Eco-friendly synthesis of silver nanoparticles using lignin peroxidase produced from <i>Caldibacillus thermoamylovorans</i> and cultivated on waste walnut shell

The preparation of silver nanoparticles (AgNPs) via an environmentally friendly green synthesis method represents an ecologically promising alternative. This research aims to develop sustainable and eco-friendly AgNPs using the lignin peroxidase (LiP) enzyme from Caldibacillus thermoamylovorans, cultivated on waste walnut shells, which are rich in lignin, to meet the growing demand for AgNPs. Among thermophilic bacteria that were isolated, the C. thermoamylovorans SA1 strain showed the highest LiP activity. The production of LiP was optimized by adding waste walnut shells and manipulating the environmental parameters. The optimal conditions were determined at 50 g/l shell amount, 96 h, pH 8, 140 rpm, and 60°C. In parallel with the increase in enzyme activity, bacterial growth also increased. As a result of the optimization, the highest enzyme activity value was 435.0 U/ml and bacterial growth was determined to be OD600: 2.09. The extracellular medium obtained from the bacteria grown in walnut shell medium was then added to an AgNO3 solution. Efficient production of AgNPs was achieved by stirring the mixture at 50°C-60°C for 4 h under optimum conditions. The synthesized AgNPs were characterized using a range of analytical techniques, including UV‒Vis spectroscopy, Fourier transform infrared spectroscopy, X-ray diffraction, and scanning electron microscopy. The biological efficacy of the synthesized AgNPs was evaluated by assessing their antibacterial activity against pathogenic bacteria, such as Escherichia coli O157:H7, Klebsiella pneumoniae, Staphylococcus aureus, Streptococcus pyogenes, and Bacillus cereus. The highest activity was observed against B. cereus (15 mm). The broad-spectrum antibacterial properties exhibited by the AgNPs synthesized in this study offer a promising and sustainable solution for diverse applications in various sectors, including the environmental, agricultural, medical, and pharmaceutical fields.