Evaluating the Environmental and Biomedical Advantages of Green Synthesised Zinc Oxide Nanoparticles Using an Aqueous Extract of Rumex hastatus Roots

Evaluating the Environmental and Biomedical Advantages of Green Synthesised Zinc Oxide Nanoparticles Using an Aqueous Extract of Rumex hastatus Roots | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Evaluating the Environmental and Biomedical Advantages of Green Synthesised Zinc Oxide Nanoparticles Using an Aqueous Extract of Rumex hastatus Roots Nidhi ., Pragti Saini, Anshul Gairola, M. C. Purohit This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9010794/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 6 You are reading this latest preprint version Abstract Zinc oxide nanoparticles are highly functional in pharmaceutical and biomedical applications, and plant-mediated nanoparticles which show good biological activities. They can be synthesised by different chemical, physical and biological methods, whereas the green method, or biosynthesis, is environment-friendly and less hazardous. In the present study, zinc oxide Nanoparticles were synthesised using an aqueous (Distilled water) extract of Rumex hastatus roots and 0.05 M zinc acetate dihydrate (Zn(CH₃COO)₂·2H₂O). Maintaining pH with a digital pH meter and using 0.1 M sodium hydroxide (NaOH), the nanoparticles synthesised, confirmed by physical characterisation using Ultraviolet-Visible spectroscopy, showed typical absorption peaks in the range of 320nm to 400nm due to their significant excitation binding energy at room temperature., X-Ray Diffraction analysis (XRD), which confirmed that the particles are crystalline and have a hexagonal wurtzite structure, ZnO-Bonding shown by the Fourier Transform Infrared Spectroscopy (FT-IR), Differential Scanning Calorimetry (DSC), Zeta potential confirmed thermal and other stability of nanoparticles and size(22-30nm) and shape (spherical to quasi spherical) confirmed by the Field Emission Scanning Electron Microscopy (FE-SEM) and Dynamic Light Scattering (DLS), Their biomedical applications include anti-microbial and antioxidant properties. In anti-microbial four pathogens were tested: two bacteria, Salmonella Typhi (S. typhi) MTCC733 and Escherichia coli (E. coli) MTCC452 and two fungi, Trichoderma reesei (T. reesei) MTCC164 and Aspergillus niger (A. niger) MTCC282.in which a 36.33% against Salmonella typhi in gram-positive bacteria, a high inhibition rate and anti-oxidant, and an IC-50 value greater than 400 µg/ml, suggests only moderate capability in scavenging free radicals when compared to potent standards like ascorbic acid, which demonstrates an IC-50 below 20 µg/ml. Biosynthesis of zinc oxide Nanoparticles characterisations antioxidants and antimicrobial activities Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 1. Introduction The biosynthesis of ZnO nanoparticles relies on natural reducing and stabilising agents derived from biological resources, such as plant extracts, microorganisms (bacteria, fungi, algae), and biomolecules (proteins, amino acids, polysaccharides). These natural agents serve as capping and stabilising agents, avoiding the use of harmful chemicals. The process generally involves the reaction of a zinc precursor (such as zinc acetate, zinc nitrate, or zinc chloride) with plant or microbial extracts under controlled conditions to form ZnONPs. Plant-mediated synthesis is the most widely used approach due to its simplicity, cost-effectiveness, and scalability ( 1 ). Various parts of plants, such as leaves, roots, stems, flowers, and fruits, contain phytochemicals like flavonoids, alkaloids, tannins, phenols, terpenoids, and proteins that act as reducing and stabilising agents. The phytochemicals facilitate the reduction of zinc salts to ZnO NPs while also providing stability to prevent aggregation. Plant-based synthesis is advantageous because it is rapid, non-toxic, and easily reproducible ( 2 ). Here, in this case, we used Rumax hastatus root aqueous extracts. Rumex hastatus is a medicinal plant easily found in the Himalayan region. Its root contains various secondary metabolites, including terpenoids, Steroids, Flavonoids, tannins, Polyphenolic Compounds, and alkaloid compounds, which serve as good capping and reducing agents in combination with zinc oxide. The various methods used for synthesising Zinc Oxide nanoparticles include sol-gel processing, homogeneous precipitation, organometallic synthesis, spray pyrolysis, thermal evaporation, microwave methods, mechanical milling, and mechanochemical synthesis ( 3 ). However, these methods are usually expensive, labour-intensive, and environmentally unsafe. The presence of certain toxic chemicals used in chemical methods may have hazardous effects in medical applications ( 4 ). Hence, there is a need for green chemistry routes which have emerged as sources of an alternate, low-cost, and eco-friendly nanoparticle production. Zinc Oxide nanoparticles are attracting attention as a promising material due to their wide range of applications in electronics, optics, optoelectronics, biomedicine, and antimicrobial therapies ( 5 – 11 ). Using plant-based extract to synthesize metal oxide is of significant advantage due to the production of various functional molecules from phytochemicals which reduce metal ions zinc oxide nanoparticles which synthesis by plant mediated or green method have significant biomedical properties,( 12 ) making them appropriate for a wide range of biomedical applications for example, previous research has shown that the antibacterial, anti-tumour and antimicrobial activities of ZnO nanoparticles generated using plant extracts are superior to those conventional medication in combating infections ( 13 ). Inorganic nanoparticles, such as silver, gold, copper oxides, and zinc nanoparticles, exhibit a range of profound biological activities. The inorganic NPs and ZnO NPs are particularly interesting because they can be easily prepared and are safe for humans and animals ( 14 ). During normal metabolic processes, the generation of free radicals and the action of antioxidants remain in equilibrium. When free radical production becomes excessive, oxidative stress occurs, leading to cellular damage and contributing to chronic health conditions such as cardiovascular diseases, diabetes, cancer, and inflammation. Consuming antioxidants helps counteract this damage by neutralising free radicals. Hence, antioxidants play a vital role in protecting the body from oxidative stress, supporting overall health, and potentially extending lifespan. Nevertheless, the use of synthetic antioxidants is limited due to concerns about their toxicity. As a result, current research increasingly focuses on natural sources of antioxidants. Among various options, zinc oxide nanoparticles (ZnO NPs) have emerged as a promising class of inorganic oxides, garnering considerable interest due to their advantageous properties and wide range of applications, including those in personal care products. ( 15 ). The zinc oxide nanoparticles not only have biomedical applications but also various significances in industrial sectors, such as the textile sector. Due to their multifunctional properties, zinc oxide nanoparticles (Zn NPs) have gained significant attention. They are commonly applied either as ZnO suspensions or through zinc salt precursors. Owing to their excellent photocatalytic efficiency, chemical stability under ultraviolet (UV) radiation, and high thermal resistance, ZnO NPs have been extensively investigated for textile finishing applications. In particular, they exhibit a broad UV absorption range, making them highly suitable for UV-protective fabric coatings. Their inherent photocatalytic activity also facilitates the degradation of dyes, surfactants, and other organic pollutants in textile effluents, making them promising candidates for wastewater treatment applications ( 16 ). 2. Materials and methods Zinc acetate salt and NaOH are used for synthesis. Distilled water and Whatman filter paper are used. The plant uses Rumax hastatus root extract, which was collected from around the BGR campus, Pauri HNBGU, Latitude 30.132719°Long 78.774266° 2.1 Sample collection and plant identification The plant material ( Rumex hastatus D. Don) was collected from the BGR Campus, Uttarakhand, India (30°13′21.9″ N latitude, 79°47′24.6″ E longitude) at an altitude of 1650 m. The plant belongs to the family Polygonaceae and is commonly found in hilly slope habitats. The plant specimen was collected by Nidhi and taxonomically identified by Dr Anant Kumar, a taxonomist and flora and herbarium expert. A voucher specimen has been prepared and deposited in the Garhwal University Herbarium (GUH), Department of Botany and Microbiology, H.N.B. Garhwal University (A Central University), Uttarakhand, India, under accession number GUH 20865. 2.2 Plant extract After collecting sample wash with distilled water and then take 2, 3 days for shady dry then grind by mixer grinder as powder form then take 20g of powder sample dissolve in 200ml distilled water and take 60–70°C temp in magnetic stirrer for one h then after cool at room temp filter with Whatman filter paper No 1. one collected filtrate part for next step. 2.3 Salt solution and synthesis of nanoparticles Take a 0.05M Zinc acetate salt in 200ml distilled water take 15min in magnetic stirrer salt solution form for synthesis nanoparticles mix salt solution and plant extract 1:1 take in magnetic stirrer for 30min with maintain pH by digital pH Meter with 0.1M NaOH solution after mixing complete and set pH then take in for one day set and aggregate reducing agent of plant part with metal and next day centrifuge 7000 rpm for 20 min and then wash as thrice with distil water then acetone or ethanol removing extra impurity. Then, take the oven-dried nanoparticles and store them in a desiccator with a seal for characterisation and biological activities. This methodology is shown in Fig. 1 3 Physical Characterisations: 3.1 UV-Visible spectroscopy analyser UV-visible spectroscopy identifies unknown compounds with respect to the UV-visible spectrum, which ranges from 190 to 1100 nm. For all analyses, it is essential to measure a reference sample commonly referred to as a blank sample, which typically consists of a cuvette containing the same solvent used to prepare the sample. This procedure ensures that any background signal is accounted for in the results. In this, we use double beam UV-visible spectroscopy, company name is LABTRONICS, model no 2201. 