Solid Calcium Phosphate Research Articles

Chemistry misconceptions associated with understanding calcium and phosphate homeostasis

Successful learning of many aspects in physiology depends on a meaningful understanding of fundamental chemistry concepts. Two conceptual diagnostic questions measured student understanding of the chemical equilibrium underlying calcium and phosphate homeostasis. One question assessed the ability to predict the change in phosphate concentration when calcium ions were added to a saturated calcium phosphate solution. Fifty-two percent of the students correctly predicted that the phosphate concentration would decrease in accord with the common ion effect. Forty-two percent of the students predicted that the phosphate concentration would not change. Written explanations showed that most students failed to evoke the idea of competing chemical equilibria. A second question assessed the predicted change in calcium concentration after solid calcium phosphate was added to a saturated solution. Only 11% of the students correctly predicted no change in calcium concentration; 86% of the students predicted an increase, and many based their prediction on a mistaken application of Le Chatelier's principle to heterogeneous equilibria. These results indicate that many students possess misconceptions about chemical equilibrium that may hamper understanding of the processes of calcium and phosphate homeostasis. Instructors can help students gain greater understanding of these physiochemical phenomena by adopting strategies that enable students achieve more accurate conceptions of chemical equilibria.

Synthesis of nanocrystalline calcium hydroxyapatite from calcium saccharates and ammonium hydrogen phosphate

The synthetic hydroxyapatite Ca 10 (PO 4 ) 6 (OH) 2 (HAP) is a mineral component of the bone tissue of mammals. HAP is biocompatible with the human body, capable of biointegration with the bone tissue, has no negative influence on the immune system, is nontoxic, and shows osteoconductive behavior. Therefore, HAP is used to design ceramic materials for bone implants [1]. The activity of powders toward sintering in the production of ceramic materials increases with a decrease in the particle size of the powder [2, 3]. For bioceramics, an important feature is the absence of acute cytotoxicity, which is determined by the chemical purity of the initial powder. To prepare HAP containing no toxic or associated products difficult to remove, the reaction of calcium hydroxide with ammonium phosphate is used. In this case, one component, Ca(OH) 2 , is taken as a suspension [4]. An obvious drawback of the synthesis of calcium phosphates from a poorly soluble compound is that dissolution of Ca(OH) 2 is the rate-limiting step, and this increases the duration of synthesis. The formation of calcium phosphates may also occur on the surface of Ca(OH) 2 particles, which shifts the reaction to a region of slower diffusion through a solid calcium phosphate bed and gives rise to an inhomogeneous target product. The use of soluble calcium compounds for preparing highly dispersed powders is preferred, because this implements the known bottom-up approach to the creation of a dispersion system (from small to large) [5]. In the presence of saccharose, the solubility of calcium hydroxide increases due to formation of soluble calcium saccharates [6]. Organic compounds being adsorbed on the surface of the inorganic crystals formed prevent their growth and thus ensure the formation of a finely crystalline powder [7]. The purpose of the present work was to prepare nanocrystalline HAP containing a biocompatible associated product, saccharose, from ammonium hydrogen phosphate and calcium saccharates. The study was carried out with solutions of calcium saccharates Ca n C 12 H 22 – 2 n O 11 at different n ( n is the Ca : C 12 H 22 O 11 molar ratio in the saccharate synthesis) and HAP samples synthesized from different calcium saccharates and ammonium hydrogen phosphate.

New apatite-based supports prepared by industrial phosphoric acid for HDS catalyst synthesis

Abstract New NiMo hydrodesulfurization catalysts supported on a variety of calcium phosphate solids made of apatite materials, have been synthesized. Apatites have been prepared by a co-precipitation method in basic medium, with the use of industrial phosphoric acid of various purities (29%, 54% or 60 wt.% P2O5) instead of research-grade ammonium hydrogenophosphate. Characterization techniques have revealed that supports obtained from industrial 60% P2O5 phosphoric acid and from ammonium hydrogenophosphate are both Ca-deficient hydroxyapatites. The presence of fluoroapatite has been evidenced on the supports prepared with industrial acid of lower purity. Molybdenum then nickel ions have been deposited on supports by successive impregnation. Catalytic activities have been measured for the hydrogenolysis reaction of dimethyldisulfide. Catalysts prepared with hydroxyapatite synthesized from ammonium hydrogenophosphate and phosphoric acid 60% present similar activities and are found to be more active than an industrial catalyst NiMoP/Al2O3 used as a reference. The higher activity is explained by the presence of superficial HPO42− species and surface cationic vacancies leading to a better dispersion of the NiMo active phase.

