Formation Of Amorphous Calcium Phosphate Research Articles

Inhibiting Pathological Calcium Phosphate Mineralization: Implications for Disease Progression.

Pathological calcifications, especially calcium phosphate microcalcifications (MCs), appear in most early breast cancer lesions, and their formation correlates with more aggressive tumors and a poorer prognosis. Hydroxyapatite (HA) is a key MC component that crystallizes in the tumor microenvironment. It is often associated with malignant breast cancer lesions and can trigger tumorigenesis in vitro. Here, we investigate the impact of additives on HA crystallization and inhibition, and how precancerous breast cells respond to minerals that are deposited in the presence of these additives. We show that nonstoichiometric HA spontaneously crystallizes in a solution simulating the tumor microenvironmental fluids and exhibits lump-like morphology similar to breast cancer MCs. In this system, the effectiveness of poly(aspartic acid) and poly(acrylic acid) (PAA) to inhibit HA is examined as a potential route to improve cancer prognosis. In the presence of additives, the formation of HA lumps is associated with the promotion or only minimal inhibition of mineralization, whereas the formation of amorphous calcium phosphate (ACP) lumps is followed by inhibition of mineralization. PAA emerges as a robust HA inhibitor by forming spherical ACP particles. When precancerous breast cells are exposed to various HA and ACP minerals, the most influential factors on cell proliferation are the mineral phase and whether the mineral is in the form of discrete particles or particle aggregates. The tumorigenic effects on cells, ranging from cytotoxicity and suppression of proliferation to triggering of proliferation, can be summarized as HA particles < HA aggregates < ACP particles < ACP aggregates. The cellular response to minerals can be attributed to a combination of factors, including mineral phase, crystallinity, morphology, surface texture, aggregation state, and surface potential. These findings have implications for understanding mineral-cell interactions within the tumor microenvironment and suggest that, in some cases, the byproducts of HA inhibition can contribute to disease progression more than HA itself.

Direct observations of nanoscale brushite dissolution by the concentration-dependent adsorption of phosphate or phytate

With the development of agricultural intensification, phosphorus (P) accumulation in croplands and sediments has resulted in the increasingly widespread interaction between inorganic and organic P species, which has been, previously, underestimated or even ignored. We quantified the nanoscale dissolution kinetics of sparingly soluble brushite (CaHPO4·2H2O, DCPD) over a broad range of phosphate and/or phytate concentrations by using in situ atomic force microscopy (AFM). Compared to water, we found that low concentrations of phosphate (1–1000 µM) or phytate (1–100 µM) inhibited brushite dissolution by slowing single step retraction. However, with increasing phosphate or phytate concentrations to 10 mM, there was a reverse effect of dissolution promotion at brushite-water interfaces. In situ observations of the coupled dissolution-reprecipitation showed that phosphate precipitated more readily than phytate on brushite surfaces, with the formation of amorphous calcium phosphate (ACP). For a fundamental understanding, zeta potential and in situ Raman spectroscopy (RS) revealed that the concentration-dependent dissolution is attributed to the reverse of outer-sphere to inner-sphere adsorption with increasing phosphate or phytate concentrations. In addition, the mineralization of phytate with outer-sphere adsorption by phytase was higher than that with inner-spere adsorption, and the presence of phytate delayed ACP phase transformation to hydroxylapatite (HAP). These in situ observations and analyses may fill the knowledge gaps of interaction between inorganic and organic P species in P-rich terrestrial and aquatic environments, thereby implicating their biogeochemical cycling and the associated availability.

Polycarboxy/Sulfo Betaine-Calcium Phosphate Hybrid Materials with a Remineralization Potential.

Biomacromolecules control mineral formation during the biomineralization process, but the effects of the organic components' functionality on the type of mineral phase is still unclear. The biomimetic precipitation of calcium phosphates in a physiological medium containing either polycarboxybetaine (PCB) or polysulfobetaine (PSB) was investigated in this study. Amorphous calcium phosphate (ACP) or a mixture of octacalcium phosphate (OCP) and dicalcium phosphate dihydrate (DCPD) in different ratios were identified depending on the sequence of initial solution mixing and on the type of the negative functional group of the polymer used. The more acidic character of the sulfo group in PSB than the carboxy one in PCB determines the dominance of the acidic solid phases, namely, an acidic amorphous phase or DCPD. In the presence of PCB, the formation of ACP with acicular particles arranged in bundles with the same orientation was observed. A preliminary study on the remineralization potential of the hybrid material with the participation of PSB and a mixture of OCP and DCPD did not show an increase in enamel density, contrary to the materials based on PCB and ACP. Moreover, the latter showed the creation of a newly formed crystal layer similar to that of the underlying enamel. This defines PCB/ACP as a promising material for enamel remineralization.

