WO2010095001A1

WO2010095001A1 - Submicron monetite powders production

Info

Publication number: WO2010095001A1
Application number: PCT/IB2009/050683
Authority: WO
Inventors: Ahmet Cuneyt Tas
Original assignee: Yeditepe Universitesi
Priority date: 2009-02-19
Filing date: 2009-02-19
Publication date: 2010-08-26

Abstract

This invention relates to the synthesis of submicron monetite powders to be used as highly soluble and resorbable bone substitute materials. Monetite (CaHPO₄) powders were prepared by reacting precipitated and submicron calcium carbonate powders together with phosphoric acid in a medium of pure ethanol. The solutions did not contain any water. Monetite synthesis reaction was performed at room temperature. Monetite synthesis reaction was typically completed in 3 hours. The obtained monetite powders were submicron and consisting of nano sheets of monetite stacked together. The mildly acidic monetite powders were aged in a calcium-containing saline solution at 37^oC for 6 days to produce neutral calcium phosphate powders with nanosize particles. The calcium-containing saline solution contained 142 mM Na⁺, 5 mM K⁺ and 50 mM Ca²⁺ ions.

Description

Title of Invention: SUBMICRON MONETITE POWDERS

PRODUCTION

[ 1 ] Field of the Invention

[2] This invention relates to the production of submicron particulated powders of monetite.

[3] Background of the Invention

[4] Monetite (CaHPO₄, dicalcium phosphate anhydrous, DCPA) is one of the mildly acidic and soluble (at the physiological pH) calcium phosphate (CaP) phases [1, 2] and currently finding a significant place to itself in the powder components of self- hardening CaP putties, pastes, and cements used for skeletal repair [3-6]. Moreover, it must hereby be noted that even the world's very first patented CaP putty-like paste development invention, by T. D. Driskell et al. [7] back in 1974, involved powders of CaHPO₄ mixed with those of self-hardening alpha-Ca₃(PO₄)₂. CaHPO₄ is also used in powder form in some toothpastes, chewing gums and in food processing industry to act as acidity regulator, anti-caking agent, dough modifier and emulsifier (food additive number: E341) [8]. CaHPO₄ has a triclinic unit cell with the following lattice parameters; £i=6.910, b=6.627, c=6.998 A, alpha=96.34°, beta =103.82°, and gamma=88.33° [9, 10], with a calculated density of 2.92 g/cm3. Its triclinic-pinacoidal structure consists of CaHPO₄ chains held together by Ca-O bonds and three types of hydrogen bonds [H]. The dissolution kinetics of CaHPO₄ in water at 37 C° was investigated by Lebugle et al. [12]. Ca²⁺ and HPO4² ions were initially released from CaHPO₄ and completely stopped after 4 days in contact with water at 37C° [12]. The Ca/P atomic ratio in water, in which these dissolution experiments were performed, also decreased with time and was 0.75 after 15 min of dissolution; it finally (after 4 d) reached a value of 0.62. The dissolution of DCPA was found to be incongruent [12]. This incongruence was explained by the formation of a thin apatitic (apatite-like) calcium phosphate (Ap-CaP) layer on the surfaces of CaHPO₄ particles through a topotactic reaction, which hindered further dissolution [12-16]. The logarithm of the thermodynamic solubility (i.e., log K_sp) of CaHPO₄ was reported by McDowell et al. [16] to be -6.90 and -7.04 at 25C° and 37C°, respectively. For comparison purposes, log K_sp values at 25C° for CaHPO₄ . 2H₂O (brushite), CaCO₃, Ca₁₀(PO₄)₆(OH)₂ (hydroxyapatite, HA), beta-Ca₃(PO₄)₂(whitlockite), and Ca₉(HPO₄)(PO₄)_S(OH) (Ca-deficient HA) are -6.60, -8.55, -117.1, -81.7, and -85.1, respectively [17]. These numbers simply relate to the solubility of a given substance; for instance, Ca-deficient HA (-85.1) is much less soluble than CaHPO₄ (-6.90). This is basically why some self- hardening CaP pastes contain, for instance, about 25 wt% CaHPO₄ [3,4] in their powder components. While CaHPO₄ powders are being produced, it is not difficult at all to convert CaHPO₄ . 2H₂O (brushite, dicalcium phosphate dihydrate, DCPD) powders into single -phase CaHPO₄ by "dehydration" at high temperatures. Two methods of dehydration can be envisaged and the previous literature provides ample examples to those. The first method of CaHPO₄ synthesis, which used brushite powders as the starting material, consisted of dehydrating 400 g of brushite (CaHPO₄ . 2H₂O) in 4 L of 0.07 M H₃PO₄ solution by boiling for 72 h, followed by filtering the resultant CaHPO₄ powders [16, 18]. The second method of dehydrating brushite powders to obtain CaHPO₄ simply consisted of heating brushite in a static air atmosphere over the temperature range of 200 to 250⁰C [19-23]. However, both of these dehydration techniques would have the following disadvantage in terms of ceramic powder science: it would be quite difficult to significantly alter the particle size and shape distribution present in the initial brushite powders. Synthetic brushite powders usually exhibit large (i.e., 10 to 130 μm) tabular, needle-like, lath-like, or rectangular prismatic crystals, and at high salt concentrations large acicular crystals are also formed [24-27]. Since such an idiomorphic tabular or acicular crystal habit in brushite is very difficult to circumvent (if not impossible), monetite synthesis procedures based on the dehydration of brushite will not be able to produce submicron particles [28].

