THE BIOLOGICAL SIGNIFICANCE OF CHEMICAL DIFFERENCES IN BILE SALTS

Summary1. The chemical nature of the bile salts is a character that must be under the control of several genes and is also affected by intestinal micro‐organisms and perhaps again by the liver in the course of enterohepatic circulation. Gall‐bladder bile contains the bile salts which are in use; bile from a fistula has only the primary bile salts actually produced by the liver.2. In the only invertebrate examined, the crab Cancer pugurus, the bile‐salt‐like substances in the digestive juices were compounds made up of sarcosine, taurine and decanoic or 5‐dodecenoic acids.3. It has been shown for at least 13 vertebrate species (7 eutherian mammals; a bird, a boid snake, an alligator, a toad, a frog and a teleost) that the characteristic bile salts are made in viuo from cholesterol; it is assumed that this is so in all vertebrates.4. In rats and man, 3α, 7α, 12α‐trihydroxycoprostanic acid, which contains all 27 atoms of cholesterol and is made from this compound, acts as an efficient precursor of the principal bile acid, cholic acid (3α, 7α12α‐trihydroxycholanic acid, C24H40O5). 3α, 7α, 12α‐trihydroxycoprostanic acid conjugated with taurine is a principal bile acid in all three crocodilians examined, is also present in the bile salts of two species of Rana and has been isolated from human fistula bile. This acid is an example of substances that are intermediates in bile salt synthesis in highly evolved vertebrates and also act as bile salts in less evolved ones.5. There is clear evidence of evolution of bile salts through the stages C27 (or C26) alcohols → C27 acids → C24 acids. Bile alcohols act as bile salts after conjugation with sulphate, C27 acids are conjugated with taurine and C24 acids exist in the bile as taurine and (in eutherians) also as glycine conjugates.6. Two species of hagfish (Myxinidae) have an alcohol, myxinol, as a disulphate of unknown chemical constitution.7. The coelacanth Latimeria chalumnae has a principal bile alcohol which is the 3β epimer of cyprinol; a little cyprinol is also present. Cyprinol, 3α, 7α, 12α,26,27‐pentahydroxycholestane, is a chief bile alcohol in the dipnoan Protopterus aethiopicus and also in the five species of Cyprinidae examined. The Latimeria alcohol is (if not an artifact of the enterohepatic circulation) more ‘primitive’ chemically (i.e. nearer to cholesterol) than is cyprinol: it could give rise to cyprinol by evolution of a method for inversion at C‐3. Thus, all these fishes are related by the possession of a bile alcohol type not yet found in other vertebrates.8. About 34 teleostean fish species, excluding Ostariophysii, have cholic and in some cases also chenodeoxycholic (3α,7α‐dihydroxycholanic) acids, conjugated with taurine, as their chief bile salts. In some species allocholic (3α,7α,12α‐trihydroxyallo(5α)‐cholanic) acid is also present: its significance is not clear. Probably most teleosts are highly evolved in their bile salt chemistry.9. Three species of sturgeons (Acipenseridae) contain cholic and allocholic acids and small amounts of bile alcohol sulphates are also present. Identification of these may be of value in elucidating sturgeon evolutionary history.10. Chimaera monstrosa has as its chief bile salt the sulphate of chimaerol, probably 3α,7α,12α,24ξ,26‐pentahydroxycoprostane. Sharks and rays may contain a little chimaerol but the principal bile salt is the sulphate of scymnol, 3α,7α,12α,24ξ,26,27‐hexahydroxycoprostane, which could arise from chimaerol by oxidation at C‐27. The cholic acid found in selachians could be a dietary artifact; the bile salts of these fishes have either not evolved so far or have reached a different evolutionary result from those of teleosts. Scymnol is not an efficient precursor of cholic acid in the rat.11. In amphibia, Rana catesbiana contains the sulphates of 5α‐ and 5β‐ranol, i.e. 3α,7α,12α,24ξ,26‐pentahydroxy‐27‐norcholestane‐27‐sulphate and its 5β epimer; in this species 