (TCE) is the γ-aminobutyric acid A
receptor (GABAA). By increasing the affinity of the GABAA
Reproduction (photocopying) of editorial content of this journal is prohibited without publisher’s permission.
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receptor for the endogenous neurotransmitter γ-aminobutyric
acid (GABA) volatile inhalants cause CNS depression, euphoria,
and muscular in-coordination (13,18,19). Compounds structurally related to DFE are also believed to sensitize myocardial
tissue to endogenous catecholamine leading to potentially fatal
arrhythmias (5,20). In one study, three of 12 dogs developed
cardiac arrhythmias and died following exposure to 0.405 g/L
of DFE and 0.008 mg/kg of epinephrine (21). Although that
study demonstrated DFE’s cardiotoxic potential, tissue DFE
levels were not given. In three postmortem cases where sudden
death was attributed to the arrhythmogenic effects of DFE,
femoral blood DFE levels were 83.5, 136.3, and 218.1 mg/L
(22). The mechanism of action which leads to arrhythmia and
cardiac failure is poorly understood. Recently Jiao et al. (23) reported a decrease in conduction velocity in cardiomyocytes
treated with CF3Br and epinephrine but not CF3Br alone. Their
results were not conclusive because they could not exclude alternate toxic mechanisms such as involvement of other ion
channels.
Given that DFE exerts toxic effects in two different organ systems it is possible that differences in relative uptake and elimination in these organ systems partly explains some of the observed effects. Although volatile toxins have been shown to
act on specific protein targets within the CNS, this does not
preclude the possibility that high concentrations of DFE and
other solvents also have effects on membrane integrity. Therefore if accumulation of DFE occurs with repeated dosing, effects on membrane integrity might predominate.
Little data exist regarding the relative uptake and elimination of DFE in the different biological compartments. Those
studies which have been performed have either used low dose
exposures similar to those which might be encountered in the
work place or have not conducted chemical analyses on relevant tissues such as brain, heart, and liver (21). For wellstudied toxins, measured tissue levels may be used to estimate
the effects, dose, time, and/or duration of exposure. With respect to DFE, such conclusions are currently not possible.
The purpose of this work was to measure DFE concentrations in different tissue compartments in the rat as a function
of time following exposure to levels of DFE consistent with
abuse. The protocol examined the uptake, distribution and
elimination phases of DFE in different body compartments. Tissues of interest included blood, because of its relevance to
postmortem and human performance toxicology, and brain
and heart because they are the primary target organ systems
for DFE. In addition, because of their importance in toxicology
and their high blood perfusion rate, liver and kidney were also
collected and analyzed. Upon completion, plots of DFE concentration over time were used to identify the toxicokinetic
model that best describes DFE uptake and elimination. Plots
also allowed for the reasonable estimation of DFE bioavailability following inhalation exposure and provided the experimental determination of DFE half-life (t½) in blood, brain and
heart.
Dosed animals were observed for evidence of intoxication in
the form of prostration and reduced motor activity. These effects were then correlated to DFE tissue concentrations in
CNS (brain), heart, and blood. Particular attention was paid to
382
the relative concentration of DFE in brain and cardiac tissue
because of the evidence that the mechanism of action of halogenated hydrocarbons in those systems may be distinct. Therefore differences in the relative uptake and/or concentration in
these tissues may be critical to development of toxic effects.
Materials and Methods
Male Sprague-Dawley (SD) rats (300 g) were obtained from
Taconic Farms (Germantown, NY). Animals were housed two
per cage for two weeks after arrival. The animals had free access
to food and water, and were kept on a 12-h light/dark cycle. All
protocols were approved by the St. John’s University Institutional Animal Care and Use Committee. 600 g of > 98% 1,1-difluoroethane was obtained from Sigma-Aldrich Chemical St.
Louis, MO. A Labcrest Flowmeter # 448-225 had been previously obtained. All other apparatus were designed in house.
