Liver cytochrome P450 system as affected by endophyte-infected tall fescue seed extracts and ergot alkaloids
ABSTRACT
Endophyte infected tall fescue (E+) is the base diet for nearly all beef cattle in the southern USA. It has been linked to a variety of toxicological conditions due to the presence of large numbers of ergot alkaloids. This study was designed to investigate the effects of E+ seed extract and selected ergot alkaloids on the detoxification pathway by cytochrome P450 (CYP3A4) enzyme system. Tests were performed using the P450- Glo CYP3A4 enzyme activity kit (Promega, WI), according to the manufacturer’s manual. Luminescence was measured using a single tube TD20/20 luminometer. Endophyte infected tall fescue seed was extracted with 50/50 methanol/25 mM ammonium carbonate, cleaned and concentrated on Strata-X reversed phase column (Phenomenex). The extracts were evaluated on an HPLC, and then tested using a serial dilution method. Commercially available ergonovine (EN), ergocorine (ER), bromocryptine (BC) and ergocryptine (EC) were tested individually using 0 to 44 nM concentrations. Seed extract of E+ produced a significant (P < 0.05) dose dependent inhibition of CYP3A4 enzyme activity similar to that produced by the commercially available ergot alkaloids EC, ER, BC and EN which inhibited CYP3A4 enzyme activity in a significant (P < 0.05) dose dependent manner with EC being most potent, followed by ER, BC, and then EN (70%, 40%, 30% and 10% at 44 nM concentration). The similarity of the inhibition curves of seed extract to that of the commercially available ergot alkaloids suggests a related mode of action and that the use of such ergot alkaloids and CYP3A4 assay is a good model to study the toxicity of tall fescue. Furthermore, it provides the foundation to identify the individual toxic components of purified endophyte infected tall fescue extract.
Endophyte infected tall fescue (E+) is the base diet for nearly all beef cattle in the southern USA. It has been linked to a variety of toxicological conditions due to the presence of large numbers of ergot alkaloids. This study was designed to investigate the effects of E+ seed extract and selected ergot alkaloids on the detoxification pathway by cytochrome P450 (CYP3A4) enzyme system. Tests were performed using the P450- Glo CYP3A4 enzyme activity kit (Promega, WI), according to the manufacturer’s manual. Luminescence was measured using a single tube TD20/20 luminometer. Endophyte infected tall fescue seed was extracted with 50/50 methanol/25 mM ammonium carbonate, cleaned and concentrated on Strata-X reversed phase column (Phenomenex). The extracts were evaluated on an HPLC, and then tested using a serial dilution method. Commercially available ergonovine (EN), ergocorine (ER), bromocryptine (BC) and ergocryptine (EC) were tested individually using 0 to 44 nM concentrations. Seed extract of E+ produced a significant (P < 0.05) dose dependent inhibition of CYP3A4 enzyme activity similar to that produced by the commercially available ergot alkaloids EC, ER, BC and EN which inhibited CYP3A4 enzyme activity in a significant (P < 0.05) dose dependent manner with EC being most potent, followed by ER, BC, and then EN (70%, 40%, 30% and 10% at 44 nM concentration). The similarity of the inhibition curves of seed extract to that of the commercially available ergot alkaloids suggests a related mode of action and that the use of such ergot alkaloids and CYP3A4 assay is a good model to study the toxicity of tall fescue. Furthermore, it provides the foundation to identify the individual toxic components of purified endophyte infected tall fescue extract.
Cite this paper
Moubarak, A. , Johnson, Z. and Rosenkrans Jr., C. (2012) Liver cytochrome P450 system as affected by endophyte-infected tall fescue seed extracts and ergot alkaloids. Agricultural Sciences, 3, 1-4. doi: 10.4236/as.2012.31001.
Moubarak, A. , Johnson, Z. and Rosenkrans Jr., C. (2012) Liver cytochrome P450 system as affected by endophyte-infected tall fescue seed extracts and ergot alkaloids. Agricultural Sciences, 3, 1-4. doi: 10.4236/as.2012.31001.
