Green Synthesis Method of ZnO Nanoparticles using Extracts of <i>Zingiber officinale</i> and Garlic Bulb (<i>Allium sativum</i>) and Their Synergetic Effect for Antibacterial Activities

Kebede Urge, Solomon; Tiruneh Dibaba, Solomon; Belay Gemta, Abebe

doi:https://doi.org/10.1155/2023/7036247

Journal of Nanomaterials

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Abstract Introduction Results and Discussion Conclusion Data Availability Conflicts of Interest Acknowledgments References Copyright Related Articles

Research Article | Open Access

Volume 2023 | Article ID 7036247 | https://doi.org/10.1155/2023/7036247

Green Synthesis Method of ZnO Nanoparticles using Extracts of Zingiber officinale and Garlic Bulb (Allium sativum) and Their Synergetic Effect for Antibacterial Activities

Solomon Kebede Urge,¹Solomon Tiruneh Dibaba,¹and Abebe Belay Gemta¹

Academic Editor: Vidya Nand Singh

Received29 Aug 2022

Revised26 Nov 2022

Accepted29 Nov 2022

Published12 Jan 2023

Abstract

In this paper, antibacterial activities of zinc oxide (ZnO) nanoparticles synthesized through green method using garlic bulb (Allium sativum), ginger (Zingiber officinale) extracts, and their mixture are reported. The synthesized ZnO NPs were characterized by X-ray diffraction (XRD), ultraviolet visible (UV–vis), photoluminescence (PL), spectroscopy and Fourier transform infrared (FTIR) spectroscopy. The crystalline sizes of ZnO NPs synthesized using garlic bulb (Allium sativum) extract, ginger (Zingiber officinale) root extract, and their mixture were 19.8, 21.94, and 23.86 nm, respectively. Similarly, the corresponding peak absorbances were 369.5, 377.5, and 374 nm, respectively. The antibacterial activities of synthesized ZnO NPs were tested against gram-negative bacteria Escherichia coli (E. coli) and Pseudomonas putida (P. putida) and gram-positive bacteria Staphylococcus aureus (S. aureus) and Streptococcus pyogenes (S. pyogenes). The ZnO NPs synthesized using the mixture of garlic bulb (Allium sativum) and ginger (Z. officinale) root extract have shown maximum inhibition zone against gram-negative bacteria (P. putida: 28.67 ± 0.82 mm) and gram-positive bacteria (S. pyogenes: 10.67 ± 0.47 mm) as compared to ZnO NPs synthesized using the two extracts separately. On the other hand, ZnO NPs obtained from garlic bulb and Z. officinale root extracts exhibited maximum inhibition zone against E. coli (19 ± 0.82 mm) and S. aureus (16.4 ± 0.47 mm), respectively.

1. Introduction

The novel properties of nanomaterials bring the rapid development of nanoscience and nanotechnology. Among nanomaterials, metal oxide nanoparticles received exceedingly attention from the scientific community because of their size-tunable several unique properties, such as high chemical stability, high electrochemical coupling coefficient, broad range of radiation absorption, high photostability, easily availability, low cost, and nontoxicity [1–7].

Among metal oxide nanomaterials, zinc oxide (ZnO) nanoparticle is one of the foremost considered metal oxide nanomaterials due to its unique chemical, physical, biological, and optical properties [8–10]. ZnO NPs have many applications in different fields like, optical, piezoelectric, mechanical, biomedical, gas sensing, catalyst, etc. [11, 12].

There are different methods of synthesizing ZnO NPs, including hydrothermal [13], sol−gel processing [14], spray pyrolysis method [15], coprecipitation [16], chemical vapor deposition (CVD) [17], green synthesis method, etc. Among these synthesis methods, green synthesis using plant extracts is commonly preferred for the synthesis of ZnO NPs due to eco-friendly, cost-effective, and nontoxicity of the resulting NPs for human therapeutic use [18].

