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Biodegradation, Antibacterial Performance and Cytocompatibility of SMLed ZK30-Cu-Mn
Table 2. The ion concentrations of SBF and human blood plasma assay. The extracts of SLMed ZK30-0.2Cu-xMn alloys
according to the ISO standard (10 mol/L) were extracted in the humidified atmosphere containing
−3
Ion SBF (pH 7.40) Blood plasma (pH 7.2–7.4) 5% CO at 37°C for 24 h using Dulbecco’s Modified
2
Na + 142 142 Eagle’s Medium as extraction medium with an extraction
2
K + 5.0 5.0 rate of 1.25 cm /ml. The supernatant was extracted,
Mg 2+ 1.5 1.5 centrifuged, and filtered to produce the extract. The
Ca 2+ 2.5 2.5 extract was refrigerated at 4°C to prepare for cell viability
Cl − 148 103 test. CCK-8 assays were used to examine the proliferation
HCO 3 − 4.2 27 of MG63 cells cultured in SLMed ZK30-0.2Cu-xMn and
Ti extracts. The fluorescence live/dead staining assay was
HPO 4 2− 1.0 1.0 performed based on the following procedures. Cells were
SO 4 2− 0.5 0.5 cultured on a 96-well plate at the density of 5 × 10 cells
3
per 100 ml for 24 h to ensure cell adherence. The medium
P =2.279V h (ii) was then replaced by 100 μL extract. After 1 day and
h
3 days of incubation, the cell viability was determined by
P =2.1 ΔW (iii) live/dead staining.
w
The surface morphologies and corrosion products 3. Results
of the specimens after immersion were characterized by
SEM with EDS and X-ray photoelectron spectroscopy 3.1. Microstructure
(XPS). The XPS measurements were achieved using Figure 1 shows optical microstructures of the SLMed
an X-ray source of Mg Kα (1253.6 eV). The binding
energy of the measurement was corrected by the binding ZK30-0.2Cu-xMn alloys. The microstructures of all
energy of C of hydrocarbons (284.6 eV) absorbed on the alloys consist of fine equiaxed grains. The grain
size decreased with increasing Mn content. The grain
1s
the surface.
size of SLMed ZK30-0.2Cu was about 5 μm, as shown
2.5. Antibacterial properties in Figure 1A, which was smaller than that obtained by
traditional casting . Increasing Mn contents decreased
[27]
Staphylococcus aureus (S. aureus, ATCC 25923) is one the grain size, as shown in Figure 1B through Figure 1E.
of the most common bacteria causing infection and was, The grain size of SLMed ZK30-0.2Cu-1.6Mn alloy was
therefore, used as a model bacterium. The preparation about 3 μm. This indicates that the incorporation of Mn
method of SLMed ZK30-0.2Cu-xMn alloy extracts for into SLMed ZK30-0.2 Cu refines grain size. The small
antibacterial test was as follows. After disinfection by grain size produced by SLM is attributed to the rapid
ultraviolet radiation, all the samples and control groups solidification of the melt pool. The additional grain
were cultured in SBF solution with S. aureus in three refinement by alloying Mn is attributed to additional
replicates and placed in a 12-well untreated polystyrene nucleation sites and grain boundary pinning effect
plate. Each well contained sample and S. aureus provided by Mn, which can inhibit grain growth.
suspension with a concentration of with a concentration Figure 2 shows the XRD patterns of the SLMed
of 1 × 10 colony-forming unit (CFU)/ml prepared using ZK30-0.2Cu-xMn alloys. The XRD patterns for
5
–10
sterile SBF solution. The ratio of the specimen surface SLMed ZK30-0.2Cu, SLMed ZK30-0.2Cu-0.4Mn, and
are (cm ) to the solution volume (mL) were 1.25 cm / SLMed ZK30-0.2Cu-0.8Mn included peaks of α-Mg,
2
2
mL. The plates were kept at constant temperature for 4, MgZnCu, and MgZn phases. For a Mn content higher than
2
12, 48, 72, and 96 h at 37 ± 0.5°C. Bacterial cell density 0.8 wt.%, SLMed ZK30-0.2Cu-xMn alloys (x = 1.2 and
in the SBF solution was evaluated by bacterial counting 1.6) also produced diffraction peaks of the α-Mn phases.
after each culture period. Before calculating the number The microstructure of SLMed ZK30-0.2Cu-1.6Mn
of colonies, the suspension was diluted to 1 × 10 CFU/ is presented at a higher magnification SEM micrograph
3
–10
ml, and 0.05 ml suspension was added to the LB nutrient in Figure 3A. Numerous irregularly shaped intermetallic
agar plate, which was carefully spread and plated and phases were distributed inside the grains and along grain
then incubated for 24 h at 37 ± 0.5°C. boundaries. The composition of the intermetallic phases
at Point 1, Point 2, and Point 3 in Figure 3A is presented
2.6. Cytocompatibility
in Figure 3B determined from the EDS spectra. Point 1
Cell compatibility, which is essential for biomedical implant (bright granular precipitate distributed along the grain
materials, was studied using the MG63 osteosarcoma boundaries) was composed of Zn and Mg; Point 2
cells. The cytocompatibility was evaluated by carrying out (short bar-shaped precipitate distributed along the grain
cell proliferation assay and fluorescence live/dead staining boundaries) was composed of Zn, Mg, and Cu; and Point

80 International Journal of Bioprinting (2021)–Volume 7, Issue 1