Gastroenterologia Japonica Vol. 24, No.

3
Copyright (~ 1989 by The Japanese Society of Gastroenterology Printed in Japan

--Case Report--

A case of Gilbert's syndrome combined with macroamylasemia

Hiroshi INOUE, Yukihiko ADACHI, Masaki YAMASHITA, Tatsuo NANNO, Hiroko KATOH,
Masakazu ENOMOTO, Masao SUWA, and Toshio YAMAMOTO
Second Department of Internal Medicine, Kinki University School of Medicine, Osakasayama 589, Japan

Summary: A 30-year-old Japanese male, who had no remarkable family history, visited our hospital
with a complaint of abdominal pain, and unconjugated hyperbilirubinemia and hyperamylasemia were
observed. He showed negative hemolysis tests, positive nicotinic acid test, low hepatic bilirubin UDP-
glucuronyltransferase activity, decreased bilirubin diglucuronide and increased bilirubin mono-
glucuronide in bile, and a decrease in serum bilirubin after phenobarbital administration. He also
showed high serum amylase level, low urine amylase level, and low amylase-creatinine clearance ratio.
Gel filtration of serum with Sephadex G-200 revealed the existance of macroamylase. Countercurrent
immunoelectrophoresis proved binding of serum amylase to lambda type IgA. From these results, the
case was diagnosed as Gilbert's syndrome combined with macroamylasemia. Gastroenterol Jpn
1989;24:320-324

Key words: bilirubin; Gilbert's syndrome; immunoglobulin A; macroamylasemia; phenobarbital

Introduction
Gilbert's s y n d r o m e is m o s t f r e q u e n t l y f o u n d in
constitutional u n c o n j u g a t e d h y p e r b i l i r u b i n -
emia 1, in w h i c h slight h y p e r b i l i r u b i n e m i a of
less t h a n 5 m g / d l 2"3is observed. Macroamylase-
m i a w a s first reported by W i l d i n g et al. 4 in 1964.
The disease s h o w s a h i g h s e r u m amylase level
b e c a u s e the s e r u m amylase is attached to h i g h -
m o l e c u l a r - w e i g h t molecules such as i m m u n o -
globulin a n d forms molecules too large to pass
t h r o u g h renal glomeruli. We recently e n c o u n -
tered a case of Gilbert's s y n d r o m e c o m b i n e d
with macroamylasemia.
Case Report
Patient: A 30-year-old b u s i n e s s m a n .
Chief complaint: Jaundice.
Family history: U n r e m a k a b l e .
History:
He suffered from left ureterolithiasis at the
age of 28; this w a s c u r e d by medical treatment.
No operative history.
Present History:
In March 1981, he visited a family doctor
because of a b d o m i n a l pain, was d i a g n o s e d as
h a v i n g acute pancreatitis a n d received oral
medication. Jaundice w a s p o i n t e d o u t at that
time, b u t was n e g l e c t e d because the s y m p t o m s
d i s a p p e a r e d . In O c t o b e r 1984, he c o n s u l t e d the
s a m e doctor again for a b d o m i n a l pain, a n d
jaundice, slight liver d y s f u n c t i o n a n d h y p e r -
a m y l a s e m i a w e r e o b s e r v e d as a result of blood
tests. He was i n t r o d u c e d to our h o s p i t a l for
further e x a m i n a t i o n .
Physical findings:
H e i g h t 166cm, w e i g h t 69kg, b l o o d pressure
130/60, heart rate 72/min. No a n e m i a or e d e m a

Received October 6, 1988. Accepted December 16, 1988.

Address for correspondence: Yukihiko Adachi, M.D., Second Department of Internal Medicine, Kinki University School of Medicine,
377-20hnohigashi, Osakasayama-shi, Osaka 589, Japan.
June 1989 Coexistance of Gilbert's syndrome and macroamylasemia 321

Table 1 Laboratory data on admission

GBC " Biochemical Examination ." Serological Examination :

