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The Candida infected, embryonated hen's egg is a realistic complement for the model of the Candida infected mouse and can be used in the search for new systemically active antimycotics. This alternative method is rapid, sensitive convincing and inexpensive. The use of the embryonated hen's egg in an anti-Candida screening can reduce the use of small laboratory animals to a considerable amount. Thus, with the help of this new method pain and suffering of animals can be reduced in a part of the biologic-medical research in the sense of the animal protection law.

Plasma dehydroepiandrosterone (DHEA) and DHEA sulfate levels were determined by an enzyme immunoassay in male and female Sprague-Dawley rats fed a diet containing 0.6% or 0.3% DHEA. A significant difference in DHEA plasma levels was observed in male and female animals. In male rats, total DHEA plasma concentrations were found in the range of 0.4-1.5 microgram/ml (0.6% DHEA chow) and 0.3-0.5 microgram/ml (0.3% DHEA chow). In female animals several times higher DHEA plasma levels were determined: 17.5-33 micrograms/ml (0.6% DHEA chow) and 8.3-14.8 micrograms/ml (0.3% DHEA chow). DHEA was present in rat plasma of both sexes preferably as the sulfate conjugate. Significant sex differences were also found in the DHEA metabolite patterns obtained by TLC separation of extracts from plasma, bile and urine following administration of 3H-DHEA. In female rats, DHEA is present predominantly as the sulfate conjugate in considerable amounts in all materials investigated, whereas in male rats polar metabolites dominate in the patterns.

Out of the group of 2-amino-oxazoles 1 was found to be the most potent antiviral compound. Following p.o. or s.c. administration to rats, the 14C-labeled 1 was quickly and completely absorbed. The TRA was eliminated mainly via the kidneys and the liver with half-lives between 32 and 42 h. The acute pharmacodynamic effects of 1 were decrease of blood pressure, bradycardia, and inhibition of both gastric emptying and acid secretion. On smooth muscles spasmolytic and alpha-anti-adrenergic actions were predominant. After single administration the following MTD's were determined: 30 (mouse), 20 (rat), 10 mg/kg i.v. (pig), and 500 (mouse, rat), greater than 100 mg/kg p.o. (pig), respectively. In a subchronic toxicity study in rats, oral doses of 1 between 15 and 240 mg/kg given daily for 4 weeks were tolerated without any severe alterations related to the drug.

The preparation of the 17 beta-carbamoyl derivatives 4 and of the 17 beta-carbonic acid esters 5 from 15 beta,16 beta-epoxy-3-methoxyoestra-1,3,5(10)-triene-17 beta-ol (3a), and the subsequent opening of the 15 beta,16 beta-epoxy group resulting in 15 beta,16 alpha,17 beta-trisubstituted oestratrienes 6 are described. The influences of several substitutents in the positions 15 and 16 and of the 17 beta-hydroxy group and derivatives thereof on the uterotropic and cholesterol-lowering activities were investigated in rats after oral application. A correlation between the cholesterol-lowering and the uterotropic activity could be established. Activity dissociation was not achieved.

A number of drugs including steroid hormones undergo enterohepatic circulation (EHC) which influences the drug disposition parameters. EHC of drugs leads to prolonged drug exposition which may enhance the risk of liver incompatibility. The extent of EHC expressed by the reabsorption rate of a given drug is of interest from the clinical and toxicological point of view. A two-compartment model with an additional time lag was realized by a hybrid computer system to study the influence of EHC on the shape of plasma concentration-time profiles and pharmacokinetic parameters. Reabsorption rates of potential steroid drugs were calculated by model-simulation and the results compared with the experimentally found ones. Although the lag time model is only a simplified approximation to the underlying physiological processes it reflects sufficiently the pharmacokinetic profile of steroid drugs subject to EHC.

Disposition and excretion of the progestin Dienogest (17 alpha-cyanomethyl-17 beta-hydroxy-estra-4,9-dien-3-one, STS 557) were investigated in female rabbits. Following single and repeated administration of the tritium-labelled compound the plasma concentration courses of total radioactivity (Dienogest + metabolites) and of the parent drug alone were estimated and also the urinary and fecal excretion of total radioactivity. From these data basic pharmacokinetic parameters were calculated. Additionally, the enterohepatic recirculation of biliary excreted metabolites was studied using bile of donors for oral administration to recipients. Following oral administration the high bioavailability of Dienogest which was already found in other animal species and in man could also be confirmed with rabbits. The parameters of Dienogest disposition do not differ significantly from those of the progestin levonorgestrel. Thus, the different effects of the both progestins in the McPhail-Clauberg assay in the rabbit cannot be attributed to differences in pharmacokinetics.