DFE exposure
The toxicokinetic study was conducted using rats (n = 30),
exposed to a fixed amount of DFE. DFE delivery was accomplished using an apparatus (Figure 1) that feed gas to a
Figure 1. Dynamic exposure chamber.
Figure 2. DFE tissue concentrations of exposed (30 s of 20 L/min) Sprague
Dawley rats.
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dynamic exposure chamber large enough to contain two animals (chamber = 2298 cm3). Gas from a 600-g canister of >
98% DFE was passed through a gas regulator equipped with a
shut off valve. The regulated shut off valve fed gas to a calibrated gas flow meter set to deliver 20 L/min of DFE. The flow
meter controlled gas flow into the exposure chamber. Gas outflow from the chamber was passed into a
charcoal filter for safety.
Animals were weighed then exposed to
30 s of 20 L/min DFE gas. Sample introduction was controlled precisely by using
the shut off valve which allowed instantaneous control of flow. After exposure
the chamber was immediately opened
and the animals were removed. Animals
were sacrificed and tissues were collected
at fixed intervals. Groups (n = 3) were
sacrificed at 0, 10, 20, 30, 45, 60, 120,
240, 480, and 900 s. Animals were killed
by decapitation, using a guillotine. Trunk
blood was collected in 20-mL headspace
vials which were crimped immediately.
Other collected tissues included brain,
heart, liver and kidney. After a satisfactory amount of each tissue was collected,
it was placed immediately into a 20-mL
headspace vial, which was sealed. All
specimens were subsequently labeled and
stored frozen at –12.8°C.
Analytical
Figure 3. Blood (A, B); brain (C, D); heart (E, F); liver (G, H); and kidney (I, J) from DFE-exposed Sprague
Dawley rats.
Specimens were analyzed for DFE
using an Agilent 6850 gas chromatograph equipped with a 7694 headspace
autosampler. The system was fitted with
a flame-ionization detector. System calibration and specimen analysis were performed using a previously reported
method (24). Further data analysis was
performed using Graph Pad Prism software (v. 4, San Diego, CA).
Prior to analysis the weight of each
collected specimen was determined and
recorded. Tissue were grouped (all
bloods, brains etc) and analyzed for DFE
content using the method previously described. After nominal tissue concentrations were determined, samples were corrected for weight and volume before
recording final concentrations. Tissue
concentrations were plotted against time.
To calculate t½ of DFE in selected compartments, concentration results were
collected from nonlinear regression analysis at points that occurred in the alpha
(α), beta (β), and gamma (γ) phases. For
the elimination phases multiple points
were determined and the results were averaged. Point selection, accomplished by
visual inspection of graphs, and toxicokinetic equations are described by
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Medinsky and Valentine (25).
Maximum DFE exposure (DoseMax) for 320 g male rats was
estimated using a respiratory minute volume of 0.21 L/min
(26). Pure DFE at 1 atm contains 2.7 g/L (24) therefore, 30 s of
2.7 g/L DFE = 283 mg or 0.884 mg/g for a 320-g animal assuming 100% absorption.
Total uptake (burden) in each tissue was calculated by taking
average tissue concentrations at time 0 (t0) and determining
total DFE amounts in each based upon average organ weights
of 320-g male Sprague Dawley rats. The blood volume was estimated to be 21.9 mL (27). Other estimated tissue weights
were, brain 2.1 g, heart 1.3 g, liver 8.6 g, and kidneys 2.5 g (28).
Tissue burdens were summed to estimate total body burden.
Percent uptake of individual tissue compartments = tissue
burden (μg)/DoseMax (μg) · 100. Total percent uptake = total
body burden/ DoseMax · 100.
The area under the curve following a single acute inhalation
exposure in blood (AUCAcInh) was estimated using the trapezoidal method. The elimination rate constant (β) was determined using data collected from 30 to 240 s using the equation:
β = ln(CBld1) – ln(CBld2)/t2 – t1.
