Synthesis of NiO Nanoparticle from L. acutangula peel
A useful method to determine if the precursors are completely reduced was UV-visible spectroscopy analysis. As the nickel oxide solution was incubated, a colour shift was discernible in the extract of L. acutangula peel. There was no confirmation of the formation of nickel oxide nanoparticles, and the pure NiNO₃ concentrate of L. acutangula peel without watery leaves did not exhibit any color variation. After the NiNo₃ and L. acutangula peel extract reacted, the colour turned mustard yellow for three hours during a stirred conditional incubation phase. After that, no significant changes happened.¹⁸ The stimulation of plasmonic resonance at the surface during the production of Nickle oxide NPs was the origin of this distinctive colour variation. Now, the optical range of 200-800 nm was employed to document the absorption NiO NPs from UV to visible light. The absorption of UV-visible spectra of NiO NPs is shown in Figure 2. The creation of NiO nanoparticles was indicated by the greatest absorption peak, which was detected at 320 nm. The absorption peak for green produced nickel nanoparticles was observed around 342 nm in earlier studies.¹⁹ The produced titanium dioxide nanoparticles exhibit a comparable absorption peak in comparison to the prior research.²⁰

FTIR Analysis
The involvement of phytochemicals found in Luffa acutangula peel extract in the formation of nanoparticles was verified by FTIR analysis. Phenols and alcohols’ O-H stretching vibrations are shown by the wide absorption band at 3456 cm^-1. Carbonyl groups’ C=O stretching vibrations are responsible for the peak at 1680 cm^-1, whereas alkanes’ C-H stretching is indicated by peaks close to 3034 cm^-1. C=C or C-N stretching vibrations may be represented by the peaks seen at 1447 cm^-1.

The C-O stretching vibrations of alcohols and carboxylic acids are represented by the peaks at 1183, 1061, and 1006 cm^-1, indicating the role of phytochemicals in the reduction and stabilization process (Figure 3). The effective production of NiO nanoparticles is confirmed by the distinctive Ni-O stretching vibration seen below 700 cm^-1.^21-23

XRD analysis
The crystalline nature of the synthesised NiO nanoparticles was verified by the XRD pattern. The (111), (200), (220), (311), and (222) crystal planes, which are typical of the face-centered cubic (fcc) rock-salt structure of NiO nanoparticles, are represented by the diffraction peaks seen at 2θ values of 37.31°, 43.35°, 62.91°, 75.53°, and 79.45° (Figure 4). The creation of phase-pure cubic NiO nanoparticles with no discernible impurities was confirmed by the diffraction data’s good agreement with standard JCPDS card No. 47-1049.

The average crystallite size was estimated using the Debye-Scherrer equation:

D = 0.94λ / βcosθ

where D is the crystallite size, λ is the wavelength of CuKα radiation (0.15406 nm), β is the full width at half maximum (FWHM), and θ is the Bragg diffraction angle. The average crystallite size was found to be approximately 15-18 nm, which agrees well with FESEM observations.^24,25

FESEM analysis
The surface morphological characteristics of the produced NPs were examined using FESEM. FESEM pictures of the NiO nanoparticle are shown in Figure 5 at a 65k magnification. Nearly spherical and clumped nanoparticles with an average size of 13-18 nm are visible in the image. The tight size distribution shown by the measured sizes (13.85 nm, 15.84 nm, and 17.68 nm) validates the successful production of NiO nanoparticles in the nanometer range.

Surface hydroxyl groups and remaining phytochemicals from the L. acutangula peel extract form hydrogen bonds, which causes the mild aggregation typical of nickel oxide nanoparticles made from the peel. The plant metabolites may have served as stabilizing and capping agents, preventing overproduction during calcination, as indicated by the tiny, porous structure.²⁶

Anti-bacterial activity
The synthesised NiO nanoparticles shown concentration-dependent antibacterial activity against Pseudomonas aeruginosa, Bacillus cereus, and Staphylococcus aureus. At a concentration of 100 µL, Bacillus cereus showed the largest zone of inhibition (32 ± SD mm). On the other hand, there was very little inhibition against Escherichia coli. (Figure 6; Table). The dosage of nanoparticles ranges from 50-100 µL, depending on how much the zone produces as the concentration increases.

Table. The zone of inhibition (mm, mean ± SD) of NiO nanoparticles synthesised from Luffa acutangula peel

The complex outer lipopolysaccharide membrane seen in Gram-negative bacteria, which limits nanoparticle penetration and lessens ROS-mediated damage, may be the cause of E. coli’s decreased susceptibility. Gram-positive bacteria, on the other hand, have a somewhat porous peptidoglycan layer that makes it easier for nanoparticles to connect with the bacterial membrane. NiO nanoparticles’ antibacterial action is primarily linked to the production of reactive oxygen species (ROS), oxidative stress, membrane rupture, denaturation of proteins, and intracellular component leakage. Additionally, the phytochemicals adsorbed on the surface of the nanoparticles may enhance the antibacterial efficiency in a synergistic way.^27-34