A biocompatible hydrogel as a template for oxidative decomposition reactions: a chemodosimetric analysis and in vitro imaging of hypochlorite

The self-assembly properties of new biocompatible, thermoreversible fluorescent hydrogels, composed of amino acid residues, e.g., l-phenylalanine (PyL-PheOx) and l-tyrosine (PyL-TyrOx), have been reported. Spectroscopic investigations indicate that PyL-PheOx forms π-stacked ‘compact’ aggregates, while ‘loose’ aggregates with stronger CT characteristics are observed for PyL-TyrOx. Both the compounds showed the presence of fibrous networks in the self-assembled state. Circular dichroism spectral studies indicate the formation of M-helical and P-helical structures for PyL-PheOx and PyL-TyrOx, respectively. A striking gel-to-sol transition, caused by oxidative decomposition, is explicitly noticed in the presence of hypochlorite. A mechanistic investigation reveals the oxidation of the acyl aroyl hydrazine core of the gelators in the presence of ClO−. In addition to this, change in the fluorescence emission intensity of the hydrogel in the presence of ClO− is utilized for the analysis of commercial bleach samples. Gel-coated paper strips are also developed for the on-site detection of ClO−. Furthermore, the system is utilized for imaging hypochlorite in live mammalian cells.


Materials and methods
All chemicals, solvents and silica gel for TLC were obtained from well-known commercial sources and were used without further purication, as appropriate. Solvents were freshly distilled and dried by a standard procedure before use. Melting points were measured in open capillaries and were uncorrected. 1 H-NMR and 13 C-NMR spectra were recorded using a Bruker-400 Advance NMR spectrometer. Chemical shis were reported in ppm downeld from the internal standard tetramethyl silane (TMS). Mass spectrometry of individual compounds was performed using a Micro Mass ESI-TOF MS instrument. Elemental analysis was performed using a Thermo Finnigan EA FLASH 1112 SERIES.

Gelation studies
The formation of the gel was conrmed using the tube inversion method. In a typical experiment, PyL-PheOx and PyL-TyrOx were taken and to this mixture 1 mL of deionized water was added, followed by heating at 80 C till the solution became transparent. The resultant mixture was sonicated for a few minutes and subsequently allowed them to cool at room temperature. If a gel was formed, it was evaluated quantitatively by determining the critical gelator concentration (CGC). For the evaluation of the CGC, a known amount of PyL-PheOx/PyL-TyrOx in pure water was heated to 75-80 C, which resulted in a slightly turbid solution and then sonicated. The resulting clear solution transformed into an optically transparent/opaque robust hydrogel within 1-2 min. The gelation propensity of PyL-PheOx/ PyL-TyrOx has been checked systematically by gradually lowering the amount of the gelator. In that way, the minimum concentration of the gelator that was required to immobilize 1 mL of water, i.e., the CGC value was determined as reported with other systems. The above measurement was carried out three times and the average CGC value has been reported. A glass tube with a capacity of 10 Â 75 mm has been used for the CGC measurement. For thixotropic properties, the gel was vortexed for 60 s for complete gel-to sol transition and kept at rest for 10 min to revert to gel.

FT-IR spectroscopy
The prepared solution and gels of individual samples were drop coated on a CaF 2 cell and dried under vacuum and FT-IR spectra were recorded using a PerkinElmer Spectrum BX FT-IR system. For solution measurements, PyL-PheOx/PyL-TyrOx was dissolved in CHCl 3 and was transferred to a constant path length cell (50 mm) containing CaF 2 windows. For baseline correction, the spectrum of neat CHCl 3 was subtracted from the sample spectrum.
UV/Vis and uorescence spectroscopy UV-vis spectroscopy and uorescence spectroscopy of the described solutions/suspensions were performed on a Shimadzu model 2100 spectrophotometer and Cary-Eclipse spec-trouorimeter respectively, both equipped with a temperature controlled bath. The slit width for the uorescence experiment was kept at 5 nm (excitation) and 5 nm (emission) and the excitation wavelength was set at 345 nm.

Circular dichroism spectroscopy (CD)
All the CD spectra of the described solutions were recorded on a JASCO instrument, Model J-815-150S. Experiments were performed by purging dry N 2 gas continuously. Data were collected in a quartz cuvette of 1 mm path length between 200 and 450 nm.

