Graphene-Based Composites for Biomedical Applications

Graphene-Based Composites for Biomedical Applications: Comparison

Please note this is a comparison between Version 1 by Rita Teixeira-Santos and Version 2 by Lindsay Dong.

The application of graphene-based materials in medicine has led to significant technological breakthroughs. The remarkable properties of these carbon materials and their potential for functionalization with various molecules and compounds make them highly attractive for numerous medical applications. To enhance their functionality and applicability, extensive research has been conducted on surface modification of graphene (GN) and its derivatives, including modifications with antimicrobials, metals, polymers, and natural compounds.

graphene-based materials
surface modification
antimicrobial activity
biocompatibility
biomedical applications

1. Introduction

In recent years, graphene materials have attracted significant interest due to their remarkable properties and various applications. Graphene (GN) is a two-dimensional carbon allotrope composed of a single layer of carbon atoms arranged in a hexagonal lattice [1]. This structure, which has a high surface area and large aspect ratio, confers high electronic and thermal conductivities to GN, as well as superior mechanical strength ^[2][3][4][2,3,4]. In addition, GN exhibits a high ability to interact with other molecules through various processes, including physical and chemical interactions [5].

Graphene derivatives can be generated by introducing oxygen-containing functional groups to the GN structure or reducing its oxide form, obtaining graphene oxide (GO) or reduced graphene oxide (rGO) [6], respectively. Additionally, GN monolayers can be modified with metals, antimicrobial drugs, polymers, and natural compounds ^{[7][8][9][10]}[7,8,9,10]. While these derivatives preserve their original properties, they also offer enhanced advantages, such as improving the GN dispersion factor in solvents or polymeric matrices and reducing GN toxicity ^[11][12][13][11,12,13]. As a result, GN has been applied in numerous industries, including construction [14], energy [15], food [16], environmental [17], and biomedical [18].

Within the medical field, there have been notable technological advances in the application of GN and its derivatives in drug/gene delivery, biosensing, bioimaging [19], wound healing [20], and tissue engineering [18]. Furthermore, due to the antimicrobial activity and biocompatibility of GN-based materials, they are deemed suitable for manufacturing implantable medical devices such as cardiovascular stents, orthopaedic scaffolds, and urinary implants ^[21][22][21,22].

2. Graphene Modified with Antimicrobials

One desirable feature of graphene is its capacity to bind to a variety of molecules, including antimicrobial agents, peptides, and biocides. This capability not only broadens the range of potential applications of GN materials but also has the potential to enhance their antimicrobial activity and biocompatibility ^[23][47]. Table 1 summarizes recent studies that assessed the biocompatibility and antimicrobial performance of GN materials modified with antimicrobial compounds, including antibiotics ^[24][48], antimicrobial peptides ^[8][25][26][8,49,50], and disinfectants ^[27][36].

Table 1.

Studies focusing on the biocompatibility and antimicrobial activity of graphene modified with antimicrobials.

Graphene Material	Biomedical Application	Biocompatibility	Microorganism	Main Conclusions	Ref.
Doxycycline (Dox)-graphene oxide (GO) immobilized on titanium (TiO₂)	Medical devices	Dox-GO/TiO₂ did not affect the viability of human fibroblasts (over 80% cell viability).	Escherichia coli Staphylococcus aureus	Dox-GO/TiO₂ reduced the viability of adhered bacteria by over 90%, whereas the GO/TiO₂ surface inactivated adhered bacteria by 40%.	^[24][48]

^[29][30]. However, their biocompatibility can vary depending on factors such as the type of metal chosen and the method of conjugation.

3. Graphene Modified with Metals

Various metals and metal oxides have been utilized to modify the surface of GN and its derivatives in order to enhance their antimicrobial activity. Metals are known for their strong antimicrobial properties against a wide range of pathogens [52,53,54]. However, their biocompatibility can vary depending on factors such as the type of metal chosen and the method of conjugation.

