Calcined low-density silica nanoparticles for HRP immobilization: catalytic dye removal and hydrogen peroxide detection in milk

Calcined low-density silica nanoparticles for HRP immobilization: catalytic dye removal and hydrogen peroxide detection in milk | 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 Calcined low-density silica nanoparticles for HRP immobilization: catalytic dye removal and hydrogen peroxide detection in milk Viswanathan kaliyaperumal, Radha perumal ramasamy, Rathinam Ganesan, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9063139/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract In this study, low density silica nanoparticles (LDS) were synthesized and used as a heterogeneous catalyst for dye removal and nanosensors for hydrogen peroxide detection. Organosilane silica nanoparticles were produced and calcinated at 400°C to produce low density particles. SEM, FTIR, DLS, and zeta potential analysis were used to characterize the generated particles. The results confirmed that the particle size, charge, and density varied with the APTES concentration. The results showed that at lower APTES concentrations, small particles (70–75 nm) were created, but at higher concentrations, larger particles (420–430 nm) were produced. The particle density changed depending on the APTES content. The UV absorption spectra of APTES-added samples varied significantly between 440 and 480 nm. The dye removal activity was measured using crystal violet dye; under UV irradiation, the particles destroyed up to 90% of the dye within 120 minutes, and the kinetics were also detailed. HRP-coated LDS was used as a nanosensor for detecting HO. Results indicated linearity of 5⋅10 ~ 1⋅10(M) (R = 0.995), and a detection limit of 5 nm mol. The milk was spiked with HO at varied concentrations and used as detections for the actual sample analysis. Finally, this paper describes the simplest way for producing tailored low-density silica, which is ideal for biomedical and drug delivery applications. low density silica Stobber method hydrogen peroxide crystal violet kinetics Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1. Introduction Silica nanoparticles are an important type of nanotool used in a variety of applications due to their ease of manufacture, high chemical stability, easy surface modification, biocompatibility, and use as a tool to control drug molecules [ 1 – 3 ]. Currently, three types of silica nanoparticles have been created: solid, nonporous, and mesoporous [ 4 – 6 ]. Several methods for synthesizing silica nanoparticles have been recorded, including plasma manufacturing, chemical vapor deposition, microemulsion synthesis, combustion processing, sol-gel synthesis, and hydrothermal reactions [ 7 ]. Recently, one-, two-, and three-dimensional silica nanoparticles, Silica nanotubes, hollow silica nanotubes, and silica nanofibers have been described [ 8 – 10 ]. Currently, silica nanoparticles are used in a variety of applications, including catalysis, drug delivery, biomedicine, environmental remediation, and wastewater treatment [ 11 – 14 ]. In biological applications, surface charge is significant, and cationic or anionic charge-based elimination was shown in animal models. Silica nano particles smaller than 6 nm are quickly removed by urine, but those larger than this accumulates in the liver [ 15 ]. Organic silica has recently been discovered to be a highly effective therapeutic agent for the treatment of bone and skin diseases. The small size and positive surface charge of organic silica-based nanoparticles make them an attractive platform for medication and gene therapy, where a negatively charged membrane improves cellular uptake [ 16 – 17 ]. These materials are also biocompatible and contain low toxicity. Silica nanoparticles were easily combined with iron oxide, quantum dots, fluorescent dye, gold, and other metal nanoparticles to form hybrid nanoparticles [ 18 – 21 ]. In this work, we developed low density silica particles based on organ silane modification and calcinations. The catalytic activity was examined using crystal violet, and the nano sensors application for hydrogen peroxide detection were made utilizing the nanoparticles. 2. Experimental procedure 2.1. Production of organosilane-modified silica nanoparticles Organosilane-modified silica nanoparticles were produced using slightly modified procedures from the previously disclosed simplified stober approach [ 22 ]. To summarize, 10 mL of ethanol, 1 mL of tetraethyl orthosilicate, 0.2 mL of (3-Aminopropyl)-triethoxysilane (APTES), and 1 mL of ammonium hydroxide solution were added. This solution was agitated for an additional 3 hours at room temperature. The particles were collected by centrifugation at 6000 rpm for 10 min, then washed with ethanol and dried in a vacuum oven at 37°C for 2 hours. 2.2. Photo catalytic activity studies Photocatalytic activity was evaluated using previously published data [ 23 ]. For this work, 5 mL of 10 mg/L crystal violet was combined with 500 mg of nanoparticles at room temperature (C), and another set of samples was exposed to a lamp (mercury vapour lamp, 40 W, 40 cm long, wavelength 253.7 nm). The sample was placed in a 10-mL tube. At regular intervals, the emission spectra were examined. The dye degradation rate was computed as C t /C 0 . The initial concentration is C 0 , the reaction concentration is C t , and the graph shows C t /C 0 vs irradiation time. 2.3. Enzyme adsorption and Assay : For horse radish peroxidase adsorption, 130 mg of organo silica nanoparticles were placed in a glass vial, followed by 1 mL of 5 mg/mL horse radish peroxidase solutions, which were incubated for 60 minutes while stirring. Following