Green synthesis of ZnO NPs has been carried out by extracting different plants such as Cassia fistula and Melia Azadarch [19], garlic (Allium sativum) [20], Z. officinale (ginger), Aloe vera [21], coffee [22], orange peel extract [23], Cardamom [24], and Lippia adoensis (koseret) [25]. However, the comparison of antimicrobial activities of ZnO synthesized using ginger (Z. officinale), garlic (Allium sativum) extract, and most importantly the antimicrobial activities of ZnO nanoparticles synthesized by the synergetic effect of the mixture of the two extracts were not performed so far to the best of our knowledge. This study was conducted to compare antibacterial activities of ZnO NPs synthesized using Z. officinale root extract, garlic bulb (Allium sativum) extract, and their mixture against the two standard pathogens, gram-positive and gram-negative bacteria.

Ginger is the most frequently and heavily consumed dietary condiments in the world. Ginger is frequently prescribed by Indian and Chinese medicine for the treatment of cough, common cold, rheumatism, snake bite, and respiratory problems [26]. It is also used in treatment of cancer, skin diseases, heart palpitation, swelling, urinary problems, stomach problems, abdomen problems, dyspepsia, and loss of appetite [27]. Chemically, ginger is rich in flavonoids and polyphenolic compounds such as gingerols, shagols, zingerone, paradol, terphineol, terpenes, borneol, geraniol, limonene, linalool, and alpha-zingiberene [28].

Similarly, garlic is used worldwide as a food additive, spice, and medicine. It has been described as a gastric stimulant. Garlic (Allium sativum) is a rich source of organosulfur compounds showing a variety of biological activities [29] such as, allyl sulfide, allicin, allyl cysteine, ajoene, and alliin [30]. Phenols, terpenoids, ketones, aldehydes, and amides in plants were responsible for the production of metal NPs [31].

2. Experimental Method

2.1. Materials and Chemicals

The materials used for the synthesis were contained: fresh root of ginger (Z. officinale), garlic bulb (Allium sativum), beaker, magnetic stirrer, hot plate, incubator, power supply, thermometer, Whatman No. 1 filter papers, digital electronic analytical balance (Model FA2104, China), furnace (Model BK-5-12GJ), ceramic crucible cups, drying oven (Model 101-0, Biobase Biodustry, Shandong Co. Ltd, China), cylinders, and centrifuge (Model AVI-558, max RPM:5,000 rpm).

The precursor chemicals used for the synthesis were included: zinc acetate (Zn(CH₃COO)₂·2H₂O) (molar: 219.49 g/mol, Merk Life Science, PLC), deionized water, ethanol, sodium hydroxide (NaOH), dimethyl sulfoxide (DMSO, (CH₃)₂OS, 99.9% pure, Loba Chemie Pvt. Ltd., India), zinc acetate dehydrate was provided as precursors and sodium hydroxide was used for controlling the pH values. All the reagents used in the experiments will be in analytical grade and used without any further purification.

2.2. Methods of Extraction

The fresh root of Z. officinale and garlic was purchased from local market (Adama, Ethiopia). Z. officinale sample was washed thoroughly and soaked in distilled water for 20 min in order to remove dirty present on the skin of sample. In the same manner, garlic was washed using distilled water after the outer skin was pilled off. The roots were completely air-dried and the moisture contents were removed. The dried roots were crushed using mortar and pestle. After that, 10 g of the powder of each sample was boiled for 30 min in 100 mL of deionized water (DW). Finally, the extracts were filtered with the Whatman No. 1 filter paper and stored at 4°C for further use. These steps are indicated in Figures 1–3.

2.3. Synthesis of ZnO NPs

Four grams of zinc acetate (Zn (CH₃COO)₂·2H₂O) was added into 50 mL of deionized water (DW) to get final concentration of zinc acetate dehydrate (Zn (C₂H₃O₂)₂·2H₂O). Ten milliliters of Z. officinale root extract and garlic bulb extract were slowly added to zinc acetate dehydrate solution in the different beaker and 10 mL of their mixtures were added to zinc acetate dehydrate solution in the same ratio. 1 M of NaOH were added drop-by-drop to the solution to control the pH of the solutionat 12 [32, 33]. The mixtures were heated and stirred on magnetic stirrer for 2 hr at 50°C until the white precipitate was observed. Then, the pellets were centrifuged three times for 10 min interval at 5,000 rpm and dried in an oven overnight. Then, the dried samples were calcined in a muffle furnace at 450°C for 2 hr and the powders were collected for characterization. Figures 1–3 represent the schematics of synthesized ZnO NPs using garlic bulb extract, Z. officinale root extract, and their mixture, respectively.