Understanding the Nanoscale Affinity between Dissolved Organic Matter and Noncrystalline Mineral with the Implication for Water Treatment.

In recent decades, the concentration of dissolved organic matter (DOM) in aquatic ecosystems has gradually increased, leading to water pollution problems. Understanding the interfacial chemical processes of DOM on natural minerals is important to the exploration of high-efficiency absorbents. However, studying DOM chemical processes and adsorption mechanisms are still challenging due to the complex DOM structure and environmental system. Hence, we characterized the microstructure changes after the formation of amorphous calcium phosphate (ACP) at the interface of montmorillonite (Mt) minerals in a simulated environment system. Combined with atomic force microscopy and density functional theory (DFT) simulation, the mechanism of interfacial interaction between Mt-ACP and DOM was characterized at the molecular level. Moreover, we further evaluated the adsorption behavior of Mt-ACP as a potential adsorbent for organic matter. The comprehensive investigation of humic acid adsorption, intermolecular force, and DFT simulation is conducive to our understanding of the interfacial interaction mechanism between organic matter and noncrystalline minerals in aquatic environments and provides new perspectives on the application of clay-based mineral materials in pollutant removal under exposure from DOM.

Bioactive membranes of tricalcium phosphate and hydroxyapatite containing manganese for bone regeneration

In this study, we produced bioactive ceramic membranes of tricalcium phosphate (TCP) and hydroxyapatite (HA) incorporated with manganese (Mn) with osteogenic and antibacterial properties for applications in bone regeneration. Membranes were prepared by tape casting technique using PVA as a binder between molecules. The incorporation of manganese favors the reticulation of the membranes, providing more excellent resistance in aqueous media. Furthermore, the HA-Mn membrane exhibits the lowest optical density values and, therefore, the most significant difference from the negative control, presenting the lowest cellular viability. Between the membranes without manganese, HA shows the highest efficiency in improving cellular viability. Through bioactivity analysis, we observed that the TCP-Mn membrane presents the most significant formation of amorphous calcium phosphate precipitates, making the material more bioactive. Thus, our results become fascinating for applications in biomaterials.

Two-dimensional solid-state NMR spectroscopy investigations of surface precipitation of phosphate onto calcite

The interaction of phosphate with typical soil minerals is important for understanding P cycling in natural and agricultural systems. We investigated the mechanisms of kinetics of phosphate uptake onto calcite using solid-state NMR spectroscopy. At a low phosphate concentration of 0.5 mM, the 31P single-pulse solid-state NMR peak revealed the formation of amorphous calcium phosphate (ACP) within the initial 30 min, which transformed to carbonated hydroxyapatite (CHAP) after 12 d. At a high phosphate concentration (5 mM), the results showed transformation from ACP to OCP, later to brushite, and eventually to CHAP. The formation of brushite is further supported by 31P{1H} heteronuclear correlation (HETCOR) spectra via a correlation of δP–31 = 1.7 ppm and the 1H peak at δH–1 = 6.4 ppm, which denotes the structure water of brushite. Furthermore, 13C NMR directly revealed both A-type and B-type CHAP. Generally, this work provides a detailed understanding of the aging effect on the phase transition scale of phosphate surface precipitation onto calcite in soil environments.

Hydrogen phosphates play a critical structural role in amorphous calcium phosphates.

Amorphous calcium phosphate (ACP) is an intriguing mineral phase of calcium phosphate in its own right, in addition to its relevance in biomineralization. We hereby demonstrate that ACPs prepared by different synthetic routes such as the crosslinking of inorganic oligomers and polymer-induced liquid precursors have distinctive relative compositions of orthophosphate and hydrogen phosphate, and the extent of their hydrogen bonding with water. For all the ACPs or ACP-derived materials studied in this work, the species of hydrogen phosphate is the most important structural element. Depending on the synthetic pathways, orthophosphate and water, as well as their associated hydrogen bonds, may also play a role in the structural formation of ACPs.

