The synthesis of C-19 steroids by guinea-pig adrenal homogenates

Guinea-pig adrenal homogenates were incubated with [³H]-pregnenolone as substrate. There was conversion to progesterone, cortisol, cortisone, 11β-hydroxyandrostenedione (11βOHA), androstenedione, adrenosterone and testosterone. Dehydroepiandrosterone could not be isolated. Of the total amount of substrate, 3.26% was converted to 11βOHA, and 4.08% was converted to cortisol. In an incubation using [¹⁴C]-cortisol as substrate, 2.34% conversion to 11βOHA was obtained. When adrenal homogenates were incubated with both [³H]-pregnenolone and [¹⁴C]-cortisol, the ³H/¹⁴C ratio in isolated 11βOHA was 3–4 fold higher than the ³H/¹⁴C ratio in isolated cortisol. These results indicate that cortisol is not the major precursor of 11βOHA in guinea-pig adrenal. It is likely that most of 11βOHA is formed from androstenedione.     

Synthesis and reduction of steroid C-20 alkylidene cyanoacetates.

The geometry of the condensation products between ethyl cyanoacetate and 20-ketosteroids (5 alpha-pregnane-20-one, 3 beta-hydroxypregn-5-en-20-one and 3 beta-acetoxypregn-5-en-20-one) was established by NMR spectra. Reduction of these steroid C-20 alkylidene cyanoacetates was shown to afford one of the two possible 20-C epimers, which seen to correspond to the 20 beta-methyl configuration.     

Side-chain cleavage and C-20 ketone reduction of hydrocortisone by a natural isolate ofChroococcus dispersus

A unicellular cyanobacterium,Chroococcus dispersus (Keissl.) Lemmermann, was isolated from paddy-field and tested in biotransformation experiments of hydrocortisone (compound 1). This strain has not been previously examined for steroid substance modification. Fermentation was carried out in BG-11 medium supplemented with 0.05% substrate at 25 °C for seven days incubation. The metabolites were chromatographically purified and characterised using spectroscopic methods. The fermentation yielded 11β,17α,20β,21-tetrahydroxypregn-4-en-3-one (compound 2), 11β,17β-dihydroxyandrost-4-en-3,17-dione (compound 3), and 11β-hydroxyandrost-4-en-3,17-dione (compound 4). Bioreaction characteristics observed were 20-ketone reduction for accumulation of compound 2 and side chain degradation of the substrate to give compounds 3 and 4. Time course study showed the accumulation of the product 2 from the second day of the fermentation and product 3 as well as product 4 from the third day. All the metabolites reached their maximum concentration in seven days. Aeration and continuous light or light duration (16/8 hours light/dark) have no effect on the transformation yield. Optimum concentration of the substrate, which gave maximum bioconversion efficiency, was 0.5 mg ml^?1 in the transformation experiment. Growth was not influenced by the addition of steroid substrate. Biotransformation was completely inhibited when steroid concentration was above 2.0 mg ml^?1.     

Urinary excretion of C-20 reduction products of corticosterone and 11-dehydrocorticosterone

Three hundred milligrams of corticosterone was ingested by a healthy man on each of 2 days, and urine was collected for 48 hr. The urine was extracted with chloroform in the presence of β-glucuronidase. Three steroids with a glycol side chain and the Δ⁴-3-ketone group have been separated from the urine extract. These were identified as (I) 20β,21-dihydroxy-4-pregnen-3,11-dione; (II) 11β,20β,21-trihydroxy-4-pregnen-3-one; and (III) 11β,20α,21-trihydroxy-4-pregnen-3-one.Evidence is presented indicating the presence of compound I in the urine of a woman with hypopituitarism following ingestion of 11-dehydrocorticosterone; and of the compound II in urine collected from two men during periods of adrenocorticotropin injections.     