2.4. Characterization Techniques

Some of the instruments used for characterization of ZnO NPs were X-ray diffraction (XRD) (X-ray target tube, Cu–K radiation, 40 kV, scanning rate 3° min⁻¹, recorded 2θ range between 10° and 80°) used to study the crystal structure the nanoparticles, UV–visible spectroscopy (UV-3600Plus series) to measure energy bandgap of the particles, photoluminescence (PL) spectroscopy (Cary Eclipse fluorescence spectrophotometer, scan rate 600 nm/min, MY18490002, Agilent Technologies, Malaysia) to determine the emission spectra of the nanoparticles and FTIR spectroscopy (IS50, APX, Germany) to characterize the functional groups attached to synthesized ZnO nanoparticles.

2.5. Antibacterial Activities

The antibacterial activities of ZnO NPs were tested against standard pathogens of gram negative (E. coli-ATCC 9637 and P. putida-ATCC 47054) and gram positive (S. aureus-ATCC 6538 and S. pyogenes-ATCC 19615) by agar well diffusion method (at concentration about 5 × 10⁵ CFU/mL). Ampicillin was used as positive control in each plate. The plates were incubated for 18 hr at 37°C. The stock solution of synthesized ZnO NPs was prepared in dimethyl sulfoxide (DMSO, (CH₃)₂OS, 99.9% pure, Loba Chemie Pvt. Ltd., India) at different concentrations: 15, 10, and 5 mg/mL.

3. Results and Discussion

3.1. XRD Analysis

X-ray diffraction patterns of synthesized ZnO NPs peaks were agreed with the ordinary data (JCPDS card no. 00-036-1451). Figure 4 shows the characteristic Bragg diffraction at 2θ values of the three samples. The first sample is ZnO NPs synthesized using Z. officinale root extract (Z-ZnO NPs: Figure 4(a)) shows the distinctive Bragg diffraction at 2θ values of 31.22, 31.78, 34.07, 34.44, 36.26, 36.93, 44.02, 47.55, 56.61, 62.28, 62.87, 64.38, 66.39, 67.96, 69.09, 76.95, and 77.46. The strongest peaks are 2θ = 36.26, 31.78, and 34.44. The second one is ZnO NPs synthesized using garlic extract (G-ZnO NPs: Figure 4(b)) indicates the distinctive Bragg diffraction at 2θ values of 31.76, 34.42, 36.24, 47.53, 56.02, 56.58, 62.86, 66.39, 67.95, 69.04, and 77.09. The strongest peaks are three peaks observed at 2θ = 36.24, 34.42, and 31.76. The third one is ZnO NPs synthesized using the mixture of Z. officinale root extract and garlic extract (GZ-ZnO NPs: Figure 4(c)) shows the distinctive Bragg diffraction at 2θ values of 31.79, 33.91, 34.45, 36.27, 36.85, 47.56, 56.06, 56.62, 57.18, 62.42, 62.88, 66.41, 67.97, and 69.09. The strongest peaks are 2θ = 34.44, 31.22, and 34.07. They were indexed to the plane (100), (002), (101), (102), (110), (103), (200), (112), and (201) that are in good agreement with wurtzite ZnO structure. The distinct and clear peaks confirmed the high purity and crystalline nature of the prepared ZnO NPs. The crystallite size has been estimated from the broad diffraction peak using the Debye–Scherrer formula.where D is the crystal size of ZnO NPs, k is Scherrer’s constant given by 0.9, λ is the wavelength of X-rays (1.541 Å), θ is the Bragg diffraction angle, and β is the full-width at half-maximum (FWHM) of the diffraction peak corresponding to plane (101). The average particle size of ZnO NPs synthesized using garlic bulb (Allium sativum) extract, Z. officinale (ginger) root extract, and their mixture were found to be 19.8, 21.94, and 23.86 nm, which were derived from the FWHM of the more intense peak (β = 0.422, 0.380, 0.350) corresponding to the (101) plane located at 2θ = 36.24°, 36.26°, and 36.27°, respectively.

