Dendronized Gelatin-Mediated Synthesis of Gold Nanoparticles

Thermoresponsive dendronized gelatins (GelG1) or gelatin methacrylates (GelG1MA) were used as precursors to modulate the efficient reduction of Au(III) to form stable gold nanoparticles (AuNPs) through UV irradiation. These dendronized gelatins were obtained through the amidation of gelatin or gelatin methacrylates with dendritic oligoethylene glycols (OEGs). Crowded OEG dendrons along the gelatin backbones create a hydrophobic microenvironment, which promotes the reduction of Au(III). Gelatin backbones act as ligands through the electron-rich groups to facilitate the reduction, while the dendritic OEGs provide shielding effects through crowding to form a hydrophobic microenvironment, which not only enhances the reduction but also stabilize the formed AuNPs through encapsulation. The effects of dendron coverage on the dendronized biomacromolecules and their thermoresponsiveness on the reduction kinetics were examined. Dendronized gelatin/AuNPs hydrogels were further prepared through the in situ photo-crosslinking of GelG1MA. The modification of natural macromolecules through dendronization presented in this report facilitates a novel platform for the environmentally friendly synthesis of noble metal nanoparticles, which may form a new strategy for developing smart nano-biosensors and nano-devices.


Introduction
The modification of natural macromolecules has attracted more and more attention in recent years to create novel environmentally friendly materials, which not only inherit excellent biofunctions from biomacromolecules, but also can be endowed with new properties through functional modification [1][2][3][4]. Natural macromolecules carry abundant functional groups, such as hydroxyl, amine and carboxyl groups, which can be readily used for chemical modification. In recent years, various strategies have been applied for the modification of biomacromolecules, aimed at different functions and properties. Modified natural macromolecules show advantages in stability, safety, responsibility, and sustainability, and have been widely used in fields from smart materials to biomedicine [5].
Gold nanoparticles (AuNPs) exhibit unique physical and chemical properties, and have attracted extensive attention continuously. They have been applied in various fields, such as optics, electronics, catalysis [6] and liquid crystal composites [7], as well as in drug delivery [8], biological imaging [9], and cancer treatment [10]. AuNPs can be prepared through the reduction of Au(III) by various agents, such as alcohols, amines, carboxylic acids or borohydride. Since the nanoparticles formed tend to aggregate in aqueous solutions, the effective strategy to avoid this is to protect them by stabilizers, such as thiols, polymers or polyelectrolytes [11]. For this purpose, many polymer stabilizers, such as microgels, dendritic macromolecules, hydrogels and latex particles, have been applied. Polymer stabilizers can control the reduction rate and sizes of formed nanoparticles [12]. Traditional synthesis methods usually use toxic chemicals, which may cause some pollution to the environment. In addition, the residues of chemical substances may affect the

Synthesis and Characterization of GelG1 and GelG1MA
Dendronized gelatins were prepared through amidation with dendritic OEG scribed previously [29]. In order to examine the dendritic effects on the modulatio reduction of AuCl4 − into AuNPs, two dendronized gelatins-GelG115:1 and Ge with an OEG dendron substitution degree of 86.7% and 54.4%, respectively, were sized and compared to the naked gelatin. In addition, GelG1MA1:1 with meth pendants was synthesized from GelG11:1 (Scheme S1), allowing for the formation dronized gelatin hydrogels via photo-crosslinking. The substitution degrees of O dron and methacryloyl moiety in GelG1MA1:1 were 45.6% and 48.5%, respectiv structures of the dendronized gelatins were confirmed with 1 H NMR spectroscopy 2 shows the assembled 1 H NMR spectra of naked gelatin, GelG11:1 and GelG1MA pared to the unmodified gelatin and GelG11:1, two new signals (a and b) at δ 6 and 5.8 ppm were observed, corresponding to the protons from the methacryloyl The intensity of signal c at δ 1.45 ppm, which corresponds to the methyl group f methacryloyl moiety, increased obviously after modification. This confirms that ryloyl groups were successfully grafted to the gelatin. These dendronized gelatins characteristic thermoresponsive properties, and the cloud points (Tcps) for G GelG15:1 and GelG1MA1:1 at a concentration of 0.5 wt% were found to be 35.7 °C and 48.6 °C, respectively ( Figure S1).

