Synthesis of trypsin protected CsPbCl3 fluorescent nanocrystals for hydroxyl radical sensing

Synthesis of trypsin protected CsPbCl3 fluorescent nanocrystals for hydroxyl radical sensing | 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 Synthesis of trypsin protected CsPbCl3 fluorescent nanocrystals for hydroxyl radical sensing Suresh Kumar Kailasa, Kartik Pankajbhai Makwana, Madhura Pradeep Deshpande, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5436012/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 10 Mar, 2025 Read the published version in Microchimica Acta → Version 1 posted 9 You are reading this latest preprint version Abstract Water-dispersible perovskite nanocrystals (PNCs) show promising applications in recognizing ionic and molecular species because of their excellent optical properties. However, lead halide PNCs have some limitations when they are used as probes for molecular species sensing in aqueous media. Here, we introduce trypsin (Try) as a bioligand for the synthesis of cesium lead chloride (CsPbCl 3 ) perovskite nanocrystals (PNCs) with high water stability. The as-fabricated Try-CsPbCl 3 PNCs show λ Em/Ex at 433/370 nm with quantum yield of 17.26%. The fluorescence emission spectral characteristics of Try-CsPbCl 3 PNCs demonstrated that water-stable Try-CsPbCl 3 PNCs acted as a promising fluorescent probe for the detection of hydroxyl radical ( • OH) via turn-off mechanism. The Try-CsPbCl 3 PNCs-based turn-off fluorescence approach displayed good selectivity for hydroxyl radical in water, showing a wider linear range (0.01–5 µM) with a remarkable detection limit of 3.10 nM for hydroxyl radical. The as-prepared Try-CsPbCl 3 PNCs were demonstrated to be a facile probe for sensing • OH in water samples, which signifies that Try-CsPbCl 3 PNCs exhibited broad applications for hydroxyl radical sensing and cell imaging. Try-CsPbCl3 PNCs Hydroxyl radical Fluorescence spectrometry HR-TEM Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1. Introduction In recent years, lead halide perovskite nanocrystals (PNCs) have emerged as outstanding materials in fabricating solar cells, sensors, light-emitting diodes and optoelectronic devices because of their remarkable characteristics including superior quantum yield (QY), narrow full width at half-maximum, impressive charge transport capabilities, high photocatalytic properties and tunable emission wavelength in complete visible and near-infrared region [ 1 – 5 ]. Interestingly, it was noticed that perovskite oxides are quite stable in water whereas lead halide PNCs are highly unstable in water and easily degraded by exposing them to heat, light and moisture [ 6 – 8 ], resulting in limiting their promising applications in multidisciplinary research. To improve lead halide PNCs water stability, several researchers have introduced different strategies to fabricate lead halide PNCs including lead chloride PNCs with high water stability. For example, perflorocompounds were used as ligands for the preparation of water-stable CsPbBr 3 @Cs 4 PbBr 6 PNCs [ 9 ]. Furthermore, various polymers (poly(l-lactide) polypropylene glycol, and polysulfone [ 10 ], NH 2 -PEG-COOH [ 11 ], NH 2 group terminated hiperbranched polymer [ 12 ], polymethyl methacrylate [ 13 ], polystyrene [ 14 ], polystyrene-cetyltrimethylammonium bromide [ 15 ] and polyethylene glycol [ 16 ]), organic molecules (ethylenediaminetetraacetic acid [ 17 ], bolaamphiphilic ligand (NKE-12) [ 18 ], adamantane-1-amine [ 19 ], (4,4'-bipyridine and 2,2'-bipyridine) [ 20 ], glycyrrhizic acid [ 21 ], 4-bromo-butyric acid-oleylamine [ 22 ], oleylamine-oleic acid [ 23 ] and oleic acid-3-bromopropionic acid [ 24 ]) and inorganic salts and compounds (cesium trifluoroacetate [ 25 ], MAPbBr 3 @lead laurate [ 26 ], mesoporous silica [ 27 ], Al 2 O 3 [ 28 ], metal-organic frameworks [ 29 ], ZrO 2 [ 30 ] and TiO 2 [ 31 ]), respectively. The above approaches were successfully produced high stability of lead halide PNCs without losing their fluorescence properties, suggesting encapsulation of lead halide PNCs with suitable ligand chemistry offers several features such as water stability, good QY and superior optical properties. Furthermore, it is a very challenging task to design water- and air-stable lead chloride PNCs without the use of complicated synthetic approaches as well as ligand chemistry. In order to alter the surface chemistry and fluorescence properties of CsPbCl 3 PNCs, trypsin (Try) was explored as a bioligand for the preparation of water-stable CsPbCl 3 PNCs in the sensing of hydroxyl radical. Reactive oxygen species (ROS) are produced by mitochondria in the cells, exhibiting a high reactive nature [ 32 ]. The ROS (oxygen-containing radicals - superoxide, peroxyl, hydroxyl ( • OH) and hydroperoxyl) and non-radical agents ozone, HOCl and H 2 O 2 which are easily converted into ROS) play a key role in numerous biochemical pathways in the cells [ 33 ]. It was observed that certain levels of ROS efficiently enhance cellular functions (migration, proliferation and differentiation) [ 34 ]. However, ROS have potentially induced oxidative stress and cell damage, which yields cell death [ 35 ]. Among ROS, hydroxyl radical is recognized as one of the highly reactive ROS, exhibiting a lifetime in the nanoseconds [ 36 – 37 ]. Usually, hydroxyl radical is produced in in vivo via oxygen molecule oxidation to superoxide, higher levels of • OH cause oxidative damage to various bio-macromolecules (nucleic acids, lipids, carbohydrates and proteins, demonstrating that significant attention must be paid to monitoring of • OH in cells and various environments. In order to identify • OH radical, several analytical techniques such as electrochemical, electron spin resonance, fluorescence, and UV–visible spectroscopic and high-performance liquid chromatographic techniques have been applied to detect ROS including • OH radical [ 38 – 46 ]. Importantly, several fluorescent probes including terbium complexes [ 36 ], Coumarin–Neutral Red [ 41 ], copper and molybdenum nanoclusters [ 47 – 48 ], Ag–Au nanocages [ 49 ], carbon dot-based hydrogel [ 50 ] and dihydroquinolines have been utilized as fluorescence probes for sensing of • OH radical with lower limit of detection (LOD) [ 51 ]. The nanomaterials- and organic molecules-based fluorescence probes have proven to be promising readers to selective assay of • OH in biological and environmental systems. Due to their complex procedures in fabricating fluorescence probes, there is a still necessity to introduce a facile and novel fluorescence probe for • OH sensing in water samples. In this work, we report a simple and selective hydroxyl radical ( • OH) sensing strategy using aqueous-stable trypsin (Try) encapsulated cesium lead chloride (CsPbCl 3 ) PNCs as a nanoprobe (Scheme 1). The as-synthesized Try-CsPbCl 3 PNCs were stable in aqueous medium and displayed spherical shape morphology with a mean size of 2.5 ± 0.5 nm. Further, Try-CsPbCl 3 PNCs exhibited blue fluorescence under UV lamp (365 nm), showing λ Em/Ex at 433/370 nm, which offers a QY of 17.26%. Noticeably, the emission intensity of Try-CsPbCl 3 PNCs was quenched by hydroxyl radical, leading to the development of a fluorescence turn-off approach for • OH assay with an LOD of 3.10 nM. The developed sensing strategy was used to detect • OH in water samples, demonstrating Try-CsPbCl 3 PNCs-based fluorescence method could be an effective tool for monitoring • OH in real samples. 2. Experimental section 2.1. Materials and instruments Lead (II) chloride (PbCl 2 , 98%) and cesium chloride (CsCl, 99.99%) were produced from SRL and BLD pharm, respectively. 1 -Octadecene (ODE), potassium superoxide (KO 2 ) were obtained from Sigma Aldrich, N-nromo succinimide (NBS), sodium hypochlorite (NaOCl) and hydrogen peroxide (H 2 O 2 ) were purchased from SRL, FINAR and SDFCL Chemicals, respectively. Deionized water was used for preparation of solutions and sensing experiments, and analytical-grade chemicals were utilized without any further purification. The as-prepared Try-CsPbCl 3 PNCs size and morphology were investigated using field emission transmission electron microscopy (FETEM) (JEOL-200, Tokyo, Japan). Fluorescence (emission and excitation) spectra of Try-CsPbCl 3 PNCs were examined by a Cary Eclipse fluorescence spectrometer (Agilent Technologies, Santa Clara, CA, USA). UV–vis absorption spectra were recorded with a Maya Pro 2000 spectrophotometer (Ocean Optics, Orlando, FL, USA). Infrared spectra of Try-CsPbCl 3 PNCs were examined an ALPHA II Fourier transform infrared (FT-IR) spectrometer (Bruker, Billerica, MA, USA). X-ray diffraction (XRD) spectrum of Try-CsPbCl 3 PNCs was recorded using a D8-Advance Instrument (Bruker AXS). X-ray photoelectron spectroscopy (XPS) (K-alpha+, Thermo Fisher Scientific, Waltham, MA, USA) was performed for the confirmation of the elemental composition of Try-CsPbCl 3 PNCs. Hydrodynamic diameter and zeta potentials of Try-CsPbCl 3 PNCs were obtained using a NanoZS90 nano-particle Size potential analyzer (Malvern, UK). 2.2. Synthesis of Try-CsPbCl 3 PNCs Blue fluorescence water-stable Try-CsPbCl 3 PNCs were prepared via a simple reaction. Firstly, 100 mM (33.6 mg) of PbCl 2 and 100 mM (55.6 mg) of CsCl precursors were dispersed in 2 mL of ODE separately and stirred for 30 minutes at room temperature. Then, the preparation of CsPbCl 3 PNCs was initiated by mixing both solutions in 10 mL of reaction flask. The mixture was stirred at room temperature for 3 h. The formed CsPbCl 3 PNCs were capped with Try by adding Try (32 mg) into CsPbCl 3 solution and then stirred for 12 h, triggering the formation of blue fluorescent Try-CsPbCl 3 PNCs. The formed Try-CsPbCl 3 PNCs were washed with hexane and then dispersed in water for sensing applications. 