5p-Cholestane

Smith LL, Gouron RE, Sterol metabolism. VI. Detection of 5P-cholestan-3P-ol... 

There have been known many examples that the hydrogenation of 3p-substituted A5-steroids yields mainly or exclusively saturated steroids of 5a series.166 Lewis and Shoppee studied the influence of various 3 a substituents on the stereochemical course of the hydrogenation of A5-steroids, and found that the 3a substituents lead to the preferential and sometimes exclusive formation of 5P-cholestane derivatives.166 The hydrogenations over platinum oxide were effectuated in methanol or ethyl acetate in the presence of traces of strong acids such as perchloric acid, sulfuric acid or hydrobromic acid. The results summarized by Lewis and Shoppee (eq. 3.34), which also include those by Haworth et al.,167 suggest that the bulkier the axial 3a substituent, the larger is the proportion of 5P steroid formed. The hydrogenation of A4-steroids usually leads to a mixture of 5a and 5P compounds and the stereochemical influence of 3a and 3P substituents is less marked than in the cases of A5-steroids.168... 

As in an example of franr-2-decalone described above, the hydrogenation of substituted cyclohexanones with platinum catalysts usually gives higher yields of axial alcohols in the presence of hydrobromic acid than in the presence of hydrochloric acid. Ruzicka et al. obtained 5a-cholestan-3a-ol in 67% yield by hydrogenating 5a-cholestan-3-one (62 g, 0.16 mol) with platinum oxide (12 g) in Bu20 (1200 ml)-HBr (48%, 3 ml) at 65-70°C. Similarly, 5P-cholestan-3P-ol was obtained in 95% yield in the hydrogenation of 5P-cholestan-3-one (40 g, 0.10 mol) over platinum oxide (8 g) in AcOH (1200 ml)-HBr (48%, 8 ml) at 60°C.155 The proportion of hydrogenolysis... 

Bile alcohols are polyhydroxy C27 sterols that serve as intermediates in the biosjmthesis of cholic acid and chenodeoxycholic acid from cholesterol (1, 2). Recently several studies have shown that increased amounts of bile alcohols namely 27-nor-5p-cholestane-3a,7a,12a,24, 25-pentol and 5P-cholestane-3a, 7a,12a,25,26-pentol are excreted (as glucuronides in urine of patients with liver diseases such as primary biliary cirrhosis (3), liver cirrhosis (4, 5) and a-antitrypsin deficiency (6). Ichimiya et. al., described the occurrence of 5P-cholestane-3a,7a,12a,26,27-pentol (5P-cyprinol) and 5P-cholestane-3a,7a,... 

Chromatographic analysis of the polar sterol fractions of the bile and feces, in conjunction with gas-liquid chromatography (GLC)-mass spectrometry indicated two major components, 5P-cholestane-3a,7a,12a,25-tetrol and 5P-cholestane-3a,7a,12a,23,25-pentol, and a minor component, 5P-cholestane-3a, 7a,12a,24,25-pentol(18-21). Only minute amounts of 5P-cholestane-3a, 7a,12a,23R-tetrol, 5P-cholestane-3a,7a,12a,24R-tetrol, 5P-cholestane-3a,7a, I2a,24S-tetrol and 5P-cholestane-3a,7a,12a,25S,26-pentol have been detected (20-24). The presence of 5P-cholestane-3a,7a,12a,25-tetrol was positively identified by comparison with the synthesized sample prepared in our laboratory (20-22,24-27) (Fig.l). The predominent bile alcohol of the pentol fraction was 5P-cholestane-3a,7a,12a,23,25-pentol, amounting to approximately 80% by weight, while 5P-cholestane-3a,7a,12a,24,25-pentol accounted for approximately 20% of this fraction. The less abundant pentol was shown to be identical with 5P-cholestane-3a,7a,12a,24a,25-pentol, which had been prepared from 5P-cholestane-3a,7a,12a,25-tetrol (20,21). 

The biosynthetic 5p-cholestanetetrol and pentol had the same melting point, TLC and GLC properties, and infrared and mass spectra as the reference compound (19-21). In addition, the recently described two-dimensional H-NMR studies (28-30) from our and other laboratories have further elucidated their structure and stereochemistry. Fig. 2 illustrates 2D-NMR H- H COSY spectra of 5P-cholestane-3a,7a, 12a 25-tetrol (for comparison 2D-NMR H- H COSY spectra of 5P-cholestane-3a,7P, 12a,25-tetrol is also provided). The 13c-DEPT spectra of (24R) and 5P-cholestane-3a,7a,12a,24a,25-pentol and (23R)-5P-cholestane-3a,7a,12a, 24a,25-pentol has been recently described (21). 