[5] Chenot [29] provided solution preparation recipes for the dehydration of the crystals of brushite to monetite in aqueous slurries heated from 85° to 98°C, and observed the formation of large monetite crystals with rhombohedral diamond, plate-like, rectangular or needle-like habits.

[6] Takagi et al. [30] simultaneously and slowly (in 5 h) added, dropwise, equimolar solutions of Ca-chloride and Na₂HPO₄ (one liter each) into one liter of water heated to 95°C, and thus obtained hexagonal prismatic crystals of CaHPO₄. Griffith and McDaniel [31] first formed CaHPO₄ . 2H₂O crystals by reacting an aqueous Ca(OH)₂ slurry with H₃PO₄ solutions and then mostly converted those into CaHPO₄ in 2 hours by increasing the temperature of the resultant suspension to about 95°C. In a very similar study, Martin and Brown [32] later reported the precipitation of CaHPO₄ at 85° C from an acid-base reaction of H₃PO₄ and Ca(OH)₂, however, the aqueous slurry containing the precipitates was continuously stirred for 16 h at 85°C to ensure complete reaction between Ca(OH)₂ and H₃PO₄. The high temperatures (85°-98°C) used in the above-mentioned studies made possible the dehydration of any brushite, which might initially form, to monetite. Louati et al. [33], on the other hand, reported the synthesis of large crystals of CaHPO₄ by slowly evaporating an aqueous solution of the stoichiometric mixture of Ca(NO₃)₂ . 4H₂O and NH₄H₂PO₄ at around 80⁰C.

[7] There were a number of attempts to develop new powder synthesis methods which would facilitate a reduction in the mean particle size of CaHPO₄. Chen et al. [34] synthesized CaHPO₄ particles in oil-in-water (o/w) system and water-in-oil (w/o) system with a novel membrane microdispersion mixing technique. In this study, sodium dodecyl sulphate (SDS)-containing Ca-acetate aqueous solutions were used as the water phase, whereas butanol-phosphoric acid mixture was the oil phase. Chen et al. [34] reported the synthesis of submicron particles of CaHPO₄. Tokuoka et al. [35] were reacting two solutions at room temperature for 24 h; triethyl phosphate solution in water and a solution of Ca-nitrate in water containing cetyltrimethylammonium bromide. However, their CaHPO₄ precipitates were not submicron and were heavily agglomerated.

[8] Kong et al. [36] prepared reverse microemulsion by adding calcium chloride and sodium hydrogen phosphate aqueous solutions into a mixture of Span 80®, Tween 80®, and n-heptane to get two kinds of emulsions, n-buranol was used to adjust the emulsions to transparent state. Calcium phosphate was prepared by adding phosphate microemulsion to the calcium microemulsion. Kong et al. [36] reported to form micron-size needles of CaHPO₄. Wei et al. [37] used reverse micelles solution of water and cyclohexane containingeither cetyltrimethylammonium bromide (CTAB) or poly- oxyethylene-8-dodecyl ether (C12E8) surfactants and n-pentanol as co-surfactant as organized reaction microenvironments for monetite precipitation. Well-crystallized CaHPO₄ nanoparticles with various morphologies such as spheres, monodisperse nanofibers and bundles of nanowires were obtained in this study. Xu et al. [38] obtained CaHPO₄ nanoparticles by spray-drying. Jinawath et al. [39] used monocalcium phosphate monohydrate (Ca(H₂PO₄)₂ . H₂O) and Ca(OH)₂ as the starting materials in an autoclave operated between 160°- 200⁰C to obtain plate-like and large crystals of monetite. Thomas and Dehbi [40] formed CaHPO4 in the solid state by ball milling Ca(H₂PO₄)₂ . H₂O and Ca₃(PO₄)₂ powders together at room temperature.