2.4. Fluorescence sensing of • OH using Try-CsPbCl 3 PNCs as a turn-off probe In the sensing study, the following reactive species solutions were prepared as follows, hydroxyl radical ( • OH) solution was prepared via the Fenton reaction ( • OH and OH ̄ were produced by the reaction between Fe 2+ and H 2 O 2 ), superoxide anion ( • O 2 ̄) was prepared by using 0.711 mg of KO 2 in 10 mL DMSO and other species (MnO 4 - , Cr 2 O 7 2- , HPO 4 2- , S 2 O 8 2- and NBS) were generated by dissolving their salts in water. For • OH sensing, the as-prepared Try-CsPbCl 3 PNCs were used to detect • OH radical and the fluorescence spectra of Try-CsPbCl 3 PNCs (1 mL) were investigated with different concentrations of • OH radical (0.5 mL). Briefly, 1.0 mM of • OH radical was generated by mixing Fe 2+ ion (1 mM) with H 2 O 2 (10%) at a volume ratio of 1:1. Different concentrations of • OH radical (0.01–500 µM, 0.5 mL) were treated separately with 1 mL of Try-CsPbCl 3 PNCs and then their emission spectra were recorded, leading to establish good calibration graph between the ratio of I 0 /I at 433 nm and concentrations of • OH radical. To ensure the selectivity of Try-CsPbCl 3 PNCs, different chemical species (MnO 4 - , Cr 2 O 7 2- , HPO 4 2- , S 2 O 8 2- , O 2 •- , and NBS) were added separately into Try-CsPbCl 3 PNCs solutions and examined their impact on the emission spectral intensities of Try-CsPbCl 3 PNCs. The fluorescence emission spectra of Try-CsPbCl 3 PNCs were recorded with λ Ex at 370 nm for • OH radical sensing and selectivity tests. The selectivity of Try-CsPbCl 3 PNCs toward • OH radical was evaluated by investigating the emission spectra of Try-CsPbCl 3 PNCs in the presence of various biomolecules (cysteine, arginine, tryptophan, and alanine, 500 µM), cations (Na + , Ca 2+ , Mg 2+ , Zn 2+ , Cu 2+ , and Fe 3+ , 500 µM) and anions (Cl - , I - , Br - , PO 4 3- , and SO 4 2- , 500 µM) with and without addition of • OH radical. 2.5. Fluorescence detection of • OH radical in water samples To apply potential application of the probe, tap and river waters from Tapi River, Surat, Gujarat, India were used in the present study. The water samples were filtered through a microfilter and then treated the sample with different concentrations (5.0, 10.0 and 25.0 µM) of • OH radical and then introduced 1 mL of Try-CsPbCl 3 PNCs, and vortexed for 2 min. The fluorescence emission intensities of Try-CsPbCl 3 PNCs at 433 nm were examined and the spectral studies were repeated three times and represented the statistical data as mean ± relative standard deviation (RSD). Scheme 1. Schematic illustration for Try-CsPbCl 3 PNCs synthesis and sensing of • OH. 3. Results and discussion 3.1. Synthesis and characterization of Try-CsPbCl 3 PNCs The synthesis pathway for the preparation of Try-CsPbCl 3 PNCs and their application for • OH radical sensing in aqueous medium were shown in Scheme 1. Initially, the influence of Try concentration (2–10 mg/mL) was studied on the fluorescence spectra of CsPbCl 3 PNCs using PbCl 3 (100 mM) and CsCl (100 mM) as precursors (Figure S1 ). Upon increasing Try concentration from 2.0 to 8.0 mg/mL, the intensity of fluorescence emission spectra of CsPbCl 3 PNCs was increased, however, the emission intensity was decreased by using 10 mg/mL of Try as an encapsulating agent. These results suggest that 8.0 mg/mL of Try was effectively enveloped CsPbCl 3 PNCs, thereby improving their dispersion ability in water with good fluorescence intensity. Similarly, we also investigated the optimum reaction time for the preparation of Try-CsPbCl 3 PNCs (Figure S2). It was clearly observed that the emission intensity of Try-CsPbCl 3 PNCs at 433 nm was increased with increasing reaction time from 3 to 12 h, after that the emission peak intensity was decreased, confirming the 12 h was found to optimum reaction time for the fabrication of Try-CsPbCl 3 PNCs. In order to confirm the origin of emission spectra, absorption and emission spectra of Try and Try-CsPbCl 3 PNCs were examined and shown in Figure S3. The spectral results demonstrated that Try did not show any emission peak, however, Try-encapsulated CsPbCl 3 PNCs displayed a characteristic emission peak at 433 nm (Figure S3a). Interestingly, pure Try and Try-CsPbCl 3 PNCs exhibited different absorption spectral characteristics (Figure S3b), indicating the formation of Try-CsPbCl 3 PNCs. After optimizing the reaction conditions, we examined the spectral characteristics, size, morphology, zeta potential and elemental composition of Try-CsPbCl 3 PNCs. The as-fabricated Try-CsPbCl 3 PNCs displayed λ max at 377 nm whereas the emission/excitation peaks exhibited at 433/370 nm (Fig. 1 ). The obtained spectral characteristics of Try-CsPbCl 3 PNCs were well matched with the reported method for CsPbCl 3 PNCs dispersed in organic solvent (hexane) [ 52 ]. Upon irradiation of Try-CsPbCl 3 PNCs solution with 365 nm of UV light, blue fluorescence was noticed, which confirms the formation of Try-CsPbCl 3 PNCs (Inset of Fig. 1 ). Furthermore, excitation-dependent emission spectral characteristics of Try-CsPbCl 3 PNCs were studied and shown in Figure S4. The emission spectral profiles of the as-prepared Try-CsPbCl 3 PNCs at different excitation wavelengths (300–400 nm) displayed the almost nonvariant nature in the emission spectra, however, the emission peak intensity was increased with increasing λ Ex from 300 to 370 nm, after that the intensity was decreased. The maximum emission intensity was noticed at λ Em = 433 nm upon excitation of Try-CsPbCl 3 PNCs at λ Ex = 370 nm. In addition, the fluorescence QY of Try-CsPbCl 3 PNCs was 17.26% and the lifetime of Try-CsPbCl 3 PNCs was τ = 1.64 ns (Figure S5). In order to apply Try-CsPbCl 3 PNCs as a promising fluorescence probe, it is essential to investigate the stability of Try-CsPbCl 3 PNCs. The stability of fabricated Try-CsPbCl 3 PNCs in water was examined by monitoring the emission spectral profiles at different time intervals (Figure S6). The emission spectral profiles of Try-CsPbCl 3 PNCs exhibited almost negligible changes up to 8 days, after that the emission peak intensity was decreased, which confirms that Try-CsPbCl 3 PNCs displayed good stability to use as a fluorescence probe for sensing applications. The FT-IR spectral profiles of pure Try and Try-CsPbCl 3 PNCs were studied and the data were depicted in Figure S7. The FT-IR spectrum of Try displayed a strong broad band in the range of 3600–3000 cm − 1 , which confirms to O-H and -N-H bonds stretching vibrations. The bands at 1650 and 1523 cm − 1 are ascribed to the Amide I and II bands of Try, respectively. Similarly, the stretching vibrations of COO − , C-N stretching and N-H bending were noticed at 1438, 1253 and 1519 cm − 1 , respectively. The characteristic FT-IR spectral profiles of Try were completely changed due to the encapsulation of CsPbCl 3 PNCs. It can be observed that a noticeable decrease in the intensity of broadband in the range of 3600 − 3000 cm − 1 for -OH and -NH 2 groups stretching and a drastic shift in the characteristic Amide bands confirm the backbone structural deformation in Try due to the formation of Try encapsulated CsPbCl 3 PNCs. Size and morphological analysis of the as-synthesized Try-CsPbCl 3 PNCs was further confirmed by using FE-TEM and dynamic light scattering (DLS) (Fig. 2 and Figure S8a). The as-prepared Try-CsPbCl 3 PNCs are nearly spherical shape with a mean size of 2.5 ± 0.5 nm, suggesting that Try-CsPbCl 3 PNCs are highly monodispersity, as it confirmed from histogram (Fig. 2 a-c). The hydrodynamic diameter of Try-CsPbCl 3 PNCs was 10.34 nm, displaying a higher size as compared to FETEM data due to the measurement of Try-CsPbCl 3 PNCs with water molecules. The as-fabricated Try-CsPbCl 3 PNCs exhibited a negative charge (-19.51 mV) (Figure S9a), which was confirmed by measuring zeta potential. The elemental spectral profile of Try-CsPbCl 3 PNCs was further investigated by XPS and the XPS survey spectrum of Try-CsPbCl 3 PNCs was shown in Fig. 3 a, indicating the as-synthesized Try-CsPbCl 3 PNCs contains C, O, Cs, Pb and Cl elements. Figure 3 b shows the Cs 3d high resolution (HR) spectrum, displaying two peaks at 726.7 eV and 740.7 eV are correspond to the Cs 3d 5/2 and Cs 3d 3/2 , respectively, confirming the presence of Cs + ion in Try-CsPbCl 3 PNCs. Figure 3 c represents the Pb 4f HR spectrum, showing the binding energies of Pb 4f 7/2 and Pb 4f 5/2 at 140.8 eV and 145.7 eV, respectively, which is indicative for the existence of Pb 2+ ion in Try-CsPbCl 3 PNCs. Similarly, the HR spectrum of Cl 2p displayed two peaks at 200.3 and 201.8 eV, indexing to the Cl 2p 3/2 and Cl 2p 1/2 binding energy levels, respectively (Fig. 3 d). Furthermore, the XRD pattern of Try-CsPbCl 3 PNCs is shown in Figure S10, exhibiting the diffraction peaks at 2θ = 15.9, 22.4, 32.2, 35.7, 43.12, 46.3 and 54.5 correspond to 100, 110, 200, 211, 220, 310 and 222 lattice planes of Try-CsPbCl 3 PNCs, which is well agreed with the XRD pattern of CsPbCl 3 PNCs [ 53 – 54 ]. The as-fabricated Try-CsPbCl 3 PNCs are in crystalline nature with high monodispersity. All the above data strongly support the formation of Try-CsPbCl 3 PNCs with monodispersity and good spectral characteristics, which explore them as potential probes for • OH sensing. 3.2. Fluorescence sensing of • OH radical To examine the fluorescence detection capability of Try-CsPbCl 3 PNCs toward • OH radical, several oxidizing and ROS (MnO 4 − , Cr 2 O 7 2− , HPO 4 2− , • OH, S 2 O 8 2− , O 2 •− , and NBS, 1 mM, 0.5 mL) were mixed with 1 mL of Try-CsPbCl 3 PNCs, separately, followed by vertexing the samples for a few minutes. The fluorescence emission spectra of the samples were evaluated (Fig. 4 ). As can be seen the emission spectra of Try-CsPbCl 3 PNCs in a significant decrease in the fluorescence emission intensity of Try-CsPbCl 3 PNCs was noticed in the presence of • OH radical, while no significant fluorescence quenching was noticed in the presence of other oxidizing and ROS. These emission spectral profiles demonstrated that the as-prepared Try-CsPbCl 3 PNCs could be utilized as a probe for fluorescence analysis of • OH radical in aqueous media. Moreover, the fluorescence color of Try-CsPbCl 3 PNCs solution with the addition of the above species was monitored under 365 nm of UV light, indicating the blue fluorescence of Try-CsPbCl 3 PNCs was almost non-fluorescent nature (Inset of Fig. 4 ), which signifies that Try-CsPbCl 3 PNCs act as a turn-off fluorescent probe for • OH radical sensing. The fluorescence emission signals of Try-CsPbCl 3 PNCs were evaluated in the presence of phosphate-buffered saline (PBS) with pH from 2.0 to 12.0 with and without • OH radical (Figure S11a), demonstrating that the addition of PBS pH (2.0–12.0) into Try-CsPbCl 3 PNCs did not affect the fluorescence spectra of Try-CsPbCl 3 PNCs. However, the maximum fluorescence emission quenching was observed with • OH radical at PBS of pHs 10 and 12 (Figure S11b), PBS of pH 10 was selected as an optimum pH for sensing of • OH radical using Try-CsPbCl 3 PNCs as a probe. 