Recent advances in the investigation of the crystal chemistry of dental enamel

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RECENT ADVANCES IN THE INVESTIGATION OF THE CRYSTAL CHEMISTRY OF DENTAL ENAMEL DUNCAN MCCONNELL College of Dentistry, Ohio State University, Columbus,

Ohio, U.S.A.

Ahdnct-In view of detailed investigatloas carried out by the writer and several other individuals, the conclusion is inescap&le that dental enamel is composed of a single “mine&” phase which is carbonate hydroxyapatite (dahllite). There is no valid reason for supposing that the crystal structure or crystal &e&try of bone or dentine is signiticantly different. Even in bovine bone no appreciable isotopic exchange with C** was obsexved by FRA~OEL(1958). so the concept of adsorption of carbonate on the surfaces of the cqstallites can be relegated to unrealistic speculation. The writer’s structural theory (1952) is essentially confirmed. INTRODUCTION

mineral, francolite, and the “mineral” phase of teeth and bones, and a fairly complete structural theory for the carbonate apatites Was presented (MCCONNELL, 1952a). The major postulate of this theory has been accepted by mineralogists and now appears in the principal reference books by STRUNZ(1957), EITEL(1954) and PALACHE, BERMAN and FRONDEL (1951). This theory, to be sure, was established on the basis of a carbonate apatite containing appreciable amounts of fluoride (francolite) rather than the carbonate hydroxyapatite (dahllite), but recent results have not only permitted the extension of this theory to include dahllite (MCCONNELL, 196Oa) but a hydrated calcium aluminum silicate (scawtite) as well (MCCONNELLand MURDOCK, 1958). The question concerning the presence of carbonate ions in apatite is not new, of course, but arose in 1822 (HAUY). IN 1952 attention

was called to the relationship between the carbonate-apatite

DISCUSSION

OF EXPERIMENTAL

RESULTS

investigations by several persons, however, have resolved this question beyond reasonable doubt with respect to the minerals dahllite and francolite. AMES (1959), for example, has demonstrated (a) that a calcium phosphate can be synthesized which contains as much as 10 per cent of carbonate, (b) that tbis substance has essentially the structure of apatite, and (c) that the carbonate is essentially within the apatite structure rather than located on the surface of the crystal&es. The significance of AMES’ contribution with respect to an understanding of teeth and bones can be appreciated only through the realization that it demonstrates the futility of attempts to learn anything about the structure of teeth and bones by such methods as dissolving them in acids or heating them. Sfia (19&I), for example, has recently made an elaborate study of the dissolution of dental enamel and found the Ca/P weight ratio in solution to range from 038 to 790 at pH 54. At lower pH values he found $e Ca/P ratios to show narrower ranges, and at pH 2-Ohe found a mean ratio of 2.30. From these data, in combination 28 Ruxnt

mz arsra

-Y

W

OF DENTAL E-

with several erroneous assumptions, he deduces that dental enamei contains a second phase, namely CaCOI in “amorphous form”, because “CaCOs is more soluble in hydrochloric acid solution than hydroxyapatite”. This experiment, of course, is comparable to one in which an attempt might be made to ascertain the metaliic composition of an alloy by analysing the solution after purriui dissolution has taken place. Obviously this could be done only after the comparative dissolution rates and their mutual influences upon each other were known with respect to each metal present and with respect to time, pH, formation of complex ions, etc. SEEL’S results, although interesting, are non-critical with respect to any conclusions related to the composition of the solid phase of dental enamel. From the viewpoint of fundamental crystal-chemical theory, the results of ADCES (1959) are not surprising, but had been theoretically anticipated, at least in a qualitative way. However, some workers are still reluctant to admit the direct applicability of these, and numerous other, data and conclusions to teeth and bones. They are perplexed by large quantities of data on Ca/P ratios, so-called exchange reactions, etc., which have been published at various times in various places, Most of these data do not disprove the close analogy between dental enamel and dahllite; in fact, in view of the discrepancies which exist, it can fairly be said that they do not prove or disprove anything, inasmuch as many of the conclusions based thereon are non sequifurs (M&o=, 1955a,b). TABLE1.

RMIOS OF LMOL CATlONSTO PHOSPHORUS FORCARDONATE -APATITEMINEJlALS (By petission from The American Mineralogist)

Locality and reference’

Mineral DabJlite

272

Dabllitc

446

Dailuitc

3.36

BluEton, Ohio McC4xIN~ 1960 Mouillac, France McCoN?JELL,1938 St. Paul’s Rocks. Atlantic

545

WASHINOTON,1929 F-is Lake, B.C.