that, the absorbed enzyme activity on nanoparticles was investigated using o-phenylenediamine (OPD). Briefly, 0.2 mL of 5mg/mL enzyme-coated organosilane silica nanoparticles were placed in an eppendorf tube. The tubes were then centrifuged at 6000 rpm for 5 minutes to separate the particles. Then, 0.1 mL of o-phenylenediamine and 0.1 mL of serially diluted hydrogen peroxide were added. The mixture was gently agitated for 20 minutes before adding 20 µL of pure sulphuric acid to terminate the enzyme reaction. The tube was then centrifuged at 6000 rpm for 10 minutes. The product was recovered from the supernatant, and the absorbance was measured at 450 nm. 2.4. Hydrogen peroxide estimation based on colorimetric assay The enzyme-immobilized nanoparticles were combined with varying concentrations of H 2 O 2 and OPD and incubated for 20 minutes to detect color. The reaction was then stopped using sulphuric acid, and the supernatant was recovered via centrifugation. The optical density (O.D) was measured at 492 nm using an enzyme linked immunosorbent assay (ELISA) reader. 100 µL of supernatant was deposited in the wells of a 96-well microplate. 3. Results and discussion In this report, we describe the characterization and adsorption properties of functionalized silica nanoparticles containing vitamin B2 and horseradish peroxide enzyme. Silica nanoparticles are particularly appealing due to their biocompatibility, ease of manufacture, and low aggregation. The wet chemical method was utilized to easily change the surface of the silica layer. In this study, organosilane-modified silica nanoparticles were produced using a modified Stober process and then calcined to generate the mesoporous structures. Figure 1 displays the findings of a research of functionalized particles in terms of their morphology, structure, and properties. The DLS, SEM, and zeta potential measurements all confirmed that the particle size varied according to the aminopropyl triethoxy siliane (APTES). After calcination, plain silica nanoparticles have a zeta potential of around 39 mV. The values of APTES modified silica nanoparticles varied based on APTES concentrations, with the maximum concentration (3.576 M) revealing a value of −10 mV and the lowest concentration (0.298 M) revealing a value of −26 mV. During calcination, the produced particles immediately lose weight due to the removal of water molecules adsorbed on the silica surface, followed by APTES degradation and -C-Si bond breaking. As a result, the density of the nanoparticles varied significantly when compared to typical silica. The APTES network generates a cage-like structure in the outer silica layer, regulating the production and density of silica nanoparticles. The density of control silica nanoparticles after calcination was around 1.986g/cm 3 , but the density of APTES modified silica nanoparticles was dramatically lowered to 0.979 g/cm 3 due to 3.56 M of APTES, and the particles' shape is very porous, as shown in Fig. 2 a. The amine functional groups were measured using the ninhydrin test, revealing a maximum of 1.81⋅10 11 NH 2 functional group/particles. Calcination caused amine groups to decompose, as evidenced by a reduction in the NH 2 group to 1.23₴ 10 4 /particle. Surface characterizations were explored using FTIR spectra, with a strong and stretching peak at 1080 cm−1 suggesting that the Si-O-Si derives from silica, and three characteristic peaks at 462, 799, and 805 cm−1 indicating that the SiO 2 arises from vibrational modes. Peaks at 3439–3450 correspond to OH stretching and bending vibrations, while peaks at 1633–1637 cm−1 correspond to the amine group. The intensity of the Si-OH absorption band reduced following calcination, according to the findings. These findings suggested that the condensation process in the sol-gel method was improved, resulting in the creation of Si-O-Si functional groups, as depicted in Fig. 2 b.After calcination, the nanoparticles showed absorbance maxima between 420 and 480 nm. The peak intensity and refractive index parameters were adjusted based on the APTES concentration. The peak shift was driven by defect states produced by surface oxidation of Si-O-Si and amine group functionalization of silica nanoparticles. The peak was quite broad, and emission mechanisms can be ruled out due to their massive size and quantum confinement effects. The maximum emission was seen at lower APTES concentrations (0.298 M), as indicated in Fig. 2 c. The refractive index studies demonstrated that the values decreased and the band gap energy changed following calcination. The photodegradation study was conducted using crystal violet dye at various time intervals. The nanoparticles may serve as a catalyst by accepting electrons, allowing for a powerful oxidizing electron hole and the subsequent transfer of trapped electrons to the absorbed O 2 . As a result, more molecules are adsorbed on nanoparticle surfaces, enhancing both photoexcited electron transport to the conduction band and electron transfer to the adsorbed O 2 . The kinetics were presented in Fig. 3 . According to this, during UV irradiation, the maximum concentration of crystal violet dye was 9 mg/ml, while without UV irradiation, the maximum concentration of nanoparticles was 5 mg/ml. UV irradiation destroyed up to 90% of the dye. The degradation research uses a pseudo-first-order reaction. Table 1 is a summary of the values. The rate constant (k) was estimated by taking the starting concentration of crystal violet solution (Co) and the concentration of crystal violet at time t (Ct). The slope of the ln Co/Ct Vs time figure indicates the rate constant k in minutes (lnCo/Ct = kt). The k values obtained through linear