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preface
Magnesium alloys are considered one of the most promising biodegradable materials due to their excellent mechanical strength and lightweight properties, especially for biodegradable implant manufacturing.
They promote bone healing and osseointegration, and maintain sufficient mechanical strength after implant placement. However, over time, magnesium alloys gradually degrade under the influence of physiological media.
In order to further improve the properties and biocompatibility of these materials, surface modification has become an important research direction. Various coating technologies have been extensively studied to improve the corrosion resistance, biocompatibility, and mechanical properties of magnesium alloys.
Our research aims to develop a polymer-based composite coating that introduces bioactive fillers into the surface of biodegradable magnesium alloys.
Research materials and methods
The surface morphology and elemental composition of the sample were examined using the QUANTA INSPECT F scanning electron microscope and the Energy Dispersive X-ray Spectrometer detector (EDAX).
Infrared spectroscopy is performed on the JASCO FTIR 6200 spectrometer using an ATR device with a spectral range of 600-4000 cm1.
Raman spectroscopy was performed using an alpha 75 RAS+ system equipped with a 300 nm laser (532 mW) in combination with a grating spectrometer (600 grooves/mm) and a microscope (50 × magnification).
For spectral acquisition, the integration time is 2 s per spectrum. To improve the signal-to-noise ratio, the spectra were averaged over 20 consecutive measurements.
Due to optical alignment and raster dispersion relative to the incident laser line, Raman spectral shifts of up to 8 cm−1 in the spectral range between 200 and 3800 cm−1.
Contact angle measurements are performed using the KRÜSS DSA30 droplet shape analysis system to determine the wettability of the sample surface. Process the obtained image by aligning tangents to the contours of the sessile droplets at the contact point with the surface. Contact angle measurements are made in triplicate and the average value is calculated.
Thermogravimetric (TGA) curves are simultaneously obtained using the TA Instruments SDT Q600 system, which obtains thermogravimetric (TGA) curves in a nitrogen working atmosphere at a constant heating rate of 600°C per minute in a temperature range of 10-10°C.
Research findings and discussion
Cellulose acetate (CA) is an absorbable, non-toxic, environmentally friendly neutral polymer that forms a transparent film. It is highly biocompatible and is made from a natural polymer called cellulose.
Due to its hydrophilicity, it can be functionalized by different chemical groups for higher biocompatibility. Membranes made of CA exhibit important chemical stability, good mechanical properties, higher hydrophilicity, excellent protein transport capacity, low protein adsorption and water affinity.
Cellulose can be prepared from plants, algae, wood and bacteria, so the annual production is estimated to be around 7×1010 tons/year. Regarding the mechanical properties of CA, we can name high modulus of elasticity, bending and tensile strength.
The polymer can be used in biomedical applications such as wound healing patches, drug delivery systems, and separation membranes. Hydroxyl groups activated in CA can be modified or replaced by other functional groups by different methods such as hydrolysis, grafting, oxidative etherification, copolymerization, crosslinking and esterification.
Today, CA polymers of different molecular weights are used between 30,000 g mol−1 and 60,000 g mol−1.
To reduce the corrosion rate of magnesium alloys, CA-based coatings are used. A coating system for laser structural surfaces consisting of a primer layer and a polymer coating was developed to improve the degradation behavior of magnesium alloys.
They used CA as a primer and deposited layers of chitosan and carboxymethylcellulose. A very low corrosion rate of 1.15 cm/year was obtained, and the implant was hydrophilic.
Mg-Ca-Mn-Zr alloy is coated with cellulose acetate using the impregnation method. The formation of polymer coatings was verified based on surface characterization methods and scanning electron microscopy.
The potentiodynamic polarization test proves that the CA coating significantly improves the corrosion properties of the alloy under study. The authors also conducted in vitro and in vivo studies showing increased bone regeneration and good cytocompatibility in MC3T3-E1 osteoblasts.
Although CA coatings have high potential for orthopedic applications, there is little research in the literature on their application to magnesium-based alloys.
Thermogravimetric analysis (TGA) is used to study the effect of filler particles on the thermal stability of polymers.
All samples showed similar characteristics with two degradation stages. The first stage of degradation is due to the loss of water in the polymer material, and the second stage is due to the loss of water in the material itself.
The cellulose acetate coating has the lowest thermal stability, and the thermal resistance increases with the addition of filler particles. Although the curves behave similarly, there are significant differences in mass loss; The lowest values were detected in the case of CA-Mg samples (composite samples containing magnesium particles).