Francolitc

Francolite

1.80

Franc&c

3.36

Francolite

4.43

Fmncoiitc

l-98

Francolitc

3.40

/

Ca’

P

Ca. Mg. Na

P

Ca, Na. Kt

P. S

ca

P. S

ca5

P

co, Me

P

1.85

co,M8

P. v

2-02

Ca. Mrs. Na

P

I.81

ca. Sr

P

l-80

co. Ms

P

wASHlNO’TON,

H. S. (1929).

PorlFwN. 1927 Busumbu, Uganda Diva, 1W7 Sta!Tel, Gemany GRUNERand MCCONNELL,1937 Richterweld. South Africa DE VurruS. 1942 Yorkshire, England Dmq1938

1

Tavistock, Devon. Eqland Suuxu. et ol., 1939

lIWirrarfcxtheTaMeangivenbyMCCCINNBI Anwr. Min. 14,369. t Oxides dipeprded f -garbi wut:

ca’/P (Atomic)

I

(196Oa). except

as “impuritia” m: Fk&*, Alp,. Sio,. FeO, Fe& Mn,Q. Al&, MpO. SiQ and organic C.

DUNCANM-

30

Of particular interest are Ca/P ratios of the solid material. Extensive anaIyticai results on teeth and bones have appeared, as well as those on various synthetic substances which gave apatite-like dilTraction patterns. With respect to bones, mm’s (1938) results are of interest because all of his molar ratios, except for three avian samples, exceed the theoretical value for hydroxyapatite (1467). Likewise all of the Ca/P ratios for mineral carbonate apatites exceed the theoretical value when magnesium and sodium equivalents are summed with calcium (MCCONNELL, 196Oa). Indeed, this ratio was found to be as high as 292 for francolite from South Africa (Table 1). All of these data are consistent with the theory on the repiacement of phosphorus by carbon in the apatite structure, but one cannot expect a direct correlation between carbonate content and the Ca/P ratio for reasons which will be discussed. TABLE2. Smuc’rtn~U -tctTtm OF m~RoxyApATm (By permission from Die Natw-wiw&m) R&ruK!e*

atA)

BALE Posta and &EPHEN..N Jmsw and MILLER THEWLIS,GLOCK and MURRAY KUMENTand DUIN HENDRlCWl: ui. Powit, Ruapr and Dmro Ww (precipitated) WA(hydrothermal) TRAUTZ -w

9.49*0-03 9-45 ~450*0902 943 9-42 9-42&O-03 9.43, 9~41O_c,O-005 9403*0901 9-421 iO-003 9.421 &O-O01 9*420*0-001 9.41, 9-q 944*0-01

PERDOK

BRARZRUR

-w WAl.UEmQ

c.G> 6-89*0-03 6-89 6-871~0901 6-88 6-94 6.94f0-03 6-88, 6-865*0-m 6-886fO+XIl 6.881 fO-003 6-882rto’oOl 6-885*0~001 6-88, 6% 6+60*0-m

Rckencu for the tabie are given by MCCONNELL (196Ob). t socplled tricalcium phosphate dihydrate. 5 Aqueous precipitated Wicakium phosphate. l

Synthetic “hydroxyapatite” (in&ding so-called tricalcium phosphate hydrate) has been reported to show CajP from 1.95 to as low as 1.33, although it is usuaily admitted that more than one phase is present when the lower ratio is obtained (ARNOLD, 1950). Re-examination of X-ray diRraction data on various “hydrcxyapatites” clearly indicates (Table 2) that these measurements could not have been made on the same pure crystalline phase because the discrepancies exceed the experimental errors by large factors (MCCONNELL, l!XOb). Under these circumstances it becomes necessary to assume not only that the unit cell dimensions fail to indicate those of pure hydroxyapatite, but that the reported Ca/P ratios are either those of mixtums (containing more than one phase) or are, those of variants containing

THE CRrsrAl.

