Acta Anaesthesiol Scand 2004; 48: 469—473
Printed in Denmark. All rights reserved
Biochemical markers for brain damage after carbon
monoxide poisoning
L. S. RASMUSSEN1, M. G. POULSEN1, M. CHRISTIANSEN2 and E. C. JANSEN1
1
Department of Anesthesia, Center of Head and Orthopedics, and 2Department of Clinical Biochemistry, Statens Serum Institut, Copenhagen,
Denmark
Background: Carbon monoxide poisoning is associated with
high mortality and a substantial risk for brain damage in sur-
vivors. Evidence for acute brain dysfunction may be obtained
by measuring concentrations of suitable biochemical markers.
We hypothesized that increased serum concentrations of
Neuron-specific enolase (NSE) and S-100b protein could be
detected after carbon monoxide poisoning and that the con-
centration would correlate with the severity of intoxication.
Methods: Prospective non-interventional study in the university
hospital. We included 20 patients admitted for hyperbaric treat-
ment due to carbon monoxide poisoning. Serum levels of NSE
and S-100b protein were measured in all patients on admission
and after 12, 24, 36 and 48 h. As a control group, we included 20
patients who underwent elective hyperbaric treatment.
Results: Serum concentrations of NSE and S-100b protein
were not significantly different from the controls, with median
values at admission being 10.6 vs. 9.7 mg l
1 and 0.15 vs.
C ARBON monoxide (CO) poisoning is a common
condition with a high mortality, and in those
surviving a serious intoxication there is a substantial
risk for long-term brain dysfunction (1, 2). Serious
poisonings are treated with hyperbaric oxygen
(HBO) and the severity of intoxication is assessed
using various parameters such as carboxyhemoglobin
concentration, level of consciousness, and other neu-
rological symptoms and signs (3, 4). Additional evi-
dence for acute brain injury could be obtained by
measuring blood concentrations of biochemical mar-
kers for brain damage. This could be helpful as a
prognostic marker and perhaps aid in deciding who
should undergo HBO.
Neuron-specific enolase (NSE), a glycolytic enzyme,
and S-100 protein, a glial cytoplasmic protein, have
been shown to be useful markers of cerebral damage
after head trauma, cerebral infarction, cardiac arrest,
and following cardiac surgery (5—8). The markers are
assumed to be released from neuronal and glial tissue
to the blood when cell membrane integrity is lost. The
markers may be used in clinical care but also in
Copyright # Acta Anaesthesiol Scand 2004
ACTA ANAESTHESIOLOGICA SCANDINAVICA
doi: 10.1111/j.1399-6576.2004.00362.x
0.13 mg l
1, respectively (P ¼ 0.82 and P ¼ 0.38). The concentra-
tions did not change significantly during the sampling period.
We were unable to show any significant relation to level of
consciousness.
Conclusion: Blood concentrations of NSE and S-100b protein
were not significantly increased after carbon monoxide
poisoning and do not seem to be related to a history of
unconsciousness.
Accepted for publication 19 December 2003
Key words: Brain damage; carbon monoxide; neuron-specific
Enolase; S-100b protein.
# Acta Anaesthesiologica Scandinavica 48 (2004)
research, for example, as surrogate for neuropsycho-
logical testing in a pretreatment assessment to group
patients or in a follow-up assessment of outcome after
an intervention.
We hypothesized that CO poisoning would be
accompanied by elevated blood concentrations of
NSE and S-100 protein and that the concentration
would correlate with a history of unconsciousness,
because this is considered to be an important measure
of the severity of the intoxication.
Materials and methods
This was a prospective observational study of adult
patients (>18 years) admitted to our unit from
September 1999 to April 2001 for HBO treatment of
CO poisoning. The hospital is a tertiary care hospital
with a hyperbaric chamber and we treat patients
from our own community as well as those transferred
from outside hospitals. The study was approved by
the Research Ethics Committee. Written informed
469
L. S. Rasmussen et al.
consent was obtained from patients or from their next
of kin.
Indications for HBO treatment after exposure to CO
were:
* Any new neurological deficit, including low level
of consciousness.
* CO-level above 25% regardless of symptoms
present.
Neurological function was assessed on admission and
we classified the patients into two groups: those in
which unconsciousness was found at the scene or
during transfer to hospital, and those who were not
unconscious. We defined unconsciousness as a
Glasgow Coma Scale score below 13.
All patients received pure oxygen by open facemask
during transfer to the hyperbaric chamber. Endo-
tracheal intubation was performed before transfer in
case of unconsciousness and ventilation with pure
oxygen was accomplished, aiming at an arterial car-
bon dioxide tension of 4.5—6 kPa. After endotracheal
intubation, sedatives or neuromuscular blocking
agents were generally not necessary.
The treatment in the hyperbaric chamber consisted of
exposing the patient to an environmental and oxygen
pressure of 2.8 bars for 90 min. The protocol applied
involves three treatments within the first 24 h and two
daily treatments on the following days until the patient
is free from neurological impairment, does not
improve, or dies. The hyperbaric chamber (Drass
Galeazzi, Zingonia, Italy) is equipped with devices to
monitor the inspired gas throughout the hyperbaric
oxygen treatment. An FiO2 of 95—100% is assured.
Oxygen is given by a respirator (Siemens 900C,
Siemens AB, Stockholm, modified by us for hyperbaric
use), face mask (Hyperlite, Divex Ltd, Aberdeen, UK)
or hood (Amron Ltd, Escondido, CA). The chamber is
equipped for full intensive care if needed.
Venous blood samples were drawn at admission
and 12, 24, 36 and 48 h later.
In addition, as a control group, we included 20
patients admitted to our department for elective
HBO treatment because of postradiation osteonecrosis
of the mandible or chronic ulcers. In these subjects we
collected blood samples at admission as well as 24
and 48 h later. All blood samples were taken as 10-ml
venous samples. After centrifugation, serum was
stored frozen at
20 C until analysis.
Blood analyses
Neuron-specific enolase was determined by a time-
resolved fluoroimmunoassay DELFIA1 NSE kit
(Wallac OY, Turku, Finland). The detection limit is
470
1.0 mg l
1 and the normal level is below 12.5 mg l
1.
All controls, internal as well as external, were within
two standard deviations (SD) of the assigned value.
Hemolyzed samples were discarded because erythro-
cyte enolase interferes with the analysis in case of
gross hemolysis.
S-100b was determined by a commercially available
monoclonal two-site sandwich immunoluminometric
method LIA-mat1 Sangtec1100 (AB sangtec Medical,
Bromma, Sweden). The sensitivity of the method is
<0.02 mg l
1 and the normal level is below 0.5 mg l
1.
During the sample run, all controls were within 0.5 SD
of the reference value.
Statistics
Demographic data and blood concentrations of NSE
and S-100b protein are reported as median values
with interquartile range (IQR). Some S-100b protein
values were below the detection limit, and in that case
the level was presumed to be 0.01 mg l
1.
We compared the concentrations of NSE and S-100b
protein between groups with Mann—Whitney’s rank-
sum test and within groups using Wilcoxon’s rank
sum test. P-values <5% were considered statistically
significant.
We calculated that a total sample size of 40 would
allow us to detect a difference in NSE of 3 mg l
1
between controls and CO-intoxicated patients, expect-
ing a standard deviation of 3 mg l
1 and accepting a
type 1 error risk of 5% and a type 2 error risk of 10%.
We aimed at 20 patients because this would allow us
to analyse changes in the concentrations of the
biochemical markers over time using paired statistics
with the same risk of type 1 and type 2 errors.
Statistical analysis was performed using the SAS for
Windows computer programme. version 6.2. (SAS
Institute, Cary, NC, USA).
Results
Of the included 20 patients (Table 1), 16 were men and
four were women. Eleven had a history of uncon-
sciousness while nine had not been unconscious.
Eight had attempted suicide. All underwent treatment
with hyperbaric oxygen for a maximum of 2 days.
Two patients died after 9 and 22 days, respectively,
without regaining consciousness. The remaining 18
patients obtained full neurological recovery and they
were discharged from the department after a median
of 3.5 days (IQR: 2—6 days). Four patients were
admitted to a department of psychiatry after a suicide
attempt and two patients were transferred to a
 Carbon monoxide and brain damage