[9] Ma et al. [41] synthesized CaHPO₄ particles consisting of nanosheets, by a one-step microwave-assisted heating method at 95°C using CaCl₂ . 2H₂O, NaH₂PO₄, and sodium dodecyl sulfate (SDS) in water/ethylene glycol (EG) mixed solvents. The synthesis procedure of this study was quite remarkable that it resulted in spindle, flower- or bundle-like CaHPO₄ particles (3 to 5 μm in diameter) consisting of stacked nanosheets.

[10] Aoki et al. [42] studied the morphology of monetite crystals formed when Ca(OH)₂ or CaCO₃ particles (ca. 20-100 μm in particle size) were slowly added to concentrated (10%) aqueous solutions Of H₃PO₄ at 40⁰C in highly acidic solutions.

[11] Eshtiagh-Hosseini et al. [43] slowly added a solution of H₃PO₄ diluted in methanol to a solution of Ca(NO₃)₂ . 4H₂O dissolved in methanol, followed by aging the mixed solutions from 24 h to 6 d in sealed glass containers to obtain heavily agglomerated and CaCO₃-containing CaHPO₄ particles. [12] The Japanese patent application No. JP 1037409 known in the state of the art discloses synthesizing monetite powders in water-based solutions, with particles larger than 10 microns in diameter at reaction temperatures over the range of 40° to 100⁰C.

[13] The USA patent application No. US3927180 known in the state of the art discloses hydrothermal conversion of brushite into monetite.

[14] The Japanese patent application No. JP59217610 known in the state of the art discloses production of calcium-phosphorus based apatite of high quality by easy method, by reacting a calcium compound with a phosphorus compound in a reaction medium containing water and a water-miscible organic solvent.

[15] In the state of the art, monetite (CaHPO₄) powders were typically produced by using high-temperature (between 75° and 98°C) synthesis procedures, and they were thus having particle sizes greater than 10 microns.

[16] In the state of the art, neutral calcium phosphate powders from monetite powders could only be prepared by immersing those powders in strongly basic solutions at high temperatures. Moreover, the monetite powders known in the art do not have submicron particles.

[17] In the previous studies, CaHPO₄ powders consisting of much larger prismatic crystals were converted into apatitic calcium phosphate in basic solutions (pH > = 10.5) of Na₂ CO₃ or NaOH at temperatures between 75° and 98°C.

[18] In the contemporary bone defect repair applications, it is no longer a valid argument to use, as an implant material, the Ca-hydroxyapatite (Ca₁₀(PO₄)₆(OH)₂) synthesized in the laboratory, because stoichiometric hydroxyapatite is not resorbable and it does not take part in the natural bone-remodeling processes even after 10 years of in vivo implantation. As a live example, the use of highly soluble CaSO₄ has been preferred by numerous orthopedic surgeons over Ca-hydroxyapatite in recent decades. In orthopedic and maxillofacial clinical applications using CaHPO₄ is a much better alternative to CaSO₄. The in vivo resorbability of CaHPO₄ is strongly affected by the particle size, shape and morphology distribution present in its powders. Currently available CaHPO₄ powders are far from having desired physico-chemical properties to be considered in biomedical applications.

[19] Summary of the Invention

[20] The objective of this invention is to realize a synthesis procedure which enables the user to produce submicron monetite powder particles at room temperature.

[21] Another objective of the present invention is to realize monetite powders that are used in developing and formulating new putties, pastes or cements for clinical use in maxillofacial and orthopedic bone substitute, bone defect repair applications.

[22] Another objective of the present invention is to produce submicron particulated powders of monetite at room temperature in pure ethanol solutions containing small amounts of orthophosphoric acid (H₃PO₄).

