Keto bile acids

Common to spectra of ketonic bile acid methyl esters lacking hydroxyl groups is that the peak at mje M-13 cl) has a considerably higher relative intensity than in spectra of derivatives of hydroxylated bile acids. The influence of keto groups on fragmentations in the side chain is also seen in the relatively pronounced loss of 32 mass units (cj) from the molecular ions of 3-keto- and 7-ketocholanoates (Fig. 9). Derivatives of hydroxylated bile acids usually give an ion at mje M-3 by loss of the ester methoxyl group. 

When a 3-keto group is the only substituent in a cholanoic acid structure the mass spectrum clearly indicates its position. The mass spectrum of the ethyl ester of 3-keto-5/3-cholanoic acid is shown in Fig. 9 and it is seen that a major fragmentation is loss of carbons 1-4 with the carbonyl group n, M-70). The ion formed is found at mje 332 and 318 in spectra of the ethyl 

One way to study the presence of a 3-keto group is to convert it to an enol trimethylsilyl ether which gives pronounced peaks at mje 142 and 143 (cf. 23). The latter ion is the base peak in the spectrum of an enol silyl ether of methyl 3-keto-12a-hydroxy-5/3-cholanoate. An intense peak is also seen at mje 316, i.e., A/-(90- -142). In view of the preferred loss of the side chain following loss of a 12-trimethylsiloxy function the fragment of mass 142 is unlikely to represent the side chain with C-16 and C-17. A prominent peak at mje 201 (Af-[90-f 1424-115]) indicates that it represents carbon atoms 1-4. 

A 3-keto group can also be converted into a 1,1-dimethylhydrazone under mild conditions (48). Spectra of compounds derivatized in this way show large molecular ion peaks (often the base peak), thermal loss of 2 and 4 hydrogens, and peaks at mje 42, 69, and 70. 

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The enzyme 3a-hydroxysteroid dehydrogenase plays a key role in this transport across the hepatocyte. A particularly elegant experiment demonstrated the role of the 3a-hydroxysteroid dehydrogenase, by using [ H] at the 3p hydrogen to show cyclical oxidation-reduction of the 3a-hydroxyl with no accumulation of 3-keto bile acids. Confirmation was obtained by use of indo-methacin, an inhibitor of 3a-hydroxysteroid dehydrogenase, which decreased... 

The most common assay uses 3a-hydroxysteroid dehydrogenase to form the 3-keto bile acid that is trapped by, for example, hydrazine hydrate, causing the reaction to go to completion. The co-factor NAD is reduced stoichiometrically and can be measured by ultraviolet absorption or more commonly by fluorescence at an activation of 345 nm and emission of 450 nm. Use of this enzyme measures all bile acids with a 3a-hydroxyl but not cholesterol, which has a 3p-hydroxyl, and does not measure bile acids with a sulphate or glucuronide group conjugated to the 3a-hydroxyl. 

Previously, some fluorocyclohexanones were used in a catalytic amount with Oxone for asymmetric epoxidation reaction, but they gave a poor ee . It was found later that chiral ketones derived from fructose work well as asymmetric epoxidation catalysts and show high enantioselectivity in reactions of /rani-disubstituted and trisubsti-tuted olefins ". Cis and terminal olefins show low ee under these reaction conditions. Interestingly, the catalytic efficiency was enhanced dramatically upon raising the pH. Another asymmetric epoxidation was also reported using Oxone with keto bile acids. ... 

Dehydrogenation of carbonyl compounds. Introduction of a 9,11-double bond into a 12-keto bile acid by selenium dehydrogenation was discovered by Schwenk and Stahl and became a key step in the commercial production of cortisone. Kendall and co-workers " studied the reaction carefully as applied to methyl 3a-benzoyloxy-12-ketocholanate, using as solvent a 4 1 mixture of chlorobenzene and acetic acid. They found that a trace of hydrochloric acid slowed down the reaction but increased the yield. Thus with no added mineral acid the yield reached a maximum of 67% in a reflux period of 24 hrs. but with the mixture 0.0006 normal in HCl refluxing for 72 hrs. alforded the product in 84% yield. The product was to be isolated after... 

Esters. Under usual conditions of experimentation, carboxylic esters are not reduced by sodium borohydride. Thus 3-keto bile acid esters can be reduced at C3 without disturbance of the ester function. Methyl 4y8-bromo-3-ketocholanate... 

In addition to bile and serum bile acids, fecal bile acids can be estimated by the enzymatic method, provided that they are first extracted to remove interfering lipids. Methods for this purpose are continually being simplified and improved (B16, D6, V3). Enzymatic determination of fecal extracts slightly underestimates total bile add excretion, since 3-keto bile acids and bile acids which are sul ted at the 3a position occur in feces and are not... 