Synthesis and Characterization of GelG1 and GelG1MA
Dendronized gelatins were prepared through amidation with dendritic OEGs, as described previously [29]. In order to examine the dendritic effects on the modulation of the reduction of AuCl 4 − into AuNPs, two dendronized gelatins-GelG1 15:1 and GelG1 5:1 -with an OEG dendron substitution degree of 86.7% and 54.4%, respectively, were synthesized and compared to the naked gelatin. In addition, GelG1MA 1:1 with methacryloyl pendants was synthesized from GelG1 1:1 (Scheme S1), allowing for the formation of dendronized gelatin hydrogels via photo-crosslinking. The substitution degrees of OEG dendron and methacryloyl moiety in GelG1MA 1:1 were 45.6% and 48.5%, respectively. The structures of the dendronized gelatins were confirmed with 1 H NMR spectroscopy. Figure 2 shows the assembled 1 H NMR spectra of naked gelatin, GelG1 1:1 and GelG1MA 1:1 . Compared to the unmodified gelatin and GelG1 1:1 , two new signals (a and b) at δ 6.19 ppm and 5.8 ppm were observed, corresponding to the protons from the methacryloyl moiety. The intensity of signal c at δ 1.45 ppm, which corresponds to the methyl group from the methacryloyl moiety, increased obviously after modification. This confirms that methacryloyl groups were successfully grafted to the gelatin. These dendronized gelatins exhibit characteristic thermoresponsive properties, and the cloud points (T cp s) for GelG1 15:1 , GelG1 5:1 and GelG1MA 1:1 at a concentration of 0.5 wt% were found to be 35.7 • C, 45.8 • C and 48.6 • C, respectively ( Figure S1).

Preparation of AuNPs in the Presence of Dendronized Gelatins with UV Irradiation
Reduction of AuCl4 -in the presence of GelG1 at room temperature through ultraviolet irradiation was examined first. Aqueous solution of GelG1 containing AuCl4 − changed from colorless to purple after 30 min and to red after 60 min of irradiation ( Figure  3a). UV/vis spectra were, therefore, used to follow the reduction process. As shown in Figure 3a, a strong absorption around 520 nm appeared and its intensities increased with irradiation time, corresponding to the surface plasma vibration from AuNP [30]. The plasma resonance peak intensities of the solution increased with irradiation time, indicating that the amount of AuNPs formed by reduction increased with time. For comparison, the reduction of Au(III) in the presence of naked gelatin was performed, and a similar plasma resonance peak appeared with its intensities also increased with irradiation time ( Figure S2a). This indicates that the reduction of Au(III) can be conducted in either dendronized gelatins or naked gelatin. We suppose the reduction was driven by phenolic hydroxyl or other reducing groups from the peptide backbone [31]. However, the reduction kinetics are quite different when comparing the naked gelatin to that of GelG1. As shown in Figure 3b,c, after irradiation for a certain time, maximum absorbance (Amax) and reduction rates ((At − A0)/Δt) in the case of GelG1 are much higher than that for gelatin, suggesting that the dendritic OEGs are supportive to enhance the reduction kinetics. Compared to that from naked gelatin, the wavelength of the maximum absorption (λmax) from GelG1/AuNPs is blue shifted (Figure 3d), indicating that the formed AuNPs should have smaller sizes. The morphology and sizes of the AuNPs were observed by a transmission electron microscope (TEM). As shown in Figure S2c, uniformed AuNPs with sizes in the range of 8-12 nm were observed from gelatin, while sizes in the range of 5-8 nm were observed from GelG1 ( Figure S2d). The AuNPs formed in the matrix of dendronized gelatins and gelatin were investigated by dynamic light scattering (DLS). The hydrodynamic radii (Rhs) of GelG1/AuNPs and Gel/AuNPs at 25 °C are plotted in Figure S2e