fitting from Table 1 were 0.025 with UV irradiation and 0.01192 without UV irradiation. The activity of the particle's adsorbed enzymes was tested using o-phenylenediamine as a substrate for 70 days, and the stability results are presented in Fig. 4 a. It was clearly demonstrated that HRP-absorbed LDS particles are very stable and had higher activity than free HRP enzyme, as illustrated in 4b. The kinetics plot for HRP absorbed LDS was created using the Michaelis-Mentan equation, which is (Km/Vmax) 1/[S] + 1/Vmax = 1/v. Lineweaver-Burk plots (1/v.1/[S]) were used to derive the Michaelis-Menten constant Km and the maximum rate Vmax, while K cat was calculated as Vmax= K cat [E0], where [E0] is the initial concentration of the enzyme entrapped on the nanoparticle surface. The K cat value represents enzyme activity by reflecting the maximum number of substrate molecules that can be converted into product by one enzyme molecule in a given amount of time, as shown in Table 1 . The influence of solution pH on the immobilized enzyme-catalyzed reaction was studied in the pH range of 3 to 9, with no other experimental conditions altered. The assay findings revealed that the maximum intensity was recorded at pH 7. As a result, the pH of the reaction solution was adjusted to 7. We optimized the catalytic reaction time by conducting the assay at various reaction periods (ranging from 5 to 30 minutes) while leaving the other experimental parameters constant. Based on these findings, the reaction time was set to 20 minutes (Fig. 4 c). Figure 4 d depicts the findings of an investigation into the impact of o-phenylenediamine concentrations on the assay. Increasing the concentration of o-phenylenediamine from 2.5 ₴10 −3 to 3.0 ₴10 −2 mol L−1 boosted the intensity of the assay system. The assay findings showed a plateau exceeding 3 ₴10 −2 mol L−1. The optimal concentration of o-phenylenediamine was determined to be 3 ₴10 −2 mol L−1. HRP enzyme linked nanoparticles served as a calorimetric enzyme probe for detecting H 2 O 2 in the presence of o-phenyl diamine as a substrate. The results showed that nanoparticles had a low detection limit of up to 10 −10 mol L −1 of H 2 O 2 , comparable to 5 nmol, and the H 2 O 2 concentrations had a linear range from 5₴10 −10 ~ 1₴10 −5 (M) (R 2 =0.995), as shown in Figure 5. The milk sample was spiked with H 2 O 2 for real-world examination, and the results were measured. For the tests, 1, 50, and 100 nmol of H 2 O 2 were produced and spiked with 1 mL of sterile milk samples, and the recovery rate was measured using colorimetric assays. The average absorption values (O.D) for the H 2 O 2 solution were 0.183 ± 0.08, 0.457 ± 0.02 and 0.501 ± 0.07. The spiked milk samples yielded levels of 0.183 ± 0.04, 0.454 ± 0.09, and 0.505 ± 0.01.‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬ 4. Conclusion Finally, this study proved the ability to produce low-density silica nanoparticles of various sizes by employing organosiliane as an amine functional source. After calcination, Si-O-Si molecules with silica and amine groups were de absorbed, affecting density and making the material more mesoporous. For the adsorption investigations, the particles were adsorbed with horse radish peroxidase enzymes, and the enzyme kinetics and adsorption isotherm of the particles were also determined. The particles were then employed as a calorimetric probe to detect H 2 O 2 in milk samples. Finally, we concluded that the described technology will provide a simple way to construct bio functional porous structures with large volumes. Declarations Author contributions Viswanathan kaliyaperumal− Complete manuscript writing, experimental design, and physicochemical characterization of LSD NPs, Over all supervision. Chitra Priya kaliyaperumal and Sri lekha Rajasekaran - catalytic and hydrogen detection assays Radha perumal ramasamy - enzyme and dye kinetics and manuscript corrections Funding No funding involved in this work. Data availability The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. Competing interests The authors declare no competing interests. Acknowledgement Author would like thanks AVC college, SAIF -IIT-Madras for lab facility and particles characterizations and analysis. References Moller K, Bein T (2019) Degradable drug carriers: vanishing mesoporous silica nanoparticles. Chem Mater 31:4364–4378 Akhter F, Rao AA, Abbasi MN et al (2022) A Comprehensive Review of Synthesis, Applications and Future Prospects for Silica Nanoparticles (SNPs). Silicon 14:8295–8310 Chen F, Hableel G, Zhao ER, Jokerst JV (2018) Multifunctional nanomedicine with silica: Role of silica in nanoparticles for theranostic, imaging, and drug monitoring. J Colloid Interface Sci 521:261–279 Qhobosheane M, Santra S, Zhang P, Tan W (2001) Biochemically functionalized silica nanoparticles. 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Nanomedicine (Lond). 