[The clinical diagnostic value of a study of the activity of enzymes of the guanyl branch of purine metabolism in patients with ankylosing spondyloarthritis].

AIM: To assess the diagnosis of the activity of a pathological process in patients with ankylosing spondyloarthritis (AS) and to reveal purine metabolic (PM) changes in relation to the clinical features of AS. MATERIALS AND METHODS: The serum activities of the PM enzymes: xanthine oxidase (XO), guanine deaminase (GDA), guanosine deaminase (GSDA), purine nucleoside phosphorylase (PNP), guanosine phosphorylase (GP), and adenosine deaminase (ADA) were determined in 55 patients (51 males and 4 females) aged 36.0 +/- 1.4 years). A control group comprised 30 apparently age- and gender-matched healthy individuals, as in the study group. RESULTS: On admission, the patients were found to have increased XO, GDA, PNP, and GD activities and decreased GSDA activity. The higher activity of the process was observed, the greater activities of GDA, XO, PNP, GP and the lower activity of GSDA and ADA were. There were the enzymatic activity differences that depended on the degree of pathological process activity, clinical form, the magnitude of X-ray changes, and the degree of joint functional insufficiency. CONCLUSION: The findings suggest that there may be PM disturbances in the immunocompetent cells.

Estimation of guanine deaminase using guanosine as a "prosubstrate".

Plasma guanine deaminase (guanase; GD) is well established as an indicator of hepatocellular disease, recently being applied in the detection of hepatitis C in donor blood and in the diagnosis of hepatoma. No totally efficient, simple method for the estimation of plasma GD activity is routine since both guanine and 8-azaguanine, the substrates of the enzyme, are scarcely soluble in water. This difficulty in preparing stable substrates of sufficient concentration has resulted in methods that are both troublesome and inaccurate. Here we describe the development of new colorimetric and high-performance liquid chromatography (HPLC) methods utilizing guanosine as a "prosubstrate." After an initial breakdown of the guanosine to guanine using purine nucleoside phosphorylase, the ammonia formed as a result of the breakdown of the guanine by GD was estimated colorimetrically by the Berthelot reaction. As an alternative or a complementary assay, the xanthine also formed was measured using an isocratic HPLC method. These methods are suitable for routine assays for measuring plasma GD over a wide range of activities.

Clinical value of the determination of serum guanase activity in patients with chronic hepatitis type C.

The study examines the clinical significance of guanase (GU) measurement in patients with hepatitis C. 688 patients in whom either ALT was abnormal, or in whom HBsAg or HCVAb was detected in the serum, were enrolled into this study. The percentage of cases in which normal ALT while elevated GU was compared among the different disease groups. Then, the percentage of cases with normal ALT but elevated GU was compared between HBV and HCV groups. For the entire population, a significant correlation was observed between ALT and GU (r=0.872). The overall percentage of cases with normal ALT but elevated GU activity was 11.4%. In HCV group, 449 cases had normal ALT. Of these cases, 20.3% had elevated GU, while ALT was normal. Before 1989, no test to check donated blood for HCV antibody was available. However, screening of donated blood for high GU was associated with a reduced incidence of post-transfusion hepatitis. This is probably because following the screening, blood donated by patients with hepatitis C who had normal ALT but elevated GU was rejected. After the introduction of HCV antibody measurement, GU measurement is still useful to reveal the pathophysiological condition in-patients with chronic hepatitis type C.

Purine metabolism in Echinococcus multilocularis.

The activities of the enzymes in Echinococcus multilocularis metacestodes involved in purine salvage were studied by HPLC. As in most parasites, this cestode relies entirely on salvage of preformed bases and nucleosides for its purine requirement. Therefore, these enzymes may be targets for drugs in the chemotherapeutic treatment of diseases caused by this parasite. The animals used in this study were gerbils (Meriones unguiculatus). Enzyme activities from sera and hepatic tissue in control and infected animals were similar with the exception of adenine phosphoribosyltransferase which showed an activity 4-fold greater in the serum from control than in serum from infected animals. In the parasite, adenine and hypoxanthine-guanine phosphoribosyltransferases and adenosine deaminase had the highest activities. Therefore, in E. multilocularis metacestodes, this pathway seems to be important for the parasite's metabolism.

Simultaneous assay for aspartate aminotransferase and guanase in human serum by high-performance liquid chromatography.

A simple high-performance liquid chromatography (HPLC) assay for the simultaneous determination of guanase and aspartate aminotransferase (AST) activities in a single serum sample is described. The method is based on direct detection of enzymatically formed products xanthine and glutamate, respectively. The procedure is sensitive, precise (C.V. below 2% for guanase and 3% for AST), suitable for routine purposes and requires only 100 microliters of sample. Kinetic measurements have shown the guanase activity to have an apparent Michaelis constant of 24.5 microM and the AST activity of 11.1 and 0.18 mM for aspartate and oxoglutarate, respectively, at 37 degrees C in Tris-HCl buffer (pH 7.5).

[The value of determining guanine deaminase in diagnosis of hepatic diseases].

We determined the Guanine Deaminase (GD) activity of 200 patients with different diseases. It was found that GD activity of hepatic patients is higher than that of health adults, while the GD activity of other patients is in the normal range. There is a linear correlation between GD activity and ALT in patients with chronic hepatitis, billiary obstruction, and between GD activity and total bilirubin in patients with chronic active hepatitis, biliary obstruction and liver cirrhosis. Moreover, the GD activity of patients positive for anti-HCV is significantly increased. So GD activity in serum is a specific and sensitive index to estimating hepatic functions and can be used in the diagnosis of acute and chronic hepatitis, cirrhosis of liver, and C virus hepatitis.