[23] Another objective of the present invention is to realize a robust synthesis technique for submicron and mildly acidic monetite powders consisting of nanosheets glued to one another, in stark contrast to available powders of much larger particles, and showed that these submicron powders do easily transform into a high surface area and neutral calcium phosphate phase, mimicking the biological bone mineral in morphology.

[24] Another objective of the present invention is that it does not employ any toxic organic or inorganic substances in decreasing the particle sizes of currently available monetite powders (typically 50 microns) to submicron sizes.

[25] The chemical synthesis technique does not employ any toxic organic or inorganic substances in decreasing the particle sizes of currently available monetite powders (typically 50 microns) to submicron sizes depicted in this invention. The ideal synthetic bone substitute material to be used by the surgeons must have a high solubility and must be able to convert itself, in vivo, into the biological calcium phosphate mineral of natural bones in less than a month.

[26] A further objective of the invention is to realize a process in which transformation of monetite powders is achieved at pH values and temperatures close to the physiological conditions.

[27] Detailed Description of the Invention

[28] "Submicron Monetite Powders" realized to fulfill the objective of the present invention is illustrated in the accompanying figures, in which,

[29] Figure 1 is the view of the SEM photomicrograph of precipitated CaCO₃ powders.

[30] Figure 2 is the view of the FTIR and XRD (inset) traces of precipitated CaCO₃ powders.

[31] Figure 3 is the view of the SEM photomicrograph of product powders obtained after reacting 2.96 g CaCO₃ and 2 mL of H₃PO₄ in 100 mL EtOH for 4h at RT.

[32] Figure 4 is the view of the FTIR spectrum of powders shown in Figure 3.

[33] Figure 5 is the view of the FTIR spectrum of single-phase CaHPO₄ powders produced by reacting 2.96 g CaCO₃ and 4 mL of H₃PO₄ in 100 mL EtOH at RT for 3h.

[34] Figure 6 is the view of the XRD trace of single-phase CaHPO₄ powders produced by reacting 2.96 g CaCO₃ and 4 mL of H₃PO₄ in 100 mL EtOH at RT for 3h

[35] Figure 7a- Figure 7d is the view of the SEM photomicrographs (at increasing magnifications) of single-phase CaHPO₄ powders produced by reacting 2.96 g CaCO₃ and 4 mL of H₃PO₄ in 100 mL EtOH at RT for 3h.

[36] Figure 8 is the view of the XRD traces showing phase development in 2.96 g CaCO₃ and 100 mL EtOH mixtures containing different H₃PO₄ amounts (RT, 3h stirring each; * calcite peak). [37] Figure 9 is the view of the XRD traces showing phase development in 2.96 g CaCO₃ and 4 mL H₃PO₄ in 100 mL EtOH mixtures as a function of increasing stirring time at RT (+ MCPM peaks).

[38] Figure 10 is the view of the Comparative FTIR spectra of single-phase monetite powders synthesized in EtOH (trace- 1) and those aged in Ca-containing saline solution at 37°C for 6 d of soaking of CaHPO₄ (trace-2).

[39] Figure 11 is the view of the XRD spectrum of submicron monetite powders aged in

Ca-containing saline solution at 37°C for 6 d.

[40] Figure 12a - Figure 12b is the view of the SEM photomicrographs of submicron monetite powders after ageing in Ca-containing saline solution at 37°C for 6 d.

[41] This invention relates to the production of monetite powders with monodisperse and submicron particles.

[42] Monetite synthesis consisted of stirring submicron CaCO₃ powders in glass containers or bottles for 3 hours together with appropriate volumes of concentrated H₃ PO₄ (ortho-phosphoric acid) solutions in pure ethanol. Submicron monetite powders obtained were single-phase with a powder particle texture comprising stacked- nanosheets, and did not contain any unreacted CaCO₃. CaHPO₄ powders of this invention were fully converted at 37°C, in 6 days, into a high surface area calcium phosphate phase in a solution comprising 142 mM Na+, 5 mM K⁺ and 50 mM Ca⁺² dissolved in water. The human blood plasma contains 142 mM Na⁺ and 5 mM K⁺, and this is only where this special saline solution has a resemblance to biological fluids. The human blood plasma contains, on the other hand, only 2.5 mM Ca²⁺.

[43] The calcium phosphate powders disclosed in this invention have high surface area, and the special saline solution developed to produce more neutral calcium phosphate powders from mildly acidic monetite powders would allow the in situ addition of a number of biopolymers, proteins, drugs or growth factors. One cannot add any of these biological or bioactive agents into Na₂CO₃ or NaOH solutions heated at high temperatures.