Preparation of AuNPs in the Presence of Dendronized Gelatins with UV Irradiation
Reduction of AuCl 4 − in the presence of GelG1 at room temperature through ultraviolet irradiation was examined first. Aqueous solution of GelG1 containing AuCl 4 − changed from colorless to purple after 30 min and to red after 60 min of irradiation ( Figure 3a). UV/vis spectra were, therefore, used to follow the reduction process. As shown in Figure 3a, a strong absorption around 520 nm appeared and its intensities increased with irradiation time, corresponding to the surface plasma vibration from AuNP [30]. The plasma resonance peak intensities of the solution increased with irradiation time, indicating that the amount of AuNPs formed by reduction increased with time. For comparison, the reduction of Au(III) in the presence of naked gelatin was performed, and a similar plasma resonance peak appeared with its intensities also increased with irradiation time ( Figure S2a). This indicates that the reduction of Au(III) can be conducted in either dendronized gelatins or naked gelatin. We suppose the reduction was driven by phenolic hydroxyl or other reducing groups from the peptide backbone [31]. However, the reduction kinetics are quite different when comparing the naked gelatin to that of GelG1. As shown in Figure 3b,c, after irradiation for a certain time, maximum absorbance (A max ) and reduction rates ((A t − A 0 )/∆t) in the case of GelG1 are much higher than that for gelatin, suggesting that the dendritic OEGs are supportive to enhance the reduction kinetics. Compared to that from naked gelatin, the wavelength of the maximum absorption (λ max ) from GelG1/AuNPs is blue shifted (Figure 3d), indicating that the formed AuNPs should have smaller sizes. The morphology and sizes of the AuNPs were observed by a transmission electron microscope (TEM). As shown in Figure S2c, uniformed AuNPs with sizes in the range of 8-12 nm were observed from gelatin, while sizes in the range of 5-8 nm were observed from GelG1 ( Figure S2d). The AuNPs formed in the matrix of dendronized gelatins and gelatin were investigated by dynamic light scattering (DLS). The hydrodynamic radii (R h s) of GelG1/AuNPs and Gel/AuNPs at 25 • C are plotted in Figure S2e, and it was found that R h of GelG1/AuNPs is approximately 35 nm, which is slightly smaller than that of Gel/AuNPs (about 38 nm).
The effect of the grafting ratio of dendritic OEGs on the modified gelatins for the preparation of AuNPs was checked. As shown in Figure 3c and Figure S2b, the reduction rates of both GelG115:1 and GelG15:1 were greater than that of the naked gelatin, and GelG115:1 exhibited the highest rate. The grafting coverage of dendritic OEGs in GelG115:1 (86.7%) was higher than that of GelG15:1 (54.4%), suggesting that abundant dendritic OEGs may have provided enhanced microconfinement to shield the solvation of metal anions, resulting in the promotion of the in situ reduction of Au(III) to form AuNPs. Therefore, an increase in the grafting ratio of dendritic OEGs is helpful to create the confined microenvironment to increase the reduction rate. Reduction of HAuCl4 to form AuNPs was also performed in the presence of GelG1MA through UV irradiation. GelG1MA1:1 carried the same dendritic OEGs as GelG1 but with a slightly lower grafting ratio (45.6%). The reduction process was followed by UV/vis spectroscopy, and the spectra are shown in Figure 4a. Similar to the case from GelG1, there was a plasma resonance absorption near 550 nm after UV irradiation and its intensity increased with irradiation time, indicative of forming AuNPs. The aqueous solution of GelG1MA1:1 containing AuCl4 − changed from colorless to purple after UV irradiation for 60 min at 25 °C (Figure 4b), which also indicates the reduction of Au(III) into Au(0). For comparison, the reduction of AuCl4 − was also performed in the presence of GMA, where gelatin carries no dendrons but only the methacrylol pendants ( Figure  S3a), and the solution turned light blue. Compared to the case of GMA/AuNPs, the The effect of the grafting ratio of dendritic OEGs on the modified gelatins for the preparation of AuNPs was checked. As shown in Figure 3c and Figure S2b, the reduction rates of both GelG1 15:1 and GelG1 5:1 were greater than that of the naked gelatin, and GelG1 15:1 exhibited the highest rate. The grafting coverage of dendritic OEGs in GelG1 15:1 (86.7%) was higher than that of GelG1 5:1 (54.4%), suggesting that abundant dendritic OEGs may have provided enhanced microconfinement to shield the solvation of metal anions, resulting in the promotion of the in situ reduction of Au(III) to form AuNPs. Therefore, an increase in the grafting ratio of dendritic OEGs is helpful to create the confined microenvironment to increase the reduction rate.
Reduction of HAuCl 4 to form AuNPs was also performed in the presence of GelG1MA through UV irradiation. GelG1MA 1:1 carried the same dendritic OEGs as GelG1 but with a slightly lower grafting ratio (45.6%). The reduction process was followed by UV/vis spectroscopy, and the spectra are shown in Figure 4a. Similar to the case from GelG1, there was a plasma resonance absorption near 550 nm after UV irradiation and its intensity increased with irradiation time, indicative of forming AuNPs. The aqueous solution of GelG1MA 1:1 containing AuCl 4 − changed from colorless to purple after UV irradiation for 60 min at 25 • C (Figure 4b), which also indicates the reduction of Au(III) into Au(0). For comparison, the reduction of AuCl 4 − was also performed in the presence of GMA, where gelatin carries no dendrons but only the methacrylol pendants ( Figure S3a), and the solution turned light blue. Compared to the case of GMA/AuNPs, the reduction rates for GelG1MA 1:1 were higher (Figure 4b) and the maximum absorption from GelG1MA 1:1 /AuNPs solution stronger (Figure 4c). These suggest that reduction efficiency for GelG1MA 1:1 was significantly enhanced when compared to that for GMA. This again supports that dendritic OEG pendants may have provided a hydrophobic microenvironment through their crowded packing along the gelatin chains, which strengthened the interactions between the polymers and Au(III) to promote the reduction.
Molecules 2022, 27, x FOR PEER REVIEW 6 of 13 reduction rates for GelG1MA1:1 were higher ( Figure 4b) and the maximum absorption from GelG1MA1:1/AuNPs solution stronger (Figure 4c). These suggest that reduction efficiency for GelG1MA1:1 was significantly enhanced when compared to that for GMA. This again supports that dendritic OEG pendants may have provided a hydrophobic microenvironment through their crowded packing along the gelatin chains, which strengthened the interactions between the polymers and Au(III) to promote the reduction.  Compared to that from GMA, the wavelength of the maximum absorption from GelG1MA 1:1 /AuNPs blue shifted (Figure 4d), indicating that the formed AuNPs should be smaller in size. The XPS spectra of the AuNPs were recorded, and the binding energies of the Au (4f 7/2 ) and Au (4f 5/2 ) peaks appeared at 84.17 eV, 87.79 eV, respectively, which can be assigned to Au(0) (Figure 4e). This further supports the reduction of Au(III) to form Au(0). The morphology and sizes of the AuNPs was further observed by TEM. As shown in Figure 4f, uniformed AuNPs with sizes in the range of 4-8 nm were observed from GelG1MA 1:1 . The lattice plane spacing was found to be 2.15 Å, which corresponds to the (111) plane of the face-centered cubic gold crystals [32]. Differently, AuNPs with much larger sizes in the range of 8-12 nm were observed from GMA ( Figure S3b). As shown in Figure S3c, the R h of GelG1MA 1:1 /AuNPs is approximately 52 nm, slightly smaller than that of GMA/AuNPs (about 55 nm). The particle size of AuNPs prepared in the presence of GelG1MA is smaller, which may be due to the hydrophobic microenvironment provided by the crowded OEG dendrons. This makes the interaction between the dendronized gelatins and Au(III) stronger, which promotes the reduction. In addition, the steric hindrance effect from the dendrons affords better stability to the formed AuNPs.

