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Acute hemolysis due to naphthalene inhalation
904
The Journal o[ P E D I A T R I C S
Acute hemolysis due to
naphthaleneinhalation
Acute hemolytic episodes with the presence of Heinz bodies and fragmented erythrocytes occurred [ollowing inhalation of naphthalene in 21 newborn Greek in[ants. Glucose-6-phosphate dehydrogenase activity was deficient in the erythrocytes of 12 in[ants. Surprisingly, however, the activity of this enzyme was normal in the other 9.
Timos Valaes, M.D., D.C.H., Spyros A. Doxiadis, M.D., "x"
and Phaedon Fessas, M.D. A T I - I E N S~ GRF, E G E
T H E medical literature contains many reports of acute hemolysis caused by the ingestion of naphthalenC -7 and other reports have suggested that hemolysis may follow the absorption of naphthalene through the skin of infants, s-~~ The biochemical abnormalities characteristic of the primaquine sensitive individuals have been found in patients with hemolysis caused by naphthalene.7, 10-17 In the present work we describe cases of naphthalene-induced hemolysis with an important and hitherto unrecognized mode of entry of this substance into the body. In addition, some of our cases developed acute
From the Departments o[ Obstetrics and Gynaecology and o[ Clinical Therapeutics, University of Athens and the Paediatric Unit of "Aghia Sophia" Children's Hospital. This work was supported by a research grant [rom Pfizer-Hellas and at its later stages by the Association for the Aid o[ Crippled Children. ~'Address, Children's Hospital "'Aghia Sophia," Athens 609, Greece.
hemolysis following exposure to naphthalene in spite of normal G6PD activity in the erythrocytes. MATERIAL The material of the present work consists of 21 newborn or young infants investigated and treated in the neonatal department of the "Alexandra" Maternity Hospital, or in the Pediatric Unit of the Children's Hospital "Aghia Sophia," Athens. All the infants were seen at the stage of acute hemolysis and most of the survivors have been followed.
METHODS Standard hematologic methods were used throughout this study. Bilirubin was measured according to the method of Malloy and Evelyn. is Methemoglobin and methemalbumin were detected by the Hartridge reversion spectroscope. For the detection of Heinz bodies a drop of whole blood was mixed with a drop of a one per cent solution of methyl violet in normal saline. The mixture was left at room
Volume 63 Number 5
temperature for 15 minutes and dry smears were prepared and examined under oil immersion. Glucose-6-phosphate dehydrogenase of the red cells was assayed by the method of Motulsky and Campbell-Kraut? 9 Because variations in the normal decolorization times were noted when different batches of reagents were used, the results on abnormal samples have been expressed by timing the delay of the onset of decolorization in comparison to controls examined simultaneously. Males with G6PD deficiency, as a rule, did not show decolorization even after incubation of 3 hours. Heterozygotic females had decolorization times ranging from normal to a delay of 190 minutes. Thus in a female even normal decolorization time does not exclude the presence of the gene for G6PD deficiency and corroborative evidence was sought by studying members of the family. In the more recent cases the G6PD activity was quantitatively assessed, but as this test did not add materially to the information already obtained by the Motulsky and Campbell test the results are omitted for the sake of uniformity. RESULTS
Table I gives most of the relevant clinical and laboratory data. Representative case histories are presented in the Appendix. There is a definite preponderance of males - - 1 6 males to only 5 females. Most of the cases were seen during the autumn and winter months. This is explained by the fact that the source of naphthalene, in most of the cases, was a blanket or woolen clothes kept with naphthalene mothballs during the summer. The common element in the histories of the cases was a period of exposure to naphthalene inhalation. This exposure was either apparent on admission when, for instance, the admitting doctor noticed the strong smell of naphthalene of a blanket with which the infant was covered or was revealed only after direct interrogation of the parents. Absorption of the naphthalene through the skin is highly improbable. Only in two
Hemolysis due to naphthalene inhalation
9 05
of the infants was the source of naphthalene a cloth worn next to the skin. We were able to exclude all other known hemolytic agents. Neither the mothers nor the infants were receiving any drugs prior to the appearance of hemolysis and none of the nursing mothers had ingested vicia fava or naphthalene mothballs. On the basis of common features o u r cases can be divided into three groups. T h e first group includes Cases 1 to 6. In all these infants exposure to naphthalene vapors took place after the sixth day of life. Before this the babies had either no jaundice (Cases 1, 4, 5, and 6) or after a period of neonatal jaundice which had already subsided exposure to naphthalene took place and jaundice reappeared (Cases 2 and 3). The infants of this group were males with G6PD deficiency. The presence of hemolysis in these infants was evident. High indirect serum bilirubin values were found in all of them at a period when other causes of neonatal jaundice cease to operate. Definite anemia was present in all cases and was marked in four (Cases 2, 3, 4, and 6). Heinz bodies and fragmentation of the red cells were observed in all infants of this group. Cases 1 and 6 are the only ones in all our material in which hemoglobinuria and methemoglobinemia were evident. Two of the infants had definite kernicterus on admission. One of them died and the diagnosis was confirmed at postmortem examination. Three of the infants were treated by simple transfusion and one infant by exchange transfusion. The second group includes Cases 7 to 13. The sequence of events and the age at exposure to naphthalene are similar as in the first group. There is, however, an important difference as G6PD deficiency could not be demonstrated in any of the infants. The three males of the group were tested during the acute stage and at a later date and normal G6PD activity was found on both occasions. The father of Infant 8 showed no G6PD activity and the mother of Infant 9 had intermediate G6PD deficiency. Whether
906
Valaes, Doxiadis, and Fessas
November 1963
Table I
Exposure to naphthalene Day o[ li[e I Source o/ naphthalene 7-8 Blanket
Case 1
Sex M
Date o[ birth 29/5/1959
2 3 4 5 6 7 8 9 10 11
M M M M M M M M F F
3/10/1959 21/9/1961 28/11/1961 26/11/1962 14/12/1962 5/12/1959 2/9/1960 26/9/1961 29/10/1961 8/10/1962
12 13
F F
2/12/1962 6/3/1963
7 2-8
14 15 16 17 18 19 20 21
F M M M M M M M
17/2/1960 25/6/1961 5/9/1961 30/9/1961 29/12/1961 25/7/1962 5/10/1961 1l/3/1963
0-3 1-5 1-4 0-1 0-4 0-3 1-6 6
7 36-39 6-9 9 7 7-13 10-17 14 5-12 13
these findings are fortuitous or connected by some u n k n o w n mechanism with the hemolysis of the infants is at present a matter of speculation. We c a n n o t exclude the possibility that the four females (10, 11, 12, and 13) whose blood gave n o r m a l decolorization time are heterozygotes. I n this group again the evidence of acute hemolysis is u n d e n i a b l e b u t the hemoglobin a n d serum bilirubin values suggest that hemolysis was not as severe as in the first group. All the infants of the second group survived without sequelae. F o u r received an exchange transfusion, one a simple transfusion, a n d two no t r e a t m e n t at all. T h e third group consists of the r e m a i n i n g patients who were exposed to n a p h t h a l e n e soon after birth at a period w h e n hemolytic j a u n d i c e from other causes also occurs. Hemolytic disease of the n e w b o r n due to isoimmunization could be excluded in all. F o r evidence of acute hemolysis in these in-
Cloths in baby's room Blanket Blanket Infant's cloths Diapers Infant's clothes Blankets in mother's bed Clothes in infant's room Clothes in infant's room Clothes and carpet on corridor outside infant's room Clothes in infant's room Blanket and clothes in infant's room Blanket Infant's clothes Sheets in mother's bed Blanket Infant's clothes Sheets in infaht's cot Blanket Infant's clothes
Age on admission (day.F) 9
Serum bilirubin (mg./lO0 ml.) 14.0
10 42 9 10 10 13 18 18 12 14
43.2 5.5 55.0 24.4 12.8 16.5 20.2 17.0 28.8 26.0
8 8
24.0 29.4
3 5 4 3 4 3 6 6
37.7 28.5 46.0 20.2 38.6 41.6 40.9 34.0
rants we relied m a i n l y on the serum bilir u b i n values. I n this group the rise in serum bilirubin was far more r a p i d t h a n could have been expected if complete block of bilirubin excretion occurred with a n o r m a l rate of hemolysis. Some of the babies did not show a n y definite anemia. T h e poor excretion of bilirubin d u r i n g the first postn a t a l days created the need for t r e a t m e n t by exchange transfusion or caused death from kernicterus before the increased rate of hemolysis h a d time to affect the hemoglobin values. I n this third group not all of the infants of which blood films were examined showed Heinz bodies o r / a n d fragm e n t a t i o n of the red cells. Of the 7 males in this group 4 had no G 6 P D activity a n d one male h a d a n intermediate degree of deficiency as is usually seen in female heterozygotes. T h e other two males had n o r m a l G 6 P D activity. T h e female had intermediate G 6 P D activity. O f the eight infants of this group 2 sur-
Volume 63
Number 5
Hemolysis due to naphthalene inhalation
907
Laboratory findings on admission emoglobin (Gin.~100 Reticuloeytes
ml.)