Charge balancing stabilized apatite enter into collagen fibers by osmotic pressure to induce the formation of intrafibrillar mineralization

Intrafibrillar mineralized collagen is the main form of natural mineralized collagen in bone matrix. However, the mechanism involved in the intrafibrillar mineralization is still unclear. Here, by measuring the changes of Zeta potential and adjusting the osmotic pressure of collagen fibers to study the mechanism of intrafibrillar mineralization. The changes in zeta potential and reduction in size during the formation of amorphous calcium phosphate (ACP) after adding negatively charged polyacrylic acid (PAA) and positively charged cationicpolyacrylamide (CPAM) indicate that the stabilization mechanism of ACP induced by polyelectrolyte compound is charge balance. Then, osmotic pressure pressures charge balancing stabilized ACP into collagen fibers to induce intrafibrillar mineralization. This discovery provides basis for further understanding the intrafibrillar mineralization, preparing new biomimetic mineralized materials and promoting the development of tissue engineering.

Evaluating the induction of antibacterial properties upon zinc incorporation in strontium phosphosilicate for bone regeneration applications

Synthesis of Pure phase and a series of Zinc substituted Strontium phosphosilicate was attempted in the present study and assessed structurally and biologically. The Sol-Gel technique ensured proper reactivity and apatite formation upon sintering. The acidic medium was more applicable for synthesis on account of the isoelectric point of silica and formation of secondary Sr2SiO4 phase, important and applicable in bioactivity apart from Sr2P2O7 as another secondary phase. Sintering formed agglomerated stable crystalline phases evident from diffraction further confirmed by infrared spectroscopy. The morphology showed minimal differences from the parent material increasing agglomeration upon the increment of dopant. Biological evaluations showed the successful formation of amorphous Calcium phosphate with two days of SBF immersion and proliferation of MG-63 Cells upon Zinc incorporation. The antibacterial properties were enhanced upon the incorporation of Zinc while Strontium phosphosilicate in the pure phase failed to produce favourable antibacterial activity.

Hydroxyapatite Growth on Amelogenin‐Amelotin Recombinamers

AbstractStrategies to improve hydroxyapatite mineralization for enamel repair are essential for tissue regeneration. Amelogenin and amelotin (AMTN) are enamel matrix proteins playing critical roles in enamel formation. Amelogenin acts as a scaffold for hydroxyapatite, while AMTN (specifically its ‘SSEEL’ domain) is necessary for proper enamel mineralization. The functional relationship between recombinant AMTN and amelogenin, and their combined ability to guide uniaxial hydroxyapatite growth in vitro has been investigated recently. However, incorporation of the active domain of AMTN within recombinant amelogenins has not been studied yet. Here we describe the synthesis of modified amelogenin by inserting, internally and at its C‐terminus, the mineralizing AMTN‐derived motif SSEEL. C‐terminus modified amelogenin promoted hydroxyapatite formation, whereas internal incorporation of the motif initially resulted in amorphous calcium phosphate formation. Here we show that modified amelogenin with the mineralizing AMTN‐motif SSEEL can promote hydroxyapatite growth. These results give new insights for mineralized tissue regeneration using recombinamer proteins.

Structure and formation of amorphous calcium phosphate and its role as surface layer of nanocrystalline apatite: Implications for bone mineralization

We provide a critical review of the chemical composition and structure of synthetic and biogenic (bone/dentin mineral) nanocrystalline hydroxy-carbonate apatite (HCA). Such particles exhibit a “core–shell” organization, where an ordered HCA core is coated by a surface layer, whose nature is best captured by amorphous calcium phosphate (ACP), which is known to be a precursor phase of synthetic HCA, but whose role of bone/dentin mineralization has remained a most controversial subject. After reviewing the structure of each HCA and ACP component, as well as the most recent findings on their in vitro formation mechanisms, we examine the core–shell HCA organization further, with a focus on the disordered surface (“shell”) domain. In most of recent literature, the surface portion is often referred to as the “hydrated surface layer”, but without identifying its shared chemical and structural features of (synthetic) ACP. Unfortunately, that missing surface-layer/ACP equivalence obscures that the surface layer at the synthetic/biogenic nanocrystallites may simply constitute a remnant of the ACP phase from which the ordered HCA “core” nucleated. Although many topics reviewed herein have been investigated for more than six decades, several remain unsettled and heavily debated. Notably, decades-old articles offer suggestions that have passed unnoticed by the younger generations of researchers; we contrast and discuss both the latest and early contributions of this field, as well as highlighting several unsettled topics that should be revisited to improve our understanding of the ACP and HCA structures and in vitro/in vivo formation mechanisms.

