Green Synthesis of ZnO and CuO Nanoparticles Using Macadamia Nut Shell Extracts: Photocatalytic and Antibacterial Activities

doi:10.21203/rs.3.rs-5265919/v1

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Green Synthesis of ZnO and CuO Nanoparticles Using Macadamia Nut Shell Extracts: Photocatalytic and Antibacterial Activities

https://doi.org/10.21203/rs.3.rs-5265919/v1

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Green synthesis methods have garnered significant attention as eco-friendly alternatives to conventional chemical approaches, which often involve hazardous substances. In this study, macadamia nut shell extract was used as a natural stabilizing and capping agent for the sustainable and environmentally friendly synthesis of zinc oxide nanoparticles (ZnO-NPs) and copper oxide nanoparticles (CuO-NPs). The natural compounds present in the macadamia nut shell extract facilitate the formation of these nanoparticles. The synthesized ZnO and CuO NPs were confirmed by various types of spectroscopic and microscopic techniques. XRD analysis confirmed that ZnO NPs exhibit a hexagonal wurtzite structure, while CuO NPs display a face-centered cubic lattice structure. The green-synthesized nanoparticles demonstrated excellent antioxidant and antimicrobial properties against both Gram-positive and Gram-negative bacteria. Moreover, the photocatalytic efficiency of the nanoparticles was assessed by examining the degradation of tetracycline, with ZnO demonstrating superior performance compared to CuO. The antimicrobial activity of ZnO and CuO NPs was tested against Staphylococcus aureus (strain 902), Escherichia coli (strain 443), Aspergillus niger, and Candida albicans, showing promising results. Furthermore, both ZnO and CuO NPs exhibited synergistic effects when combined with antibiotics. The newly developed green-synthesized ZnO and CuO NPs proved to be simple, cost-effective, and environmentally friendly, and utilized for various applications, including food packaging, biosensors, and catalysis in pharmaceutical reactions.

Zinc oxide

Copper oxide

Macadamia nut shells

Antioxidant

Antibacterial

Hydrogenation reaction

Over the past few years, material scientists have increasingly prioritized the exploration of environmentally conscious methods in synthesizing nanoscale materials [1, 2]. The synthesis of nanoparticles involves various methods, such as chemical precipitation, sol-gel processes, mechanical milling, vapor-phase deposition, and biological methods [3, 4]. All the methods offer distinct advantages and is chosen based on the specific requirements of the desired nanoparticles and their intended applications [5]. Lately, the plant-based synthesis of nanoparticles has emerged as a rising trend in green chemistry, known for being straightforward, cost-effective, and environmentally friendly [6–8]. Over the few decades, the noble metal nanoparticles exhibit remarkable properties and utilized for the various application including medicine, electronics, and materials science [9–11]. But they also come with certain limitations, more economic, size uniformity, surface stability and so on. The scientist are actively working on overcoming these limitations through innovations in synthesis method, alternative cheapest nanomaterials with similar properties, surface modification techniques etc.,[12, 13] In this regard, metal oxide and green synthetic methods are gaining importance in various fields, including nanotechnology materials science and pharmaceuticals, as researchers and industries seek sustainable alternatives to traditional synthetic approaches [14, 15]. Islam et al reported the green synthesis of ZnO NPs using Allium cepa L. waste peel extracts, the obtained ZnONPs shows outstanding antioxidant and antibacterial activities [16]. The aforementioned methods contribute to the development of environmentally friendly technologies with a smaller ecological footprint.