[
(%)
Heinz bodies
(%.)
12.0
--
4.6 5.3 8.3 12.2 7.2 10.3 ll.5 9.1 9.6 14.8
6 14 7.5 2.2 6 7 5.5 3 -2.8
++ 70 20 4 15 70 70 40 No No
12.5 12.0
1.6 1.2
No 11
16.0 11.7 15.3 18.1 14.4
6.0 6.0 -3.3 7.2
12.5
4
14.8 17.2
5.4 3.2
-No -No 10 No 95 23
I00
Fragmented erythrocytes Treatment +++ Methylene blue Vitamin C Tranfusion +++ None ++++ Transfusion ++ None + Transfusion +++ Exchange transfusion + Exchange transfusion ++++ Transfusion ++ None ++ Exchange transfusion + Exchange transfusion
Outcome Survived
G6PD activity Deficient
Kernicterus-Died Survived Kernicterus-Survived Survived Survived Survived Survived Survived Survived Survived
Deficient Deficient Deficient Deficient Deficient Normal Normal Normal Normal Normal
+ +
None Survived Exchange transfusion Survived
Normal Normal
+
Exchange Exchange Exchange Exchange None Exchange Exchange Exchange
Intermediate Deficient Deficient Deficient Intermediate Deficient Normal Normal
No +
No + +
No
vived a n d have escaped central nervous system damage after exchange transfusion. Five infants were admitted with signs of kernicterus, b u t it was t h o u g h t that exchange transfusion might p r e v e n t further damage a n d they were treated with this procedure. O n e died shortly after a n d at autopsy the diagnosis of kernicterus was confirmed. Last, one baby h a d very definite signs of C.N.S. damage on admission a n d as his serum bilir u b i n was declining at this time n o treatm e n t was given. DISCUSSION I n the thirteen n e w b o r n infants of Groups 1 a n d 2 the occurrence of hemolysis after the first week, at a time when most other hemolytic factors cease to operate, and the certain exposure to n a p h t h a l e n e inhalation leave no doubt that this substance was the cause of hemolysis. I n the r e m a i n i n g cases ( G r o u p 3) both n a p h t h a l e n e i n h a l a t i o n a n d hemolysis, manifesting m a i n l y as severe
transfusion transfusion transfusion transfusion
Kernicterus-Died Survived Kernicterus-Survived Survived Kernicterus-Survived transfusion Kernicterus-Survived transfusion Kernieterus-Survived transfusion Kernicterus-Survived
n e o n a t a l jaundice, took place d u r i n g the first week of life; however hemolytic disease of the n e w b o r n due to Rhesus or A B O incompatibility could be excluded in all these infants. It is true that cases of severe neon a t a l j a u n d i c e have been described in infants deficient in G 6 P D in the absence of any known extrinsic hemolytic a g e n t ? 7, 20-27 I t could be argued, therefore, that in the babies of the third group there was no etiologic connection between n a p h t h a l e n e i n h a l a t i o n a n d n e o n a t a l jaundice. However, the indisputable hemolytic action of n a p h thalene i n h a l a t i o n in the babies of the first 2 groups makes it very likely that the same m e c h a n i s m also operated in the babies of the third group. T h e possibility of n a p h t h a l e n e acting through i n h a l a t i o n has scarcely been mentioned in the medical l i t e r a t u r e ? s, 29 I n the reported cases of hemolysis occurring after i n f a n c y the n a p h t h a l e n e h a d been ingested, while in n e w b o r n infants absorption from
908
Valaes, Doxiadis, and Fessas
November 1963
the skin had been considered responsible. However, our experience makes it likely that even in the latter cases naphthalene acted after inhalation. This route of entry is not restricted to the neonatal period. Younis and colleagues 3~ documented 6 cases of severe hemolysis (one of them fatal) in young recruits in a training camp where the men were covered with blankets strongly smelling of naphthalene. Isolated cases of hemolysis in infants through inhalation of naphthalene have also been recorded in the Greek literature.l?, 31 The collection of 21 cases within a fairly short time and by one team of workers shows that severe hemolysis caused by naphthalene inhalation is not very rare in infancy. It is known that the toxic agent responsible for hemolysis is not naphthalene but its metabolic products alpha- and betanaphthol and naphthoquinone. These substances require conjugation with glucuronic
- E2 0
o Exchange 9 No
o
K
15
_z
O
~Ko
m
Q
8 9149 O
O
~o
o L0
,,
K Kernicterus
o
o
TranSl.
,,
5
"T
I
I
I
,,~
~
I
8 6~
50 o K
E Z
40
+~
m
gE _.1
30
0
O
@
I[
20
bJ ul
10
0
O"
9
00
I
I
I
I
5
10
15
20
AGE
IN
I ,42
DAYS
Fig. 1. Serum bilirubin and hemoglobin values on adnfission.
acid for their excretionY 2 I t is reasonable, therefore, to assume that newborn infants, because of their limited conjugating capacity, are more susceptible to the toxic action of these substances. Moreover, the same limitation of conjugation i n the neonatal period increases the deleterious effects of hemolysis. Thus most of our cases developed severe icterus before severe anemia became manifest. In fact, the jaundice was the main and usually the only reason for sending the infant to the hospital. Infants exposed later, because of the better conjugating capacity of their liver, did not develop severe jaundice early in their illness and thus were sent to us at a time when anemia was already present (Fig. 1). It is important to note that of the twentyone infants of this series, 8 developed kernicterus, in 2 this was fatal. All eight were under 10 days of age and had serum bilirubin values above 30 rag. per 100 inl. In another 7 infants timely exchange transfusion probably prevented this development. This is the first time that hemolysis from naphthalene has been considered responsible for kernicterus. Finally the experience of the present series shows that kernicterus can occur even after the first week of life. This should lead to a reconsideration of the significance of the maturation of the bloodbrain barrier in the path 9 of kernicterus? 3 The present series shows that in most cases of naphthalene-induced hemolysis G6PD deficiency is the predisposing factor. However, in 9 babies with identical histories and findings the activity of this enzyme was normal. One can assume that in these infants either the amount of the toxic agent absorbed was so large as to affect even normal erythrocytes or that another, as yet undetected, abnormality of the red blood cells was present. The danger of naphthalene inhalation, particularly in the newborn period, should be recognized not only in the countries where G6PD deficiency is present but in all places where naphthalene is used. Such a possibility should be investigated in all infants with
Volume 63 Number 5
acute hemolysis by searching for a history of exposure to naphthalene and for the presence of fragmented erythrocytes and Heinz bodies in the blood. The present report is limited to the clinicai aspects of hemoIysis in infancy due to naphthalene inhalation. However, m a n y and complex problems are raised from our observations. The histories of m a n y of our cases lead us to think that even minimal exposure to naphthalene was enough to injure the red cells. The sequence of events in these infants was more like the explosive hemolysis in favlSrn than the gradual and dose-dependant hemolysis induced by primaquine. Another important question is whether this extreme sensitivity to naphthalene is present in all infants (and perhaps adults) with G6PD deficiency or is it limited to some of the racial groups with this deficiency as seems to be the case with neonatal jaundice resulting from this defect. The paucity of reports of similar cases from other countries would suggest that this might be true although at present two imponderables should be taken into consideration: 1. We have no information about the extent of the household use of naphthalene mothballs in other countries in comparison to Greece. 2. We cannot be sure that in the history taking of infants with unexplained hemolysis exposure to naphthalene inhalation was considered. Our experience with these cases has persuaded us that, at least in Greece, extreme sensitivity to naphthalene is present in some infants. With the consideration that vitamin K analogues are structurally related to the toxic metabolites of naphthalene, the possibility that some infants are sensitive to even small doses of these compounds seems logical. T h a t this might be so is also suggested by a number of cases of severe hemolysis following small doses of vitamin K analogues which we have observed. In spite of other studies a4 that 2.5 to 7.5 mg. of vitamin K analogues are safe for infants with G6PD deficiency, we cannot accept as proved the absolute safety of this amount for newborn
Hemolysis due to naphthalene inhalation
909
Fig. 2. Case 1. Fragmented and pyknotic erythrocytes. infants in Greece and perhaps for those of some other areas. SUMMARY
Acute hemolysis following, naphthalene inhaIatlon is described in 21 Greek newborn infants--16 were males and 5 females. Severe jaundice, anemia in some and methemoglobinemia and hemoglobirmria in 2 were the main clinical manifestations. Kernicterus developed in 8 infants. The presence of Heinz bodies and fragmentation of the red cells were prominent features in most of the cases. A deficiency of the red cell enzyme glucose-6-phosphate dehydrogenase was detected in 12 of the infants. In the remaining 9 infants the activity of this enzyme was found to be normal.