3.3. Fluorescence sensing mechanism In order to evaluate the fluorescence sensing mechanism of • OH radical using Try-CsPbCl 3 PNCs as a probe, several analytical techniques (TEM, DLS, zeta potential and lifetime) were examined. As can be seen in the FETEM image of Try-CsPbCl 3 PNCs with the addition of • OH radical (Fig. 2 d), the morphology and size of Try-CsPbCl 3 PNCs were drastically changed by the addition of • OH radical, indicating the deformation of Try-CsPbCl 3 PNCs by • OH radical, which leads to form Try-CsPbCl 3 nanoaggregates. Similarly, the hydrodynamic diameter of Try-CsPbCl 3 PNCs was significantly increased to 21.5 nm by introducing • OH radical (Figure S8b), leading to destabilization of Try-CsPbCl 3 PNCs, which resulted in quenching the fluorescence of Try-CsPbCl 3 PNCs. The zeta potential of Try-CsPbCl 3 PNCs was − 19.51 mV, however, it was increased to -28.23 mV. Interestingly, the negligible change (from 1.64 to 1.51 ns) in the lifetime of Try-CsPbCl 3 PNCs was observed with the addition of • OH radical, indicating the static quenching mechanism (Figure S5). Further, the characteristic FT-IR spectral profiles of Try-CsPbCl 3 PNCs were significantly changed upon the addition of • OH radical (Figure S12), suggesting the structural changes in the Try-CsPbCl 3 PNCs by • OH radical. To further evaluate the inner filter effect (IFE), the fluorescence (excitation and emission) spectra of Try-CsPbCl 3 PNCs and the absorption spectrum of • OH radical were studied (Figure S13), revealing the overlapping of absorption spectrum of • OH radical with excitation spectrum of Try-CsPbCl 3 PNCs, which confirms the IFE. Thus, the as-prepared Try-CsPbCl 3 PNCs act as a turn-off fluorescence probe for the detection of • OH radical. 3.4. Sensitivity Under the optimal conditions, the variations in the fluorescence intensity of Try-CsPbCl 3 PNCs were investigated by adding different concentrations of • OH radical (0.01–500 µM). As can be noticed in Fig. 5 , the fluorescence emission intensity of Try-CsPbCl 3 PNCs centered at 433 nm was gradually quenched along with increasing concentration of • OH radical. Then, the emission intensity of Try-CsPbCl 3 PNCs was quenched 77% when • OH radical at 500 µM. Figure S14 displayed the constructed plot of the ratio I 0 /I (where I 0 and I represent the emission intensity of Try-CsPbCl 3 PNCs in the absence and presence of • OH radical, respectively) against • OH radical concentration (0.01–500 µM). Furthermore, with • OH radical concentration in the range of 0.01-5.0 µM, the fluorescence quenching efficiency showed the linear fitting equation of y = 0.2596x + 1.2061 ( R 2 = 0.9795). The LOD was 3.10 nM (3σ/s, where “s” is the slope of the calibration curve and “σ” is the standard deviation of the blank) for • OH radical. The analytical characteristics of the developed Try-CsPbCl 3 PNCs-based fluorescence approach were compared with other reported methods [ 47 , 55 – 62 ] for • OH radical sensing (Table S1 ), revealing the developed probe exhibited superior and a comparable analytical performance with other analytical techniques for • OH radical sensing. Furthermore, the developed Try-CsPbCl 3 PNCs-based turn-off fluorescence strategy exhibits several analytical features such as free surface modification, simple chemical routes, good selectivity and sensitivity, and good stability in aqueous phase, which allows for use as a promising fluorescence probe for • OH radical sensing. 3.5. Selectivity The sensing selectivity of Try-CsPbCl 3 PNCs toward • OH radical was investigated in the presence of various metal ions, ROS, and biomolecules phosphate buffer (20 mM, pH 7.4). As shown in Figure S15, the fluorescence behavior of Try-CsPbCl 3 PNCs in the presence of biomolecules (cysteine, arginine, tryptophan, and alanine, 500 µM), cations (Na + , Ca 2+ , Mg 2+ , Zn 2+ , Cu 2+ , and Fe 3+ , 500 µM) and anions (Cl − , I − , Br − , PO 4 3− , and SO 4 2− , 500 µM) is almost same and did not show any significant changes. As anticipated, Try-CsPbCl 3 PNCs exhibited a remarkable fluorescence quenching only with • OH radical even in the existence of the other interfering chemical species, revealing that the as-prepared Try-CsPbCl 3 PNCs stand out as a highly selective turn-off fluorescence probe for • OH radical sensing. 3.6. Analysis of • OH radical in water samples To evaluate the practical application of Try-CsPbCl 3 PNCs in monitoring • OH radical in real samples, the collected water (tap and river) samples were filtered and subsequently added different concentrations of • OH radical (5.0, 10.0 and 25.0 µM). Then, • OH radical treated water samples were added into Try-CsPbCl 3 PNCs, and their concentrations were estimated by the aforesaid procedure. From Table S2, it can be noticed that the recovery rates of • OH radical in water samples were 99.00–101.40% with a relative standard deviation of < 2.0%. The results demonstrate that Try-CsPbCl 3 PNCs could be used as the potential fluorescence turn-off probe for the detection of • OH radical in real samples. 4. Conclusions In summary, a simple analytical tool was developed for sensing • OH radical using water-dispersible Try-CsPbCl 3 PNCs as a turn-off fluorescence probe. The as-fabricated Try-CsPbCl 3 PNCs exhibited blue fluorescence under UV irradiation (λ ∼ 365 nm) and displayed λ Em/Ex = 433/370 nm. The developed Try-CsPbCl 3 PNCs-based fluorescence approach has a highly selective and sensitive response toward • OH radical with a good linearity in the concentration range of 0.01-5.0 µM, which achieves the LOD of 3.10 nM. Importantly, Try-CsPbCl 3 PNCs exhibit superior selectivity for • OH radical with virtually no fluorescence quenching by other interfering chemical species (ROS, metal ions, anions and biomolecules). Furthermore, Try-CsPbCl 3 PNCs-based analytical approach was successfully applied to detect • OH radical in water samples. Thus, the as-synthesized Try-CsPbCl 3 PNCs could be successfully integrated with fluorescence spectrometry for the detection of • OH radical in real samples. Declarations Acknowledgements SKK thanks the Director, Sardar Vallabhbhai National Institute of Technology, Surat for grating the sabbatical leave for Brainpool Programme at Chung-Ang University, South Korea. This research was supported by the Brain Pool program funded by the Ministry of Science and ICT through the National Research Foundation of Korea (RS-2023-00221526). Ethical Approval Not applicable Consent to participate Not applicable Consent to publish Not applicable Funding SKK thanks the Director, Sardar Vallabhbhai National Institute of Technology, Surat for grating the sabbatical leave for Brainpool Programme at Chung-Ang University, South Korea. This research was supported by the Brain Pool program funded by the Ministry of Science and ICT through the National Research Foundation of Korea (RS-2023-00221526). Availability of data and materials Data will be available from corresponding author on demand. Consent for publication Not Applicable. Conflict of interest All the authors declare that there is no conflict of interest. Authors' contributions Suresh Kumar Kailasa : Methodology, Formal analysis, Writing—original draft. Kartik Pankajbhai Makwana : Material preparation, Formal analysis, Writing—Review and Editing. Madhura Pradeep Deshpande : Formal analysis, Data collection and analysis, Writing—Review and Editing. 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Anal Chem 86(3):1829–1836 Xie Y, Xianyu Y, Wang N, Yan Z, Liu Y, Zhu K, Hatzakis NS (2018) Jiang Functionalized gold nanoclusters identify highly reactive oxygen species in living organisms. Adv Funct Mater 28:1702026 Scheme Scheme 1 is available in the Supplementary Files section. Additional Declarations No competing interests reported. Supplementary Files SureshetalSupportingInformationofTrypsinCsPbCl3FNCs.pdf floatimage1.png Scheme 1. Schematic illustration for Try-CsPbCl 3 PNCs synthesis and sensing of • OH. Cite Share Download PDF Status: Published Journal Publication published 10 Mar, 2025 Read the published version in Microchimica Acta → Version 1 posted Editorial decision: Revision requested 15 Dec, 2024 Reviews received at journal 12 Dec, 2024 Reviews received at journal 30 Nov, 2024 Reviewers agreed at journal 28 Nov, 2024 Reviewers agreed at journal 24 Nov, 2024 Reviewers invited by journal 18 Nov, 2024 Editor assigned by journal 13 Nov, 2024 Submission checks completed at journal 13 Nov, 2024 First submitted to journal 11 Nov, 2024 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. 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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-5436012","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":384798763,"identity":"911483f9-2a09-4c8e-a39f-a332b94343e6","order_by":0,"name":"Suresh Kumar Kailasa","email":"","orcid":"","institution":"Sardar Vallabhbhai National Institute of Technology","correspondingAuthor":false,"prefix":"","firstName":"Suresh","middleName":"Kumar","lastName":"Kailasa","suffix":""},{"id":384798764,"identity":"19096f12-b4bc-44fe-bb89-e04ad7a9214e","order_by":1,"name":"Kartik Pankajbhai Makwana","email":"","orcid":"","institution":"Sardar Vallabhbhai National Institute of Technology","correspondingAuthor":false,"prefix":"","firstName":"Kartik","middleName":"Pankajbhai","lastName":"Makwana","suffix":""},{"id":384798767,"identity":"e76b4e84-c549-4c41-a040-8f9184d0bc95","order_by":2,"name":"Madhura Pradeep Deshpande","email":"","orcid":"","institution":"Chung-Ang University","correspondingAuthor":false,"prefix":"","firstName":"Madhura","middleName":"Pradeep","lastName":"Deshpande","suffix":""},{"id":384798772,"identity":"8415af83-3462-448b-b47e-5ce13d92aaed","order_by":3,"name":"Yoojin Choi","email":"","orcid":"","institution":"Chung-Ang University","correspondingAuthor":false,"prefix":"","firstName":"Yoojin","middleName":"","lastName":"Choi","suffix":""},{"id":384798773,"identity":"e552e71d-6c72-4d97-92b4-0ed9d42fe6b1","order_by":4,"name":"Tae Jung Park","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAwElEQVRIiWNgGAWjYFCCAyBsA+MlENbAA9GSRpIWsE2HSdBiz3j24OeKM+fl+WckMH74wZCWT4Qt55Ilz9y4bTjjRgKzZA9DjmUDYS1nDCQbPtxOYLiRwCDNwFBhQIQtZ4x/Nnw4lyAPtOU3sVrMJBtuHEgwuJHABrQlhwgtB86lWTacSTbceOZhm2WPQRphLewzzh6+2XDMTl7uePLhGz8qkglrYZA4A2MxNjAwEKGBgYG/hxhVo2AUjIJRMKIBAE2nPiyfRkGKAAAAAElFTkSuQmCC","orcid":"","institution":"Chung-Ang University","correspondingAuthor":true,"prefix":"","firstName":"Tae","middleName":"Jung","lastName":"Park","suffix":""}],"badges":[],"createdAt":"2024-11-12 04:23:09","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5436012/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5436012/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s00604-025-07070-8","type":"published","date":"2025-03-10T15:57:46+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":71687070,"identity":"2f30aaef-5ce5-4869-8533-2c81483ccda4","added_by":"auto","created_at":"2024-12-17 17:33:53","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":145351,"visible":true,"origin":"","legend":"\u003cp\u003eAbsorption (λ\u003csub\u003emax\u003c/sub\u003e = 377 nm), fluorescence excitation and emission (λ\u003csub\u003eEx \u003c/sub\u003e= 370 nm; λ\u003csub\u003eEm\u003c/sub\u003e = 433 nm) spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs (Inset: Digital images of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in daylight and under UV light at 365 nm).