Distribution Analysis of Nanoparticle Architectures
Hydroxyapatite Description and Uses
Hydroxyapatite (HA) is a bioactive calcium-based mineral with the chemical formula Ca10(PO4)6(OH)21,2,4. Along with protein-based polymers such as collagen, this mineral forms the basis of all hard tissues in the human body, with as much as 50% of bone and over 90% of enamel (T. Cate Oral Histology, 1998). In the case of bone, the HA particles are primarily poorly crystalline, rod shaped nanoparticles2. HA Nanoparticles have been shown to exhibit more bioactivity than macroscopic or micron-scale particles, with greater adhesion to osteoblasts and higher levels of bioresorbability1. HA-based biomaterials are currently used in the treatment of bone injuries resulting from injuries or trauma2,4. It is important, therefore, to produce biomaterials which mimic the natural properties of bone as closely as possible. Perhaps due to natural bone being a composite of HA with protein (natural polymers) research into bone grafts appears to focus of HA particles embedded into a polymeric matrix.
Most commercial applications currently used consist of macro-particles or blocks of material to be placed on or in the bone flaw, and only recently have injectable forms begun to find clinical use2. Block and particulate devices are harder to place than injectable bone graft materials2, increasing the risk of injury during surgery and likelihood of lamination, this makes injectable bone grafts an attractive proposition, as they are less invasive than traditional bone grafts and can more easily be made to fit the specific wound of the patient.
In the 1980s, injectable bone grafts were developed in which one or more Ca compounds were mixed with water or another liquid and this mixture was injected into the bone. Once injected the mixture underwent a self-setting reaction and transformed into monolithic HA2. Such Ca-phosphate cements are injectable and hence minimally invasive, but can suffer from serious biocompatibility issues2, often resulting in the need for further surgery.
Later approaches involved the use of HA powder mixed with a dispersant. These injectable bone grafts did not undergo a setting reaction however a high pH was required to maintain a stable suspension2.
HA materials delivered in the form of a paste are considered more convenient than those coming in the form of a powdered material which needs to be combined with a liquid immediately before surgery2.
Bioactivity is key in bone graft materials as full incorporation into the bone tissues is the ultimate goal of bone graft research. Bone is a naturally dynamic material which restructures itself in response to the applied stresses. Porosity and pore size is an important factor for proliferation and differentiation of bone cells, as well as exchange of both nutrients and metabolites1. The ideal bone graft material is therefore one which has appropriate porosity to allow these processes to take place. This would allow the host’s bone cells to migrate into the bone where they may begin to restructure the component.
Biomaterials properties of HA pastes
As noted previously, desirable properties for bone graft materials include bioactivity, injectability and porosity, and ideally the material would be as simple as possible for ease of production. A class of materials which seems to match all of these properties is hydroxyapatite paste, in particular that composed of just HA and water. Size, shape, aspect all determine the rheological behaviour, but have no impact on biocompatibility, suggesting that morphology has no role in osteoblast cytotoxicity2.
Nano HA is used as a drug-delivery agent for anti-tumour drugs and antibodies in treatment of osteomyelitis5.
Especially notable is the bioactivity of HA nanorod particles, which have been shown to be more bioactive than other types of HA.
(write about the rabbits)
Physical Properties of HA pastes
A number of papers exist describing the preparation of nanohydroxyapatite and properties. Rheology of a bone-graft material is crucial for surgical delivery2,3, and as such most studies have focused on compositions of HA with water which have additives to improve flowability. This reduces the force required to inject the material into the bone defect.
HA pastes in water are shear-thinning, non-Newtonian liquids2. HA pastes with a range of physical properties have been demonstrated, with seemingly small differences in nanoparticle shape and size appearing to cause significant changes in macroscopic properties such as the initial viscosity. Unusually, although a mixture of two pastes with well-defined viscosities would be expected to follow the rule of mixtures (predicting overall viscosity to be a weighted average of the viscosities of the two pastes), this does not appear to be the case. Instead, the pastes seem to obey a “majority rule”, wherein the properties of the mixture are much closer to the properties of the majority paste2.
Steric stabilisation is often necessary to achieve sufficiently low viscosity for injection and is achieved by addition of a dispersant which adsorbs to particle surfaces4.
Flow behaviour of aqueous suspensions of HA have been investigated, showing that rheological behaviour of the suspensions is strongly dependent on both dispersant concentration and phase volume4. Small changes in either condition can result in large changes in the viscosity of the system. For example, according to Gardini and Galassi, increasing the solids content of the suspension from 22% to 25% while keeping content of dispersants the same increased the initial viscosity by 3 orders of magnitude. The implication of this is that the viscosity of a HA suspension could be easily altered to suit the application.
At very low shear, the elementary solid particles group together in aggregates that can form a 3D network involving the whole volume of suspension. This network imparts elastic properties to the material and is responsible for an apparent yield strength.4 This apparent yield strength allows the material to behave as a solid at low stresses, meaning that the paste could be applied in non-load bearing applications without the need for stabilising plates.
At higher shear rates, the aggregates break down into smaller units which flow more easily, causing the rheological properties to change over time4. This also means that the elastic properties disappear at high shear and hence in load-bearing applications the paste could only be used as a complement to more dimensionally stable components.
Microstructural properties of HA pastes
Few detailed studies exist which combine rheological and morphological investigations to understand the structure-property relationships2.
Variation in particle architectures is well-documented, with larger, more crystalline particles being formed at higher temperatures2,3,5.
Smaller particles have a larger specific area, and hence experience larger van der waals forces, and hence exhibit higher initial viscosity2,4,. More elongated particles also have higher specific area2.
Both HA and H2O can be both H donors or acceptors, suggesting that H-bonding may play a part in the rheology of the system.2
Agglomeration phenomena are governed by forces involving Brownian, hydrodynamic and interparticular forces. Van der Waals forces between particles are the primary forces responsible for flocculation.4
At high particle concentration elastic effects appear4.
At high particle concentration and low dispersant concentration a transition in the viscoelastic properties occurs which resembles the sol-gel transition associated with chemical or physical cross-linking.4
The rebuilding of particle agglomerates depends on the initial state of agglomeration of the particles, so HA suspensions are very sensitive to shear history. This contributes to the time-dependent effects observed.4