Lignin Nanosphere-Supported Cuprous Oxide as an Efficient Catalyst for Huisgen [3+2] Cycloadditions under Relatively Mild Conditions

In this work, low-cost lignin nanospheres were fabricated and further applied as an efficient and sustainable support for preparing cuprous oxide (Cu2O) “green” catalyst by using electrospraying technology. The unalloyed lignin, a special three-dimensional molecular structure, was successfully processed into uniform nanospheres under an electrospraying condition. The synthesized lignin-supported Cu2O catalyst had a well-defined nanosphere structure, and Cu2O nanoparticles with sizes less than 30 nm were supported by exposed layers of lignin nanospheres. There were C–O–Cu bonds formed between the lignin nanospheres and the metallic nanoparticles. The lignin nanospheres and the lignin nanosphere-supported catalyst werfe characterized by utilizing XRD, SEM, TEM, XPS, EDS, and TGA. The immobilization of Cu2O nanoparticles on the lignin nanospheres was beneficial for dispersion of the Cu2O nanoparticles and preventing their aggregation, which could cause catalyst deactivation, which favored the Huisgen [3+2] cycloaddition reaction. The triazole synthesis results indicated that the lignin nanosphere-supported Cu2O catalyst had a high catalytic performance with 99% yield under solvent-free conditions. Furthermore, the as-synthesized catalyst could be recycled for four times without significantly losing its catalytic activity.


Introduction
"Click" chemistry has a wide scope, gives high yields, and forms irreversible carbon-heteroatom and carbon-carbon bonds [1]. It uses only the most practical and reliable chemical reactions to connect a diversity of structures bearing a wide variety of functional groups [2]. Therefore, it has attracted much attention in many research areas, such as catalyst design, polymer synthetization, material science, synthesizing libraries of compounds, and drug development [3]. Monovalent copper-mediated azide-alkyne Huisgen [3+2] cycloaddition is a highly typical and important "click" reaction [4]. It involves the ligation of azides and terminal alkynes to generate triazoles, and usually needs a copper salt in conjunction with a base. The catalyst can be a Cu(I) salt or Cu(I) generated in situ by the reduction of Cu(II) salts, usually in organo-aqueous media [5], and allows facile and reliable production of 1,4-disubstituted 1,2,3-triazoles.
Nowadays, there is a growing interest in the synthesis of environmentally benign catalysts [6] because of their added advantages, such as requiring mild reaction conditions and being simple to recover and regenerate, as well as being environmentally friendly [7]. It is desirable to develop high-performance and "green" heterogeneous catalysts for "click" chemistry. Recently, the development of Cu-based heterogeneous catalysts with the presence of copper or cuprous oxide nanoparticles has significantly increased catalytic activity for the construction of triazole molecules, owing to the enhanced surface area of copper or cuprous oxide and unique catalyst structure-activity relationships [8][9][10]. The supported catalysts are simple to prepare. The inexpensive catalysts are practical for their applications. Moreover, the immobilization of small Cu or Cu 2 O nanoparticles on supports makes the catalyst easily reusable with excellent recyclability performance, largely reduces nanoparticle leakage, and avoids environmental hazards. Currently, the most commonly used solid supports for heterogeneous catalysts include silica [11], zeolites [12], graphene [13], magnetic materials [14], and soluble and insoluble polymers [15]. Furthermore, various other supports have been used in different catalytic reactions [16][17][18][19][20][21]. However, some of these supports of the heterogeneous catalysts may not be sufficient in the near future. As a result, it is crucial to switch to cheap and biorenewable resources. Natural biopolymers [22][23][24] have been considered as attractive candidates to create high-performance biobased catalysts due to their abundance, renewability, biodegradability, and low cost [25]. Lignin is one of the most important biopolymers from renewable resources, owing to its abundance (being the second most plentiful plant biopolymer on the earth, after cellulose). The production of lignin is over 50 million tons worldwide annually, while its utilization is rather limited except for its use in biochar or activated carbon [26]. Therefore, lignin is a desirable raw material for fabricating value-added products for various applications. Recently, the synthesis of lignin-based functional materials and their potential applications have attracted much attention, such as anode materials for lithium-ion batteries [27], supercapacitors [28], and electrocatalysts [29]. Furthermore, lignin derivatives exhibit interesting and attractive abilities in improving the performance of photovoltaic devices [30]. As a heterogeneous and rigid polymer of phenolic nature, it is the only biomass that is based on aromatic units. It is composed of phenylpropane, including p-coumaryl alcohol, coniferyl alcohol, and sinapyl alcohol, which makes it highly polar with a large amount of hydroxyl groups. Moreover, typical C-O links of the lignin are β-O-4, α-O-4, and 4-O-5; and C-C links are β-5, 5-5, β-1, and β-β linkages [31][32][33][34]. It has hydroxylic groups and aromatic methoxy, carboxylic, carbonylic, and ethereal moieties. Most importantly, lignin owns three-dimensional molecular structures. These make lignin an efficient and stable ligand for the preparation of biocatalysts.
Electrospinning is a relatively convenient and versatile strategy to produce nanofibers from polymer solutions or blends. However, lignin possesses complex three-dimensional structures with various types of functional groups. The lignin is formed by oxidative coupling of the main three kinds of lignin precursors; the reaction sites abound upon forming of the body polymers. The special architectural structures of lignin have led to little success in creating absolute lignin electrospinning nanofibers. Current strategies for using lignin in electrospinning nanofibers focus on integrating it with other natural or synthetic polymers; for instance, Kai et al. [35] engineered nanofibrous composites with different lignin mass fractions into lignin-polymethylmethacrylate by electrospinning technology. Interestingly, we found that the special molecular structure of lignin made it easy to be processed into nanospheres when electrospraying lignin precursor solution at an appropriate concentration in DMF solvent under a determined voltage condition. The inherent phenolic hydroxyl, carboxylic, and carbonylic groups in lignin molecules are beneficial for the adsorption of Cu ions, among others, to load and coordinate [36,37], and the three-dimensional molecular structure contributes to making lignin a stable and favorable support for catalysts.
Herein, we report a novel straightforward method to fabricate lignin nanospheres in quantity (in the grams scale) by utilizing electrospraying technology. Then, we successfully exploit it as an effective support for Cu 2 O nanoparticles for preparing a high-performance cuprous oxide heterogeneous catalyst. Above all, lignin and monovalent copper could form Cu 2 O@L composites owing to the abundant carbon-oxygen bonds in lignin molecules. Meanwhile, the strongly polar phenolic hydroxyl and alcoholic hydroxyl groups contributed to its reducing property [38,39], which favored the reduction of cupric sulfate in aqueous solution for the preparation of cuprous oxide catalyst. In particular, Cu 2 O@L was investigated to catalyze a series of Huisgen "click" reactions and showed excellent activity for 1,2,3-triazole syntheses. The reactions proceeded under air conditions in the absence of organic solvents, without adding any base or external heating, which made the method to be a simple, powerful, and environmentally benign alternative. Furthermore, the catalytic performance of the Cu 2 O@L was researched in terms of a series of azides and terminal alkynes. As compared with other copper-mediated Huisgen "click" cycloaddition reactions, "click" reactions using the Cu 2 O@L exhibited a higher yield and required a shorter reaction time and relatively mild reaction conditions.