Formation of hydroxyapatite–polyphosphazene polymer composites at physiologic temperature

Aspects of the formation of bone analog composites at 37 degrees C are described. The composites are composed of hydroxyapatite (HAp) and the calcium salt of a biocompatible polymer and are capable of forming under in vivo conditions. Composite formation involves the formation of monolithic HAp from particulate calcium phosphate precursors while Ca ions liberated to the aqueous medium in which this reaction is occurring form crosslinks with the acidic polymer. The reactants are poly[bis(carboxylatophenoxy)phosphazene] (acid-PCPP), tetracalcium phosphate [Ca4(PO4)2O, TetCP], and anhydrous dicalcium phosphate (CaHPO4, DCPA). The effects of the proportion of polymer (5, 10, or 15 wt %) on the kinetics of HAp formation were studied. Compositional evolution of the solid calcium phosphates present was followed by X-ray diffraction and infrared spectroscopy analyses. HAp formation through a dissolution-precipitation process provided a mildly alkaline medium suitable for deprotonation of the acid-PCPP and for the formation of the calcium crosslinks, as monitored by infrared spectroscopy. Concurrence of crosslinking of the polymer and HAp formation was established, indicating true composite formation can be realized at physiologic temperature.

Brain calcifications induce neurological dysfunction that can be reversed by a bone drug

Perivascular calcifications within the brain form in response to a variety of insults. While considered by many to be benign, these calcium phosphate deposits or “brain stones” can become large and are associated with neurological symptoms that range from seizures to parkinsonian symptoms. Here we hypothesize that the high concentrations of calcium in these deposits produce reversible, toxic effects on neurons that can be overcome with “bone” drugs that chelate solid phase calcium phosphates. We present preliminary findings that suggest a direct association between progressive neurological symptoms and brain calcification and the symptomatic improvement of seizures, headaches, and parkinsonian symptoms in patients treated with the bisphosphonate drug disodium etidronate, normally used to treat bone diseases. Future, longitudinal epidemiological studies and randomized trials will be needed to determine the true relationship between brain stones and neurological disorders as well as the utility of bisphosphonates in their prevention and treatment.

Antibody-conjugated soybean oil-filled calcium phosphate nanoshells for targetted delivery of hydrophobic molecules

Hollow calcium phosphate nanoparticles capable of encapsulating poorly water-soluble molecules were produced by self-assembly. Previously reported were solid calcium phosphate nanoparticles and water-filled calcium phosphate nanocapsules suited for encapsulating mostly hydrophilic, but not hydrophobic compounds. Here, calcium phosphate was deposited around 100 nm diameter, 1,2-dioleoyl-sn-glycero-3-phosphate stabilized soybean oil nanoemulsions using either calcium chloride or NaOH titrations to achieve shell thickness between 20–70 nm. The surface was functionalized with carboxylic acid via the addition of carboxyethylphosphonic acid to attach Molecular Probes AB-594C antibody using sulpho-n-hydroxysuccinimide and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride with an efficiency of ∼70%, while retaining near complete antibody function. Hydrophobic pyrene was encapsulated with an efficiency of 95%, at concentrations much higher than its water solubility limit, and exhibited spectral features characteristic of a hydrophobic environment. These materials can be used in the targeted delivery of many useful, yet poorly water-soluble pharmaceutical and nutraceutical compounds.

Accumulation of nitrite in denitrifying barriers when phosphate is limiting

Permeable in situ denitrifying barriers can remove nitrate from groundwater. Barriers may be constructed by filling an excavated area with a porous mixture of sand, fine gravel, and substrate or by the injection of a nonaqueous phase substrate into an aquifer. The substrate stimulates the development of a denitrifying microbial community by providing an electron donor. The objective of this study was to determine the ability of denitrifying barriers to function under low-phosphate conditions. Sand columns injected with a soybean oil emulsion were used as laboratory models of denitrifying barriers. When a natural groundwater containing 17 mg l −1 nitrate-N and 0.009 mg l −1 phosphate-P was pumped through the columns, only a small amount of nitrate was removed from the water and, in some effluent fractions, 52% to 88% of the influent nitrate had converted to nitrite. Nitrite also accumulated when the phosphate concentration of the groundwater was increased to 0.040 or 0.080 mg l −1 phosphate-P. Only when a 0.160 mg l −1 phosphate-P supplement was added to the groundwater was there a loss of nitrate without a large accumulation of nitrite. The addition of solid calcium phosphate or rock phosphate to the sand columns was found to provide adequate phosphate for denitrification in short-term studies. These studies point out the need to ensure that adequate phosphate is present in denitrifying barriers especially when such barriers are used beneath phosphate-binding soils.