APPENDIX REPRESENTATIVE
CASE REPORTS
Case 1. A male infant was born in the "Alexandra" Maternity Hospital on M a y 29, 1959, with a birth weight of 4,200 grams. He was the first child of healthy parents. A maternal uncle had died at 9 years of age from favism. T h e newborn during his 7 day stay in "Alexandra" did not have jaundice. T h e day
9 10
Valaes, Doxiadis, and Fessas
after his discharge the mother noticed icterus and dark red urine staining the diapers. On admission (ninth day) he was found to be moderately jaundiced (serum bilirubin 14 mg. per 100 ml.) and cyanosed. Spectroscopically methemoglobin was detected. His hemoglobin was 12.0 Gm. per 100 ml., and multiple Heinz bodies were present in all the red cells which were also fragmented (Fig. 2). The blanket with which the infant was covered since going home had a strong smell of naphthalene. Because of the presence of methemoglobin it was decided to give the infant methylene blue. Inadvertently 1.5 ml. of a 5 per cent solution instead of the correct 1.5 ml. of 1 per cent solution was given intravenously. This was followed by extreme cyanosis and distress of the infant. The distress lasted for few hours but cyanosis did not disappear until 2 days later. Six hours after the administration of the methylene blue 500 mg. of ascorbic acid was given intramuscularly. The following day the hemolysis continued. Thus in the morning the hematocrit was 33 per cent while in the evening it was 20 per cent; the hemoglobin was 8.0 Gm. per 100 ml., and the white blood cell count 64,000 per cubic millimeter. All the red cells still contained Heinz bodies and were fragmented. A blood transfusion of 80 ml. compatible blood was given. On the eleventh day his hemoglobin was 7.8 Gm. per 100 ml. and Heinz bodies were present in 30 per cent of the red cells. The plasma contained large amounts of methemalbumin. He was given another transfusion of 90 ml. The urine on admission was of dark red color and was positive for hemoglobin. Following the administration of methylene blue the color of urine turned to green-blue. On the twelfth day it was realized that the infant was oliguric and uremic, the fluid intake was restricted, and he was given terramycin orally. During the oliguric stage which lasted for 3 days, the urine had a most bizzare greenish red color. Diuresis was followed by return of the normal color of urine and by the disappearance of jaundice. The infant was quite well to be discharged on
November 1963
the eighteenth day of life (9 days after admission). At the age of 1 year he was thriving and had no anemia. Motulsky's test. This showed that the father had normal decolorization, the mother, a delay of 10 minutes, and the patient (at the age of l year), no decolorization at 3 hours. Case 2. This male infant who weighed 3,650 grams at birth was delivered on Nov. 3, 1959, at a private nursing home. He was the first child of healthy parents. Jaundice appeared at 36 hours of life but during the 5 day stay of the infant in the nursing home it did not become deep enough for the attending pediatrician to ask for any laboratory investigation. O n the seventh day of life, according to the parents, the infant was .still jaundiced but active and feeding very well. On this day winter clothes kept with naphthalene mothballs were taken into the room where the infant was sleeping. Next day the jaundice was deeper. When admitted at 10 days of age he had definite signs of kernicterus, severe jaundice, hepatosplenomegaly, and anemia. The infant died on the fourteenth day. A postmortem examination confirmed the diagnosis of kernicterus. Laboratory results. The mother was Group A, Rhesus positive and the infant was Group O, Rhesus positive. T h e direct Coombs test was negative. T h e course of serum bilirubin values is shown in Fig. 3. Motulsky's test. The father and mother gave normal decolorization. T h e patient gave no decolorization at 3 hours. A male sibling born recently had normal G6PD activity and had no jaundice. Case 4. A male infant was born in a private nursing home on Nov. 28, 1961. The birth weight was 4,500 grams. He was the second child of healthy parents. The mother had an episode of jaundice during the spring when she was 15. T h e firstborn, also a male, never had jaundice. The infant was discharged home on the sixth day having no trace of jaundice. On the seventh day slight jaundice appeared. In the morning of the next day the jaundice was intense and in the evening opisthotonus and inability to
Volume
63
Number 5
Hemolysis due to naphthalene inhalation
1•
.= 5O E
9 11
TotQI BRirubin J Direct //
E _z "n IE
~20
I0
2
o
o
o
"d ~
J:
,g2
a. w->{ o E
IS
F i g . 3. C a s e 2. C o u r s e o f s e r u m bilirubin values.
I
I
I
8
9
10
1 I
AGE
11 IN DAYS
I
I
I
12
13
14
[ ~
Tot(][ Bilirubin
l
%
I'-
Direc,
..
~50
Do
E40 z
o
"G E
.,~ .
.
.
.
.
.
.
.
o x p o s u r e to napht h(:l[ene ( >
F i g . 4. C a s e 4. C o u r s e o f s e r u m bilirubin and hemoglobin values.
I 6
I 8
,0~o 5 i
I 10
I I 12 14 AGE IN DAYS
I 16
I 18
I 20
E,T.
~
2o-~
20
o
g
z o
o
~I0
10o o
"~" "~ I :E D
F i g . 5. C a s e 7. C o u r s e o f s e r u m bilirubin and hemoglobin values. E x c h a n g e t r a n s f u s i o n is represented by the arrow.
i
....
exposure to naphtha[ene
S.B.
Hb
2
5
l l l l
3
4
5
6
7 ~
13 14 I"5
A G E IN DAYS
I'6 I'7 I'8
9 12
Valaes, Doxiadis, and Fessas
November 1963
nurse appeared. He was admitted in the "Alexandra" on the ninth day with very marked jaundice and unmistakable signs of kernicterus. The blanket with which the infant was covered since his return home smelled strongly of naphthalene. Examination at the age of 11 months showed definite sequelae of kernicterus. The course of serum bilirubin and hemoglobin is depicted in Fig. 4. Motulsky's test. The father showed normal decolorization; the mother, a delay of 30 to 40 minutes. The patient and his elder brother showed no decolorization at 3 hours. Case 6. This male infant was born at home Dec. 14, 1962. He was the third child of healthy parents. The previous two males had slight neonatal jaundice. No jaundice appeared during the first week of life. On the seventh day 3 or 4 diapers strongly smelling of naphthalene were used. The smell was quite strong as the father commented to the mother that it might " h a r m the baby." In the morning of the eighth day the infant was "blue." On the ninth day the infant was both "yellow and blue." Finally o n the tenth day the jaundice was intense and the infant's urine had a dark red color. On admission in the evening of the same day the infant was much distressed, cyanotic, anemic, and jaundiced. The spleen was palpable 2 cm. below the costal margin. Urine collected on admission had a dark red color and was positive for hemoglobin. He was placed in an oxygen tent and a transfusion of sedimented red
cells was started immediately. Three hours after admission an exchange transfusion of 500 ml. was started and was followed by a transfusion of 60 ml. of sedimented red cells. An infusion of 5 per cent glucose was given for the next 18 hours. Next morning his general condition was much improved, he was free from cyanosis, and had only moderate jaundice. On the twelfth day only a trace of jaundice was present and the spleen was no more palpable. He was discharged on the fifteenth day. Urine was collected for the whole 24 hours following his admission. There was a dramatic change in color following the exchange transfusion and 12 hours after the exchange the urine contained no hemoglobin. Motulsky's test. Mother and father had normal decolorization. The patient and his two brothers showed no decolorization at 3 hours.
Comment. This infant presented all the clinical and laboratory manifestations of acute intravascular hemolysis plus methemoglobinemia. Apart from the danger of hyperbilirubinemia his life was endangered from a much reduced oxygen-carrying capacity (anemia plus methemoglobinemia). Moreover 60 per cent of his red cells had morphologic evidence of impeding lysis. It was feared that this amount of hemoglobin when released would produce blockage of renal tubules. By choosing exchange transfusion instead of a simple transfusion we were able to correct rapidly the abnormal state.
Table II. Laboratory investigations Hematocrit
Sample
Serum bili- Hemoglobin Methemorubin (rag.~ (Gm./lO0 globin (%) 100 ml.) ml.)
O n admission ( 10th day, 19.00 hr.)
12.8
7.2
23
Pre-exchange (22.25 hr.) (following transfusion of packed cells)
25.6
10.2
14
Post-exchange
19.0
14.2
llth
--
14.8
day
Packed red cells
Column of stromata
. Heinz bodies (%)
Pyknocytes an~d [ragmented R.B.C.s
(%) 60
23
5
15
--
0.16
46
0
--
--
--
46
0
rare
rare
Volume 63 Number 5
Case 7. This infant, a male, was born in "Alexandra" Dec. 5, 1959. Birth weight was 3,200 grams. He was the second child of healthy parents. Jaundice was noticed before the end of the first 24 hours of life. The course of serum bilirubin is depicted in Fig. 5. The mother was Group O Rhesus positive while the father and the infant were Group B Rhesus positive. T h e direct Coombs test was negative. A diagnosis of O-B hemolytic disease was made and this was further substantiated when on the eighteenth day postpartum anti-B hemolysins were detected in the mother up to a dilution of 1:8 (anti-A hemolysins were also present up to 1:4). The infant was discharged on the seventh day still having slight jaundice. H e was fully breast fed and mother and infant received no drugs. The infant was brought to the outpatient department on the thirteenth day of life on account of increasing jaundice for the past 48 hours. Since the discharge from the Maternity Hospital, the infant had been dressed with woolen clothes which were previously kept with naphthalene mothballs and which at the time of his examination in outpatient were still strongly smelling of naphthalene. The infant was given an exchange transfusion on the fourteenth day of his life. The exchange transfusion was performed through the umbilical vein exposed above the umbilicus. Group B Rhesus positive blood was used after removal of the supernatant plasma; 290 ml. were introduced and 250 ml. removed. Exchange transfusion was used in preference to simple transfusion as the removal of bilirubin and damaged red cells was considered essential in addition to the correction of anemia. In the pre-exchange specimen 70 per cent of the red cells contained Heinz bodies (Fig. 6). Repeated testing of urine failed to detect any hemoglobin. The infant was discharged on the twentythird day of life free of jaundice. Follow-up to the age of 3 years failed to reveal any blood abnormality apart from moderate anemia which had responded to iron at the age of 11 months.