3α,7α,12α‐trihydroxycoprostanic acid was also found. In R. temporaria 5α‐ranol sulphate is almost the only bile salt. R. nigromaculata has the trihydroxy‐coprostanic acid; bile alcohols were not found. These findings put the three species in the evolutionary order R. temporaria, R. catesbiana, R. nigromaculata. Bufo vulgaris japonicus has C27 or C28 bile acids with the cholic acid nucleus and also the sulphate of 3α,7α,12α,25ξ,26‐pentahydroxycoprostane. This alcohol is quite different from those found in Ranidae. Its 5α epimer has been reported in the newt Diemyctylus phyrrho‐gaster and, if this is the case, it suggests a possible link between ancestors of this animal and of Bufonidae.12. Bile alcohols have not been found in reptiles or higher vertebrates. Chelonians and platynotan lizards have (probably) 3α,7α,12α,x‐tetrahydroxycoprostanic acids that may be unique to each group; 3α,7α,12α‐trihydroxycoprostanic acid is a chief bile acid of crocodilians. C24 bile acids may be general in the higher lizards and in snakes. Boid snakes have pythocholic (3α, 12α, 16α‐trihydroxycholanic) acid, formed by rehydr‐oxylation in the liver of the deoxycholic (3α,12α‐dihydroxycholanic) acid made by intestinal micro‐organisms from the primary cholic acid. 3α,7α,12α,23‐tetrahydr‐oxycholanic acid is found in some snakes as well as allocholic acid; the latter also occurs in some lizards.13. The few birds examined contained cholic, allocholic and chenodeoxycholic acids. In penguins the amount of cholic‐allocholic acid is almost the same as that of chenodeoxycholic acid, but in a few other birds examined the latter is the principal bile acid. The germ‐free domestic fowl also has allocholic acid.14. Monotremes contain cholic, chenodeoxycholic and perhaps deoxycholic acids, as do some marsupials. Glycine conjugates have not been found in these mammals or in any lower group. Koala bile salts are almost entirely taurine‐conjugated 3α‐hydroxy‐7‐oxocholanic acid.15. Eutherian mammals usually have cholic and chenodeoxycholic as primary bile acids. Herbivores (except bovids) often have a preponderance of dihydroxy acids, frequently as glycine conjugates; omnivores have a mixture of tri‐ and dihydroxy acids as glycine and taurine conjugates and carnivores have taurine‐conjugated trihydroxy acids. Glycine conjugation in some species is apparently less well established than taurine conjugation; but dietary deficiencies can increase glycine conjugates. Unique bile acids, certainly or probably primary, have been found in Murinae (3α,6β,7α‐ and 3α,6β,7β‐trihydroxycholanic acids), Sus (3α,6α,7α‐trihydroxycholanic acid) and all Pinnipedia (3α,7α,23‐trihydroxycholanic acid). Other substances, such as ursode‐oxycholic (3α,7β‐dihydroxycholanic) acid, may be wholly or partly artifacts of the enterohepatic circulation; they may nevertheless be physiologically important. Deoxycholic acid (as its glycine conjugate) is normally the chief bile acid in rabbits, although it is an artifact, but Murinae re‐hydroxylate it to cholic acid. The biochemical status of 3α‐hydroxy‐7‐oxocholanic acid is disputed.16. Animals with primitive bile salts often also contain small amounts of more evolved types; the beginnings of bile salt evolution can be detected long before it appears likely to affect the physiological behaviour of the bile salts as a whole.17. The physiological functions of the bile are not sufficiently understood to permit of speculation about the advantages of any particular type of bile salt.18. Biochemical studies show that there are even more interspecific differences between bile salts than the chemistry alone suggests. Such essentially enzymic studies approach an understanding of the genes controlling the chemical characters reviewed here and may eventually throw light on fundamental questions such as the biochemical nature of vigour and senescence in evolutionary processes and the reasons why almost all classes of highly evolved vertebrates have the same C24 bile acids.