Cholestane-triol

Sevanian et al. (1994) applied GLC and LC/TS/MS for the analysis of plasma cholesterol-7-hydroperoxides and 7-ketocholesterol. Analysis of human and rabbit plasma identified the commonly occurring oxidation products, yet dramatic increases in 7-ketocholesterol and cholesterol-5p, 6P-epoxide were observed. The study failed to reveal the presence of choles-terol-7-hydroperoxides, which were either too unstable for isolation, metabolized or further decomposed. The principal ions of cholesterol oxides monitored by LC/TS/MS were m/z 438 (cholestane triol) m/z 401 (cholesterol-7-hydroperoxide) m/z 401 (7-ketocholesterol) m/z 367 (7a-hydroxycholesterol) m/z 399 (cholesta-3,5-dien-7-one) and m/z 385 (choles-terol-5a,6a-epoxide). The major ions were supported by minor ions consistent with the steroid structure. Kamido et al. (1992a, b) synthesized the cholesteryl 5-oxovaleroyl and 9-oxononanoyl esters as stable secondary oxidation products of cholesteryl arachidonate and linoleate, respectively. These compounds were identified as the 3,5-dinitrophenylhydrazone (DNPH) derivatives by reversed-phase LC/NICI/MS. These standards were used to identify cholesteryl and 7-ketocholesteryl 5-oxovaleroyl and 9-oxononanoyl esters as major components of the cholesteryl ester core aldehydes generated by copper-catalysed peroxidation of low-density lipoprotein (LDL). In addition to 9-oxoalkanoate (major product), minor amounts of the 8, 9, 10, 11 and 12 oxo-alkanoates were also identified among the peroxidation products of cholesteryl linoleate. Peroxidation of cholesteryl arachidonate yielded the 4, 6, 7, 8, 9 and 10 oxo-alkanoates of cholesterol as minor products. The oxysterols resulting from the peroxidation of the steroid ring were mainly 7-keto, 7a-hydroxy and 7P-... 

The presence of an a-bromo substituent may cause anomalies. With NaBH4, 2a-bromo-5a-cholestan-3-one gives a mixture of epimers, in which the 3p-o predominates. 4 -Bromo-17)5-hydroxy-5)5-androstan-3-one acetate gives 25% of the 315,4 -bromohydrin and 34% of the 3a,4)5-compound. Reduction of 7a-bromo-3)5,5a-diacetoxycholestan-6-one gives exclusively 7a-bromocholestane-3)5,5a,6a-triol 3,5-diacetate,whereas reduc-... 

Preparation—Cholestane-3(i,5a,6P-triol 3-Cathylate A solution of choles-tane-3/ ,5a,6j5-triol (1 g) in dioxane (10 ml) and pyridine (1.6 ml) is cooled to 25° and treated dropwise and with cooling with 2 ml of ethyl chlorocarbonate. After 1 hr, 25 ml of water and 1 ml of 36 % hydrochloric acid are added and the mixture is heated for 30 min on the steam bath and cooled. The product, a granular white solid, is filtered to yield 1.19 g mp 180-182°. Crystallization from methanol (75 ml) gives 0.62 g or prisms, mp 184.5-185°, and a second crop, 0.38 g, mp 183-184° (total yield 83%). Two recrystallizations of the 1st crop material yield prisms mp 184-185° [ ] —16° (CHCI3). 

It is clear then that more than one mechanism is operative for glycol fission. In the case of c -cyclopentanediols and camphanediols a cyclic ester is a necessary intermediate. For tra/js-decalin-9,10-diol a non-cyclic mechanism must operate which cannot function for cholestane-3/ ,6j8,7a-triol and is inefficient for /rans-camphanediols. It is pertinent that while the fission of glycols capable of forming cyclic esters proceeds several hundred times faster in benzene than in acetic acid, the reactions of trans-decalin-9,10-diol and tra/ij-hydrindane-l,6-diol are 4-5-fold slower in benzene . ... 

As mentioned earlier, oxidation of LDL is initiated by free radical attack at the diallylic positions of unsaturated fatty acids. For example, copper- or endothelial cell-initiated LDL oxidation resulted in a large formation of monohydroxy derivatives of linoleic and arachi-donic acids at the early stage of the reaction [175], During the reaction, the amount of these products is diminished, and monohydroxy derivatives of oleic acid appeared. Thus, monohydroxy derivatives of unsaturated acids are the major products of the oxidation of human LDL. Breuer et al. [176] measured cholesterol oxidation products (oxysterols) formed during copper- or soybean lipoxygenase-initiated LDL oxidation. They identified chlolcst-5-cnc-3(3, 4a-diol, cholest-5-ene-3(3, 4(3-diol, and cholestane-3 3, 5a, 6a-triol, which are present in human atherosclerotic plaques. 