Table 2 presents studies that have evaluated the antimicrobial activity and biocompatibility of GN materials modified with metals or metal oxides. Several authors have focused on surface-modifying GO ^[31][32] or rGO ^[33][34] with silver nanoparticles (AgNPs). The resulting composites exhibited higher inactivation rates against both Gram-positive and Gram-negative bacteria, with the exception of the composite synthesized and tested by Wierzbicki et al. ^[31] against

presents studies that have evaluated the antimicrobial activity and biocompatibility of GN materials modified with metals or metal oxides. Several authors have focused on surface-modifying GO [55,56] or rGO [57,58] with silver nanoparticles (AgNPs). The resulting composites exhibited higher inactivation rates against both Gram-positive and Gram-negative bacteria, with the exception of the composite synthesized and tested by Wierzbicki et al. [55] against

Salmonella enteritidis (approximately 50% reduction. The modification of GN-based materials with AgNPs results in a synergistic effect, as they inactivate bacteria by interacting with proteins and enzymatic thiol groups ^[33]. Furthermore, composites containing AgNPs appear safe for medical use.

(approximately 50% reduction. The modification of GN-based materials with AgNPs results in a synergistic effect, as they inactivate bacteria by interacting with proteins and enzymatic thiol groups [57]. Furthermore, composites containing AgNPs appear safe for medical use.

Table 2. Studies addressing the biocompatibility and antimicrobial activity of graphene modified with metals.

Graphene Material	Biomedical Application	Biocompatibility	Microorganism	Main Conclusions	Ref.
Silver nanoparticles (AgNPs)-reduced graphene oxide (rGO)	Medical textiles	NP	Escherichia coli	AgNPs-rGO composites exhibited enhanced activity against E. coli (100% inactivation) compared to rGO (82.5% inactivation).	^[33][57]
Antimicrobial peptide (CATH-2)–reduced graphene oxide (rGO)	Medical devices	Functionalized rGO induced low cytotoxicity towards erythrocytes in comparison to rGO alone.	E. coli	Peptide-functionalized rGO exhibited higher antimicrobial activity compared to rGO (13.3- and 21.8-mm inhibition halo).	^[25][49]
AgNPs-graphene oxide (GO)	NE	The viability of human cells was not changed when incubated on nanoplatforms coated with AgNPs-GO.	Salmonella enteritidis	AgNPs-GO nanoplatform significantly inhibited S. enteritidis growth (over 50% cell inactivation).	^[31][55]	Antimicrobial peptide (ponericin G1)/growth factor (bFGF)/poly(lactide-co-glycolide (PLGA)-GO composite
AgNPs-rGO immobilized into polyurethane/cellulose acetate matrix	Wound healing	Wound healingProduced composite increased cell proliferation compared to PLGA (p < 0.05).	E. coli	In vivo data demonstrated that AgNPs-rGO-based film significantly promoted the wound healing process.	Pseudomonas aeruginosaS. aureus	Staphylococcus aureusPonericin G1/PLGA-GO reduced bacteria growth compared to PLGA or PLGA-GO composite (p < 0.05).	[8]
The produced film exhibited an inactivation rate of 100% for Gram-negative bacteria and 95% against Gram-positive bacteria.	^[	³⁴	^]	[	58] *	Antimicrobial peptide (OH30)/polyethylene glycol (PEG)-GO	Wound healing	OH30/PEG-GO had high cell viability (over 80%) and low toxicity.	S. aureus	In vitro data demonstrated that OH30 released by the synthesized composite inhibited S. aureus growth by up to 95% after 3 h. In vivo data indicated that, on day 7, the number of S. aureus in wounds containing the composite was 6 times less than OH30 or PEG-GO (p < 0.05).	^[26][50] *
AgNPs-GO deposited on nickel-titanium alloy	Medical devices	NP	Streptococcus mutans	AgNPs-GO reduced the number of S. mutans viable cells by up to 5 Log.	N-halamine-GO fibrous membrane	NS	NP	E. coli	Synthesized composite exhibited high biocidal activity against E. coli (>90%).	^[27][36]

NP, Not Performed; NS, Not Specified; *, in vivo study.