This fact can be explained by the cross-linking effects that occur between magnesium atoms and non-participating electrons of oxygen atoms in polymer components, which are much stronger than the vander wall forces that occur between polymers and hydroxyapatite particles.
It has previously been demonstrated that the electrostatic interaction of cellulose acetate with magnesium is strong and stable enough to increase the thermal resistance of the composite, and the atoms implicit in these interactions are oxygen atoms from acetyl groups with complexing ability.
As a result, a crosslinking effect that stabilizes the entire matrix occurs. In addition, HAp particles form agglomerates, reducing the amount of interaction between filler and polymer.
Samples containing two types of particles (hydroxyapatite and magnesium) CA-HAp-Mg have intermediate behavior, and thermal stability is given by competition between the two interaction formation mechanisms.
Fourier transform infrared (FTIR) spectroscopy is used to identify specific functional groups or chemical bonds found in experimental samples, highlighting subtle differences between cellulose acetate sample spectra and composite coating sample spectra.
The spectra showed that cellulose acetate had a characteristic band −1 (mC=O) assigned to the stretching of acetyl carbonyl esters (C=O) at ~1745 cm.
−1 (m C-O) at ~1230 cm is attributed to the C-O stretching pattern of the acetyl group, and −1 and ~1430 cm −1 (dC-H) bands at ~1370 cm −1 (dC-H) due to the bending vibration of C-H in -CH.
At 1040 cm−1, a band assigned to the C-O tensile vibration of the pyranose ring appears. The strap −1 at ~1163 cm is assigned to the COC antisymmetric bridge stretch.
CA and CA-Mg samples have similar morphology. This indicates that Mg is evenly distributed throughout the coating and is directly related to the aspects highlighted by scanning electron microscopy.
Conversely, the addition of HAp particles results in the separation of domains, showing a high degree of agglomeration. This fact may be related to HAp crystallization and separation from the solution. CA-HAp-Mg samples show some degree of segregation, although the chemical composition may differ due to the presence of Mg.
Raman spectra of hydroxyapatite (HAp) show the main features associated with PO 43− and Ohio− ions. Purchase order 43− is characterized by a symmetrical stretch (P–O) mode −1 at 973 cm.
Bending (O–P–O) mode −1 at 430–450 cm−1, antisymmetric stretching (P–O) mode −1 at 1020–1080 cm, and bending (O–P–O) mode −1 at 585–610 cm. The presence of OH − results in a tensile pattern −1 at 3600 cm and a vibrating band at 630 cm−1.
Pan mode at 340 cm−1. OH- contributed weakly to HAp Raman spectroscopy. The symmetrical stretching of the (P-O) mode moves to a higher value (i.e., from 960 cm−1 to 973 cm−1) compared to other HAp.
The Raman spectra of CA correlated with the vibrating band at 2945 cm−1 and 1129 cm−1, which are attributed to C-H stretching and asymmetric tensile vibrations of C-O-C glycosidic bonds, respectively.
The contribution of the pyranose ring −1 was observed at 1081 cm. The vibration band −1 associated with the C-OH bond was observed at 1272 cm.
The characteristic Raman signal of the acetyl group can be observed at −1 at 1744, 1443, and 1390 cm, corresponding to the vibration of the carbonyl group (C = O) and the asymmetric and symmetric vibration of the C-H bond present in the acetyl group. The vibration band −1 at 986, 914, 842, and 667 cm is associated with C-O, C-H, O-H, and C-OH bonds, respectively.
Therefore, adding HAp and Mg to CA causes subtle changes in the corresponding vibration bands. Adding HAp to CA causes weakening of C-H bonds and some vibrations associated with the pyranose ring.
In addition, the symmetrical stretching (P-O) pattern of PO43− in HAp was exhibited at 973 cm. −1. These characteristics indicate that HAp is crystallized and separated CA. Similar results were found in samples containing Mg and CA, except that the contribution of C-H bonds to vibrations is enhanced, while some of the vibrations associated with the pyranose ring are suppressed.
However, the addition of Mg and HAp to the CA coating results in a dramatic change in Raman spectroscopy, with the overall characteristics exhibited by CA being suppressed above 1100 cm−1, indicating a dramatic change in the backbone chemistry of the membrane.
The absence of O-H tensile vibration−1 of pure water in the range of 3600-3800 cm indicates membrane dehydration. The main Raman vibration bands of the CA-Mg-HAp membrane were shown at 127, 275 and 380 cm, respectively.