ciS&USRY

Of

DENTAL

ENAUEL

31

isomorphic combinations, such as: Ca,,,(OH),(PO,)r(COsOH) and/or ~IO(H,O~(PO~XO,HJ, which have Ca: P: HtO equal to 2.00: 1: 0.30 and 250: 1: 1.50, respectively. It is to be noted that although both of these hypothetical compounds have Ca/P ratios which exceed 1.67, only one of them contains carbon dioxide. Consequently no correlation between the Ca/P ratio and the carbon dioxide content would be expected, and none is observed. In general, it can be assumed that Ca/P ratios less than the theoretical value are to be accounted for either (a) because of failure to include Na and Mg with Ca, or (b) because of the presence of other extraneous phases, such as brush& or octacalcium phosphate (BROWN efof., 1957). The near coincidence of most of the more intense X-ray diffraction maxima of these substances with those of apatite could readily prevent detection of small amounts of these substances. So-called alpha tricalcium phosphate hydrate not only fails to show consistent analyses with respect to the amount of water present, but it merely approximates Ca/P- 15. Reports on the u perk&city of this precipitate are 944 r~O.Ol, according to WALWYS (1952) and 9-40, A according to BRU~EUR(19511). it should be obvious that the compositions of these two substances must be significantly different. Equally obvious is the futility of attempting to reconcile something with Ca/P= 15 with the apatite structure, because (a) not only does this supposition require vacancy ‘among one-fourth of the calcium positions of one symmetrical sort, but (b) it requires that these vacancies must be random, because their symmetrical location would surely give rise to recognizable structural differences which have not been observed. Indeed, inasmuch as this proposal involves omission of one calcium atom of the fourfold position (l/3,2/3,0), the planes of symmetry at l/4 and 3/4, as well as the 6* axis and the two-fold axes, would necessarily become inoperative, and the space-group symmetry could no longer be that of hydroxyapatite (P6Jm). An alternative is to assume that every fourth unit cell has all four calcium positions vacant, a proposition which becomes equally untenable. It is deduced, then, that such a substance as Ca#O&~2H10 cannot possibly be isostructural with hydroxyapatite ?n the basis of fundamental crystallographic considerations. Since all substances with this alleged composition give the diffraction pattern of hydroxyapatite, there is no valid evidence for the existence of any such compound as tricalcium phosphate hydrate. In a recent attempt to obtain an explanation of the isostructural character of hydroxyapatite and tricalcium phosphate hydrate, BRGSEUR (1958) calls upon the results of m and WONDRA~SCHEK (1957) who prepared some lead-containing apatites with the supposed composition Pb,A(YO&, where A Na, K, Rb, Cs, Ti; and Y-P, As, V. On this basis BRUSEUR assumes that Ca,H(PO& also would be isostructurai, but this assumption violates one of the principal rules of crystal chemistry: hydrogen, in general, will not isomorphously replace alkali ions, and vice versa. Thus his theoretical explanation-he gives no experimental evidence-is quite unacceptable. It must be admitted that knowledge of the crystal chemistry of bone is considerably complicated by the extremely small size of the crystallite% but the inherent

32

DuNcN4MCCONNELL

natural d%icultics have been made appreciably

worse by misguided interpretations of non-critical experiments. However, the relationship between dental enamel and dahllite is a straightforward one which is not confused by the size of the crystallites. Thus we have, in terms of the X-ray diffraction data, most excellent correlation between human dental enamel and a fossil dental enamel of a mastodon (MC~DNNELL, 196Oa). Both of these substances produce very good, if not excellent, diffraction patterns (Figs. 1 and 2). Of course, both contain appreciable amounts of carbon dioxide but give no evidence whatever of consisting of more than one crystalline phase. The suggestion has been made that an amorphous phase might be present, and might contain both carbonate and phosphate ions in addition to calcium. This suggestion surely deserves consideration in view of a vast accumulation of geochemical knowledge. There is no amorphous mineral substance known to occur in nature which contains any such quantity of carbon dioxide as occurs in bone. Furthermore, there is no naturally occurring amorphous mineral substance which consists essentially of calcium phosphate. With respect to bone, it can be concluded that the amount of carbon dioxide present probably could not be accounted for even if the entire substance were amorphous. Certainly in the case of dental enamel, which appears to be essentially crystalline by both optical and X-ray methods, the carbonate content cannot be present as a constituent of an amorphous phase. The suggestion has been made that the carbonate is adsorbed on the surfaces of the crystal&es of bone. Again, these conclusions were base-d on non-critical experiments, which ‘are not subject to unique interpretation. Recent experiments using G4 as a tracer have indicated that no appreciable amount of the carbonate is subject to isotopic exchange. FRANCIS (1958) found this to be true for bovine bone, and AMES(1959) found it to be true of his synthetic carbonate apatite. AMESshould be quoted in this connection; he states: “. . . this carbouatf2 can be entirely present within the apatite lattice when the apatite contains less than 10 per cent by weight COs*‘*. This amount, to be sure, is more than twice the quantity present in bone or dental enamel. AhLEsstates .further: “Structural formulae, equilibrium data and P tracing show no indications of the carbonate being present in Hendricks’ ‘voids’ or adsorbed to any great extent on apatite surfaces . . .‘* Another conclusion of AMES is interesting: “The variable composition of the apatite phase, even in this relatively simple system, shows the fallacy of attempting to apply the laws of sparingly soluble compounds to apatite”. In 1952, at the Josiah Macy, Jr. Confemnce on Metabolic Interrelations, the “incongruent solubility of silicates” was refernd to by me (MCCONNELL,1952b, p. 171) in an attempt to point out that one would not expect bone or tooth substance to behave differently from most silicates. One versed in silicate chemistry surely would not expect to be able to write a “solubility product” for bone mineral. Although I may have introduced the phrase, “incongruent solubility” to emphasize the important similarities, nevertheless, such principles are well known to silicate chemists who surely know that feldspar can be dissolved in water or acid, but nothing remotely resembling feldspar will m-precipitate from this same solution at ordinary temperature within a reasonable length of time-perhaps IO years.