The occurrence of bile alcohols hydroxylated at position 25 in CTX patients indicated the presence of an alternate pathway of bile acid synthesis from cholesterol (via 25-hydroxylated intermediates ). In particular, the identification of 5P-cholestane-3a,7a,12a,24a,25-pentol indicated that cholic acid arised from the cleavage of a 24,25- glycol. 

During the course of these studies, the two 5P-cholestane-3a,7a, 12a,24,25-pentols, epimeric at C-24, and 5P-cholestane-3a,7a,12a,23,25-pentol epimeric at C-23 were further resolved by analytical and preparative TLC, and were either synthesized or isolated and characterized. In addition, the absolute stereochemistry at C-23 of 5P-cholestane-3a,7a, 12a,23,25-pentol and at C-24 of 5P-cholestane-3a,7a,12a,24,25-pentol (39,40) was established by circular dichroism (CD) studies employing the method of Nakanishi (41,42). These experiments conclusively defined the chirality of these pentahydroxy bile alcohols having 1,2 and 1,3 glycol systems in the side chain. Using Eu(fod)3... 

The insertion of hydroxyl groups into the 23- or 24-position of 5P-cholestane-3a,7a,12a,25-tetrol was found to be stereospecific. Although all these compounds were potential precursors of bile acid, studies in vivo and in vitro experiments using [3P- H] and (24- C) 5P-cholestane-3a,7a,12a,25-tetrol (46) (Figs.6, 7), (24- C) 5p-cholestane-3a,7a,12a,24R,25-pentol and (24- C) 5P-cholestane-3a,7a,12a,24S,25-pentol demonstrated the existence of a new 25-hydroxylation pathway for the transformation of cholesterol to cholic acid in these patients (2,10). The reaction sequence involved the stereospecific formation of a 24S-hydroxy pentol, 5P-cholestane-3a,7a,12a,24S,25-pentol, 3a7a,12a,25-tetrahydroxy-5P-cholestan-24-one and did not involve SP-cholestanoic acids as intermediates (Fig. 8). The two bile pentols, SP-cholestane-3a,7a,12a,24R, 25-pentol and 5P-cholestane-3a,7a,12a,23R,25-... 

Hvdroxvlation pathway This pathway has been demonstrated in both rat and human liver (2,10,56). It involves the 25-hydroxylation of 5P-cholestane-3a,7a,12a-triol to give 5P-choiestane-3a,7a,12a,25- tetrol (XIV) followed by stereospecific 24S-hydroxylation to yield 5P-cholestane-3a,7a,12a,24S,25-pentol (XV, Fig. 9). The pentoi is then oxidized to 5P-choiestane-3a,7a,12a, 25-tetrahydroxy-5P-cholestan-24-one (XVI) (59,60), which is degraded by... 

Hvdroxvlation pathway An alternative explanation for the bile acid synthetic defect in CTX has been proposed by Oftebro and colleagues which starts via 26-hydroxylation of 5P-cholestane-3a,7a,12a-triol (IX, Fig. lOa and 10b). In this pathway the mitochondrial fraction of both human and rat liver contains a 26-hydroxylase enzyme (63) which can convert 5P-cholestane-3a,7a,12a-triol (IX ) to 5P-cholestane-3a,7a,12a,26-tetrol (XI) (Fig. 10a and 10b ). This tetrol is oxidized to 3a,7a,12a-trihydroxy-5P-cholestan-26-oic acid (THCA, XII) by liver cytosol (2,64). Further hydroxylation at C-24 forms varanic acid (XIV) and its side chain is shortened with oxidation at C-24 to yield cholic acid (X,Fig. 10 a). These investigators demonstrated diminished mitochondrial 26-hydroxylation of 5p-cholestane-3a,7a,12a-triol and 5P-cholestane-3a,7a-diol, possible precursors for cholic acid and chenodeoxycholec acid in CTX liver. As a consequence, neither 26-hydroxylated intermediates can be formed so that total primary bile acid synthesis would be diminished. Accordingly, the accumulation of 5P-cholestane-3a,7a,12a,25-tetrol arises from 25-hydroxylation of 5P-cholestane-3a,7a,12a-triol by the alternative microsomal 25-hydroxylation mechanism. 

Fig. 10 (b). The sequence leading to the oxidation and cleavage of the side chain of 5P-cholestane-3a,7a,12a-triol pathway for side-chain cleavage in cholic acid biosynthesis. 

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