Nephrol Dial Transplant (2003) 18: 2354–2358
DOI: 10.1093/ndt/gfg395
Original Article
Methaemoglobinaemia and haemolysis associated with hydrogen
peroxide in a paediatric haemodialysis centre: a warning note
Miriam Davidovits1, Ayala Barak2, Roxana Cleper1, Irit Krause1, Zahava Gamzo1 and
Bella Eisenstein1
1
Nephrology Clinic and Dialysis Unit, Schneider Children’s Medical Center of Israel, Petah Tikva and
Sackler School of Medicine, Tel Aviv University, Tel Aviv and 2Aylab Ltd, Tel Aviv, Israel
Abstract
Background. Haemodialysis exposes patients to contaminants in the dialysate. The AAMI standards deal
only with two disinfectants, chlorine and chloramine.
We report an event of methaemoglobinaemia and
haemolysis related to an unsuspected disinfection
agent.
Methods. Nine children aged 3–17 years undergoing
dialysis after reconstruction of our paediatric dialysis
unit developed methaemoglobinaemia of 3.1–11%,
with a mean reduction in haemoglobin levels of
11.9 ± 5.9% (P < 0.001). Air bubbles were noted in
the bloodlines. The water treatment system (WTS) of
the dialysis unit is disinfected when necessary by adding
concentrated hydrogen peroxide (HP) to the storage
tank and circulating it through the re-circulation loop
with draining and subsequent flushings. Total chlorine
analysis of the water is performed by DPD-iodide
colorimetric method.
Results. Dialysis water testing yielded a high chloramine concentration in the storage tank and pointsof-use stations (3.08 and 2.06 p.p.m., respectively).
However, this finding was not true for the tap water,
and it also failed to explain the air bubbles in the
dialysis tubing. The concentration of free chlorine was
within the recommended range. Further investigation
revealed that the WTS was disinfected by the service
company during remodelling of the unit, without
notification of the hospital staff. Since the DPDiodide test is not specific, and in effect detects not
only total chlorine, but all oxidants capable of oxidizing
iodide, we assumed the culprit was residual HP that was
inadequately flushed from the water system.
Conclusions. HP used for disinfection of the WTS can
pose a serious dialysis risk if not flushed out properly.
Correspondence and offprint requests to: Miriam Davidovits, MD,
Nephrology Clinic and Dialysis Unit Schneider Children’s Medical
Center of Israel, 14 Kaplan Street, Petah Tikva, Israel 49202.
Email: [email protected]
Total chlorine analysis should be performed before
every dialysis session, and positive results should
prompt further work-up for other oxidants. The clinical
staff must always be involved in decisions regarding any
intervention in the dialysis water system.
Keywords: children; haemodialysis; haemolysis;
hydrogen peroxide; methaemoglobinaemia
Introduction
The ancient Latin saying Aqua turbida non lavat (dirty
water does not wash clean) is particularly relevant to
haemodialysis patients. The vast quantity of water in
the dialysate originates from drinking water. Any
contaminants present in the water come into close
contact with the patients’ blood within the dialyser
across a thin non-selective membrane that has none of
the ‘wisdom’ of the gastrointestinal tract. Moreover,
owing to their compromised renal function, haemodialysis patients lack the ability to excrete toxins efficiently,
such that alterations in the chemical composition of the
water may cause acute and chronic complications.
Adverse effects have been reported in patients exposed
to dialysis water containing high concentrations of
aluminium, zinc, fluoride, copper, calcium or magnesium, sodium, nitrate, chloramine and sodium azid
[1]. Haemodialysis patients also risk exposure to the
chemicals used for the periodic disinfection of the fluid
pathways of the water treatment system (WTS) [2].
In 1981, the Association for the Advancement of
Medical Instrumentation (AAMI) established comprehensive water quality standards for the maximal concentration of minerals and heavy metals [3]. Most
WTSs are constructed to provide water that meets
the recommended standard. However, in the absence of
appropriate treatment of the system with regular and
stringent monitoring, the water used for preparing
Nephrol Dial Transplant 18(11) ß ERA–EDTA 2003; all rights reserved
Hydrogen peroxide-induced methaemoglobinaemia and haemolysis during HD
dialysate can pose a serious and even fatal risk.