Effects of the Thermoresponsive Properties of the Dendronized Gelatins on the Formation of AuNPs
Since these dendronized gelatins are thermoresponsive, their modulation of the reduction at elevated temperature was examined. The effect of the thermoresponsive properties of dendronized gelatins on the preparation of AuNPs can be investigated by UV irradiation above the cloud point (50 • C), compared to that below the cloud point (25 • C). From the UV/vis spectra in Figure 5a, when irradiated at 50 • C, which is above the T cp of GelG1, plasma resonance with much stronger intensities than that at room temperature appeared (Figure 5b). This indicates that reduction kinetics can be greatly enhanced by increases in temperature. To verify whether this enhancement was originated from temperature increase itself or the combined effects from the thermally induced dehydration and collapse of the dendritic OEGs, the reduction of Au(III) at elevated temperature in the presence of naked gelatin, which is not thermoresponsive, was performed and compared ( Figure S4). As shown in Figure 5c, reduction kinetics increased in both naked gelatin and GelG1 with temperature, with the latter more pronounced. This indicates again that crowded dendritic OEGs are supportive to the reduction. Furthermore, the maximum absorption wavelength of AuNPs blue shifted to 525 nm from 575 nm (Figure 5d), indicating that the size of AuNPs gradually decreased at elevated temperature.
Similar to the case at room temperature, the aqueous solution of GelG1MA 1:1 containing AuCl 4 − irradiation with UV light at 50 • C transformed from a colorless solution into purple, as shown in Figure S5a. From the UV/vis spectra in Figure 6a, when irradiated at 50 • C, which is above the T cp of GelG1MA 1:1 , the maximum absorption showed much enhanced intensities at the same UV irradiation time when compared to the case at room temperature (Figure 6b), indicating that the reduction rate was obviously accelerated by the increased temperature. However, the reduction rate of AuNPs by GMA at elevated temperature remained relatively slow, as shown in Figures 6c and S5b. This again suggests that the enhanced hydrophobic microenvironment from the dehydrated dendritic OEGs at elevated temperatures enhanced the reduction. Furthermore, when irradiated at 50 • C, the maximum absorption wavelength of AuNPs blue shifted to 525 nm from 560 nm (Figure 6d), indicating that the size of AuNPs gradually decreased at elevated temperature.

Stability of AuNPs Formed in the Matrices of Dendronized Gelatins
The above results show that dendronized gelatins can reduce Au(III) to form welldefined AuNPs through UV irradiation. This method does not need the addition of any external reducing agent. To explore the feasibility of the methodology developed in present work, the stability of the AuNPs formed in the matrix of dendronized gelatins were investigated. After storage at 25 • C for 30 days, the pink solutions from GelG1 15:1 /AuNPs or GelG1MA 1:1 /AuNPs remained clear and homogeneous, as shown in Figure 7a, indicating their high stability. In contrast, Gel/AuNPs became turbid and precipitation was observed. UV/vis spectroscopy was also applied to follow the stability of the AuNPs. After 20 days of storage at room temperature, A max from the dendronized gelatins did not change significantly (Figure 7b), while A max from the Gel/AuNPs showed the most obvious change. The above results indicate that dendronized gelatins act as reducing agents for the transformation of Au(III) into Au(0), and at the same time, as a stabilizer to the formed nanoparticles. The electrostatic or coordination interactions between the dendronized gelatins and the formed AuNPs should have contributed to their stability in aqueous media. On the other hand, crowding from the dendritic units in the dendronized gelatins may have also provided an enveloping effect to prevent the AuNPs from agglomeration, making them dispersed with high stability [28].

Stability of AuNPs Formed in the Matrices of Dendronized Gelatins
The above results show that dendronized gelatins can reduce Au(III) to form welldefined AuNPs through UV irradiation. This method does not need the addition of any external reducing agent. To explore the feasibility of the methodology developed in present work, the stability of the AuNPs formed in the matrix of dendronized gelatins were investigated. After storage at 25 °C for 30 days, the pink solutions from GelG115:1/AuNPs or GelG1MA1:1/AuNPs remained clear and homogeneous, as shown in Figure 7a, indicating their high stability. In contrast, Gel/AuNPs became turbid and precipitation was observed. UV/vis spectroscopy was also applied to follow the stability of the AuNPs. After 20 days of storage at room temperature, Amax from the dendronized gelatins did not change significantly (Figure 7b), while Amax from the Gel/AuNPs showed the most obvious change. The above results indicate that dendronized gelatins act as reducing agents for the transformation of Au(III) into Au(0), and at the same time, as a stabilizer to the formed nanoparticles. The electrostatic or coordination interactions between the dendronized gelatins and the formed AuNPs should have contributed to their stability in aqueous media. On the other hand, crowding from the dendritic units in the dendronized gelatins may have also provided an enveloping effect to prevent the AuNPs from agglomeration, making them dispersed with high stability [28].

Fabrication of Dendronized Gelatin/AuNP Hydrogels
GelG1MA contains unsaturated double bonds. Through UV irradiation, they can act as a matrix for in situ reduction to form AuNPs, and simultaneously transform into hydrogels via photo-crosslinking polymerization (Scheme S2). Therefore, a mixture of GelG1MA1:1 (10 wt%), HAuCl4 (2.5 mmol/L) and photo-initiator 2959 (2 wt%) was illuminated by ultraviolet light (30 W, 365 nm), which changed from colorless solution into dark purple hydrogel (Figure 8a). For comparison, GelG1MA1:1 hydrogel was prepared by the same method without HAuCl4, which exhibited a light-yellow color. Both hydrogels can keep their shape when heated to 50 °C, indicating chemical crosslinking. The reduction of HAuCl4 in the presence of GelG1MA1:1 to form AuNPs and transform into hydrogel was traced by UV/vis spectroscopy. The plasma resonance absorption peak of AuNPs appeared at 520 nm after UV irradiation for 5 min (Figure 8a), and the intensity of the absorption peak remained unchanged after about 15 min (Figure 8b), indicating that GelG1MA1:1 had reduced most of Au(III) into AuNPs at that condition. The λmax exhibited a blue shift to 505 nm from about 508, indicating that the elongation of irradiation afforded AuNPs with slightly smaller sizes. The dendronized gelatin/gold nanoparticle composite

Fabrication of Dendronized Gelatin/AuNP Hydrogels
GelG1MA contains unsaturated double bonds. Through UV irradiation, they can act as a matrix for in situ reduction to form AuNPs, and simultaneously transform into hydrogels via photo-crosslinking polymerization (Scheme S2). Therefore, a mixture of GelG1MA 1:1 (10 wt%), HAuCl 4 (2.5 mmol/L) and photo-initiator 2959 (2 wt%) was illuminated by ultraviolet light (30 W, 365 nm), which changed from colorless solution into dark purple hydrogel (Figure 8a). For comparison, GelG1MA 1:1 hydrogel was prepared by the same method without HAuCl 4 , which exhibited a light-yellow color. Both hydrogels can keep their shape when heated to 50 • C, indicating chemical crosslinking. The reduction of HAuCl 4 in the presence of GelG1MA 1:1 to form AuNPs and transform into hydrogel was traced by UV/vis spectroscopy. The plasma resonance absorption peak of AuNPs appeared at 520 nm after UV irradiation for 5 min (Figure 8a), and the intensity of the absorption peak remained unchanged after about 15 min (Figure 8b), indicating that GelG1MA 1:1 had reduced most of Au(III) into AuNPs at that condition. The λ max exhibited a blue shift to 505 nm from about 508, indicating that the elongation of irradiation afforded AuNPs with slightly smaller sizes. The dendronized gelatin/gold nanoparticle composite hydrogels were successfully obtained through in situ reduction and photo crosslinking polymerization, prompted by UV irradiation. This method has the advantages of fast photo crosslinking to form hydrogels and the high efficiency of Au(III) reduction into gold nanoparticles. This work provides a simple, rapid and in situ reduction method for the preparation of gold nanoparticles' composite hydrogels, which may have broad application prospects in biomedical fields such as bioimaging, biosensing and drug delivery. We believe these thermoresponsive dendronized gelatin hydrogel combined AuNP composites could be important in promising bioapplications.
Molecules 2022, 27, x FOR PEER REVIEW 10 of 13 hydrogels were successfully obtained through in situ reduction and photo crosslinking polymerization, prompted by UV irradiation. This method has the advantages of fast photo crosslinking to form hydrogels and the high efficiency of Au(III) reduction into gold nanoparticles. This work provides a simple, rapid and in situ reduction method for the preparation of gold nanoparticles' composite hydrogels, which may have broad application prospects in biomedical fields such as bioimaging, biosensing and drug delivery. We believe these thermoresponsive dendronized gelatin hydrogel combined AuNP composites could be important in promising bioapplications.

Instrumentation and Measurements
1 H NMR spectra were recorded on a Bruker AV 500 (500 MHz) spectrometer. UV/vis measurements were conducted on a JASCO V-750 spectrophotometer equipped with a thermostatically regulated bath. Aqueous polymer solutions were placed in the spectrophotometer (path length 1 cm). Fluorescence (FL) spectra were recorded in a fluorescence spectrometer (Horiba Jobin Yvon Fluorolog-3) equipped with a temperature control device. Dynamic light scattering (DLS) measurements were performed on a DynaPro Nanostar. X-Ray photoelectron spectroscopy (XPS) spectra were recorded on an AXIS Supra+ (Shimadzu, Manchester, UK). This system uses a focused monochromatic Al Kα X-ray (1486.7 eV) source for excitation. The binding energies were referenced to C1s at 284.8 eV from hydrocarbon to compensate for the charging effect. The micromorphology of in situformed spherical AuNPs was imaged by using a field-transmission electron microscope (JEM-2100, INCAX-Max80) with an accelerating voltage of 160 kV. Sample preparation: An appropriate amount of solution was taken into the centrifuge tube, and centrifuged with a rotating speed 10,000 RPM for 15 min. The supernatant was discarded, and ultrapure water was added to the ultrasonic dispersion and centrifuged twice. After

Instrumentation and Measurements
1 H NMR spectra were recorded on a Bruker AV 500 (500 MHz) spectrometer. UV/vis measurements were conducted on a JASCO V-750 spectrophotometer equipped with a thermostatically regulated bath. Aqueous polymer solutions were placed in the spectrophotometer (path length 1 cm). Fluorescence (FL) spectra were recorded in a fluorescence spectrometer (Horiba Jobin Yvon Fluorolog-3) equipped with a temperature control device. Dynamic light scattering (DLS) measurements were performed on a DynaPro Nanostar. X-ray photoelectron spectroscopy (XPS) spectra were recorded on an AXIS Supra+ (Shimadzu, Manchester, UK). This system uses a focused monochromatic Al K α X-ray (1486.7 eV) source for excitation. The binding energies were referenced to C1s at 284.8 eV from hydrocarbon to compensate for the charging effect. The micromorphology of in situformed spherical AuNPs was imaged by using a field-transmission electron microscope (JEM-2100, INCAX-Max80) with an accelerating voltage of 160 kV. Sample preparation: An appropriate amount of solution was taken into the centrifuge tube, and centrifuged with a rotating speed 10,000 RPM for 15 min. The supernatant was discarded, and ultrapure water was added to the ultrasonic dispersion and centrifuged twice. After discarding the supernatant, absolute ethanol was added to the ultrasonic dispersion, which was then dispersed on copper wire.

Fabrication of AuNPs in the Matrix of Gelatin or Modified Gelatins
Gelatin or modified gelatins (4 mg) were mixed with HAuCl 4 ·4H 2 O (0.2 mg) in aqueous solution (1 mL). The mixture was stirred by a vortex oscillator, then irradiated by ultraviolet light (365 nm, 30 W) for a designed time period (60-90 min). The distance between the lamp and the sample was fixed at 10 cm. UV/vis spectroscopy was applied to follow the reduction process. AuNPs obtained from the matrix of naked gelatin, gelatin methacryloyl, dendronized gelatin, and dendronized gelatin methacryloyl are named as Gel/AuNPs, GMA/AuNPs, GelG1/AuNPs, and GelG1MA/AuNPs, respectively.

Fabrication of GelG1MA/AuNP Hydrogel
The mixture of GelG1MA 1:1 (100 mg/mL), HAuCl 4 (1 mg/mL) and photo-initiator 2959 (20 mg/mL) was oscillated for 30 s in the scroll oscillator to form a uniformly mixed solution. The mixture was illuminated for 30 min by ultraviolet lamp (30 W, 365 nm), and the mixture changed from a colorless solution into a dark purple hydrogel.

Conclusions
We have developed a green and convenient method in the present work to generate stable AuNPs by using dendronized gelatins or dendronized gelatin methacrylates as precursors, triggered by UV irradiation. The reduction can be processed efficiently without any additional reducing chemicals. Dendronized gelatins act as both a reducing agent for the transformation of Au(III) into Au(0) and a stabilizer for the formed nanoparticles through crowded dendritic OEG units. The dendritic OEGs provide a hydrophobic microenvironment to promote the formation of AuNPs, and the reduction rate can be enhanced with an increase in grafting coverage of the dendritic OEGs, as well as the thermally induced aggregation of polymer chains. The high stability of the nanoparticles in the matrix of dendronized gelatins has been verified through long-term storage. Therefore, the formation of AuNPs within the envelope of dendronized gelatins avoids cumbersome operation and environmental pollution. Dendronized gelatin/AuNP composite hydrogels can be readily prepared from dendronized gelatin methacryloyl via UV irradiation through simultaneous reduction and photo-crosslinking polymerization. This clean and environmentally friendly method for the synthesis of AuNPs may be extended for the fabrication of other noble metal nanoparticles and shed light on promising applications in nano biosensors and nano devices.