The volume of distribution (VdAcInh) following a single acute
inhalation exposure was determined using VdAcInh = Dose
(mg)/(β · AUCAcInh). Clearance (CL) from blood was determined
using CL = β · VdAcInh. As a confirmatory method t½ in blood
was also calculated using the equation t½ = 0.693/β.
Results
During exposure all animals showed signs of intoxication
manifested as sedation which began at about 20 s and rapidly
progressed to more profound intoxication. At the end of the exposure period animals were prostrate and had lost all righting
reflex. Animals remained visibly intoxicated until about 4 min
as evidenced by either an absence of ambulation or staggered
gait. From 4 to 8 min, animals showed signs of sedation in the
form of reduced aggression and excitement during handling.
Recovery was rapid, and at 8 min post exposure, animals
showed no signs of obvious intoxication.
Point to point plots of all tissue results are presented in
(Figure 2). Individual tissue data following nonlinear analysis
of time versus concentration, plotted both standard and logarithmically (log C) are presented in (Figure 3A–3J). Correlation
analysis expressed as r2 was 0.9329, 0.9571, 0.9371, 0.8393, and
0.9060 for blood, brain, heart, liver, and kidney, respectively.
The graphed results of log C versus concentration produced
curved lines characteristic of compounds that exhibit multicompartment toxicokinetics. Of particular interest was the
relative concentration of DFE in the target tissues, brain, and
heart so these tissues were graphed together as shown in
(Figure 4).
The results from regression analysis (data not shown) of
Table II. Total DFE Tissue Burden (µg) and Uptake (%) in
DFE-Exposed Sprague Dawley Rats
Estimated
Tissue Amount
in 320-g Rat
Tissue
Figure 4. Regression analysis of brain and heart DFE levels (log C) from
Sprague Dawley rats.
Blood
Brain
Heart
Liver
Kidney
21.9 mL
2.1 g
1.3 g
8.6 g
2.5 g
Total
Table I. DFE Distribution and Elimination Rates in DFEExposed Sprague Dawley Rats
Average DFE
Conc. at t0
(mg/L or mg/kg)
Total Tissue
Burden
(µg)
Uptake
(%)
352
519
338
136
364
7708
1090
439
1169
910
2.72
0.38
0.15
0.41
0.32
11,316
Table III. Toxicokinetic Parameters Determined
Experimentally from Sprague Dawley Rats Acutely
Exposed to DFE
Tissue
α
(seconds)
β
(seconds)
γ
(seconds)
Blood
9
86
range 75–99
240
range 200–284
Brain
18
110
range 54–157
340
range 272–405
β
231
range 224–242
VdAcInh
0.06 L
CL
0.03 L/min
Heart
384
27
168
range 97–218
Parameter
Value
AUCAcInh
409.8 mg · min/L
0.48/min
4.0
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blood, brain, and heart were used to calculate t½ during the
distribution and elimination phases. The initial distribution
phase (α) for blood, brain, and heart was much shorter than either of the other elimination phases. As a consequence only a
single point could be calculated starting at t0. The half time of
all calculated specimen results in the various phases of distribution and elimination are summarized in Table I.
DoseMax for 320-g male SD rats was estimated to be 283 mg
(283,000 μg). Total tissue burdens were calculated by multiplying the tissue concentration at t0 by the estimated tissue
weight. Total body burden from these compartments = 11,316
μg. Total percent uptake = 11,316 μg/283,000 μg · 100 = 4.0%.
Percent uptake of individual compartments was calculated by
dividing individual tissue burden by the calculated total dose.
Results are shown in Table II.
The AUCAcInh was found to be 24,588 mg · s/L (409.8 mg ·
min/L). β = ln(115) – ln(21)/240 – 30 s = 0.008/s (0.48/min).
VdAcInh = 11.3 mg/(0.48/min · 409.8 mg · min/L) = 0.06 L. CL
= 0.48/min · 0.06 L = 0.03 L/min and blood half life calculated
using the equation t½ = 0.693/β = 86.6 s. Summary results are
presented in Table III.