3:703 – 17 Ilhan-Ayisigi E, Yesil-Celiktas O (2018) Silica-based organic-inorganic hybrid nanoparticles and nanoconjugates for improved anticancer drug delivery. Eng Life Sci 18:882–892 Kumar R, Roy I, Ohulchanskky TY, Vathy LA, Bergey EJ, Sajjad M, Prasad PN (2010) In vivo biodistribution and clearance studies using multimodal organically modified silica nanoparticles. ACS Nano 4:699–708 Chircov C, Matei MF, Neacșu IA, Vasile BS, Oprea OC, Croitoru AM, Trușcă RD, Andronescu E, Sorescu I, Barbuceanu F (2021) Iron Oxide-Silica Core-Shell Nanoparticles Functionalized with Essential Oils for Antimicrobial Therapies. Antibiot (Basel) 10:1138 Kobayashi Y, Matsudo H, Li TT, Shibuya K, Kubota Y, Oikawa T, Nakagawa T, Gonda K (2016) Fabrication of quantum dot/silica core–shell particles immobilizing Au nanoparticles and their dual imaging functions. Appl Nanosci 6:301–307 Jankiewicz BJ, Jamiola D, Choma J, Jaroniec M (2012) Silica–metal core–shell nanostructures. Adv Colloid Interface Sci 170:28–47 Akhter F, Rao AA, Abbasi MN, Wahocho SA, Mallah MA, Anees-ur-Rehman H, Chandio ZA (2022) A Comprehensive Review of Synthesis, Applications and Future Prospects for Silica Nanoparticles (SNPs). Silicon 14:8295–8310 Han Y, Lu Z, Teng Z, Liang J, Guo Z, Wang D, Han MY, Yang W (2017) Unraveling the Growth Mechanism of Silica Particles in the Stöber Method. Situ Seeded Growth Model Langmuir 33:5879–5890 Ahmad R, Ansari K (2022) Fabrication of alginate@ silver nanoparticles (Alg@ AgNPs) bionanocomposite for the sequestration of crystal violet dye from aqueous solution. Int J Biol Macromol 218:157–167 Tables Tables are available in the Supplementary Files section. Additional Declarations No competing interests reported. Supplementary Files table.docx floatimage1.png Graphical abstract Cite Share Download PDF Status: Posted Version 1 posted 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. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-9063139","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":603003959,"identity":"b0db163c-b6ce-4c13-bcb6-e638c0a0dce1","order_by":0,"name":"Viswanathan kaliyaperumal","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA90lEQVRIiWNgGAWjYDACZihtAGaz2TAwSBCpRQKqJY0ILQyoWg4T1mJwnP2ZxM8ddnXm7GePPS4oO5/YP7v54AOGGptonFoO85hJ9p5JlrDsyUs3nnHuduKMO8eSDRiOpeU24NAi2czDdoO3jVnC4ECOmTRv2+3Ehhs5ZhKMDYfxaGF/dvNvW72Ewfk3IC3nEucT0sLPzGB2m7ftsITBDbAtBxI3ENbCY/5btu245IYbb8yNec4lG2+8kZZskIDHL2z8xx8bvm2r5jc4n2P2mKfMTnbejeSDDz7U2ODUgqIdRDiCVSYQoRyuxZ5IxaNgFIyCUTCCAABDuVjLtfew1wAAAABJRU5ErkJggg==","orcid":"","institution":"Saveetha Institute of Medical and Technical Sciences","correspondingAuthor":true,"prefix":"","firstName":"Viswanathan","middleName":"","lastName":"kaliyaperumal","suffix":""},{"id":603003960,"identity":"20aee3bc-c822-4496-88f8-8e5917ed176f","order_by":1,"name":"Radha perumal ramasamy","email":"","orcid":"","institution":"Anna University, Chennai","correspondingAuthor":false,"prefix":"","firstName":"Radha","middleName":"perumal","lastName":"ramasamy","suffix":""},{"id":603003961,"identity":"12b2cb18-da70-4fc1-b3d3-847cc1e50d54","order_by":2,"name":"Rathinam Ganesan","email":"","orcid":"","institution":"siddha central research institute","correspondingAuthor":false,"prefix":"","firstName":"Rathinam","middleName":"","lastName":"Ganesan","suffix":""},{"id":603003962,"identity":"6b65096e-1731-4f44-a799-7cf80243d81f","order_by":3,"name":"kaliyaperumal Chitrapriya","email":"","orcid":"","institution":"AVC college of arts and science","correspondingAuthor":false,"prefix":"","firstName":"kaliyaperumal","middleName":"","lastName":"Chitrapriya","suffix":""}],"badges":[],"createdAt":"2026-03-08 09:23:04","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-9063139/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9063139/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":104296536,"identity":"d09fd700-8a37-47ae-aef5-dd0f2f1a631a","added_by":"auto","created_at":"2026-03-10 07:59:51","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1141147,"visible":true,"origin":"","legend":"\u003cp\u003ePhysicochemical characterization of low-density silica nanoparticles. The SEM and particle size measurement results based on APTES concentrations, as well as the correlation analysis of size and zeta potential values.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/2e61dffc642a942586040b5a.png"},{"id":104296531,"identity":"489659e8-51e2-4ae7-bd0a-31ccaebbca4d","added_by":"auto","created_at":"2026-03-10 07:59:51","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":443763,"visible":true,"origin":"","legend":"\u003cp\u003eResults of the APTES impacts (2a) Density variation (2b) Results of FTIR-based surface characterizations (2c) UV absorption spectrum alterations arise.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/6c7547a73f4371ca0672ece8.png"},{"id":104296537,"identity":"019b976d-8fb2-45b6-8622-4c41173a8e64","added_by":"auto","created_at":"2026-03-10 07:59:52","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":222136,"visible":true,"origin":"","legend":"\u003cp\u003eCatalytic activity study results, the dye concentration is 10 mg/L, the concentration of low-density silica nanoparticles is 500 mg, and the maximum irradiation time is 150 minutes (3a). Spectra changes during and without UV irradiation (3b) Pseudo first-order kinetics for photocatalytic degradation in the presence and absence of UV (3c).\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/b21a29c503b60a81ca02b1a4.png"},{"id":104296534,"identity":"374b64e4-37ca-4e67-a4ba-40dbfdf2a4db","added_by":"auto","created_at":"2026-03-10 07:59:51","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1043720,"visible":true,"origin":"","legend":"\u003cp\u003eEnzymes absorbed LSD nanoparticles. (4a). Stability investigations of free HRP and enzyme-absorbed LSD nanoparticles (4b) HRP-absorbed LSD particles-based H2O2 detection reaction scheme (4c) optimization findings of reaction time (4d) substrate concentrations\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/8dbe1c549e96c4db13d1da9f.png"},{"id":104296533,"identity":"22322001-5655-4805-b68f-b6cdf5114f76","added_by":"auto","created_at":"2026-03-10 07:59:51","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1215400,"visible":true,"origin":"","legend":"\u003cp\u003eResults of detecting hydrogen peroxide in milk samples\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/bb8c8ef32904b76d6e730aac.png"},{"id":104786352,"identity":"54e7a85c-aa9b-48ba-8035-3261bb1293e4","added_by":"auto","created_at":"2026-03-17 