Materials and Methods
The alkali lignin (M W = 6000) was purified from poplar wood pulping black liquor using acid treatment; the black liquor was provided by Shuntai Co., Ltd. (Changsha, Hunan, China). Cu 2 O (~30 nm) for comparison was purchased from Shandong Xiya Chemical Reagent Co., Ltd. (Linyi, Shandong, China). All the chemicals were used as received without further purification.

Electrospraying Lignin Nanopheres
In a typical process, lignin solution with a concentration of 35 wt % was prepared by dissolving 0.9 g lignin into 1.1 g DMF liquid solution in a flask. The mixture liquid was then stirred at room temperature for 2 h until the lignin was completely dissolved. The electrospraying setup used in this study consisted of a syringe and needle (ID = 0.84 mm), high voltage supply, a ground electrode, and a collector (stainless steel sheet). A syringe pump connected to the syringe controlled the flow rate. The lignin nanospheres were obtained by electrospraying at a positive voltage of 15 kV and a negative voltage of 1 kV, with a tip-to-collector distance of 15 cm and a solution flow rate of 0.75 mL/h. The procedure was carried out at 25 • C and 60% ambient humidity. The lignin nanospheres were stripped down from aluminum paper and then dried in an oven. Furthermore, the electrosprayed lignin nanospheres were preliminarily heated at 280 • C in air for 2 h and then were heated to 800 • C with a heating rate of 1 • C/min. Finally, the nanospheres were held for 30 min under N 2 atmosphere in a tube furnace to obtain lignin carbon nanospheres.

Preparation of Cu 2 O@L
The preparation of Cu 2 O@L used the methods as follows: In a 50 mL test tube, 10 mL of double-distilled water was added into the test tube and stirred at room temperature. 500 mg lignin nanospheres were added to form a suspension. Then, 10 mL (2%, w/v) of aqueous copper sulphate solution was added into the suspension with stirring for an hour. This process was carried out in N 2 atmosphere to eliminate the dissolved oxygen present in the solution. An aqueous solution of hydrazine hydrate (4 M) as the main reducing agent was added to reduce the copper sulfate to Cu 2 O nanoparticles sufficiently. After 2 h of continuous stirring, the catalyst was separated using a centrifuge (10,000 rpm, 10 min) and washed with double-distilled water for three times to remove the unentrapped particles and ions. The wet sample was then dried in an oven to obtain Cu 2 O@L.