Discussion
Given that, in 2005, over 25% of reported inhalant abuse involved misuse of duster products, research into DFE toxicology
is needed. DFE has been described as practically nontoxic (21),
and when used as intended, it poses little risk of adverse outcome. Unfortunately, DFE-containing products have been diverted from their original intended use, and DFE has become
a substance of abuse. When abused, substantially greater exposure to DFE occurs, and as a result, this volatile gas has entered the arena of medicolegal investigation.
Toxicokinetic parameters for DFE are unreported (21). The
studies into DFE toxicology have focused primarily on DFE
toxicodynamics and have not correlated response to tissue
levels. This research set out to determine some of the fundamental kinetic parameters using a protocol mimicking DFE
abuse, an experimental design that has not been attempted.
The protocol allowed for the in vivo determination DFE during
the distribution and elimination phases and for the reasonable
estimation of bioavailability. Tissue plots of log concentration
versus time from 0 to 900 s resulted in curved lines typical of
compounds that distribute according to a multi compartment
toxicokinetic model (25).
Inhaled, DFE is unlikely to be significantly absorbed in either
the nasopharyngeal region or the trachea, even in obligatory
nose breathers like the rat, because of its low solubility in
water (0.017 g/mL) (21). As a result the majority of DFE absorption likely occurs in the alveoli, where it equilibrates with
the blood of the pulmonary capillaries. With a favorable concentration gradient, DFE diffuses into blood and is distributed.
Distribution of gases like DFE are limited by the perfusion
and solubility in the various tissues (29). Accordingly, a physiologically based toxicokinetic (PBTK) model developed for
1,1,1,2-tetrafluoroethane found that inhalation uptake was
still 2–3% 2 h after initiation of exposure due to continued uptake into adipose (30). Considering the much shorter time of
DFE exposure used here (30 s), poorly perfused tissues like adipose are unlikely to have accumulated substantial DFE levels.
Multiple doses of DFE are typical in abuse and are likely to increase DFE body burden due to continued uptake into lowly
perfused tissues.
Additional body burden would alter the toxicokinetic parameters determined in this experiment. Vd and other toxicokinetic data are ideally determined using bolus amounts of
drug or chemical administered instantaneously via intra venous injection. In addition, DFE is rapidly eliminated by exhalation. Therefore, the Vd derived here from acute exposure
(0.06 L) using AUC as a measure of blood concentration may
underestimate DFE kinetic parameters associated with chronic
use. Calculating the initial volume of distribution using blood
concentration at t0 and animal weight provided an estimated
Vd = 0.10 L/kg (range 0.08–0.12) compared to 0.06 L which was
determined using AUC. Further experiments are required to assess the impact multiple doses would have on distribution and
elimination.
After cessation of DFE exposure, the animals remained visibly intoxicated until about 4 min, and blood levels averaged 21
mg/L. Two reports of fatal MVA involving DFE intoxication reported blood levels of 29.8 and 78 mg/L (9,10) which correlate
well to the observed effects of DFE intoxication in this study.
During the period from about 4–8 min, animals remained sedated, then recovered quickly at about 8 min when blood levels
had fallen to 10 mg/L.
Adult exposure following a single dose of DFE can be estimated by multiplying the concentration of DFE at 1 atm (2.7
g/L) by lung capacity 6 L (31). Using this calculation, an estimated 16.2 g of DFE may be inhaled in a single dose, equivalent to 0.231 mg/g for a 70 kg individual. In contrast the estimated maximum DFE dose in this study was 283 mg or 0.884
mg/g in 320-g rats. The aforementioned body burden calculations assume 100% absorption but the actual bioavailability of
inhaled DFE and other gases are much less than 100%. The uptake of the structurally similar compound 1,1,1,2-tetrafluoroethane was estimated to be 3.7% (30) and only 1.6% for inhaled 1,1,1-trifluoroethane (32), however both of these reports
had to rely on estimated tissue partition coefficients for toxicokinetic modeling because true coefficients were unavailable.