Hemolysis due to naphthalene inhalation
9 13
Fig. 6. Case 7. Coarse Heinz bodies in the red ceils.
Motulsky's test. The infant and both his parents gave normal decolorization. Also no abnormality was detected by hemoglobin electrophoresis in the patient and his parents. Case 20. On Oct. 5, t961, a male infant was born in a private nursing home in Athens. Birth weight was 3,800 grams. He was the second child of healthy parents. The first child, a girl, never had jaundice. Jaundice was noticed on the third day and gradually became deeper. On the sixth day the infant stopped sucking and was irritable. On admission the same day he had marked jaundice, no hepatosplenomegaly, and equivocal signs of kernicterus. The infant's blanket was smelling of naphthalene. He was treated with two exchange transfusions on the sixth and seventh day of life following which his jaundice gradually subsided. Laboratory investigations. T h e mother and infant were Group A Rhesus positive. The direct Coombs test was negative. O n the sixth day at 12:30 noon the serum bilirubin was 33.1 nag. per 100 ml. At 3 P.M. (preexchange) serum bilirubin was 40.9 mg. per 100 ml. and hemoglobin 14.8 Gin. per 100 mL Reticulocytes were 5.4 per cent. Heinz bodies were present in 95 per cent of the red ceils. The smear showed marked poikilocyto-
9 14
Valaes, Doxiadis, and Fessas
sis, a n d fragmentation of the red cells, anisocytosis, polychromasia, a n d spherocytosis were also present. Eighteen hours following the first exchange transfusion his serum b i l i r u b i n was 35.8 mg. per 100 ml. a n d a second exchange was given (seventh day). Motulsky's test. T h e infant, the parents, the m a t e r n a l grandparents, a n d two brothers of the mother showed n o r m a l decolorization.
Copies of the remaining case reports may be obtained by writing to the authors.
REFERENCES 1. Zuelzer, W. W., and Apt, L.: Acute hemolytic anemia due to naphthalene poisoning, J. A. M. A. 141" 185, 1949. 2. Newns, G.: Moth-ball anaemia, Lancet 2: 964, 1949. 3. Mackell, J. V., Rieders, F., Brleger, I-I., and Bauer, E, L.: Acute haemolytic anemia due to ingestion of naphthalene moth-balls, Pediatrics 7: 722, 1951. 4. MacGregor, R. R.: Naphthalene poisoning from the ingestion of moth-balls, Canad. M. A. J. 70: 313, 1954. 5. Chusld, E., and Fried, C. T.: Acute hemolytic anemia due to naphthalene ingestion, A. M. A. Am. J. Dis. Child. 89: 612, 1955. 6. Gidron, E., and Leurer, J.: Naphthalene poisoning, Lancet 1: 228, 1956. 7. Sansone, G., and Segni, G: Difetto biochimico eritrocitario a carattere genetico in un bambino con anemia emolitiea da naftalina, Boll. Soc. ital. biol. sper. 34: 516, 1958. 8. Schafer, W. B.: Acute hemolytic anemia related to naphthalene. Report of a case in a newborn infant, Pediatrics 7: 172, 1951. 9. Cock, T. C.: Acute hemolytic anemia in the neonatal period, A. M. A. Am. J. Dis. Child. 94: 77, 1957. 10. Dawson, J. P., Thayer, W. W., and Desforges, J.F.: Acute hemolytic anemia in the newborn infant due to naphthalene poisoning, Blood 13: 1113, 1958. 11. Zinkham, W. H., and Childs, B.: Effect of naphthalene derivatives on glutathione metabolism of erythrocytes from patients with naphthalene hemolytic anemia, J. Clin. Invest. 36: 938, 1957. 12. Zinkham, W. It., and CMlds, B.: The effect of vitamin K and naphthalene metabolites on the glutathione metabolism of erythroeytes from normal newborns and patients with naphthalene hemolytic anemia, A. M. A. Am. J. Dis. Child. 94: 120, 1957.
November 1963
13. Zinkham, W. If., and Childs, B.: A defect of glutathione metabolism in erythrocytes from patients with naphthalene induced hemolytic anemia, Pediatrics 22: 461, 1958. 14. Gross, R. T., Marks, P. A., Ifurwitz, R. E., Beasley, J., and Sui, M. Studies on the hereditary enzymatic defect in certain drug induced hemolytic anemias, Society for Pediatric Research. Twenty-eighth Meeting, 1958. 15. McGovern, J. J., Isselbacher, K., Rose, P. S., and Grossman, M. S.: Observations on the glutathione stability of red blood cells, Society for Pediatric Research. Twenty-eighth Meeting, 1958. 16. Sansone, G.: L'anemia emolitica acuta da ingestione accidentale di naftalina nel bambino. Contributo clinico e considerazioni patogenetiche, I-Iaemat. Lat. 1: 45, 1958. 17. Zannos, L.: Biochemical study of favism and hemolysis induced by drug and chemicals in Greece, Thesis, Athens, 1961. 18. Malloy, It. T., and Evelyn, K. A.: The determination of bilirubin with the photoelectric colorimeter, J. Biol. Chem. 119: 481, 1937. 19. Motulsky, A. G., and Campbell-Kraut J . M . : Proceedings of the conference on genetic polymorphlsms and geographic variations in disease, New York and London, 1961, Grune & Stratton Company, Inc. 20. Doxiadis, S., Valaes, T.: The problem of kernicterus in Greece, Proc. M. Soe., Athens, 1960, p. 507. 21. Doxiadis, S., Fessas, Ph., and Valaes, T.: Erythrocyte enzyme deficiency in unexplained kernicterus, Lancet 2: 44, 1960. 22. Doxiadis, S., Fessas, Ph., and Valaes, T., and Mastrokalos, N.: Glucose-6-phosphate dehydrogenase deficiency: A new aetiological factor of severe neonatal jaundice, Lancet 2: 297, 1961. 23. Panizon, F.: L'ictfre grave du nouveau-n6 associ6 ~ une defieience en glucose-6-phosphate dehydrogenase, Biologia Neonatorum 2: 167, 1960. 24. Weatherall D. J.: Enzyme deficiency in haemolytic disease of the newborn, Lancet 2: 835, 1960. 25. Gaburro, D., Volpatos, S., Gianqinto, M.: Ict~re nucleaire du nouveau-n6 par d6faut de la G-6-PD, Semaine H6p. (Ann. Ped.) 37: 69, 1961. 26. Valaes, T., Fessas,, Ph., and Doxiadis, S. A . : Kernicterus in full-term infants without isoimmunization, Proc, Roy. Soc. Med. 54: 331, 1961. 27. Fessas, Ph., Valaes, T., and Doxiadis, S. A.: Study of new aetiological factors of icterus gravis neonatorum, Proc. Med. Soc. Athens, 1961, p. 117. 28. Fabian, P.: Toxicka hemolyticka anemia u novoredence po otrove naptalinovymi parami, Clesk. pediat. 12: 833, 1957. 29. Betke, K., and Schall, I-I.: Akute h~imolytische Innenk6rperan~imie mit Meth~imoglobin~imie und Meth~imalbuminSmie bei einem Neuge-
Volume 63 Number 5
borenen, Wahrscheimlich bedingt durch Naphthalin, Zeitschr. Kinderh. 81" 379, 1958. 30. Younis, D., Veltsos, A., Platakos, Th., and Vafiadis, S.: Clinical and laboratory study of cases of naphthalene intoxication, Arch. Inst. Pasteur Hellenique, 1957, p. 521. 31. Kassimos, Ch., Vlachos-Anastasea, K., Koutsoukos K., and Ginieris, D.: Acute haemolytie anaemia in the neonatal period due to naphthalene, Arch. Paediat. Clinic University of Athens 5: 313, 1958.
Hemolysis due to naphthalene inhalation
9 15
32. Boyland, E., and Wihshire, G. H.: The metabolism of polycyclic compounds, Biochem. J. 53: 636, 1953. 33. Valaes, T.: Red-cell enzymes and severe neonatal jaundice, Cerebral palsy Bull. 3: 431, 1961. 34. Zinkham, W. H.: Peripheral blood and bilirubin values in normal full-term primaquinesensitive Negro infants: Effect of vitamin K, Pediatrics 31: 983, 1903.