\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/a4b7afa71fa9aeba67a2958d.png"},{"id":71686580,"identity":"80eca19c-9007-4feb-9366-8cc22ca39390","added_by":"auto","created_at":"2024-12-17 17:25:54","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1249333,"visible":true,"origin":"","legend":"\u003cp\u003eHR-TEM pictures of Try-CsPbCl\u003csub\u003e3 \u003c/sub\u003ePNCs with scale bars of (a) 50 nm and (b) 20 nm. (c) The size distribution of Try-CsPbCl\u003csub\u003e3 \u003c/sub\u003ePNCs. (d) HR-TEM picture of Try-CsPbCl\u003csub\u003e3 \u003c/sub\u003ePNCs with \u003csup\u003e•\u003c/sup\u003eOH.\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/346f1e3d1f092e7399dc0e78.png"},{"id":71686581,"identity":"da5d8168-7640-4e00-b2a2-5d8f7b7f3641","added_by":"auto","created_at":"2024-12-17 17:25:54","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":69725,"visible":true,"origin":"","legend":"\u003cp\u003eXPS characterization of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs: (a) survey spectrum, HR spectra of (b) Cs 3d, (c) Pb 4f, and (d) Cl 2p.\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/fecc91a0904d80abd816fc84.png"},{"id":71686583,"identity":"37cd29bc-ac01-443d-a9b4-4d5c1aa55d5f","added_by":"auto","created_at":"2024-12-17 17:25:54","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":194777,"visible":true,"origin":"","legend":"\u003cp\u003eFluorescence emission spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs after adding different ROS (MnO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e\u003csub\u003e, \u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, HPO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, \u003csup\u003e•\u003c/sup\u003eOH, S\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, O\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e •-\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e and NBS). Inset picture of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs after the addition of various ROS (1. MnO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e\u003csub\u003e, \u003c/sub\u003e2. Cr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, 3. HPO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, 4. \u003csup\u003e•\u003c/sup\u003eOH, 5. S\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, 6. O\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e •-\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e and 7. NBS) under UV light (365 nm).\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/1cc2ec01c8faa751e8e6f6a5.png"},{"id":71686584,"identity":"d2e13ed5-1b6a-4059-b6c8-4b260830d9ee","added_by":"auto","created_at":"2024-12-17 17:25:54","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":239937,"visible":true,"origin":"","legend":"\u003cp\u003eFluorescence spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in the existence of various concentrations of \u003csup\u003e•\u003c/sup\u003eOH (0.01–500 µM). Inset picture demonstrates the variations in the blue fluorescence of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs with the addition of \u003csup\u003e•\u003c/sup\u003eOH (0.01–500 µM) under UV light at 365 nm.\u003c/p\u003e","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/04a408e891b729b3d6d14863.png"},{"id":78689209,"identity":"37d7d7b4-f28e-4059-b927-8574de69d837","added_by":"auto","created_at":"2025-03-17 16:12:22","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2951489,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/e99ad5e2-1a7e-4199-bfc9-fa758d138759.pdf"},{"id":71686631,"identity":"7e981aa0-52cd-41a2-a48c-c2b8ecf7ac7a","added_by":"auto","created_at":"2024-12-17 17:26:03","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":731970,"visible":true,"origin":"","legend":"","description":"","filename":"SureshetalSupportingInformationofTrypsinCsPbCl3FNCs.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/85bd3b0ba453b06bc9137021.pdf"},{"id":71686579,"identity":"6c536805-d7a9-4b6d-ab2a-96fa42892d90","added_by":"auto","created_at":"2024-12-17 17:25:54","extension":"png","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":649654,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 1. \u003c/strong\u003eSchematic illustration for Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs synthesis and sensing of \u003csup\u003e•\u003c/sup\u003eOH.\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-5436012/v1/4cd35481728b0b00af5819d7.png"}],"financialInterests":"No competing interests reported.","formattedTitle":"Synthesis of trypsin protected CsPbCl3 fluorescent nanocrystals for hydroxyl radical sensing","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eIn recent years, lead halide perovskite nanocrystals (PNCs) have emerged as outstanding materials in fabricating solar cells, sensors, light-emitting diodes and optoelectronic devices because of their remarkable characteristics including superior quantum yield (QY), narrow full width at half-maximum, impressive charge transport capabilities, high photocatalytic properties and tunable emission wavelength in complete visible and near-infrared region [\u003cspan additionalcitationids=\"CR2 CR3 CR4\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Interestingly, it was noticed that perovskite oxides are quite stable in water whereas lead halide PNCs are highly unstable in water and easily degraded by exposing them to heat, light and moisture [\u003cspan additionalcitationids=\"CR7\" citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e], resulting in limiting their promising applications in multidisciplinary research.\u003c/p\u003e \u003cp\u003eTo improve lead halide PNCs water stability, several researchers have introduced different strategies to fabricate lead halide PNCs including lead chloride PNCs with high water stability. For example, perflorocompounds were used as ligands for the preparation of water-stable CsPbBr\u003csub\u003e3\u003c/sub\u003e@Cs\u003csub\u003e4\u003c/sub\u003ePbBr\u003csub\u003e6\u003c/sub\u003e PNCs [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Furthermore, various polymers (poly(l-lactide) polypropylene glycol, and polysulfone [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e], NH\u003csub\u003e2\u003c/sub\u003e-PEG-COOH [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e], NH\u003csub\u003e2\u003c/sub\u003e group terminated hiperbranched polymer [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e], polymethyl methacrylate [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], polystyrene [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e], polystyrene-cetyltrimethylammonium bromide [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e] and polyethylene glycol [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]), organic molecules (ethylenediaminetetraacetic acid [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e], bolaamphiphilic ligand (NKE-12) [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], adamantane-1-amine [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e], (4,4'-bipyridine and 2,2'-bipyridine) [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e], glycyrrhizic acid [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e], 4-bromo-butyric acid-oleylamine [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e], oleylamine-oleic acid [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e] and oleic acid-3-bromopropionic acid [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]) and inorganic salts and compounds (cesium trifluoroacetate [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e], MAPbBr\u003csub\u003e3\u003c/sub\u003e@lead laurate [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e], mesoporous silica [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e], Al\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e3\u003c/sub\u003e [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e], metal-organic frameworks [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e], ZrO\u003csub\u003e2\u003c/sub\u003e [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] and TiO\u003csub\u003e2\u003c/sub\u003e [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]), respectively. The above approaches were successfully produced high stability of lead halide PNCs without losing their fluorescence properties, suggesting encapsulation of lead halide PNCs with suitable ligand chemistry offers several features such as water stability, good QY and superior optical properties. Furthermore, it is a very challenging task to design water- and air-stable lead chloride PNCs without the use of complicated synthetic approaches as well as ligand chemistry. In order to alter the surface chemistry and fluorescence properties of CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, trypsin (Try) was explored as a bioligand for the preparation of water-stable CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in the sensing of hydroxyl radical.