08:16:45","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":5888956,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/4af4256d-9000-475a-9159-512ac402ba72.pdf"},{"id":104296532,"identity":"4524fce4-8b30-48f8-ac69-e3ba256d7b5a","added_by":"auto","created_at":"2026-03-10 07:59:51","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":15087,"visible":true,"origin":"","legend":"","description":"","filename":"table.docx","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/522e03fc5426af6fd115fd38.docx"},{"id":104296535,"identity":"946f2e6d-f856-411b-bc35-84486fc5dfaf","added_by":"auto","created_at":"2026-03-10 07:59:51","extension":"png","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":161625,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGraphical abstract\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-9063139/v1/a3c8d39946dd997d3144e9da.png"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eCalcined low-density silica nanoparticles for HRP immobilization: catalytic dye removal and hydrogen peroxide detection in milk\u003c/p\u003e","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eSilica nanoparticles are an important type of nanotool used in a variety of applications due to their ease of manufacture, high chemical stability, easy surface modification, biocompatibility, and use as a tool to control drug molecules [\u003cspan additionalcitationids=\"CR2\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Currently, three types of silica nanoparticles have been created: solid, nonporous, and mesoporous [\u003cspan additionalcitationids=\"CR5\" citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Several methods for synthesizing silica nanoparticles have been recorded, including plasma manufacturing, chemical vapor deposition, microemulsion synthesis, combustion processing, sol-gel synthesis, and hydrothermal reactions [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Recently, one-, two-, and three-dimensional silica nanoparticles, Silica nanotubes, hollow silica nanotubes, and silica nanofibers have been described [\u003cspan additionalcitationids=\"CR9\" citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Currently, silica nanoparticles are used in a variety of applications, including catalysis, drug delivery, biomedicine, environmental remediation, and wastewater treatment [\u003cspan additionalcitationids=\"CR12 CR13\" citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. In biological applications, surface charge is significant, and cationic or anionic charge-based elimination was shown in animal models. Silica nano particles smaller than 6 nm are quickly removed by urine, but those larger than this accumulates in the liver [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Organic silica has recently been discovered to be a highly effective therapeutic agent for the treatment of bone and skin diseases. The small size and positive surface charge of organic silica-based nanoparticles make them an attractive platform for medication and gene therapy, where a negatively charged membrane improves cellular uptake [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. These materials are also biocompatible and contain low toxicity. Silica nanoparticles were easily combined with iron oxide, quantum dots, fluorescent dye, gold, and other metal nanoparticles to form hybrid nanoparticles [\u003cspan additionalcitationids=\"CR19 CR20\" citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. In this work, we developed low density silica particles based on organ silane modification and calcinations. The catalytic activity was examined using crystal violet, and the nano sensors application for hydrogen peroxide detection were made utilizing the nanoparticles.\u003c/p\u003e"},{"header":"2. Experimental procedure","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Production of organosilane-modified silica nanoparticles\u003c/h2\u003e \u003cp\u003eOrganosilane-modified silica nanoparticles were produced using slightly modified procedures from the previously disclosed simplified stober approach [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. To summarize, 10 mL of ethanol, 1 mL of tetraethyl orthosilicate, 0.2 mL of (3-Aminopropyl)-triethoxysilane (APTES), and 1 mL of ammonium hydroxide solution were added. This solution was agitated for an additional 3 hours at room temperature. The particles were collected by centrifugation at 6000 rpm for 10 min, then washed with ethanol and dried in a vacuum oven at 37\u0026deg;C for 2 hours.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Photo catalytic activity studies\u003c/h2\u003e \u003cp\u003ePhotocatalytic activity was evaluated using previously published data [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. For this work, 5 mL of 10 mg/L crystal violet was combined with 500 mg of nanoparticles at room temperature (C), and another set of samples was exposed to a lamp (mercury vapour lamp, 40 W, 40 cm long, wavelength 253.7 nm). The sample was placed in a 10-mL tube. At regular intervals, the emission spectra were examined.\u003c/p\u003e \u003cp\u003eThe dye degradation rate was computed as C\u003csub\u003et\u003c/sub\u003e/C\u003csub\u003e0\u003c/sub\u003e. The initial concentration is C\u003csub\u003e0\u003c/sub\u003e, the reaction concentration is C\u003csub\u003et\u003c/sub\u003e, and the graph shows C\u003csub\u003et\u003c/sub\u003e/C\u003csub\u003e0\u003c/sub\u003e vs irradiation time.