Further, ZnO NPs exhibit diverse advantages as inorganic materials, encompassing outstanding optical, mechanical, thermal, semiconducting, and piezoelectric properties, as well as chemo-sensor capabilities [17]. These attributes position ZnO-NPs for utilization in a wide array of industries such as cosmetics, paramedical, energy storage, textile, and healthcare [18, 19]. Moreover, CuO-NPs have garnered significant attention due to their unique properties and multifunctional applications [20, 21]. CuONPs exist in various forms, including cuprous oxide (Cu₂O) and cupric oxide (CuO) and they exhibit distinctive optical, electrical, and catalytic characteristics, making them valuable in several fields [22, 23]. Green synthesis methods have gained prominence as researchers and industries seek alternatives that are both sustainable and economically viable. Baglari et al. explored the green synthesis of CuO nanoparticles using citrus maxima peel extract [24]. These approaches aim to create materials with minimal environmental impact while addressing the growing demand for eco-friendly technologies across various sectors. For the green synthesis process, various plant extract have been utilized to preparation of nanoparticles [25]. Various plants are utilizing for the green synthesis processes to produce nanoparticles[26, 27]. The extracts or biomolecules derived from these plants serve as reducing agents, stabilizers, or capping agents in the formation of nanoparticles. The plant extract was derived from various plants including aloe vera, neem leaf, turmeric, green tea, cinnamon, tulsi, mint leaf and tea leaf extract [3, 28, 29]. Macadamia nut shell extract contains various bioactive compounds that can serve as reducing and capping agents in the synthesis of nanoparticles. In the course of this study, we employed green synthetic methods to synthesize ZnO and CuO NPs, making use of macadamia nut shell extract. The resulting nanosized ZnO and CuO particles, stabilized by the macadamia nut shell extract, were evaluated for their antibacterial properties against E. coli and S. aureus bacteria. Furthermore, these nanoparticles exhibited effective photocatalytic capabilities in degrading various organic pollutants, including methylene blue, methyl orange, and tetracycline. Further, mechanism of the photocatalytic performance and antibacterial study was investigated.

2.1 Synthesis of macadamia nuts-shell extract

The macadamia nut shells were coarsely powdered using a mechanical blender machine [30–32]. The obtained powder (100 g) was placed in a round-bottomed flask, and 200 mL of ethanol and water (1:1 ratio) was added. The extraction was performed at 100°C for 5 h with constant stirring. Then, extraction was repeated until getting clear solution using water solvent. The extract was filtered and evaporate the ethanol solvent. Then remaining solvent was removed by freeze drying process. The macadamia nut shells derived crude extract was stored in airtight container bottles at 4 ºC.

2.2 Synthesis of ZnO and CuO NPs

ZnO and CuO NPs were prepared by eco-friendly green synthetic route using macadamia nuts cell extract [2, 33]. Initially, 0.2 g of macadamia nuts shell extract was dissolved in 25 mL of distilled water in two different beakers. Then stoichiometric amount of Zn(NO₃)₃.6H₂O and Cu(NO₃)₂.6H₂O were added into the respective beaker then adjust the pH around 11. The resulting white precipitate was kept in a pre-heated using muffle furnace at 400 ºC. The resulting fine powdered milky white and black coloured ZnO and CuO was formed.

2.3 Photocatalytic degradation of dye

The photodegrading efficacy of the prepared ZnO and CuO NPs was examined through degradation tetracycline (TC) under visible irradiation. The photo catalytic degradation of organic wastes was carried out using a 200 x 100 mm batch reactor at room temperature [34]. The reaction mixture was prepared by adding 20 mg of the catalyst to 50 mL of a 10 ppm organic pollutant solution. The catalyst-containing solution was stirred under dark conditions to establish the adsorption-desorption equilibrium. Afterward, the photocatalytic reaction mixture was exposed to UV-Visible light irradiation. During the irradiation, the concentration of TC was monitored by measuring the absorbance intensity of the solution at regular intervals. Before UV-visible analysis, the catalyst particles were removed by centrifugation to avoid interference in the UV spectrum. The photocatalytic degradation efficiency of methylene blue was then determined using the following equation:

$$\:Degradation\:efficiency=\frac{C0-C}{C0}*100\dots\:\dots\:\dots\:\dots\:\dots\:\dots\:\dots\:\dots\:\dots\:\dots\:\left(1\right)$$

Here C₀ and C is the original and residual concentration of organic pollutant. The stability of the photocatalyst was carried out using recyclability test.

2.4 Antimicrobial study

The detailed experimental procedure for antimicrobial study was provided in supplementary information (SI file).

