Citrate Stabilizes Hydroxylapatite Precursors: Implications for Bone Mineralization

Abstract

Mineralization of hydroxylapatite (HAp), the main inorganic phase in bone, follows nonclassical crystallization routes involving metastable precursors and is strongly influenced by organic macromolecules. However, the effect of small organic molecules such as citrate on the formation of HAp is not well constrained. Using potentiometric titration experiments and titration calorimetry, in combination with a multianalytical approach, we show that citrate stabilizes prenucleation species as well as a liquid-like calcium phosphate precursor formed before any solid phase nucleates in the system. The stabilization of a liquid-like precursor phase could facilitate infiltration into the cavities of the collagen fibrils during bone mineralization, explaining the enhancement of collagen-mediated mineralization by citrate reported in previous studies. Hence, citrate can influence bone mineralization way before any solid phase (amorphous or crystalline) is formed. We also show that HAp formation after amorphous calcium phosphate (ACP) in the absence and presence of citrate results in nanoplates of about 5–12 nm thick, elongated along the c axis. Such nanoplates are made up of HAp nanocrystallites with a preferred c axis orientation and with interspersed ACP. The nanoplatelet morphology, size, and preferred crystallographic orientation, remarkably similar to those of bone HAp nanocrystals, appear to be an intrinsic feature of HAp formed from an amorphous precursor. Our results challenge current models for HAp mineralization in bone and the role of citrate, offering new clues to help answer the long-standing question as to why natural evolution favored HAp as the mineral phase in bone.

Mineralized Enzyme-Based Biomaterials with Superior Bioactivities for Bone Regeneration

“…This process is tightly influenced by various molecules such as ALP in this study. 4 The morphology and size of CaP nanomaterials can be regulated by molecules with carboxylic acid 28 and/or phosphorus-containing groups. 29,30 Our previous studies have also illustrated that the −COOH groups and the N-containing ring structures coordinate with Ca 2+ ; 31 ALP which consists of a series of amino acids tends to hinder the crystal formation or growth of CaP nanoparticles due to the interaction with Ca 2+ ions; thus, the size of MALPNs is smaller than that of control CaP nanoparticles.…”

Development of Biodegradable Osteopromotive Citrate‐Based Bone Putty

“…[56][57][58] Further, binding of citrate to specific surfaces of HA as it undergoes crystallization from early amorphous phases serves to regulate crystal size/shape to the preferred nanocrystal morphology, preventing bulk crystal fusion and brittle mechanics. [58][59][60][61] Inspired by the dual-phase bridging roles of in vivo soluble citrate, we theorized that polymers directly incorporating citrate could intrinsically recapitulate the strong organic/inorganic phase binding in a biomaterial platform.…”

Potential Load-Bearing Bone Substitution Material: Carbon-Fiber-Reinforced Magnesium-Doped Hydroxyapatite Composites with Excellent Mechanical Performance and Tailored Biological Properties

“…Hydroxyapatite [HA; Ca 10 (OH) 2 (PO 4 ) 6 ], which has been widely studied as a bone substitute in recent years, is similar to the main component of natural bone apatite. − HA has been used in various medical applications because of its mechanical stability, high activity, and good biocompatibility . However, the application of pure HAs is greatly limited to small or unloaded implants because of their low strength and fracture toughness .…”