Preparation of high-performance silico-aluminophosphate geopolymers using fly ash and metakaolin as raw materials

Silico-aluminophosphate (SAP) geopolymers were synthesized by using metakaolin (MK) and a high content of fly ash (FA) for the first time. With the increase in the FA mass fraction in the raw materials from 50% to 100%, the setting time of the SAP geopolymers was shortened and their compressive strength was decreased. The effects of the FA content on the SAP geopolymers composition and microstructure were investigated. The results showed that a high FA proportion caused the lack of active aluminum, which led to excess phosphate acid reacting with the calcium from FA. The fast formation of amorphous calcium phosphate (ACP) shortened the initial setting time through the consumption of water. However, after long-term aging, ACP transformed into needle- and slice-like nanoparticles, which decreased the compressive strength of the resulting geopolymers. Moreover, MK supplied active aluminum to increase the water-containing -Si-O-Al-O-P- network (SAP gel). The SAP gel contributed to the compressive strength enhancement and prevented the ACP transformation; however, it prolonged the final setting time, due to the slow polycondensation of SAP gel. These results indicate that a desirable setting time and compressive strength of the SAP geopolymers can be achieved by adjusting the FA and MK contents.

Effectiveness of antiscalants in preventing calcium phosphate scaling in reverse osmosis applications

Antiscalants are well known to prevent the precipitation of carbonate and sulphate scales of calcium in reverse osmosis (RO) applications, but according to literature their inhibitory ability against calcium phosphate is not clear. The objective of this study was to investigate if antiscalants, without acid addition, can prevent calcium phosphate scaling in RO systems. Eight calcium phosphate antiscalants from different manufacturers spanning a range of concentrations were tested in batch (in glass reactors) experiments to inhibit the formation of calcium phosphate in synthetic concentrate corresponding to 85% recovery (Ca2+ = 765 mg/L, PO43− = 13–15 mg/L and pH = 7.6) of a groundwater RO in the Netherlands. Additionally, once-through lab-scale RO tests were conducted where an RO element was fed with synthetic concentrate and the performance of antiscalants was evaluated from the rate of flux-decline in the RO element. Without antiscalant addition, a substantial flux-decline was observed due to the deposition of amorphous calcium phosphate (ACP) on the RO membrane. The tested antiscalants were unable to inhibit the formation of ACP and were incapable of preventing the deposition of the formed ACP particles, since with each antiscalant, the flux of the RO element decreased at least 15% in a 3-h period. Briefly, the available antiscalants, tested in this study, did not provide acceptable inhibition of calcium phosphate scaling in RO applications.

Defluoridation by fluorapatite crystallization in a fluidized bed reactor under alkaline groundwater condition

Fluoride removal using fluidized bed crystallization (FBC) technology with phosphate rock (PR) and calcite as seed particles under an alkaline groundwater condition was investigated. The results indicated that the optimized operating parameters were: upflow velocity = 7.21 m h−1, Ca:P:F molar ratio = 10:4:1, and initial bed height = 120 cm. The turbidity of FBC reactor effluent was 1.5–5.9 NTU at the first phase, and then reduced to less than 1.0 NTU at the second phase, while the effluent fluoride level remained at a constant level (0.4–0.9 mg L−1) during the entire procedure. The exfoliation and dissociation of newly-generated nuclei from the seed particles surfaces after aging and drying for three days could serve as an additional filtration layer (located on the top of the expanded bed with a thickness of 1.0 cm) to intercept the fine particles in the second phase. Consequently, a rational strategy for fluidized bed operation was provided, which focused on turbidity control and fluoride removal. FBC performance can be divided into ripening and running periods according to the evolution of turbidity in effluent. Fluoride removal was achieved by the formation of amorphous calcium phosphate and then recrystallization into fluorapatite and/or FCO3-apatite and induced crystallization of fluorapatite and/or FCO3-apatite. The FBC process with additional filtration function of dissociated newly-generated nuclei could be regarded as a cleaner technology for fluoride removal; additionally, the spent seed particles could be utilized as resources in the phosphatic fertilizer industry. However, the fertilizer industry should continue to recover fluoride for further economic benefits.

Remineralization of enamel caries by an amelogenin-derived peptide and fluoride in vitro.