The X-ray diffraction (XRD) patterns of ZnO and CuO show definite line broadening of the diffraction peaks, indicating that the green-synthesized particles are in the nanoscale range, as shown in Fig. 1a. The XRD peaks for ZnO are located at 31.81°, 34.49°, 36.25°, 47.59°, 56.58°, 62.84°, 66.36°, 67.92°, and 69.09°, corresponding to the (100), (002), (101), (102), (110), (103), (200), and (112) planes, respectively [17, 30]. The crystalline diffraction peaks have been indexed to the hexagonal phase of ZnO, with a high degree of crystallinity, according to JCPDS card number 36-1451. Similarly, the diffraction peaks for CuO are located at 32.51°, 35.44°, 38.76°, 48.12°, 53.62°, 58.31°, 61.63°, 66.14°, 68.04°, 72.39°, and 75.13°, corresponding to the (110), (002), (111), (202), (020), (127), (113), (022), (220), (222), and (313) planes. These diffraction peaks confirm that the prepared CuO has a monoclinic structure, matching well with JCPDS card number 89-5895[33, 35]. Furthermore, the diffraction peaks revealed that all the characteristic peaks correspond to ZnO and CuO NPs. Additionally, no extra peaks appeared in the ZnO and CuO NPs, indicating that there are no impurities present in the ZnO and CuO NPs. The particle diameters of the ZnO and CuO crystallites were calculated using the Debye-Scherrer formula. Based on Bragg's diffraction angle (θ) and the full width at half maximum (FWHM) of the more intense ZnO peaks corresponding to the (101) planes located at 36.25°, the crystalline size is 24 nm, while the average crystalline size is around 35.16 nm. Similarly, for the CuO NPs, the most intense peaks correspond to the (002) planes at 35.44°, with a crystalline size of 62.8 nm and an average size of around 68.54 nm. The obtained indexing confirms the standard hexagonal wurtzite structure (JCPDS file no. 00-036-1451) for ZnO NPs and the monoclinic structure (JCPDS card number 89-5895) for CuO NPs, as previously reported in other studies.

Furthermore, the surface functional groups of macadamia nut shell extract (Mac) and Mac-stabilized ZnO and CuO NPs were characterized by FTIR analysis, as shown in Fig. 1b. The broad peak observed at 3350 cm^− 1 is attributed to -OH functional groups, indicating the presence of polyphenols, flavonoids, reducing sugars, alkaloids, and tannins in the Mac extract[31]. Additionally, the band at 1628 cm^− 1 is attributed to N-H bending in amines (alkaloids) or C = O stretching in polyphenols and flavonoids. These functional groups play a crucial role in the formation of ZnO and CuO NPs. Moreover, a metal-oxygen bond vibration was observed at 598 cm^− 1, indicating their participation in the formation of the nanoparticles (Fig. 3b) [36]. Specifically, the metal-oxygen bond vibrations were observed at 605 cm^− 1 for Zn-O and 674 cm^− 1 for Cu-O, respectively, further confirming the formation of ZnO and CuO NPs.

The surface morphology of the as-prepared ZnO and CuO NPs is shown in Fig. 2a and 3a. Figure 2 shows SEM images of pure ZnO NPs. It was observed that the particles in pure ZnO exhibit a spherical-like shape with diameters in the range of 50 to 72 nm. Similarly, in Fig. 3a, the SEM images of pure CuO NPs show a spherical shape with sizes ranging from 40 to 65 nm [37]. The elemental mapping images of ZnO NPs reveal that Zn, O, C, and N elements exist and are uniformly dispersed in the matrix (Fig. 2b-f). Furthermore, Fig. 3(b-f) shows the elemental mapping analysis of CuO NPs, indicating that Cu, O, C, and N elements are evenly distributed in the matrix. Additionally, the ZnO and CuO NPs were confirmed by elemental dispersive spectroscopy (EDS) analysis, with the corresponding images shown in Fig. 4(a-b), respectively. In Fig. 4a, it is revealed that the prepared ZnO NPs contain 68.25%, 18.17%, 0.6%, and 13.52% of Zn, O, N, and C elements, respectively. Similarly, Fig. 4b shows that the prepared CuO NPs contain 65.46%, 18.66%, 0.11%, and 15.78% of Cu, O, N, and C elements, respectively. These experimental results confirm that the ZnO and CuO NPs were successfully formed.