[44] This invention discloses a robust synthesis technique for submicron and mildly acidic monetite powders consisting of nanosheets glued to one another, in stark contrast to available powders of much larger particles, and showed that these submicron powders do easily transform into a high surface area and neutral calcium phosphate phase, mimicking the biological bone mineral in morphology and chemical composition, over a short period of few days when soaked at 37°C in solutions similar to a physiological saline solution.

[45] Calcium carbonate of the calcite form (CaCO₃, Fisher Scientific, Fair Lawn, NJ,

Catalog No: C63, precipitated), orthophosphoric acid solution (H₃PO₄: Merck KGaA, Darmstadt, Germany, Catalog No: 100573, 85%) and absolute ethanol (Merck KGaA, Darmstadt, Germany, Catalog No: 100983, > 99.9%) were used to synthesize monetite (CaHPO₄) powders of this invention.

[46] Produced samples were characterized by powder X-ray diffraction (XRD; D8-

Advance, Bruker AXS GmbH, Karlsruhe, Germany), scanning electron microscopy (SEM; S-4700, Hitachi, Tokyo, Japan), and Fourier-transform infrared spectroscopy (FTIR; Nicolet 550, Thermo-Nicolet, Woburn, MA). The BET surface area of powder samples was determined by applying the standard Brunnauer-Emmet-Teller method to the nitrogen adsorption isotherm obtained at -196°C by using a Micromeritics ASAP 2020 instrument. Powder samples for the XRD analyses were first ground in an agate mortar by using an agate pestle and then sprinkled onto ethanol-damped single-crystal quartz sample holders to form a thin layer, followed by tapping to remove the excess of powder. The X-ray diffractometer was operated at 40 kV and 30 mA by using monochromated Cu K_alpha radiation. XRD data (over the typical range of 10 to 50° 20) were collected with a step size of 0.02° and a preset time of 1 sec at each step. FTIR samples were first ground in a mortar, in a manner similar to that described for the preparation of XRD samples, then mixed with pure and moisture-free KBr powders in a ratio of 1:100, followed by forming a 1 cm-diameter thin pellet by using a uniaxial cold press. 128 scans were performed at a resolution of 3 cm ¹. As obtained (i.e., without any grinding in mortars) powders examined with the scanning electron microscope (SEM) were sputter-coated with a thin Au-Pd layer prior to imaging, to impart surface conductivity to the samples.

[47] Calcite (CaCO₃) powders used in this invention were selected on the basis of their mean particle size (i.e., being submicron) and morphology. It should be emphasized that the resultant CaHPO₄ particle size and nano-textured morphology observed in this invention could not be obtained if we were using other calcite powders which have much larger rectangular prismatic or rhombohedral particles. The SEM photomicrograph depicted the particle size and shape distribution in the starting calcite powders (Figure 1). These powders comprised spindle-like particles, 1 to 1.5 μm in length and 500 nm in width. Although these spindles were sometimes forming hard agglomerates, they had uniform dimensions throughout the powder body. The FTIR data of these calcite powders showed a weak band at around 3643 cm ¹, which was indicating that these commercial powders were actually produced from the aqueous car- bonation of a slurry of fine particles of Ca(OH)₂ (Figure 2) [44]. The remaining IR bands were characteristic of the calcite phase [45, 46]. The XRD trace depicted that these powders were single-phase calcite (Figure X).

[48] CaHPO₄ Synthesis: In a successful synthesis run to yield only single-phase CaHPO₄,

100 mL of ethanol was first placed into a 100 mL-capacity Pyrex® media bottle (Fisher Scientific, Catalog No: 06-423-3B) containing a Teflon®-coated magnetic stir bar. 2.96 g of CaCO₃ powder (=0.0296 mol Ca) was added into ethanol and the formed opaque suspension was magnetically stirred at room temperature (RT: 22+1⁰C) for 5 minutes. Finally, 4 mL of H₃PO₄ (=0.0592 mol P) was added into this calcite suspension, the glass bottle was tightly capped and the bottle contents were stirred at RT for 3 h. At the end of 3 h, the bottle was opened; the precipitates were filtered (Whatman, No. 42 paper), washed with 25 mL of ethanol, the filter paper having the precipitates was finally dried overnight at 37°C in a static air oven.

