Introduction

1. In 1967, Mayaharaet al. reported the localization of Alkaline phosphatase in the enamel organs. Sapir-Koren and Livshits explained about Pi/PPiratio, regulation of linked bone mineralization and phosphate homeostasis by inorganic forms of phosphate itself. Pyrophosphate (PPi) consisted of two molecules of inorganic phosphate (Pi) joined by a hydrolyzablehighenergyester bond. 　High-polymeric inorganic polyphosphates were found in living organisms by L. Liberman in 1888. These compounds are linear polymers containing a few to several hundred residues of orthophosphate linked same as PPi. And, polyphosphate is food addictive for unfavorable metal ion chelater. These findings can be concluded. 1. Phosphatase is localized at the place that calcifying. 2. Polyphosphate could be hydrolyzed to phosphate by the enzyme. 3. Polyphosphate can hold calcium ions by their chelate ability. Therefore, Polyphosphate seems tobe a key substance of calcification. In spite of abundunt reports about preparation of Hydroxyapatite sinceMoreno et al. in 1968, we could not find the explanation about polyphosphate except for calcification inhibitor. Preparation of Calcium OxideCalcium Oxide (Tateho Chemical Co.) were sintered 800ﾟ Cin electron furnace, Confirmed by X-Ray Diffraction Pattern. Preparation of Polyphosphate 85% Ortho Phosphate (TOSOH NIKKEMI Co.） was placed in Silicon Carbide pot, stored in 200ﾟ C for few days, then 300ﾟ Cfor a week. Synthesize 2g of Polyphosphate and 1.5 to 3.5g of Calcium oxide were placed in a pressure vessel(Fig.3), mixed and added 10ml of water, then stored in 200ﾟ C in electro furnace for 24 hours. The chamber was cooled by current water, PH ware measured by indicator (PH Indecator Strip 109535, Merk) The slurry was washed until the PH goes neutralized, and dried. Hydrothermal hydoxyapatite growth from Calcium oxide and Polyphosphate Biomaterials and Technology Department of Conservative Dentistry, Showa University School of Dentistry NARUSAWA Hideaki, OWADA Hiroyuki, TAKIGUCHI Yuichi, MIYAZAKI Takashi Hypothesis and Objectives References MayaharaH, Hirano H, Saito T, Ogawa K. The new lead citrate method for the ultracytochemical demonstration of activity of non-specific alkaline phosphatase (orthophosphoric monoester phosphohydrolase). Histochemie. [Internet]. 1967 Jan;11(1):88–96. Available from: http://www.ncbi.nlm.nih.gov/pubmed/5589645 Sapir-koren R, Livshits G. Bone Mineralization and Regulation of Phosphate Homeostasis. 2011;8(6):286–300. Lieberman, L.: Uber das Nuclein der Here und KunstlicheDarstellungeines Nucleus Eiweiss und Metaphosphatsaure. Bet. Chem.-Ges., 21 (1888), 598-607. Moreno EC, Gregory TM, Brown WE. Preparation and Solubility of Hydroxyapatite. J. Res. Natl. Bur. Stand. - A. Phys. Chem. 1968;72(6):773–82. ArendsJ, Schuthof J, Van der Linden WH. Bennema P. Van den Berg PJ (1979) reparation of pure hydroxyapatite single crystals by hydrothermal recrystallization. J Crystal Growth 46:213-220 Roy DM (1971)Ctystal growth of hydroxyapatile. Mat Res Bull 6:1337-1340 MengeotM. Hlarvill ML (1973) Gilliain OR. hydrothermal growth of calcium hydroxyapatite single crystals. I Crystal Growth 19:199-203 Conclusion Results and Discussions Materials and Method As preliminary experiment, we mixed calcium oxide and polyphosphate, after exothermic reaction, it becomes the dry clot. The clots were moved to beaker and added a half-cup of water, then the water came to a boil again. After a few minutes, upper clear water indicates PH 3. By XRD analysis, precipitated powder assigned as Monetite and Calcium hydroxide mixture. No significant change was found at PH and chemical after a week.Calcium hydroxide should be reacted with water as usual; it seemed poly phosphate kept calcium hydroxide stable in acidic water by chelation. Introduction Acknowledgement • A picture of dried sample is shown at Fig.6. fibrous object can be seen by unaided visions. (red circle, magnified to right pictures) • Rod like crystals can be seen low magnification SEM.