References
[1] Porter, T.D. and Coon, M.J. (1991) Cytochrome P-450. Multiplicity of isoforms, substrates, and catalytic and regulatory mechanisms. Journal of Biological Chemistry, 266, 13469-13472. [2] Pollock, B.G. (1994) Recent developments in drug metabolism of relevance to psychiatrists. Harvard Review of Psychiatry, 2, 204-213. doi:10.3109/10673229409017138 [3] Ball, S.E., Maurer, G., Zollinger, M., Ladona, M. and Vicker, A.E. (1992) Characterization of the cytochrome P-450 gene family responsible for the N-dealkylation of the ergot alkaloid CQA 206-291 in humans. Drug Metabolism and Disposition, 20, 56-63. [4] Moochhala, S.M., Lee, E.J., Hu, G.T., Koh, O.S. and Becket, G. (1989) Effects of bromocryptine on hepatic cytochrome P-450 monooxygenase system. Japanese Journal of Pharmacology, 49, 285-291. doi:10.1254/jjp.49.285 [5] Witkamp, R.F., Nijmeijer, S.M., Monshouwer, M. and Van Miert, A.S. (1995) The antibiotic tiamulin is a potent inducer and inhibitor of cytochrome P4503A via the formation of a stable metabolic intermediate complex. Studies in primary hepatocyte cultures and liver microsomes of the pig. Drug Metabolism and Disposition, 23, 542- 547. [6] Moubarak A.S., Rosenkrans, C.F. Jr. and Johnson, Z.B. (2003) Modulation of Cytochrome P450 Metabolism by Ergonovine and Dihydroergotamine. Veterinary & Human Toxicology, 45, 6-9. [7] Strickland, J.R, Cross, D.L, Jenkins, T.C., Petroski, R.J. and Powell, R.G (1992) The effect of alkaloids and seed extracts of endophyte-infected tall fescue on prolactin secretion in an in vitro rat pituitary perfusion system. Journal of Animal Science, 70, 2779-2786.
[1] Porter, T.D. and Coon, M.J. (1991) Cytochrome P-450. Multiplicity of isoforms, substrates, and catalytic and regulatory mechanisms. Journal of Biological Chemistry, 266, 13469-13472. [2] Pollock, B.G. (1994) Recent developments in drug metabolism of relevance to psychiatrists. Harvard Review of Psychiatry, 2, 204-213. doi:10.3109/10673229409017138 [3] Ball, S.E., Maurer, G., Zollinger, M., Ladona, M. and Vicker, A.E. (1992) Characterization of the cytochrome P-450 gene family responsible for the N-dealkylation of the ergot alkaloid CQA 206-291 in humans. Drug Metabolism and Disposition, 20, 56-63. [4] Moochhala, S.M., Lee, E.J., Hu, G.T., Koh, O.S. and Becket, G. (1989) Effects of bromocryptine on hepatic cytochrome P-450 monooxygenase system. Japanese Journal of Pharmacology, 49, 285-291. doi:10.1254/jjp.49.285 [5] Witkamp, R.F., Nijmeijer, S.M., Monshouwer, M. and Van Miert, A.S. (1995) The antibiotic tiamulin is a potent inducer and inhibitor of cytochrome P4503A via the formation of a stable metabolic intermediate complex. Studies in primary hepatocyte cultures and liver microsomes of the pig. Drug Metabolism and Disposition, 23, 542- 547. [6] Moubarak A.S., Rosenkrans, C.F. Jr. and Johnson, Z.B. (2003) Modulation of Cytochrome P450 Metabolism by Ergonovine and Dihydroergotamine. Veterinary & Human Toxicology, 45, 6-9. [7] Strickland, J.R, Cross, D.L, Jenkins, T.C., Petroski, R.J. and Powell, R.G (1992) The effect of alkaloids and seed extracts of endophyte-infected tall fescue on prolactin secretion in an in vitro rat pituitary perfusion system. Journal of Animal Science, 70, 2779-2786.