Hemostatic, biocompatible, and antibacterial non-animal fungal mushroom-based carboxymethyl chitosan-ZnO nanocomposite for wound-healing applications
Introduction
There is increasing interest in the use of biomedical inorganic nanomaterials in biomedical applications because of their unique biological characteristics [1]. During the development of various inorganic nanomaterials, metal oxide nanoparticles have shown promise for use in biomedical applications, especially for antibacterial, drug delivery, biosensing, and cell imaging applications [[2], [3], [4]]. Several types of metal oxides have been synthesized, such as TiO2, ZnO, and CuO, for use in various applications [[2], [3], [4]]. Among these, ZnO nanoparticles are of particular interest in biomedical applications because they are safe to use and inexpensive to produce easily [5]. Moreover, ZnO nanoparticles have been generally recognized as safe and approved for use by the US-FDA [6]. ZnO has been included in a wide range of applications in the biomedical field due to its anti-cancer, drug delivery, antifungal, antibacterial, and agricultural properties [2,5,6]. Interactions between ZnO nanoparticles and bacteria are mostly toxic, a property that has been exploited in antibacterial applications in the biomedical and food industry fields [7]. Also, it has been used to enhance the healing of both acute and chronic wounds because of its bacteriostatic and epithelization properties [8]. ZnO nanoparticles with varying sizes and morphologies exhibit significant antibacterial properties across a wide spectrum of bacteria [9]. The toxic nature of ZnO nanoparticles in that bacterial spectrum has been mainly attributed to their high specific surface area-to-volume ratios and their distinctive physicochemical properties [9].
ZnO nanoparticles with different morphologies, such as nanowire, nanoflake, a nanobelt, nanoflower, nanorod, and spherical have been reported [10]. Various methods, including sol-gel, thermal decomposition of organic precursor, microemulsion, ultrasonic, electrodeposition, microwave-assisted technique, hydrothermal, chemical vapor deposition, and precipitation, have been employed for the synthesis of ZnO nanoparticles with various morphological structures [10]. Recently, natural extracts created from plant parts such as stem, root, leaf, seed, and fruit have been used in the synthesis of ZnO nanoparticles due to the specific or exclusive phytochemicals they contain [11]. Using natural extracts is eco-friendly, and the extraction process is relatively inexpensive; moreover, it does not involve the use of intermediate base groups. Bio-reduction of zinc ions to ZnO nanoparticles with the help of phytochemicals, such as polyphenolic compounds, vitamins, amino acids, alkaloids and terpenoids secreted from the plant parts, have been reported [11]. Polysaccharides, as well as some other ‘green’ sources, have been used for the synthesis of ZnO nanoparticles and are reported to potentially have antibacterial properties [[12], [13], [14]]. Moreover, combining polymers with ZnO nanoparticles is one of the most reliable approaches for obtaining polymer composites with application potential in wound care.
Recently, advanced wound dressing products using biopolymers, and/or synthetic polymers with the combination of nanofillers have shown potential to promote progressive events in wound healing [15,16]. However, the development of synergistic combination of materials have remains a challenge for wound healing. Many biopolymers have already been used in the preparation of combination ZnO nanoparticles; however, the antibacterial property of the product arises from the ZnO nanoparticles, not from the polymer. Antibacterial synergism is a challenge in the development of effective treatment modalities that do not affect normal cells. Thus, combinatorial therapies that can improve antibacterial activity are being investigated. Chitosan (CS) is a linear polysaccharide composed of randomly distributed beta-(1-4)-linked D-glucosamine (deacetylated) and N-acetyl-D-glucosamine (acetylated). It is widely used in various biomedical applications due to its biocompatibility, its mucoadhesion property, and its ability to form gels [17]. However, a drawback in using CS for biomedical purposes is due to its insolubility in water, although