3.2 X-ray powder Diffraction The diffracted X-rays are captured, analysed, and recorded. By scanning the sample across a range of 2θ angles, the random orientation of the powdered material ensures that all possible lattice diffraction directions are covered. This is made with Rigaku Ultima IV (Japan) with a copper target (λ = 1.5414 Å) and a goniometer angular range of 2° to 140°, capable of wide-angle (2°–140°) and small-angle (0.25°–5°) measurements. 3.3 FT-IR spectroscopy FT-IR spectroscopy identifies functional groups from phytochemicals responsible for nanoparticle reduction and stabilisation. Analysis uses powder samples in KBr pellets. PerkinElmer FTIR spectrometers, featuring lithium tantalate MIR detectors and operating from 4000 to 400 cm⁻¹ within a temperature range of 5–45°C, enable precise routine testing. 3.4 FE-SEM Analyser FE-SEM imaging (TESCAN MIRA 3 LMH) scans samples with electrons in a zigzag pattern, providing high-resolution surface morphology and chemical composition images at magnifications up to 1,000,000x. 3.6 Differential Scanning Calorimetry Analyser Differential Scanning Calorimetry (TA Instruments DSC25) measures heat flow during phase transitions over − 80 to 375°C with high temperature precision, analysing solids, liquids, and powders. 3.7 DLS/Zeta potentials analyser Dynamic Light Scattering and Zeta Potential (Brookhaven Zeta Plus) quickly assess nanoparticle size (0.3 nm–3 µm) and stability in various liquids, with high repeatability and mobility range of 10⁻¹¹ to 10⁻⁷ m²/V·s. 3.8 Biological applications Antimicrobial activities The antimicrobial efficacy of the samples was tested against Salmonella typhi (MTCC 733), Escherichia coli (MTCC 452), Trichoderma reesei (MTCC 164), and Aspergillus niger(MTCC 282) using the well diffusion method. Mueller Hinton Agar (38 g/L) and Savoured Dextrose Agar (65 g/L) were prepared, sterilised, and poured into sterile petri dishes. After solidification, microbial inoculum was evenly spread, and wells were created to introduce 50 µL of samples and controls (ciprofloxacin for bacteria, itraconazole for fungi, and methanol as a negative control). Plates were incubated at 37°C for bacteria and 25°C for fungi for 24 hours. Antimicrobial activity was determined by measuring the diameter of inhibition zones. Antioxidant activities Antioxidant activity was evaluated via the 1-diphenyl-2-picrylhydrazyl DPPH radical scavenging assay. Samples (1 mg/mL in methanol) at concentrations ranging from 62.5 to 1000 µg/mL were mixed with DPPH solution and incubated in the dark for 30 minutes at 20°C. Absorbance was measured at 517 nm, and percentage radical scavenging activity (%RSA) was calculated relative to the control. The IC50 values were derived from the dose-response curve. 4 Result and Discussion 4.1 UV-Visible Spectroscopy UV-visible spectroscopy analysis of ZnONPs synthesised from Rumax hastatus root shows an absorption band at 348 nm, as seen in Fig. 2 (a). This indicates successful formation of ZnONPs. The broad peak, ranging from 320 nm to 400 nm, further confirms the presence of zinc oxide nanoparticles ( 18 ). In Fig. 2 (b) , which presents a comparative analysis of the UV-visible spectra of a zinc salt solution, Rumax root aqueous extract, and ZnONPs, distinct peaks are clearly visible, indicating the formation of nanoparticles. 4.2 X-ray diffraction The Zinc oxide nanoparticles phase crystallinity verified their XRD pattern in this following Fig. (3) in which show Hexagonal wurtzite structure the Diffraction peaks at 31.6°,34.5°,36.5°,47.6°,56.9°,62.6°and 68.7° which set by Origin pro and their lattice pattern miller indices h k l are respectively are (100),(002),(101),(102),(110),(103)and(112) in these analysis confirm ZnONPs is hexagonal (wurtzite) structure based on 2θ 31.6° hkl (100) θ°is 15.8° d(Ǻ) 2.83, a(Ǻ)3.25,c(Ǻ)5.21, which is calculated by adding the high score. The ZnO nanoparticles exhibited strong crystallinity, indicated by pronounced peak intensities in the XRD pattern. The crystallite size (D)( 19 ) was estimated using the Scherrer equation: $$\:D=\frac{K\lambda\:}{\beta\:\text{c}\text{o}\text{s}\theta\:}$$ where \(\:D\) Is the crystallite size in nanometres, \(\:K\) Is the Scherrer constant, \(\:\lambda\:\) represents the X-ray wavelength, \(\:\beta\:\) Is the full width at half maximum (FWHM) of the diffraction peak, and \(\:\theta\:\) is the diffraction angle. ( 20 ) The calculated crystallite size ranged between 22 and 32 nm. 4.3 FTIR analysis Fourier transform infrared (FTIR) spectroscopy was employed to identify the biomolecules in Rumex hastatus root extract involved in the reduction and stabilisation of ZnO nanoparticles. The range from 400 to 600cm⁻¹ indicated metal–oxygen bonds. In this study, two sharp peaks at 631cm⁻¹ and 604 cm⁻¹ were observed, indicating that metal oxide nanoparticles had formed, as shown in Fig. 4 (b). Additionally, a broad absorption band was observed between 3000 cm⁻¹ and 3600 cm⁻¹, attributed to O–H stretching from surface hydroxyl groups and residual organic compounds. ( 21 ) Peaks around 2883 cm⁻¹, 1496cm⁻¹, and 1394 cm⁻¹ were assigned to -C-H stretching, C = C stretching and C-H bending vibrations, respectively. One peak is observed at 842 cm⁻¹, indicating an Aromatic derivative. These functional groups suggest the attachment of phytochemical compounds acting as capping and stabilising agents, thereby preventing nanoparticle agglomeration. ( 22 ) which is shown in Fig. 4 (a). 4.4 FE-SEM Analysis FE-SEM analysis data were used to analyse the information about the morphology of nanoparticles in this study. The following data clearly show a distinct formation of nanoparticles with a flower-like structure, as seen in the high-resolution images. ( 23 ). The nanoparticles were observed to be predominantly spherical to quasi-spherical in shape, with an estimated average size of 20–30nm. The observed aggregation at lower magnifications suggests a tendency for nanoparticles to form clusters, which is typical for materials synthesised via chemical or green methods due to van der Waals interactions and surface energy effects. Nevertheless, individual nanoparticles within agglomerates could be resolved, confirming the nanoscale dimensions. Therefore, SEM analysis confirms the successful formation of nanoparticles with an average size of ~ 20–30 nm, which is consistent with the expected nanoscale morphology for this class of materials. Which shows the following Fig. 5 . 4.5 Differential Scanning Calorimetry Differential scanning calorimetry (DSC) curve of ZnONPs recorded from 25 to 350°C at a heating rate of 10°C/min. The initial endothermic transition below 100°C corresponds to the evaporation of surface-adsorbed moisture. ( 24 ). A sharp endothermic peak near 110°C is attributed to the removal of hydroxyl groups and volatile residues. A broad exothermic event between 150–250°C indicates the decomposition of residual organic capping agents and possible crystallisation of ZnONPs. Beyond 250°C, no significant transitions are observed, confirming the high thermal stability of the ZnONPs, shown in Fig. 6 . 4.6 DLS Analysis and zeta potential Dynamic Light Scattering (DLS) measurements of the samples. The cumulative distribution profiles, which reach 100% at higher size ranges, confirm the predominance of large agglomerates over well-dispersed nanoscale entities. Such results suggest poor colloidal stability and highlight the likelihood of particle agglomeration driven by insufficient electrostatic or steric stabilisation. Diameter 1,133.88 nm, polydispersity 0.072, baseline index 3.4, count rate 401.2 96.85, diffusion coefficient 4.328e-09. Based on this data of DLS, particle size and diameter are very large as comparative to SEM data, which means particle agglomeration is large ( 25 ) (Fig. 7 a ). 4.7 Zeta Potential Analysis The zeta potential of the synthesised sample Rumax hastatus Root was measured using phase analysis light scattering (PALS) in triplicate. The obtained values ranged from − 4.59 mV to − 8.51 mV, with an average potential of − 6.40 mV (± 1.98 mV, SD). The corresponding electrophoretic mobility values ranged from − 0.36 to − 0.67 (µ/s)/(V/cm), with a mean value of − 0.50 ± 0.15 (SD). The RMS residual values ranged between 0.050 and 0.071, with a mean of 0.0586, indicating a good fit of the experimental data. According to colloidal stability criteria, dispersions with zeta potentials less than ± 10 mV are generally considered unstable due to insufficient electrostatic repulsion, which increases the likelihood of particle aggregation. In contrast, values above ± 30 mV usually indicate moderate to high stability, which is more significant than the 60mV ( 26 ). Based on these criteria, the measured zeta potential of the Rumax hastatus roots sample (–6.40 mV) suggests low surface charge and poor electrostatic stabilisation, implying that the nanoparticles may exhibit aggregation tendencies in aqueous suspension( 27 ) Fig. 7 (b). Table 1 Zeta potential comparative data with the previous study. Zeta Potential Range (mV) Stability Interpretation references 0 to ± 10 Highly unstable (rapid aggregation) ± 10 to ± 30 Relatively stable (limited aggregation) ( 27 ) > ±30 Moderately stable > ±60 Highly stable (strong repulsion) ( 26 ) Table 1 shows a mean zeta potential of − 6.40 mV, the Rumax hastatus roots sample falls within the 0 to ± 10 mV range, indicating highly unstable dispersions and a strong tendency toward aggregation. 