C.d. curves have been suggested as a means for distinguishing between keto-bile acids with the keto-group at C-3, C-7, or C-12. The c.d. curves of a series of oestr-4-en-3-ones (21), -4,9-dien-3-ones (22), and -4,9,ll-trien-3-ones (23), including compounds substituted in ring A and elsewhere, show temperature sensitivity indicative of conformational equilibria between normal and inverted forms. Some apparent anomalies in earlier c.d. data for compounds with 2/3-or 1/8-methyl substituents are explained in terms of equilibria involving predominant inverted conformations, rather than single twist or boat conformers... 

Several other groups have reported effective dioxirane systems employing Oxone as the terminal oxidant. For example, Armstrong et al. have developed a spirocyclic iV-carbethoxy-azabicyclo[3.2.1]octanone precatalyst, which affords up to 91.5% ee in the epoxidation of stilbenes (eq 102). Shing et al. have developed an arabinose-derived ketone and employed this in the enantioselective synthesis of the Taxol side chain however, enantioselectivities for the epoxidation were only up to 68% (eq 103). Bortolini et al. have also described the epoxidation of alkenes with the stoichiometric keto bile acid-Oxone system, a range of ee values were observed over several substrate types but up to 98% was observed for the epoxidation of tran -stilbene, although the yield was only 50% (eq 104). ... 

Confirmative evidence of the proposed structure was obtained from partial synthesis of hyocholic acid [Hsia et al. (30)]. An important intermediate in the synthesis was 3a,6a-dihydroxy-7-keto-5 -cholanic acid (VII, Fig. 2), first prepared by Takeda et al. (35). The 3 - and 6a-hydroxyl groups in VII were established by the formation of hyodeoxycholic acid (IX) after hydrogenolysis of the ethylenedithioketal derivative (VIII) with Raney nickel. Hyocholic acid was obtained from VII either by reduction with sodium borohydride or by hydrogenation in the presence of platinum both methods were known to produce the axially oriented 7a-hydroxy from 7-keto bile acids [Mosbach et al. (36) Iwasaki (37)]. More direct evidence for the la-hydroxyl group in hyocholic acid was found in a later study [Hsia et al. (8)], when hyocholic acid was derived from bromohydrin acetate XII (Fig. 3),... 

Partial (3, 97) or complete (98) silylation may be obtained. Hydroxyl groups at C-3, 6a, Ip, and 12 are transformed into silyl ethers when treated with hexamethyldisilazane, 0.03 ml, in dry dimethylformamide, 0.06 ml, at 50 °C for 3hr (77). Complete silylation, both of equatorial and axial hydroyxl groups, is obtained in pyridine(dry)-hexamethyldisilazane-trimethyl-chlorosilane, 10 5 2 (v/v), 15 min, room temperature (98). Prolonged reaction times may give enol silyl ethers with keto bile acids. In an analogous reaction VandenHeuvel and Brady have prepared chloromethyldimethylsilyl ethers (99). Silyl ethers are very readily hydrolyzed. 

The following discussion is based on spectra of the bile acid derivatives listed in Table I. This is not a complete list of all published spectra but the results are representative of general fragmentation patterns. The molecular weights of bile acid derivatives can be calculated from the group contributions listed in Table II. This table also shows how the molecular weight changes when a hydroxy and/or keto bile acid methyl ester is converted into the derivatives. 

Ring A of an enol trimethylsilyl ether of a 3-keto bile acid. 

The intensity of the peak at mje 292 depends on the stereochemistry of A/B ring junction. It is very low in the spectrum of methyl 3,7-diketo-5a-cholanoate (36) and is about 30% of the base peak in that of the 5 -isomer. The same difference is seen for the peak at mje 111. Comparisons of spectra of different 7-keto bile acid derivatives indicate that the latter ion may represent loss of side chain from the ion of mass 292. 

In 2001, Bortolini et al. described a series of keto bile acids 410 as dioxirane precursors for the asymmetric epoxidation [289, 290]. Trans-stilhem can be epoxi-dized in up to 98% ee (Fig. 7.21). 

In 2001, Bortolini used keto bile acids 431 which epoxidized p-methylcinnamic acid in good yields (45-94%) and high ee s (26-95%) (Fig. 7.24) [300, 301]. 

The 3p-HSDH-catalyzed reductions of the 3-keto bile acids 2, 32, 35, 38, 41, 44, 47, and 50 to the corresponding 3p-hydroxy derivatives were also obtained with quantitative yields and practically absolute stereospecificity. M this case, all of the various substrates except 32 (about 50% slower) were transformed by the enzyme with similar rates. 

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