Furthermore, the AAMI standards concern mostly
inorganic agents; chlorine and chloramine are the only
disinfectants listed, with suggested maximal levels of
0.5 and 0.1 p.p.m., respectively.
We report on an outbreak of methaemoglobinaemia
and haemolysis in a paediatric dialysis unit due
to contamination of the dialysate with hydrogen
peroxide (HP).
Subjects and methods
The Hemodialysis Unit of Schneider Children’s Medical
Center of Israel was opened in October 1995 with five stations
and enlarged to 11 stations in December 1997. At the
completion of reconstruction, nine patients aged 3–17 years
were being treated in the unit. Dialysis was performed with
Centry 3 and Gambro AK 100 machines and hollow fibre
polysulphon dialysers (Fresenius). No reuse was practiced. At
the beginning of the first dialysis session in the remodelled
unit, air bubbles were noted in the bloodlines. Assuming that
the problem was in the tubing sets or dialysers, we changed the
equipment. However, the problem persisted. Subsequently,
one patient turned grey and her blood in the tubing turned
chocolate brown. Methaemoglobinaemia was suspected
and confirmed by a high level of methaemoglobin (11%) in
this patient. All haemodialysis treatments were immediately
stopped; blood tests for haemoglobin, methaemoglobin and
haptoglobin were taken before disconnection. The one patient
with hypoxic symptoms and anaemia was immediately treated
with methylene blue. The children were transferred to another
hospital for continuation of dialysis.
Fig. 1. Scheme of the WTS of the dialysis unit.
2355
WTS (Figure 1)
City water is supplied to the dialysis unit from the hospital’s
water distribution system after chlorination. It flows through
a softener, an activated carbon filter (to absorb the chlorine
and halogenated disinfection by-products) and a reverse
osmosis unit. The treated water then passes to a 5000 l storage
tank and into a closed re-circulation loop. The stored water
circulates continuously, passing through two UV light treatment units and two microfilters before reaching the dialysis
stations.
Chlorine analysis of water
Chlorine and chloramine in the water are analysed by
DPD-iodide colorimetric method [4]. DPD (N,N-diethyl-Pphenilenediamine) is oxidized instantly by free chlorine to a
relatively stable free radical producing a red colour (Wurster
dye). Absorption is measured by colorimeter at 530 nm.
Because chloramines react slowly with DPD, they are
quantified by the subsequent addition of potassium iodide.
The iodide ion acts catalytically causing colour production by
monochloramine and dichloramine. The iodide-DPD test
method is typically used for measuring total chlorine, which is
the sum of free (chlorine reacting with DPD directly) and
combined (chlorine derivatives which react with DPD only in
the presence of iodide).
Method of disinfection
To disinfect the water, the reverse osmosis unit is shut off and
the UV light sources are disconnected. HP 30% is added
directly to the storage tank and diluted with dialysis water to
a final concentration of 1%. The HP is circulated in the
2356
M. Davidovits et al.
re-circulation loop for 2 h. Thereafter, the circulation pump is
stopped, the tank is emptied, the reverse osmosis system is
switched on and the storage tank is refilled with fresh dialysis
water. The re-circulation pump is operated for half an hour.
The storage tank and re-circulation loop are then drained
by opening all the point-of-use stations. Following several
flushing cycles, a test for oxidative substances is performed by
the DPD method at the points-of-use stations. Additional
flushing cycles are performed until the residue measures
<0.1 p.p.m.
Statistics
The data were analysed using the Student’s t-test (paired).
A P-value of <0.05 was considered significant. Correlations
were calculated using Spearman’s method for non-parametric
data.
Results
All patients showed methaemoglobinaemia with levels
ranging from 3.1 to 11% (normal <1%), and a mean
reduction in haemoglobin levels of 11.9 ± 5.9%
(P < 0.001) from the last measured value. A decrease
in haptoglobin level (normal >50 mg/dl) was noted in
two cases (Table 1). The methaemoglobin levels were
positively correlated with dialysis duration (r ¼ 0.8).
