Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1989 Feb;83(2):688–697. doi: 10.1172/JCI113933

Erythromycin breath test as an assay of glucocorticoid-inducible liver cytochromes P-450. Studies in rats and patients.

P B Watkins 1, S A Murray 1, L G Winkelman 1, D M Heuman 1, S A Wrighton 1, P S Guzelian 1
PMCID: PMC303730  PMID: 2913056

Abstract

The major P-450IIIA gene family member present in human liver is HLp which, like its rat liver orthologue P-450p, is inducible by glucocorticoids and catalyzes erythromycin N-demethylation. To develop a practical method to estimate the amounts of HLp in patients [14C]N-methyl erythromycin was injected into rats that had been pretreated with dexamethasone or with inducers of other forms of cytochrome P-450. The rate of demethylation of this substrate, measured simply as 14CO2 in the breath, correlated well with the concentrations of immunoreactive P-450p protein (r = 0.70), holocytochrome P-450p (r = 0.70), or with erythromycin N-demethylase activity (r = 0.90) determined in the liver microsomes prepared from each rat. Next, [14C]N-methyl erythromycin was administered to 30 patients and there was a sixfold interindividual variation in breath 14CO2 production seemingly unrelated to medications, smoking status or age. However, the average breath test values were twofold greater in female as compared to male patients (P less than 0.01). Breath 14CO2 production rose in patients retested after treatment with the P-450IIIA inducers dexamethasone (P less than 0.05) or rifampicin (P less than 0.05) and was decreased after treatment with the HLp inhibitor triacetyloleandomycin (P less than 0.05). We conclude that the erythromycin breath test provides a convenient assay of P-450IIIA cytochromes in rats and in some patients.