The size and morphology of the ZnO and CuO nanoparticles (NPs) derived from macadamia extract were analyzed using HRTEM, as shown in Fig. 5a and Fig. 5c, respectively, with the corresponding SAED patterns displayed in Fig. 5b and Fig. 5d. The ZnO nanoparticles in Fig. 5a exhibit an average size of approximately 28.34 nm, as indicated by the scale measurement. The particles appear aggregated but maintain a distinguishable shape, suggesting that the synthesized ZnO NPs possess good crystallinity and nanoscale dimensions. The slight agglomeration is typical of ZnO due to its high surface energy. In contrast, Fig. 5c shows that the CuO NPs have a larger average size of about 78.2 nm. The CuO particles display a distinct spherical morphology with more uniform dispersion compared to the ZnO sample. Additionally, the label "ME" likely refers to the macadamia extract, which helps stabilize the ZnO and CuO NPs. The SAED pattern of ZnO, shown in Fig. 5b, displays distinct diffraction rings, confirming the crystalline nature of the ZnO NPs. The sharp and clear diffraction spots suggest a high degree of crystallinity, consistent with the aggregated nanoparticle structure observed in the TEM image. Similarly, the SAED pattern of CuO (Fig. 5d) reveals well-defined diffraction rings, indicating a polycrystalline structure. The pattern is characteristic of CuO's monoclinic phase, suggesting that the synthesized CuO particles maintain a robust crystal structure.

The diffuse reflectance UV-Vis (DRS UV) spectra presented in the image compare the optical properties of ZnO and CuO, highlighting their distinct absorbance behaviors and band gap energies (Fig. 6a). In Fig. 5a, the ZnO absorbance sharply drops around the UV region, corresponding to a steep cutoff near 370 nm. This suggests that ZnO primarily absorbs UV light while allowing visible light to pass through, making it an efficient UV-blocking material. The corresponding Tauc plot (Fig. 6b) indicates a direct optical band gap of approximately 2.72 eV for ZnO, which is consistent with previous literature for bulk ZnO, typically in the 3.2 eV range [38]. This slightly lower value may suggest some degree of defect states or a slight reduction in crystallinity. Additionally, in Fig. 6c, shows the absorbance spectrum of CuO, exhibits broader absorbance extending into the visible region, which is characteristic of materials with narrower band gaps. The CuO NPs shows a more significant absorption over a broader wavelength range, including both UV and visible regions. The Tauc plot analysis for CuO reveals an optical band gap of 1.32 eV, which aligns with the known semiconductor properties of CuO (Fig. 6c). This lower band gap compared to ZnO indicates that CuO can absorb lower-energy visible light, making it suitable for applications like solar energy harvesting and photocatalysis in the visible spectrum.

The photocatalytic activities of ZnO NPs and CuO NPs were evaluated for the degradation of tetracycline (TC) under visible light, with the corresponding results illustrated in Fig. 7a. The UV-Vis absorption spectrum of methylene blue (MB) displayed a strong peak at 663 nm. Upon exposure to visible light in the presence of the catalysts, the absorbance of the MB solution decreased progressively over time, indicating successful photodegradation. When comparing the photocatalytic performances of ZnO and CuO NPs, ZnO NPs demonstrated superior efficiency, attributed to its higher electron mobility, better stability, and lower charge carrier recombination rates (Fig. 7b). The rate constants (k) derived from the linear fits of ln(Ct/C0) vs. time (Fig. 7c) were 0.0311 min⁻¹ for ZnO NPs and 0.0162 min⁻¹ for CuO NPs, respectively [39]. As shown in Fig. 7d, the photocatalytic degradation efficiencies of ZnO NPs and CuO NPs were 98.13% and 74.23%, respectively. The photocatalytic efficiency of the ZnO NPs shows better catalytic activity than that of CuO NPs. As indicated by the TEM results, the smaller size of ZnO NPs compared to CuO NPs contributes to the enhanced photocatalytic performance.