\u003c/p\u003e \u003cp\u003eReactive oxygen species (ROS) are produced by mitochondria in the cells, exhibiting a high reactive nature [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. The ROS (oxygen-containing radicals - superoxide, peroxyl, hydroxyl (\u003csup\u003e\u0026bull;\u003c/sup\u003eOH) and hydroperoxyl) and non-radical agents ozone, HOCl and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e which are easily converted into ROS) play a key role in numerous biochemical pathways in the cells [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. It was observed that certain levels of ROS efficiently enhance cellular functions (migration, proliferation and differentiation) [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. However, ROS have potentially induced oxidative stress and cell damage, which yields cell death [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. Among ROS, hydroxyl radical is recognized as one of the highly reactive ROS, exhibiting a lifetime in the nanoseconds [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Usually, hydroxyl radical is produced in \u003cem\u003ein vivo\u003c/em\u003e via oxygen molecule oxidation to superoxide, higher levels of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH cause oxidative damage to various bio-macromolecules (nucleic acids, lipids, carbohydrates and proteins, demonstrating that significant attention must be paid to monitoring of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH in cells and various environments. In order to identify \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, several analytical techniques such as electrochemical, electron spin resonance, fluorescence, and UV\u0026ndash;visible spectroscopic and high-performance liquid chromatographic techniques have been applied to detect ROS including \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical [\u003cspan additionalcitationids=\"CR39 CR40 CR41 CR42 CR43 CR44 CR45\" citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e]. Importantly, several fluorescent probes including terbium complexes [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e], Coumarin\u0026ndash;Neutral Red [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e], copper and molybdenum nanoclusters [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e], Ag\u0026ndash;Au nanocages [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e], carbon dot-based hydrogel [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e] and dihydroquinolines have been utilized as fluorescence probes for sensing of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical with lower limit of detection (LOD) [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e]. The nanomaterials- and organic molecules-based fluorescence probes have proven to be promising readers to selective assay of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH in biological and environmental systems. Due to their complex procedures in fabricating fluorescence probes, there is a still necessity to introduce a facile and novel fluorescence probe for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH sensing in water samples.\u003c/p\u003e \u003cp\u003eIn this work, we report a simple and selective hydroxyl radical (\u003csup\u003e\u0026bull;\u003c/sup\u003eOH) sensing strategy using aqueous-stable trypsin (Try) encapsulated cesium lead chloride (CsPbCl\u003csub\u003e3\u003c/sub\u003e) PNCs as a nanoprobe (Scheme 1). The as-synthesized Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were stable in aqueous medium and displayed spherical shape morphology with a mean size of 2.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.5 nm. Further, Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited blue fluorescence under UV lamp (365 nm), showing λ\u003csub\u003eEm/Ex\u003c/sub\u003e at 433/370 nm, which offers a QY of 17.26%. Noticeably, the emission intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was quenched by hydroxyl radical, leading to the development of a fluorescence turn-off approach for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH assay with an LOD of 3.10 nM. The developed sensing strategy was used to detect \u003csup\u003e\u0026bull;\u003c/sup\u003eOH in water samples, demonstrating Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs-based fluorescence method could be an effective tool for monitoring \u003csup\u003e\u0026bull;\u003c/sup\u003eOH in real samples.\u003c/p\u003e"},{"header":"2. Experimental section","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Materials and instruments\u003c/h2\u003e \u003cp\u003eLead (II) chloride (PbCl\u003csub\u003e2\u003c/sub\u003e, 98%) and cesium chloride (CsCl, 99.99%) were produced from SRL and BLD pharm, respectively. 1 -Octadecene (ODE), potassium superoxide (KO\u003csub\u003e2\u003c/sub\u003e) were obtained from Sigma Aldrich, N-nromo succinimide (NBS), sodium hypochlorite (NaOCl) and hydrogen peroxide (H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e) were purchased from SRL, FINAR and SDFCL Chemicals, respectively. Deionized water was used for preparation of solutions and sensing experiments, and analytical-grade chemicals were utilized without any further purification.\u003c/p\u003e \u003cp\u003eThe as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs size and morphology were investigated using field emission transmission electron microscopy (FETEM) (JEOL-200, Tokyo, Japan). Fluorescence (emission and excitation) spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were examined by a Cary Eclipse fluorescence spectrometer (Agilent Technologies, Santa Clara, CA, USA). UV\u0026ndash;vis absorption spectra were recorded with a Maya Pro 2000 spectrophotometer (Ocean Optics, Orlando, FL, USA). Infrared spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were examined an ALPHA II Fourier transform infrared (FT-IR) spectrometer (Bruker, Billerica, MA, USA). X-ray diffraction (XRD) spectrum of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was recorded using a D8-Advance Instrument (Bruker AXS). X-ray photoelectron spectroscopy (XPS) (K-alpha+, Thermo Fisher Scientific, Waltham, MA, USA) was performed for the confirmation of the elemental composition of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. Hydrodynamic diameter and zeta potentials of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were obtained using a NanoZS90 nano-particle Size potential analyzer (Malvern, UK).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Synthesis of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs\u003c/h2\u003e \u003cp\u003eBlue fluorescence water-stable Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were prepared \u003cem\u003evia\u003c/em\u003e a simple reaction. Firstly, 100 mM (33.6 mg) of PbCl\u003csub\u003e2\u003c/sub\u003e and 100 mM (55.6 mg) of CsCl precursors were dispersed in 2 mL of ODE separately and stirred for 30 minutes at room temperature. Then, the preparation of CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was initiated by mixing both solutions in 10 mL of reaction flask. The mixture was stirred at room temperature for 3 h. The formed CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were capped with Try by adding Try (32 mg) into CsPbCl\u003csub\u003e3\u003c/sub\u003e solution and then stirred for 12 h, triggering the formation of blue fluorescent Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. The formed Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were washed with hexane and then dispersed in water for sensing applications.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e\u003cb\u003e2.4. Fluorescence sensing of\u003c/b\u003e \u003csup\u003e\u003cb\u003e\u0026bull;\u003c/b\u003e\u003c/sup\u003e\u003cb\u003eOH using Try-CsPbCl\u003c/b\u003e\u003csub\u003e\u003cb\u003e3\u003c/b\u003e\u003c/sub\u003e \u003cb\u003ePNCs as a turn-off probe\u003c/b\u003e\u003c/h2\u003e \u003cp\u003eIn the sensing study, the following reactive species solutions were prepared as follows, hydroxyl radical (\u003csup\u003e\u0026bull;\u003c/sup\u003eOH) solution was prepared \u003cem\u003evia\u003c/em\u003e the Fenton reaction (\u003csup\u003e\u0026bull;\u003c/sup\u003eOH and OH ̄ were produced by the reaction between Fe\u003csup\u003e2+\u003c/sup\u003e and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e), superoxide anion (\u003csup\u003e\u0026bull;\u003c/sup\u003eO\u003csub\u003e2\u003c/sub\u003ē) was prepared by using 0.711 mg of KO\u003csub\u003e2\u003c/sub\u003e in 10 mL DMSO and other species (MnO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e Cr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, HPO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, S\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e and NBS) were generated by dissolving their salts in water. For \u003csup\u003e\u0026bull;\u003c/sup\u003eOH sensing, the as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were used to detect \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical and the fluorescence spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs (1 mL) were investigated with different concentrations of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (0.5 mL). Briefly, 1.0 mM of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical was generated by mixing Fe\u003csup\u003e2+\u003c/sup\u003e ion (1 mM) with H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e (10%) at a volume ratio of 1:1. Different concentrations of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (0.01\u0026ndash;500 \u0026micro;M, 0.5 mL) were treated separately with 1 mL of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs and then their emission spectra were recorded, leading to establish good calibration graph between the ratio of I\u003csub\u003e0\u003c/sub\u003e/I at 433 nm and concentrations of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical. To ensure the selectivity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, different chemical species (MnO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e Cr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, HPO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, S\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e, O\u003csub\u003e2\u003c/sub\u003e \u003csup\u003e\u0026bull;-\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e and NBS) were added separately into Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs solutions and examined their impact on the emission spectral intensities of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. The fluorescence emission spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were recorded with λ\u003csub\u003eEx\u003c/sub\u003e at 370 nm for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing and selectivity tests. The selectivity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs toward \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical was evaluated by investigating the emission spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in the presence of various biomolecules (cysteine, arginine, tryptophan, and alanine, 500 \u0026micro;M), cations (Na\u003csup\u003e+\u003c/sup\u003e, Ca\u003csup\u003e2+\u003c/sup\u003e, Mg\u003csup\u003e2+\u003c/sup\u003e, Zn\u003csup\u003e2+\u003c/sup\u003e, Cu\u003csup\u003e2+\u003c/sup\u003e, and Fe\u003csup\u003e3+\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e 500 \u0026micro;M) and anions (Cl\u003csup\u003e-\u003c/sup\u003e, I\u003csup\u003e-\u003c/sup\u003e, Br\u003csup\u003e-\u003c/sup\u003e, PO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e3-\u003c/sup\u003e, and SO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2-\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e 500 \u0026micro;M) with and without addition of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.5. Fluorescence detection of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in water samples\u003c/h2\u003e \u003cp\u003eTo apply potential application of the probe, tap and river waters from Tapi River, Surat, Gujarat, India were used in the present study. The water samples were filtered through a microfilter and then treated the sample with different concentrations (5.0, 10.0 and 25.0 \u0026micro;M) of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical and then introduced 1 mL of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, and vortexed for 2 min. The fluorescence emission intensities of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs at 433 nm were examined and the spectral studies were repeated three times and represented the statistical data as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;relative standard deviation (RSD).