Vitellogenin induction by a mixture of steroidal estrogens in freshwater fishes and relevant risk assessment.

The study method on combined effects of environmental contaminant mixture and ecological risk assessment was discussed. Batch tests were conducted to assess the in vivo potency of binary mixtures of estrogens using plasma vitellogenin concentrations in male crucian carp as the endpoint. The estrogenic potencies of 17beta-estradiol (E(2)) and 17alpha-ethynylestradiol (EE(2)) were determined following 14 day exposure to the individual chemicals and equipotent binary mixtures. A Nonlinear regression was obtained and 95% confidence limits of effect concentration were achieved using the bootstrap method. Concentration-response curve for fixed ratio binary mixtures of E(2) and EE(2) was compared with those for individual chemicals, using the biomathematical models of concentration addition (CA) and independent action (IA). A complete overlap was found for the CA predictions with the 95% confidence interval of the best-fit regression line of the observed responses, and the IA predictions was shown lower than the observations. The observed mixture effects were considerably higher than those of the hormone alone and far exceeded the 95% confidence interval of the estrogen regression lines. The predicted effects of binary mixtures at different mixture ratios indicated that the potential impact of components on mixture would depend predominantly on its concentration, the mixture ratio and its relative potency. Results suggested that E(2) and EE(2) acted together in an additive manner and the combined effects can be accurately predicted in whole range of exposure concentration by the models of CA and IA, the model of CA might be realistic, but more useful for ecological risk assessment.

CD spectrometric methods for the simultaneous determination of ethisterone and its delta5-isomer.

Quick and accurate direct and indirect circular dichroism (CD) spectrometric methods were developed for the simultaneous determination of ethisterone (17alpha-ethinyl-17-hydroxy-4-androstene-3-one) and its delta(5)-isomer (delta(5)-ethisterone). The direct method is based on the selective negative Cotton effect of the delta(4)-3-oxo group in ethisterone (negative maximum at 348 nm in dioxan) and measurement of the ellipticity at 296 nm (positive maximum of delta(5)-ethisterone), where the measured ellipticity is the sum of those of the two isomers. In the indirect procedure delta(5)-ethisterone is transformed to ethisterone by base-catalysed isomerization and the ellipticities are measured at 339 nm in ethanol before and after isomerization. Preliminary experiments show the usefulness of CD detector in the HPLC determination of the mixture of the isomers. A major advantage of the direct CD spectrometric and the HPLC/CD methods is that the delta(5)-isomer with extremely low UV activity can also be directly measured with high sensitivity.

Analysis of steroids Part 50. Derivatization of ketosteroids for their separation and determination by capillary electrophoresis.

4-Ene-3-ketosteroids and 17-ketosteroids were quantitatively transformed into the corresponding hydrazones using Girard P and T reagents, respectively. The positively charged derivatives were separated by capillary electrophoresis. The spectrophotometric characteristics of the derivatives permitted their sensitive detection in the 230-280 nm range. The steroids investigated included nortestosterone and its phenylpropionate, norethisterone and its oenanthate, d,l-norgestrel, dehydroepiandrosterone, androstenedione and ethisterone.

[Analysis of steroids. Part 42. By-products of the ethynylation of 17-ketosteroids (isolation, identification and determination)]. / Adatok a szteránvázas vegyületek analitikájához 42. 17-Ketoszteroidok etinilezésének melléktermékei (elválasztás, azonosítás, meghatározás).

The therapeutically very important 17 alpha-ethynyl steroids are prepared from 17-keto steroids by means of addition of acetylene. Two important side reactions of this procedure are known: the formation of the isomeric beta-ethynyl derivative and the formation of a dimeric product with acetylene bridge. The aim of this paper is to approach this problem from the point of view of impurity profiling of 17 alpha-ethynyl steroids (norethisterone, ethisterone, norgestrel and delta 9(11)-ethisterone) i.e. isolation, identification and quantification of the above mentioned by-products as impurities in the bulk drugs. Capillary gas chromatography is an ideal tool for the separation and quantitative determination of the beta-ethynyl derivatives (20 m long fused silica capillary, I.D 0.2 mm; stationary phase Ultra-2: 5% phenylmethyl silicon gum phase with a film thickness of 0.33 mu; column temperature 240-250 degrees C). The dimeric impurity cannot be determined directly by gas chromatography as it decomposes in the flash heater to the 17 alpha-ethynyl and the 17-keto derivatives. For this reason reversed-phase HPLC was preferred for their separation and quantitation; (column: 250 x 4 mm LiChrosorb RP-18, 10 microns; eluent methanol-water 7:3; UV detector 240 nm). The HPLC method is suitable for the separation and determination of the beta-ethynyl impurities, too. The chemical shifts of the protons and carbon atoms in the vicinity of C-17 in the 1H and 13C NMR spectra of the epimeric 17-ethynyl steroids greatly depend on the configuration of the ethynyl group and for this reason they (especially that of the C-18 are eminently suitable for the characterization of the isomers.(ABSTRACT TRUNCATED AT 250 WORDS)