R BC 5.36 x 10 s Glucose 83mg/d I CR P (-)
Hb 16.2 g / d l Cholesterol 17 l m g / d I ASLO (-)
Ht 48 % T.P. 7.3 g / d l RA (-)
P latlet 19.1 x 104 Albumin 5.76 g / d I Coombs' test (--)
Reticulocyte 19 %0 Oh-Globulin 0.41 g J d l antinuclear antibody (--)
WBC 5.6x103 Q'2-Globul in 0.77 g / d l anti - D N A (-)
Stab 1% #-Globulin 0.65 g / d l haptoglobin 241 m g / d I
Seg 40 % T-Globulin 1.06 g / d l I mmunogl obul ins "
Lymph 46 % T . B i l irubin 2.7 m g / d I I gG 970mg/d I
Mono 10 Yo D. Bilirubin 0.3 m g / d l I gA 28ling/all
Eos 3 % I .Bil irubin 2.4mg/dl IgM 79mg/d I
Bas 0 % GOT 29 U / L I gD 16.gmg/d I
Urinalysis : GPT 71 U / L Others
protein (-) AL P 87 U / L I C G (15") 10%
glucose (--) L DH 167 U / L PFD test 80.2 %
urobi I inogen normal BUN 16 m g / d l CPC pattern normal
sediment normal Great inine 1.0 m g / d l creatinine clearance 80.4m l / m i n .
Feces : Gh - E 1.15 -~pH s--Amylase 1030 I U / L
Occult blood (--) LAP 160 G.U. u--Amylase T87 I U / L
r -GTP 11 mU/ml s--Elastarse I 120 m g / d l
Na 143 m E q / L s - - Lipase 0.1 UJml
K 3.9 mEq/L Cam/Oct 0.6%
CI 108 m E q / L

Fig. 1 Electrophoretic mobility of patient's amylase on agarose
gel film.
Serum amylase of the patient was stained diffusely be-
tween bands S and P, with slight tailing of staining from
band S to anode.
V: The position where the samples were applied.
was found. Heart-lung observations w e r e un-
remarkable. The a b d o m e n was flat w i t h un-
palpable liver and spleen. No abnormality was
f o u n d on neurological examination.
Laboratory examinations:
As s h o w n in Table 1, slight unconjugated hy-
p e r b i l i r u b i n e m i a was observed with slightly
elevated GPT. No data suggestive of hemolysis
were obtained: negative C o o m b s ' test, normal
haptoglobin level, normal pattern of hemolysis
with a coil planet centrifuge, and normal reticu-
locyte count. Renal function test and pancreatic
exocrine function test (PFD test) were normal.
Macroamylasemia was d i a g n o s e d by the
high s e r u m amylase level, low urine amylase
level, and a low amylase-creatinine clearance
ratio of 0.6%. Electrophoresis of the patient's
serum on agarose gel film 5 s h o w e d diffuse con-
tinuous staining from b a n d s P to S with slight
tailing from b a n d S toward the anode (Fig. 1).
On thin layer gel filtration, control serum and
the patient's urine s h o w e d a single b a n d of
amylase activity at almost the same position,
while the patient's serum s h o w e d bands at the
low-molecular-weight area and near the IgG
322 H. Inoue et al. Vol. 24, No. 3

(mg/dl) : - T. Bil.
e.----e L Bil.
6.0- , - - - e D. Bil.
,~ 50mg of Nicotinic Acid.
- - ~,..,,,,.,,~
5.0-

Fig. 2 Thin layer gel chromatogram of serum and urinary amyl-

ase of the patient.
4.0-

3.0-

2.0"
/
/
Serum amylase of the patient was stained at the IgG
(and IgA) elution band and at the normal amylase band, 1.0"

with light staining between them. Urinary amylase of the

patient was stained at the normal amylase band.
(~ 610 1"~0 I~ 2~'0 (rain)

Fig. 4 Nicotinic acid test.

Serum total and indirect-reacting bilirubins increased after
intravenous nicotinic acid administration, and they
reached their peaks at more than twice the preadministra-
tion levels after 120 min. Serum direct-reacting bilirubin
did not change throughout the test.

