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Serum Biotinidase Activity in Children With Chronic Liver Disease and Its Clinical Significance

Pabuçcuoğlu, Aysun*; Aydoğdu, Sema; Baş, Meral*

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Journal of Pediatric Gastroenterology and Nutrition: January 2002 - Volume 34 - Issue 1 - p 59-62
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Biotin, a member of the B complex group of vitamins, acts as a prosthetic group in each of the four carboxylases in humans: pyruvate carboxylase, propionyl-CoA carboxylase, β-methylcrotonyl-CoA carboxylase, and acetyl-CoA carboxylase. These carboxylases participate in gluconeogenesis, fatty acid synthesis, and amino acid catabolism. Biotinidase (E.C is an amidohydrolase that catalyzes the cleavage of biotin from biocytin or biotinylated peptides that are formed during proteolytic turnover of biotin-dependent carboxylases. This enzyme is important not only for recycling endogenous biotin, but also appears to play an important role in processing dietary protein–bound biotin (1–3). Individuals lacking biotinidase activity become biotin deficient, and if untreated, these individuals usually exhibit seizures, hypotonia, hearing loss, alopecia, skin rash, and developmental delay often accompanied by metabolic acidosis and organic aciduria. Pharmacologic doses of biotin can alleviate symptoms and prevent neurologic damage if treatment is initiated early (4–7). In animals, the highest specific activity of biotinidase is found in the serum (8). The liver is the major source of plasma biotinidase (9). Serum biotinidase activity was shown to decrease in carbon tetrachloride–treated and partially hepatectomized rats (10). In addition, a few clinical reports suggest that serum biotinidase activity decreases in adult patients with chronic liver diseases (9,11,12).

This study was conducted to determine the serum biotinidase activity values in children with chronic liver disease and to investigate the relation among enzyme activity, certain liver function tests, and degree of liver damage.


The patient group consisted of 62 children with chronic liver disease who were treated at Ege University, Faculty of Medicine, Department of Pediatric Gastroenterology and Nutrition, and whose ages ranged from 8 months to 18 years (median age, 9.73 years). Diagnosis of the patient group was as follows: noncirrhotic chronic HBV infection (n = 12), metabolic disease (n = 16; Wilson disease, n = 7; glycogen storage disease type I, n = 5; Gaucher disease, n = 2; cystic fibrosis, n = 2), autoimmune hepatitis (n = 6), cholestatic liver diseases (n = 14), cryptogenic cirrhosis (n = 5), fulminant hepatitis (n = 5), and prehepatic portal hypertension (n = 4). All of the patients underwent liver biopsy, and their hepatocellular functions were assessed by Child-Pugh (C-P) classification (13).

The control group consisted of 27 healthy subjects (17 adults and 10 children). The mean age of the 17 healthy adults (10 women and 7 men) was 39.7 ± 10.7 years, ranging from 20 to 60 years. The mean age of the healthy children (5 girls and 5 boys) was 7.4 ± 4.7 years, ranging from 2 to 14 years.

Fasting sera were obtained from the whole blood of these patients and controls. The serum samples were frozen at −20°C until the time of assay (14–16). Biotinidase activity was determined spectrophotometrically in all specimens according to the modified method of Wolf et al. (8), in which biotinidase is assayed by measuring the hydrolysis of n-biotinyl-p-aminobenzoate. p-Aminobenzoic acid thus released is diazotized and coupled to a naphthol reagent, and determined by its absorbance at 546 nm. Enzyme activity is expressed as the released p-aminobenzoic acid in nmol · min −1 · mL −1 of serum.

Albumin, total bilirubin, alkaline phosphatase, alanine aminotransferase, and γ-glutamyltransferase levels, and prothrombin time were determined for each patient using a Falcor 300 autoanalyzer (The Menarini Group, Menarini Diagnostics, Italy), and the results were correlated with serum biotinidase activity.

Statistical Analysis

The results reported represent the mean ± SD. Differences between the groups were evaluated using Student t test. Correlation of parameters in the patient group was performed using linear regression analysis. A P value of less than 0.05 was considered statistically significant.


Table 1 shows the mean serum biotinidase activity values of healthy adults and children. Serum biotinidase activities of healthy adults and children were similar. Therefore, all values were combined in the same control group. Furthermore, there was no significant difference in the serum biotinidase activity between female and male subjects (P > 0.05).

Serum biotinidase activity in control group

Table 2 shows the mean serum biotinidase activity values of all patient groups. There was significant difference between the mean enzyme activity of the controls and of all patients with chronic liver disease (P < 0.05).

Serum biotinidase activity in patient group

Within the patient group, patients with noncirrhotic chronic viral hepatitis, metabolic liver diseases, autoimmune hepatitis, cryptogenic cirrhosis, and prehepatic portal hypertension displayed serum biotinidase activity similar to that of the control group (P > 0.05). In the metabolic group, patients with Wilson disesase had lower serum biotinidase activity values than did controls (P < 0.05). Of the seven patients with Wilson disease, two were accepted as fulminant Wilson disease and underwent transplantation. However, serum biotinidase activities of patients with glycogen storage disease type I, Gaucher disease, and cystic fibrosis were similar to that of the control group. Serum biotinidase activity values in cholestatic hepatic diseases and the fulminant hepatitis group were significantly lower than those of the control group (P < 0.05). In the cholestasis group, seven patients had biliary atresia, two had Alagille syndrome, and five had nonsyndromic biliary hypoplasia. There was significant difference between the mean enzyme activity of the biliary atresia group and that of the control group (P < 0.05).