\u003c/p\u003e \u003cp\u003e \u003cb\u003eScheme 1.\u003c/b\u003e Schematic illustration for Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs synthesis and sensing of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results and discussion","content":"\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e3.1. Synthesis and characterization of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs\u003c/h2\u003e \u003cp\u003eThe synthesis pathway for the preparation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs and their application for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing in aqueous medium were shown in Scheme 1. Initially, the influence of Try concentration (2\u0026ndash;10 mg/mL) was studied on the fluorescence spectra of CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs using PbCl\u003csub\u003e3\u003c/sub\u003e (100 mM) and CsCl (100 mM) as precursors (Figure \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). Upon increasing Try concentration from 2.0 to 8.0 mg/mL, the intensity of fluorescence emission spectra of CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was increased, however, the emission intensity was decreased by using 10 mg/mL of Try as an encapsulating agent. These results suggest that 8.0 mg/mL of Try was effectively enveloped CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, thereby improving their dispersion ability in water with good fluorescence intensity. Similarly, we also investigated the optimum reaction time for the preparation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs (Figure S2). It was clearly observed that the emission intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs at 433 nm was increased with increasing reaction time from 3 to 12 h, after that the emission peak intensity was decreased, confirming the 12 h was found to optimum reaction time for the fabrication of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. In order to confirm the origin of emission spectra, absorption and emission spectra of Try and Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were examined and shown in Figure S3. The spectral results demonstrated that Try did not show any emission peak, however, Try-encapsulated CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs displayed a characteristic emission peak at 433 nm (Figure S3a). Interestingly, pure Try and Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited different absorption spectral characteristics (Figure S3b), indicating the formation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs.\u003c/p\u003e \u003cp\u003eAfter optimizing the reaction conditions, we examined the spectral characteristics, size, morphology, zeta potential and elemental composition of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. The as-fabricated Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs displayed λ\u003csub\u003emax\u003c/sub\u003e at 377 nm whereas the emission/excitation peaks exhibited at 433/370 nm (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The obtained spectral characteristics of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were well matched with the reported method for CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs dispersed in organic solvent (hexane) [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. Upon irradiation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs solution with 365 nm of UV light, blue fluorescence was noticed, which confirms the formation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs (Inset of Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Furthermore, excitation-dependent emission spectral characteristics of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were studied and shown in Figure S4. The emission spectral profiles of the as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs at different excitation wavelengths (300\u0026ndash;400 nm) displayed the almost nonvariant nature in the emission spectra, however, the emission peak intensity was increased with increasing λ\u003csub\u003eEx\u003c/sub\u003e from 300 to 370 nm, after that the intensity was decreased. The maximum emission intensity was noticed at λ\u003csub\u003eEm\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;433 nm upon excitation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs at λ\u003csub\u003eEx\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;370 nm. In addition, the fluorescence QY of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was 17.26% and the lifetime of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was τ\u0026thinsp;=\u0026thinsp;1.64 ns (Figure S5). In order to apply Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs as a promising fluorescence probe, it is essential to investigate the stability of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. The stability of fabricated Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in water was examined by monitoring the emission spectral profiles at different time intervals (Figure S6). The emission spectral profiles of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited almost negligible changes up to 8 days, after that the emission peak intensity was decreased, which confirms that Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs displayed good stability to use as a fluorescence probe for sensing applications.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe FT-IR spectral profiles of pure Try and Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were studied and the data were depicted in Figure S7. The FT-IR spectrum of Try displayed a strong broad band in the range of 3600\u0026ndash;3000 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, which confirms to O-H and -N-H bonds stretching vibrations. The bands at 1650 and 1523 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e are ascribed to the Amide I and II bands of Try, respectively. Similarly, the stretching vibrations of COO\u003csup\u003e\u0026minus;\u003c/sup\u003e, C-N stretching and N-H bending were noticed at 1438, 1253 and 1519 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, respectively. The characteristic FT-IR spectral profiles of Try were completely changed due to the encapsulation of CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. It can be observed that a noticeable decrease in the intensity of broadband in the range of 3600\u0026thinsp;\u0026minus;\u0026thinsp;3000 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e for -OH and -NH\u003csub\u003e2\u003c/sub\u003e groups stretching and a drastic shift in the characteristic Amide bands confirm the backbone structural deformation in Try due to the formation of Try encapsulated CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. Size and morphological analysis of the as-synthesized Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was further confirmed by using FE-TEM and dynamic light scattering (DLS) (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e and Figure S8a). The as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs are nearly spherical shape with a mean size of 2.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.5 nm, suggesting that Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs are highly monodispersity, as it confirmed from histogram (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea-c). The hydrodynamic diameter of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was 10.34 nm, displaying a higher size as compared to FETEM data due to the measurement of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs with water molecules. The as-fabricated Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited a negative charge (-19.51 mV) (Figure S9a), which was confirmed by measuring zeta potential.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe elemental spectral profile of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was further investigated by XPS and the XPS survey spectrum of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea, indicating the as-synthesized Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs contains C, O, Cs, Pb and Cl elements. Figure\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb shows the Cs 3d high resolution (HR) spectrum, displaying two peaks at 726.7 eV and 740.7 eV are correspond to the Cs 3d\u003csub\u003e5/2\u003c/sub\u003e and Cs 3d\u003csub\u003e3/2\u003c/sub\u003e, respectively, confirming the presence of Cs\u003csup\u003e+\u003c/sup\u003e ion in Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. Figure\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec represents the Pb 4f HR spectrum, showing the binding energies of Pb 4f\u003csub\u003e7/2\u003c/sub\u003e and Pb 4f\u003csub\u003e5/2\u003c/sub\u003e at 140.8 eV and 145.7 eV, respectively, which is indicative for the existence of Pb\u003csup\u003e2+\u003c/sup\u003e ion in Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. Similarly, the HR spectrum of Cl 2p displayed two peaks at 200.3 and 201.8 eV, indexing to the Cl 2p\u003csub\u003e3/2\u003c/sub\u003e and Cl 2p\u003csub\u003e1/2\u003c/sub\u003e binding energy levels, respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ed). Furthermore, the XRD pattern of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs is shown in Figure S10, exhibiting the diffraction peaks at 2θ\u0026thinsp;=\u0026thinsp;15.9, 22.4, 32.2, 35.7, 43.12, 46.3 and 54.5 correspond to 100, 110, 200, 211, 220, 310 and 222 lattice planes of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, which is well agreed with the XRD pattern of CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e]. The as-fabricated Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs are in crystalline nature with high monodispersity. All the above data strongly support the formation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs with monodispersity and good spectral characteristics, which explore them as potential probes for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH sensing.