3. Graphene Modified with Metals

^[
³²
^]
[
56	]
Gold (Au)-decorated amine-functionalized graphene oxide (NH₂-GO)	Implant devices	Au-NH₂-GO did not affect the viability of human cells (approximately 100% viability).	Bacillus subtilis E. coli
₂
-GO nanocomposite inhibited
E. coli
,
P. aeruginosa	,	S. aureus	, and	S. typhi biofilms by 38, 40, 31, and 35%, respectively.	^[37][61]

NP, Not Performed; NS, Not Specified; *, in vivo study.

4. Graphene Modified with Polymers

Graphene surface modification with polymers is an interesting approach that can offer several advantages, including enhanced GN dispersion and improved mechanical properties and biocompatibility ^[38][39]. In addition, the association with polymers can potentially increase the antimicrobial activity of GN-based materials, as the polymers serve as a matrix that helps GN dispersion ^[40], thus promoting contact with microorganisms. Furthermore, some polymers have inherent antimicrobial properties because they contain cationic groups that can facilitate interactions with bacteria ^[41][42].

Graphene surface modification with polymers is an interesting approach that can offer several advantages, including enhanced GN dispersion and improved mechanical properties and biocompatibility [62,63]. In addition, the association with polymers can potentially increase the antimicrobial activity of GN-based materials, as the polymers serve as a matrix that helps GN dispersion [35], thus promoting contact with microorganisms. Furthermore, some polymers have inherent antimicrobial properties because they contain cationic groups that can facilitate interactions with bacteria [64,65].

Table 3

summarizes studies addressing the biocompatibility and antimicrobial activity of GN materials modified with natural and non-natural polymers. Both pristine GN and GO have been modified with various polymers for potential medical applications (e.g., tissue engineering, wound dressing, or implantable medical devices).

Table 3. Studies reporting the biocompatibility and antimicrobial activity of graphene modified with polymers.

Graphene Material	Biomedical Application	Biocompatibility	Microorganism	Main Conclusions	Ref.
Non-natural polymers
Polyoxyalkyleneamine (POAA)-graphene oxide (GO)	Surface coatings	NP	Bacillus subtilis Escherichia coli	After 3 h, bacteria exposed to POAA-GO decreased their viability to at least 75%.
E. coli
and
S. aureus
(up to 35 and 32 mm inhibition halo, respectively).
[
10
]
CS/polyethylene glycol (PEG)-decorated GO biocomposite	Wound healing	Cell survival on CS/PEG-GO was 95.4%.	E. coli S. aureus	CS, 1 wt% CS/GO and 1 wt% CS/PEG-GO were able to inactivate S. aureus by 80, 85, and 100% and E. coli by 65, 85, and 95%, respectively.	^[48][42]
Carboxymethyl Chitosan (CC)-GO-based Sponge	Wound healing	CC/L-cysteine-GO sponge showed a high cell viability rate, as demonstrated by Live/Dead staining.	E. coli S. aureus	In vivo data indicated that the CC/L-cysteine-GO sponge had a faster wound-healing rate than CC/GO. In vitro tests revealed that the addition of L-cysteine-GO and GO to CC increased sponges’ antimicrobial activity.	^[49][43] *
Folic acid (FA)/silk fibroin (SF)-GO	Wound healing Tissue engineering	The viability of fibroblast cells exposed to FA/SF-GO for 24 h was 97%.	P. aeruginosa	After 24 h, FA/SF-GO film reduced biofilm formation by 80% compared to control (polystyrene).	^[50][73]

NP, Not Performed; NS, Not Specified; VBNC, viable but non-culturable; *, in vivo study.