The Journal o[ P E D I A T R I C S
Acute hemolysis due to
naphthaleneinhalation
Acute hemolytic episodes with the presence of Heinz bodies and fragmented erythrocytes occurred [ollowing inhalation of naphthalene in 21 newborn Greek in[ants. Glucose-6-phosphate dehydrogenase activity was deficient in the erythrocytes of 12 in[ants. Surprisingly, however, the activity of this enzyme was normal in the other 9.
Timos Valaes, M.D., D.C.H., Spyros A. Doxiadis, M.D., "x"
and Phaedon Fessas, M.D. A T I - I E N S~ GRF, E G E
T H E medical literature contains many reports of acute hemolysis caused by the ingestion of naphthalenC -7 and other reports have suggested that hemolysis may follow the absorption of naphthalene through the skin of infants, s-~~ The biochemical abnormalities characteristic of the primaquine sensitive individuals have been found in patients with hemolysis caused by naphthalene.7, 10-17 In the present work we describe cases of naphthalene-induced hemolysis with an important and hitherto unrecognized mode of entry of this substance into the body. In addition, some of our cases developed acute
From the Departments o[ Obstetrics and Gynaecology and o[ Clinical Therapeutics, University of Athens and the Paediatric Unit of "Aghia Sophia" Children's Hospital. This work was supported by a research grant [rom Pfizer-Hellas and at its later stages by the Association for the Aid o[ Crippled Children. ~'Address, Children's Hospital "'Aghia Sophia," Athens 609, Greece.
hemolysis following exposure to naphthalene in spite of normal G6PD activity in the erythrocytes. MATERIAL The material of the present work consists of 21 newborn or young infants investigated and treated in the neonatal department of the "Alexandra" Maternity Hospital, or in the Pediatric Unit of the Children's Hospital "Aghia Sophia," Athens. All the infants were seen at the stage of acute hemolysis and most of the survivors have been followed.
METHODS Standard hematologic methods were used throughout this study. Bilirubin was measured according to the method of Malloy and Evelyn. is Methemoglobin and methemalbumin were detected by the Hartridge reversion spectroscope. For the detection of Heinz bodies a drop of whole blood was mixed with a drop of a one per cent solution of methyl violet in normal saline. The mixture was left at room
Volume 63 Number 5
temperature for 15 minutes and dry smears were prepared and examined under oil immersion. Glucose-6-phosphate dehydrogenase of the red cells was assayed by the method of Motulsky and Campbell-Kraut? 9 Because variations in the normal decolorization times were noted when different batches of reagents were used, the results on abnormal samples have been expressed by timing the delay of the onset of decolorization in comparison to controls examined simultaneously. Males with G6PD deficiency, as a rule, did not show decolorization even after incubation of 3 hours. Heterozygotic females had decolorization times ranging from normal to a delay of 190 minutes. Thus in a female even normal decolorization time does not exclude the presence of the gene for G6PD deficiency and corroborative evidence was sought by studying members of the family. In the more recent cases the G6PD activity was quantitatively assessed, but as this test did not add materially to the information already obtained by the Motulsky and Campbell test the results are omitted for the sake of uniformity. RESULTS
Table I gives most of the relevant clinical and laboratory data. Representative case histories are presented in the Appendix. There is a definite preponderance of males - - 1 6 males to only 5 females. Most of the cases were seen during the autumn and winter months. This is explained by the fact that the source of naphthalene, in most of the cases, was a blanket or woolen clothes kept with naphthalene mothballs during the summer. The common element in the histories of the cases was a period of exposure to naphthalene inhalation. This exposure was either apparent on admission when, for instance, the admitting doctor noticed the strong smell of naphthalene of a blanket with which the infant was covered or was revealed only after direct interrogation of the parents. Absorption of the naphthalene through the skin is highly improbable. Only in two
Hemolysis due to naphthalene inhalation
9 05
of the infants was the source of naphthalene a cloth worn next to the skin. We were able to exclude all other known hemolytic agents. Neither the mothers nor the infants were receiving any drugs prior to the appearance of hemolysis and none of the nursing mothers had ingested vicia fava or naphthalene mothballs. On the basis of common features o u r cases can be divided into three groups. T h e first group includes Cases 1 to 6. In all these infants exposure to naphthalene vapors took place after the sixth day of life. Before this the babies had either no jaundice (Cases 1, 4, 5, and 6) or after a period of neonatal jaundice which had already subsided exposure to naphthalene took place and jaundice reappeared (Cases 2 and 3). The infants of this group were males with G6PD deficiency. The presence of hemolysis in these infants was evident. High indirect serum bilirubin values were found in all of them at a period when other causes of neonatal jaundice cease to operate. Definite anemia was present in all cases and was marked in four (Cases 2, 3, 4, and 6). Heinz bodies and fragmentation of the red cells were observed in all infants of this group. Cases 1 and 6 are the only ones in all our material in which hemoglobinuria and methemoglobinemia were evident. Two of the infants had definite kernicterus on admission. One of them died and the diagnosis was confirmed at postmortem examination. Three of the infants were treated by simple transfusion and one infant by exchange transfusion. The second group includes Cases 7 to 13. The sequence of events and the age at exposure to naphthalene are similar as in the first group. There is, however, an important difference as G6PD deficiency could not be demonstrated in any of the infants. The three males of the group were tested during the acute stage and at a later date and normal G6PD activity was found on both occasions. The father of Infant 8 showed no G6PD activity and the mother of Infant 9 had intermediate G6PD deficiency. Whether
906
Valaes, Doxiadis, and Fessas
November 1963
Table I
Exposure to naphthalene Day o[ li[e I Source o/ naphthalene 7-8 Blanket
Case 1
Sex M
Date o[ birth 29/5/1959
2 3 4 5 6 7 8 9 10 11
M M M M M M M M F F
3/10/1959 21/9/1961 28/11/1961 26/11/1962 14/12/1962 5/12/1959 2/9/1960 26/9/1961 29/10/1961 8/10/1962
12 13
F F
2/12/1962 6/3/1963
7 2-8
14 15 16 17 18 19 20 21
F M M M M M M M
17/2/1960 25/6/1961 5/9/1961 30/9/1961 29/12/1961 25/7/1962 5/10/1961 1l/3/1963
0-3 1-5 1-4 0-1 0-4 0-3 1-6 6
7 36-39 6-9 9 7 7-13 10-17 14 5-12 13
these findings are fortuitous or connected by some u n k n o w n mechanism with the hemolysis of the infants is at present a matter of speculation. We c a n n o t exclude the possibility that the four females (10, 11, 12, and 13) whose blood gave n o r m a l decolorization time are heterozygotes. I n this group again the evidence of acute hemolysis is u n d e n i a b l e b u t the hemoglobin a n d serum bilirubin values suggest that hemolysis was not as severe as in the first group. All the infants of the second group survived without sequelae. F o u r received an exchange transfusion, one a simple transfusion, a n d two no t r e a t m e n t at all. T h e third group consists of the r e m a i n i n g patients who were exposed to n a p h t h a l e n e soon after birth at a period w h e n hemolytic j a u n d i c e from other causes also occurs. Hemolytic disease of the n e w b o r n due to isoimmunization could be excluded in all. F o r evidence of acute hemolysis in these in-
Cloths in baby's room Blanket Blanket Infant's cloths Diapers Infant's clothes Blankets in mother's bed Clothes in infant's room Clothes in infant's room Clothes and carpet on corridor outside infant's room Clothes in infant's room Blanket and clothes in infant's room Blanket Infant's clothes Sheets in mother's bed Blanket Infant's clothes Sheets in infaht's cot Blanket Infant's clothes
Age on admission (day.F) 9
Serum bilirubin (mg./lO0 ml.) 14.0
10 42 9 10 10 13 18 18 12 14
43.2 5.5 55.0 24.4 12.8 16.5 20.2 17.0 28.8 26.0
8 8
24.0 29.4
3 5 4 3 4 3 6 6
37.7 28.5 46.0 20.2 38.6 41.6 40.9 34.0
rants we relied m a i n l y on the serum bilir u b i n values. I n this group the rise in serum bilirubin was far more r a p i d t h a n could have been expected if complete block of bilirubin excretion occurred with a n o r m a l rate of hemolysis. Some of the babies did not show a n y definite anemia. T h e poor excretion of bilirubin d u r i n g the first postn a t a l days created the need for t r e a t m e n t by exchange transfusion or caused death from kernicterus before the increased rate of hemolysis h a d time to affect the hemoglobin values. I n this third group not all of the infants of which blood films were examined showed Heinz bodies o r / a n d fragm e n t a t i o n of the red cells. Of the 7 males in this group 4 had no G 6 P D activity a n d one male h a d a n intermediate degree of deficiency as is usually seen in female heterozygotes. T h e other two males had n o r m a l G 6 P D activity. T h e female had intermediate G 6 P D activity. O f the eight infants of this group 2 sur-
Volume 63
Number 5
Hemolysis due to naphthalene inhalation
907
Laboratory findings on admission emoglobin (Gin.~100 Reticuloeytes
ml.)