Thin film of low-crystalline calcium phosphate apatite formed at low temperature

Surface modification of biomaterials to improve biocompatibility without changing their bulk properties is desired for many clinical applications and has become an emerging technology in biomaterial research and industry. In the present study, a simple method of coating the solid surfaces of metals, organic tissue matrices, glasses, inorganic ceramics as well as organic polymers with a thin film of low-crystalline apatite crystals (LCA) was developed. Acidic solution containing calcium and phosphate ions was neutralized with alkaline solution to form calcium phosphate precipitates at low temperature. Precipitates of solid calcium phosphate particles were, then, removed by filtration. Concentration of free ions in the filtered ion solution which were not involved in the formation of calcium phosphate precipitate was high enough to induce the heterogeneous nucleation on the solid surfaces at low temperature. Thin layers of calcium phosphate crystals were formed on the surfaces of metals, glasses, inorganic ceramics, organic polymers including hydrophobic ones, and biological tissue matrices with this solution. The thin layer of crystals consisted of poorly crystalline calcium phosphate apatite crystals which contain high amount of labile ions like bone crystals and did not dissolve in the physiologic solutions. Various cells attached to this crystal layer and proliferated well.

Tale of three cratons: tectonostratigraphic anatomy of the Damara Orogen in northwestern Namibia and the assembly of Gondwana : Prave A.R., Geology, 1996, 24/12 (1115–1118)

Modern phosphogenesis occurs on continental margins influenced by upwelling and high primary productivity. The formation of phosphatic sediments is coupled to global climate fluctuations, biological cycling of phosphorus and local redox conditions. Although the processes involved in phosphogenesis are well described, high-resolution data on the redox and stable isotope systematics in Recent in-situ phosphorites are scarce. In this contribution, we investigate the trace element and sulfur, nitrogen and organic carbon stable isotope composition of Recent in-situ phosphatic sediments off the coast of Namibia. Also, we examine the reliability of different widely used geochemical proxies in phosphatic sediments. Our results suggest a shift from sulfidic to suboxic conditions, coinciding with the maximum in solid calcium phosphate mineral concentration. This shift is accompanied by unidirectional changes in Mo and Re enrichments and TOC abundance. Relatively low pyrite δ34S values (ca −20‰) of phosphatic sediments indicate open system fractionation during phosphogenesis. The initiation of phosphogenesis is also accompanied by negative shifts in sedimentary δ13Corg and δ15N values. Phosphate associated sulfate (PAS) δ34S values are lower than modern seawater sulfate values, suggesting the involvement of chemolithotrophic sulfur oxidation. Our results show a shift in redox conditions from sulfidic to (sub)oxic, coupled with active sulfur cycling are prerequisites for phosphogenesis. Phosphatic sediments show substantial enrichments in U and V highlighting the complexity of using these elements, as well as V/(V + Ni) and V/Cr, as redox proxies particularly in phosphorites and phosphatic sediments.

Evidence of calcium phosphate supersaturation in the loop of Henle.

We have used published rat micropuncture data to construct a matrix of ion concentrations along the rat nephron. With an iterative computer model of known ion interactions, we calculated relative supersaturation ratios in all nephron segments. The collecting ducts and urine showed expected supersaturation with stone-forming salts. Fluid in the thin segment of the loop of Henle may be supersaturated with calcium carbonate and calcium phosphate under certain conditions. Because calculations cannot predict the actual course of crystallization, we made solutions to mimic, in vitro, presumed conditions in the loop of Henle. The solid phases that formed were analyzed by X-ray powder diffraction, electron microprobe, and infrared spectroscopy. All samples were identified as poorly crystallized or immature apatite. The descending limb of Henle's loop creates a unique condition as it extracts water but not sodium, bicarbonate, calcium, or phosphate, giving a calcium concentration at the bend of 3 mM, pH 7.4, and a phosphate concentration that varies from 0.8 to 48 mM, depending on parathyroid hormone and dietary phosphate. We conclude that conditions in the thin segment potentially could create a solid calcium phosphate phase, which may initiate nucleation of calcium oxalate salts in the collecting ducts, potentiating nephrolithiasis and nephrocalcinosis.

