Fabrication and Characterization of Copper Nanoparticles by Green Synthesis Approach Using Plectranthus Amboinicus Leaves Extract

The eld of nanotechnology is gaining interest among the researchers towards the eco-friendly way of synthesis of nanoparticles. In this project, green synthesis technique was employed to induce the synthesis of copper nanoparticles using Plectranthus amboinicus, i.e. Mexican mint, identied as Coleus amboinicus leaf extract. We report an eco-friendly synthesis of copper nanoparticle using Plectranthus amboinicus leaf extract, which is a simple and an ostentatiously rapid method which produces stable nanoparticles. The copper sulphate solution was naturally employed as a precursor for synthesizing the copper nanoparticles. The extract of the plant Plectranthus amboinicus was found to showcase excellent reducing and stabilizing properties. By using Ultraviolet-Visible spectroscopy, Zeta Potential, and X-Ray Diffraction (XRD) studies, it was conrmed that copper nanoparticles have been synthesized. The UV-Spectrometer analysis shows the characteristic peak indicating the synthesis of copper nanoparticles. The pattern of XRD analysis showed particle size of 16 - 25 nm and it reveals high crystallinity of the copper nanoparticles. Zeta potential was done to nd the charge of the nanoparticles and size distribution which showed to have signicant stability. This method proves to be cost-effective, can be performed at ease, and it’s also free of pollutants.

nanoparticles. The extract of the plant Plectranthus amboinicus was found to showcase excellent reducing and stabilizing properties. By using Ultraviolet-Visible spectroscopy, Zeta Potential, and X-Ray Diffraction (XRD) studies, it was con rmed that copper nanoparticles have been synthesized. The UV-Spectrometer analysis shows the characteristic peak indicating the synthesis of copper nanoparticles. The pattern of XRD analysis showed particle size of 16 -25 nm and it reveals high crystallinity of the copper nanoparticles. Zeta potential was done to nd the charge of the nanoparticles and size distribution which showed to have signi cant stability. This method proves to be cost-effective, can be performed at ease, and it's also free of pollutants.

Introduction:
Copper is a cost-effective material and has gathered attention due to its extensive applications in nanoparticle synthesis for novel catalytic antimicrobial and other optical properties [1]. To gain the complete bene ts of copper nanoparticles, an ecofriendly and economically suitable, synthesis is desired. Copper is less toxic and; it is an important element in human health [2].
Copper is a less-expensive metal than other metals which are often utilized as Nano-sized particles for increasing antibiotic effectiveness [3]. The ratio between area and volume is high for copper nanoparticles. The antibacterial activity, catalytic potency, magnetic and optical characteristics are higher for copper when compared to other metals like gold and silver [4]. The important applications of copper nanoparticles are to act as an antibacterial compound [5]. The application of copper nanoparticles towards health-related processes is important because of its antifungal and antibacterial activities [6]. The applications of copper nanoparticles are wide, as it is used in many elds like cosmetics for nano pigment purpose and in pharmaceutical industries for drug-eluting [7]. In food packages, to reinforce the antibacterial system, the laser-generated copper nanoparticles were used [8]. By the reduction process using CuSO4 as the precursor, the copper nanoparticles are synthesized [9]. For synthesizing copper nanoparticles there are three methods, namely chemical, physical, and biological methods [10].
The biological method using extracts of the plant has been put forward for consideration as a possible environmental-friendly method than physical and chemical methods [11]. The advantage of plants concentrate on their biosynthetic reaction is one of the important factors in the biosynthesis of nanoparticles [12]. Synthesis of copper nanoparticles in solutions showed the absorption peak at 574 nm [13]. Copper nanoparticles synthesized by the electro-reduction process showed XRD patterns peaks very sharply due to high nano-crystalline property of the metal where 2θ values are 43.4°, 50.5° and 74.2° [14]. Zeta potential was used to evaluate the morphological properties of nanoparticles. A study revealed that the zeta potential for copper nanoparticle at a concentration of 20 mg/L was − 9.25 mV [15]. Materials And Methods:

Preparation of Plectranthus amboinicus leaf extract:
Leaves of Plectranthus amboinicus were washed and cleaned thoroughly with sterile water, nely cut and soaked in 100 ml of distilled water, and boiled on a hot plate. The leaves were strained using a lter cloth and the leaf extract was cooled at room temperature and was stored in the deep freezer.