4.8 Anti-microbial Activities In these studies, antimicrobial activities of Rumax hastatus root were evaluated using the well diffusion method. Four pathogens were tested: two bacteria Salmonella Typhi (S. typhi) MTCC733 and Escherichia coli (E. coli) MTCC452 and two fungi Trichoderma reesei (T. reesei) MTCC164 and Aspergillus niger (A. niger) MTCC282. For antibacterial tests, Ciprofloxacin served as the positive control and methanol as the negative control. For antifungal tests, itraconazole served as the positive control, and methanol as the negative control. The samples were prepared at various concentrations and applied to agar plates inoculated with the respective pathogens. All test plates were incubated. After incubation, zones of inhibition were measured to assess antimicrobial activity, using different concentrations: 100mg/mL, 50mg/mL, 25mg/mL, gram-positive and gram-negative as shown in Fig. 9 . Anti-fungal, and Fig. 10 anti-bacterial in which the plate mention clearly mentions C1, C2, C3 as above concentration respectively and V+, V- for gram positive and gram negative respectively. Interaction between Zinc oxide nanoparticles (ZnONPs) and microbial cells likely occurs at the plasma membrane, resulting in inhibitory effects ( 28 ). Inhibition rates ranged from 8.5% to 36.33%, with the highest observed against S. typhi, which is clearly shown in Fig. 8 , and Table 2 presents the detailed inhibition rates for each pathogen with respect to different concentrations and Gram-positive and Gram-negative( 29 ) In the case of following Fig. 11 , which illustrates how metallic nanoparticles interact with a bacterial cell and exhibit their own activities, the mechanism is depicted in Fig. 11 , copied from the internet. Table 2 Antimicrobial activity inhibition against four different pathogens with respect to different concentrations. Microorganism Con.100mg/ml Con.50mg/ml Con.25mg/ml Gram positive Gram Neg. E. coli 14.66 ± 0.577 13.16 ± 0.28 11.83 ± 0.76 24.66 ± 1.04 Nil S. typhi 24.16 ± 2.020 20.33 ± 0.57 19.33 ± 1.52 36.33 ± 0.57 Nil T. reesei 8.5 ± 0.86 Nil Nil 13.83 ± 1.04 Nil A. niger 13.00 ± 1.5 11.83 ± 1.04 10.66 ± 0.57 12.83 ± 0.28 Nil 4.8 Anti-oxidant activity The DPPH assay results indicate that the RHR extract exhibits a significantly weaker antioxidant potential than ascorbic acid, as evidenced by its higher IC50 value. An IC-50 value greater than 400 µg/ml suggests only moderate capability in scavenging free radicals when compared to potent standards, such as ascorbic acid, which demonstrates an IC-50 below 20 µg/ml. The observed trend of decreasing radical scavenging activity with lowering concentrations is in agreement with established antioxidant behaviours. For comparison, the standard antioxidant ascorbic acid exhibited an IC50 of 18.09 ± 0.51 µg/mL, confirming its stronger radical-quenching ability at lower concentrations. ( 30 ) The observed antioxidant activity of RHR can be attributed to the phytochemical constituents present on the nanoparticle surface, which effectively donate electrons or hydrogen atoms to neutralise DPPH radicals. ( 31 ). These findings align with literature reports indicating that natural extracts often display weaker antioxidant activities compared to pure compounds such as ascorbic acid ( 32 – 33 ). This difference could be attributed to the complex composition of plant extracts, the presence of synergistic or antagonistic constituents, and the extraction efficiency. ( 32 – 33 ) Although RHR displayed relatively moderate antioxidant efficiency compared to pure ascorbic acid, these results demonstrate that the biosynthesised ZnONPs possess significant radical scavenging capability. This property supports their potential use in biomedical and food preservation applications where mitigation of oxidative stress is critical. Table 3 Observed Ascorbic acid equivalent antioxidant capacity by the DPPH method of the sample and the standard. S. No Test Sample Code Observation 1 DPPH IC-50 (ug/ml) Ascorbic acid (Standard) 18.09 ± 0.51 2 DPPH IC-50 (ug/ml) RHR 422.25 ± 2.61 5. Conclusion In this research, zinc oxide nanoparticles (ZnONPs) were successfully synthesised using Rumex hastatus root extract through a green synthesis approach. The nanoparticles were characterised using ultraviolet-visible spectroscopy (UV-Vis), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), and field emission scanning electron microscopy (FE-SEM). The UV-Vis spectrum exhibited an absorption peak at 348 nm, indicating the formation of nanoparticles. XRD analysis confirmed the hexagonal wurtzite crystal structure, with crystallite sizes ranging from 20 to 25 nm as determined by the Scherrer equation. Thermal behaviour and stability assessments were performed using differential scanning calorimetry (DSC), dynamic light scattering (DLS), and zeta potential measurements, which also reflected interactions with phytochemicals present in the root extract. Biologically, the synthesised ZnONPs showed antimicrobial efficacy against four pathogenic strains, demonstrating notable inhibition against Salmonella typhi and Gram-positive bacteria, along with moderate antioxidant properties. These findings underscore the eco-friendly and potentially safe nature of biogenic ZnONPs, supporting their future application as antimicrobial agents against drug-resistant pathogens. Abbreviations ZnONPs- Zinc oxide nanoparticles RHR - Rumex hastatus roots FE-SEM- Field Emission Scanning Electron Microscopy XRD- X-ray diffraction FT-IR – Fourier Transform Infrared Spectroscopy ZP- Zeta potential DLS- dynamic light scattering DSC-Differential Scanning Calorimetry R&D- Research and Development Declarations Data availability statement The data generated during this study are available from the authors upon reasonable request. Funding sources This research was conducted without any funding or financial support from institutions or organisations. Declaration of Competing Interest. The authors affirm that they have no financial conflicts or personal connections that might have influenced the research presented in this paper. Author Contribution Declaration The first author and corresponding author (Nidhi) wrote the manuscript and interpreted all the data The Second author (Pragti Siani) reviewed the manuscript The third (Anshul Gairola) and fourth (M.C Purohit) authors reviewed the Manuscript. Consent to Publish declaration Not applicable Ethics and Consent to Participate declarations The plant material (Rumex hastatus D. Don) used in this study was collected from the BGR Campus, Uttarakhand, India. The plant was taxonomically identified by Dr Anant Kumar, a Taxonomist, Flora and Herbarium expert, and a voucher specimen (Accession No. GUH 20865) has been deposited in the Garhwal University Herbarium (GUH), on 12 March 2025, Department of Botany and Microbiology, H.N.B. Garhwal University (A Central University), Uttarakhand, India, for future reference. The collection of plant material was carried out in accordance with applicable local and national guidelines. The plant was collected from a natural (wild) habitat, and no endangered or protected species were involved in this study. All procedures complied with institutional and national regulations for plant research. Acknowledgement The authors express their gratitude to the Department of Chemistry, HNBGU, BGR Campus, Pauri, for providing laboratory support, and the plant identified by Dr Anant kumar who is a taxonomist of Garhwal University Herbarium, Uttarakhand Department of Botany, HNBGU, for authentication of plants, IIT Roorkee for instrumental characterisation, and R&D Cytogeny Company for assistance with biological testing. References Ahmed S., Annu, Chaudhry S.A., Ikram S.: A review on biogenic synthesis of ZnO nanoparticles using plant extracts and microbes: A prospect towards green chemistry. 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New J Chem 41:10347-10356(2017) Jamdagni P., Khatri P., Rana J.S.: Green synthesis of zinc oxide nanoparticles using flower extract of Nyctanthes arbor-tristis and their antifungal activity. J King Saud Univ Sci 30:168-175(2018) Datta A., Patra C., Bharadwaj H., Kaur S., Dimri N, Khajuria R.: Green Synthesis of Zinc Oxide Nanoparticles Using Parthenium hysterophorus Leaf Extract and Evaluation of their Antibacterial Properties. J Biotechnol Biomater 7(2017) Kedare S.B., Singh R.P.: Genesis and Development of DPPH Method of Antioxidant Assay. J Food Sci Technol 48:412-422(2011) Baliyan S., Mukherjee R., Priyadarshini A., Vibhuti A., Gupta A., Pandey RP., Chang CM.: Determination of Antioxidants by DPPH Radical Scavenging Activity and Quantitative Phytochemical Analysis of Ficus religiosa. Molecules 27:1326(2022) Parvin M.N.: Evaluation of Phytochemical Screening and DPPH Free-Radical Scavenging Activity of Leaves Extract of Pouzolzia zeylanica (L.) Benn (Family: Urticaceae). J Pharmacognosy Phytochem 14:578-582(2025) Flieger J., Flieger W., Baj J., Maciejewski R.: Antioxidants: Classification, Natural Sources, Activity/Capacity Measurements, and Usefulness for the Synthesis of Nanoparticles. Materials 14(2021) Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Reviewers agreed at journal 28 Apr, 2026 Reviewers agreed at journal 31 Mar, 2026 Reviewers invited by journal 31 Mar, 2026 Editor assigned by journal 24 Mar, 2026 Submission checks completed at journal 20 Mar, 2026 First submitted to journal 20 