Endothelin-1 assay

This assay procedure has been applied to various haptens having a primary amino group, including leukotriene C4 (VI), T4 (P2), substance P (P2), endothelin (P2), and AGII (G2) (for structures, see Fig. 5A and 5C). These noncompetitive assays were 10-300-fold more sensitive than corresponding competitive assays (G2). The minimal detectable concentrations described in these papers were as follows 2 pg/ml ( 0.3 fmol/assay) for leukotriene C4, 14 pg/ml ( 0.2 fmol/assay) for T4, 6 pg/ml ( 0.4 fmol/assay) for substance P, and 20 pg/ml ( 0.8 fmol/assay) for endothelin. 

Plumpton, C., Kalinka, S., Martin, R.C., Horton, J.K., and Davenport, A.P. (1994). Effects of phosphoramidon and pepstatin-a on the secretion of endothelin-1 and big endothelin-1 by human umbilical vein endothelial-cells—measurement by 2-site enzyme-linked immunosorbent assays. Clin. Sci., 87, 245-251. 

The sterol sulfate, halistanol disulfate B (544) was isolated from a South African Pachastrella sp. The structure and stereochemistry of compound 544 were established mainly by interpretation of spectral data. Halistanol disulfate B (544) was active in the endothelin converting enzyme (ECE) assay at a micromolar concentration [451]. Three sterol trisulfates (545-547) have been isolated from the sponges Trachyopsis halichondrioides and Cymbastela coralliophila [452]. 

The use of specific quenched fluorescent substrates (QFS) provides a rapid and sensitive method to measure peptidase activity, and is readily adaptable to high-throughput screening of potential peptidase inhibitors. In this chapter, we discuss general considerations for the development of QFS assays, and describe in detail an assay protocol for the mammalian metallopeptidase, endothelin-converting enzyme. 

WS009 A (220, = FR 901366) and B (221, = FR 901367), isolated from Strep-tomyces sp. No. 89009, were discovered during a screening course for endothelin receptor antagonists using ET-1, the most potent vasoconstrictor to date [160-162]. These compounds show a selective activity in an endothelin binding assay (IC50 5.8 x 10 M and 6.7 x 10" M, respectively). The structures feature a unique C-6 a linked N-acetylcysteine moiety which may have arisen biosyn-thetically via an attack of cysteine on a 6a -12 a epoxide structure, such as exemplified in A-683 (210) or SF 2315 B or elmycin C [3]. 

The first compound with published detailed information which supports the claims for a specific ETg antagonist is IRL 1038, (20) [Cys -Cys ] ET-1 (11-21). This compound showed specific antagonist activity both in binding assays and in endothelin-induced contraction models. With ET-3 as agonist, 3 /M IRL 1038 antagonized the ETb receptor-mediated contraction of guinea-pig ileal and tracheal smooth muscle but did not affect the ET receptor-mediated contraction of rat aortic tissue [110]. 

Most of the other compounds which have been discovered to have endothelin antagonist properties were identified as a result of the random screening of natural products in endothelin binding assays. 

Claims that compounds with activity in the angiotensin area also inhibit the actions of endothelin have appeared in the literature [157]. The only compounds patented for both these properties are three series of compounds from Roussel-Uclaf [158-160]. Details of activity are shown for only one compound (36) which has very weak activity, IC50 = 14,000 /tM in the rat cortical membrane binding assay. It is claimed to be active at 1 mg/kg i.v. in the rat against an ET-l-induced vasoconstriction [160] which could... 

We developed a simultaneous bioluminescent assay of acetate kinase (AK) and pyruvate phosphate dikinase (PPDK). In the method, the detection limits (blank + 3SD) of AK and PPDK were 1.03x10 and 2.05x10 mol/assay, respectively. Previously, we successfully applied a tandem bioluminescent enzyme immunoassay (BL-EIA) for simultaneous measurement of insulin and c-peptide. In this study, we also applied the method to tandem BL-EIA for Angiotensin I and Endothelin-1, which are hypertension related peptides. The tandem BL-EIA used a competitive immuno-reaction for Angiotensin I and sandwich inununo-reaction for Endothelin-1. 

Using the proposed BL-EIA, the measurable range for Angiotensin I and Endothelin-1 were 7.81 - 1000 pg/mL and 15.63 - 1000 pg/mL, respectively. The intra-assay coefficients of variation of Angiotensin I and Endothelin-1 at each standard point were below 9.7% and 11%, respectively. (Fig. 2)... 

In an in vitro assay for antithrombin activity, a methylene chloride extract fraction of M. cerifera showed greater than 80% activity. Myricitrin has shown choleretic, bactericidal, and paramecicidal activities myricadiol has shown mineralocorticoid activity. Myriceric acid A is a selective endothelin A receptor antagonist. ... 

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