The 1 alpha-hydroxylation of 24-hydroxyvitamin D3 by chick kidney homogenates

Tritium-labeled 24(R)-hydroxyvitamin D3 and 24(S)-hydroxyvitamin D3 were chemically synthesized and the 1 alpha-hydroxylation of these compounds by chick kidney homogenates was studied. A marked stereospecific preference with regard to the orientation of the hydroxyl functionality on carbon-24 was noted: while the 24(R)-epimer could be 1 alpha-hydroxylated in readily detectable amounts, the 24(S)-epimer was not hydroxylated. Thus, 1.2 micrograms of 1 alpha,24(R)-dihydroxyvitamin D3 was isolated and its structure confirmed by mass spectrometry. The relative rate of 1 alpha-hydroxylation of 125 nM substrate tritiated 24(R)-hydroxyvitamin D3 and 25-hydroxyvitamin D3 (the presumed natural substrate for the renal 1 alpha-hydroxylase) was 1:6.7.     

The gas-liquid chromatographic analysis of some C-20, 21-dihydroxycorticosteroids as C-20, 21-acetonides

The preparation and gas Chromatographic behaviour of 20, 21-isopropylidenedioxy derivatives of some C-20,21-dihydroxycorticosteroids is described. These ‘acetonide’ derivatives have been used for the separation of the epimeric cortols and cortolone in β-glucuronidase hydrolysed urine and for the quantitative estimation of the following urinary unconjugated corticosteroids: -20α and 20β-dihydrocortisol, 20-dihydrocortisone, 20α and 20β-dihydroprednisolone and 20-dihydroprednisone.     

Stereoselective reduction of non-cyclic carbonyl compounds by some eumycetes

Summary Twelve strains of Eumycetes were able to perform the reduction of 2-pentanone, acetophenone and ethyl acetoacetate, sometimes in a yield suitable for preparative work. For each substrate, preferential reduction to the R-configurated alcohol was observed with one or more strains.     

Steric effects at C-20 and C-24 on the metabolism of sterols by Tetrahymena pyriformis

Cultures of Tetrahymena pyriformis were incubated with various sterols and the extent of dehydrogenation at C-7 and C-22 was determined. The sterols incubated were desmosterol, 22-dehydrodesmosterol, 24-methyldesmosterol, 24 alpha-methylcholesterol (campesterol), 24-methylene-cholesterol, isohalosterol (26,27-bisnorcampesterol, also known as 24,24-dimethylchol-5-en-e beta-ol, a naturally occurring C26-sterol), and 20-isohalosterol. 20-Isohalosterol was not metabolized, while products with delta 7- and delta 22-bonds were formed from isohalosterol and all of the other sterols studied. This confirms an earlier conclusion, based on results with 20-isocholesterol and cholesterol, that inversion of the configuration from 20(R) to 20(S) completely prevents metabolism both in the nucleus and the side chain. On the other hand, changes in the electronics or stereochemistry at C-24 had a direct affect only on metabolism in the side chain. The presence of a methyl group at C-24 reduced the yield of metabolites with a delta 22-bond relative to those with a delta 7-bond producing an accumulation of 7-dehydro metabolite. A double bond at position-24 counteracted this steric effect, presumably by enhancing the rate of dehydrogenation, and a delta 24(28)-bond was more effect than was a delta 24(25)-bond.     

The effect of zinc on the oxidation of krebs cycle intermediates by rat liver and kidney homogenates

Zinc ions in small concentrations (2 × 10^?5M) inhibit rat liver and kidney succinoxidase systems by forming an inactive complex with the enzyme. Under conditions which favor oxidative phosphorylation (isotonic KCl homogenates, Mg ion, and muscle adenylate or adenosine triphosphate), inhibition by zinc occurs, but this inhibition is transitory. The rate of oxidation subsequently rises to a level even higher than shown by the control data. A later fall to a lower level of activity can be attributed to the zinc inhibition of other Krebs cycle intermediates, particularly the oxidation of α-ketoglutarate and citrate. The addition of a protein such as insulin has no protective action against zinc inhibition, and crystalline zinc insulin causes the same inhibition as does zinc sulfate. With the exception of succinoxidase, the oxidizing enzymes of rat kidney are more readily inhibited by zinc ions than are those of rat liver.Amorphous insulin, in itself, inhibits citrate oxidation by rat kidney homogenates.     