ml!. CRYSTAL CHEMHTUY OF DeMM

ENAMEL

33

The fallacy of depending solely upon X-ray diffraction data as a baais for deciding whether dental enamel, for example, is composed of hydroxyapatite or a carbonate hydroxyapatite was demonstrated by O~MONDand SAWS (1959). They accurately measured forty samples of recent and fossil teeth of vertebrate animals_ Some representative measurements of II periodicities were as follows (in AngstrUm units): 9.45 (human), 940 (Miocene rodent), 9.42 (recent lunghsh), 9.37 (Permian lungfish), 9.39 (recent shark), 9.37 (P&n shark). They conclude “that fluoridization alone is not the cause of the change with time of vertebrate tooth apatite unit-ceil dimensions”. These findings and conclusions, including the smaller u dimension for a recent shark than for a Miocene rodent, are entirely consistent with the theory (MCCONNELL, 1960b) that the variations in carbon dioxide and hydroxyl contents influence these dimensions, as well as the fluorine content. Thus it is completely meaningless to state that human dental enamel is hydroxyapatite because unit cell dimensions closely resemble those of “hydroxyapatitr”. The question immediately arises: which “hydroxyapatite” in Table 2? The fundamental emphasis, to be sure, must be placed on the fact that all of these substances contain significant, although variable, amounts of carbon dioxide. Such differences in structural periodicities as those observed by OSMOND and SAWIN (1959) cannot he explained on the basis of variations in the amount or composition of some admixed amorphous constituent. These crystal-chemical differences must be produced by compositional variations among these specimens.

CONCLUSIONS Biochemists do not seem to be reluctant to call the carbonate hydroxyapatite of bone and teeth a “mineral” substance, and they refer to its formation as “mineralization”in this sense meaning merely the formation of mineral matter, apparently. However, if they are quite willing to borrow the terms-and there are good reasons for their doing so-one might raise the important question: why ;Lre they reluctant to borrow the fundamental principles. which apply to mineralogical chemistry also? At this juncture it might be mentioned that the term, mineralization, implies something beyond the mere formation of a mineral to most mineralogists, but this is a minor distinction over which we can pass without further consideration. However, the complete disregard of applicable mineralogical and crystal-chemical concepts cannot be considered a trivial matter. The composition of teeth and bones cannot be correctly expressed as hydroxyapatite. These substances are carbonate hydroxyapatites or,.if one wants a good mineralogical term, dahilite. We have made no attempt to discuss the details of several new lines of evidence which indicate the carbonate-apatite composition of teeth and bones. My own results have been integrated with those of Aws (1959), FRANCIS (1958) and MASLENNIKOVand KAV~~SKAYA (1956). These results were concerned with fossil dental enamel, synthesis of carbonate apatite, bovine bone, and the phosphorite minerals. All of these data point toward the principle that the carbonate component is within the lattice of the apatite structure, and, furthermore, that no significant amount of carbonate is adsorbed on the surface of the crystallitcs. All of these investigations,