Full text

PDF
688

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arlotto M. P., Sonderfan A. J., Klaassen C. D., Parkinson A. Studies on the pregnenolone-16 alpha-carbonitrile-inducible form of rat liver microsomal cytochrome P-450 and UDP-glucuronosyltransferase. Biochem Pharmacol. 1987 Nov 15;36(22):3859–3866. doi: 10.1016/0006-2952(87)90450-3. [DOI] [PubMed] [Google Scholar]
  2. Austin K. L., Mather L. E., Philpot C. R., McDonald P. J. Intersubject and dose-related variability after intravenous administration of erythromycin. Br J Clin Pharmacol. 1980 Sep;10(3):273–279. doi: 10.1111/j.1365-2125.1980.tb01755.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baker A. L., Kotake A. N., Schoeller D. A. Clinical utility of breath tests for the assessment of hepatic function. Semin Liver Dis. 1983 Nov;3(4):318–329. doi: 10.1055/s-2008-1040784. [DOI] [PubMed] [Google Scholar]
  4. Beaune P. H., Kremers P. G., Kaminsky L. S., De Graeve J., Albert A., Guengerich F. P. Comparison of monooxygenase activities and cytochrome P-450 isozyme concentrations in human liver microsomes. Drug Metab Dispos. 1986 Jul-Aug;14(4):437–442. [PubMed] [Google Scholar]
  5. Beaune P. H., Umbenhauer D. R., Bork R. W., Lloyd R. S., Guengerich F. P. Isolation and sequence determination of a cDNA clone related to human cytochrome P-450 nifedipine oxidase. Proc Natl Acad Sci U S A. 1986 Nov;83(21):8064–8068. doi: 10.1073/pnas.83.21.8064. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brüch M., Kling L., Legrum W., Maser E. The 14CO2 breath test: facilities and limitations of a rapid and noninvasive method for in vivo evaluation of modified hepatic cytochrome P-450--a critique. Arch Toxicol. 1987;60(1-3):81–85. doi: 10.1007/BF00296953. [DOI] [PubMed] [Google Scholar]
  7. Distlerath L. M., Reilly P. E., Martin M. V., Davis G. G., Wilkinson G. R., Guengerich F. P. Purification and characterization of the human liver cytochromes P-450 involved in debrisoquine 4-hydroxylation and phenacetin O-deethylation, two prototypes for genetic polymorphism in oxidative drug metabolism. J Biol Chem. 1985 Jul 25;260(15):9057–9067. [PubMed] [Google Scholar]
  8. Duane W. C., Levitt D. G., Mueller S. M., Behrens J. C. Regulation of bile acid synthesis in man. Presence of a diurnal rhythm. J Clin Invest. 1983 Dec;72(6):1930–1936. doi: 10.1172/JCI111157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Elshourbagy N. A., Guzelian P. S. Separation, purification, and characterization of a novel form of hepatic cytochrome P-450 from rats treated with pregnenolone-16 alpha-carbonitrile. J Biol Chem. 1980 Feb 25;255(4):1279–1285. [PubMed] [Google Scholar]
  10. Fonne-Pfister R., Bargetzi M. J., Meyer U. A. MPTP, the neurotoxin inducing Parkinson's disease, is a potent competitive inhibitor of human and rat cytochrome P450 isozymes (P450bufI, P450db1) catalyzing debrisoquine 4-hydroxylation. Biochem Biophys Res Commun. 1987 Nov 13;148(3):1144–1150. doi: 10.1016/s0006-291x(87)80252-8. [DOI] [PubMed] [Google Scholar]
  11. Fujino T., Park S. S., West D., Gelboin H. V. Phenotyping of cytochromes P-450 in human tissues with monoclonal antibodies. Proc Natl Acad Sci U S A. 1982 Jun;79(12):3682–3686. doi: 10.1073/pnas.79.12.3682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gonzalez F. J., Jaiswal A. K., Nebert D. W. P450 genes: evolution, regulation, and relationship to human cancer and pharmacogenetics. Cold Spring Harb Symp Quant Biol. 1986;51(Pt 2):879–890. doi: 10.1101/sqb.1986.051.01.101. [DOI] [PubMed] [Google Scholar]
  13. Gonzalez F. J., Skoda R. C., Kimura S., Umeno M., Zanger U. M., Nebert D. W., Gelboin H. V., Hardwick J. P., Meyer U. A. Characterization of the common genetic defect in humans deficient in debrisoquine metabolism. Nature. 1988 Feb 4;331(6155):442–446. doi: 10.1038/331442a0. [DOI] [PubMed] [Google Scholar]