Following administration of 14 alpha, 15 alpha-3H-Dienogest (STS 557, 17 alpha-cyanomethyl-17 beta-hydroxy-4, 9-estradien-3-one) to female rabbits, extracts from urine, bile and plasma were separated by means of HPLC. Urinary and biliary metabolites are characterized by patterns of high complexity. From the mass spectra and UV absorption data of the urinary Dienogest metabolites a variety of biotransformation reactions has been derived like: hydroxylation in different positions of the Dienogest molecule, among these the 11-position; reduction of the 3-oxo group to 3-hydroxy; introduction of 2 and 4 hydrogen atoms; aromatization of ring A; transformation of 17 alpha-CH2CN to CH2OH, and formation of compounds with a 5(10), 9(11)-diene structure. Some of these reactions occur simultaneously resulting in a very complicated metabolite spectrum. Possibly the multiple effects of Dienogest established in animals are partially caused by metabolites.

In mice and rabbits of both sexes the acute toxicity of STS 557 (17 alpha-cyanomethyl-17 beta-hydroxy-estra-4, 9-dien-3-one) was determined after its oral or parenteral (i.p., s.c.) administration. In rabbits increasing lethality was observed following STS 557 suspended in tylose solution at the dose range of 1.0 to 3.0 g/kg p.o. or i.p. The approximate LD50-values were 1.0 to 1.5 g/kg for the i.p. route and 1.0 to 2.0 g/kg for the oral route. Levonorgestrel injected i.p. did not cause any lethality up to the dose of 3.0 g/kg. After oral or s.c. administration to mice, doses of 4.0 g/kg STS 557 were well tolerated. A dose-related toxicity occurred only after i.p. doses between 0.5 and 1.0 g/kg (STS 557), and between 2.0 and 4.0 g/kg (levonorgestrel), respectively. Using an oily vehicle for the oral route in mice, the lethal threshold dose for STS 557 was lowered to about 2.0 g/kg. In conclusion, a low oral acute toxicity was determined for STS 557 corresponding to that of other progestagens like levonorgestrel or norethisterone.

The toxicity of 17 alpha-cyanomethyl-17 beta-hydroxy-estra-4, 9-dien-3-one (STS 557) was studied by its oral administration of 0.1, 1.0 or 10.0 mg/kg/day to Wistar rats for six months, and of 0.01, 0.1 or 1.0 mg/kg/day to beagle dogs for six months, respectively. Levonorgestrel at a dose of 1.0 mg/kg/day was used as the standard in the dog study. With respect to the progestational activity of the compound the main target organs were the hypophysis, the reproductive organs and the adrenals. Mammary hyperplasia was observed in dogs treated with STS 557 or levonorgestrel at the dose of 1.0 mg/kg/day, but in no case mammary nodules could be detected. At the dose of 1.0 mg/kg/day STS 557 and levonorgestrel were found to increase the plasma insulin response to i.v. glucose in bitches, but neither the mean blood glucose levels nor the glucose utilization were affected. Moreover, during administration of both steroids to dogs temporary changes in serum concentrations of triglycerides and total cholesterol were noted. The results obtained in rats and dogs from functional and morphological investigations did not reveal any toxic side effects of STS 557 on the liver, the kidneys, the bone marrow or on blood coagulation. The effects on the reproductive organs observed following STS 557 especially in dogs are related to both the hormonal effects of the compound and the specific response of the dog to potent progestagens.

The influence of progesterone (P) and synthetic gestagens given alone or in combination with estradiol (E2) on triglyceride (TG) concentration in serum of adult female rats was studied. Norethisterone acetate (NEA), levonorgestrel (LNG), dienogest (DEG), chlormadinone acetate (CMA) and P were administered orally at a dose of 10 mg/kg for three times. E2 was given chronically by s.c. implants. Synthetic gestagens and P were without any effect on the TG serum concentrations but E2 elevated the TG level by 168%. This E2-induced TG increase was not reversed by synthetic gestagens or P given orally but only by P s.c. after combined treatment with E2 plus gestagens. In a second experiment the hepatic TG release into the blood was studied using the model of Triton WR-1339 induced hypertriglyceridemia. E2 as well as the synthetic gestagens stimulated the TG release while the natural P failed to produce this effect. Following treatment with E2 plus gestagens, the E2 stimulated TG release was not significantly influenced by the gestagens. It is concluded that both the hepatic TG release into and the TG removal from the circulation may be stimulated by gestagens.

In female rats the influence of progestins of the estrane type [norethisterone acetate (NEA), norgestrel (NG) and dienogest (DEG)] and of the pregnane type [chlormadinone acetate (CMA), progesterone (P)] on total cholesterol (TC) and high-density-lipoprotein cholesterol (HDL-C) in serum has been investigated in combination with estradiol (E). E increased TC (45%) and HDL-C (12%). NEA, NG and DEG alone decreased TC maximally by 58% and HDL-C maximally by 67%, CMA and P were without influence. In combination with E, the gestagens NEA, NG and DEG suppressed the E-induced increase of TC and HDL-C, whereas CMA and P did not counteract the TC increase. The effects of estrogen/gestagen combinations on HDL-C in the serum of rats are similar to those in men, but species differences in the lipoprotein metabolism must be taken into consideration.