Mar, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9010794","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":615604695,"identity":"9339f438-550b-4705-a0dd-7df40e70b891","order_by":0,"name":"Nidhi .","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABAElEQVRIiWNgGAWjYBACA2YGNiDJwMNwg7HhwIcKG6AYY+MBgloOQLQ0PpxxJg2kpQG/FgaQFhDrBgOzMW/LYbAoXi3m7OzPHn8ouCPDd7u5TXJmw3m7te2HgbbU2ETj0mLZzGNucMDgGY/knYNtEh933E7ediYRqOVYWm4DLocd5mGTOAAkDW4kAm05czvZ7ABQC2PDYTxa2J/BtUjztp1LNjv/kJAWBjOYlmZj3rYDdmY3CNgC9IuZxBmgFskbiaBATk4wuwG0JQGPX8z5jz+TqPhz2J7vRvoDYFTa2ZudT3/44EONDU4tGCARrDKBWOUgYE+K4lEwCkbBKBgZAAAqrG0lCXCQ/AAAAABJRU5ErkJggg==","orcid":"","institution":"H.N.B. Garhwal Central University","correspondingAuthor":true,"prefix":"","firstName":"Nidhi","middleName":"","lastName":".","suffix":""},{"id":615604698,"identity":"2d8c0c1f-bafd-4f65-916d-d6bb7bd6e9b6","order_by":1,"name":"Pragti Saini","email":"","orcid":"","institution":"Indian Institute of Technology Roorkee","correspondingAuthor":false,"prefix":"","firstName":"Pragti","middleName":"","lastName":"Saini","suffix":""},{"id":615604699,"identity":"88d031ab-b5a5-45da-b849-d21fb3522e77","order_by":2,"name":"Anshul Gairola","email":"","orcid":"","institution":"Hemwati Nandan Bahuguna Garhwal University","correspondingAuthor":false,"prefix":"","firstName":"Anshul","middleName":"","lastName":"Gairola","suffix":""},{"id":615604701,"identity":"f0329271-ff91-46e2-ade8-ce586d81047a","order_by":3,"name":"M. C. Purohit","email":"","orcid":"","institution":"H.N.B. Garhwal Central University","correspondingAuthor":false,"prefix":"","firstName":"M.","middleName":"C.","lastName":"Purohit","suffix":""}],"badges":[],"createdAt":"2026-03-02 13:38:29","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-9010794/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9010794/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":106402517,"identity":"f6dfa5de-fef9-4bfb-b39e-4ca6b670beb2","added_by":"auto","created_at":"2026-04-08 09:12:12","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":295221,"visible":true,"origin":"","legend":"\u003cp\u003eMethodology of synthesis of zinc oxide nanoparticles.\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/0f41e2650bf0e18fdc8b4db9.jpeg"},{"id":106245783,"identity":"45223d77-c97b-45d4-adf2-ad130bcc7226","added_by":"auto","created_at":"2026-04-06 15:56:33","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":101710,"visible":true,"origin":"","legend":"\u003cp\u003e(a) UV-visible spectra of ZnONPs (b) comparative analysis of zinc salts, plant extract and nanoparticles\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/0ef89d15218f300ef1df2acd.jpeg"},{"id":106245748,"identity":"88f4b70f-7b48-459e-aa11-e8c7bd41a406","added_by":"auto","created_at":"2026-04-06 15:56:19","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":107052,"visible":true,"origin":"","legend":"\u003cp\u003eX-ray Diffraction: all peaks shown with respect to Miller indices.\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/8e28388f53634d3679e58f17.jpeg"},{"id":106245761,"identity":"f8d8eb0f-4e0f-4c99-981f-d265cfe9c896","added_by":"auto","created_at":"2026-04-06 15:56:26","extension":"jpeg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":122750,"visible":true,"origin":"","legend":"\u003cp\u003e(a) FTIR data of transmittance, which spans 400-4000 cm-1. (b) shows the metal oxide functional group.\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/a2e5914bbee7cb4d2fd44ff0.jpeg"},{"id":106245766,"identity":"fd12dc0f-a3ae-4fa0-ae5b-84e61e3574e7","added_by":"auto","created_at":"2026-04-06 15:56:28","extension":"jpeg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":337465,"visible":true,"origin":"","legend":"\u003cp\u003eFE SEM image(A) In 5000nm (B) 2000nm, and (C) 500nm (D) is clearly show particle size.\u003c/p\u003e","description":"","filename":"floatimage5.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/729e1a9889e384b20cce8329.jpeg"},{"id":106245745,"identity":"656a38d5-94b0-467d-995a-3b1dc097ceb4","added_by":"auto","created_at":"2026-04-06 15:56:19","extension":"jpeg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":673027,"visible":true,"origin":"","legend":"\u003cp\u003eDSC of nanoparticles showing exothermic and endothermic temperature and stability.\u003c/p\u003e","description":"","filename":"floatimage6.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/7fb77a9a0ef2f48c88d90e59.jpeg"},{"id":106245763,"identity":"107e4ffc-25dc-4e47-b630-dfb4e0916462","added_by":"auto","created_at":"2026-04-06 15:56:28","extension":"jpeg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":74260,"visible":true,"origin":"","legend":"\u003cp\u003e(a) is the DLS, and (b) is the zeta potential of ZnONPs\u003c/p\u003e","description":"","filename":"floatimage7.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/23d361612702f68de485ccdc.jpeg"},{"id":106245816,"identity":"ce1a1b09-b539-4c49-af60-88bd5e815a13","added_by":"auto","created_at":"2026-04-06 15:56:36","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":60536,"visible":true,"origin":"","legend":"\u003cp\u003eInhibition % of using pathogen in anti-microbial activities with respect to different concentrations and gram-positive and gram-negative bacteria.\u003c/p\u003e","description":"","filename":"floatimage8.png","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/3c67fb189ec38e9cfebec327.png"},{"id":106245818,"identity":"f96cc1cd-98e4-4e6a-856f-0b7d385c7b7f","added_by":"auto","created_at":"2026-04-06 15:56:36","extension":"jpeg","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":581455,"visible":true,"origin":"","legend":"\u003cp\u003eTesting plate dorsal and ventral apprences of Anti-fungal Aspergillus.nigar, Trichoderma nesseri.\u003c/p\u003e","description":"","filename":"floatimage9.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/b05476ca4939943c883dacb4.jpeg"},{"id":106245779,"identity":"8a0aa0b3-f7ff-426c-8671-6eec820d2c30","added_by":"auto","created_at":"2026-04-06 15:56:32","extension":"jpeg","order_by":10,"title":"Figure 10","display":"","copyAsset":false,"role":"figure","size":429482,"visible":true,"origin":"","legend":"\u003cp\u003eTesting plate dorsal and ventral apprences of anti-bacterial Salmonella typhi and Escherichia coli.\u003c/p\u003e","description":"","filename":"floatimage10.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/09043fe59f71c505fed9a9d0.jpeg"},{"id":106245777,"identity":"bd385aef-74bb-4df8-b3ab-8e3bac9876b0","added_by":"auto","created_at":"2026-04-06 15:56:32","extension":"png","order_by":11,"title":"Figure 11","display":"","copyAsset":false,"role":"figure","size":171698,"visible":true,"origin":"","legend":"\u003cp\u003eMechanism of anti-microbial activities of Zinc oxide nanoparticles\u003c/p\u003e","description":"","filename":"floatimage11.png","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/8fd2d1582f6e854b07b79537.png"},{"id":106245765,"identity":"792dca9b-713b-487a-876b-68f488bd4da7","added_by":"auto","created_at":"2026-04-06 15:56:28","extension":"jpeg","order_by":12,"title":"Figure 12","display":"","copyAsset":false,"role":"figure","size":180876,"visible":true,"origin":"","legend":"\u003cp\u003e(a) Ascorbic acid, (b) Rumax hastatus roots, and (c) is the combined data of ascorbic acid and zinc oxide nanoparticles RHR% % RAS with respect to different concentrations.\u003c/p\u003e","description":"","filename":"floatimage12.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/866923826284cbea33be94e8.jpeg"},{"id":106406686,"identity":"2f0a4833-3108-4332-90d8-1d47ab675050","added_by":"auto","created_at":"2026-04-08 09:33:34","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4184678,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9010794/v1/02310ff3-73b6-4a0e-a88e-11fbff30bd42.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Evaluating the Environmental and Biomedical Advantages of Green Synthesised Zinc Oxide Nanoparticles Using an Aqueous Extract of Rumex hastatus Roots","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eThe biosynthesis of ZnO nanoparticles relies on natural reducing and stabilising agents derived from biological resources, such as plant extracts, microorganisms (bacteria, fungi, algae), and biomolecules (proteins, amino acids, polysaccharides). These natural agents serve as capping and stabilising agents, avoiding the use of harmful chemicals. The process generally involves the reaction of a zinc precursor (such as zinc acetate, zinc nitrate, or zinc chloride) with plant or microbial extracts under controlled conditions to form ZnONPs. Plant-mediated synthesis is the most widely used approach due to its simplicity, cost-effectiveness, and scalability (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). Various parts of plants, such as leaves, roots, stems, flowers, and fruits, contain phytochemicals like flavonoids, alkaloids, tannins, phenols, terpenoids, and proteins that act as reducing and stabilising agents. The phytochemicals facilitate the reduction of zinc salts to ZnO NPs while also providing stability to prevent aggregation. Plant-based synthesis is advantageous because it is rapid, non-toxic, and easily reproducible (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e). Here, in this case, we used Rumax \u003cem\u003ehastatus\u003c/em\u003e root aqueous extracts. \u003cem\u003eRumex hastatus\u003c/em\u003e is a medicinal plant easily found in the Himalayan region. Its root contains various secondary metabolites, including terpenoids, Steroids, Flavonoids, tannins, Polyphenolic Compounds, and alkaloid compounds, which serve as good capping and reducing agents in combination with zinc oxide. The various methods used for synthesising Zinc Oxide nanoparticles include sol-gel processing, homogeneous precipitation, organometallic synthesis, spray pyrolysis, thermal