  14. Gonzalez F. J., Song B. J., Hardwick J. P. Pregnenolone 16 alpha-carbonitrile-inducible P-450 gene family: gene conversion and differential regulation. Mol Cell Biol. 1986 Aug;6(8):2969–2976. doi: 10.1128/mcb.6.8.2969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Guengerich F. P., Martin M. V., Beaune P. H., Kremers P., Wolff T., Waxman D. J. Characterization of rat and human liver microsomal cytochrome P-450 forms involved in nifedipine oxidation, a prototype for genetic polymorphism in oxidative drug metabolism. J Biol Chem. 1986 Apr 15;261(11):5051–5060. [PubMed] [Google Scholar]
  16. Guengerich F. P., Müller-Enoch D., Blair I. A. Oxidation of quinidine by human liver cytochrome P-450. Mol Pharmacol. 1986 Sep;30(3):287–295. [PubMed] [Google Scholar]
  17. Gut J., Gasser R., Dayer P., Kronbach T., Catin T., Meyer U. A. Debrisoquine-type polymorphism of drug oxidation: purification from human liver of a cytochrome P450 isozyme with high activity for bufuralol hydroxylation. FEBS Lett. 1984 Aug 6;173(2):287–290. doi: 10.1016/0014-5793(84)80792-9. [DOI] [PubMed] [Google Scholar]
  18. Hardwick J. P., Song B. J., Huberman E., Gonzalez F. J. Isolation, complementary DNA sequence, and regulation of rat hepatic lauric acid omega-hydroxylase (cytochrome P-450LA omega). Identification of a new cytochrome P-450 gene family. J Biol Chem. 1987 Jan 15;262(2):801–810. [PubMed] [Google Scholar]
  19. Henry D. A., Sharpe G., Chaplain S., Cartwright S., Kitchingman G., Bell G. D., Langman M. J. The [14C]-aminopyrine breath test. A comparison of different forms of analysis. Br J Clin Pharmacol. 1979 Dec;8(6):539–545. doi: 10.1111/j.1365-2125.1979.tb01041.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hepner G. W., Vesell E. S. Assessment of aminopyrine metabolism in man by breath analysis after oral administration of 14C-aminopyrine. Effects of phenobarbital, disulfiram and portal cirrhosis. N Engl J Med. 1974 Dec 26;291(26):1384–1388. doi: 10.1056/NEJM197412262912605. [DOI] [PubMed] [Google Scholar]
  21. Heuman D. M., Gallagher E. J., Barwick J. L., Elshourbagy N. A., Guzelian P. S. Immunochemical evidence for induction of a common form of hepatic cytochrome P-450 in rats treated with pregnenolone-16 alpha-carbonitrile or other steroidal or non-steroidal agents. Mol Pharmacol. 1982 May;21(3):753–760. [PubMed] [Google Scholar]
  22. Hoensch H. P., Balzer K., Dylewizc P., Kirch W., Goebell H., Ohnhaus E. E. Effect of rifampicin treatment on hepatic drug metabolism and serum bile acids in patients with primary biliary cirrhosis. Eur J Clin Pharmacol. 1985;28(4):475–477. doi: 10.1007/BF00544371. [DOI] [PubMed] [Google Scholar]
  23. Hostetler K. A., Wrighton S. A., Kremers P., Guzelian P. S. Immunochemical evidence for multiple steroid-inducible hepatic cytochromes P-450 in the rat. Biochem J. 1987 Jul 1;245(1):27–33. doi: 10.1042/bj2450027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jacqz E., Hall S. D., Branch R. A. Genetically determined polymorphisms in drug oxidation. Hepatology. 1986 Sep-Oct;6(5):1020–1032. doi: 10.1002/hep.1840060534. [DOI] [PubMed] [Google Scholar]
  25. Kronbach T., Fischer V., Meyer U. A. Cyclosporine metabolism in human liver: identification of a cytochrome P-450III gene family as the major cyclosporine-metabolizing enzyme explains interactions of cyclosporine with other drugs. Clin Pharmacol Ther. 1988 Jun;43(6):630–635. doi: 10.1038/clpt.1988.87. [DOI] [PubMed] [Google Scholar]
  26. LEE C. C., ANDERSON R. C., CHEN K. K. The excretory products of N-methyl-C14-erythromycin in rats. J Pharmacol Exp Ther. 1956 Jul;117(3):274–280. [PubMed] [Google Scholar]
  27. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  28. Mahgoub A., Idle J. R., Dring L. G., Lancaster R., Smith R. L. Polymorphic hydroxylation of Debrisoquine in man. Lancet. 1977 Sep 17;2(8038):584–586. doi: 10.1016/s0140-6736(77)91430-1. [DOI] [PubMed] [Google Scholar]
  29. Mao J. C., Tardrew P. L. Demethylation of erythromycins by rabbit tissues in vitro. Biochem Pharmacol. 1965 Jul;14(7):1049–1058. doi: 10.1016/0006-2952(65)90033-x. [DOI] [PubMed] [Google Scholar]