Effects of temperature on the formation of hydroxyapatite

The formation of hydroxyapatite by an acid-base reaction between solid calcium phosphates at temperatures from 5 to 60 °C was examined. The basic reactant used is Ca4(PO4)2O, while the acidic reactants include CaHPO4, CaHPO4 · H2O, and Ca(H2PO4)2 · H2O. Rates of heat evolution during reaction were determined by isothermal calorimetry. The variations in the proportions of reactants and hydroxyapatite and the formation of intermediate products were assessed by x-ray diffraction. Development of microstructure was observed. Generally, hydroxyapatite formation occurs by rapid initial reaction followed by a period during which reaction occurs slowly. Apparent activation energies calculated for the reaction when CaHPO4 is the acidic reactant show differing values depending on its surface area. When the acidic reactant is Ca(H2PO4)2 · H2O, intermediate products are formed. At low temperatures the intermediate is CaHPO4 · 2H2O, while at higher temperatures it is CaHPO4. Above 38 °C, the rate during the period of slow reaction decreases with increasing temperature. This appears to be related to the retrograde solubilities of the reactants, CaHPO4 · 2H2O and CaHPO4, and of HAp.

Physicochemical Background for Ambiguity of Clinical Recommendations in Treating Phosphate Nephrohthiasis

Efficiency of clinical recommendations for curing and prevention of phosphate nephrolithiasis founded on physicochemical investigations is unexpectedly low and apparently did not exceed 50%. We tried to consider possible physicochemical causes of this fact. For this purpose the influence of changes of urine pH and of calcium, phosphate and complexon concentrations on the probability of different precipitation phases of solid calcium phosphate has been reviewed. The importance of tendencies of changing the urine dynamic physicochemical parameters such as the precipitation induction period and the amplitude of urine pH variations in time has been shown. It was established that the probability of the existence of Ca3(PO4)2, Ca8H2(PO4)6.5H2O, CaHPO4.2 H2O and MgNH4PO4.6 H2O in urine depends on the time of its transition from the supersoluble state and is determined in the final analysis by the relationship of formation rates and solid phase dissolution rates. This was verified by experiments and by data obtained in the clinical laboratory.

Solid state31NMR studies of the conversion of amorphous tricalcium phosphate to apatitic tricalcium phosphate

The hydrolytic conversion of a solid amorphous calcium phosphate of empirical formula Ca9 (PO4)6 to a poorly crystalline apatitic phase, under conditions where Ca2+ and PO4(3-) were conserved, was studied by means of solid-state magic-angle sample spinning 31P-NMR (nuclear magnetic resonance). Results showed a gradual decrease in hydrated amorphous calcium phosphate and the formation of two new PO4(3-)-containing components: an apatitic component similar to poorly crystalline hydroxyapatite and a protonated PO4(3-), probably HPO4(2-) in a dicalcium phosphate dihydrate (DCPD) brushite-like configuration. This latter component resembles the brushite-like HPO4(2-) component previously observed by 31P-NMR in apatitic calcium phosphates of biological origin. Results were consistent with previous studies by Heughebaert and Montel [18] of the kinetics of the conversion of amorphous calcium phosphate to hydroxyapatite under the same conditions.

Kinetics of Heterogeneously Initiated Precipitation of Calcium Phosphates1

The precipitation of calcium phosphate from stable supersaturated solutions was studied under conditions of pH/Ca-stat and pH-stat. The pH and Ca2+ activity of the titrand were kept constant by means of specific ion electrodes coupled to automatic titrators. The precipitation was initiated by the addition of well-characterized Domino soil, which had no identifiable calcium phosphate solids. The precipitate was characterized as CaHPO4·2H2O (DCPD) by x-ray diffraction, scanning electron microscopy, energy dispersive x-ray, chemical analysis, and equilibrium solution data. The rate of precipitation was compared using H+, Ca2+, and H2PO-4 data. The rate of H+ release was shown to be the most reliable indicator of the rate of precipitation under the chosen experimental conditions. The proton release data were used to calculate the empirical order of reaction (n) and the rate constant (log K). Under conditions of pH/Ca-stat, n was found to be 1.73 ± 0.17 and log K was found to be 2.60 ± 0.60. Under conditions of pH-stat, n was found to be 1.27 ± 0.12 and log K was found to be 0.69 ± 0.44. No relationship was found between the amount of seed material used and the kinetic parameters under the chosen experimental conditions.

Ethanol-induced formation of solid calcium phosphates.

Conventional transmission electron microscopy requires an effective dehydration of the tissue specimens under investigation. This dehydration, for which ethanol most often is used, may influence the solubility of the slightly soluble calcium phosphates. To study the behavior of calcium phosphates in ethanol, stable neutral aqueous solutions containing the ions were mixed with ethanol in various ratios at 20 degrees C. After 24 hours, the precipitates formed wee identified by x-ray diffraction to be either apatite, brushite, or amorphous calcium phosphate. The results are discussed with reference to possible formation of calcium phosphate artifacts.