evaporation, microwave methods, mechanical milling, and mechanochemical synthesis (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). However, these methods are usually expensive, labour-intensive, and environmentally unsafe. The presence of certain toxic chemicals used in chemical methods may have hazardous effects in medical applications (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). Hence, there is a need for green chemistry routes which have emerged as sources of an alternate, low-cost, and eco-friendly nanoparticle production. Zinc Oxide nanoparticles are attracting attention as a promising material due to their wide range of applications in electronics, optics, optoelectronics, biomedicine, and antimicrobial therapies (\u003cspan additionalcitationids=\"CR6 CR7 CR8 CR9 CR10\" citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e). Using plant-based extract to synthesize metal oxide is of significant advantage due to the production of various functional molecules from phytochemicals which reduce metal ions zinc oxide nanoparticles which synthesis by plant mediated or green method have significant biomedical properties,(\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e) making them appropriate for a wide range of biomedical applications for example, previous research has shown that the antibacterial, anti-tumour and antimicrobial activities of ZnO nanoparticles generated using plant extracts are superior to those conventional medication in combating infections (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e). Inorganic nanoparticles, such as silver, gold, copper oxides, and zinc nanoparticles, exhibit a range of profound biological activities. The inorganic NPs and ZnO NPs are particularly interesting because they can be easily prepared and are safe for humans and animals (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e). During normal metabolic processes, the generation of free radicals and the action of antioxidants remain in equilibrium. When free radical production becomes excessive, oxidative stress occurs, leading to cellular damage and contributing to chronic health conditions such as cardiovascular diseases, diabetes, cancer, and inflammation. Consuming antioxidants helps counteract this damage by neutralising free radicals. Hence, antioxidants play a vital role in protecting the body from oxidative stress, supporting overall health, and potentially extending lifespan. Nevertheless, the use of synthetic antioxidants is limited due to concerns about their toxicity. As a result, current research increasingly focuses on natural sources of antioxidants. Among various options, zinc oxide nanoparticles (ZnO NPs) have emerged as a promising class of inorganic oxides, garnering considerable interest due to their advantageous properties and wide range of applications, including those in personal care products. (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). The zinc oxide nanoparticles not only have biomedical applications but also various significances in industrial sectors, such as the textile sector. Due to their multifunctional properties, zinc oxide nanoparticles (Zn NPs) have gained significant attention. They are commonly applied either as ZnO suspensions or through zinc salt precursors. Owing to their excellent photocatalytic efficiency, chemical stability under ultraviolet (UV) radiation, and high thermal resistance, ZnO NPs have been extensively investigated for textile finishing applications. In particular, they exhibit a broad UV absorption range, making them highly suitable for UV-protective fabric coatings. Their inherent photocatalytic activity also facilitates the degradation of dyes, surfactants, and other organic pollutants in textile effluents, making them promising candidates for wastewater treatment applications (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e"},{"header":"2. Materials and methods","content":"\u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eZinc acetate salt and NaOH are used for synthesis. Distilled water and Whatman filter paper are used. The plant uses \u003cem\u003eRumax hastatus root\u003c/em\u003e extract, which was collected from around the BGR campus, Pauri HNBGU, Latitude 30.132719\u0026deg;Long 78.774266\u0026deg;\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Sample collection and plant identification\u003c/h2\u003e \u003cp\u003eThe plant material (\u003cem\u003eRumex hastatus\u003c/em\u003e D. Don) was collected from the BGR Campus, Uttarakhand, India (30\u0026deg;13\u0026prime;21.9\u0026Prime; N latitude, 79\u0026deg;47\u0026prime;24.6\u0026Prime; E longitude) at an altitude of 1650 m. The plant belongs to the family Polygonaceae and is commonly found in hilly slope habitats.\u003c/p\u003e \u003cp\u003eThe plant specimen was collected by Nidhi and taxonomically identified by Dr Anant Kumar, a taxonomist and flora and herbarium expert. A voucher specimen has been prepared and deposited in the Garhwal University Herbarium (GUH), Department of Botany and Microbiology, H.N.B. Garhwal University (A Central University), Uttarakhand, India, under accession number GUH 20865.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Plant extract\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eAfter collecting sample wash with distilled water and then take 2, 3 days for shady dry then grind by mixer grinder as powder form then take 20g of powder sample dissolve in 200ml distilled water and take 60\u0026ndash;70\u0026deg;C temp in magnetic stirrer for one h then after cool at room temp filter with Whatman filter paper No 1. one collected filtrate part for next step.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Salt solution and synthesis of nanoparticles\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eTake a 0.05M Zinc acetate salt in 200ml distilled water take 15min in magnetic stirrer salt solution form for synthesis nanoparticles mix salt solution and plant extract 1:1 take in magnetic stirrer for 30min with maintain pH by digital pH Meter with 0.1M NaOH solution after mixing complete and set pH then take in for one day set and aggregate reducing agent of plant part with metal and next day centrifuge 7000 rpm for 20 min and then wash as thrice with distil water then acetone or ethanol removing extra impurity. Then, take the oven-dried nanoparticles and store them in a desiccator with a seal for characterisation and biological activities. This methodology is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"3 Physical Characterisations:","content":"\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e3.1 UV-Visible spectroscopy analyser\u003c/h2\u003e \u003cp\u003eUV-visible spectroscopy identifies unknown compounds with respect to the UV-visible spectrum, which ranges from 190 to 1100 nm. For all analyses, it is essential to measure a reference sample commonly referred to as a blank sample, which typically consists of a cuvette containing the same solvent used to prepare the sample. This procedure ensures that any background signal is accounted for in the results. In this, we use double beam UV-visible spectroscopy, company name is LABTRONICS, model no 2201.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e3.2 X-ray powder Diffraction\u003c/h2\u003e \u003cp\u003eThe diffracted X-rays are captured, analysed, and recorded. By scanning the sample across a range of 2θ angles, the random orientation of the powdered material ensures that all possible lattice diffraction directions are covered. This is made with Rigaku Ultima IV (Japan) with a copper target (λ\u0026thinsp;=\u0026thinsp;1.5414 \u0026Aring;) and a goniometer angular range of 2\u0026deg; to 140\u0026deg;, capable of wide-angle (2\u0026deg;\u0026ndash;140\u0026deg;) and small-angle (0.25\u0026deg;\u0026ndash;5\u0026deg;) measurements.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e3.3 FT-IR spectroscopy\u003c/h2\u003e \u003cp\u003eFT-IR spectroscopy identifies functional groups from phytochemicals responsible for nanoparticle reduction and stabilisation. Analysis uses powder samples in KBr pellets. PerkinElmer FTIR spectrometers, featuring lithium tantalate MIR detectors and operating from 4000 to 400 cm⁻\u0026sup1; within a temperature range of 5\u0026ndash;45\u0026deg;C, enable precise routine testing.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e3.4 FE-SEM Analyser\u003c/h2\u003e \u003cp\u003eFE-SEM imaging (TESCAN MIRA 3 LMH) scans samples with electrons in a zigzag pattern, providing high-resolution surface morphology and chemical composition images at magnifications up to 1,000,000x.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e3.6 Differential Scanning Calorimetry Analyser\u003c/h2\u003e \u003cp\u003eDifferential Scanning Calorimetry (TA Instruments DSC25) measures heat flow during phase transitions over \u0026minus;\u0026thinsp;80 to 375\u0026deg;C with high temperature precision, analysing solids, liquids, and powders.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e3.7 DLS/Zeta potentials analyser\u003c/h2\u003e \u003cp\u003eDynamic Light Scattering and Zeta Potential (Brookhaven Zeta Plus) quickly assess nanoparticle size (0.3 nm\u0026ndash;3 \u0026micro;m) and stability in various liquids, with high repeatability and mobility range of 10⁻\u0026sup1;\u0026sup1; to 10⁻⁷ m\u0026sup2;/V\u0026middot;s.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e3.8 Biological applications\u003c/h2\u003e \u003cp\u003e \u003cb\u003eAntimicrobial activities\u003c/b\u003e \u003c/p\u003e \u003cp\u003eThe antimicrobial efficacy of the samples was tested against Salmonella typhi (MTCC 733), Escherichia coli (MTCC 452), Trichoderma reesei (MTCC 164), and Aspergillus niger(MTCC 282) using the well diffusion method. Mueller Hinton Agar (38 g/L) and Savoured Dextrose Agar (65 g/L) were prepared, sterilised, and poured into sterile petri dishes. After solidification, microbial inoculum was evenly spread, and wells were created to introduce 50 \u0026micro;L of samples and controls (ciprofloxacin for bacteria, itraconazole for fungi, and methanol as a negative control). Plates were incubated at 37\u0026deg;C for bacteria and 25\u0026deg;C for fungi for 24 hours. Antimicrobial activity was determined by measuring the diameter of inhibition zones.