There was no correlation between duration of dialysis
and decrease in haemoglobin levels.
The findings of methaemoglobinaemia and haemolysis raised suspicions of a problem with the water
system. Results of water nitrate testing were negative,
but total chlorine analysis yielded a high concentration
of chloramines: 3.08 p.p.m. in the storage tank and 2.06
p.p.m. at the point-of-use stations. However, a negligible concentration of 0.02 p.p.m. was found in the tap
water (Table 2). The concentration of free chlorine in
the tap water was 0.33 p.p.m. (within the recommended
range of up to 0.5 p.p.m.) [5] and, as expected, the levels
in the storage tank and point-of-use stations were low.
Further investigation revealed that during the reconstruction and enlargement of the unit, the water system
was disinfected by the maintenance company without
notifying the clinical and technical staff of the hospital.
On consultation with water chemistry specialists, we
learned that the iodide-DPD method is not specific for
chlorine and chloramine and all oxidants capable of
oxidizing iodide, including HP, react with iodide-DPD
and are measured as total chlorine [6].
To correct the problem, additional flushings of the
WTS were performed until no oxidant residue was
detected by the DPD-iodide method at any of the pointof-use stations. No test for measurement of HP was
available at our unit at the time.
The next dialysis session was performed uneventfully.
Repeated blood tests for methaemoglobin showed
normal levels in all children.
Discussion
Nitrates or chloramine presenting in the dialysate water
have been implicated in methaemoglobinaemia and
haemolytic anaemia in dialysis patients [1,7,8]. The
investigations of Eaton et al. in 1973 [7] proved the
direct relationship between the oxidative effect (severity
of methaemoglobinaemia) and chloramine concentration in the water. The authors demonstrated that only
carbon filter is effective in removing chloramines [7,8].
Nowadays, chloramines are still frequently used as a
bactericidal agent to purify urban water. In most cases,
the municipal supplier does not inform the dialysis
unit staff about the time of chloramine addition,
which poses a danger of overwhelming the carbon
filter capacity and causing methaemoglobinaemia and
haemolysis in the treated patients [9].
In our event, although the measurement of total
chlorine seemed to indicate a high chloramine residue, it
did not account for the appearance of air bubbles in the
bloodlines. Moreover, even assuming complete failure
of the carbon filters, the chloramine level in the storage
tank cannot be higher than the level in the tap water.
We also failed to observe any increase in conductivity
of the reverse osmosis water due to deterioration of
the reverse osmosis membranes, which would almost
certainly have occurred had the chloramine concentration been high.
Since the disinfection of the water system with HP
added directly to the tank was performed just before the
Table 1. Duration of dialysis methaemoglobin levels and parameters of haemolysis during the event
Patient
Duration of dialysis (min)
Methaemoglobin (%)
Hb (g%)
Hb (%)
Haptoglobin (mg/dl)
1
2
3
4
5
6
7
8
9
Mean ± SD
120
100
70
110
105
5
50
35
10
11
6.9
5
5
4.8
4.6
4
3.4
3.1
5.3 ± 2.4
6.7
8.4
9.7
11.9
9.4
6.9
7.7
10.8
9.5
9 ± 1.7
5.6
16.8
12.3
10.9
17.5
4.2
20
14.8
5.2
11.9 ± 5.9a
101
–
11
68.6
–
116
–
74.2
43.2
a
P < 0.001.
Methaemoglobin, normal level <1%; haptoglobin, normal level > 50 mg/dl.
Hydrogen peroxide-induced methaemoglobinaemia and haemolysis during HD
Table 2. Chlorine and chloramine concentrations in the different
parts of the WTS
Tap water
Storage tank
Point-of-use stations
Chlorine (p.p.m.)
Chloramine (p.p.m.)
0.33
0.02
0.03
0.02
3.08
2.06
event, and the oxidant was detected only in the tank and
not in the tap water, it seemed reasonable to attribute
the observed clinical phenomena to the presence of HP.