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3. \u003cem\u003eEnzyme adsorption and Assay\u003c/em\u003e:\u003c/h2\u003e \u003cp\u003eFor horse radish peroxidase adsorption, 130 mg of organo silica nanoparticles were placed in a glass vial, followed by 1 mL of 5 mg/mL horse radish peroxidase solutions, which were incubated for 60 minutes while stirring. Following that, the absorbed enzyme activity on nanoparticles was investigated using o-phenylenediamine (OPD). Briefly, 0.2 mL of 5mg/mL enzyme-coated organosilane silica nanoparticles were placed in an eppendorf tube. The tubes were then centrifuged at 6000 rpm for 5 minutes to separate the particles. Then, 0.1 mL of o-phenylenediamine and 0.1 mL of serially diluted hydrogen peroxide were added. The mixture was gently agitated for 20 minutes before adding 20 \u0026micro;L of pure sulphuric acid to terminate the enzyme reaction. The tube was then centrifuged at 6000 rpm for 10 minutes. The product was recovered from the supernatant, and the absorbance was measured at 450 nm.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4. Hydrogen peroxide estimation based on colorimetric assay\u003c/h2\u003e \u003cp\u003eThe enzyme-immobilized nanoparticles were combined with varying concentrations of H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e and OPD and incubated for 20 minutes to detect color. The reaction was then stopped using sulphuric acid, and the supernatant was recovered via centrifugation. The optical density (O.D) was measured at 492 nm using an enzyme linked immunosorbent assay (ELISA) reader. 100 \u0026micro;L of supernatant was deposited in the wells of a 96-well microplate.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results and discussion","content":"\u003cp\u003eIn this report, we describe the characterization and adsorption properties of functionalized silica nanoparticles containing vitamin B2 and horseradish peroxide enzyme. Silica nanoparticles are particularly appealing due to their biocompatibility, ease of manufacture, and low aggregation. The wet chemical method was utilized to easily change the surface of the silica layer. In this study, organosilane-modified silica nanoparticles were produced using a modified Stober process and then calcined to generate the mesoporous structures. Figure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e displays the findings of a research of functionalized particles in terms of their morphology, structure, and properties. The DLS, SEM, and zeta potential measurements all confirmed that the particle size varied according to the aminopropyl triethoxy siliane (APTES). After calcination, plain silica nanoparticles have a zeta potential of around 39 mV. The values of APTES modified silica nanoparticles varied based on APTES concentrations, with the maximum concentration (3.576 M) revealing a value of \u0026minus;10 mV and the lowest concentration (0.298 M) revealing a value of \u0026minus;26 mV. During calcination, the produced particles immediately lose weight due to the removal of water molecules adsorbed on the silica surface, followed by APTES degradation and -C-Si bond breaking. As a result, the density of the nanoparticles varied significantly when compared to typical silica. The APTES network generates a cage-like structure in the outer silica layer, regulating the production and density of silica nanoparticles. The density of control silica nanoparticles after calcination was around 1.986g/cm\u003csup\u003e3\u003c/sup\u003e, but the density of APTES modified silica nanoparticles was dramatically lowered to 0.979 g/cm\u003csup\u003e3\u003c/sup\u003e due to 3.56 M of APTES, and the particles' shape is very porous, as shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea. The amine functional groups were measured using the ninhydrin test, revealing a maximum of 1.81\u0026sdot;10\u003csup\u003e11\u003c/sup\u003e NH\u003csub\u003e2\u003c/sub\u003e functional group/particles. Calcination caused amine groups to decompose, as evidenced by a reduction in the NH\u003csub\u003e2\u003c/sub\u003e group to 1.23₴ 10\u003csup\u003e4\u003c/sup\u003e/particle. Surface characterizations were explored using FTIR spectra, with a strong and stretching peak at 1080 cm\u0026minus;1 suggesting that the Si-O-Si derives from silica, and three characteristic peaks at 462, 799, and 805 cm\u0026minus;1 indicating that the SiO\u003csub\u003e2\u003c/sub\u003e arises from vibrational modes. Peaks at 3439\u0026ndash;3450 correspond to OH stretching and bending vibrations, while peaks at 1633\u0026ndash;1637 cm\u0026minus;1 correspond to the amine group. The intensity of the Si-OH absorption band reduced following calcination, according to the findings. These findings suggested that the condensation process in the sol-gel method was improved, resulting in the creation of Si-O-Si functional groups, as depicted in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb.After calcination, the nanoparticles showed absorbance maxima between 420 and 480 nm. The peak intensity and refractive index parameters were adjusted based on the APTES concentration. The peak shift was driven by defect states produced by surface oxidation of Si-O-Si and amine group functionalization of silica nanoparticles. The peak was quite broad, and emission mechanisms can be ruled out due to their massive size and quantum confinement effects. The maximum emission was seen at lower APTES concentrations (0.298 M), as indicated in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec. The refractive index studies demonstrated that the values decreased and the band gap energy changed following calcination. The photodegradation study was conducted using crystal violet dye at various time intervals. The nanoparticles may serve as a catalyst by accepting electrons, allowing for a powerful oxidizing electron hole and the subsequent transfer of trapped electrons to the absorbed O\u003csub\u003e2\u003c/sub\u003e. As a result, more molecules are adsorbed on nanoparticle surfaces, enhancing both photoexcited electron transport to the conduction band and electron transfer to the adsorbed O\u003csub\u003e2\u003c/sub\u003e. The kinetics were presented in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e. According to this, during UV irradiation, the maximum concentration of crystal violet dye was 9 mg/ml, while without UV irradiation, the maximum concentration of nanoparticles was 5 mg/ml. UV irradiation destroyed up to 90% of the dye. The degradation research uses a pseudo-first-order reaction. Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e is a summary of the values. The rate constant (k) was estimated by taking the starting concentration of crystal violet solution (Co) and the concentration of crystal violet at time t (Ct). The slope of the ln Co/Ct Vs time figure indicates the rate constant k in minutes (lnCo/Ct\u0026thinsp;=\u0026thinsp;kt). The k values obtained through linear fitting from Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e were 0.025 with UV irradiation and 0.01192 without UV irradiation. The activity of the particle's adsorbed enzymes was tested using o-phenylenediamine as a substrate for 70 days, and the stability results are presented in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea. It was clearly demonstrated that HRP-absorbed LDS particles are very stable and had higher activity than free HRP enzyme, as illustrated in 4b. The kinetics plot for HRP absorbed LDS was created using the Michaelis-Mentan equation, which is (Km/Vmax) 1/[S]\u0026thinsp;+\u0026thinsp;1/Vmax\u0026thinsp;=\u0026thinsp;1/v. Lineweaver-Burk plots (1/v.1/[S]) were used to derive the Michaelis-Menten constant Km and the maximum rate Vmax, while K\u003csub\u003ecat\u003c/sub\u003e was calculated as Vmax= K\u003csub\u003ecat\u003c/sub\u003e [E0], where [E0] is the initial concentration of the enzyme entrapped on the nanoparticle surface. The K\u003csub\u003ecat\u003c/sub\u003e value represents enzyme activity by reflecting the maximum number of substrate molecules that can be converted into product by one enzyme molecule in a given amount of time, as shown in Table \u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. The influence of solution pH on the immobilized enzyme-catalyzed reaction was studied in the pH range of 3 to 9, with no other experimental conditions altered. The assay findings revealed that the maximum intensity was recorded at pH 7. As a result, the pH of the reaction solution was adjusted to 7. We optimized the catalytic reaction time by conducting the assay at various reaction periods (ranging from 5 to 30 minutes) while leaving the other experimental parameters constant. Based on these findings, the reaction time was set to 20 minutes (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec). Figure\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed depicts the findings of an investigation into the impact of o-phenylenediamine concentrations on the assay. Increasing the concentration of o-phenylenediamine from 2.5 ₴10\u003csup\u003e\u0026minus;3\u003c/sup\u003e to 3.0 ₴10\u003csup\u003e\u0026minus;2\u003c/sup\u003emol L\u0026minus;1 boosted the intensity of the assay system. The assay findings showed a plateau exceeding 3 ₴10\u003csup\u003e\u0026minus;2\u003c/sup\u003emol L\u0026minus;1. The optimal concentration of o-phenylenediamine was determined to be 3 ₴10\u003csup\u003e\u0026minus;2\u003c/sup\u003e mol L\u0026minus;1. HRP enzyme linked nanoparticles served as a calorimetric enzyme probe for detecting H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e in the presence of o-phenyl diamine as a substrate. The results showed that nanoparticles had a low detection limit of up to 10\u003csup\u003e\u0026minus;10\u003c/sup\u003e mol L\u003csup\u003e\u0026minus;1\u003c/sup\u003e of H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e, comparable to 5 nmol, and the H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e concentrations had a linear range from 5₴10\u003csup\u003e\u0026minus;10\u003c/sup\u003e ~ 1₴10\u003csup\u003e\u0026minus;5\u003c/sup\u003e(M) (R\u003csup\u003e2\u003c/sup\u003e=0.995), as shown in Figure 5. The milk sample was spiked with H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e for real-world examination, and the results were measured. For the tests, 1, 50, and 100 nmol of H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e were produced and spiked with 1 mL of sterile milk samples, and the recovery rate was measured using colorimetric assays. The average absorption values (O.D) for the H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e solution were 0.183 \u0026plusmn; 0.08, 0.457 \u0026plusmn; 0.02 and 0.501 \u0026plusmn; 0.07. The spiked milk samples yielded levels of 0.183 \u0026plusmn; 0.04, 0.454 \u0026plusmn; 0.09, and 0.505 \u0026plusmn; 0.01.‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬\u003c/p\u003e "},{"header":"4. Conclusion","content":"\u003cp\u003eFinally, this study proved the ability to produce low-density silica nanoparticles of various sizes by employing organosiliane as an amine functional source. After calcination, Si-O-Si molecules with silica and amine groups were de absorbed, affecting density and making the material more mesoporous. For the adsorption investigations, the particles were adsorbed with horse radish peroxidase enzymes, and the enzyme kinetics and adsorption isotherm of the particles were also determined. The particles were then employed as a calorimetric probe to detect H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e in milk samples. Finally, we concluded that the described technology will provide a simple way to construct bio functional porous structures with large volumes.