Isolation and Properties of Porcine Thyrotropin-releasing Hormone

Abstract Isolation on a preparative scale of thyrotropin-releasing hormone from porcine hypothalami is described. The yields from two batches of 100,000 and 165,000 hypothalami were 2.8 mg and 4.4 mg, respectively. The material with thyrotropin-releasing hormone activity, calculated to be purified about 570,000 times, was active in mice at doses of 1 ng in vivo, and stimulated the secretion of thyroid-stimulating hormone from rat pituitaries in vitro at doses of 10 pg. Thin layer chromatography, electrophoresis, and paper chromatography indicated that the material was homogeneous. Histidine, glutamic acid, and proline, which were present in an equimolar ratio, accounted for about 32% of the dry weight of the active material. However, eight synthetic peptides containing histidine, proline, and glutamic acid or glutamine had no thyrotropin-releasing hormone activity at doses as large at 10 µg. Biological activity of thyrotropin-releasing hormone was abolished by diazotized sulfanilic acid, N-bromosuccinimide, or acid hydrolysis, but was not affected by periodate or by incubation with proteolytic enzymes.