Unfortunately, we could not directly measure HP
because the dialysis unit was not equipped with the
specific reagents at that time. (As the DPD-iodide
method cannot be used to quantify HP, the measured
values of 3.08 or 2.06 p.p.m. did not represent the actual
concentration of peroxide in the dialysis water.) Our
suspicions, however, were supported by the ample
evidence of the effects of HP in the literature.
HP, an oxidative substance, is used extensively in
medicine for its antiseptic qualities due to its ready
release of oxygen. It is produced endogenously during
electron transport processes in all aerobic cells and in
the blood from auto-oxidation of haemoglobin [10].
Studies have shown that the in vitro exposure of
normal human erythrocytes to increasing concentrations of HP is associated with methaemoglobin formation, lipid peroxidation and alterations in membrane
function in a clear dose-dependent fashion [11]. This
process is mediated by hydroxyl and ferrylhaemoglobin
radicals created from interaction of HP with iron and
other redox metals in a Fenton reaction [10,12]. Cellular
defence mechanisms against the potential damage by
HP consist of glutathione peroxidase and, especially,
erythrocyte catalase. Catalase decomposes HP to water
and oxygen without the generation of free radicals [13].
The amount of methaemoglobin generated on exposure
of red cells to HP is inversely proportional to the cell
catalase content [14].
The use of HP within enclosed body cavities or
irrigation under pressure in surgical procedures has
resulted in gas embolism and local emphysema due
to the liberation of oxygen: 1 ml of 3% HP solution
liberates 10 ml of O2 [15]. Colonic irrigation with HP,
recommended in the past for meconium plug syndrome,
was abandoned because of complications of gas
embolism to the mesenteric and portal vessels, gangrenous bowel and subsequent bowel perforation [16].
Portal venous embolism was reported in a 2-year-old
boy following ingestion of 3% HP solution [17]. The
trial use of i.v. infusion of HP for extrapulmonary
oxygenation in an animal study led to severe methaemoglobinaemia [18]. I.v. administration of HP solution
as unconventional therapy for AIDS resulted in severe
acute haemolysis [19].
In our case, we speculate that the full activation of
catalase on exposure to high concentrations of HP led
to the formation of oxygen bubbles in the blood lines,
but saturation of the finite amount of catalase left it
2357
incapable of preventing the oxidative effect and
haemolysis.
The variability in severity of methaemoglobinaemia
in our patients is largely explained by the different
duration of exposure to contaminated water (Table 2).
The presence of HP in the dialysis water was
apparently caused by insufficient washing out of the
system after disinfection. To the best of our knowledge,
there is only one report of the exposure of dialysis
patients to HP in the water [20]; this event was also
recorded by CDC [21]. In this case, three children
developed anaemia with an increased need for blood
transfusions during 11 days following disinfection of
the WTS with HP. As the test kit was not sufficiently
sensitive to identify the scant remains of HP in the
water, the washing of the system was inadequate.
A similar danger is posed by the peracetic acid–HP
mixtures (Dialox, Puristeril), which are still being used
to disinfect parts of dialysis systems [22].
This report highlights the critical need for stringent
monitoring for disinfectants in haemodialysis to insure
patient’s safety. Total chlorine analysis performed at
the onset of each dialysis session is an efficient tool for
determining the presence of oxidants in the dialysis
water, provided the test is performed accurately. Special
attention should be directed to the contact time between
the reactants and the water sample (at least 3 min) [6]
and the sensitivity of the method must be determined
before testing. The method used must be adequate to
measure low oxidant concentrations (0.1 p.p.m. total
chlorine). If the results for total chlorine are positive,
further analysis for HP residues must be performed
with peroxide test strips [23]. This method is based on
the oxygen transfer by peroxidase from the peroxide to
an organic redox indicator, which is then converted to a
blue-coloured product. The test is specific for HP and
has a sensitivity of 0.5–25 p.p.m.
When changes in the WTS are being considered, all
components of the system and their interaction should
first be evaluated. The clinical staff should be well
informed about the various breakdowns that can occur
in the water system and their clinical repercussions.
Ultimately, the responsibility for adequate water
quality and the patients’ well-being is in the hands of the
clinical staff.
Conflict of interest statement. None declared.
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Received for publication: 11.9.02
Accepted in revised form: 4.6.03