it is soluble under acidic pH conditions [17]. The modified CS such as quaternized CS, and N-carboxyethyl CS (NCS) have improved water solubility and a broader pH solubility and showed excellent biological performance for wound dressing applications [[18], [19], [20], [21], [22]]. Carboxymethyl chitosan (CMCS) is also modified CS and can interact with cells and has been associated with successful cell growth and wound-healing results [23,24]. CMCS is also used in cosmetics, as it has stabilizing agent moisture absorption and retention properties and antimicrobial characteristics [[25], [26], [27]]. Thus, it may be expected that a combination of CMCS and ZnO could have a synergetic antibacterial effect without a toxic effect on normal cells, potentially enhancing wound healing. Recently, non-animal CS-derived CMCS has been used in biomedical applications due to its non-allergic responses and the renewable, inexhaustible, biodegradable, non-toxic, and non-ecotoxic properties of non-animal CS [[28], [29], [30]]. Moreover, it is safe for people who are vegan or are allergic to crustacean-sourced CS. In the present work, a simple method was employed for the preparation of combination CMCS-ZnO nanoparticles by using a non-animal fungal mushroom-derived CMCS (NAM-CMCS) as a stabilizer and reducing agent under ultrasonic treatment and ambient laboratory temperature conditions. We expected the combination of NAM-CMCS and ZnO (NAM-CMCS-ZnO) would produce a synergistic antibacterial product with biocompatibility and hemostatic properties as compared to NAM-CMCS and ZnO. To the best of our knowledge, there are no previous reports on a hemostatic, biocompatible, and antibacterial ZnO nanocomposite created using NAM-CMCS. The structural characteristics of the NAM-CMCS-ZnO nanocomposite were investigated by various characterization techniques. Furthermore, the biological properties of the NAM-CMCS-ZnO nanocomposite were assessed to determine the hemostatic, biocompatibility, and antibacterial properties when used in a wound dressing application.
Section snippets
Materials
Mushroom Agaricus Bisporus, Aspergillus Niger derived NAM-CMCS (MW = 200,000–2000000DA with deacetylation 80–98%; Viscosity 20–1000 mpas; randomly distributed N, O-carboxymethyl-β-(1 → 4)-linked D-glucosamine (deacetylated unit) and N-acetyl-D-glucosamine (acetylated unit)) was received as a gift from the Endovision Company, Daegu, South Korea. Zinc nitrate (Zn(NO3)2) was purchased from Sigma Aldrich. All other chemicals were used as received. Double distilled water was used as needed.
Synthesis of NAM-CMCS-ZnO
The
Synthesis and conformation of NAM-CMCS-ZnO
After considering the potential excellent properties of NAM-CMCS, in this paper, we enhanced ZnO nanoparticle characteristics by developing a hemostatic, antibacterial, and biocompatible NAM-CMCS-ZnO nanocomposite via an ultrasonic method under ambient laboratory temperature conditions (Scheme 1). The formation of NAM-CMCS-ZnO nanoparticles was confirmed by XRD and XPS techniques. The XRD patterns of the pristine NAM-CMCS and NAM-CMCS-ZnO nanoparticles are shown in Fig. 1a. The XRD peaks of
Conclusion
A NAM-CMCS-ZnO nanocomposite was synthesized by applying an ultrasonic-assisted method under ambient laboratory temperature conditions. The FTIR, XRD, and XPS results confirmed that the formation and composition of the NAM-CMCS-ZnO nanoparticles. SEM and SEM-EDS elemental mapping confirmed the spherical shape of NAM-ZnO nanoparticles and indicated an average size of 18 ± 3.6 nm. The enhanced antibacterial activity of NAM-CMCS capped ZnO NPs toward S. aureus was ascribed to synergetic activities
Authorship contribution statement
Kummara Madhusudana Rao: Conceptualization, Methodology, Validation, Investigation, Resources, review & editing.
Maduru Suneetha: Methodology, Validation, Investigation, Writing - original draft.
Gyu Tae Park: Methodology, Validation.
Anam Giridhar Babu: Methodology, Validation.
Sung Soo Han: Resources, review & editing, Supervision.
Declaration of competing interest
The authors declare that they have no competing interests.
Acknowledgements
K.M. Rao would like to acknowledge the NRF of South Korea funded by the Ministry of Education, Science, and Technology, grant number NRF-2 2019R1I1A3A01063627.
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