The average particle size of synthesized ZnO NPs obtained from XRD data shows good results compared to the previous report [34]. Vidya et al. [10] synthesized ZnO NPs by using Calotropis gigantea and reported the average grain size as 30–35 nm. Yuvakkumar et al. [35] synthesized ZnO NPs using noble plant rambutan (Nephelium lappaceum L) peel extract and has reported the average crystalline size as 50.95 nm.

3.2. Photoluminescence (PL) Spectrum

Figure 5 indicates the PL spectrum (peak emission) of ZnO NPs synthesized using Z. officinale root extract, garlic extract, and their mixture. A broad blue emission peak centered at (421.05 nm), (393 and 420 nm), and (421.95 nm) can be observed for ZnO NPs synthesized using Z. officinale root extract, garlic extract, and their mixtures, respectively. From the evidence of given data (Figure 5), the different peaks exists at different wavelength, which were normally associated with Zn defects, such as transitions of an electron from conduction band to the valance band and O₂ vacancies [36]. The O₂ vacancies change the properties of oxide materials by producing F centers of ionic oxides which may cause the formation of metal−metal bonds in covalent bond. In other case, O₂ vacancies change the oxidation state of compounds of transition metal atoms with empty d orbitals, which can eventually alter the local electronic structure of atoms [37]. In n-type ZnO, acceptor-like defects (O_i) are easier to form, which enables the transition between different bandgap energies [38]. The PL emission rate is directly proportional to the photo-excited load carrier’s recombination probability. Higher PL strength advanced e⁻/h⁺ pair (electron−hole pair) recombination and guides to the lower photocatalytic activity [39].

3.3. UV–Vis Absorbance

Figure 6 shows the UV–vis absorbance of ZnO NPs synthesized by using Z. officinale root extract, garlic extract, and their mixture. The optical bandgap energy of ZnO NPs synthesized using Z. officinale root extract, garlic bulb extract, and their mixture calculated using Tauc formula was found to be 3.2, 3.27 and 3.24 eV at maximum wavelength of 377.5, 369.5, and 374 nm, respectively.

The calculated bandgap energy (E_g) of synthesized ZnO NPs were performed in good agreement with the optical bandgap energy of standard ZnO NPs.

To evaluate the energy band gap we used the Tauc equation:where E_g is energy bandgap, h is Planks constant, α is absorption coefficient, v is the frequency given by (v = c/λ), γ is the nature of electronic transition which equal to 1/2 for direct bandgap, and A is constant, usually taken as 1 for amorphous materials [40].

3.4. FTIR Analysis

FTIR spectroscopy was performed to identify the functional groups associated with the ZnO NPs formation. Figure 7(a)–7(c) represents the FTIR spectrum of ZnO NPs synthesized using garlic bulb extract, Z. officinale root extract, and their mixture, respectively. ZnO NPs absorption band was obtained within the region 4,000–400 cm⁻¹. The bands at 913.09, 919.16, and 915.55 cm⁻¹ in FTIR spectrum confirm the C–H bending of alkenes in ZnO NPs synthesized using garlic bulb extract, Z. officinale root extract, and their mixtures, respectively [41]. The bands that are near to 1,000 cm⁻¹ are associated with metal−oxygen tension and bending, in this case, Zn–O bonding was formed [42]. Broadband at 3,400–3,200 cm⁻¹ showed characteristic OH group. Bands at 1,331.37, 1,391.83, and 1,394.66 cm⁻¹ confirmed the presence of carboxylic acid and aromatic C–H bending [43]. Therefore, the present studies agreed with the previous report.

3.5. Antibacterial Activities

The bactericidal activities of ZnO NPs synthesiszed using garlic bulb extract, Z. officinale root extract, and their mixtures were tested against standard pathogenes gram-negative (G–ve) and gram-positive (G+ve) bacteria. From G–ve, Escherichia coli (E. coli) and Pseudomonas putida (P. putida) and from G+ve, Staphylococcus aureus (S. aureus), and Streptococcus pyogenes (S. pyogenes) were grown to test the antimicrobial activities of synthesized ZnO NPs. Figures 8–10 show zone of inhibition of the four bacteria against synthesized ZnO NPs at the concentration of 15, 10 and 5 mg/mL. The mean values of zone of inhibition of three replicates are shown in Tables 1–3. As the concentration of synthesized NPs increased, the zone of inhibition also increased. The small particle size and the high surface area of ZnO NPs can increase antimicrobial activity, causing an improvement in surface reactivity [44].