[49] When the Ca/P molar ratios in ethanol (EtOH) solutions were made equal to 1.0 (i.e.,

100 mL EtOH + 2.96 g CaCO₃ + 2 mL cone. H₃PO₄), the resultant powders, even after 4 h of stirring at RT, were not single-phase CaHPO₄. The SEM photomicrograph showed that the CaCO₃ spindles of the starting powder were mostly intact, but occasional flower-like nanosheets of CaHPO₄ were observed to form (Figure 3). As expected, the FTIR trace of these samples exhibited a biphasic material (Figure 4), i.e., calcite and CaHPO₄.

[50] The synthesis recipe described in the first paragraph did only differ from the above recipe in terms of doubling the H₃PO₄ concentration in the solution, bringing the Ca/P molar ratio in the synthesis vessel to 0.50. Figures 5 showed the FTIR and XRD traces obtained, respectively, for the sample described in the first paragraph. The FTIR trace showed only the characteristic IR bands of CaHPO₄ [47-49]. The IR bands observed were matching very well with those reported in the previous literature (Figure 5) [47-49]. The XRD trace conformed to all the diffraction peaks of CaHPO₄ listed in ICDD PDF 70-360 over the 2Θ range of 10 to 50° (Figure 6) [50]. These samples were produced by stirring at RT for only 3 h, stirring for 4 h did not produce any noticeable changes in Figure 5 and Figure 6. This meant that stirring the CaHPO₄ suspension in absolute ethanol beyond 3 hours was not necessary. At the end of 3 h of stirring at RT, the pH of the mother liquors (prior to the filtration of the CaHPO₄ particles) were found to be constant at around 3.25 + 0.2.

[51] The SEM photomicrographs of Figure 7 through 3f displayed the particle morphology of CaHPO4 powders produced in 100 mL absolute EtOH at RT by using 0.0296 mol of precipitated CaCO₃ powder and 0.0592 mol H₃PO₄ (85% solution) as the starting materials. Especially, Figures 7c and 7d clearly exhibited the nanosheets of CaHPO₄ stacked together to form the ellipsoidal particles.

[52] Effect of H₃PO₄ volume: The XRD traces depicted the influence of different volumes of H₃PO₄ added to the suspensions created by mixing, in each case, 2.96 g of precipitated CaCO₃ and 100 mL EtOH, following 3 h of stirring at RT after the addition of H₃PO₄ (Figure 8). It was thus apparent that when the Ca/P molar ratio in the reaction vessels at the start of stirring was made equal to 0.50 (i.e., 4 mL H₃PO₄ addition), single-phase CaHPO₄ powders were obtained at the end of 3 h. Lesser volumes of H₃ PO₄ additions resulted in the observation of unreacted CaCO₃ (Figure 8), identified by the most intense diffraction peak of the calcite phase (ICDD PDF 5-586) observed at 29.415° 2Θ. [53] Effect of stirring time: The XRD traces showed the effect of stirring time on the phase development (Figure 9). All of the samples of Figure 9 were produced according to the synthesis recipe of "100 mL EtOH + 2.96 g CaCO₃ + 4 mL H₃PO₄.". [54] The stirring times employed were 0.5, 1, 2, and 3 h, respectively. The experimental

X-ray diffraction peaks of the monocalcium phosphate monohydrate (MCPM, Ca(H₂ PO₄)₂ . H₂O) phase matched well with those of ICDD PDF 75-1521 for MCPM (Figure 9). Therefore, synthesis of monetite in ethanol at RT, as described in this invention, took place according to the below reactions in two distinct steps, in accord with reactions (1) and (2):

[55] CaCO₃ (s) + 2H₃PO₄ (aq) -> Ca(H₂PO₄), . H₂O (s) + CO₂ (g) (1)

[56] Ca(H₂PO₄), . H₂O (s) -> CaHPO₄ (s) + HPO₄ ²" (aq) + 3H÷(aq) + OH (aq) (2)

