ARS/SCA: Tall Fescue Characteristics

Ergot alkaloid purification We have the capability to extract tall fescue alkaloids from powdered seed in 30 cm x 80 cm columns and in large tube extractor (300+ lbs each batch) with 80% ethanol as the first step in purification. With removal of the ethanol and water the product yields about a 20-fold increase in ergovaline concentration. Further purification by removal of lipids from a water solution and extraction from the water yields about a 400-fold increase in ergovaline concentration with about a 50% recovery. The partially purified ergovaline from seed to this point has biological activities in vivo and in vitro bovine bioassays that approximate that of synthetic ergovaline. This basic protocol continues to be modified to utilize less organic solvents. It is this product that will be further purified on the Waters autopurification system consisting of a Waters binary gradient module, sample manager and a photodiode array detector. In addition, we will extract ergotamine and ergocryptine from Claviceps purpurea sclerotia. There are 3 other ergot alkaloids in the sclerotia but we will extract and partially purify these main alkaloids initially. We have the sclerotia in the laboratory. In addition we will (i) continue to cooperate as we have in the recent past with the groups in the College and FAPRU. However, the laboratory cannot do an infinite number of analysis or different types of analysis. (ii) do lysergic acid, ergovaline, ergovalinine, ergotamine, ergotaminine, N-acetylloline, N-formylloline and N-acetylnorloline determinations of forage and seed as necessary. Additional analyses could be added as demand and as results are validated. The first protocol to be added will be peramine. We have measured the pyrrolizidne alkaloids in the soil but have not detected ergovaline. To accomplish these activities the lab has two HPLC’s with florescence and uv/vis detectors for ergot and some of the diazaphrenthrene alkaloid analysis. HPLC/ms will be used for some analysis, especially minor ergot alkaloids and peramine. Also, the lab has gc’s with FID and ms detectors for the pyrrolizidine alkaloid analysis. Ergot alkaloid purification for analytical standards and small amounts for cell culture bioassays will be done on our Waters autopurification system. We propose to purify ergovaline and ergovalinine initially and then ergotamine and ergocryptine as they are required for analytical purposes