Atomic force microscopy (AFM)
A dilute solution of the sample of gel was deposited on a freshly cleaved mica surface and then allowed carefully to freeze-dry. Each of the samples was analysed using a JPK 00901 AFM instrument and Nano-Wizard soware: tapping mode off 10 nm tip radius, silicon tip, 292 kHz resonant frequency, 0.7-1 Hz scan speed, 256 Â 256 and 512 Â 512 pixels.

Scanning electron microscopy (SEM)
The gels or the specied solution was carefully drop coated onto brass stubs and was allowed to freeze-dry. The samples were then coated with gold vapor and analysed on a Quanta 200 SEM operated at 10-15 kV.

X-ray diffraction (XRD)
The gel samples were frozen using liquid N 2 , followed by further drying in a lyophilizer to obtain a xerogel for X-ray diffraction study. The dried powder was then placed on a glass plate. These samples were examined using a Bruker D8 Advance instrument (q, 2q geometry with scintillation detectors). The X-ray beam generated with a rotating Cu anode at a wavelength of the KR beam of 1.5418 was directed toward the lm edge and scanning was done up to a 2q value of 30 . Data were then analysed and interpreted using the Bragg equation.

Energy minimization
The energy minimization of the hydrogelator molecules was performed using the B3LYP/6-31G* level of computations.

Rheological studies
Rheology experiments using the hydrogels were carried out using an Anton Paar MCR 52 with a cone and plate geometry (CP 25-2) having an adjustable Peltier temperature controlling system. The distance between the cone and plates was kept xed at 0.105 mm for all the measurements. The gels were taken on the plate of the rheometer. An oscillatory strain amplitude sweep experiment was performed at a constant oscillation frequency of 1 Hz for the applied strain range 0.01-100% at 20 C. The rheometer has inbuilt soware that converts torque measurements into either G 0 (the storage modulus) and G 00 (the loss modulus), which is represented either as strain or shear stress. Oscillatory frequency sweep experiments were performed in the linear viscoelastic region (strain 0.01%) to ensure that calculated parameters correspond to an intact network structure. The thixotropic properties were studied to examine the alteration of the rheological properties of the hydrogel material under the application and the release of shear stress. This process includes two steps: (i) deformation of the hydrogel with increasing shear stress from 0.01 to 80 Pa for 6.14 min and (ii) recovery of the sol state to the gel state under low shear stress (0.01 Pa) for 6 min. In these two steps, the frequency was kept constant at 1 Hz. Four consecutive cycles were applied on the hydrogels to compare the extent of gel strength recovery.

Measurement of uorescence lifetime
Fluorescence lifetimes were measured by using a time--correlated single-photon-counting uorimeter (Horiba Jobin Yvon). The system was excited with nano-LED light (Horiba-Jobin Yvon) with a pulse duration of 1.2 ns (slit width: 2/2 and l em ¼ 345 nm). The average uorescence lifetimes (s av ) for the exponential iterative tting were calculated from the decay times (s i ) and the relative amplitudes (a i ) by using eqn (1), where a 1-3 are the relative amplitudes and s 1-3 are the lifetime values.

Preparation of test strips
For the preparation of gel-coated paper strips, lter paper (Whatman 40) strips were cut into a square shape (1 Â 2 cm). To these square shaped paper strips, 30 mL of the pre-prepared gel sample (2.5 mM) was applied on the surface so that it could spread uniformly. The gel coated on the paper strip was completely absorbed within 15 min under an ambient open atmosphere and then it was air-dried naturally. These gel coated paper strips were then used for checking ClO À .

Analysis of hypochlorite in different water samples
Tap water samples were collected from the Department of Organic Chemistry Laboratory, IISc. Pool water samples were collected from the swimming pool of the Indian Institute of Science campus, Bangalore, India. Both tap water and pool water samples were separately used for analysis as received. Seawater samples were collected from the Arabian Sea water (near Mangalore). To remove the insoluble and suspended dirt particles from the sea water, it was ltered through a 0.22 mm membrane lter cartridge. To these water samples, different amounts of ClO À were added more than 30 min before the analysis.

Cell culture
HeLa cells were cultured in Dulbecco's Modied Eagle's Medium (DMEM, CELL clone, India) containing 10% FBS (Invitrogen, USA) and antibiotics (100 units per mL penicillin and 100 mg mL À1 streptomycin). The cells were incubated in a humidied 5% CO 2 incubator (Sanyo, UK) at 37 C.