The Raman band −1 at 573 and 952 cm is weak and wide. This makes it difficult to quantify the chemistry of the specimen. According to literature data, Raman zones between 100 and 210 cm −1 are lattice (phonon) vibrations with strong intensity.
This may be related to the precipitation of nano-Mg(OH)2 or MgO phases with reduced crystal symmetry. However, the C-C fat chain has a strong vibration−1 in the range of 250-400 cm.
This explains that the weak vibration−1 at −1.573 cm at Raman peaks at 275 and 380 cm may be related to the C-(I, Cl, or Br) group (i.e., vibrations between 490 and 790 cm−1).
Broadband −1 at 952 cm may be associated with the C-O-C group (i.e., vibration −1 at 800–950 cm) and/or carboxylic acid dimer (i.e., vibration −1 at 910–960 cm).
An increase in swelling capacity was observed with the addition of hydroxyapatite and magnesium particles. This is partly due to the porosity of the composite sample allowing more water molecules to be captured. This leads to more intense degradation processes.
In addition, measuring the contact angle to determine the hydrophilic or hydrophobic properties of the sample surface highlights its hydrophilicity. With the addition of hydroxyapatite and magnesium particles to the polymer matrix, the hydrophilic properties increase, increasing the swelling capacity of the sample. The hydrophilic properties facilitate better interaction between the sample and the PBS solution, mainly water.
During the first 45 minutes, the swelling rate increased rapidly for all samples, but was more pronounced for samples containing magnesium particles (CA-Mg and CA-HAp-Mg).
The sample reached a steady state after 90 minutes, with CA-Mg sample expansion by about 48%, CA-HAp-Mg sample expansion by about 38%, and CA-HAp sample expansion by 32%.
Surface wetting quantified by contact angle measurements is an important factor in the functionality and biocompatibility of implantable devices. Thus, a small contact angle will improve cell adhesion, while a hydrophobic surface (contact angle > 90°) may affect cell adhesion, resulting in rejection of the implant material.
For samples containing magnesium particles (CA-HAp-Mg and CA-Mg), the lowest contact angle values were obtained, which demonstrated that by adding them, hydrophilic surfaces conducive to biointegration could be obtained.
It is well known that complex contact angle behavior can be observed when droplets come into contact with a mixture of different particles, such as in our case (HAp and Mg particles).
As an analytical tool, Cassie-Baxter theory can be used. They showed that in the case of mixtures formed by larger particles (i.e., Mg powder) and small particles (i.e., HAp powder), if the larger particle diameter is above the critical volume fraction, the surface wettability may change greatly because small particles appear partially overriding the larger particles.
For larger particle diameters less than or equal to the critical value, full surface coverage of larger particles occurs, and the mixture receives the surface characteristics of small particles.
In our study, in the case of CA-HAp-Mg, the contact angle is almost equal to the contact angle obtained by CA-HAp. We can conclude that since HAp small particles readily form agglomerates, HAp particles massively cover the surface of Mg particles, as demonstrated by the SEM study. This fact determines the increase in surface contact angle compared to CA-Mg samples.
conclusion
The main problem with magnesium-based biomaterials is their inhomogeneity and rapid degradation. To counteract this shortcoming, this study aims to obtain composite polymer coatings that are stable in physiological environments and exhibit good osteoblast responses in terms of cell adhesion and viability.
Studies have shown that coatings based on cellulose acetate and hydroxyapatite and/or magnesium particles can be obtained by solvent evaporation. TGA analysis confirmed the stability of the sample at up to 200 °C and highlighted that in the temperature range of 5-9 °C, the mass loss was between 25% and 250% due to the presence of water in the material structure.
After 250°C, the coating shows significant weight loss due to polymer degradation. The addition of hydroxyapatite to the cellulose acetate polymer matrix affects the topography of the composite sample.
Scanning electron microscopy highlights the formation of hydroxyapatite crystals in the material structure (CA-HAp and CA-HAp-Mg) due to the poor dispersion of inorganic fillers in polymer solutions.
In the case of CA-HAp and CA-Mg composite samples, a homogeneous structure and cell viability values above 80% were obtained. Analysis of experimental composite samples highlighted the positive effects of magnesium and hydroxyapatite particles when used alone.
Two types of granules (hydroxyapatite and magnesium) are not recommended as mixed fillers. In future studies, we will use only inorganic fillers to obtain CA-based composite coatings on magnesium alloys, as these composite coatings show better results from an in vitro testing perspective for future potential orthopedic biodegradable implants for trauma.