The fate of C-20 in C19 gibberellin biosynthesis

Experiments with ent-kaur-16-ene-[¹⁴C], prepared biosynthetically from sodium acetate-[2-¹⁴C], have shown that the C-20 carbon atom of the C₂₀ gibberellins is evolved as carbon dioxide during the biosynthesis of the C₁₉ gibberellins by Gibberella fujikuroi.     

面包酵母催化羰基不对称还原合成手性醇的研究

以2-辛酮和4-氯乙酰乙酸乙酯(COBE)为模型底物分别考察了酵母细胞对直链甲基酮和陆羰基酯中的羰基不对称还原情况。实验发现不对称还原2-辛酮的产物主要是S型的2-辛醇,且对映体选择性很高。不对称还原COBE生成的主要是S(D)-型产物,反应COBE的转化率、光学选择性都比较高。同时发现COBE的浓度和产物对不对称还原都有一定负面的影响。     

Kinetics of the reduction of C-9 substituted acridinium cations by 1,4-dihydronicotinamides

Second-order rate constants (k₂) are reported for the reduction of 9-R-10-methylacridinium cations (5:R = H, CH₃, CH₃CH₂, C₆H₅CH₂, (CH₃)₂CH, C₆H₅, 4-(CH₃)₂NC₆H₄) by 1-benzyl-1,4-dihydronicotinamide (2:R = C₆H₅CH₂) in 20% CH₃CN-80% H₂O at 25°C. All 5:R ≠ H are reduced in the range 20- to 140-fold more slowly than 5:R = H. However, there is no simple relationship between k₂ and the nature of R, nor between k₂ and the second-order rate constant for hydroxide ion attack at C-9 of these cations in pseudobase formation. Rates of reduction of 5 by 1-benzyl-4,4-dideuterio-1,4-dihydronicotinamide allow the calculation of the following kinetic isotope effects in this solvent medium: 5:R, $\text{k}{}^{\mathrm{<mtext>H</mtext>}}\text{k}{}^{\mathrm{<mtext>D</mtext>}}\text{:}\text{H}$, 1.56; C₆H₅CH₂, 2.7; C₆H₅, 5.4. Substituent effects upon k₂ were evaluated for the reduction of 5 by 1-(X-benzyl)-1,4-dihydronicotinamides, and lead to the following Hammett ? parameters: 5:R, ?: H, ?0.68; C₆H₅CH₂, ?0.92; C₆H₅, ?0.96. The latter two values require essentially complete unit positive charge generation on the nicotinamide moiety in the rate-determining transition state. It is shown that these Hammett ? values and the above isotope effects can only be rationalized by a two-step e^? + H^? mechanism for hydride transfer from 2 to 5 in this solvent system. This result contrasts with our earlier conclusion of direct, one-step hydride transfer in the reduction of isoquinolinium cations by 2, but is consistent with our observation that acridinium cations are reduced 37500-fold faster by 2 than predicted on the basis of the relative rates of nucleophilic attack (hydroxide ion) on acridinium and isoquinolinium cations. It is suggested that the availability of both Hammett ? values and primary kinetic isotope effects will generally allow the establishment of the mechanism of hydride transfer in these systems. Application of these ideas to literature data suggests that 5:R = H is reduced by direct hydride transfer in acetonitrile solution, in contrast to the above result in predominantly aqueous solution. The ready formation of acridanyl radicals by electron transfer to acridinium cations is demonstrated by the formation of Wurster's Blue radical cation upon mixing solutions of acridinium cations with N,N,N′,N′-tetramethyl-p-phenylenediamine.