[
(%)
Heinz bodies
(%.)
12.0
--
4.6 5.3 8.3 12.2 7.2 10.3 ll.5 9.1 9.6 14.8
6 14 7.5 2.2 6 7 5.5 3 -2.8
++ 70 20 4 15 70 70 40 No No
12.5 12.0
1.6 1.2
No 11
16.0 11.7 15.3 18.1 14.4
6.0 6.0 -3.3 7.2
12.5
4
14.8 17.2
5.4 3.2
-No -No 10 No 95 23
I00
Fragmented erythrocytes Treatment +++ Methylene blue Vitamin C Tranfusion +++ None ++++ Transfusion ++ None + Transfusion +++ Exchange transfusion + Exchange transfusion ++++ Transfusion ++ None ++ Exchange transfusion + Exchange transfusion
Outcome Survived
G6PD activity Deficient
Kernicterus-Died Survived Kernicterus-Survived Survived Survived Survived Survived Survived Survived Survived
Deficient Deficient Deficient Deficient Deficient Normal Normal Normal Normal Normal
+ +
None Survived Exchange transfusion Survived
Normal Normal
+
Exchange Exchange Exchange Exchange None Exchange Exchange Exchange
Intermediate Deficient Deficient Deficient Intermediate Deficient Normal Normal
No +
No + +
No
vived a n d have escaped central nervous system damage after exchange transfusion. Five infants were admitted with signs of kernicterus, b u t it was t h o u g h t that exchange transfusion might p r e v e n t further damage a n d they were treated with this procedure. O n e died shortly after a n d at autopsy the diagnosis of kernicterus was confirmed. Last, one baby h a d very definite signs of C.N.S. damage on admission a n d as his serum bilir u b i n was declining at this time n o treatm e n t was given. DISCUSSION I n the thirteen n e w b o r n infants of Groups 1 a n d 2 the occurrence of hemolysis after the first week, at a time when most other hemolytic factors cease to operate, and the certain exposure to n a p h t h a l e n e inhalation leave no doubt that this substance was the cause of hemolysis. I n the r e m a i n i n g cases ( G r o u p 3) both n a p h t h a l e n e i n h a l a t i o n a n d hemolysis, manifesting m a i n l y as severe
transfusion transfusion transfusion transfusion
Kernicterus-Died Survived Kernicterus-Survived Survived Kernicterus-Survived transfusion Kernicterus-Survived transfusion Kernieterus-Survived transfusion Kernicterus-Survived
n e o n a t a l jaundice, took place d u r i n g the first week of life; however hemolytic disease of the n e w b o r n due to Rhesus or A B O incompatibility could be excluded in all these infants. It is true that cases of severe neon a t a l j a u n d i c e have been described in infants deficient in G 6 P D in the absence of any known extrinsic hemolytic a g e n t ? 7, 20-27 I t could be argued, therefore, that in the babies of the third group there was no etiologic connection between n a p h t h a l e n e i n h a l a t i o n a n d n e o n a t a l jaundice. However, the indisputable hemolytic action of n a p h thalene i n h a l a t i o n in the babies of the first 2 groups makes it very likely that the same m e c h a n i s m also operated in the babies of the third group. T h e possibility of n a p h t h a l e n e acting through i n h a l a t i o n has scarcely been mentioned in the medical l i t e r a t u r e ? s, 29 I n the reported cases of hemolysis occurring after i n f a n c y the n a p h t h a l e n e h a d been ingested, while in n e w b o r n infants absorption from
908
Valaes, Doxiadis, and Fessas
November 1963
the skin had been considered responsible. However, our experience makes it likely that even in the latter cases naphthalene acted after inhalation. This route of entry is not restricted to the neonatal period. Younis and colleagues 3~ documented 6 cases of severe hemolysis (one of them fatal) in young recruits in a training camp where the men were covered with blankets strongly smelling of naphthalene. Isolated cases of hemolysis in infants through inhalation of naphthalene have also been recorded in the Greek literature.l?, 31 The collection of 21 cases within a fairly short time and by one team of workers shows that severe hemolysis caused by naphthalene inhalation is not very rare in infancy. It is known that the toxic agent responsible for hemolysis is not naphthalene but its metabolic products alpha- and betanaphthol and naphthoquinone. These substances require conjugation with glucuronic
- E2 0
o Exchange 9 No
o
K
15
_z
O
~Ko
m
Q
8 9149 O
O
~o
o L0
,,
K Kernicterus
o
o
TranSl.
,,
5
"T
I
I
I
,,~
~
I
8 6~
50 o K
E Z
40
+~
m
gE _.1
30
0
O
@
I[
20
bJ ul
10
0
O"
9
00
I
I
I
I
5
10
15
20
AGE
IN
I ,42
DAYS
Fig. 1. Serum bilirubin and hemoglobin values on adnfission.
acid for their excretionY 2 I t is reasonable, therefore, to assume that newborn infants, because of their limited conjugating capacity, are more susceptible to the toxic action of these substances. Moreover, the same limitation of conjugation i n the neonatal period increases the deleterious effects of hemolysis. Thus most of our cases developed severe icterus before severe anemia became manifest. In fact, the jaundice was the main and usually the only reason for sending the infant to the hospital. Infants exposed later, because of the better conjugating capacity of their liver, did not develop severe jaundice early in their illness and thus were sent to us at a time when anemia was already present (Fig. 1). It is important to note that of the twentyone infants of this series, 8 developed kernicterus, in 2 this was fatal. All eight were under 10 days of age and had serum bilirubin values above 30 rag. per 100 inl. In another 7 infants timely exchange transfusion probably prevented this development. This is the first time that hemolysis from naphthalene has been considered responsible for kernicterus. Finally the experience of the present series shows that kernicterus can occur even after the first week of life. This should lead to a reconsideration of the significance of the maturation of the bloodbrain barrier in the path 9 of kernicterus? 3 The present series shows that in most cases of naphthalene-induced hemolysis G6PD deficiency is the predisposing factor. However, in 9 babies with identical histories and findings the activity of this enzyme was normal. One can assume that in these infants either the amount of the toxic agent absorbed was so large as to affect even normal erythrocytes or that another, as yet undetected, abnormality of the red blood cells was present. The danger of naphthalene inhalation, particularly in the newborn period, should be recognized not only in the countries where G6PD deficiency is present but in all places where naphthalene is used. Such a possibility should be investigated in all infants with
Volume 63 Number 5
acute hemolysis by searching for a history of exposure to naphthalene and for the presence of fragmented erythrocytes and Heinz bodies in the blood. The present report is limited to the clinicai aspects of hemoIysis in infancy due to naphthalene inhalation. However, m a n y and complex problems are raised from our observations. The histories of m a n y of our cases lead us to think that even minimal exposure to naphthalene was enough to injure the red cells. The sequence of events in these infants was more like the explosive hemolysis in favlSrn than the gradual and dose-dependant hemolysis induced by primaquine. Another important question is whether this extreme sensitivity to naphthalene is present in all infants (and perhaps adults) with G6PD deficiency or is it limited to some of the racial groups with this deficiency as seems to be the case with neonatal jaundice resulting from this defect. The paucity of reports of similar cases from other countries would suggest that this might be true although at present two imponderables should be taken into consideration: 1. We have no information about the extent of the household use of naphthalene mothballs in other countries in comparison to Greece. 2. We cannot be sure that in the history taking of infants with unexplained hemolysis exposure to naphthalene inhalation was considered. Our experience with these cases has persuaded us that, at least in Greece, extreme sensitivity to naphthalene is present in some infants. With the consideration that vitamin K analogues are structurally related to the toxic metabolites of naphthalene, the possibility that some infants are sensitive to even small doses of these compounds seems logical. T h a t this might be so is also suggested by a number of cases of severe hemolysis following small doses of vitamin K analogues which we have observed. In spite of other studies a4 that 2.5 to 7.5 mg. of vitamin K analogues are safe for infants with G6PD deficiency, we cannot accept as proved the absolute safety of this amount for newborn
Hemolysis due to naphthalene inhalation
909
Fig. 2. Case 1. Fragmented and pyknotic erythrocytes. infants in Greece and perhaps for those of some other areas. SUMMARY
Acute hemolysis following, naphthalene inhaIatlon is described in 21 Greek newborn infants--16 were males and 5 females. Severe jaundice, anemia in some and methemoglobinemia and hemoglobirmria in 2 were the main clinical manifestations. Kernicterus developed in 8 infants. The presence of Heinz bodies and fragmentation of the red cells were prominent features in most of the cases. A deficiency of the red cell enzyme glucose-6-phosphate dehydrogenase was detected in 12 of the infants. In the remaining 9 infants the activity of this enzyme was found to be normal.