(mgldl)

Fig. 3 Detection of immunoglobulin bonded to amylase by elec- 4.0. T. Bil.

troimmunosyneresis. e-...e I, Bil.
The amylase of the patient's serum was precipitated by 3.o-
anti-lgA antibody and anti-K chain antibody. On the right
side the precipitated bands are schematically drawn by 2.0-
pencil.
1.0.

area, with slight activity between them (Fig. 2). before after

Sorting by countercurrent immunoelectro- Fig. 5 Effect of phenobarbital administration on serum bilirubin

phoresis 6 proved that the amylase-binding im-
munoglobulin was lambda type IgA (Fig. 3).
For the diagnosis of jaundice, nicotinic acid
(50mg) was injected intravenously and serum
bilirubin was monitored over time (nicotinic
acid test) (Fig. 4). Total and indirectreacting
bilirubin were 2.3 and 1.8 mg/dl, respectively,
before the nicotinic acid test. Bilirubin began to
rise and reached a peak 120 min after loading,
with total bilirubin of 5.2 mg/dl and indirect-
reacting bilirubin of 4.8 mg/dl: more than twice
of the values before loading. Lowering of the
serum bilirubin level was observed after oral
phenobarbital administration (100 mg per day
for 14 days) (Fig. 5). Serum bilirubin decreased
level.
Serum total and indirect-reacting bilirubins fell to the up-
per limit of the normal values on the 8th day after oral
phenobarbital treatment (100 mg/day).
Their decreased levels continued for at least 4 days after
discontinuation of the treatment.
to the normal level a week later, remained low at
least for 4 days after the discontinuation of
phenobarbital and then rose to the initial level
after a week. Bilirubin in the C bile obtained by
the Meltzer-Lyon procedure was fractionated
by high performance liquid chromatography
(HPLC) 7 to estimate in vivo hepatic bilirubin-
conjugating ability (Table 2). Decreased biliru-
June 1989 Coexistance of Gilbert's syndrome and macroamylasemia 323

76.5+4.8% (mean+SD) for BDG, 9.8_+2.5% for

BMG and 0.2+0.2% for UB 8. Phenobarbital ad-
ministration increased the proportion of BDG
and decreased those of UB and BMG to the
normal levels in the C bile.
Laparoscopy revealed the normal appearance
and color of the surface of the liver. Histology of
the liver was compatible w i t h that of Gilbert's
s y n d r o m e : No fibrosis in lobules, no remark-
able inflammation, and slight fatty infiltration
(Fig. 6). Bilirubin-UDP-glucuronyltransferase
(BGT) activity of the liver h o m o g e n a t e 9 was 0.65
nmol/mg-protein/15 min, less than 1/4 of the
m e a n normal value of 4.10+0.30 (SD) n m o l / m g -
protein/15 m i n obtained in our department.

Fig. 6 Histology of the biopsied liver specimen. Hematoxylin and
eosin staining. (x400)
bin diglucuronide (BDG) and increased biliru-
bin m o n o g l u c u r o n i d e (BMG) and unconju-
gated bilirubin IXa (UB) were found in the C
bile in comparison with the following percen-
tages of the fractions in normal subjects:
Discussion
Gilbert's s y n d r o m e is reported to be present in
2 to 5% of the population 2'3. Powell et al. l~ re-
ported that the s y n d r o m e was hereditary and
that the h o m o z y g o t e m i g h t develop Crigler-
Najjar s y n d r o m e type II. Patients with Gilbert's
s y n d r o m e are reported to have lower BGT ac-
tivity in the liver than normal subjects ii, and
this was true in the present case. Fractionation
of bilirubin in bile may allow estimation of the
in vivo conjugating ability of bilirubin in hep-
atocytes, since UB and BMG in bile are re-
ported to be clearly increased in Gilbert's syn-
d r o m e s. As illustrated in Table 2, the present
case s h o w e d higher levels of UB and BMG and
lower level of BDG in d u o d e n a l aspirate than
controls. In this case, phenobarbital increased
BDG and reduced UB and BMG to the normal
values. The increased BDG was thought to be

Table 2 Effect of phenobarbital administration on the bilirubin fractions in duodenal aspirates

Total bilirubin BDG GG GX BMG Bilirubin IX,8 UB

(mg/dl) &/or IXa
Before phenobarbital 9.6 66.2 9.1 4.5 16.7 2.5 1.0
After Phenobarbital 7.9 77.8 6.5 2.3 10.3 2.5 0.6
Normal controls
(mean+SD, n=7) 8 19.4+13.7 76.5+4.8 8.7_+2.0 3.7_+1.1 9.8_+2.5 1.1_+0.7 0.2_+0.2