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e3.2. Fluorescence sensing of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical\u003c/h2\u003e \u003cp\u003eTo examine the fluorescence detection capability of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs toward \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, several oxidizing and ROS (MnO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e Cr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003csup\u003e2\u0026minus;\u003c/sup\u003e, HPO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2\u0026minus;\u003c/sup\u003e, \u003csup\u003e\u0026bull;\u003c/sup\u003eOH, S\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e8\u003c/sub\u003e\u003csup\u003e2\u0026minus;\u003c/sup\u003e, O\u003csub\u003e2\u003c/sub\u003e \u003csup\u003e\u0026bull;\u0026minus;\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e and NBS, 1 mM, 0.5 mL) were mixed with 1 mL of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, separately, followed by vertexing the samples for a few minutes. The fluorescence emission spectra of the samples were evaluated (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). As can be seen the emission spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in a significant decrease in the fluorescence emission intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was noticed in the presence of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, while no significant fluorescence quenching was noticed in the presence of other oxidizing and ROS. These emission spectral profiles demonstrated that the as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs could be utilized as a probe for fluorescence analysis of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in aqueous media. Moreover, the fluorescence color of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs solution with the addition of the above species was monitored under 365 nm of UV light, indicating the blue fluorescence of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was almost non-fluorescent nature (Inset of Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e), which signifies that Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs act as a turn-off fluorescent probe for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing. The fluorescence emission signals of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were evaluated in the presence of phosphate-buffered saline (PBS) with pH from 2.0 to 12.0 with and without \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (Figure S11a), demonstrating that the addition of PBS pH (2.0\u0026ndash;12.0) into Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs did not affect the fluorescence spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. However, the maximum fluorescence emission quenching was observed with \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical at PBS of pHs 10 and 12 (Figure S11b), PBS of pH 10 was selected as an optimum pH for sensing of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical using Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs as a probe.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e3.3. Fluorescence sensing mechanism\u003c/h2\u003e \u003cp\u003eIn order to evaluate the fluorescence sensing mechanism of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical using Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs as a probe, several analytical techniques (TEM, DLS, zeta potential and lifetime) were examined. As can be seen in the FETEM image of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs with the addition of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ed), the morphology and size of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were drastically changed by the addition of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, indicating the deformation of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs by \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, which leads to form Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e nanoaggregates. Similarly, the hydrodynamic diameter of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was significantly increased to 21.5 nm by introducing \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (Figure S8b), leading to destabilization of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, which resulted in quenching the fluorescence of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs. The zeta potential of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was \u0026minus;\u0026thinsp;19.51 mV, however, it was increased to -28.23 mV. Interestingly, the negligible change (from 1.64 to 1.51 ns) in the lifetime of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was observed with the addition of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, indicating the static quenching mechanism (Figure S5). Further, the characteristic FT-IR spectral profiles of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were significantly changed upon the addition of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (Figure S12), suggesting the structural changes in the Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs by \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical. To further evaluate the inner filter effect (IFE), the fluorescence (excitation and emission) spectra of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs and the absorption spectrum of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical were studied (Figure S13), revealing the overlapping of absorption spectrum of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical with excitation spectrum of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, which confirms the IFE. Thus, the as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs act as a turn-off fluorescence probe for the detection of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e3.4. Sensitivity\u003c/h2\u003e \u003cp\u003eUnder the optimal conditions, the variations in the fluorescence intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were investigated by adding different concentrations of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (0.01\u0026ndash;500 \u0026micro;M). As can be noticed in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e, the fluorescence emission intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs centered at 433 nm was gradually quenched along with increasing concentration of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical. Then, the emission intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs was quenched 77% when \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical at 500 \u0026micro;M. Figure S14 displayed the constructed plot of the ratio I\u003csub\u003e0\u003c/sub\u003e/I (where \u003cem\u003eI\u003c/em\u003e\u003csub\u003e0\u003c/sub\u003e and \u003cem\u003eI\u003c/em\u003e represent the emission intensity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in the absence and presence of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical, respectively) against \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical concentration (0.01\u0026ndash;500 \u0026micro;M). Furthermore, with \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical concentration in the range of 0.01-5.0 \u0026micro;M, the fluorescence quenching efficiency showed the linear fitting equation of y\u0026thinsp;=\u0026thinsp;0.2596x\u0026thinsp;+\u0026thinsp;1.2061 (\u003cem\u003eR\u003c/em\u003e\u003csup\u003e\u003cem\u003e2\u003c/em\u003e\u003c/sup\u003e\u0026thinsp;=\u0026thinsp;0.9795). The LOD was 3.10 nM (3σ/s, where \u003cem\u003e\u0026ldquo;s\u0026rdquo;\u003c/em\u003e is the slope of the calibration curve and \u0026ldquo;σ\u0026rdquo; is the standard deviation of the blank) for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical. The analytical characteristics of the developed Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs-based fluorescence approach were compared with other reported methods [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e, \u003cspan additionalcitationids=\"CR56 CR57 CR58 CR59 CR60 CR61\" citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e] for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing (Table \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e), revealing the developed probe exhibited superior and a comparable analytical performance with other analytical techniques for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing. Furthermore, the developed Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs-based turn-off fluorescence strategy exhibits several analytical features such as free surface modification, simple chemical routes, good selectivity and sensitivity, and good stability in aqueous phase, which allows for use as a promising fluorescence probe for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e3.5. Selectivity\u003c/h2\u003e \u003cp\u003eThe sensing selectivity of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs toward \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical was investigated in the presence of various metal ions, ROS, and biomolecules phosphate buffer (20 mM, pH 7.4). As shown in Figure S15, the fluorescence behavior of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in the presence of biomolecules (cysteine, arginine, tryptophan, and alanine, 500 \u0026micro;M), cations (Na\u003csup\u003e+\u003c/sup\u003e, Ca\u003csup\u003e2+\u003c/sup\u003e, Mg\u003csup\u003e2+\u003c/sup\u003e, Zn\u003csup\u003e2+\u003c/sup\u003e, Cu\u003csup\u003e2+\u003c/sup\u003e, and Fe\u003csup\u003e3+\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e 500 \u0026micro;M) and anions (Cl\u003csup\u003e\u0026minus;\u003c/sup\u003e, I\u003csup\u003e\u0026minus;\u003c/sup\u003e, Br\u003csup\u003e\u0026minus;\u003c/sup\u003e, PO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e3\u0026minus;\u003c/sup\u003e, and SO\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e2\u0026minus;\u003c/sup\u003e\u003csub\u003e,\u003c/sub\u003e 500 \u0026micro;M) is almost same and did not show any significant changes. As anticipated, Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited a remarkable fluorescence quenching only with \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical even in the existence of the other interfering chemical species, revealing that the as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs stand out as a highly selective turn-off fluorescence probe for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical sensing.