A number of tests are used to evaluate the liver; however, none are sensitive and specific for assessment of hepatic function. Abnormalities detected during abdominal palpation and radiography may indicate changes in liver size, but do not provide any indication of hepatic function.1 Although ultrasonography of the liver may reveal morphologic changes in hepatic structure, it does not permit definitive assessment of hepatic function.2
Other modalities commonly used to evaluate the liver include the measurement of serum hepatic enzyme activities and serum bile acid concentrations. Alterations in serum hepatic enzyme activities may indicate hepatocellular damage, which may be associated with a decrease in hepatic function. However, some of these enzymes are released from other tissues during nonhepatic disorders and make interpretation of hepatic function difficult.3 Presently, measurement of pre- and postprandial serum bile acid concentrations is the most clinically useful hepatic function test available; however, this test is neither sensitive for compromised hepatic function nor specific for identifying a particular hepatic disease.4 Patients with mild liver disease may have serum bile acids concentrations within the reference range. Additionally, serum bile acids concentrations may be increased in patients with cholestasis in which hepatic function is initially normal.4 In addition, serum bile acids concentrations do not correlate with the severity of hepatic disease.4
The indocyanine green, sulfobromophthalein, and ammonia tolerance tests have been developed to evaluate hepatic function. The indocyanine green and sulfobromophthalein tests evaluate hepatic perfusion and hepatobiliary function, but they use compounds with several disadvantages and are therefore rarely used.5 The ammonia tolerance test measures the ability of the liver to extract and detoxify ammonia.6 This test requires rapid processing of samples, and its clinical use is limited by its insensitivity. Therefore, a new test that is both sensitive and specific for assessment of hepatic function in dogs and cats is clearly needed.
In humans, the aminopyrine breath test has been developed to quantify hepatic microsomal enzyme function.7–11 Results of numerous studies9–12 indicate that this test is clinically useful and that test results correlate with disease severity as assessed by histopathologic evaluation of biopsy specimens obtained from patients with hepatic cirrhosis and chronic hepatitis.
In contrast to humans, breath tests are difficult to perform in dogs and cats, and thus a blood-based test would be preferable for use in these animals. The principles of the carbon 13 (13C)-labeled aminopyrine demethylation blood test have previously been reported13 and are similar to those of the breath test used in humans. Aminopyrine is administered orally or IV and is demethylated in the liver by microsomal enzymes. As a result, the methyl groups produced are then oxidized to CO2. The CO2 diffuses into the vascular space, is eventually carried to the pulmonary alveoli, and is released into the expiratory air.8,14,15 By use of aminopyrine labeled with various carbon isotopes, either 13C or carbon 14 (14C), the amount of 13C or 14C derived from the labeled aminopyrine can be measured in the expired air (breath test) as either 13CO2 or 14CO2, respectively.16 For the breath test, the amount of 13CO2 or 14CO2 is determined as a percentage of the oral or IV dose of 13C-aminopyrine or 14C-aminopyrine administered that is recovered in expiratory air.16 Similarly, the blood test involves measuring the amount of 13CO2 extracted from blood samples.13 Results of a preliminary study13 indicated that a 13C-aminopyrine demethylation blood test is technically feasible in dogs. In that study, 13C-aminopyrine administered orally at a dose of 2 mg/kg to healthy dogs resulted in a detectible increase in the PCD in all dogs. Another study17 was performed to evaluate the demethylation kinetics of 13C-aminopyrine administered IV in healthy dogs and determine an appropriate parameter for quantification of aminopyrine demethylation. Results of that study indicated that IV administration of 13C-aminopyrine, as previously detected for oral administration, does not result in any gross clinical adverse effects. In addition, PCD was found to be an appropriate parameter for the quantification of aminopyrine demethylation. Results of that study17 also indicated that a single blood sample collected 45 minutes after IV administration of 13C-aminopyrine is sufficient for assessment of hepatic demethylating capacity in dogs.
The purpose of the study reported here was to determine the optimal dose of 13C-aminopyrine for use in a 13C-aminopyrine demethylation blood test in healthy dogs. We used a dose of 2 mg/kg as an arbitrary starting dose because this dose is commonly used for the aminopyrine breath test in humans. The ideal dose would be one that gives the lowest variability of PCD values in healthy dogs and is cost-effective for use in a clinical setting.
Percentage dose of carbon 13 administered as carbon 13-aminopyrine and recovered in gas extracted from blood samples
Cumulative PCD values up to a given sampling time
Coefficient of variation
Nutritional Research Center, The IAMS Co, Lewisburg, Ohio.
BD vacutainer, Preanalytical Solutions, Franklin Lakes, NJ.
4-dimethyl-13C2-aminoantipyrine, Isotech Inc, Miamisburg, Ohio.
Gelman Sciences, Supor Acrodisc, 0.1 μm, sterile, VWR Scientific Products Corp, West Chester, Pa.
Hydrochloric acid, Sigma Chemical Co, St Louis, Mo.
Automated breath carbon analyzer, Europa House, Crewe, UK.
GraphPad Prism, version 3.0, GraphPad Software Inc, San Diego, Calif.
Center SA, ManWarren T & Slater MR, et al. Evaluation of twelve-hour preprandial and two-hour postprandial serum bile acids concentrations for diagnosis of hepatobiliary disease in dogs. J Am Vet Med Assoc 1991;199:217–226.
Center SA. Serum bile acids in companion animal medicine. Vet Clin North Am Small Anim Pract 1993;23:625–657.
Center SA, Bunch SE & Baldwin BH, et al. Comparison of sulfobromophthalein and indocyanine green clearances in the cat. Am J Vet Res 1983;44:727–730.
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Henry DA, Kitchingman G, Langman MJ. [14C]Aminopyrine breath analysis and conventional biochemical tests as predictors of survival in cirrhosis. Dig Dis Sci 1985;30:813–818.
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Beyeler C, Reichen J & Thomann SR, et al. Quantitative liver function in patients with rheumatoid arthritis treated with low-dose methotrexate: a longitudinal study. Br J Rheumatol 1997;36:338–344.
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Moeller EM, Steiner JM & Williams DA, et al. Preliminary studies of a canine 13C-aminopyrine demethylation blood test. Can J Vet Res 2001;65:45–49.
Schoeller DA, Kotake AN & Lambert GH, et al. Comparison of the phenacetin and aminopyrine breath tests: effect of liver disease, inducers and cobaltous chloride. Hepatology 1985;5:276–281.
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Moeller EM, Steiner JM & Williams DA, et al. Kinetic analysis of demethylation of 13C-aminopyrine in healthy dogs. Am J Vet Res 2004;65:159–162.
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Chiaramonte D, Steiner JM & Broussard JD, et al. Use of a 13Caminopyrine blood test: first clinical impressions. Can J Vet Res 2003;67:183–188.