\u003c/p\u003e \u003cp\u003e \u003cb\u003eAntioxidant activities\u003c/b\u003e \u003c/p\u003e \u003cp\u003eAntioxidant activity was evaluated via the 1-diphenyl-2-picrylhydrazyl DPPH radical scavenging assay. Samples (1 mg/mL in methanol) at concentrations ranging from 62.5 to 1000 \u0026micro;g/mL were mixed with DPPH solution and incubated in the dark for 30 minutes at 20\u0026deg;C. Absorbance was measured at 517 nm, and percentage radical scavenging activity (%RSA) was calculated relative to the control. The IC50 values were derived from the dose-response curve.\u003c/p\u003e \u003c/div\u003e"},{"header":"4 Result and Discussion","content":"\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e4.1 UV-Visible Spectroscopy\u003c/h2\u003e \u003cp\u003eUV-visible spectroscopy analysis of ZnONPs synthesised from Rumax \u003cem\u003ehastatus\u003c/em\u003e root shows an absorption band at 348 nm, as seen in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e \u003cb\u003e(a).\u003c/b\u003e This indicates successful formation of ZnONPs. The broad peak, ranging from 320 nm to 400 nm, further confirms the presence of zinc oxide nanoparticles (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e). \u003cb\u003eIn\u003c/b\u003e Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e\u003cb\u003e(b)\u003c/b\u003e, which presents a comparative analysis of the UV-visible spectra of a zinc salt solution, Rumax root aqueous extract, and ZnONPs, distinct peaks are clearly visible, indicating the formation of nanoparticles.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e4.2 X-ray diffraction\u003c/h2\u003e \u003cp\u003eThe Zinc oxide nanoparticles phase crystallinity verified their XRD pattern in this following Fig.\u0026nbsp;(3) in which show Hexagonal wurtzite structure the Diffraction peaks at 31.6\u0026deg;,34.5\u0026deg;,36.5\u0026deg;,47.6\u0026deg;,56.9\u0026deg;,62.6\u0026deg;and 68.7\u0026deg; which set by Origin pro and their lattice pattern miller indices h k l are respectively are (100),(002),(101),(102),(110),(103)and(112) in these analysis confirm ZnONPs is hexagonal (wurtzite) structure based on 2θ 31.6\u0026deg; hkl (100) θ\u0026deg;is 15.8\u0026deg; d(Ǻ) 2.83, a(Ǻ)3.25,c(Ǻ)5.21, which is calculated by adding the high score. The ZnO nanoparticles exhibited strong crystallinity, indicated by pronounced peak intensities in the XRD pattern. The crystallite size (D)(\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e) was estimated using the Scherrer equation:\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$\\:D=\\frac{K\\lambda\\:}{\\beta\\:\\text{c}\\text{o}\\text{s}\\theta\\:}$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003ewhere \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:D\\)\u003c/span\u003e\u003c/span\u003e Is the crystallite size in nanometres, \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:K\\)\u003c/span\u003e\u003c/span\u003e Is the Scherrer constant, \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\lambda\\:\\)\u003c/span\u003e\u003c/span\u003e represents the X-ray wavelength, \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\beta\\:\\)\u003c/span\u003e\u003c/span\u003e Is the full width at half maximum (FWHM) of the diffraction peak, and \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\theta\\:\\)\u003c/span\u003e\u003c/span\u003e is the diffraction angle. (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e) The calculated crystallite size ranged between 22 and 32 nm.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e4.3 FTIR analysis\u003c/h2\u003e \u003cp\u003eFourier transform infrared (FTIR) spectroscopy was employed to identify the biomolecules in \u003cem\u003eRumex hastatus\u003c/em\u003e root extract involved in the reduction and stabilisation of ZnO nanoparticles. The range from 400 to 600cm⁻\u0026sup1; indicated metal\u0026ndash;oxygen bonds. In this study, two sharp peaks at 631cm⁻\u0026sup1; and 604 cm⁻\u0026sup1; were observed, indicating that metal oxide nanoparticles had formed, as shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e\u003cb\u003e(b).\u003c/b\u003e Additionally, a broad absorption band was observed between 3000 cm⁻\u0026sup1; and 3600 cm⁻\u0026sup1;, attributed to O\u0026ndash;H stretching from surface hydroxyl groups and residual organic compounds. (\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e) Peaks around 2883 cm⁻\u0026sup1;, 1496cm⁻\u0026sup1;, and 1394 cm⁻\u0026sup1; were assigned to -C-H stretching, C\u0026thinsp;=\u0026thinsp;C stretching and C-H bending vibrations, respectively. One peak is observed at 842 cm⁻\u0026sup1;, indicating an Aromatic derivative. These functional groups suggest the attachment of phytochemical compounds acting as capping and stabilising agents, thereby preventing nanoparticle agglomeration. (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e) which is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e \u003cb\u003e(a).\u003c/b\u003e\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003e4.4 FE-SEM Analysis\u003c/h2\u003e \u003cp\u003eFE-SEM analysis data were used to analyse the information about the morphology of nanoparticles in this study. The following data clearly show a distinct formation of nanoparticles with a flower-like structure, as seen in the high-resolution images. (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e). The nanoparticles were observed to be predominantly spherical to quasi-spherical in shape, with an estimated average size of 20\u0026ndash;30nm. The observed aggregation at lower magnifications suggests a tendency for nanoparticles to form clusters, which is typical for materials synthesised via chemical or green methods due to van der Waals interactions and surface energy effects. Nevertheless, individual nanoparticles within agglomerates could be resolved, confirming the nanoscale dimensions. Therefore, SEM analysis confirms the successful formation of nanoparticles with an average size of ~\u0026thinsp;20\u0026ndash;30 nm, which is consistent with the expected nanoscale morphology for this class of materials. Which shows the following Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003e4.5 Differential Scanning Calorimetry\u003c/h2\u003e \u003cp\u003eDifferential scanning calorimetry (DSC) curve of ZnONPs recorded from 25 to 350\u0026deg;C at a heating rate of 10\u0026deg;C/min. The initial endothermic transition below 100\u0026deg;C corresponds to the evaporation of surface-adsorbed moisture. (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). A sharp endothermic peak near 110\u0026deg;C is attributed to the removal of hydroxyl groups and volatile residues. A broad exothermic event between 150\u0026ndash;250\u0026deg;C indicates the decomposition of residual organic capping agents and possible crystallisation of ZnONPs. Beyond 250\u0026deg;C, no significant transitions are observed, confirming the high thermal stability of the ZnONPs, shown \u003cb\u003ein\u003c/b\u003e Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003e4.6 DLS Analysis and zeta potential\u003c/h2\u003e \u003cp\u003eDynamic Light Scattering (DLS) measurements of the samples. The cumulative distribution profiles, which reach 100% at higher size ranges, confirm the predominance of large agglomerates over well-dispersed nanoscale entities. Such results suggest poor colloidal stability and highlight the likelihood of particle agglomeration driven by insufficient electrostatic or steric stabilisation. Diameter 1,133.88 nm, polydispersity 0.072, baseline index 3.4, count rate 401.2 96.85, diffusion coefficient 4.328e-09. Based on this data of DLS, particle size and diameter are very large as comparative to SEM data, which means particle agglomeration is large (\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e) (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ea\u003cb\u003e).\u003c/b\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec21\" class=\"Section2\"\u003e \u003ch2\u003e4.7 Zeta Potential Analysis\u003c/h2\u003e \u003cp\u003eThe zeta potential of the synthesised sample Rumax \u003cem\u003ehastatus\u003c/em\u003e Root was measured using phase analysis light scattering (PALS) in triplicate. The obtained values ranged from \u0026minus;\u0026thinsp;4.59 mV to \u0026minus;\u0026thinsp;8.51 mV, with an average potential of \u0026minus;\u0026thinsp;6.40 mV (\u0026plusmn;\u0026thinsp;1.98 mV, SD). The corresponding electrophoretic mobility values ranged from \u0026minus;\u0026thinsp;0.36 to \u0026minus;\u0026thinsp;0.67 (\u0026micro;/s)/(V/cm), with a mean value of \u0026minus;\u0026thinsp;0.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15 (SD). The RMS residual values ranged between 0.050 and 0.071, with a mean of 0.0586, indicating a good fit of the experimental data. According to colloidal stability criteria, dispersions with zeta potentials less than \u0026plusmn;\u0026thinsp;10 mV are generally considered unstable due to insufficient electrostatic repulsion, which increases the likelihood of particle aggregation. In contrast, values above \u0026plusmn;\u0026thinsp;30 mV usually indicate moderate to high stability, which is more significant than the 60mV (\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e). Based on these criteria, the measured zeta potential of the Rumax \u003cem\u003ehastatus\u003c/em\u003e roots sample (\u0026ndash;6.40 mV) suggests low surface charge and poor electrostatic stabilisation, implying that the nanoparticles may exhibit aggregation tendencies in aqueous suspension(\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e) Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e\u003cb\u003e(b).