To better understand the active radicals involved in the photodegradation process, benzoquinone (BQ), isopropanol (IPA), and ammonium oxalate (AO) were introduced into the system as radical scavengers (Fig. 8a). These scavengers were employed to specifically target ˙O₂⁻, ˙OH, and h⁺ radicals, respectively. Following the addition of BQ, IPA, and AO, the degradation efficiency of MB by ZnO decreased to 20.3%, 67.4%, and 86.8%, respectively. This indicates that the ˙O₂⁻ radical is the primary active species in the photodegradation process, while ˙OH and h⁺ species play secondary, yet significant, roles.

Additionally, the stability, activity, and reusability of the photocatalyst are crucial factors for its potential in industrial applications, such as in the dyeing industry, ceramic antioxidants, and natural-origin antioxidants, among others. For the recycling experiments, the photocatalyst was subjected to five consecutive cycles under the same conditions. Before each cycle, the ZnO and CuO particles were washed with deionized water and methanol, then dried for reuse. As shown in Fig. 8b, c, the results indicate that even after multiple uses, the free radical scavenging capacity remained similar. However, a significant decrease in activity was observed for ZnO, with a drop from 97.12–90% and from 74–50% over the cycles. The recycling efficiency results reveal that ZnO NPs exhibit better catalytic and recycling efficiency compared to CuO NPs.

3.1 Antibacterial and antifungal activity

The antibacterial activity of green synthesized ZnO-ME and CuO-ME was evaluated against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) using the agar diffusion method, as illustrated in the provided in Fig. 9. From In the tests involving ZnO (Fig. 9a-b), a clear zone of inhibition is visible around the ZnO discs, indicating significant antibacterial activity[40]. In Fig. 9a shows the results for S. aureus, where ZnO exhibits effective inhibition at concentrations of 50 µg/mL, 100 µg/mL, and 250 µg/mL. Similarly, Fig. 9b demonstrates ZnO's antibacterial effects against E. coli, with evident inhibition zones at the same concentrations. The presence of inhibition zones suggests that ZnO effectively disrupts bacterial growth through mechanisms such as the generation of reactive oxygen species (ROS) and the release of Zn²⁺ ions, which damage bacterial cell walls and interfere with vital cellular functions.

Conversely, CuO NPs (Fig. 9c-d) also displays notable antibacterial properties. In Fig. c shows the impact of CuO NPs on S. aureus, with discernible inhibition zones at concentrations of 50 µg/mL, 100 µg/mL, and 250 µg/mL. Figure 9d presents the results for E. coli, revealing similar inhibitory effects. The antibacterial activity of CuO NPs can be attributed to the release of Cu²⁺ ions and ROS production, which cause oxidative stress, damage cellular structures, and ultimately lead to bacterial cell death. Additionally, the macrostructures of CuO may enhance its interaction with bacterial cells, thereby improving its antibacterial efficacy. Comparatively, both ZnO and CuO NPs demonstrate strong antibacterial activities, with clear inhibition zones indicating effective bacterial suppression. The results underscore the potential application of these materials in antimicrobial coatings, water treatment, and medical devices to prevent bacterial infections and promote hygiene. In summary, the antibacterial activity of ZnO and CuO NPs against S. aureus and E. coli is evident from the significant inhibition zones observed in the agar diffusion assays. These findings highlight the effectiveness of ZnO and CuO NPs as potent antibacterial agents, offering promising prospects for various applications requiring antimicrobial properties.

3.2 Antifungal Activity of ZnO NPs (a-b) and CuO NPs (c-d)

ZnO NPs shows significant antifungal activity, as evidenced by the clear zones of inhibition around the discs in the agar plates. In Fig. 10a, displays the results for A. niger, where ZnO NPs effectively inhibits fungal growth at concentrations of 50 µg/mL, 100 µg/mL, and 250 µg/mL. Figure 10b demonstrates similar antifungal effects against C. albicans, with visible inhibition zones at the same concentrations. The antifungal activity of ZnO NPs can be attributed to several mechanisms, including the generation of reactive oxygen species (ROS), which cause oxidative damage to fungal cells, and the release of Zn²⁺ ions, which disrupt cellular functions and integrity. The macrostructure of ZnO enhances its surface area, providing more active sites for interaction with fungal cells, thereby improving its efficacy.