Synthesis of Copper nanoparticles using Plectranthus amboinicus leaf extract:
The leaf extract of Plectranthus amboinicus has inherent reducing properties, thereby aiding in the reduction of aqueous copper sulphate into copper nanoparticles. The aqueous CuSO4 solution with concentration 1mM was prepared and the leaf extract of Plectranthus amboinicus that was prepared and was mixed in 1:5 ratio (v/v), i.e., 2ml of the plant extract was added to 10 ml of copper sulphate solution, dropwise on a magnetic stirrer. A positive control solution as shown in the Fig 1(a) using distilled water and plant extract was prepared and a negative control solution as shown in the Fig 1(b) that contained only copper sulphate solution was also prepared. The solutions were incubated overnight in the darkness at room temperature and monitored constantly. After 24 hours, the change in colour of the solution from light green to dark green was observed. By centrifuging the solution at 12000 rpm for 10 -15 min, the copper nanoparticle was obtained. The pellet was stored at 4 o C for the characterization and analysis [16].

Characterization Of Copper Nanoparticles:
The important factor of metal nanoparticles is that their optical properties change due to the morphological characteristics of the nanoparticles [17]. The change in color from light green to dark green denoted the copper nanoparticles formation as shown in the Fig 2(a) and 2(b) respectively. UV-Vis spectral analysis was done by using Shimadzu UV-Visible spectrophotometer with a resolution operating at 1 nm used and the UV spectrum was recorded. The powder X-Ray Diffraction analysis was done using Malvern PANanalytical instrument with 15 mA, 40 kV using Cu -Kα radiation in θ -2θ con gurations. Dynamic Light Scattering analysis was deployed to decode the mean grain size of the nanoparticles by zeta potential analysis using HORIBA.
Results And Discussion:

Ultraviolet Visible Spectrophotometer:
The copper nanoparticles formation is usually con rmed by UV-Vis spectroscopy studies of colored solution [18]. The concept of obtaining the peak in the UV-Vis spectrum is that, on the surface of metal nanoparticles the free electrons will be present which undergoes strong repulsion with the light of speci c wavelength [19]. Higher the conversion of copper sulphate solution to copper nanoparticles, higher will be the absorbance which corresponds to higher concentration of copper nanoparticles [20].
The spectroscopy studies of the solution were recorded after 24 hours. The peak of absorbance from the absorption spectra was between the ranges of 230 nm -270 nm as shown in Fig. 3 pinpointing copper nanoparticles formation.

X-Ray Diffraction (XRD):
X-Ray Diffraction is an analytical method for determining the size and crystalline structure of the nanoparticles [21]. The high degree of crystallinity of nanoparticles is re ected by the intensity and sharpness of peaks [22]. There is an undeniable role played by the secondary metabolites in the plant extract even if the clear mechanism is unknown [23]. The average size of the copper nanoparticles was obtained using Scherrer's Formula, Where Dp depicts the average grain size, K is Scherrer's constant which is 1.54060, λ denotes the X-Ray diffraction wavelength which has a value of 1.54178 nm, B is FWHM i.e, Full Width at Half Maximum derived from the peak of XRD. The observed XRD peak broadening is said to come from the crystalline size. θ is the XRD peak position (i.e.,) the Bragg's angle obtained from the half of 2θ, where 2θ is in the range of 30 -80 o [24].

Zeta Potential:
The Z-average was estimated as 5.3 nm as shown in the Fig 5(a) using DLS. The charge obtained using zeta analysis was -25.2mV as shown in the Fig 5(b). Particles that have charges of or above -25 and +25 tend to be more stable according to the previous research documents [25]. Particles that have negative charges tend to have a strong force of repulsion between them.
This indicates the properties of stability and quality [26].

Conclusion:
The environment friendly methods for the fabrication of copper nanoparticles can be developed by applying knowledge from the areas of nanotechnology and incorporated into use with the plant extract of Plectranthus amboinicus. The copper nanoparticles were produced by the plant extract which functions both as a reducer and a stabilizer. Thereby, the copper sulphate solution was reduced into copper ions without employing harsh man-made chemicals. From this study, it is evident that the reaction is completely a green chemical process favorable for nanoparticle synthesis. We used UV-visible spectroscopy, Zeta potential analysis, and XRD analysis techniques for characterization of formed copper nanoparticles. UV absorbance peaks were observed between 230 nm-270 nm, therefore con rming the presence of Copper nanoparticles. The copper nanoparticles were analyzed using XRD and the average grain size was calculated to be 21.47 nm. DLS assessment showed the average diameter i.e., Z-average of the Copper nanoparticles to be 5.3 nm and the surface charge of -25.2 mV indicating its high stability. The use of Plectranthus amboinicus is bene cial for the production of copper nanoparticles because it is readily available, less expensive, has a simple method that makes it easy to perform, acts as reducing and stabilizing agents, contributes in producing nanoparticles that are stable and crystalline in structure. Thus, proves to be one of the best plants for the production of the copper nanoparticle by the eco-friendly green synthesis method. All data generated or analyzed during this study are included in this manuscript. Figure 1 A positive control solution as shown in the Fig 1(a) using distilled water and plant extract was prepared and a negative control solution as shown in the Fig 1(b) that contained only copper sulphate solution was also prepared.

Supplementary Files
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