Cell viability assay
The cytotoxicity of the compounds was evaluated in HeLa and MCF7 cells by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. In a typical experiment, the cells were seeded at a density of $1 Â 10 4 cells per well in 96 well plates and incubated for 24 h. Then, the cells were treated with different concentrations of the probe. Finally, the cells were washed, and 20 mL of MTT (5 mg mL À1 in DMEM) was added into each well and incubated for 3 h in the dark. Finally, the whole medium was removed from the wells, and DMSO (100 mL) was added. The cells were incubated for 5 min in the dark, and then absorbance was measured at 560 nm.

Cell imaging
For blank reading, the HeLa cells were incubated with 20 mM PyL-TyrOx for 30 min at 37 C. Then, the medium was removed, and the cells were washed multiple times with pH 7.4 DMEM buffer.

Synthesis of PyL-PheOx
To the solution of PyL

Results and discussion
Design and synthesis of uorescent hydrotropes Herein, we have synthesized two different uorescent hydrotropes, attached with a gallate derivative (Fig. 1) and linked via aromatic amino acids namely phenylalanine (PyL-PheOx) and tyrosine (PyL-TyrOx). Both the compounds were synthesized following a conventional peptide coupling protocol and characterized thoroughly using 1 H-NMR, 13 C NMR, ESI-MS, FT-IR, and elemental analysis (for details see the Experimental section and ESI †).

Self-assembly and gelation properties in aqueous medium
Both PyL-PheOx and PyL-TyrOx belong to the class of supergelators as the former formed a stable transparent hydrogel (CGC: 0.52 mM) in water, while an opaque hydrogel (CGC: 0.82 mM) was found with the latter (Fig. 1a). The strong gelation ability of PyL-PheOx may presumably have originated from the enhanced p-p stacking interactions attributed to unsubstituted phenyl residues. As expected, FT-IR analysis indicated that in both cases, the C]O (Dl $ 15 cm À1 ) and N-H (Dl $ 45 cm À1 ) stretching frequencies got shied to higher energy regions upon the formation of the hydrogel (in comparison to that in CHCl 3 ). The extent of shi was found to be larger for PyL-PheOx, and this might be due to the stronger H-bonding interactions between the acyl aroyl hydrazine (AAH) units, promoted by enhanced p-p stacking interactions (Fig. S1 †). Owing to the presence of AAH units, both the gel samples were found to be susceptible to pH change as well (Fig. S2 †). Interestingly, on applying mechanical force (shaking), a gel-to-sol transformation was observed, which on keeping at rest reverted to the native gel state in a few minutes (thixotropic behavior, Fig. S2 †).

Spectroscopic studies of hydrogelators in the solution phase
Concentration-dependent UV-visible studies indicate the broadening of the pyrene-characteristic absorption peaks in the higher concentration range (Fig. S3 †), probably due to the pstacking interactions between such residues. 69,70 Similarly, concentration-dependent uorescence emission spectral studies showed the formation of a broad band in the red-shied region (at 422 nm for PyL-PheOx and at 460 nm for PyL-TyrOx) in addition to pyrene-characteristic vibronic bands (Fig. S4 †). Excitation spectral analysis indicates that this newly formed structureless band on the longer-wavelength side is originated from the aggregated species (ground-state interaction) and not due to the formation of photo-excited dimers (Fig. S5a †). The larger uorescence intensity of the aggregated band apparently indicates the effective prevalence of p-stacking interactions in the case of PyL-PheOx compared to the case of PyL-TyrOx. However, a closer look reveals that the aggregate band positioning was rather red shied for PyL-TyrOx (Fig. 2a). We presume that the relatively poorer alignment of the pyrenyl residues in PyL-TyrOx might have impeded the formation of a compact packing structure, but it could initiate chargetransfer interactions due to the presence of the electrondonating -OH group in it. 71 The time-dependent uorescence decay prole shows a shorter decay time for PyL-TyrOx (s av ¼ 0.85 AE 0.03 ns) in comparison to that of PyL-PheOx (s av ¼ 1.32 AE 0.02 ns). The formation of such short-lived excited state species further conrms the existence of intermolecular CT interactions in the self-assembled state of PyL-TyrOx (Fig. 2a, inset). Similarly, the large steady-state uorescence anisotropy values indicate a rather rigid microenvironment around the pyrene residues in both the cases (Fig. S5b †). When PyL-PheOx (Fig. S6a †) and PyL-TyrOx (Fig. S6b †) were subjected to variable temperature uorescence spectral studies, the diminution of the aggregate emission band was observed, probably due to the heat-induced 'dissolution' of the aggregate species. Similarly, a urea-induced shi in the aggregate to monomer emission was observed for both PyL-PheOx and PyL-TyrOx perhaps due to the dissociation of the hydrogen-bonded networks.