[57] According to the XRD data of Figure 9, Rxⁿ(l) was taking place immediately after adding the concentrated H₃PO₄ solution into the CaCO₃-EtOH suspension. However, at the end of 0.5 h of stirring at RT the resultant solids were biphasic (Figure 9), i.e., consisting of about 70% MCPM and 30% CaHPO₄. This meant that Rxⁿ(2) was also taking place within the first 30 minutes. By the end of 2 h of stirring, the material formed was again biphasic, but this time comprising 70% CaHPO₄ and 30% MCPM. Three hours of stirring was obviously enough to convert all MCPM into CaHPO₄. [58] The comparative FTIR and XRD traces of Figure 10 and Figure 11, respectively, showed the transformation of mildly acidic CaHPO₄ into neutral calcium phosphate within 6 d of ageing at 37°C in the calcium-containing saline solution of this invention. The symmetric and anti- symmetric stretching of the PO₄ ³" group were observed at 1090, 1025, 958, 602 and 560 cm ¹. The weak shoulder at around 1297-1300 cm ¹ was attributed to the smaller presence of HPO₄ ²" ions [52]. HPO₄ ²" ions do also have a band at 870 cm¹. The stretching mode of the O-H group was observed in Fig. 5a at 3570 cm- i

[59] The XRD data of Figure 11, on the other hand, depicted a calcium phosphate pattern resembling to that of bone mineral, contradicting what was previously reported by Lebugle et al. [12]. Since the XRD data were generated from ground powders, it negated the assertion (Figure 11) (by Lebugle et al. [12]) that the CaHPO₄ particles will be covered by a thin external layer of apatitic CaP in water at 37°C and thus it will hinder the further transformation of monetite. This was certainly not observed in the current invention. The XRD data of Figure 11 is almost identical with that of the natural bone mineral, i.e., the X-ray diffraction data obtained from human bones is very similar to this. [60] The rather smooth conversion of mildly acidic CaHPO₄ into a neutral calcium phosphate phase similar to that of bone mineral was quite an expected result because CaHPO₄ is one of the very interesting compounds of the calcium phosphate family which is not thermodynamically stable in solutions with pH values greater than 5.5. This was why CaHPO₄ has been one of the indispensable compounds used in self- hardening orthopedic and maxillofacial cement development [3-6].

[61] The pH values of the aging solutions (at the end of every 24 hours, just prior to full replenishment of the used solutions with fresh solutions) were recorded as 5.7 +0.1 at 36°-37°C.

[62] The SEM photomicrographs of Figure 12a - Figure 12b the nanosize particles of the calcium phosphate phase obtained from the starting monetite powders of CaHPO₄ of this invention. The entire process reported in this invention can also be regarded as a low-cost method of producing calcium phosphates similar to the biological bone mineral powders free of any ionic impurities, such as NO₃ ²", NH₄ ⁺, etc.

[63] The previous literature gave numerous examples on the hydrolysis of CaHPO₄ powders in hot and strongly basic solutions, such as NaHCO₃, Na₂CO₃, (NH₄)₂CO₃, or NaOH at temperatures between 75° and 100⁰C [53-58].

[64] In this invention, hydrolysis temperature was chosen to be 37°C, which is the physiological temperature. Again in this invention preferred to develop a hydrolysis solution (for monetite) to be as benign as possible so that in the follow-up applications, for instance, one could easily consider adding certain proteins or growth factors to such hydrolysis solutions.

[65] Working Example- 1: In a successful synthesis run to yield only single-phase CaHPO

₄, 100 rnL of ethanol was first placed into a 100 mL-capacity Pyrex® media bottle (Fisher Scientific, Catalog No: 06-423-3B) containing a Teflon®-coated magnetic stir bar. 2.96 g of CaCO₃ powder (=0.0296 mol Ca) was added into ethanol and the formed opaque suspension was magnetically stirred at room temperature (RT: 22+1⁰C) for 5 minutes. Finally, 4 mL of H₃PO₄ (=0.0592 mol P) was added into this calcite suspension, the glass bottle was tightly capped and the bottle contents were stirred at RT for 3 h. At the end of 3 h, the bottle was opened; the precipitates were filtered (Whatman, No. 42 paper), washed with 25 mL of ethanol, the filter paper having the precipitates was finally dried overnight at 37°C in a static air oven.