(Fig7, X100) • Fig.8 showed Picked up “large crystal” under magnification glass. (Macrophotography) • Fig.9: Large crystal mounted on SEM sample stage.( Diameter 14mm)Biggest crystal ( at least 5mm length) is mounted vertically to take A plane picture. • Fig10 and 11Compare SEM and light microscopy, same individual, similar angle. • Left: SEM Right: Light microscope view. • The crystals are shown straight, hexagonal profile. • Diameter of A plane is about 150μm. • We successfully synthesized good apatite crystals from Poly phosphate and Calcium oxide under hydro thermal method. The finding shows a possibility; Poly phosphate is relevant to the calcification in a human body.And the method has three advantages, 1. Free from elements except Calcium Phosphorous Oxygen Hydrogen.2. Free from carbonate contamination.3. Extremely lower pressure and temperature compare to previous report. • These should be useful for experimental or industrial apatite production. However, growth factors, conditions were sills have uncertainty, and could not assign cubic and square crystals. These may investigate in future work. • Fig.6 • Fig.7 I would like to thank RyojiTanaka, Sagami Chemical Research Institute in Kanagawa, Japan. He took beautiful crystal picture by Nikon digital SLR cameras with macro and ultra macro lenses. • Fig.8 • Fig.3 • 4. Observation • Dried powder ware observed by SEM ( Hitachi TM-3000 ) EPMA analysis (SWIFTED,Oxfordinstruments, UK), XRD analysis (ShimadzuXRD-6100, Shimadzu, Kyoto, Japan) and Light microscope (Keyence VHX-1000, Tokyo,Japan) • Fig.9 • Fig.4 showed SEM and X-Ray diffraction pattern of 3.5 (upper) 2.5 (middle) 1.5 (lower), respectively. Upper and Middle sample were assigned as Calcium hydroxide and Hydroxyapatite mixture by XRD pattern. Lower showed monetite pattern. Note the extremely low magnifications. • Fig.5 result of EPMA mapping. Total Ca/P molar ratio is exceed stoichiometry, however, we found big difference between the measured points. Elementally mapping clearly showed Phosphorous deficient cubic crystal precipitated around hexagonal crystal. • The result is concluded in Table 1.Plate like crystal (non apatite Calcium Phosphate) is not shown higher than 2.0 calcium oxide. Best result should be obtained between 2.0 to 2.5 Chemical reaction from polyphosphate and calcium oxide and hydroxide to apatite is shown in fig. 1. According to preliminary experiment, hydrolysis of polyphosphate doesn’t occurred at room temperature. • Fig.10 • Fig.4 Fig.1 • Fig.11 • Therefore, we speculated that breaking the polyphosphate chain causes freeing phosphate ion and chelated calcium ion, it could precipitate apatite.( Fig.2) • The objective of this study is to validate above hypothesis; whether hydrothermal condition force to the apatite precipitation or not. • Several researchscientists were challenged to grow larger apatite. Arends et al reported 0.1- 3.0 mm length 430-500ﾟ C 200Mpa in 1971, Roy (103Mpa, 745-900ﾟ C) Mangeot(310Mpa, 390ﾟ C). No big apatite products are available; the reason should be the cost to generate ff extreme high temperature and pressure. In contrast, our condition is only 1.3 MPa (vapor pressure at 200ﾟ C) • The result of this study was to prepare the large apatite crystals in lower temperature and pressure in comparison to previous work. In addition, it could be synthesized without other ions, such as sodium, potassium, ammonium and chloride. And starts from acidic solution, it may avoid carbonate contamination. The result indicates polyphosphate is suitable for apatite crystal growth under hydrothermal condition. • Fig.5 Nikon ECLIPSE ME600 X100 Polarized ？ • Fig.2 • Table.１