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United States Department of Agriculture

Agricultural Research Service

Title: Development of Enzyme-Linked Immunosorbent Assays for the Hepatotoxic Alkaloids Riddelliine and Riddelliine N-Oxide

Authors
item LEE, STEPHEN
item Schoch, Thomas
item STEGELMEIER, BRYAN
item GARDNER, DALE
item Than, Khin - CSIRO
item Molyneux, Russell

Submitted to: Journal of Agricultural and Food Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 20, 2000
Publication Date: April 20, 2001
Citation: LEE, S.T., SCHOCH, T.K., STEGELMEIER, B.L., GARDNER, D.R., THAN, K.A., MOLYNEUX, R.J. DEVELOPMENT OF ENZYME-LINKED IMMUNOSORBENT ASSAYS FOR THE HEPATOTOXIC ALKALOIDS RIDDELLIINE AND RIDDELLIINE N-OXIDE. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY. 2001.

Interpretive Summary: Pyrrolizidine alkaloid containing plants are widely distributed throughout the world and are particularly common in the genus Senecio. Many pyrrolizidine alkaloids are toxic and cause poisoning in livestock and in humans. The type and concentrations of the alkaloids vary between plant species. In addition, within a species of plant, concentrations vary with environment and location. Rapid, sensitive and specific diagnostic techniques are needed to identify poisoned animals and to determine the particular plants and conditions under which livestock are likely to be poisoned. In this study, two competitive inhibition enzyme-lined immunosorbent assays (CI-ELISA) for riddelliine, riddelliine N oxide and other closely related pyrrolizidine alkaloids were developed. One assay is class specific toward the free base forms of the pyrrolizidine alkaloids, the other assay showed cross-reactivity to both the free base and N-oxide forms of the alkaloids. The assay with the lowest limit of detection had limit of detection of 47.5 pg for riddelliine. Studies using these assays demonstrated that riddelliine could be detected in blood and allowed us to estimate the total pyrrolizidine alkaloid content in Senecio riddellii/alfalfa mixtures. These findings suggest that these techniques will be excellent tools to diagnose poisoned animals and identify highly toxic plants.

Technical Abstract: Pyrrolizidine alkaloid containing plants are widely distributed throughout the world and are particularly common in the genus Senecio. The structural types and concentrations of the alkaloids vary between plant species. In addition, within a species of plant, concentrations vary with environment and location. Many pyrrolizidine alkaloids are toxic and cause poisoning in livestock and in humans. Rapid, sensitive and specific diagnostic techniques are needed to identify poisoned animals and to determine the particular plants and conditions under which livestock are likely to be poisoned. In this study, two competitive inhibition enzyme-lined immunosorbent assays (CI-ELISA) for riddelliine, riddelliine N oxide and other closely related pyrrolizidine alkaloids were developed using polyclonal antibodies. One assay is class specific toward the free base forms of the pyrrolizidine alkaloids, the other assay showed cross- reactivity to both the free base and N-oxide forms of the alkaloids. The assay with the lowest limit of detection had an I50 of 803.9 pg with a limi of detection of 47.5 pg for riddelliine. Spike and recovery studies for riddelliine in bovine blood ranged from 45-47%. The assay that showed cross-reactivity between the N-oxide and free base forms of the pyrrolizidine alkaloids allowed us to estimate the total pyrrolizidine alkaloid content in Senecio riddellii in a mixture with alfalfa. These findings suggest that these techniques will be excellent tools to diagnose poisoned animals and identify highly toxic plants.

Last Modified: 9/29/2014
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