Bile Acids and Their Salts

The taurine conjugates of di- and trihydroxy bile acids are very strong acids and are very soluble down to pH = 1 (125-127). 

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Much of the cholesterol synthesis in vertebrates takes place in the liver. A small fraction of the cholesterol made there is incorporated into the membranes of he-patocytes, but most of it is exported in one of three forms biliary cholesterol, bile acids, or cholesteryl esters. Bile acids and their salts are relatively hydrophilic cholesterol derivatives that are synthesized in the liver and aid in lipid digestion (see Fig. 17-1). Cholesteryl esters are formed in the liver through the action of acyl-CoA-cholesterol acyl transferase (ACAT). This enzyme catalyzes the transfer of a fatty acid from coenzyme A to the hydroxyl group of cholesterol (Fig. 21-38), converting the cholesterol to a more hydrophobic form. Cholesteryl esters are transported in secreted lipoprotein particles to other tissues that use cholesterol, or they are stored in the liver. 

A compilation of critical physical constants for the common bile acids and their salts is given in Table 1. The vast majority of literature values for melting points (m.p.) fall between the limits given in this table [34,35]. However, certain bile acids are notorious in forming crystalline polymorphs [36-38], and most dihydroxy bile acids form stable crystalUne solvates with a wide variety of solvent molecules... 

The first volume in this series was primarily concerned with the physio-chemical aspects of bile acids and their salts. In keeping with the original organizational format of this work, Volume 2 focuses our attention on the physiological and metabolic aspects of these substances. 

The topic of bile acids has been the subject of a number of general reviews and books (28, 29, 30, 33, 34). Certain reviews have dealt, in part, with physicochemical aspects of the bile salts (35-39) or have mentioned physicochemical properties only briefly (28, 30, 34, 40). A review dealing specifically with the physicochemical properties of bile salts and their relation to physiologic function was published in 1967 (1). This chapter will be limited to a discussion of the physicochemical properties of bile acids and their salts. (For physiologic correlations, see 1, 8, 9, 10, 41, and 197.)... 

Bile salts secreted into the intestine are efficiently reabsorbed (greater than 95 percent) and reused. The mixture of primary and secondary bile acids and bile salts is absorbed primarily in the ileum. They are actively transported from the intestinal mucosal cells into the portal blood, and are efficiently removed by the liver parenchymal cells. [Note Bile acids are hydrophobic and require a carrier in the portal blood. Albumin carries them in a noncovalent complex, just as it transports fatty acids in blood (see p. 179).] The liver converts both primary and secondary bile acids into bile salts by conjugation with glycine or taurine, and secretes them into the bile. The continuous process of secretion of bile salts into the bile, their passage through the duodenum where some are converted to bile acids, and their subsequent return to the liver as a mixture of bile acids and salts is termed the enterohepatic circulation (see Figure 18.11). Between 15 and 30 g of bile salts are secreted from the liver into the duodenum each day, yet only about 0.5 g is lost daily in the feces. Approximately 0.5 g per day is synthesized from cholesterol in the liver to replace the lost bile acids. Bile acid sequestrants, such as cholestyramine,2 bind bile acids in the gut, prevent their reabsorption, and so promote their excretion. They are used in the treatment of hypercholesterolemia because the removal of bile acids relieves the inhibition on bile acid synthesis in the liver, thereby diverting additional cholesterol into that pathway. [Note Dietary fiber also binds bile acids and increases their excretion.]... 

There are several methods available for the extraction of bile salts from serum or plasma. The most convenient methods utilize some form of liquid-solid extraction. An early procedure involved the anion-exchange resin, Amberlyst A-26 (S8), but considerable time and effort was required to perform column chromatography and to concentrate the eluate from the column. The introduction in 1972 of the neutral resin, Amberlite XAD-2, improved the ease of extracting bile acids and their conjugates from serum samples (M6). Further improvement occurred in 1977 with the description of a batch extraction technique using the related neutral resin, Amberlite XAD-7 (B5). With this technique, serum is diluted in 0.1 M sodium hydroxide to release bile acids from albumin and mixed with resin for 1 hour. After washing the resin in dilute alkali, bile acids are eluted with methanol, which cdn be removed on a rotary evaporator (B5). 

Physical constants of bile acids and their alkali salts... 

The purpose of this chapter is to cover the physicochemical properties of cholanic acids and their salts. Some 150 cholanic acids are mentioned in Elsevier s Encyclopedia of Organic Chemistry (27) and about 440 monocar-boxylic, unsubstituted hydroxyl, and carbonyl-substituted bile acids are cataloged by Sobotka (28). Obviously, it would be impossible to deal specifically with each one. Many of these acids are by-products of chemical reactions and are not naturally occurring compounds. Further, meaningful physicochemical data are not available for the great majority of bile acids. 