Furster C, WikvaU K. 1999. Identification of CYP3A4 as the major enzyme responsible for 25-hydroxylation of 5beta-cholestane-3alpha,7alpha,12alpha-triol in human liver microsomes. Biochim Biophys Acta 1437 46-52. 

Wilson, A.M. Sisk, R.M. O Brien, N.M. 1997. Modulation of cholestan-3beta, 5alpha,6beta-triol toxicity by butylated hydroxytoluene, alpha-tocopherol and beta-carotene in newborn rat kidney cells in vitro. Br. J. Nutr. 78 479 92. 

Steroidal alcohols have been oxidised with a variety of other reagents. Sources of positive" halogen, such as N-bromoamides [30-32], isocyanuric chloride [33], and tert-butyl hypochlorite [34], readily convert many alcohols into ketones. Axial alc idls are again the most probably for reasons similar to those discussed above [31]. This reactivity difference has been exploited, for example, in the selective oxidation of 5a-cholestane-3j5,5a,6jS"triol (5) to give the 3 S,3a"dihydroxy-6-ketone (6) [35]. Equatorial alcohols display differences in rates of oxidation the more... 

Angyal and Young report second-order kinetics for the oxidation of the camphane-2,3-diols. The cis- isomers are oxidised much more rapidly than the trans- isomers, and a temperature of 80 °C had to be used for kinetic measurements on the latter. It seems likely that the rigidity of the camphane skeleton prevents the formation of a cyclic diol-periodate ester from the trans- isomers. Possibly the reaction at 80° is completely different in nature from the normal oxidation of 1,2-diols by periodate. The same workers report that cholestane-3j8,6j8,7a-triol, in which the and 7a hydroxyl groups are axial-axial, is inert towards periodate. 

C,2iS- H,3R]mevalonate, Bloxham and Akhtar showed that a tritium atom was lost whereas when [3a- H,26,27- C2]lanosterol was used the tritium was retained. The latter result was also observed by Hornby and Boyd. Presumably NAD is necessary for the oxidation at C-3 to a ketone prior to decarboxylation. Similarly, Miller and Gaylor showed that 4a-methyl-5a-cholest-7-en-3) -ol was oxidized only as far as the 4a-carboxylic acid, with retention of tritium at C-3 but loss from a 4a-C H3 group. In the latter case, the recovered 4a-methyl sterol showed no sign of tritium enrichment due to isotope effects. In banana, alkylation at C-24 seems to precede loss of the 4a-methyl groups. When the rat liver system was inhibited by cholestane-3, 5a,6 -triol, sterols accumulated which retained a methyl group at C-4. Both 4,4-dimethyl- and 4 -methyl-cholest-8-en-3/I-ol Uere isolated, and were shown to be converted into cholesterol under normal conditions. 

Several COPs have been identified as having cytotoxic, angiotoxic, carcinogenic, and mutagenic bioactivities (280, 281), all or some of which may play a role in the initiation or proliferation of atherosclerotic plaque. Specifically, 25-hydroxycholesterol and cholestane-36,5a,66-triol are particularly toxic to cultured rabbit aortic smooth muscle cells (278). The 25-hydroxycholesterol... 

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. 

Rat liver microsomes hydroxylate 5/8-cholestane-3a ,7a,12Q -triol at C-25 and C-26 both activities are dependent on cytochrome P450 and there is some evidence that different types of the latter are involved. A mitochondrial steroid 24-hydroxylase that accepts 3a,7a,12a-trihydroxy-5/3-cholestanoic acid has been extracted from rat liver apparently this is not a mixed-function oxidase although the presence of oxygen was obligatory for its action. Bile acids hydroxylated at C-23 have been formed from sodium cholate and deoxycholate in preparations from Viperinae species and a steroid-12ct-hydroxylase from liver microsomes has been studied.Sitosterol has been confirmed to be a precursor of C24 and C29 bile acids in mammalian liver, and here hydroxylation at C-26 precedes that at C-7. ° "... 

Both mesyloxy and tosyloxy are efficient leaving groups. For example, mesylation of cholestane-3, 5o ,6/3-triol 3-acetate (1) followed by reaction of the 6-mesylate... 

A solution of 1 g. of cholesterol a-oxide in 30 ml. of hot acetone is treated with a solution of 0.625 g. of periodic acid dihydrate in 10 ml. of water." Before all the precipitated oxide has redissolved, thin plates of cholestane-3/3, 5a,6(8-triol begin to separate. The mixture is refluxed for one half hour, cooled, and the product collected yield 0.83 g. (81%), m.p. 231-232°. A second crop of material (0.14 g.) melted at 225-226°. 

Acetylation. Cholestane-3 3,5a,6/8-triol on acetylation with acetic anhydride in pyridine affords the 3,6-diacetate the triacetate can be prepared by brief heating... 

Oxidation of secondary alcohols. In the presence of water, N-bromosuccinimide is a highly selective oxidizing agent. Thus oxidation of cholestane-3/3,5a,6/3-triol with... 

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