Amino acid-directed distinctive supramolecular chirality
To unravel whether chiral organizations from the selfassembled aggregates were formed in aqueous media, both PyL-PheOx and PyL-TyrOx were subjected to circular dichroism (CD) spectral analysis. The CD spectrum of PyL-PheOx showed a bisignated negative Cotton signal in the aggregated state, indicating the formation of a le-handed M-helical structure. In contrast, the self-assembly of PyL-TyrOx (Fig. 2b) exhibited a bisignated positive Cotton signal, suggesting the formation of a right-handed P-helical organisation. [72][73][74][75] However, the gradual diminution of CD signals was observed in both the cases with rising temperature, leading to CD-silent situation beyond $70 C (Fig. S7 †). This indicates the 'disintegration' of the P and M-helical aggregates of PyL-PheOx and PyL-TyrOx respectively at higher temperatures, attributed to thermal-agitation.

Viscoelastic properties of uorescent hydrogels
The nanostructures formed via 3D-assembly of the gelator molecules were also studied using both Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). In both the cases, the presence of a brillar network was observed indicating a strong intermolecular interaction. However, the brils appeared to be more 'densely-packed' in the case of PyL-PheOx, conrming the formation of 'compact' aggregates as described earlier (Fig. 1b).
The mechanical strengths of the hydrogel samples (PyL-PheOx and PyL-TyrOx) were evaluated using rheological studies. [76][77][78][79] Oscillatory frequency (Fig. S8a †) and amplitude (Fig. S8b †) sweep experiments suggest that these hydrogels are highly viscoelastic in nature, and the viscoelasticity increases with increasing gelator concentration. Also, the viscoelasticity of the hydrogels was maintained over a frequency range of 1-100 Hz under a constant applied strain of 0.01%. Interestingly, the hydrogel sample derived from PyL-PheOx exhibited greater mechanical stiffness than the PyL-TyrOx based hydrogel.
The critical stress required to disturb the solid-like behavior was found to be larger ($53 Pa) in the former case than what was needed for the latter ($41 Pa). To conrm the thixotropic behavior of the hydrogels, we performed the hysteresis loop test. The mechanical stiffness could be recovered to >98% of the original value within 30 s upon removal of the induced strain ( Fig. 3a and S8c †). Keeping this in mind, we had further explored the injectability of the hydrogel sample, where shapepersistent alphabetical letters could be written using the gel samples (inset Fig. 3a).

3D arrangement and proposed nanomolecular model
The XRD studies of dried gel of PyL-PheOx (Fig. 3b) showed the presence of reection peaks at 2q ¼ 2.  The molecular lengths calculated along the long molecular axis using the B3LYP/6-31G* level of theory were found to be 2.7 nm and 3.1 nm (inset Fig. 3b) for PyL-PheOx and PyL-TyrOx, respectively. The repetitive distance obtained for the lamellar structure from the XRD studies was less than double the full lengths of the minimized structure. The observation suggests that the gelator molecules undergo substantial interdigitation via the pyrenyl unit in the gel state. Thus, we believe that in the gel state the hydrophobic pyrene residues will form p-stacked arrays, while the oxyethylene units, being hydrophilic, remain exposed to bulk water. On the other hand, the AAH units of the adjacent molecules participate in the complementary Hbonding interactions. The formation of bre like structures, as proposed in this model, will presumably undergo entanglement further to eventually form gels (Fig. 3c). 70,80 Chemodosimetric analysis of hypochlorite using a hydrogel nano-template Considering the presence of redox sensitive AAH units, PyL-PheOx and PyL-TyrOx were exposed to various reactive oxidative species in aqueous media. In both the cases, the addition of ClO À could specically lead to a change in the solution color from colorless to yellow, which eventually became transparent soon. At the same time, the enhancement of uorescence intensity in the $390-420 nm region was observed at the expense of the aggregate emission (ratiometric response) ( Fig. 4a and S9a †). Irrespective of the nature of the amino acid residue, in both the cases, the saturation in the optical signal required an incubation time of $20 min for the completion of the event (Fig. S9b †). Time-course studies indicate that it followed a pseudo-rst-order kinetic behavior in the presence of excess ClO À . As expected, both at elevated temperature (Fig. S10 †) and at higher concentration (Fig. S9b-d †) of ClO À , the reaction rates accelerated. Selectivity is another key parameter to evaluate the performance of a new probe.