This paper reports on pharmacological properties of 17 alpha-cyano-methyl-17 beta-hydroxy-estra-4, 9-dien-3-one (STS 557), which were obtained from basic screening investigations. At high doses (100 mg/kg i.p.) STS 557 was sedatively active on the CNS, but did not show any narcotic or anticonvulsant activities. A hypothermic effect of STS 557 was found at non-sedative i.p. or oral doses in mice and rats. This was also shown for the standard levonorgestrel, doubtlessly to a lesser extent. The cardiovascular system as well as the renal excretion of urine and electrolytes were unaffected by the test compound. STS 557 was found to stimulate the spontaneous motility of the isolated rat uterus dose-dependently; an antiprogesteronic activity seems to be unlikely. STS 557 did not show any glucocorticoid or antiexudative properties. At oral dose levels of 10 and 50 mg/kg (X 4 days), STS 557 and levonorgestrel caused similar changes in the serum lipids of normolipidemic and hyperlipidemic rats. The concentration of total cholesterol was decreased, that of the FFA was increased. The triglycerides were lowered only in hyperlipidemic animals. All the pharmacological effects of STS 557 appeared at high doses, much higher than te proposed therapeutic ones.

Disposition and excretion of the estrogen derivative STS 593 [1,3-methoxy-14 beta,15 beta-methylenestra-1,3,5(10)-triene-17 beta-ol] with antifertility activity were investigated in Beagle dogs after single and repeated administration of the tritium-labelled compound. The plasma concentration vs. time courses of total radioactivity (TRA) and of the parent drug alone were estimated in conscious animals simultaneously with the urinary and fecal excretion data. From this the basic pharmacokinetic parameters were calculated and experimentally confirmed by multiple-dosing data. STS 593 differs from estradiol with respect to its higher bioavailability after oral administration; it amounts to 34-62% depending on the administration formulation. The compound is metabolized rapidly. The accumulation factor of TRA resulting from a multiple once-a-day dose regimen of STS 593 was quantified with 3.1.

Four newly synthesized estra-1,3,5(10)-triene-derivatives, --3-methoxy-17 alpha-cyanomethyl-estra-1,3,5(10)-triene-17 beta-ol (I), --3-methoxy-17 alpha-rhodanomethyl-estra-1,3,5(10)-triene-17 beta-ol-tetrahydropyranylether (II), --17 beta-(N',N'-dimethylhydrazinocarbonyloxy)-estra-1,3,5(10)-triene-3-ol (III), --3-methoxy-17 beta-(N-phenylaminocarbonyloxy)-estra-1,3,5(10)-triene (IV), were studied in subchronic toxicity tests in female rats. The drugs were given orally and compared to mestranol (V) which was used as a reference. Special histopathologic examination was performed in each rat with respect to pituitary morphology. Different pituitary responses to the applied estrogens were found with regard to hyperplasia of the adenohypophysis and interstitial haemosiderosis: 17 beta-hydroxy-17 alpha-alkylsubstituted derivatives (I, II, V) were less effective than 17 beta-carbamyloxy-derivatives (III, IV). These findings may be related to differences in pituitary accumulation of the investigated compounds, indicating any disposition-effect-relationships.

In adult female rats, the effects of progesterone (P), norethisterone acetate (NEA), levonorgestrel (LNG), dienogest (DEG) and chlormadinone acetate (CMA) given alone at doses of 2.5 or 10 mg/kg p.o. or in combination with s.c. implanted estradiol (E2) on total cholesterol (TC), high-density-lipoprotein cholesterol (HDL-C), low-density-lipoprotein cholesterol (LDL-C) and very-low-density-lipoprotein cholesterol (VLDL-C) were investigated. Additionally, the lipoprotein pattern was determined using agarose gel electrophoresis. Synthetic gestagens derived from nortestosterone (NEA, LNG, DEG) lowered TC by reduction of HDL-C and LDL-C, whereas E2 induced an increase of these lipids. The decrease of HDL-C and LDL-C caused by the gestagens was also found in E2 pretreated rats. In contrast, the pregnane related CMA and P given alone did not diminish TC, HDL-C or LDL-C. But they partly reversed the enhancing effect of E2 on the HDL-C fraction following their simultaneous administration. The results suggest that the cholesterol lowering effects of gestagens are mediated rather via androgen than via progesterone receptors.

Following oral and i.v. administration of [14 alpha, 15 alpha-3H]-STS 557 to beagle dogs, baboons, rats and female volunteers, plasma level courses of total radioactivity and STS 557, and radioactivity excretion in urine and feces have been investigated. Bioavailability of orally administered STS 557 was found to be 80--90% in man and beagle dog, 70--80% in baboon and rat. Concerning the systemic availability following oral administration of equivalent doses, the following order was established: beagle dog greater than man greater than baboon greater than rat. Equilibrium dialysis indicates species differences in plasma protein binding and a considerable part of STS 557 to be present in plasma unbound. STS 557 is rather rapidly eliminated from the plasma compartment of all species investigated with half lives less than or equal to 10 h. As an additional time parameter of pharmacokinetics the "mean residence time" was used. Urinary excretion of STS 557 metabolites is dominant in all species, including the rat. In contrast to the great part of STS 557 in plasma total radioactivity, only small amounts of unchanged STS 557 are excreted in urine. First results of current studies in rabbits are presented, too.