Dental caries is one of the most common oral diseases in the world. This study was tantamount to investigate the combinatory effects of an amelogenin-derived peptide (called QP5) and fluoride on the remineralization of artificial enamel caries. The peptide QP5 was synthesized and characterized, and the binding capability of the peptide on hydroxyapatite (HA) and demineralized tooth enamel surface was analysed. Then, the mineralization function of the peptide and fluoride was studied through the spontaneous mineralization testing and remineralization on enamel caries in vitro. First, the novel peptide QP5 could bind on the hydroxyapatite and demineralized tooth enamel surfaces. Second, QP5 can transitorily stabilize the formation of amorphous calcium phosphate and direct the transformation into hydroxyapatite crystals alone and in combination with fluoride. In addition, compared to blocks treated by peptide QP5 alone or fluoride, the sample blocks showed significantly higher surface microhardness, lower mineral loss and shallower lesion depth after treatment with a combination of QP5 and fluoride at high or low concentrations. The peptide QP5 could control the crystallization of hydroxyapatite, and combinatory application of peptide QP5 and fluoride had a potential synergistic effect on the remineralization of enamel caries.

Preparation of Laser-Modified Ti-15Mo Surfaces With Multiphase Calcium Phosphate Coatings

Multiphasic bioceramic scaffolds has been enhanced for dental and orthopedic applications. In this perspective, the laser surface texturing of metallic surfaces combined to bioactive calcium phosphate coatings have shown to be promising and economically feasible for biomaterial clinical applications. Ti-15Mo alloy samples were irradiated by pulsed Yb: YAG laser beam. The formation of HA and other calcium phosphates phases by biomimetic method should occur in the presence of Ca2+, PO43-, Mg2+, HCO3-, K+ and Na+. The modified surfaces were submitted to thermal treatment at 380 and 580°C. The results showed the processes of fusion and fast solidification from the laser beam irradiation, inducing the formation of stoichiometric α-Ti, TiO2 and non-stoichiometric titanium oxides, Ti3O and Ti6O with different oxide percentages depending on applied fluency (fluency of 0.023, 0.033, 0.040 and 0.048 J/mm2). The morphological and physicochemical properties have indicated the formation of a multiphase bioceramic coatings. It was observed the formation of amorphous calcium phosphate (ACP), octacalcium phosphate (OCP), and magnesium phosphate (Mg3(PO4)2) phases at 380°C, whereas β-TCP (tricalcium phosphate), OCP, and substituted β-TCP with Ca2,589Mg0,41(PO4)2 were obtained at 580°C. Therefore, the multiphasic bioceramic modified Ti-15Mo surface could enhance osteointegration for bone regeneration.

Unexpected Favorable Role of Ca2+ in Phosphate Removal by Using Nanosized Ferric Oxides Confined in Porous Polystyrene Beads

Polystyrene-based nanoferric oxide composite is a representative nanomaterial successfully applied in scale-up water decontamination for arsenic and phosphorus. However, little is available on the effect of solution chemistry (for instance, the coexisting Ca2+) on the long-term performance of the nanocomposite. In this study, we carried out 20 cyclic runs of phosphate adsorption-desorption on a polymer-supported ferric nanocomposite HFO@201. Unexpectedly, an enhanced phosphate removal was observed in the presence of Ca2+, which is quite different from its adverse effect on phosphate capture by granular ferric oxide. Further mechanistic studies revealed that enhanced phosphate removal was mainly realized via the Ca-P coprecipitation inside the networking pores of HFO@201 as well as the possible formation of the multiple Fe-P-Ca-P complex. The complex formation led to a distinct increase in P adsorption, and the coprecipitation, driven by the accumulated OH- in confined pores during phosphate adsorption and alkaline regeneration, favored P removal via the formation of amorphous calcium phosphate (ACP) and hydroxyapatite inside. TEM-EDS spectra indicated that coprecipitation did not occur on the surface of loaded nano-HFO, greatly mitigating its adverse effect on P adsorption on the surface of nano-HFO. Fixed-bed column study showed that the presence of Ca2+ increased the effective treatable volume of HFO@201 toward P-containing influents by ∼70%. This study is believed to shed new insights into the effect of solution chemistry on similar nanocomposites for advanced water treatment.

Ultrafast bone-like apatite formation on bioactive tricalcium silicate cement using mussel-inspired polydopamine