APPENDIX REPRESENTATIVE
CASE REPORTS
Case 1. A male infant was born in the "Alexandra" Maternity Hospital on M a y 29, 1959, with a birth weight of 4,200 grams. He was the first child of healthy parents. A maternal uncle had died at 9 years of age from favism. T h e newborn during his 7 day stay in "Alexandra" did not have jaundice. T h e day
9 10
Valaes, Doxiadis, and Fessas
after his discharge the mother noticed icterus and dark red urine staining the diapers. On admission (ninth day) he was found to be moderately jaundiced (serum bilirubin 14 mg. per 100 ml.) and cyanosed. Spectroscopically methemoglobin was detected. His hemoglobin was 12.0 Gm. per 100 ml., and multiple Heinz bodies were present in all the red cells which were also fragmented (Fig. 2). The blanket with which the infant was covered since going home had a strong smell of naphthalene. Because of the presence of methemoglobin it was decided to give the infant methylene blue. Inadvertently 1.5 ml. of a 5 per cent solution instead of the correct 1.5 ml. of 1 per cent solution was given intravenously. This was followed by extreme cyanosis and distress of the infant. The distress lasted for few hours but cyanosis did not disappear until 2 days later. Six hours after the administration of the methylene blue 500 mg. of ascorbic acid was given intramuscularly. The following day the hemolysis continued. Thus in the morning the hematocrit was 33 per cent while in the evening it was 20 per cent; the hemoglobin was 8.0 Gm. per 100 ml., and the white blood cell count 64,000 per cubic millimeter. All the red cells still contained Heinz bodies and were fragmented. A blood transfusion of 80 ml. compatible blood was given. On the eleventh day his hemoglobin was 7.8 Gm. per 100 ml. and Heinz bodies were present in 30 per cent of the red cells. The plasma contained large amounts of methemalbumin. He was given another transfusion of 90 ml. The urine on admission was of dark red color and was positive for hemoglobin. Following the administration of methylene blue the color of urine turned to green-blue. On the twelfth day it was realized that the infant was oliguric and uremic, the fluid intake was restricted, and he was given terramycin orally. During the oliguric stage which lasted for 3 days, the urine had a most bizzare greenish red color. Diuresis was followed by return of the normal color of urine and by the disappearance of jaundice. The infant was quite well to be discharged on
November 1963
the eighteenth day of life (9 days after admission). At the age of 1 year he was thriving and had no anemia. Motulsky's test. This showed that the father had normal decolorization, the mother, a delay of 10 minutes, and the patient (at the age of l year), no decolorization at 3 hours. Case 2. This male infant who weighed 3,650 grams at birth was delivered on Nov. 3, 1959, at a private nursing home. He was the first child of healthy parents. Jaundice appeared at 36 hours of life but during the 5 day stay of the infant in the nursing home it did not become deep enough for the attending pediatrician to ask for any laboratory investigation. O n the seventh day of life, according to the parents, the infant was .still jaundiced but active and feeding very well. On this day winter clothes kept with naphthalene mothballs were taken into the room where the infant was sleeping. Next day the jaundice was deeper. When admitted at 10 days of age he had definite signs of kernicterus, severe jaundice, hepatosplenomegaly, and anemia. The infant died on the fourteenth day. A postmortem examination confirmed the diagnosis of kernicterus. Laboratory results. The mother was Group A, Rhesus positive and the infant was Group O, Rhesus positive. T h e direct Coombs test was negative. T h e course of serum bilirubin values is shown in Fig. 3. Motulsky's test. The father and mother gave normal decolorization. T h e patient gave no decolorization at 3 hours. A male sibling born recently had normal G6PD activity and had no jaundice. Case 4. A male infant was born in a private nursing home on Nov. 28, 1961. The birth weight was 4,500 grams. He was the second child of healthy parents. The mother had an episode of jaundice during the spring when she was 15. T h e firstborn, also a male, never had jaundice. The infant was discharged home on the sixth day having no trace of jaundice. On the seventh day slight jaundice appeared. In the morning of the next day the jaundice was intense and in the evening opisthotonus and inability to
Volume
63
Number 5
Hemolysis due to naphthalene inhalation
1•
.= 5O E
9 11
TotQI BRirubin J Direct //
E _z "n IE
~20
I0
2
o
o
o
"d ~
J:
,g2
a. w->{ o E
IS
F i g . 3. C a s e 2. C o u r s e o f s e r u m bilirubin values.
I
I
I
8
9
10
1 I
AGE
11 IN DAYS
I
I
I
12
13
14
[ ~
Tot(][ Bilirubin
l
%
I'-
Direc,
..
~50
Do
E40 z
o
"G E
.,~ .
.
.
.
.
.
.
.
o x p o s u r e to napht h(:l[ene ( >
F i g . 4. C a s e 4. C o u r s e o f s e r u m bilirubin and hemoglobin values.
I 6
I 8
,0~o 5 i
I 10
I I 12 14 AGE IN DAYS
I 16
I 18
I 20
E,T.
~
2o-~
20
o
g
z o
o
~I0
10o o
"~" "~ I :E D
F i g . 5. C a s e 7. C o u r s e o f s e r u m bilirubin and hemoglobin values. E x c h a n g e t r a n s f u s i o n is represented by the arrow.
i
....
exposure to naphtha[ene
S.B.
Hb
2
5
l l l l
3
4
5
6
7 ~
13 14 I"5
A G E IN DAYS
I'6 I'7 I'8
9 12
Valaes, Doxiadis, and Fessas
November 1963
nurse appeared. He was admitted in the "Alexandra" on the ninth day with very marked jaundice and unmistakable signs of kernicterus. The blanket with which the infant was covered since his return home smelled strongly of naphthalene. Examination at the age of 11 months showed definite sequelae of kernicterus. The course of serum bilirubin and hemoglobin is depicted in Fig. 4. Motulsky's test. The father showed normal decolorization; the mother, a delay of 30 to 40 minutes. The patient and his elder brother showed no decolorization at 3 hours. Case 6. This male infant was born at home Dec. 14, 1962. He was the third child of healthy parents. The previous two males had slight neonatal jaundice. No jaundice appeared during the first week of life. On the seventh day 3 or 4 diapers strongly smelling of naphthalene were used. The smell was quite strong as the father commented to the mother that it might " h a r m the baby." In the morning of the eighth day the infant was "blue." On the ninth day the infant was both "yellow and blue." Finally o n the tenth day the jaundice was intense and the infant's urine had a dark red color. On admission in the evening of the same day the infant was much distressed, cyanotic, anemic, and jaundiced. The spleen was palpable 2 cm. below the costal margin. Urine collected on admission had a dark red color and was positive for hemoglobin. He was placed in an oxygen tent and a transfusion of sedimented red
cells was started immediately. Three hours after admission an exchange transfusion of 500 ml. was started and was followed by a transfusion of 60 ml. of sedimented red cells. An infusion of 5 per cent glucose was given for the next 18 hours. Next morning his general condition was much improved, he was free from cyanosis, and had only moderate jaundice. On the twelfth day only a trace of jaundice was present and the spleen was no more palpable. He was discharged on the fifteenth day. Urine was collected for the whole 24 hours following his admission. There was a dramatic change in color following the exchange transfusion and 12 hours after the exchange the urine contained no hemoglobin. Motulsky's test. Mother and father had normal decolorization. The patient and his two brothers showed no decolorization at 3 hours.
Comment. This infant presented all the clinical and laboratory manifestations of acute intravascular hemolysis plus methemoglobinemia. Apart from the danger of hyperbilirubinemia his life was endangered from a much reduced oxygen-carrying capacity (anemia plus methemoglobinemia). Moreover 60 per cent of his red cells had morphologic evidence of impeding lysis. It was feared that this amount of hemoglobin when released would produce blockage of renal tubules. By choosing exchange transfusion instead of a simple transfusion we were able to correct rapidly the abnormal state.
Table II. Laboratory investigations Hematocrit
Sample
Serum bili- Hemoglobin Methemorubin (rag.~ (Gm./lO0 globin (%) 100 ml.) ml.)
O n admission ( 10th day, 19.00 hr.)
12.8
7.2
23
Pre-exchange (22.25 hr.) (following transfusion of packed cells)
25.6
10.2
14
Post-exchange
19.0
14.2
llth
--
14.8
day
Packed red cells
Column of stromata
. Heinz bodies (%)
Pyknocytes an~d [ragmented R.B.C.s
(%) 60
23
5
15
--
0.16
46
0
--
--
--
46
0
rare
rare
Volume 63 Number 5
Case 7. This infant, a male, was born in "Alexandra" Dec. 5, 1959. Birth weight was 3,200 grams. He was the second child of healthy parents. Jaundice was noticed before the end of the first 24 hours of life. The course of serum bilirubin is depicted in Fig. 5. The mother was Group O Rhesus positive while the father and the infant were Group B Rhesus positive. T h e direct Coombs test was negative. A diagnosis of O-B hemolytic disease was made and this was further substantiated when on the eighteenth day postpartum anti-B hemolysins were detected in the mother up to a dilution of 1:8 (anti-A hemolysins were also present up to 1:4). The infant was discharged on the seventh day still having slight jaundice. H e was fully breast fed and mother and infant received no drugs. The infant was brought to the outpatient department on the thirteenth day of life on account of increasing jaundice for the past 48 hours. Since the discharge from the Maternity Hospital, the infant had been dressed with woolen clothes which were previously kept with naphthalene mothballs and which at the time of his examination in outpatient were still strongly smelling of naphthalene. The infant was given an exchange transfusion on the fourteenth day of his life. The exchange transfusion was performed through the umbilical vein exposed above the umbilicus. Group B Rhesus positive blood was used after removal of the supernatant plasma; 290 ml. were introduced and 250 ml. removed. Exchange transfusion was used in preference to simple transfusion as the removal of bilirubin and damaged red cells was considered essential in addition to the correction of anemia. In the pre-exchange specimen 70 per cent of the red cells contained Heinz bodies (Fig. 6). Repeated testing of urine failed to detect any hemoglobin. The infant was discharged on the twentythird day of life free of jaundice. Follow-up to the age of 3 years failed to reveal any blood abnormality apart from moderate anemia which had responded to iron at the age of 11 months.