From this research, the uptake of DFE from a single dose was
estimated to be 4.0%. Therefore, given a 16.2-g dose and the
apparent bioavailability, the uptake in grams in human adults
is approximated to be 0.648 g.
The % uptake into blood in the rat was 2.7%. Assuming a
16.2-g dose, similar uptake and the blood volume of human
adults (5 L), the estimated human blood concentration would
be 87.5 mg/L. Remarkably the blood concentration in rats was
highly correlated to this estimation for human subjects. The
experimentally determined rat blood peak concentration was
about four times higher (352 mg/L) than the calculated estimation of peak blood concentration in humans (87.5 mg/L).
The rats however received a dose that on a mg/g basis is also estimated to be nearly four times greater than the estimated
human dose (0.231 mg/g human vs. 0.884 mg/g rat).
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Documented human exposures have been reported (33). The
blood estimate of 87.5 mg/L is in agreement with some of the
reported cases. Other blood results, however, have been higher
(> 300 mg/L). Possible explanations are that multiple doses
were taken prior to death or that the direct consumption and
the forcible holding of breath during exposure caused DFE
pressure to be higher than 1 atm. Higher pressures result in
lung concentrations greater than the assumed 2.7 g/L subjecting individuals to the concentration effect as described by
Wollman and Dripps (34). Combined with high pressure this
condition would more effectively drive DFE into the blood.
Brain levels were highest at t0 and at peak, were highest of
all tissues analyzed. Even though brain levels were at least
167 mg/(L or kg) higher than any other measured tissue at
peak, by 60 s concentrations had dropped below all others and
remained lower until the final group when all tissues approached unity. This finding is similar to that observed for
toluene where clearance from mouse brain was most rapid of
all measured tissues (35). Rapid clearance from brain is unexpected when considering the lipophilicity of many abused inhalants and the brains limited capacity for metabolism. This
phenomenon has been explained by the high perfusion rate of
the brain (750 mL/min) (36).
CNS effects were profound at t0 and visible intoxication
lasted until approximately 4 min when brain DFE levels were
17–18 mg/kg. Sedation lasted until about 8 min, when brain
DFE levels were about 6 mg/kg. One case study that reported
brain DFE levels after fatal MVA and intoxication appeared to
be a factor in the crash found DFE brain levels of 11.7 mg/kg
(9). Similar DFE levels occurred in rats at 5.5 min using this
protocol, a point where animals were still showing signs of sedation. In another unreported case involving intoxication (unpublished data) leading to fatal MVA, brain levels were 55.2
mg/kg. An uninvolved motorist observed the deceased driving
erratically at a high rate of speed. During this study similar
brain concentrations occurred in the animals at about 1 min
and 15 s after withdrawal of DFE exposure, a time when animals were still showing gross signs of intoxication.
Heart DFE levels following DFE exposure are unreported.
The study protocol employed here allowed heart DFE levels to
be simultaneously determined along with other tissues. Previous reports (33,37) speculated that if DFE elimination proceeded more rapidly in brain versus heart, then abusers might
re-dose as CNS effects wore off while cardiac tissue levels remained high. Multiple doses of DFE might then lead to the accumulation of DFE in cardiac tissue. In this experiment, DFE
levels in cardiac tissue were reduced by half at a rate 9 s slower
than brain during the α phase (27 vs. 18 s), and t½ was nearly
a minute longer (168 vs. 110 s) in heart versus brain during β
elimination. Consequently, heart levels did remain higher than
brain tissue after about 60 s post DFE withdrawal. The relationship between the relative amounts of DFE in these target
tissues can be clearly seen in Figure 4.
This is the first evidence that DFE levels in cardiac tissue are
higher than brain following abuse at a time when CNS effects
are subsiding. Volatile toxins have been shown to act through
specific targets but that evidence does not negate the possibility
that high levels of DFE cause adverse effects on membrane in-
386
tegrity and accumulation of DFE within membranes may be
critical to the development of toxic effects. Supportive evidence is found in one unpublished case of apparent intentional overdose using DFE where cardiac tissue contained
492.4 mg/kg and brain contained 224.1 mg/kg (unpublished
data).
DFE levels in liver peaked 10 s after cessation of exposure
(187 mg/kg), the only tissue in which this occurred. Following
inhalation exposure dissolved gases are initially only supplied
to the liver through the hepatic artery. Blood flow to the liver
via the hepatic artery occurs after passing through the lipid
rich brain. Liver DFE levels peaked lowest of all measured tissues and had the second lowest concentrations after 60 s, following brain, throughout the 15 min duration of the study. The
observed levels probably reflect these physiologic factors.
DFE and similar compounds are expected to be only sparingly metabolized (< 1%), (38) and the majority of elimination
occurs by exhalation of gas through the lungs. A proposed
mechanism of DFE oxidation (39) in part proceeds through
CYP-2E1 forming intermediate aldehydic metabolites. It has
been suggested that as fluoride substitution increases in hydrofluorocarbons, there is an increase in the accumulation of
aldehyde metabolites due to a decrease in alcohol dehydrogenase activity (40).
The accumulation of an electrophillic species capable of covalently binding cellular macromolecules could reasonably be
argued to provide a mechanism for DFE toxicity. However,
chronic dosing studies have not shown evidence of hepatic fibrosis or other organ damage that could be related to accumulation of aldehydic products of DFE metabolism (21). Nevertheless the levels of exposure to DFE as seen during abuse
were largely unanticipated by manufacturers and researchers
and toxic metabolites may yet prove to be problematic to heavy
abusers. In fact, in one two-year study where rats were exposed to DFE doses 6 h/day, 5 day/week, there was a dosedependent increase in urinary fluoride suggestive of DFE
metabolism (21).
Kidney DFE peak concentrations (364 mg/kg) were observed
at t0 and were similar in magnitude to those seen in blood and
heart. Kidney DFE concentrations were higher than all other
measured tissues from about 130 s until the end of the study.
In fact kidney DFE levels were twice that of the next highest
tissue (heart 15, kidney 32) at 480 s and are estimated by regression analysis to be almost 3 times higher than heart at
about 12 min. This observation could mean that kidney is susceptible to DFE accumulation. In addition aliphatic hydroxylation of DFE to hydrophilic alcohols and subsequent formation aldehydic species could uniquely predispose kidney to
DFE toxicity but again, no statistically significant lesions in
kidney were observed in chronic dosing studies (21).
A single 10-oz canister contains 283.5 g of DFE. Using the
toxicokinetic calculations determined in this research, one
canister holds enough gas for almost 18 “hits” of inhalant in a
fully developed adult. Based upon the observations of the intoxicated animals used in this research, one could speculate
that, to maintain intoxication, an abusing individual would
re-dose approximately every 10 min, and therefore, one canister contains enough DFE for 3 h of intoxication. Using this
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scenario, it is likely that minimally perfused tissue compartments would continuously uptake DFE, and chronic abusers
may therefore have slower elimination rates and experience
longer lasting effects when compared to acute abusers. Additional studies are required to verify if continued accumulation
of DFE in heart versus brain occurs after multiple doses as is
suggested by this research.
Conclusions
Using the animal exposure protocol and data presented, the
author’s successfully calculated some standard toxicokinetic
parameters following acute exposure to DFE. The dose was
selected to mimic exposure during intentional abuse, a protocol which has previously not been reported. Measured animal
tissue results were correlated to the observed effects and were
compared to results from medical examiner case work providing a basis for interpreting DFE tissue levels. In addition this
experiment found that DFE levels in cardiac tissue remained
higher than those in brain at a time when the intoxicating effects were subsiding. This finding suggests that DFE may accumulate in cardiac tissue with repeated exposure during
abuse.
References
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samhsa.gov/nsduh/reports.htm (2006).
2. National Inhalant Prevention Coalition. Guidelines for Medical
Examiners, Coroners and Pathologists: Determining Inhalant
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Manuscript received January 7, 2010;
revision received April 19, 2010.