Table 1
Characteristics for 20 patients admitted with carbon monoxide poisoning.
Age, years Mechanism History of unconsciousness Initial carbon monoxide percentage
60 Fire in building Yes 40.2
26 Fire in building Yes 27.8
50 Fire in building Yes 46.6
31 Exhaust from engine Yes 23.1
36 Exhaust from engine Yes 25.0
28 Fire in building Yes 15.6
31 Fire in building No 3.2
33 Exhaust from engine No 32.0
38 Exhaust from engine Yes 44.0
45 Exhaust from engine No 37.0
45 Fire in building No 31.0
52 Fire in building No 18.0
60 Fire in building No 12.2
31 Fire in building Yes 24.0
51 Exhaust from engine Yes 30.4
34 Use of charcoal inside No 23.6
72 Fire in building Yes 42.0
54 Fire in building No 34.2
42 Exhaust from engine No 28.2
37 Fire in building Yes 53.0

medical department for further rehabilitation. The Discussion

controls were free of neurological disease and they
had a median age of 61 years (IQR: 52—67 years); 15
were men and five were women.
Serum concentrations of NSE and S-100b protein are
shown in Table 2. The serum concentrations of NSE
and S-100b protein were not significantly different
from the values in the control group at any time
point. The concentrations did not change significantly
during the sampling period (all P-values >0.30). The
duration of exposure to CO was very difficult to
establish but the time from end of exposure to the
time for the first blood sample was a median of 3.6 h
(IQR: 2.1—6 h). Values of NSE and S-100b protein were
not significantly higher in patients who had been
unconscious (Table 3).
This study used two markers of neuronal injury,
which has not been described in CO-intoxicated
patients before. Increasing levels of S-100b protein
have been found after several types of brain damage
and a significant correlation was found between the
S-100b protein level and the chance of regaining con-
sciousness after cardiac arrest (7). After coronary
artery bypass grafting, increasing serum levels of
both NSE and S-100b protein have been reported,
and a significant correlation was found between the
increase in the serum level of NSE and cognitive
dysfunction (8). Brain imaging studies have shown
that CO causes damage of the hippocampus, ganglia,
and white matter. Carbon monoxide exerts its toxic

