Summary
Considerable evidence suggests that an acidic calcium phosphate, such as octacalcium phosphate (OCP) or brushite, is involved as a precursor in enamel and other hard tissue formation. Additionally, there is in vitro evidence suggesting that fluoride accelerates and magnesium inhibits the hydrolysis of OCP to hydroxyapatite (OHAp). As the amount of OCP or brushite in enamel cannot be measured directly in the presence of an excess of hydroxyapatite, a procedure was developed that allows for their indirect in vivo quantification as pyrophosphate. This permits study of the effects of fluoride and magnesium ions on enamel mineral synthesis. Rat incisor calcium phosphate was labeled by intraperitoneal injection of NaH2 32PO4. The rats were then subjected to various fluoride and magnesium treatments with subcutaneous implanted osmotic pumps. They were then killed at predetermined intervals; the nascent sections of the incisors were collected, cleaned, and pyrolyzed at 500°C for 48 hours to convert acidic calcium phosphates to calcium pyrophosphate; the pyrophosphate was separated from orthophosphate by anion-exchange chromatography; and the resulting fractions were counted by liquid scintillation spectrometry. The activities of the pyro- and orthophosphate fractions were used to calculate the amount of acidic calcium phosphate present in the nascent mineral. The results demonstrated that the percentage of radioactive pyrophosphate in nascent incisors decreased with time, with increasing serum F- concentration, and with decreasing serum magnesium content. The technique described here should prove to be a powerful new tool for studying the effects of various agents on biological mineral formation.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Brown WE, Smith JP, Lehr JR, Frazier AW (1962) Crystallographic and chemical relations between octacalcium phosphate and hydroxyapatite. Nature (Lond) 196:1050–1054
Brown WE (1962) Crystal structures of octacalcium phosphate and hydroxyapatite. Nature (Lond) 196:1048–1050
Brown WE (1973) Solubility of phosphate and other sparingly soluble compounds. In: Griffith EJ, Beeton A, Spencer JM, Mitchell DT (eds) Environmental phosphorous handbook. Wiley & Sons, New York, pp 203–239
Brown WE, Nylen MU (1964) Role of octacalcium phosphate in formation of hard tissues. J Dent Res 43:751–752
Eanes ED, Meyer JL (1977) The maturation of crystalline calcium phosphates in aqueous suspensions at physiologic pH. Calcif Tissue Res 23:259–269
Eidelman N, Chow LC, Brown WE (1987) Calcium phosphate saturation levels in ultrafiltered serum. Calcif Tissue Int 40:71–78
Eidelman N, Chow LC, Brown WE (1987) Calcium phosphate phase transformations in serum. Calcif Tissue Int 41:18–26
Tomazic BB, Tung MS, Gregory TM, Brown WE (1989) Mechanism of hydrolysis of octacalcium phosphate. Scanning Microsc 3:119–127
Nylen MU, Eanes ED, Omnell KA (1963) Crystal growth in rat enamel. J Cell Biol 18:109–123
Weiss MP, Vogel JC, Frank RM (1981) Enamel crystallite growth: width and thickness study related to possible presence of octacalcium phosphate during amelogenesis. J Ultrastruct Res 76:286–292
Tomazic BB, Brown WE, Queral LA, Sadovnik M (1988) Physicochemical characterization of cardiovascular calcified deposits. I. Isolation, purification and instrumental analysis. Atherosclerosis 69:5–19
Brown WE, Eidelman N, Tomazic B (1987) Octacalcium phosphate as a precursor in biomineral formation. Adv Dent Res 1:306–313
Brown WE, Chow LC, Siew C, Gruninger S (1984) Acid calcium phosphate precursors in formation of enamel mineral. In: Fearnhead RW, Suga S (eds) Tooth enamel IV. Elsevier Science Publishers, Amsterdam, pp 8–13
Francois P (1961) Etude de la variation de composition de l'os de rat avec l'age. J de Physiologie (Paris) 53:343–344
Lim JK, Jensen GK, King OH (1975) Some toxicological aspects of stannous fluoride as a clear precipitate-free solution compared to sodium fluoride. J Dent Res 54:615–625
Robinson C, Hiller CR, Weatherell JA (1974) Uptake of 32P-labelled phosphate into developing rat incisor enamel. Calcif Tissue Res 15:143–152
Fowler BL, Moreno EC, Brown WE (1966) Intra-red spectra of hydroxyapatite, octacalcium phosphate and pyrolyzed octacalcium phosphate. Arch Oral Biol 11:447–491
Chow LC, Brown WE (1975) Formation of CaHPO4·2H2O in tooth enamel as an intermediate product in topical fluoride treatments. J Dent Res 54:65–76
Nakamura T, Yano T, Fujita A, Ohashi S (1977) Anionexchange chromatographic separation of linear phosphates with eluent containing a chelating agent. J Chromatog 130:384–386
Dawson JB, Heaton FW (1961) The determination of magnesium in biological materials by atomic absorption spectrophotometry. Biochemistry 80:99
Brown WE, Tung MS, Chow LC (1981) Role of octacalcium phosphate in the incorporation of impurities into apatites. Trans 2nd Intl Congress on Phosphorus Compounds, Boston, April 21–15, 1980, pp 59–71
Tung MS, Chickerur NS, Brown WE (1979) Studies on the hydrolysis of octacalcium phosphate. J Dent Res (Special Issue A) 58:367, Abst. No. 1112
Tung MS, Chow LC, Brown WE (1985) Hydrolysis of dicalcium phosphate dihydrate in the presence or absence of calcium fluoride. J Dent Res 64:2–5
Greenfield DJ, Eanes ED (1972) Formation chemistry of amorphous calcium phosphates prepared from carbonate containing solutions. Calcif Tissue Res 9:152–162
Schroeder HE, Bambauer HU (1966) Stages of calcium phosphate crystallization during calculus formation. Arch Oral Biol 11:1–20
Boskey AL, Posner AS (1974) Magnesium stabilization of amorphous calcium phosphate: a kinetic study. Mat Res Bull 9:907–916
Brown WE (1966) Crystal growth of bone mineral. Clin Orthop 44:205–220
Dickens B, Schroeder LW (1980) Investigation of epitaxy relationships between Ca5(PO4)3OH and other calcium orthophosphates. J Res NBS 85:347–362
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Siew, C., Gruninger, S.E., Chow, L.C. et al. Procedure for the study of acidic calcium phosphate precursor phases in enamel mineral formation. Calcif Tissue Int 50, 144–148 (1992). https://doi.org/10.1007/BF00298792
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00298792