Of our 62 patients, 23 did not have cirrhotic disease (12 had chronic HBV, 4 had prehepatic portal hypertension, 5 had glycogen storage disease, 2 had Gaucher disease). Of the other 39 patients, 22 had compensated cirrhosis (C-P score < 9, class A or B), 12 had decompensated cirrhosis (C-P score > 9, class C), and 5 had fulminant hepatitis (C-P score > 9, class C). Serum biotinidase activity values in the patients without cirrhosis and those with compensated cirrhosis were comparable with those of the control group (P > 0.05). Among the patients with decompensated cirrhosis, 10 were transplanted and 2 died while waiting for transplantation. Serum biotinidase activity values in these patients were significantly lower than those of the control group (P < 0.05) (Table 3).

The mean serum biotinidase activity in patient group in regard to the hepatocellular functions and compared with that of the controls

The lowest enzyme activity was found in individuals with fulminant hepatitis (1.8 ± 0.5 nmol · min −1 · mL −1 ) (n = 5). The mean enzyme activity of these patients was 24% of the mean control activity. In this group, two patients with fulminant Wilson disease had undergone transplantation. Of the remaining three patients who did not have hepatitis A, B, or C, one could be transplanted and the others died of the disease.

In the patient group, serum biotinidase activity correlated positively with albumin concentration (P = 0.030, r = 0.276) and correlated negatively with total bilirubin concentration (P = 0.000, r = −0.504), prothrombine time (P = 0.009, r = −0.327), alkaline phosphatase concentration (P = 0.017 r = −0.301), γ-glutamyltransferase concentration (P = 0.049, r = −0.251), and C-P score (P = 0.000, r = −0.654).


The major function of biotinidase is to recycle biotin from biocytin or biotinylated peptides (17). Therefore, low biotinidase activity can cause secondary biotin deficiency (5,6). Human serum biotinidase is primarily synthesized by the liver (9). Pispa et al. (10) showed that serum biotinidase activity decreased in carbon tetrachloride–treated and partially hepatectomized rats. Clinical reports had shown that adult patients with chronic liver diseases had decreased serum biotinidase activity and concluded that the patients with low biotinidase activity may develop symptoms of biotin deficiency (9,11,12).

Serum biotinidase activity increases throughout late gestation and the newborn period to reach mean adult levels about 3 months after birth (18). Our results for the enzyme activities of healthy adults and children support those reported in the literature (2,7,18). Therefore, the values for healthy adults and children were combined in the same control group.

In this study, we aimed to determine serum biotinidase activity in children with chronic liver disease. Serum biotinidase activity decreased in children with decompensated cirrhotic disease and fulminant hepatitis. However, enzyme activity was normal in children with noncirrhotic chronic liver disease. Meanwhile, the serum biotinidase activity in those with compensated cirrhosis was similar to that of the control group. The patients with end-stage liver disease on the awaiting list for liver transplantation had low serum biotinidase activity. The lowest enzyme activities were found in patients with fulminant hepatitis. This finding supports previous reports that suggest that the liver is the major source of serum biotinidase. Because low enzyme activity has occurred only in end-stage liver disease, serum biotinidase activity is not a prognostic criterion in chronic liver disease.

Our patients with low enzyme activity did not show any clinical symptoms related to biotin deficiency. However, Grier et. al (9) mentioned that patients who had biotinidase activity below 30% of mean normal activity could become symptomatic when stressed by an infection. Wolf et al. (19) suggested that children with biotinidase deficiency could not maintain homeostasis under the stress of fever, injury, infection, starvation, or rapid growth and that subsequently acidosis, coma, or death may occur. Biotin deficiency may affect immune function. Impaired immunoregulatory function or immunodeficiency has occurred in children with biotin-responsive multiple carboxylase deficiency (20,21). However, a novel glycosylphosphatidylinositol-anchored protein (GPI-80) has been described on human leukocytes (22). This novel protein GPI-80 and Vanin-1, a GPI-anchored protein expressed on perivascular thymic stromal cells, have been found 40% homologous with human biotinidase. This report suggested that a biotinidase superfamily of molecules may regulate leukocyte trafficking along the endothelium toward the extravascular tissues (22).

Of our patients with decompensated cirrhosis, 12 had biotinidase activities below 30% of mean control activity. These patients may become symptomatic under stressful conditions, e.g., infection.

Earlier studies reported a significant correlation between serum biotinidase activity and albumin concentration and prothrombine time (9,12). In the patient group with chronic liver disease, a significant positive correlation was observed between serum biotinidase activity and serum albumin level, and a significant negative correlation was observed between serum biotinidase activity and total bilirubin level, prothrombine time, alkaline phosphatase level, γ-glutamyltransferase level, and C-P score. These values point to impairment of hepatic biosynthetic function and to hepatic excretory failure.

In conclusion, serum biotinidase activity is significantly lower in patients with decompensated liver cirrhosis, Wilson disease, and fulminant hepatitis. Although these patients did not exhibit clinical findings related to biotin deficiency, the presence of stress conditions could trigger symptoms. As a further precaution, biotin supplement therapy may benefit patients with end-stage liver disease.


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Biotin deficiency; Biotinidase activity; Chronic liver diseases

© 2002 Lippincott Williams & Wilkins, Inc.