The growing bone-like apatite layer at the tissue-implant interface is the essential condition for materials to bond robustly to surrounding bone and may provide a favorable environment for living bone formation. Inspired by versatility of mussel adhesive proteins, we developed an ultrafast and accessible route to accelerate effectively the formation of amorphous calcium phosphate (ACP), the precursor phase of bone-like apatite, on the surface of polydopamine (PDA)-coated tricalcium silicate (TCS) within 5 min in simulated body fluid (SBF). The key of the method lies in successful preparation of PDA coating on the surface of hydrated TCS by simple dip-coating of hydrated TCS in an aqueous solution of dopamine. A strong adsorption between PDA coating and surface of hydrated TCS could be formed via bidentate hydrogen and electrostatic bonds. After 7 d of soaking in SBF, the bone-like apatite layer on the surface of PDA-coated TCS disk, about 91.1 µm in height was thicker than that on the surface of pristine TCS disk, determining about 49.5 µm. The results indicate that PDA coating can facilitate the kinetic deposition of bone-like apatite on its surface. The abundant Ca2+ ions and the lower interface energy of ACP at the interface between ACP and surfaces of PDA-coated TCS disks are responsible for the ultrafast precipitation of ACP and formation of bone-like apatite layer which is carbonated hydroxyapatite (HA) confirmed by different analytical tools. The route can open avenues for development of PDA-coated TCS biomaterials for hard tissue repair and substitution.

Contrasting In Vitro Apatite Growth from Bioactive Glass Surfaces with that of Spontaneous Precipitation.

Body-fluid-exposed bioactive glasses (BGs) integrate with living tissues due to the formation of a biomimetic surface layer of calcium hydroxy-carbonate apatite (HCA) with a close composition to bone mineral. Vast efforts have been spent to understand the mechanisms underlying in vitro apatite mineralization, as either formed by direct precipitation from supersaturated solutions, or from BG substrates in a simulated body fluid (SBF). Formally, these two scenarios are distinct and have hitherto been discussed as such. Herein, we contrast them and identify several shared features. We monitored the formation of amorphous calcium phosphate (ACP) and its crystallization into HCA from a NaO–CaO–SiO–PO glass exposed to SBF for variable periods out to 28 days. The HCA growth was assessed semi-quantitatively by Fourier transform infrared spectroscopy and powder X-ray diffraction, with the evolution of the relative apatite content for increasing SBF-exposure periods evaluated against trends in Ca and P concentrations in the accompanying solutions. This revealed a sigmoidal apatite growth behavior, well-known to apply to spontaneously precipitated apatite. The results are discussed in relation to the prevailing mechanism proposed for in vitro HCA formation from silicate-based BGs, where we highlight largely simultaneous growth processes of ACP and HCA.

Remineralization Efficacy of an Amelogenin-Based Synthetic Peptide on Carious Lesions.

Objective: The aim of this study was to evaluate the remineralization efficacy of an amelogenin-based peptide on initial enamel carious lesions in vitro. Furthermore, we attempted to provide insights into the possible mechanism of the remineralization, including determining the calcium-binding properties of the peptide and its effects on calcium phosphate mineralization.Methods: The peptide comprising the N-terminus and the C-terminus of porcine amelogenin was synthesized by Synpeptide Co., Ltd. Fifty specimens were randomly assigned to five immersing treatment groups for 12 days: remineralizing medium only; 12.5 μg/mL peptide + remineralizing medium; 25 μg/mL peptide + remineralizing medium; 50 μg/mL peptide + remineralizing medium; fluoride + remineralizing medium. After immersion, mean mineral loss before and after remineralization of each specimen was determined using micro-CT. Mean mineral gain after remineralization was calculated. Calcium binding properties were measured by Isothermal titration calorimetry (ITC). TEM and Fourier transform-infrared were used to determine the effects of the peptide on calcium phosphate mineralization.Results: A significant decrease in mineral loss after remineralization process in all groups was observed (p < 0.05). Treatment in remineralizing medium resulted in the lowest mineral gain while the fluoridated treatment exhibited the highest mineral gain among all groups. Inclusion of synthetic peptide in the remineralizing medium exhibited a higher mineral gain and the gain of 50 μg/mL group was greater than that of the 25 μg/mL group. No significant difference in mineral gain was observed between the remineralizing medium only group and the 12.5 μg/mL peptide group (p > 0.05). ITC values showed that the Ca2+-binding affinity of the peptide is about 9.914 × 104M−1. Furthermore, the peptide was found to inhibit calcium phosphate precipitation and stabilize amorphous calcium phosphate formation for more than 2 h and finally transform into ordered hydroxyapatite crystals.Conclusion: Specific concentrations of the amelogenin-based synthetic peptide promoted in vitro remineralization, with higher concentrations exhibiting significantly greater remineralization. This study presented evidence suggesting that the peptide may act as a Ca2+carrier as well as a regulating factor. When the stabilizing calcium and phosphorus ions bind with the peptide they become biologically available for the remineralization of deeper carious lesions, while also regulated by the peptide to transform into ordered hydroxyapatite crystals.