Solid-state phosphorus-31 nuclear magnetic resonance studies of synthetic solid phases of calcium phosphate: potential models of bone mineral.

Phosphorus-31 NMR spectra have been obtained from a variety of synthetic, solid calcium phosphate mineral phases by magic angle sample spinning. The samples include crystalline hydroxyapatite, two type B carbonatoapatites containing 3.2 and 14.5% CO3(2-), respectively, a hydroxyapatite in which approximately 12% of the phosphate groups are present as HPO4(2-), an amorphous calcium phosphate, monetite, brushite, and octacalcium phosphate. Spectra were observed by the standard Bloch decay and cross-polarization techniques, as well as by a dipolar suppression sequence, in order to distinguish between protonated and unprotonated phosphate moieties. The spectra of the synthetic calcium phosphates provide basic information that is essential for interpreting similar spectra obtained from bone and other calcified tissues.

Recent studies of bone mineral: Is the amorphous calcium phosphate theory valid?

Recent studies on bone mineral are reviewed, with particular emphasis on the nature of the initial mineral deposited and the changes occuring during further mineralization and maturation. It is shown that amorphous calcium phosphate, which has been proposed as the precursor of hydroxyapatite in bone mineral, cannot be detected in significant quantity in the earliest bone mineral formed, and furthermore that there is evidence that no progressive decrease in amount of any amorphous phase occurs with bone maturation. We conclude in general that the X-ray difraction pattern and non-stoichiometric composition of bone mineral, and the changes in these characteristic observed with age and maturation cannot be explained by the presence of a second phase in progressively decreasing proportion whether it be amorphous calcium phosphate, octacalcium phosphate, brushite or some other distinct phase. The structural and compositional changes observed with age and maturation in bone mineral are consistent with a general increase in the degree of crystal perfection of a calcium phosphate phase best described as a poorly crystalline hydroxyapatite, although the exact details of the stuctural and compositional modification taking place with maturation must await further research. The role of brushite, which has been proposed as the earliest solid calcium phosphate phase deposited in at least one species, is reviewed.

Empirical Border Lines of the Apatite Stability Field in the System CaO—P2O5—H2O —Na2O—CO2—MX at Room Temperature

Abstract Literature data on equilibria between solid calcium phosphates having the apatite structure and aqueous solutions containing Na+ and CO3 2- ions were reexamined in view of the present data about the complexation of ions in such systems. Recalculations showed that all solutions were undersaturated or just saturated with respect to either brushite or octocalciumphosphate or more or less finely dispersed calcite and supersaturated with respect to pure hydroxylapatite. Sections through the quasi-sextary phase diagram were constructed which are representative for the system under simulated in vivo conditions.

Stabilization of amorphous calcium phosphate by Mg and ATP.

A synergistic effect has been demonstrated when magnesium and adenosine triphosphate (ATP) are used together in solution to delay the conversion of a slurry of amorphous calcium phosphate (ACP) to crystalline hydroxyapatite (HA). Conversion is delayed in some instances more than 10 times as long as with either ATP or Mg alone. In all experiments conversion did not begin until ATP in solution had decreased through hydrolysis to an undetectable level. The effect of Mg is to decrease substantially the rate at which ATP hydrolysis occurs. Once conversion began it proceeded more slowly in the presence of both Mg and ATP than with Mg or ATP alone. ATP was also found to prevent the formation of HA from metastable solutions of calcium and phosphate which did not contain any solid phase. Over the time period of these experiments, ATP hydrolyzed to a negligible extent in Tris-HCl buffer and in solutions containing Ca, PO4, and Ca plus PO4 ions. Hydrolysis of ATP does occur in the presence of ACP or HA, presumably by transphosphorylation on the surface of the solid calcium phosphate phase. It was concluded that ATP stabilized ACP, not by affecting its dissolution, but either by poisoning heteronuclear growth sites, or by poisoning the growth of embryonic HA nuclei (formed heterogeneously or homogeneously) before their critical size is reached, or by poisoning both. In the case of embryonic HA nuclei, the poisoned nuclei would go back into solution preventing HA crystal formation. In addition, it was found that the neutral Ca9(PO4)6 clusters, which are believed to be the basic structural unit of ACP, break down into individual Ca and PO4 ions when ACP dissolves in aqueous medium.

Global atmospheric monitoring

This discussion informally introduces several of the specific problems involved in the formation of solid phase calcium phosphates, their stability and their dissolution. Briefly discussed are aspects of the crystalline structure of bone, initiation mechanisms, non-growth and systemic regulation mechanisms, and dissolution and removal mechanisms. (JWP)