2À
) and intracellular bioreductants (GSH). No analyte, other than ClO À , could produce any detectable change in the uorescence signal ( Fig. 4b and S11 †). Although a weak signal in the presence of ROOc could be observed, it was negligible in comparison to that observed in the presence of ClO À (Fig. S12b †). Titrimetric studies further indicate that the present systems could detect ClO À as low as 0.04 ppm with PyL-TyrOx and 0.12 ppm with PyL-PheOx gel (Fig. 4a). A comparison table for different hypochlorite sensors with their detection limits (especially for the hydrazone moiety, which underwent a chemodosimetric reaction on addition of hypochlorite) is also given in Table S1. †

Hypochlorite triggered oxidative decomposition reaction
The hypochlorite anion (ClO À ) is known to selectively oxidize dibenzoyl hydrazine to dibenzoyl diimide, which undergoes further decomposition in nucleophilic solvents like water. Analogously considering the presence of AAH units, both in PyL-PheOx and PyL-TyrOx molecules, these are susceptible to get oxidized to acyl aroyl diimides on exposure to ClO À . These intermediates thus formed undergo spontaneous hydrolysis in water to the corresponding acids I and II with the concomitant release of N 2 (Scheme is given in Fig. 4c). 44,81 Though we couldn't detect the presence of diimide derivative in the  reaction media, one can assume that it might be responsible for the immediate yellow coloration. However, to ascertain and conrm the identity of various products upon treatment of ClO À with the hydrogel of PyL-TyrOx, the reaction mixture was subjected to ESI-MS analysis. The experimental gure (see Fig. S13 in the ESI †) shows that, besides the peak due to the probe PyL-TyrOx (m/z: 1078.4889), there are two more peaks that appear at m/z 474.1617 (peak I, calculated M + Na + ) and 631.2368 (peak II, calculated M + Na + ) respectively ( Fig. S13a and b †). Also, the uorescence spectra of PyL-PheOx and PyL-TyrOx in the presence of ClO À showed a close resemblance to the uorescence spectra of free PyL-PheOH and PyL-TyrOH amino acid conjugates (synthesized separately, Fig. 5b and S13c †). Based on our data and previous observation reported in the literature, we conclude that ClO À selectively oxidizes acyl aroyl hydrazine of PyL-TyrOx to the corresponding acyl aroyl diimide, which then undergoes spontaneous decomposition in aqueous media. We propose that the uorescence turn on reaction in this system proceeds primarily through the route that is depicted in Fig. 4c. These two observations conrm the oxidative dissociation reaction in the presence of ClO À . However, the extent of uorescence enhancement in the $395 nm region was different for PyL-TyrOx vs. PyL-PheOx, and thus an $10-fold rise was observed for the former and only $4-fold for the latter. It is possible since the stability of the diimide derivative is known to be dependent on the nature of the adjacent aromatic residues. The formation of PyL-PheOH or PyL-TyrOH was also conrmed by HPLC studies (Fig. 5a). As shown in Fig. S13d, † the chromatographic peak due to PyL-TyrOx appeared at 12.3 min. Aer reaction with ClO À , two additional chromatographic peaks emerged at $8.2 and 4.8 min, respectively. The new chromatographic peak that appeared at $8.2 min was assigned to PyL-TyrOH, formed via oxidative hydrolysis of PyL-TyrOx. This was also conrmed by injecting authentic PyL-TyrOH into the above mentioned HPLC column.
Gel-to-sol transition induced by hypochlorite Furthermore, we studied the effect of ClO À on the stability of hydrogel samples. In both the cases, the mechanical strength of the hydrogels decreased remarkably in the presence of ClO À (Fig. 6c and S14b †). Finally, when the ClO À concentration exceeded $2 mM, a visible transition from solid gel to uid sol could be seen ( Fig. 5c and S12a †). As expected, such phase transition led to changes in the microscopic structures, and the brillar morphology of gels changed to irregularly shaped particles in the presence of ClO À (Fig. 6b and S14a †). A decrease in the hydrodynamic diameter and lowering of the uorescence anisotropy values with ClO À further conrm the dissociation of the preformed self-assembled structure (Fig. S14c, d and S15 †). Also, the analysis of CD reveals the loss in helical chirality upon addition of ClO À (Fig. 6a).

