Assembly of Layered Monetite-Chitosan Nanocomposite and Its Transition to Organized Hydroxyapatite

Abstract

Bioinspired synthesis of hierarchically structured calcium phosphate (CaP) material is a highly promising strategy for developing improved bone substitute materials. However, synthesis of CaP materials with outstanding mechanical properties still remains an ongoing challenge. Inspired by the formation of lamellar structure in nacre, we designed an organic matrix composed of chitosan and cis-butenediolic acid (maleic acid, MAc) that could assemble into a layered complex and further guide the mineralization of monetite crystals, resulting in the formation of organized and parallel arrays of monetite platelets with a brick-and-mortar structure. Using the layered monetite-chitosan composite as a precursor, we were able to synthesize hydroxyapatite (HAp) with multiscale hierarchically ordered structure via a topotactic phase transformation process. On the nanoscale, needlelike HAp crystallites assembled into organized bundles that aligned to form highly oriented plates on the microscale. On the large-scale level, these plates with different crystal orientations were stacked together to form a layered structure. The organized structures and composite feature yielded CaP materials with improved mechanical properties close to those of bone. Our study introduces a biomimetic approach that may be practical for the design of advanced, mechanically robust materials for biomedical applications.

Keywords: bioinspired matrix; chitosan; hierarchical structure; hydroxyapatite; maleic acid; monetite.

Figure 1

Schematic representation of the pathway followed to fabricate multilevel organized CaP-based composite.

Figure 2

SEM images of (a, b) pure chitosan matrix and (c, d) organized chitosan-MAc matrix: (a) top view, (b) side view. (e) XRD pattern of chitosan-MAc matrix. (f) Schematic representation of interaction between chitosan and MAc. The molecular model was established by ChemBio Ultra 11.0. The distance between the molecules was estimated using the typical bond lengths and angles. Blue in the chitosan chain and Red in the MAc molecule represent the charged amine and carboxyl groups, respectively.

Figure 3

(a, b) FTIR spectra of chitosan (blue), MAc (red), and chitosan-MAc matrix (gray) in the absence of calcium in (a) the C=O stretch and (b) the N–H bending regions. (c) CD spectra of chitosan and chitosan-MAc matrix at pH 3.5.

Figure 4

(a, b) AFM and (c, d) TEM images of (a, c) pure chitosan and (b, d) organized chitosan-MAc matrix.

Figure 5

(a–c) SEM images of the orderly layered monetite-chitosan-MAc composite. (d) TEM and (e) HRTEM images of a monetite nanosheet. Inset: FFT pattern corresponding to e. (f) XRD pattern and (g) TG curve of the layered monetite-chitosan-MAc composite.

Figure 6

(a) pH values of reaction solution without (dotted line) and with (solid line) Ca²⁺ at different reaction times. (b) pH difference (ΔpH) between the solutions with and without Ca²⁺ at different time points. (c, d) SEM images of the products collected at (c) 1 and (d) 2 h.

Figure 7

SEM images of the layered monetite-chitosan-MAc composites formed in chitosan-MAc matrix with different concentrations: (a) 0.5% (m/v) chitosan and 0.2% (m/v) maleic acid; and (b) 3% (m/v) chitosan and 1.2% (m/v) maleic acid.

Figure 8

FTIR spectrum of the chitosan-MAc-Ca²⁺ complex.

Figure 9

Organized HAp-chitosan-MAc composite formed from phase transformation of layered monetite-chitosan-MAc composite. (a–c) SEM images of HAp-chitosan-MAc composite. Inset image in b shows the SEM image of an organized bundle. Arrows in (c) indicate different layers (stacks of HAp plates) in the HAp-chitosan-MAc composite. (d) HRTEM images of an isolated crystal from the HAp-chitosan-MAc composite. Inset: FFT pattern corresponding to d.