  30. Molowa D. T., Schuetz E. G., Wrighton S. A., Watkins P. B., Kremers P., Mendez-Picon G., Parker G. A., Guzelian P. S. Complete cDNA sequence of a cytochrome P-450 inducible by glucocorticoids in human liver. Proc Natl Acad Sci U S A. 1986 Jul;83(14):5311–5315. doi: 10.1073/pnas.83.14.5311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nebert D. W., Adesnik M., Coon M. J., Estabrook R. W., Gonzalez F. J., Guengerich F. P., Gunsalus I. C., Johnson E. F., Kemper B., Levin W. The P450 gene superfamily: recommended nomenclature. DNA. 1987 Feb;6(1):1–11. doi: 10.1089/dna.1987.6.1. [DOI] [PubMed] [Google Scholar]
  32. OMURA T., SATO R. THE CARBON MONOXIDE-BINDING PIGMENT OF LIVER MICROSOMES. II. SOLUBILIZATION, PURIFICATION, AND PROPERTIES. J Biol Chem. 1964 Jul;239:2379–2385. [PubMed] [Google Scholar]
  33. Ryan D. E., Thomas P. E., Reik L. M., Levin W. Purification, characterization and regulation of five rat hepatic microsomal cytochrome P-450 isozymes. Xenobiotica. 1982 Nov;12(11):727–744. doi: 10.3109/00498258209038947. [DOI] [PubMed] [Google Scholar]
  34. Schuetz E. G., Wrighton S. A., Safe S. H., Guzelian P. S. Regulation of cytochrome P-450p by phenobarbital and phenobarbital-like inducers in adult rat hepatocytes in primary monolayer culture and in vivo. Biochemistry. 1986 Mar 11;25(5):1124–1133. doi: 10.1021/bi00353a027. [DOI] [PubMed] [Google Scholar]
  35. Shreeve W. W., Shoop J. D., Ott D. G., McInteer B. B. Test for alcoholic cirrhosis by conversion of [14C]- or [13C]galactose to expired CO2. Gastroenterology. 1976 Jul;71(1):98–101. [PubMed] [Google Scholar]
  36. Thomas P. E., Bandiera S., Maines S. L., Ryan D. E., Levin W. Regulation of cytochrome P-450j, a high-affinity N-nitrosodimethylamine demethylase, in rat hepatic microsomes. Biochemistry. 1987 Apr 21;26(8):2280–2289. doi: 10.1021/bi00382a031. [DOI] [PubMed] [Google Scholar]
  37. Wang P. P., Beaune P., Kaminsky L. S., Dannan G. A., Kadlubar F. F., Larrey D., Guengerich F. P. Purification and characterization of six cytochrome P-450 isozymes from human liver microsomes. Biochemistry. 1983 Nov 8;22(23):5375–5383. doi: 10.1021/bi00292a019. [DOI] [PubMed] [Google Scholar]
  38. Watkins P. B., Wrighton S. A., Maurel P., Schuetz E. G., Mendez-Picon G., Parker G. A., Guzelian P. S. Identification of an inducible form of cytochrome P-450 in human liver. Proc Natl Acad Sci U S A. 1985 Sep;82(18):6310–6314. doi: 10.1073/pnas.82.18.6310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Watkins P. B., Wrighton S. A., Schuetz E. G., Maurel P., Guzelian P. S. Macrolide antibiotics inhibit the degradation of the glucocorticoid-responsive cytochrome P-450p in rat hepatocytes in vivo and in primary monolayer culture. J Biol Chem. 1986 May 15;261(14):6264–6271. [PubMed] [Google Scholar]
  40. Waxman D. J., Attisano C., Guengerich F. P., Lapenson D. P. Human liver microsomal steroid metabolism: identification of the major microsomal steroid hormone 6 beta-hydroxylase cytochrome P-450 enzyme. Arch Biochem Biophys. 1988 Jun;263(2):424–436. doi: 10.1016/0003-9861(88)90655-8. [DOI] [PubMed] [Google Scholar]
  41. Wrighton S. A., Maurel P., Schuetz E. G., Watkins P. B., Young B., Guzelian P. S. Identification of the cytochrome P-450 induced by macrolide antibiotics in rat liver as the glucocorticoid responsive cytochrome P-450p. Biochemistry. 1985 Apr 23;24(9):2171–2178. doi: 10.1021/bi00330a010. [DOI] [PubMed] [Google Scholar]
  42. Wrighton S. A., Schuetz E. G., Watkins P. B., Maurel P., Barwick J., Bailey B. S., Hartle H. T., Young B., Guzelian P. Demonstration in multiple species of inducible hepatic cytochromes P-450 and their mRNAs related to the glucocorticoid-inducible cytochrome P-450 of the rat. Mol Pharmacol. 1985 Sep;28(3):312–321. [PubMed] [Google Scholar]
  43. Wrighton S. A., Thomas P. E., Willis P., Maines S. L., Watkins P. B., Levin W., Guzelian P. S. Purification of a human liver cytochrome P-450 immunochemically related to several cytochromes P-450 purified from untreated rats. J Clin Invest. 1987 Oct;80(4):1017–1022. doi: 10.1172/JCI113154. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES