A ring‐controlled fluorescent platform for visualizing polymer degradation

The creation of a new photoluminescent platform to study their photo physics and further applications is important in the fields of chemistry, material, physics, etc. In this work, we developed benzene‐based fluorophores generated by chemical reactions on conjugate acceptor precursors. Their optical properties have been studied, and the large Stokes shifts, high molar extinction coefficients and quantum yields have been revealed. The starting precursors containing bis‐intercalated vinyl thioester shows no luminescence, which is turned on by the chemically triggered cyclization. From spectroscopic, crystallographic, and computational studies, cyclization‐induced emission enhancement is proposed as a mean to explain the photoluminescent performances, a principle extended in the substituent incorporated fluorophores. Furthermore, they were applied to the chemically triggered degradation of a polymer and achieved visual tracking, quantifying, and downcycling of the processes. The method for designing and developing series fluorophores and visually tracking polymer degradation represents a new photoluminescent platform, allowing its further applications.

been studied and applied to the development of optical materials [5] and chemosensors. [6] This mechanism is derived from restricting the rotation of bonds within the emitter to improve structural rigidity. Thus, it would be interesting to combine luminescence with dynamic covalent bonding to the ketene-conjugated dimethylthioacetal acceptors to achieve the goal of tracking macroscopic metathesis on stimuliresponsive materials, including the tracking and quantitation of polymer degradation, further downcycling of the polymers (see below).
Previously, we developed thiol-initiated self-propagating cascades based on indanedione-derived conjugated receptor, and achieved the dual functions of material degradation and visualization tracking through indanonalkene indicator. [7] However, the optical mechanism and real quantitation of macromolecular alteration or degradation monitored by fluorescence signal have not been achieved. Bearing these in mind, we made studies, further explored, and discovered in this work: (i) cyclization-induced emission and enhancement on the indanedionalkene chromophores and ring size effect S C H E M E 1 General schematic of coupling reactions and photoluminescence properties exploited herein. a a Structure of the conjugate acceptor (CAs) as fluorophore precursor. The non-luminescent CAs can be scrambled by reagents containing HS-R-SH, HS-R-OH with varying R-groups through releasing methyl mercaptan, which generated the heterocyclic fluorophores As and Bs. on the luminescence properties; (ii) the enhanced luminescence was attributed to conformational rigidity and reduced vibrational energy loss; (iii) the strategy was generalized and extended in the conjugate acceptors (CAs) with different substituents; (iv) tracking and quantification of the polymer degradation through fluorescence indicator were studied.
In brief, the CAs containing bis-vinylogous thioesters were showing non-luminescence but chemically triggered cyclization led to fluorescence turn-on (Scheme 1). Further, cyclization with smaller ring sizes (i.e., five members versus six/seven members) resulted in stronger fluorescence enhancement. The approach for modulating optical responses was then investigated via experimental, computational, and single crystal analysis. Cyclization-induced emission enhancement on the fluorophores was explained toward the photoluminescence in the aqueous solution. The methodology was then extended on substituted conjugate acceptors (CA-X), where luminescence was also switched on upon β-mercaptoethanol (BME) induced cyclization, while the photophysical performances were improved. In a further application, a linear non-luminescent polymer linked with CA and 1,6-hexanedithiol through thiol-thiol exchanging was degraded rapidly via the "declick" reaction triggered by BME. More importantly, the degradation process can be tracked and quantified through nuclear magnetic resonance (NMR) and fluorescence spectroscopy. And finally, the downcycling procedures were performed to obtain isolation of the various components of the reaction, then purification and repolymerization. Herein, the design and development of new fluorophores via a general strategy and its application in macromolecular degradation through luminescence in spatial and temporal scales herald a new generation of functional molecules.

S,S-/S,O-cyclic fluorophores
The CA contains bis-vinylogous methanethioesters which can be decoupled through releasing methyl mercaptan. As depicted in Scheme 1, products As and Bs were obtained through reacting CA with chemical reagents containing HS-R-SH (Both "HS" and "SH" represent sulfhydryl functional groups), HS-R-OH which undertook addition-elimination and subsequent intramolecular cyclization. The synthetic routes and molecular characterizations through NMR and high-resolution mass spectroscopy (HRMS) can be seen from the Supporting information. We then investigated the photophysical properties of the fluorophores (S,S-/S,O-cyclic molecules), respectively. Further, structural comparisons, experimental operations, single crystals analysis, and density functional theory calculations (DFT) were carried out and discussed in each group of these fluorophores (see below).

Cyclization-induced emission on S,S-cyclic fluorophores
In the group of S,S-cyclic fluorophores ( Figure 1A), the normalized UV-Vis spectra ( Figure 1B) demonstrated blueshifts from CA at λ abs = 400 nm, seven-membered molecule A1 at λ abs = 413 nm, to six-membered molecule A2 at λ abs = 386 nm, and five-membered molecule A3 at λ abs = 370 nm, respectively, in phosphate buffered saline (PBS, pH = 7.4, 1% dimethyl sulfoxide (DMSO) as cosolvent) (Table S1). There was 30 nm wavelengh blueshifts in absorbance due to cyclization and reduced ring size on CA. While there was no fluorescence observed for CA, slight emission for A1, and a 2-fold increase for A2, but a big enhancement up to 88-fold growth for the five-membered A3 ( Figure 1C) in PBS buffer (pH = 7.4, 1% DMSO). These data indicated that the optical properties for the chromophores were modulated by the cyclization and ring size. In addition, there were large Stokes shifts (>100 nm) for the fluorophores, and a wide range of fluorescence lifetimes from 0.6 to 87 ns (Table S1), which indicated that due to the charge transfer state lowering the bond order of the alkene, caused a 90 • rotation in the excited state (Scheme 2).
To understand these physicochemical properties, we calculated the energy gap (E g ) between the highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) levels using DFT ( Figure 1D). They followed the role of small E g for CA (3.90) and A1 (3.78), increasing with A2 (4.01) and A3 (4.09), which matches the trend of absorbance blueshift and fluorescence increase with the reduced ring sizes. So, it is reasoned that the difference caused by the ring-induced state of charge transfer, resulting in a more π→π* to n→π* transition variation on the alkene bond of the S,S-cyclic fluorophors ( Figure 2). Infrared and Raman spectroscopy were then performed to investigate the vibrational states of these molecules ( Figures S1 and S2). The five-membered ring A3 had an increase in the π-bond order of the exocyclic double bond over A1, A2, and CA, resulting in a redshift of the associated stretching vibration peak in both infrared and Raman spectroscopy ( Figures S1 and S2).
Next, we explored further the fluorescence properties of A3 in experiments of the solvent effect, water fraction titration, and temperature effect. As was revealed from the solvent effect, A3 exhibited strong emission under aqueous condition over other common organic solvents ( Figure  S3). While there was a fluorescence enhancement observed in A3 (10 μM) with water fraction increasing (0%-99%) in DMSO, accompanied with 42 nm emission wavelength redshift (λ em = 450-492 nm) ( Figures 1E and S4). Furthermore, upon decreasing the temperature from 60 • C to 0.5 • C in PBS buffer (pH = 7.4, 1% DMSO), the emission intensity of A3 demonstrated a 4-fold enhancement ( Figure 1F), which can be ascribed to the less conformational vibration. There was a good linear relationship (R 2 = 0.988) formed between λ em = 494 nm and temperature ranges (20 • C-50 • C) ( Figure 1F, inset). Furthermore, the six-membered A2 also displayed a 3-fold fluorescence increase when decreasing the solvent temperature from 60 • C to 0.5 • C ( Figure S5). Above all, lowering the temperature, adding poor solvent (water) or increasing the viscosity were all methods to limit the intramolecular vibrations of the molecule to enhance fluorescence. Next, single crystals of CA, A1-A3 were obtained by the slow volatilization from ethanol or dichloromethane ( Figures 1G and S6). It can be seen that CA, A1, and A2 showed intermolecular stacking mode with torsion configurations in the alkyl ring part in each single cell. The interlaced packing mode and J-aggregate states with vertical distance between the two planes were 4.145 Å (CA) > 3.308 Å (A1) > 2.629 Å (A2), respectively ( Figure S6). Thus, the reason to explain the low quantum yields or fluorescence quenching of CA, A1, A2 was not the formation of a stacking pattern that quenched the luminescence through π-π interaction. Now, it turns to the fundamental question: what is the difference among these structures? While comparing the molecules, the S,S-cyclization and then the ring size in the structures are believed to make the difference and the ring-determined conformational changes are affecting the excited state energy relaxation and regulating the optical behaviors on the fluorophores. That is to say small-sized rigid ring is the key to fluorescent switch.

2.3
Proposed mechanism for the photoluminescence As was discussed above, the approach for modulating optical responses of luminophores was via chemically triggered cyclization on CA. The charge separation state was occurred on the cyclized molecule As which accounts to explain the UV-Vis blueshifts over CA. While the energy relaxation due to the ring vibration on As toward the excited state, especially in the six-and seven-membered ring structures led to fluorescence quenching ( Figure 2). In terms of the five-membered fluorophore, that is, A3, the less vibration and inhibition of intermolecular π-π interaction due to its more planar configuration suppressed the signal quenching as the source of luminescence. From the Jablonski diagram, it was predicated that the chromophores with S,S-cyclization resulted in conformational constraints and structural rigidity with ring sizes reducing, that regulated the change in the conformational freedom of the vibrational and rotational motion of the emitter and should be responsible for the electron transition mode of the emitter in the excited state. Thus, rigid cyclization-induced emission enhancement due to reduced conformational energy loss was explained for the photoluminescence.

Ring-controlled fluorescent switch on S,O-cyclic fluorophores
Next, as a proof-of-concept mechanism, we studied S,Oheterocyclic molecules B1-B3 ( Figure 3A). The same optical phenomenon as with the S,S-cyclic fluorophores was also observed, from CA to B1-B3, showing UV-Vis absorbance blueshifts at λ abs = 400, 347, 342, 333 nm, respectively ( Figure 3B, Table S1). In the fluorescence spectra, the five-membered molecule B3 had the strongest fluorescence emission at λ em = 480 nm, while the six-membered molecule B2 was only half of that intensity (at λ em = 494 nm) in the aqueous condition ( Figure 3C, Table S1). While the fluorescence of the seven-membered molecule B1 could be hardly detected ( Figure 3C, Table S1), as like CA. In addition, large Stokes shifts (>140 nm) were also observed in the S,O-cyclic fluorophores which indicated the charge transfer state in the excited state (Table S1, Scheme 2). From the DFT calculation ( Figure 3D), the HOMO/LUMO gaps (E g ) followed the trend of B3 (4.34) > B2 (4.256) ≈ B1 (4.261) > CA (3.90), which matched the optical differences in absorbance and emission ( Figure 3B,C).
Next choosing B3 as a model molecule, we explored further the effect of water, temperature, viscosity, and single crystal of the molecules. First, fluorescence of B3 showed significant enhancement with water over the range of 0%-90% in DMSO (Figures 3E and S9). [7] Further, emission intensity of B3 at λ em = 471 nm was enhanced with more than 4-fold when decreasing the temperature from 50 • C to −15 • C in methanol, resulted in a good linear relationship between fluorescence and temperature (R 2 = 0.97) ( Figure 3F, inset). Furthermore, the viscosity of the solution containing B3 was increased tuning with glycerol (0%-50% volume), which led to nearly 2-fold fluorescence growth and a linear curve with R 2 = 0.996 in the relationship ( Figure S10). In the crystal structure of B3, the adjacent indanedionalkene planes completely overlap with each other, and the vertical distance between the stacks was measured to be approximately 3.459 Å ( Figure 3G), which corresponds to the formation of the weak intermolecular π-π interaction. [8] Thus, the Htype aggregation that occurs along the x-axis with a coplanar geometry explains the emission redshift ( Figure 3G). [3b,9] So far, the hypothesis of rigid cyclization-induced emission enhancement due to the reduced conformational energy loss was conformed on both the S,S-/S,O-heterocyclic indanedionalkene fluorophores which was envisioned in the future applications.

Fluorescent switch on the substituted fluorophores
To generalize this approach for the development of new luminogens and improving the photophysical performances, we then extended the working principles on the CA with different substituents ( Figure 4A). The substituents we used included bromo, chloro, nitro, methoxy, and amino groups (Figure 4), which were incorporated onto the skeleton of the structure (CA-X). In the UV-Vis spectrum, that 1,2-ethanedithiol (EDT) and BME cyclized chromophores showed blueshifts in absorbance and the enhanced fluorescence intensity under neutral conditions, over the started CA-X ( Figure 4B, Figures S13-S15 and Table S2). Again, rigid ring-controlled fluorescent switch successfully applied to fluorophores substituted with electron-withdrawing and electron-donating groups (i.e., A3-X and B3-X), except for B3-NO 2 bearing strong fluorescence quenching group typical of nitro ( Figures S14 and S15). Noticeably, fluorescence emission for B3-NH 2 demonstrated a 360-fold enhancement over CA-NH 2 ( Figure 4B). In addition, fluorophore B3-NH 2 , with a strong electron donating group, led to an emission redshifting to λ em = 556 nm over other B3-X ( Figure 4C). Interestingly, B3-Br and B3-Cl, with heavy atoms imparted by halide ions, displayed strong emission under aqueous condition ( Figure S16 and Table S2). This is interesting because the heavy atom effect usually induces the spin-orbit coupling and enhances intersystem crossing, thus quenching the emission. [10] The introduction of an amine group into the donor-πacceptor motif not only enhanced the fluorescence intensity, but also gave a larger Stokes shift (196 nm) and higher quantum yield (5.15%) (Table S2). It was revealed from a water titration that the fluorescence intensity of B3-NH 2 (10 μM, λ ex = 360 nm) increased and then decreased in the range of 20%-99%, while the wavelength continuously redshifted from λ em = 455 to 556 nm ( Figure 4E). The photo images of the B3-NH 2 emission changed from blue to yellow with water in the mixture under 365 nm UV lamp ( Figure 4D). In addition, the emission displayed different colors with maximum wavelengths ranging from 430 to 556 nm among a variety of common solvents (Figures 3J and 4F) due to the polarity effect on the donor-π-acceptor system. Moreover, the bathochromic shift of B3-NH 2 in protonic solvents (EtOH, MeOH, and H 2 O) indicates that the formation of hydrogen bonds allows for more effective dipole-dipole interactions between the excited B3-NH 2 and the protonic solvents and leads to enhanced stabilization effects, reducing the energy level of the excited B3-NH 2 to produce a greater bathochromic shift. [11] In addition, decreasing temperature from 40 • C to −15 • C in dichloromethane led to a fluorescence enhancement and a 10 nm bathochromic shift (λ em = 465 to 475 nm) for B3-NH 2 . This is probably due to a reduced ring motion in low temperatures ( Figure 4H). Furthermore, plotting absorption wavelengths of B3-X (X = H, Cl, Br, and NO 2 ) against the substituent's Hammett σ m values resulted in linear relationships, with R 2 = 0.98 for absorption ( Figure  S17). This correlation between substituent and optical performance might provide a tool for predicting and designing fluorophore structures to achieve the desired optical property. So far, it can be seen that comparing with the started molecule CA-NH 2 with free methyl sulfides, the S,O-cyclized B3-NH 2 with rigid five-membered ring demonstrated superior optical properties ( Figure 4I).
Combining all these optical results at the small molecule level (i.e., CA, CA-X, As, Bs, A3-X, and B3-X), we can conclude that the bis-vinylogous in such molecules acts as a fluorescent switch, which can cause the fluorescence to turn on when it reacts with the nucleophilic substrate to generate a rigid small size ring. More importantly, the obtained fluorophores are sensitive to the microenvironment of temperature, viscosity, polarity, and hydrogen bonding, and these properties bode well for their multiple applications.

Fluorescence tracking polymer degradation
In certain scenarios, rapid degradation properties would be in high demand in plastics, [12] hydrogels, [13] as well as other categories of medical polymers. [14] Visualization and quantitation of polymer degradation extent are of great importance for tracking and control of material properties. Hence, as a proof-of-concept application, we then investigated the chemically triggered degradation of a non-luminescent polymer PHE, accompanied by changes in fluorescence behavior of the generated B3 with rigid ring ( Figure 5A). The decoupling reaction between the CA and BME could cleave the backbone of the linear polymer to release the original coupling partner and the optical indicator, a process exhibiting fluorescence turn-on.
The polymer PHE was synthesized by polycondensation reaction between CA (1 equiv., 26.5 wt%) and 1,6-hexanedithiol (1 equiv., 15.9 wt%) through thiol-thiol scrambling in chloroform at 70 • C for 1 h (see the Supporting information). The resulting mixture was purified to generate a yellow powder with a single peak recorded by the gel permeation chromatography (GPC; Figure S19). And then, the kinetic experiments of PHE (20 mg, 4.7 μmol, calculated from 1 H NMR quantitative analysis, Figure S21) degraded by BME (10 μL, 286 mM) in the presence of trimethylamine (TEA) were carried out in chloroform, which was tracked by 1 H NMR. It can be seen, as the peak intensity of protons (peak a) in the methylene at 3.08-3.11 ppm and terminal methyl protons (peak d) at 2.61 ppm ( Figure 5A,B) on the polymer PHE rapidly decreased within 13 min, in the meantime, the peak intensity of methylene protons (peak b) at 4.84 ppm and methylene protons (peak c) at 3.42 ppm belonging to indicator B3 (Figure 5A,B) dramatically enhanced ( Figure S20). The continuous monitoring showed that the degradation of the polymer and generation of the indicator was completed within 60 min.
Next, the UV-Vis spectra to monitor PHE degradation were carried out for the samples processed in chloroform after dilution 1,000 times to DMSO. The intensity of absorbance at peak λ abs = 332 nm increased and λ abs = 370 nm decreased as a function of time ( Figure 5D) with an isosbestic point at 345 nm. The ratio of absorbance value A 332 nm /A 370 nm reached a plateau within 60 min ( Figure 5D, inset). While the fluorescence of the solutions containing degraded polymer and indicators were detected at certain times after dilution 1,000 times in PBS (pH = 7.4, 20% DMSO) ( Figure 5E). As can be revealed from the fluorescence photo images monitored at different time points, the signal was turn-on at the initial 5 min and then enhanced with time over a range of 120 min (Figure 5E), which can be observed by the naked eye ( Figure 5F). In the spectroscopy, the fluorescence at λ em = 476 nm increased as a function of the reaction time due to generation of indicator B3 and the degradation finished within 60 min. When using different concentration of BME (0, 28, 42, 72, 114, 144, 172, 200, 216, 286 mM, respectively) to decompose PHE (20 mg, 4.7 μmol), we observed dose-dependent fluorescence enhancement ( Figure 5G). The fluorescence intensity at λ em = 476 nm reached a maximum at around 60 min with a good linear relationship between fluorescence intensity and BME concentrations (R 2 = 0.989) ( Figures S22 and S23).
To achieve quantitative tracking of the polymer degradation, we then built up the relationships between molecule weight-fluorescence intensity and BME concentrations. In the presence of BME (72, 144, 172, 200, 216 mM), the molecular weight of degraded PHE was recorded by the 1 H NMR spectra (Figures S24-S28, Table S3) while the fluorescence signals were scanned, after processing 60 min for each of the sample. Linear calibration curves were achieved between molecular weight of PHE and the fluorescence intensity (476 nm) of the degradation solution as a function of BME concentrations ( Figure 5H), which reveals good relationships between the molecular weight/fluorescence intensity and BME concentrations (R 2 = 0.9 and 0.99). In addition, based on the fluorescence signal as above, we then built up the relationship between BME as a trigger and B3 as a product for the polymer degradation ( Figure 5I), through a prepared calibration curve between fluorescence and doses of B3 with the known standard samples ( Figure  S29). The linear correlation coefficient (R 2 = 0.989) was formed between concentration of BME and B3, which was an important reference for the chemically triggered stoichiometry in polymer degradation between decoupling reagent and resulted product. So far, through chemically triggered cyclization and induced emission enhancement, we achieved the rapid degradation of non-fluorescent chain polymer into original partner and fluorescent indicator, along with the real-time photoluminescence tracking and quantifying.
Additionally, to investigate the polymer degradation visualizing in the solid state, the polymer PHE (M n ∼ 4.0 kDa, 25 mg) was added into an acetonitrile/water (1/1 in volume) mixture. As was seen in Figure S30A, there was no degradation and fluorescence occurring on the sample ( Figure S30), which indicate the stability of the solid PHE in the absence of chemical trigger BME. While the solid of PHE sample was gradually degraded and dissolved within 48 h, triggered by BME (286 mM) ( Figure S30C). The fluorescence of the supernatant was increased over the time range of 0-48 h  Figures 5G and S29 F I G U R E 6 Schematic diagram of "depolymerization-recycling-repolymerization" process and photo images of recycled B3 and repolymerized PHE illustrated by day light and 365 nm UV light, respectively observed by the naked eye, and the fluorescence intensity reached to its maximum within 24 h ( Figure S30D). Furthermore, the degradation solution (200 mg PHE) was diluted with dichloromethane and washed three times with 0.1 M HCl to remove triethylamine and residual BME, the organic phase was concentrated and the residue was washed with cyclohexane to obtain B3 (106 mg, 85% yield) (Figures 6  and S31), while the filtrate was concentrated and dried to obtain 1,6-hexanedithiol (45 mg, 57% yield, Figure S32). And then, another 75 mg of CA was used to react with the recovered 1,6-hexanedithiol (45 mg) re-prepare the polymer PHE in CHCl 3 , after purification, 83 mg repolymerized PHE was obtained and characterized by NMR, which indicated the effectivity of the recycled monomer and successful downcycling operation (Figures 6 and S33).

CONCLUSION
To summarize, we developed a series of benzene-based indanonalkene fluorophores through coupling reaction on a conjugated ketene dimethylthioacetal acceptor. Cyclizationinduced emission and enhancement through reduced vibrational energy loss were explained to the photoluminescence in the different ring-sized luminogens. The general methodology was extended in the development of fluorophores with different substituents on the indanonalkene skeleton with improved photophysical properties. The chemical reaction, along with the photoluminescence, was successfully applied in the degradation of a linear chain polymer and achieved the tracking and quantitation of depolymerization process, as well as the recovery and repolymerization of degraded monomer. Based on the methodology, more fluorogenic tools would be developed and some under investigation in our group for further applications, such as visualization of polymeric topological changes, biomolecular labeling, cellular microenvironment sensing, etc.

Materials E T H I C S S TAT E M E N T
There are no ethical issues in this work.