CuO NPs also exhibits strong antifungal properties. Figure 10c shows the inhibition of A. niger by CuO NPs, with clear zones of inhibition at concentrations of 50 µg/mL, 100 µg/mL, and 250 µg/mL. Similarly, Fig. 10d presents the results for C. albicans, indicating effective fungal suppression at the same concentrations. The antifungal activity of CuO NPs is primarily due to the release of Cu²⁺ ions, which penetrate the fungal cell membranes, causing structural damage and interfering with enzymatic processes. Additionally, the generation of ROS by CuO NPs induces oxidative stress, leading to cell death. The macrostructure of CuO provides a larger surface area for interaction, enhancing its antifungal performance. The efficiency of the both ZnO and CuO NPs demonstrate substantial antifungal activities, as indicated by the inhibition zones. The size of these zones suggests the materials are effective at relatively low concentrations. While both materials rely on ion release and ROS generation, their structural differences and elemental compositions might influence their specific interactions with fungal cells.

In this study, ZnO and CuO NPs were successfully synthesized using a green approach with macadamia nut shell extracts serving as both reducing and stabilizing agents. The eco-friendly synthesis method yielded nanoparticles with effective photocatalytic and antibacterial activities. ZnO NPs demonstrated superior photocatalytic performance in degrading tetracycline (TC), achieving higher degradation efficiency compared to CuO, particularly under solar light irradiation. ZnO and CuO NPs photocatalysts achieved 97.74% and 74% TC degradation, respectively, within 120 minutes, confirming the superior photocatalytic efficiency of ZnO NPs. Additionally, both ZnO and CuO NPs exhibited strong antibacterial properties against E. coli and S. aureus, with ZnO showing slightly higher activity. Recycling experiments further confirmed that ZnO NPs possess greater stability and reusability, making them promising candidates for applications in wastewater treatment and antibacterial coatings. This green synthesis route not only provides an eco-friendly alternative for nanoparticle production but also emphasizes the potential of waste biomass in the development of functional nanomaterials.

Acknowledgements:

The Authors greatly acknowledge the financial support provided by Faculty of Science and Technology, Thammasat University, Contact No. SciGR29/2567. Additionally, the author extend their sincere appreciation to FOODMATRIX GLOBAL CO., LTD and DOI TUNG DEVELOPMENT PROJECT (Mae Fah Luang Foundation under Royal Patronage), for their continuous financial support, assistance with characterization, and provision of raw materials.

Author contributions:

Yodchai Tangjaideborisu: Conceptualization, Methodology, Formal Analysis, Investigation, Prema Yugala: Conceptualization, methodology; Choowin Phanawansombat: Conceptualization, Methodology, data curation, Paramasivam Shanmugam: Supervision, Writing original draft, Writing—Review & Editing, Supakorn Boonyuen: Supervision, Writing—Review & Editing, Pariya Na Nakorn: Investigation, resources, Writing—Review & Editing

Funding: Not applicable

Data availability

Data will be made available on request

Ethical approval: Not applicable

Consent to participate: Not applicable

Consent for publication: Not applicable

Competing interests: the authors declare no competing interests

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No competing interests reported.

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Green Synthesis of ZnO and CuO Nanoparticles Using Macadamia Nut Shell Extracts: Photocatalytic and Antibacterial Activities

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Abstract

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1 Introduction

2. Experimental methods

2.1 Synthesis of macadamia nuts-shell extract

2.2 Synthesis of ZnO and CuO NPs

2.3 Photocatalytic degradation of dye

2.4 Antimicrobial study

3. Results and discussion

3.1 Antibacterial and antifungal activity

3.2 Antifungal Activity of ZnO NPs (a-b) and CuO NPs (c-d)

4 Conclusion

Declarations

References

Additional Declarations

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