5. Graphene Modified with Natural Compounds

Owing to their biodegradability, renewability, and biocompatibility, there has been growing interest in composites that incorporate natural compounds. In recent years, several natural compounds, including vivianite ^[51], usnic acid (UA) ^[52], quercetin ^[53], and juglone ^[53], have been studied in conjugation with GN composites.

Owing to their biodegradability, renewability, and biocompatibility, there has been growing interest in composites that incorporate natural compounds. In recent years, several natural compounds, including vivianite [74], usnic acid (UA) [75], quercetin [76], and juglone [76], have been studied in conjugation with GN composites.

Table 4

presents the biocompatibility and antimicrobial activity of these modified GN materials against several Gram-positive and Gram-negative bacterial species.

Table 4. Studies demonstrating the biocompatibility and antimicrobial activity of graphene modified with natural compounds.

Graphene Material	Biomedical Application	Biocompatibility	Microorganism	Main Conclusions	Ref.
Hydroxyapatite/Vivianite-GO	NS	Cell viability of osteoblasts in the presence of this composite was 98%.	E. coli S. aureus	Composite exhibited activity against E. coli and S. aureus after 24 h (14.5 and 13.4 mm inhibition halo, respectively).	^[51][74]
Usnic acid (UA)-GN	Medical devices	NP	S. aureus Staphylococcus epidermidis	After 24 h, UA-GN inhibited S. aureus and S. epidermidis biofilms by 3 Log at 25, 50, 100, and 200 µg/mL AU/GO compared to GN films and glass, except for S. aureus growing on 25 µg/mL AU-GN. After 96 h, staphylococcal biofilms were reduced by 5 Log compared to the control (glass).	^[43][67]
^[	⁵²	^]	[	75	]	Poly(ε-caprolactone) (PCL)-GO	Tissue engineering	Human fibroblasts kept their culturability and proliferation for up to 14 days.	E. coli Staphylococcus epidermidis	PCL-GO composites inactivated S. epidermidis and E. coli adhered cells by 80% after 24 h.	^[44][68]
Quercetin-GO	Drug delivery systems	GO-based materials showed a biocompatible behavior at lower concentrations (>70% cell viability).	E. coli S. aureus	Quercetin/GO composites reduced S. aureus culturability by 1 Log and E. coli culturability by 5 Log.	^[53][76]	PCL-graphene (GN)
Juglone-GO	Nasal implants	Drug delivery systemsNP	Materials showed a biocompatible behavior at lower concentrations (>70% cell viability).E. coli Staphylococcus aureus	The efficacy of PCL-GN against	E. coli S. aureusS. aureus was about 90%. In contrast, this composite did not exhibit activity against E. coli.	Juglone/GO composites reduced S. aureus culturability by 3 Log and E. coli culturability by 5 Log.^[41][64]
^[	⁵³	^]	[	76	]	P. aeruginosa S. aureus	The synthesized material exhibited a higher (5-fold more) antibacterial activity against Gram-positive and Gram-negative bacteria than bare Au or NH₂-GO material.	Epoxy-rich-GO (er-GO)	Wound dressing	Human cells exposed to er-GO exhibited viability ratios greater than 100%.	E. coli S. aureus	er-GO composite decreased in vitro E. coli and S. aureus viability by up to 57 and 97%, respectively. In vivo data indicated that E. coli and S. aureus viability was reduced by 47 and 68%, respectively, in presence of er-GO.^[35][59]
^[	⁴⁵	^]	[	69	] *	Copper oxide (CuO)-GO nanohybrids into bacterial cellulose (BC) matrix	NS	CuO-GO/BC film exhibited excellent biocompatibility towards fibroblast cells (>100%).	B. subtilis E. coli P. aeruginosa S. aureus	After 3 h, CuO-GO/BC films completely inactivated Gram-positive bacteria while only reducing the viability of Gram-negative bacteria by 20%.	^[28]