Real-life application of hypochlorite sensing
Due to superior sensitivity, further we employed PyL-TyrOx in the detection of ClO À in different natural water sources such as tap, pond, sea, etc. 82 In all the cases, one could observe almost quantitative recovery values with relatively small standard deviations (<5%) (Fig. 7a and Table S2 †). Furthermore, the estimation of ClO À was performed in commercial bleach samples with a high degree of accuracy (Fig. S16a †). We have calculated % recovery values and relative standard deviations for the estimation of ClO À in diluted bleach solution. The nearly 100% recovery values with a relatively small standard deviation (<5%) indicate that our system is indeed suitable for analyzing hypochlorite in real-life samples, such as bleach (Table S3 †).
Gel-coated portable paper strips 83 were developed for the detection of ClO À at remote locations without involving sophisticated analytical tools; for this a lter paper (Whatman 40) strip was cut into a rectangular shape (1 Â 2 cm). 30 mL of the prepared gel sample (2.5 mM) was slowly applied on the surface of the paper strips using a micropipette such that it spreads uniformly. The gel coated on the paper strip was apparently totally absorbed within 15 min and then it was kept overnight to air-dry naturally. The paper strips coated with PyL-TyrOx showed strong cyan uorescence under a >365 nm UV lamp. Before the sensing experiments, we also checked the stability of the paper strips under ambient conditions. No detectable change in the  uorescence emission intensity was observed even aer two weeks. This suggests that the strips could easily be stored at room temperature. Aer this, the gel-coated paper strips were exposed to different amounts of ClO À (0 to 25 mM). A concentration-dependent gradual quenching of cyan uorescence was observed (Fig. 7c). The changes in intensity could be quantied using readily available image processing soware, Image J (Fig. S16b †). Because of the operational procedure mentioned here, this is very simple, and even the users with no technical knowledge can use them without any difficulty.

In vitro imaging of hypochlorite
Since both PyL-PheOx and PyL-TyrOx gelators showed high biocompatibility even at $30 mM concentration, the gelators could be used for the imaging of ClO À in HeLa and MCF7 cells ( Fig. 7b and S17a †). 84 Dose-dependent kinetic studies indicated that with 20 mM PyL-TyrOx a 30 min incubation time (at 37 C) was enough for the complete cellular uptake. The stability of the gelator molecules was then monitored from time to time using HPLC and ESI-MS mass spectrometry. No signicant degradation of PyL-TyrOx was observed even when incubated in the presence of proteinase K (5 mg mL À1 ) for more than 35 h at physiological pH 7.4 (Fig. S17b †). As expected, the cells treated with PyL-TyrOx showed blue uorescence in the 450-600 nm range due to cellular internalization (Fig. S17c †). However, upon exposure to aqueous ClO À , the cells exhibited dose-dependent diminution of uorescence, which might be due to the oxidative-dissociation reactions inside the living cells (Fig. 7d). These results demonstrate that PyL-TyrOx could also be used for the ratiometric uorescence imaging of ClO À in living cells.

Conclusions
In conclusion, we have demonstrated the design and synthesis of two novel pyrene-based biocompatible hydrotropes, PyL-PheOx and PyL-TyrOx, which can readily form biocompatible, thermoreversible thixotropic hydrogels in aqueous media.
Spectroscopic investigations indicate the formation of p-stacked 'compact' aggregates in the case of PyL-PheOx, while the aggregates with stronger CT characteristics were formed from PyL-TyrOx. Though both the compounds showed the presence of brous networks in the self-assembled state, CD spectral investigations indicate the formation of M-helical and P-helical structures for PyL-PheOx and PyL-TyrOx respectively. Furthermore, the compounds showed clearly visible solid gel-to-uid sol transition in the presence of ClO À via oxidative hydrolysis. The change in the uorescence intensity of the hydrogel in the presence of ClO À was utilized for the analysis of commercial bleach samples and live-cell imaging. Finally, gel-coated paper strips were also developed for the on-site detection of ClO À . Such ndings may inspire widespread analytical use of hydrogels and related so matter and also encourage developing newer designs or assays of reactive oxygen species and other analytes.

Author contributions
S. B. conceptualized the project; experimental investigation and data collection were done by DB and ND under supervision of SB. The original dra of the manuscript was prepared by DB, its review, and editing were done by DB, ND and SB. The infrastructure and funding of the research were provided by SB.

Conflicts of interest
There are no conicts to declare.