Garlic-mediated ZnO NPs were more significant on G–ve bacteria P. putida (21.67 ± 0.88 mm) and E. coli (19 ± 0.82 mm) compared to G+ve bacteria S. aureus (17.7 ± 0.47 mm) and S. pyogenes (7 ± 0.82 mm) as shown in Figure 8 and Table 1. ZnO NPs synthesized using Z. officinale root extract were more effective on G+ve bacteria S. aureus (16.4 ± 0.47 mm) and S. pyogenes (10 ± 0.82 mm) compared to G−ve bacteria P. putida (7.7 ± 0.78 mm) and E. coli (4 ± 1.00 mm) as shown in Figure 9 and Table 2. However, it is possible to say that the zone of inhibition of the mixture of garlic bulb and Z. officinale (GZ)-mediated ZnO NPs was greater than that of garlic (G)-mediated ZnO NPs and Z. officinale root extract (Z)-mediated ZnO NPs against G−ve bacteria like P. putida. These results suggest that the use of ZnO NPs synthesized using the mixture of garlic bulb (Allium sativum) and Z. officinale root extract can be more efficient against gram-negative bacteria (G−ve). This might be due to the presence of higher number of phenolic compounds and rare secondary metabolites present in both garlic and Z. officinale. Dobrucka and Długaszewska [45] reported that reactive oxygen species (ROS) such as superoxide and hydroxyl radicals produced by ZnO NPs were caused damage to bacterial cell membranes.

According to the evidence of zone of inhibition of the recorded data shown in Figure 10 and Table 3, ZnO NPs synthesized using the mixtures of garlic bulb and Z. officinale root extract (GZ) was more effective on the G−ve bacteria P. putida (28.67 ± 0.82 mm) and E. coli (15.3 ± 0.94 mm) compared to G+ve bacteria S. pyogenes (10.67 ± 0.47 mm) and S. aureus (15.67 ± 0.82 mm).

In general, ZnO NPs synthesized using the mixtures of garlic bulb and Z. officinale root extract (GZ) showed the highest inhibition zone, this is maybe due to interaction of ZnO NPs with cell membrane of the bacteria (E. coli, P. putida, S. pyogenes, and S. aureus). The ZnO particles entered into the bacterial cell creates reactive oxygen species (ROS) that react with cysteinyl protein and iron inside the cells and cause DNA and cell wall damage. Besides, the garlic bulb (Allium sativum) extract and Zingiber officinale (ginger) root extract that were used as reducing agent have phytochemicals that are toxic to bacteria. As a result the synergetic effect enhances inhibition zones of the bacteria. The comparison of our results with some previously reported results is shown in Table 4.

4. Conclusion

In this work, ZnO NPs using Z. officinale root extract, garlic bulb extract, and their mixture were successfully synthesized. The XRD examinations of synthesized ZnO NPs were confirmed the formation of wurtzite shape NPs. The absorption spectra of ZnO NPs synthesized using garlic bulb extract, Z. officinale root extract, and their mixture were found to be 369.5, 377.5, and 374 nm, respectively. The bands at 913.09, 919.16, and 915.55 cm⁻¹ in FTIR spectrum confirms the formation of ZnO NPs and the PL spectrum centered at 421.06, 421.96 and 421.95 nm shows the peak emission of ZnO NPs synthesized using garlic bulb extract, Z. officinale root extract, and their mixture, respectively. According to the evidence shown in Figure 10 and Table 3, we observed that ZnO NPs synthesized using the mixture of garlic bulb and Z. officinale root extract, reported for the first time, exhibited high inhibition zone against gram-negative bacteria, P. putida (28.67 ± .0.82). This might be due to ROS generated by the ZnO nanoparticles entered to the bacteria cell and the presence of higher number of phenolic compounds and rare secondary metabolites present in both garlic bulb and Z. officinale root. Further study may be needed to utilize this kind of synergistic green synthesis methods for antifungal and anticancer application in the future.

Data Availability

All the necessary data are already included in the main manuscript.

Conflicts of Interest

The authors declare that they have no conflicts of interest.

Acknowledgments

This work is financially supported by the Adama Science and Technology University.

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Copyright © 2023 Solomon Kebede Urge et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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