Therefore, I shall concentrate on only those bile acids that have been reasonably well studied from a physicochemical point of view and which have some relation to physiology and biochemistry of living things. Because the specific physical characteristics of the bile acids and their alkaline metal salts vary considerably with the number of hydroxyl groups present on the steroid nucleus, I will present a fairly detailed description of the physicochemical properties of cholanic acid (no hydroxyl groups), monohydroxy, dihydroxy, and trihydroxy bile acids. Since the triketo bile acid (dehydrocholic acid) has been used widely as a choleretic, its properties will also be discussed. Unfortunately, many interesting bile acids and bile alcohols isolated from a variety of vertebrates (29-32) have not been studied physicochemical ly. However, knowing their molecular structure, many of the properties of these compounds can be deduced by comparison with the known properties of bile acids discussed in this chapter. 

The optical activity of bile acids and their sodium salts depends on their purity, the wavelength of light employed, the solvent, and the concentration. The values given in standard references (27, 74) are often not comparable. Josephson (75) showed that increased concentration of bile salts in water substantially decreases the optical activity. The concentration, however, made little difference if the salts were studied in alcohol. Since the bile salts do not form micelles in alcohol (unpublished observations by the author), the variations noted in water may be related to the formation of micelles. The optical rotation of the acids in other organic solvents is not affected appreciably by concentration. The reader is referred to the earlier work of Sobotka (28) and Josephson (75). 

The structures of certain uncommon bile acids and their ethyl esters have abo been studied (79, 81), and incomplete crystal data of certain bile acid derivatives given in a brief note (72). The crystalline structures of most of the common bile acids and their alkaline salts have not yet been defined. 

The effect of pH on the physical state of bile acids and their sodium salts is perhaps best explained by examining a titration curve of typical bile salt obtained by titrating an alkaline bile salt solution with hydrochloric acid. 

Bile acids and salts have been found to enhance the absorption of both calcium and vitamin D hence, to increase calcium absorption both directly and indirectly (3,37). However, the ability of some dietary fibers such as lignin and pectin to absorb conjugated and deconjugated bile salts onto their surfaces to be excreted in the feces (a mechanism credited to the hypocholesterolemic effect of some dietary fibers) may result in an overall decrease in calcium absorption from the gastrointestinal tract (7,33,38-40). 

Eatty acids such as oleic, capric, linoleic acids, and their monoglycerides Bile salts such as cholate, taurocholate and derivatives, UDCA, CDCA, SCG, STDHF Homovanilate... 

Beside the bile salts, the anionic surfactants investigated for enhanced intestinal delivery were mainly sodium salts of fatty acids and their derivatives. These include sodium salts of saturated and unsaturated fatty acids (C8-Ci8), SLS, dioctyl sodium sulfossuccinate (DOSS), and others. 

Bile salts are substances derived from sterols, which make up a substantial part of the solid matter in bile and which play a central role in lipid absorption, by virtue of their surface-active properties. The structure and properties of these salts have been reviewed by Haslewood (305) and Heaton (316). Bile salts essentially have molecules of detergent type hydrocarbon, with a fat-dissolving part and a polar, water-attracting part. The fat-dissolving part consists of the bulk of the steroid nucleus. The hydroxyl groups are so distributed that hydration can readily take place the remainder of the molecule will dissolve the fatty phase. Emulsification of fat/water complexes can thus occur easily. The terms bile acid and bile salt are used somewhat interchangeably in the literature. 

Cholestyramine or colestipol form an insoluble complex with the bile acids and salts, preventing their reabsorption from the intestine. 

The orientation of the mobile side chain at C-17 has not been unambiguously resolved for alkali salts of bile acids in aqueous solutions. However, evidence adduced from the differences in calculated molecular areas of surface-adsorbed bile salts versus spread monomolecular layers of insoluble bile acids at their collapse pressures [12] suggest that the side chain, when ionized, lies a-axial with respect to the plane of the molecule, but when undissociated lies parallel to the surface. Effects of systematic variations in pH between 1 and 13 on the surface areas of spread and adsorbed glycocholate molecules suggest that the difference between the two surface... 

The terms bile acid" and bile salt" refer to the un-ionized and ionized forms, respeefively, of these compounds. For illustrative purposes only. Figure 30.3 shows cholic acid as a un-ionized bile acid and glycocholate as an ionized bile salt (as the sodium salt). At physiologic and intestinal pFI values, both compounds would exist almost exclusively in their ionized forms. 

Fig. 9. Phase equilibria for the bile salt (bile acid)-fatty acid-water system at constant water concentration in relation to temperature (see Fig. 5). Six mixtures varying in molar ratios of bile salt (bile acid) and palmitic acid with total concentration of micellar bile acid plus palmitic acid equal to 40 mM were examined. Fatty acid has a finite solubility in the micellar bile acid solution, the excess being crystalline at body temperature. At 50-60 C, there is a marked increase in micellar solubility, and the fatty acid melts. At higher fatty acid/bile acid ratios, the micellar solubility is exceeded, and an immiscible oil phase occurs. The melting point of fatty acid in the presence of water is nearly identical to that in the anhydrous state (38), in contrast to the behavior of monoglyceride (Table I). As shown in Fig. 3, the size of the micellar area decreases with increasing chain length. Unsaturated fatty acids (not shown) behave similarly to saturated fatty acids, but their micellar solubility is greater, and at most experimental temperatures a crystalline phase will not occur.

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