\u003c/b\u003e\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eZeta potential comparative data with the previous study.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eZeta Potential Range (mV)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eStability Interpretation\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ereferences\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e0 to \u0026plusmn;\u0026thinsp;10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eHighly unstable (rapid aggregation)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026plusmn;\u0026thinsp;10 to \u0026plusmn;\u0026thinsp;30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eRelatively stable (limited aggregation)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e(\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026gt; \u0026plusmn;30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eModerately stable\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026gt; \u0026plusmn;60\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eHighly stable (strong repulsion)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e(\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eTable\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e shows a mean zeta potential of \u0026minus;\u0026thinsp;6.40 mV, the Rumax hastatus roots sample falls within the 0 to \u0026plusmn;\u0026thinsp;10 mV range, indicating highly unstable dispersions and a strong tendency toward aggregation.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec22\" class=\"Section2\"\u003e \u003ch2\u003e4.8 Anti-microbial Activities\u003c/h2\u003e \u003cp\u003eIn these studies, antimicrobial activities of Rumax hastatus root were evaluated using the well diffusion method. Four pathogens were tested: two bacteria Salmonella Typhi (S. typhi) MTCC733 and Escherichia coli (E. coli) MTCC452 and two fungi Trichoderma reesei (T. reesei) MTCC164 and Aspergillus niger (A. niger) MTCC282. For antibacterial tests, Ciprofloxacin served as the positive control and methanol as the negative control. For antifungal tests, itraconazole served as the positive control, and methanol as the negative control. The samples were prepared at various concentrations and applied to agar plates inoculated with the respective pathogens. All test plates were incubated. After incubation, zones of inhibition were measured to assess antimicrobial activity, using different concentrations: 100mg/mL, 50mg/mL, 25mg/mL, gram-positive and gram-negative as shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003e. Anti-fungal, and Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e10\u003c/span\u003e anti-bacterial in which the plate mention clearly mentions C1, C2, C3 as above concentration respectively and V+, V- for gram positive and gram negative respectively. Interaction between Zinc oxide nanoparticles (ZnONPs) and microbial cells likely occurs at the plasma membrane, resulting in inhibitory effects (\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e). Inhibition rates ranged from 8.5% to 36.33%, with the highest observed against S. typhi, which is clearly shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e, and Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e presents the detailed inhibition rates for each pathogen with respect to different concentrations and Gram-positive and Gram-negative(\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e) In the case of following Fig.\u0026nbsp;\u003cspan refid=\"Fig11\" class=\"InternalRef\"\u003e11\u003c/span\u003e, which illustrates how metallic nanoparticles interact with a bacterial cell and exhibit their own activities, the mechanism is depicted in Fig.\u0026nbsp;\u003cspan refid=\"Fig11\" class=\"InternalRef\"\u003e11\u003c/span\u003e, copied from the internet.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAntimicrobial activity inhibition against four different pathogens with respect to different concentrations.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMicroorganism\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCon.100mg/ml\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCon.50mg/ml\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eCon.25mg/ml\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eGram positive\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eGram Neg.\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eE. coli\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e14.66\u0026thinsp;\u0026plusmn;\u0026thinsp;0.577\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13.16\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e11.83\u0026thinsp;\u0026plusmn;\u0026thinsp;0.76\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e24.66\u0026thinsp;\u0026plusmn;\u0026thinsp;1.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eNil\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eS. typhi\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003e24.16\u0026thinsp;\u0026plusmn;\u0026thinsp;2.020\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e20.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.57\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e19.33\u0026thinsp;\u0026plusmn;\u0026thinsp;1.52\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e36.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.57\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eNil\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT. reesei\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e8.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.86\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eNil\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNil\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e13.83\u0026thinsp;\u0026plusmn;\u0026thinsp;1.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eNil\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eA. niger\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e \u003cp\u003e13.00\u0026thinsp;\u0026plusmn;\u0026thinsp;1.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11.83\u0026thinsp;\u0026plusmn;\u0026thinsp;1.04\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10.66\u0026thinsp;\u0026plusmn;\u0026thinsp;0.57\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c5\"\u003e \u003cp\u003e12.83\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eNil\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec23\" class=\"Section2\"\u003e \u003ch2\u003e4.8 Anti-oxidant activity\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eThe DPPH assay results indicate that the RHR extract exhibits a significantly weaker antioxidant potential than ascorbic acid, as evidenced by its higher IC50 value. An IC-50 value greater than 400 \u0026micro;g/ml suggests only moderate capability in scavenging free radicals when compared to potent standards, such as ascorbic acid, which demonstrates an IC-50 below 20 \u0026micro;g/ml. The observed trend of decreasing radical scavenging activity with lowering concentrations is in agreement with established antioxidant behaviours. For comparison, the standard antioxidant ascorbic acid exhibited an IC50 of 18.09\u0026thinsp;\u0026plusmn;\u0026thinsp;0.51 \u0026micro;g/mL, confirming its stronger radical-quenching ability at lower concentrations. (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e) The observed antioxidant activity of RHR can be attributed to the phytochemical constituents present on the nanoparticle surface, which effectively donate electrons or hydrogen atoms to neutralise DPPH radicals. (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e). These findings align with literature reports indicating that natural extracts often display weaker antioxidant activities compared to pure compounds such as ascorbic acid (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e). This difference could be attributed to the complex composition of plant extracts, the presence of synergistic or antagonistic constituents, and the extraction efficiency. (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e) Although RHR displayed relatively moderate antioxidant efficiency compared to pure ascorbic acid, these results demonstrate that the biosynthesised ZnONPs possess significant radical scavenging capability. This property supports their potential use in biomedical and food preservation applications where mitigation of oxidative stress is critical.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eObserved Ascorbic acid equivalent antioxidant capacity by the DPPH method of the sample and the standard.