Determination of guanase activity in normal and pathological sera by high-performance liquid chromatography.

A simple HPLC assay for serum guanase based on the direct determination of enzymatically formed xanthine was applied to normal and pathological sera. The procedure is sensitive, precise (CV below 5%) and suitable for routine purposes, and the method requires only 50 microL of sample. Using this method the reference range as determined from the sera of 40 healthy adult controls is 0-1.1 U/L. In patients with various liver diseases serum guanase activities were found to be increased 5- to 50-fold compared with the normal mean value.

New method for guanase activity measurement by high-performance liquid chromatography.

A rapid isocratic high-performance liquid chromatographic method for the determination of guanase (EC 3.5.4.3) activity is proposed. The method is highly reproducible, with a coefficient of variation of less than 1%, and requires only ca. 10 min for a complete chromatographic separation of the enzyme reaction mixture. The method allows the detection of nanomolar changes in the concentrations of both the substrate and the product, and does not require additional reactions or sample pretreatment. Kinetic studies with the proposed method showed the guanase activity to have an apparent Michaelis constant of 13.3 and 8.5 microM, and a maximum rate of 1.95 and 3.84 pmol/min per mg protein at 37 degrees C, in Tris-HCl and phosphate buffer, respectively.

Effect of prostaglandin E1 on preservation injury of canine liver grafts preserved in UW solution.

This study investigated whether prostaglandin E1 (PGE1) could reduce hepatic injury to the liver graft caused by harvesting and 24-h preservation in University of Wisconsin (UW) solution in a canine model. The PGE1-treated group was intravenously administered 0.5 microgram/kg per minute of PGE1 for 30 min before harvesting, as well as a concentration of 1 mg/l PGE1 in the washout and UW solutions. In both the PGE1-treated and the control group, all recipients survived for 1 week or more after transplantation. Arterial ketone body ratio (AKBR) remained over 1.0 in the early postoperative period. The PGE1 group showed significant reductions in guanase, GOT, and LDH during the early postoperative period compared to the untreated control group. Histological examination disclosed partial mitochondrial swelling, hepatocyte vacuolation, and necrosis in the control group, while such abnormalities were rarely seen in the PGE1 group. These results suggest that PGE1 can effectively reduce hepatic injury to liver grafts preserved in UW solution prior to transplantation.

[Activity of guanase in serum and liver function tests in HBe antigen-positive chronic hepatitis].

Forty-one patients with chronic hepatitis were divided into an HBe antigen-positive group (n = 13) and an HBe antigen-negative group (n = 28) to clarify the relationship between the presence of HBe antigen and liver function. In the HBe antigen-positive group, the activities of serum alanine aminotransferase (p less than 0.01), aspartate aminotransferase (p less than 0.05) and guanase (p less than 0.01) were significantly higher than those in the HBe antigen-negative group. The correlation coefficient between the HBe antigen titer and guanase activity was 0.528, which was higher than the corresponding values for alanine aminotransferase and aspartate aminotransferase. The determination of guanase activity in serum may be useful for evaluating the clinical severity of HBe antigen-positive chronic hepatitis.

Epidemiological study of occupational exposure to hepatitis B virus and liver function tests.

A total of 1020 hospital employees were divided into an exposure group (n = 725) and a non-exposure group (n = 295), based on whether they had been exposed to blood from patients. The HBsAg-positive rates for the exposure and the non-exposure groups were 2.48% and 1.02%, respectively. In the exposure group, the minimal exposure rate increased with age from the twenties. The odds ratios were 7.39 in the technicians, 4.38 in physicians and 1.32 in nurses. Using age-sex matched pairs from the exposure and non-exposure groups, comparison of aspartate aminotransferase, alanine amino-transferase and guanase activities showed that there were significantly higher values in HBsAg-positive subjects (n = 18) from the exposure group than in HBsAg- and HBsAb-negative subjects from the non-exposure group (p less than 0.05-0.01). However, no significant differences were found in the enzyme activities in the matched pairs (n = 89) of HBsAb-positive subjects from the exposure and non-exposure groups.

Clinical significance of serum guanase activity in various liver diseases.

Serum guanase activity was measured using a sensitive colorimetric method in patients with liver diseases. Guanase activity was correlated with GPT, GOT in acute viral hepatitis and chronic hepatitis, however, in liver cirrhosis and hepatocellular carcinoma it was correlated with total bilirubin as well as aminotransferases. In addition, the GPT-to-guanase ratio differed chronic hepatitis from liver cirrhosis. These findings suggest that determination of guanase and aminotransferases in useful in differentiation of liver diseases as well as assessing liver damage.

Simple ultraviolet spectrophotometric method for the determination of serum guanase activity.

We describe a simple, kinetic method for the determination of serum guanase activity that involves enzymatic coupling to xanthine oxidase and measurement of the rate of uric acid formation by spectrophotometric monitoring of the absorption at 300 nm. At this wavelength, the absorption of uric acid is about 80% of its maximal absorption at 293 nm, but the difference in molar extinction coefficient between guanine and uric acid is similar (9,000 at 293 nm vs 8,400 at 300 nm). There are three advantages to the use of the higher wavelength: first, the absorption of serum proteins is only one third of the absorption at 293 nm resulting in a significant reduction in noise level. Second, the lower absorption of serum proteins allows increasing sensitivity of the assay by increasing the amount of serum in the reaction mixture. Third, the higher wavelength allows the use of automated centrifugal analyzers that are generally not designed for measurements below 300 nm. The between-day coefficient of variation was 5.8% (n = 27) at an activity of 17 U/L and 8.2% at an activity of 2 U/L. The reference range for 50 sera from males and females was 0.4 to 1.8 U/L (n = 50; mean +/- 2SD). The method is linear to 40 U/L.