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eViswanathan \u0026nbsp;kaliyaperumal\u0026minus; Complete manuscript writing, experimental design, and physicochemical characterization of LSD NPs, Over all supervision.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eChitra Priya kaliyaperumal and Sri lekha Rajasekaran - catalytic and hydrogen detection assays\u003c/p\u003e\n\u003cp\u003eRadha perumal ramasamy - enzyme and dye kinetics and manuscript corrections\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003eNo funding involved in this work.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u0026nbsp;\u003c/strong\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAuthor would like thanks AVC college, SAIF -IIT-Madras for lab facility and particles characterizations and analysis. \u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eMoller K, Bein T (2019) Degradable drug carriers: vanishing mesoporous silica nanoparticles. Chem Mater 31:4364\u0026ndash;4378\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAkhter F, Rao AA, Abbasi MN et al (2022) A Comprehensive Review of Synthesis, Applications and Future Prospects for Silica Nanoparticles (SNPs). Silicon 14:8295\u0026ndash;8310\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen F, Hableel G, Zhao ER, Jokerst JV (2018) Multifunctional nanomedicine with silica: Role of silica in nanoparticles for theranostic, imaging, and drug monitoring. J Colloid Interface Sci 521:261\u0026ndash;279\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eQhobosheane M, Santra S, Zhang P, Tan W (2001) Biochemically functionalized silica nanoparticles. Analyst 126:1274\u0026ndash;1278\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMehmood A, Ghafar H, Yaqoob S, Gohar UF, Ahmad B (2017) Mesoporous Silica Nanoparticles: A Review. 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Appl Nanosci 6:301\u0026ndash;307\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJankiewicz BJ, Jamiola D, Choma J, Jaroniec M (2012) Silica\u0026ndash;metal core\u0026ndash;shell nanostructures. Adv Colloid Interface Sci 170:28\u0026ndash;47\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAkhter F, Rao AA, Abbasi MN, Wahocho SA, Mallah MA, Anees-ur-Rehman H, Chandio ZA (2022) A Comprehensive Review of Synthesis, Applications and Future Prospects for Silica Nanoparticles (SNPs). Silicon 14:8295\u0026ndash;8310\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHan Y, Lu Z, Teng Z, Liang J, Guo Z, Wang D, Han MY, Yang W (2017) Unraveling the Growth Mechanism of Silica Particles in the St\u0026ouml;ber Method. Situ Seeded Growth Model Langmuir 33:5879\u0026ndash;5890\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAhmad R, Ansari K (2022) Fabrication of alginate@ silver nanoparticles (Alg@ AgNPs) bionanocomposite for the sequestration of crystal violet dye from aqueous solution. Int J Biol Macromol 218:157\u0026ndash;167\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTables are available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"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":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"low density silica, Stobber method, hydrogen peroxide, crystal violet, kinetics","lastPublishedDoi":"10.21203/rs.3.rs-9063139/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9063139/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"In this study, low density silica nanoparticles (LDS) were synthesized and used as a heterogeneous catalyst for dye removal and nanosensors for hydrogen peroxide detection. Organosilane silica nanoparticles were produced and calcinated at 400\u0026deg;C to produce low density particles. SEM, FTIR, DLS, and zeta potential analysis were used to characterize the generated particles. The results confirmed that the particle size, charge, and density varied with the APTES concentration. The results showed that at lower APTES concentrations, small particles (70\u0026ndash;75 nm) were created, but at higher concentrations, larger particles (420\u0026ndash;430 nm) were produced. The particle density changed depending on the APTES content. The UV absorption spectra of APTES-added samples varied significantly between 440 and 480 nm. The dye removal activity was measured using crystal violet dye; under UV irradiation, the particles destroyed up to 90% of the dye within 120 minutes, and the kinetics were also detailed. HRP-coated LDS was used as a nanosensor for detecting HO. Results indicated linearity of 5\u0026sdot;10 \u0026thinsp;~\u0026thinsp;1\u0026sdot;10(M) (R\u0026thinsp;=\u0026thinsp;0.995), and a detection limit of 5 nm mol. The milk was spiked with HO at varied concentrations and used as detections for the actual sample analysis. Finally, this paper describes the simplest way for producing tailored low-density silica, which is ideal for biomedical and drug delivery applications.","manuscriptTitle":"Calcined low-density silica nanoparticles for HRP immobilization: catalytic dye removal and hydrogen peroxide detection in milk","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-03-10 07:59:46","doi":"10.21203/rs.3.rs-9063139/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"87b6ea99-4593-4eb5-a8d0-f353b4f2889a","owner":[],"postedDate":"March 10th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-03-17T07:44:13+00:00","versionOfRecord":[],"versionCreatedAt":"2026-03-10 07:59:46","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9063139","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9063139","identity":"rs-9063139","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}