Hemolysis due to naphthalene inhalation
9 13
Fig. 6. Case 7. Coarse Heinz bodies in the red ceils.
Motulsky's test. The infant and both his parents gave normal decolorization. Also no abnormality was detected by hemoglobin electrophoresis in the patient and his parents. Case 20. On Oct. 5, t961, a male infant was born in a private nursing home in Athens. Birth weight was 3,800 grams. He was the second child of healthy parents. The first child, a girl, never had jaundice. Jaundice was noticed on the third day and gradually became deeper. On the sixth day the infant stopped sucking and was irritable. On admission the same day he had marked jaundice, no hepatosplenomegaly, and equivocal signs of kernicterus. The infant's blanket was smelling of naphthalene. He was treated with two exchange transfusions on the sixth and seventh day of life following which his jaundice gradually subsided. Laboratory investigations. T h e mother and infant were Group A Rhesus positive. The direct Coombs test was negative. O n the sixth day at 12:30 noon the serum bilirubin was 33.1 nag. per 100 ml. At 3 P.M. (preexchange) serum bilirubin was 40.9 mg. per 100 ml. and hemoglobin 14.8 Gin. per 100 mL Reticulocytes were 5.4 per cent. Heinz bodies were present in 95 per cent of the red ceils. The smear showed marked poikilocyto-
9 14
Valaes, Doxiadis, and Fessas
sis, a n d fragmentation of the red cells, anisocytosis, polychromasia, a n d spherocytosis were also present. Eighteen hours following the first exchange transfusion his serum b i l i r u b i n was 35.8 mg. per 100 ml. a n d a second exchange was given (seventh day). Motulsky's test. T h e infant, the parents, the m a t e r n a l grandparents, a n d two brothers of the mother showed n o r m a l decolorization.
Copies of the remaining case reports may be obtained by writing to the authors.
REFERENCES 1. Zuelzer, W. W., and Apt, L.: Acute hemolytic anemia due to naphthalene poisoning, J. A. M. A. 141" 185, 1949. 2. Newns, G.: Moth-ball anaemia, Lancet 2: 964, 1949. 3. Mackell, J. V., Rieders, F., Brleger, I-I., and Bauer, E, L.: Acute haemolytic anemia due to ingestion of naphthalene moth-balls, Pediatrics 7: 722, 1951. 4. MacGregor, R. R.: Naphthalene poisoning from the ingestion of moth-balls, Canad. M. A. J. 70: 313, 1954. 5. Chusld, E., and Fried, C. T.: Acute hemolytic anemia due to naphthalene ingestion, A. M. A. Am. J. Dis. Child. 89: 612, 1955. 6. Gidron, E., and Leurer, J.: Naphthalene poisoning, Lancet 1: 228, 1956. 7. Sansone, G., and Segni, G: Difetto biochimico eritrocitario a carattere genetico in un bambino con anemia emolitiea da naftalina, Boll. Soc. ital. biol. sper. 34: 516, 1958. 8. Schafer, W. B.: Acute hemolytic anemia related to naphthalene. Report of a case in a newborn infant, Pediatrics 7: 172, 1951. 9. Cock, T. C.: Acute hemolytic anemia in the neonatal period, A. M. A. Am. J. Dis. Child. 94: 77, 1957. 10. Dawson, J. P., Thayer, W. W., and Desforges, J.F.: Acute hemolytic anemia in the newborn infant due to naphthalene poisoning, Blood 13: 1113, 1958. 11. Zinkham, W. H., and Childs, B.: Effect of naphthalene derivatives on glutathione metabolism of erythrocytes from patients with naphthalene hemolytic anemia, J. Clin. Invest. 36: 938, 1957. 12. Zinkham, W. It., and CMlds, B.: The effect of vitamin K and naphthalene metabolites on the glutathione metabolism of erythroeytes from normal newborns and patients with naphthalene hemolytic anemia, A. M. A. Am. J. Dis. Child. 94: 120, 1957.
November 1963
13. Zinkham, W. If., and Childs, B.: A defect of glutathione metabolism in erythrocytes from patients with naphthalene induced hemolytic anemia, Pediatrics 22: 461, 1958. 14. Gross, R. T., Marks, P. A., Ifurwitz, R. E., Beasley, J., and Sui, M. Studies on the hereditary enzymatic defect in certain drug induced hemolytic anemias, Society for Pediatric Research. Twenty-eighth Meeting, 1958. 15. McGovern, J. J., Isselbacher, K., Rose, P. S., and Grossman, M. S.: Observations on the glutathione stability of red blood cells, Society for Pediatric Research. Twenty-eighth Meeting, 1958. 16. Sansone, G.: L'anemia emolitica acuta da ingestione accidentale di naftalina nel bambino. Contributo clinico e considerazioni patogenetiche, I-Iaemat. Lat. 1: 45, 1958. 17. Zannos, L.: Biochemical study of favism and hemolysis induced by drug and chemicals in Greece, Thesis, Athens, 1961. 18. Malloy, It. T., and Evelyn, K. A.: The determination of bilirubin with the photoelectric colorimeter, J. Biol. Chem. 119: 481, 1937. 19. Motulsky, A. G., and Campbell-Kraut J . M . : Proceedings of the conference on genetic polymorphlsms and geographic variations in disease, New York and London, 1961, Grune & Stratton Company, Inc. 20. Doxiadis, S., Valaes, T.: The problem of kernicterus in Greece, Proc. M. Soe., Athens, 1960, p. 507. 21. Doxiadis, S., Fessas, Ph., and Valaes, T.: Erythrocyte enzyme deficiency in unexplained kernicterus, Lancet 2: 44, 1960. 22. Doxiadis, S., Fessas, Ph., and Valaes, T., and Mastrokalos, N.: Glucose-6-phosphate dehydrogenase deficiency: A new aetiological factor of severe neonatal jaundice, Lancet 2: 297, 1961. 23. Panizon, F.: L'ictfre grave du nouveau-n6 associ6 ~ une defieience en glucose-6-phosphate dehydrogenase, Biologia Neonatorum 2: 167, 1960. 24. Weatherall D. J.: Enzyme deficiency in haemolytic disease of the newborn, Lancet 2: 835, 1960. 25. Gaburro, D., Volpatos, S., Gianqinto, M.: Ict~re nucleaire du nouveau-n6 par d6faut de la G-6-PD, Semaine H6p. (Ann. Ped.) 37: 69, 1961. 26. Valaes, T., Fessas,, Ph., and Doxiadis, S. A . : Kernicterus in full-term infants without isoimmunization, Proc, Roy. Soc. Med. 54: 331, 1961. 27. Fessas, Ph., Valaes, T., and Doxiadis, S. A.: Study of new aetiological factors of icterus gravis neonatorum, Proc. Med. Soc. Athens, 1961, p. 117. 28. Fabian, P.: Toxicka hemolyticka anemia u novoredence po otrove naptalinovymi parami, Clesk. pediat. 12: 833, 1957. 29. Betke, K., and Schall, I-I.: Akute h~imolytische Innenk6rperan~imie mit Meth~imoglobin~imie und Meth~imalbuminSmie bei einem Neuge-
Volume 63 Number 5
borenen, Wahrscheimlich bedingt durch Naphthalin, Zeitschr. Kinderh. 81" 379, 1958. 30. Younis, D., Veltsos, A., Platakos, Th., and Vafiadis, S.: Clinical and laboratory study of cases of naphthalene intoxication, Arch. Inst. Pasteur Hellenique, 1957, p. 521. 31. Kassimos, Ch., Vlachos-Anastasea, K., Koutsoukos K., and Ginieris, D.: Acute haemolytie anaemia in the neonatal period due to naphthalene, Arch. Paediat. Clinic University of Athens 5: 313, 1958.
Hemolysis due to naphthalene inhalation
9 15
32. Boyland, E., and Wihshire, G. H.: The metabolism of polycyclic compounds, Biochem. J. 53: 636, 1953. 33. Valaes, T.: Red-cell enzymes and severe neonatal jaundice, Cerebral palsy Bull. 3: 431, 1961. 34. Zinkham, W. H.: Peripheral blood and bilirubin values in normal full-term primaquinesensitive Negro infants: Effect of vitamin K, Pediatrics 31: 983, 1903.