34

DUNCANMCCONNELL

except the work of F~AN$KB, were concerned directly with the question of the isomorphic substitution of carbon for phosphorus, and in all cases this fundamental premise was ver&d-completely vindicating those mineralogical authorities who had already accepted tlris theoretical concept on the basis of earlier evidences. The textbooks which refer to the substance of dental enamel and bone as “hydroxyapatite” contain a fundamental error, which now has been repeated so often, and in so many places, that it will probably require many years to correct this error. Twentythree years have already elapsed since GRUNEX,M&oand ARMSTRONG (1937) pointed out that this substance is a carbonate hydroxyapatite. BOGERT and HASDNGS (1931) had done so earlier, but could not tioncile their formula with the crystal structum of apatite. Acknowledgenrenf-This work was carried out under a special research felfowship of the National Institute of Dental Research, U.S. Public Health Service. REFERENCES Ahnq L. L., Jr. 1959. The 8enesis of carbonate apatites. EWR. Geol. 54, 829-841. mt.n, W. P. 1950. The nature of precipitated calcium phosphates. Trans. Faraday Sot. 46, lQ$il-1072. &GERT, I.. r. and HAJfpIos, A. B. 1931. The calcium salt of bono. 1. biol. Gem. 94.473481. Wwautt, H. 1958. Consid&ations notwelles sur la constitution possible du phosphate tricalcique hydtatt. B&J. aced. Brig. Cl. Sci. (ser. 5) 44, 5U7-513. BROWN,W. E., Lwt, J. R, Sbnr~, J. P. and Fm, A. W. 1957. Crystallography of octacllcium pltosphate. 1. Amer. dwm. Sec. 79, 5318. Enw, W. 1954. Tke Physical Chemisfry of the Silicnres. University of Chicago Press. Fa.wwts, C. 1958. La localiwion de l’anhydride carbonique darts les sels osseux et dentaints. Etudi& par la m&ode des &hanges isotopiques. &II. Sac. Chim. biol.. Parii 40. 1341-1347. GRUNER,J. W., M&~NNEI&D. and hthSTRmG, W. D. 1937. The relationship between crystal structure and chemical composition of enamel and dentin /. biol. Gem. 121, 771-781. H&Y, L’Aaat. 1822. Trait& de Midrafogie, Vol. I. p. 503. Bach&r. Paris. KLEMErrr, R. 1938. Die anorganische Skeletsubstam. Ihre Zusamt&nsetxung, natiirliche und kiinstli& Bilduntt. Nururwissensc~fie~~ X145-152. McCotwu~~,~D. 195k. The crystal &e&is&y of carbonate apatites and their relations&ip to the cotnposrtton of calci&d tissues. J. &m. Res. X,5363. MCCONNELL, D. 1952b. The crystal c&mistry of francoIite and its relationship to calctid animal tissues. T-ctionv of the Macy Conference on Metabolic interreIa&zs, Vol. 4. pp 166-184. Josiah Macy Jr. Foundation, New York. MCCONNELL, D. 1955a. The application of mineralogical theories to the “mineral” phase of teeth and bones. Biochim. biophys. Acta 17, 450-451. Mw, D. 1955b. The problem of the carbonate apatites. J. Amer. &em. Sot. 77,2344. McCoy D. 196Oa. The crystal chemistry of dahllite. Amer. Min. 45,209-216. MCCONNEU, D. 196Ob. The stoichiometry of hydroxyapatite. Naturwissenschqften 47, 227. McCo-, D. and MURDOCH,J. 1958. The crystal chemistry of scawtite. Amer. Min. 43,498-502. hhsLENNl?COV, B. M. and KNlTsKAYA, F. A. 1956. Fosfatnorn wshchewe fosforitov. me phosphate substance of phoaphorites). Dokl. Akad. Nauk SSSR 109. -992. Marutaa, L. and Wo~~twscnart, H. 1957. Neue Verbindungen mit apatitartiger Struktur-H. Die Gruppe die Alkali-Blei-Verbindungen. Z. K&wti&sr. 109. I I&l 14. O~MOND,J. K. and SAGAN,H. J. 1959. Unit-cell dimensions of recent and fossil tooth apatitm Bu/X geoi. Sot. Amer. 70,95A (Abstract). Pm. C., BEIUW, H. and FROC. 1951. Dana’s System of Mineraiogy, Vol. 2. John

Wiley, New York. STEEL,& $fV&he solution of calcium and phosphorus from the intact enamel surface. /. dent.

* , STRUNZ.H. 1957. MInrrologirchr Tobclen, 3rd ed. Geest anti Port@, Leipzig. Wm. R. 1952. Contribution a 1’6tude des apatites phosphocalciques. Ann. Chim. 7.808-W.

THE

CRYSTAL

CHEMISTRY

OF DENTAL

ENAMEL

(b)

(4

FIG. 1. X-ray diffraction diagrams for several substances. Filtered FeK radiation, r=57*3 mm. (a) Francolite. (b) Dental enamel. Cc)Oral calculus. (d) In ~Trrocaiculus. (e) Dentine.

PLATE

I

DUNCAN MCCONNELL

FIG. 2. Comparison of X-ray diffraction diagrams of in vitro calculus and fossil dental enamel (dahllite). Filtered CuK radiation. r=28*7 mm. (For detailed measurements on spacings and intensities for dahllite see MCCONNELL, 196Oa.) (a) In vitro calculus. (b) Fossil dental enamel.

PLATE 2