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e\u003cb\u003e3.6. Analysis of\u003c/b\u003e \u003csup\u003e\u003cb\u003e\u0026bull;\u003c/b\u003e\u003c/sup\u003e\u003cb\u003eOH radical in water samples\u003c/b\u003e\u003c/h2\u003e \u003cp\u003eTo evaluate the practical application of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs in monitoring \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in real samples, the collected water (tap and river) samples were filtered and subsequently added different concentrations of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical (5.0, 10.0 and 25.0 \u0026micro;M). Then, \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical treated water samples were added into Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs, and their concentrations were estimated by the aforesaid procedure. From Table S2, it can be noticed that the recovery rates of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in water samples were 99.00\u0026ndash;101.40% with a relative standard deviation of \u0026lt;\u0026thinsp;2.0%. The results demonstrate that Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs could be used as the potential fluorescence turn-off probe for the detection of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in real samples.\u003c/p\u003e \u003c/div\u003e"},{"header":"4. Conclusions","content":"\u003cp\u003eIn summary, a simple analytical tool was developed for sensing \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical using water-dispersible Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs as a turn-off fluorescence probe. The as-fabricated Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited blue fluorescence under UV irradiation (λ \u0026sim; 365 nm) and displayed λ\u003csub\u003eEm/Ex\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;433/370 nm. The developed Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs-based fluorescence approach has a highly selective and sensitive response toward \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical with a good linearity in the concentration range of 0.01-5.0 \u0026micro;M, which achieves the LOD of 3.10 nM. Importantly, Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibit superior selectivity for \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical with virtually no fluorescence quenching by other interfering chemical species (ROS, metal ions, anions and biomolecules). Furthermore, Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs-based analytical approach was successfully applied to detect \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in water samples. Thus, the as-synthesized Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs could be successfully integrated with fluorescence spectrometry for the detection of \u003csup\u003e\u0026bull;\u003c/sup\u003eOH radical in real samples.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSKK thanks the Director, Sardar Vallabhbhai National Institute of Technology, Surat for grating the sabbatical leave for Brainpool Programme at Chung-Ang University, South Korea. This research was supported by the Brain Pool program funded by the Ministry of Science and ICT through the National Research Foundation of Korea (RS-2023-00221526).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical Approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to participate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to publish\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSKK thanks the Director, Sardar Vallabhbhai National Institute of Technology, Surat for grating the sabbatical leave for Brainpool Programme at Chung-Ang University, South Korea. This research was supported by the Brain Pool program funded by the Ministry of Science and ICT through the National Research Foundation of Korea (RS-2023-00221526).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData will be available from corresponding author on demand.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot Applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll the authors declare that there is no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026apos; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSuresh Kumar Kailasa\u003c/strong\u003e: Methodology, Formal analysis, Writing\u0026mdash;original draft. \u003cstrong\u003eKartik Pankajbhai Makwana\u003c/strong\u003e: Material preparation, Formal analysis, Writing\u0026mdash;Review and Editing. \u003cstrong\u003eMadhura Pradeep Deshpande\u003c/strong\u003e\u003cstrong\u003e:\u0026nbsp;\u003c/strong\u003eFormal analysis, Data collection and analysis, Writing\u0026mdash;Review and Editing.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eYoojin Choi\u003c/strong\u003e\u003cstrong\u003e:\u0026nbsp;\u003c/strong\u003eFormal analysis, Data collection and analysis, Writing\u0026mdash;Review and Editing.\u003cstrong\u003e\u0026nbsp;Tae Jung Park\u003c/strong\u003e: Conception and design, Data collection and analysis, Formal analysis and Investigation, Software, Funding, Writing\u0026mdash;review \u0026amp; editing. 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Adv Funct Mater 28:1702026\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Scheme","content":"\u003cp\u003eScheme 1 is available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"microchimica-acta","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"miac","sideBox":"Learn more about [Microchimica Acta](https://link.springer.com/journal/604)","snPcode":"604","submissionUrl":"https://submission.springernature.com/new-submission/604/3","title":"Microchimica Acta","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Try-CsPbCl3 PNCs, Hydroxyl radical, Fluorescence spectrometry, HR-TEM","lastPublishedDoi":"10.21203/rs.3.rs-5436012/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5436012/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eWater-dispersible perovskite nanocrystals (PNCs) show promising applications in recognizing ionic and molecular species because of their excellent optical properties. However, lead halide PNCs have some limitations when they are used as probes for molecular species sensing in aqueous media. Here, we introduce trypsin (Try) as a bioligand for the synthesis of cesium lead chloride (CsPbCl\u003csub\u003e3\u003c/sub\u003e) perovskite nanocrystals (PNCs) with high water stability. The as-fabricated Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs show λ\u003csub\u003eEm/Ex\u003c/sub\u003e at 433/370 nm with quantum yield of 17.26%. The fluorescence emission spectral characteristics of Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs demonstrated that water-stable Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs acted as a promising fluorescent probe for the detection of hydroxyl radical (\u003csup\u003e\u0026bull;\u003c/sup\u003eOH) \u003cem\u003evia\u003c/em\u003e turn-off mechanism. The Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs-based turn-off fluorescence approach displayed good selectivity for hydroxyl radical in water, showing a wider linear range (0.01\u0026ndash;5 \u0026micro;M) with a remarkable detection limit of 3.10 nM for hydroxyl radical. The as-prepared Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs were demonstrated to be a facile probe for sensing \u003csup\u003e\u0026bull;\u003c/sup\u003eOH in water samples, which signifies that Try-CsPbCl\u003csub\u003e3\u003c/sub\u003e PNCs exhibited broad applications for hydroxyl radical sensing and cell imaging.\u003c/p\u003e","manuscriptTitle":"Synthesis of trypsin protected CsPbCl3 fluorescent nanocrystals for hydroxyl radical sensing","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-12-17 17:25:46","doi":"10.21203/rs.3.rs-5436012/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-12-15T15:48:23+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-12-12T15:36:16+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-11-30T08:36:54+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"120997717891649353288519159929260156799","date":"2024-11-29T00:16:21+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"142928529735927493712265646022387078169","date":"2024-11-24T07:36:47+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-11-18T08:44:05+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-11-13T23:08:29+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-11-13T23:08:05+00:00","index":"","fulltext":""},{"type":"submitted","content":"Microchimica Acta","date":"2024-11-12T04:08:37+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"microchimica-acta","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"miac","sideBox":"Learn more about [Microchimica Acta](https://link.springer.com/journal/604)","snPcode":"604","submissionUrl":"https://submission.springernature.com/new-submission/604/3","title":"Microchimica Acta","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"15e39ea6-642e-49fa-b939-dc77457f941f","owner":[],"postedDate":"December 17th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-03-17T16:07:01+00:00","versionOfRecord":{"articleIdentity":"rs-5436012","link":"https://doi.org/10.1007/s00604-025-07070-8","journal":{"identity":"microchimica-acta","isVorOnly":false,"title":"Microchimica Acta"},"publishedOn":"2025-03-10 15:57:46","publishedOnDateReadable":"March 10th, 2025"},"versionCreatedAt":"2024-12-17 17:25:46","video":"","vorDoi":"10.1007/s00604-025-07070-8","vorDoiUrl":"https://doi.org/10.1007/s00604-025-07070-8","workflowStages":[]},"version":"v1","identity":"rs-5436012","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5436012","identity":"rs-5436012","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}