[66] Working Example-2: To test the transforming ability of the mildly acidic single- phase CaHPO₄ powders of this invention into a neutral calcium phosphate phase similar to that of the bone mineral, a calcium-containing saline solution with a starting pH of 10 was prepared and the CaHPO₄ powders were aged at 37°C in this solution for 6 d. This saline solution was prepared as follows. 0.1118 g of KCl (Merck, Catalog No: 104933) and 2.49 g of NaCl (Merck, Catalog No: 106404) were first dissolved, re- spectively, in 300 niL of distilled water. The solution thus contained 5 rnM K⁺ and 142 rnM Na⁺, identical with the K⁺ and Na⁺ concentrations of human blood plasma. 2.205 g of CaCl₂ . 2H₂O (Merck, Catalog No: 102382), corresponding to 50 mM Ca²⁺, was then dissolved in this solution at RT. This solution was a calcium-containing saline solution and had an initial pH of 6.1 + 0.1 at RT. Its pH was raised to around 10 by adding a 0.01 mL aliquot of concentrated NH₄OH (Merck, Catalog No: 105423). 250 mL portions of the solution were placed into 250 mL-capacity glass media bottles together with 0.5 g of the single-phase CaHPO₄ powders as synthesized above, the bottles were tightly capped, placed into a 37°C oven and kept undisturbed over the 6-day aging period. The only exception to this was the quick replenishment of the mother liquors (ca. 250 mL) at every 24 hours. At the end of 6 d of aging at 37°C, the recovered powders were filtered (Whatman, No. 42 paper) and washed with 750 mL of distilled water, followed by drying at 37°C in a static air oven for 24 h. Calcium phosphate powders with a BET surface area of 55+3 m²/g were obtained.

[67] Monetite powders currently available do have particle sizes in the range of 5 to 50 microns, and such commercial powders are extremely sluggish in transforming into the biological bone mineral when implanted in the form of calcium phosphate putties or pastes.

[68] Monetite (CaHPO₄) is also used in powder form in some toothpastes, chewing gums and in food processing industry to act as acidity regulator, anti-caking agent, dough modifier and emulsifier (food additive number: E341).

[69] Submicron monetite powders can also be used as resorbable scaffolds and carriers for drug- and biomolecule-delivery applications.

[70] Precipitated CaCO₃ (calcium carbonate) powders having submicron particles were used as the starting material in producing submicron monetite powders. Monetite in powder form is used in orthopedic and dental repair cement, toothpaste, and chewing gum formulations. Especially, the high in vivo resorption rate of monetite is the reason for its preference in orthopedic and dental cement formulations designed for use as bone defect fillers or bone substitutes

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Claims

[Claim 1] A process for producing submicron monetite which comprises reacting precipitated calcium carbonate powder having submicron particles together with phosphoric acid in a medium of pure ethanol; and characterized in that reaction temperature is from 18° to 30⁰C, most preferably 21-22°C.

[Claim 2] A process according to Claim 1 characterized in that reaction time is from 2 to 4 hours.

[Claim 3] A process according to Claim 1 and 2 characterized in that calcium carbonate powder is mixed with ethanol at a concentration from 2 to 5 grams per 100 mL of ethanol.

[Claim 4] A process according to Claim 1-3 characterized in that phosphoric acid is mixed with calcium carbonate-ethanol mixture at a concentration from 2.7 to 6.8 mL per 100 mL of ethanol.

[Claim 5] A process according to Claim 1 characterized by a calcium phosphate phase similar to that of bone mineral, which comprises reacting submicron monetite powders by immersion in an aqueous solution containing only Na⁺, K⁺, Cl" and Ca²⁺ ions in distilled water.

[Claim 6] A process according to Claim 5 characterized in that reaction temperature is 36.5° to 37°C. [Claim 7] A process according to Claim 5-6 characterized in that immersion time is from 4 to 6 days. [Claim 8] A process according to Claim 5-7 characterized in that concentration of Na⁺ is between 130 and 170 mM, most preferably 142 mM. [Claim 9] A process according to Claim 5-8 characterized in that concentration of K⁺ is between 3 and 7 mM, most preferably 5 mM. [Claim 10] A process according to Claim 5-9 characterized in that concentration of Ca²⁺ is between 25 and 75 mM, most preferably 50 mM. [Claim 11] A process according to Claim 5-9 characterized in that concentration of Cl- is between 200 and 300 mM. [Claim 12] A process according to Claim 5 characterized in that sodium salt is NaCl. [Claim 13] A process according to Claim 5 characterized in that potassium salt is KCl. [Claim 14] A process according to Claim 5 characterized in that calcium salt is either CaCl₂ or CaCl₂ . 2H₂O. [Claim 15] A process according to Claim 1, characterized in that the monetite is produced in the form of submicron ellipsoidal particles, consisting of monetite nanosheets tightly stacked on top of one another.