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS. No\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTest\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSample Code\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eObservation\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eDPPH\u003c/p\u003e \u003cp\u003eIC-50 (ug/ml)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAscorbic acid (Standard)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e18.09\u0026thinsp;\u0026plusmn;\u0026thinsp;0.51\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eDPPH\u003c/p\u003e \u003cp\u003eIC-50 (ug/ml)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eRHR\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e422.25\u0026thinsp;\u0026plusmn;\u0026thinsp;2.61\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"5. Conclusion","content":"\u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eIn this research, zinc oxide nanoparticles (ZnONPs) were successfully synthesised using \u003cem\u003eRumex hastatus\u003c/em\u003e root extract through a green synthesis approach. The nanoparticles were characterised using ultraviolet-visible spectroscopy (UV-Vis), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), and field emission scanning electron microscopy (FE-SEM). The UV-Vis spectrum exhibited an absorption peak at 348 nm, indicating the formation of nanoparticles. XRD analysis confirmed the hexagonal wurtzite crystal structure, with crystallite sizes ranging from 20 to 25 nm as determined by the Scherrer equation. Thermal behaviour and stability assessments were performed using differential scanning calorimetry (DSC), dynamic light scattering (DLS), and zeta potential measurements, which also reflected interactions with phytochemicals present in the root extract. Biologically, the synthesised ZnONPs showed antimicrobial efficacy against four pathogenic strains, demonstrating notable inhibition against Salmonella typhi and Gram-positive bacteria, along with moderate antioxidant properties. These findings underscore the eco-friendly and potentially safe nature of biogenic ZnONPs, supporting their future application as antimicrobial agents against drug-resistant pathogens.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eZnONPs- Zinc oxide nanoparticles\u003c/p\u003e\n\u003cp\u003eRHR\u003cem\u003e- Rumex hastatus\u0026nbsp;\u003c/em\u003eroots\u003c/p\u003e\n\u003cp\u003eFE-SEM-\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eField Emission Scanning Electron Microscopy\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eXRD- X-ray diffraction\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFT-IR \u0026ndash; Fourier Transform Infrared Spectroscopy\u003c/p\u003e\n\u003cp\u003eZP- Zeta potential\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eDLS- dynamic light scattering\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eDSC-Differential Scanning Calorimetry\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eR\u0026amp;D- Research and Development\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eData availability statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data generated during this study are available from the authors upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding sources\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was conducted without any funding or financial support from institutions or organisations.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclaration of Competing Interest.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors affirm that they have no financial conflicts or personal connections that might have influenced the research presented in this paper.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contribution Declaration\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe first author and corresponding author (Nidhi)\u003c/strong\u003e wrote the manuscript and interpreted all the data\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe Second author (Pragti Siani)\u003c/strong\u003e reviewed the manuscript\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003cstrong\u003eThe third (Anshul Gairola) and fourth (M.C Purohit)\u003c/strong\u003e authors reviewed the Manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to Publish declaration\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics and Consent to Participate declarations\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe plant material (Rumex hastatus D. Don) used in this study was collected from the BGR Campus, Uttarakhand, India. The plant was taxonomically identified by Dr Anant Kumar, a Taxonomist, Flora and Herbarium expert, and a voucher specimen (Accession No. GUH 20865) has been deposited in the Garhwal University Herbarium (GUH), on 12 March 2025, Department of Botany and Microbiology, H.N.B. Garhwal University (A Central University), Uttarakhand, India, for future reference.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe collection of plant material was carried out in accordance with applicable local and national guidelines. The plant was collected from a natural (wild) habitat, and no endangered or protected species were involved in this study. All procedures complied with institutional and national regulations for plant research.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;Acknowledgement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;The authors express their gratitude to the Department of Chemistry, HNBGU, BGR Campus, Pauri, for providing laboratory support, and the plant identified by Dr Anant kumar who is a taxonomist of Garhwal University Herbarium, Uttarakhand Department of Botany, HNBGU, for authentication of plants, IIT Roorkee for instrumental characterisation, and R\u0026amp;D Cytogeny Company for assistance with biological testing.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eAhmed S., Annu, Chaudhry S.A., Ikram S.: \u0026nbsp;A review on biogenic synthesis of ZnO nanoparticles using plant extracts and microbes: A prospect towards green chemistry. 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Materials 14(2021)\u0026nbsp;\u003c/li\u003e\n\u003c/ol\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"discover-applied-sciences","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Discover Applied Sciences](https://link.springer.com/journal/42452)","snPcode":"42452","submissionUrl":"https://submission.springernature.com/new-submission/42452/3","title":"Discover Applied Sciences","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Discover Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Biosynthesis of zinc oxide Nanoparticles, characterisations, antioxidants, and antimicrobial activities","lastPublishedDoi":"10.21203/rs.3.rs-9010794/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9010794/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eZinc oxide nanoparticles are highly functional in pharmaceutical and biomedical applications, and plant-mediated nanoparticles which show good biological activities. They can be synthesised by different chemical, physical and biological methods, whereas the green method, or biosynthesis, is environment-friendly and less hazardous. In the present study, zinc oxide Nanoparticles were synthesised using an aqueous (Distilled water) extract of \u003cem\u003eRumex hastatus\u003c/em\u003e roots and 0.05 M zinc acetate dihydrate (Zn(CH₃COO)₂\u0026middot;2H₂O). Maintaining pH with a digital pH meter and using 0.1 M sodium hydroxide (NaOH), the nanoparticles synthesised, confirmed by physical characterisation using Ultraviolet-Visible spectroscopy, showed typical absorption peaks in the range of 320nm to 400nm due to their significant excitation binding energy at room temperature., X-Ray Diffraction analysis (XRD), which confirmed that the particles are crystalline and have a hexagonal wurtzite structure, ZnO-Bonding shown by the Fourier Transform Infrared Spectroscopy (FT-IR), Differential Scanning Calorimetry (DSC), Zeta potential confirmed thermal and other stability of nanoparticles and size(22-30nm) and shape (spherical to quasi spherical) confirmed by the Field Emission Scanning Electron Microscopy (FE-SEM) and Dynamic Light Scattering (DLS), Their biomedical applications include anti-microbial and antioxidant properties. In anti-microbial four pathogens were tested: two bacteria, \u003cem\u003eSalmonella Typhi\u003c/em\u003e (S. typhi) MTCC733 and \u003cem\u003eEscherichia coli\u003c/em\u003e (E. coli) MTCC452 and two fungi, \u003cem\u003eTrichoderma reesei\u003c/em\u003e (T. reesei) MTCC164 and \u003cem\u003eAspergillus niger\u003c/em\u003e (A. niger) MTCC282.in which a 36.33% against Salmonella typhi in gram-positive bacteria, a high inhibition rate and anti-oxidant, and an IC-50 value greater than 400 \u0026micro;g/ml, suggests only moderate capability in scavenging free radicals when compared to potent standards like ascorbic acid, which demonstrates an IC-50 below 20 \u0026micro;g/ml.\u003c/p\u003e","manuscriptTitle":"Evaluating the Environmental and Biomedical Advantages of Green Synthesised Zinc Oxide Nanoparticles Using an Aqueous Extract of Rumex hastatus Roots","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-04-06 15:55:57","doi":"10.21203/rs.3.rs-9010794/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewerAgreed","content":"313577122355654286525760044610871354694","date":"2026-04-28T07:35:46+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"122077986874405387677250891531209414019","date":"2026-04-01T02:55:25+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-04-01T00:04:18+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-03-25T03:03:41+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-03-20T04:35:11+00:00","index":"","fulltext":""},{"type":"submitted","content":"Discover Applied Sciences","date":"2026-03-20T04:16:18+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"discover-applied-sciences","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Discover Applied Sciences](https://link.springer.com/journal/42452)","snPcode":"42452","submissionUrl":"https://submission.springernature.com/new-submission/42452/3","title":"Discover Applied Sciences","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Discover Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"23d20b1b-5141-4871-9d42-76a774861e0f","owner":[],"postedDate":"April 6th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-04-06T15:55:58+00:00","versionOfRecord":[],"versionCreatedAt":"2026-04-06 15:55:57","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9010794","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9010794","identity":"rs-9010794","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}