Data of each fraction are expressed as the percentage of total bilirubin

GG: Bilirubin monoglucuronide monoglucoside diester
GX: Bilirubin monoglucuronide monoxyloside diester
324 H. Inoue et al.
caused by BGT induction in hepatocytes after
administration of the drug, although BGT was
not determined. On the other hand, Felscher et
aP 2 described that phenobarbital had no effect
on bilirubin fractions in bile from patients with
Gilbert's syndrome. The difference between
our results and theirs may indicate hetero-
geneity in Gilbert's syndrome.
Macroamylasemia is found in 0.21% of the
male population 13. Macroamylasemia some-
times shows no clinical manifestations and
should be kept in mind in cases of incompre-
hensible hyperamylasemia. Berk et al. 14 has
reported macroamylasemia in cases of autoim-
m u n e diseases such as rheuamatoid arthritis,
multiple sclerosis, and systemic lupus erythe-
matosus, but there are no such reports in this
country. In Japan, many cases have been re-
ported in association with pancreatic disease or
pancreatitis-like pain. This may be simply be-
cause amylase is tested more frequently in these
conditions. The combination of Gilbert's syn-
drome with macroamylasemia has not been re-
ported. The present case was found by chance.
The frequency of complication of macroamyl-
asemia with Gilbert's syndrome is estimated to
be less than 0.011% by simple calculation on the
assumption that putative genes for these dis-
eases are independent of each other. Gilbert's
syndrome and macroamylasemia both have
good prognoses and our patient has been fol-
lowed up as an outpatient with no special treat-
ment.
Vol. 24, No. 3
References
1. Adachi Y, Yamashita M, Ozaki K, et al: Constitutional jaun-
dice. Kantansui 1984;9:363-372 (in Jpn)
2. Berk PD, Wolkoff AW, Berlin Nh Inborn errors of bilirubin
metabolism. Med Clin North Am 1975;59:803-816
3. Namihisa T, Yamaguchi K: Constitutional hyperbilirubin-
emia in Japan. Studies on 139 cases reported during the
period from 1963 to 1969. Jpn J Med 1971;60:518-527 (in Jpn)
4. Wilding P, Cooke WT, Nicholson Gh Globulin-bound amyl-
ase. A Cause of persistently elevated levels in serum. Annal
Int Med 1964;60:1053-1059
5. Matsumaru K, Saitoh I, Kawahara T, et al: A new amylase-
fractionating method using cellulose acetate membrane.
Physico-chemical Biology 1979;24:266 (in Jpn)
6. Horii K and Kano S: Detection by electroimmunosyneresis
without deproteinization (in combination with orthogonal
electrophoresis). Physico-chemical Biology 1983;27:315-320
(in Jpn)
7. Yamashita M, Adachi Y, Yamamoto T: Analysis of bilirubin
conjugated in human bile by column liquid chromatography.
J Chromatogy 1986;375:386-391
8. Yamashita M, Adachi Y, Yamamoto T: Analysis of bilirubin
conjugates in human bile---changes of their composition in
hepatobiliary diseases. J Gastroenterology Hepatol 1987;2:
181-190
9. Black M, Billing BH, Heiwegh KPM: Determination of bili-
rubin UDP-glucuronyltransferase activity in needle-biopsy
specimens of human liver. Clin Chim Acta 1970;29:27-35
10. Powell LW, Hemingway E, Billing BH: Idiopatic unconjuga-
ted hyperbilirubinemia (Gilbert's syndrome). A study of 42
families. N Engl J Med 1967;277:1108-1112
11. Adachi Y, Yamamoto T: Hepatic Bilirubin-conjugating en-
zymes of man in the normal state and in liver disease. Gastro-
enterol Jpn 1982;17:235-240
12. Felsher BF, Craig JR, Carpio N: Hepatic Bilirubin glucuronid-
ation in Gilbert's syndrome. J Lab Clin Med 1973;6:829-837
13. Tozawa T: Amylase-binding immunoglobulin. Jpn J Clin
Pathol 1984;60(Suppl):98-104
14. Berk JE, Kizu H, Take S, et al: Macroamylasemia: clinical and
laboratory features. Am J Gastoroenterology 1970;53:211-222