Poly(Lactic-co-Glycolic Acid) (PLGA)-graphene nanoplatelets (GNP)	NE	NP	E. coli	At 15 MHz, PLGA-GNP composites reduced E. coli viability by 33%, while at lower frequencies (10 and 5 MHz), these films decreased bacteria viability by up to 60%.	[52]
[	9	]	CuO-rGO	NS	NP	P. aeruginosa	CuO-rGO composites led to complete bacterial inactivation (7 Log reduction).
Polydimethylsiloxane (PDMS)-GNP	Implantable medical devices	NP	Pseudomonas aeruginosa S. aureus	The PDMS-GNP reduced the number of total (57%), viable (69%), culturable (55%), and VBNC cells (85%) of S. aureus biofilms. A decrease of 25% in total cells and about 52% in viable, culturable, and VBNC cells was observed for P. aeruginosa biofilms.	^[29][53]
^[	⁴⁶	^]	[	66	]	Copper nanoparticles (CuNPs)-graphene (GN) supported on silicon (Si) wafers	NS	CuNPs-GN/Si showed slight toxicity for human cells (15% reduction in cell viability).	E. coli S. aureus	In the presence of CuNPs-GN/Si films, S. aureus growth was completely inhibited, and E. coli
Natural polymers	viability was reduced by 87%.		^[	³⁰	^][54]
				Palladium (Pd)/polypyrrole (PPy)-rGO composite	Tissue engineering	Pd/PPy-rGO (<100 µg/mL) did not substantially affect osteoblast viability (>80%).	B. subtilis E. coli Klebsiella pneumoniae P. aeruginosa	Pd/PPy-rGO nanocomposite significantly inhibited the biofilm formation of B. subtilis (72%), E. coli (90%), K. pneumoniae (89%), and P. aeruginosa (83%).	^[36][60]
Chitosan (CS)-graphene oxide (GO)	Surface coatings	NP	B. subtilis E. coli	After 3 h, bacteria exposed to CS-GO composite decreased their viability to less than 10%.	^[43][67]	Cerium oxide (CeO₂)-GO	Wound healing	NP
CS/poly(vinyl alcohol) (PVA)-GO nanocomposites	Tissue engineering	After 30 days of film implantation, the absence of injuries in the intervened areas with normal healing was observed.	E. coli P. aeruginosa S. aureus	Bacillus cereus S. aureusSalmonella typhi	E. coli Salmonella spp.CeO	Biocomposites containing 0.75 and 1 wt.% GO completely inhibited pathogen growth.	^[47][70] *
CS/PVA-GO	Wound healing	CS/PVA-GO hydrogels showed nontoxicity towards pre-osteoblast cells (>70% cell viability).	E. coli S. aureus	Hydrogels exhibited high antimicrobial activity against

NP, Not Performed; NS, Not Specified.

6. Conclusions

Biocompatible and antibacterial nanomaterials are in high demand for a variety of medical applications. The surface modification of GN and its derivatives (e.g., graphene oxide or reduced graphene oxide) with antimicrobials (e.g., antimicrobial peptides or biocides), metals or metal oxides, polymers, and natural compounds has enhanced the functionality and applicability of these materials, resulting in improved antimicrobial performance and increased biocompatibility towards human tissues. The combination of graphene materials with agents that possess intrinsic antimicrobial properties, such as antimicrobial peptides, metals (silver or copper), or chitosan, enhances the effectiveness of GN materials in inactivating bacteria, especially Gram-positive bacteria, because of their less complex membrane structure. Conversely, although promising for a wide range of applications, the use of non-natural polymers for GN surface modification results in composites with lower antimicrobial activity than those obtained through the modifications mentioned above. However, GN–polymer composites exhibit superior biocompatibility compared to antimicrobial or metal-based GN composites. In the latter case, adverse effects on human cells are highly dependent on the type of metal used and the methodology employed for their production.