Table 2
Serum concentrations (mg l
1) of Neuron specific enolase (NSE) and S-100b protein in 20 patients with carbon monoxide poisoning (CO)
and in 20 patients undergoing elective hyperbaric treatment (controls). Median values with interquartile range (IQR). No significant
difference was found between the two groups (Mann—Whitney’s test).
NSE mg l
1 S-100b protein mg l
1
CO Controls CO Controls
On admission 10.6 (7.7—13.8) 9.7 (8.3—11.7) 0.15 (0.06—0.25) 0.13 (0.04—0.22)
12 h 10.3 (7.9—18.0) 0.09 (0.04—0.29)
24 h 10.1 (7.2—16.1) 9.9 (8.5—12.1) 0.19 (0.07—0.27) 0.18 (0.09—0.50)
36 h 10.2 (8.0—12.9) 0.12 (0.05—0.33)
48 h 10.8 (7.6—13.9) 12.1 (8.8—14.8) 0.04 (0.02—0.23) 0.11 (0.05—0.30)

471
L. S. Rasmussen et al.
Table 3

1
Peak serum concentrations (mg l ) of Neuron specific enolase
(NSE) and S-100b protein after carbon monoxide poisoning
according to level of consciousness. No significant difference
was found (P ¼ 0.40 and P ¼ 0.29, respectively, Mann—Whitney’s
test).
Unconscious (n ¼ 11) Not unconscious (n ¼ 9)
NSE S-100 protein NSE S-100 protein
Median 14.7 0.26 10.8 0.20
Mean 17.7 0.44 14.5 0.27
IQR 9.4—23.5 0.11—0.71 9.1—11.9 0.07—0.25
SD 11.1 0.51 9.1 0.32
effects by inhibition of oxygen release from hemoglo-
bin in peripheral tissues, by blocking oxidative meta-
bolism, and by lipid peroxidation (1,3). The brain is
particularly sensitive and, in addition, to acute neuro-
logical symptoms, late brain dysfunction is common
with personality changes, Parkinsonism, and cogni-
tive impairment in approximately 30% of the patients
(9,10).
We were unable to demonstrate that carbon
monoxide poisoning is associated with high blood
concentrations of biochemical markers for brain
damage. The level was only half of the level found
24 h after cardiac arrest in individuals who did not
regain consciousness (7).
The blood concentrations remained stable for as
much as 36—48 h after arrival, so it seems that we did
not miss a secondary increase.
Two of our 20 patients died and very high levels of
NSE and S-100b protein were found in one of these
(44.0 mg l
1 and 1.82 mg l
1, respectively, after 12 h) but
in the entire group, no significant difference was
found according to the level of consciousness.
HBO treatment per se did not seem to induce any
release in the biochemical markers based on the data
from the group undergoing elective HBO.
There are a number of limitations in our study. The
time profile of NSE and S-100b protein were difficult
to interpret due to insufficient data on duration of
exposure and to a high variability in time from end
of exposure to arrival in our facility. The half life of
NSE is approximately 24 h and 2 h for S-100b protein
but in cases of brain damage the kinetics are compli-
cated due to different patterns of release (focal/dif-
fuse lesions) and clearance.
Other factors may have explained the level of con-
sciousness, such as intoxication and especially cya-
nide in house-fire victims. It should also be taken
into account that other results may have been
obtained if the cerebrospinal fluid was analyzed.
472
We were unable to find a significant difference in
NSE or S-100b protein between the CO-intoxicated
patients and the control subjects but in such a small
number of patients this may partly be explained by
the sample size. However, the difference in NSE
median values at admission was less than 1 mg l
1 and
the calculated SD for NSE is approximately 5 mg l
1. The
detection of a difference of 0.2 SD would require
approximately 700 patients if the statistical power is
set to 80%.
Acute brain dysfunction after CO poisoning may be
detected as neurologic or neuropsychological dys-
function. Detailed neurological studies may predict
outcome but require some patient cooperation and
are difficult to quantify. Neuropsychological evalu-
ation is not possible if the patient is unconscious,
and in awake patients requiring hyperbaric treatment,
it may delay treatment. Thus, the use of biochemical
markers of neuronal injury could be of great value
in providing objective data on brain dysfunction
when deciding duration and number of hyperbaric
treatment sessions. Unfortunately, we were not able
to show that blood concentrations of NSE and S-100b
protein are useful for this purpose after carbon mon-
oxide poisoning and neither could we demonstrate a
difference in the release of brain-specific proteins
between conscious and unconscious patients. If a
difference does exist, it is probably small and a large
number of patients will be needed to document such a
difference.
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