Catalytic Performance Test
The catalytic ability of Cu 2 O@L was evaluated by the synthesis of triazoles from alkyl azide and alkyne. 1.2 mmol of alkyne, 1.0 mmol of alkyl azide, and 20 mg (1.2 mol %) Cu 2 O@L were added into a reaction tube under organic solvent-free conditions. Then, the reaction mixture was stirred for 3 h at room temperature. After the reaction, the product was separated by ethyl acetate extraction from Cu 2 O@L, and the catalyst was recovered by filtration or centrifugation (8000 rpm, 10 min) for reuse. The products were further purified by column chromatography on silica gel using ether/ethyl acetate (5:1) as the eluent. The triazoles were obtained after solvent evaporation and the yield of the products was calculated.
For the practical applications of such heterogeneous systems, the lifetime of the catalyst is a very significant factor for evaluation of catalyst performance. Therefore, the number of cycles and the catalytic activity in terms of reaction yield using the recycled catalyst were also studied. After the completion of the first reaction, the product was extracted using ethyl acetate and the catalyst was recovered by simple centrifugation and dried at room temperature. Then, a fresh reaction was performed with new reactants, under the same conditions.

Analyses
All 1 H spectra of the products were recorded on a FT-NMR (600 MHz, Bruker Corp., Karlsruhe, German) spectrometer, and 13 C spectra were recorded on a Bruker FT-NMR (151 MHz). NMR chemical shifts are given as δ values (ppm) with reference to TMS as the internal standard. FTIR spectra were recorded on a Bruker Nicolet 6700 FTIR spectrophotometer in the range of 400-4000 cm −1 using KBr pellets. The Cu content was determined by a Hitachi Z-2000 AAS (Tokyo, Japan). SEM images were recorded on a Zeiss EVO-18 (Oberkochen, Germany) operating at 10 kV, and TEM images were recorded on a JEM-2100 (HR) TEM (JEOL, Tokyo, Japan) working at 200 kV. The fluorescence images were acquired using a LSCM (Leica TCS SP5, Wetzlar, Germany) equipped with a diode laser (405 nm), using a 100× oil objective with numerical aperture of 1.40, and three samples were observed under the same conditions. The UV-vis-NIR absorption spectrum of Cu 2 O was acquired with a PerkinElmer Lambda 750 spectrophotometer (Waltham, MA, USA). FL were taken on a FLS-980 spectrometer (Edinburgh Instruments, Ltd., Edinburgh, UK). The samples were excited by a pulse laser (405 nm). An appropriate filter was also used before the PMT to cut off any other stray light. Topographic and phase images of samples were obtained using an AFM (Bruker Dimension Fastscan) in tapping mode. XRD measurements were carried out at room temperature using a Bruker D8 Advance X-ray powder diffractometer with Ni-filtered Cu-Kα radiation (λ = 0.154 nm) from 5 • -80 • . XPS measurements were performed on an Axis Ultra DLD instrument using Al-Kα radiation (hν = 1486.6 eV) with contaminated C as an internal standard (C 1s = 284.6 eV). EDS was measured with a Horiba XMAX X-act energy dispersive spectroscope that was attached to the Oxford Instruments (Abingdon, UK). The size and the size distribution of Cu 2 O@L were measured by dynamic light scattering on a Nano-Zetasizer (Zetasizer NanoZS, Malvern Instruments Ltd., Worcestershire, UK) at 25 • C under a scattering angle of 173 • at λ = 633 nm. TGA (Q500, TA Instruments, New Castle, DE, USA) was carried out in an aluminum crucible by heating to 650 • C at a heating rate of 20 • C min −1 with a nitrogen flow of 25 mL min −1 .

Characterization of the Electrosprayed Lignin Nanosphere and Cu 2 O@L
As shown in Figure S1, the concentration of the lignin solution shows a significant influence on the lignin morphology under the electrospraying condition. When the concentration of the lignin solution is 25%, 35%, or 45% (w/w), electrosprayed lignin exhibits a nanosphere shape (Figure S1a-c). Under these concentration conditions, the surface tension is greater than the electric force, which plays a key role on the nanospherical shape of lignin. However, the electrosprayed lignin exhibits the similar fiber morphology when the lignin concentration is higher than 55%, due to lignin molecular chains twining around each other ( Figure S1d). As compared with the electrosprayed lignin at other concentrations, the lignin nanosphere at the concentration of 35% solution had a more uniform morphology and therefore was chosen for further studies.
The as-prepared electrosprayed lignin nanospheres show a regular spherical morphology with a diameter less than 1 µm for the most part ( Figure 1a). From the SEM images, the sizes of lignin nanospheres show the average size of 605 ± 5 nm. As shown in Figure 1b, Cu 2 O@L shows a "raspberry" morphology. Furthermore, the spherical morphology of the electrosprayed lignin nanosphere still remains after carbonization at 800 • C under N 2 atmosphere (Figure 1c). The reason for this is that the good rigid and three-dimensional molecular structure obviously maintains its volume unchanged before and after the carbonization process. The EDS of Cu 2 O@L illustrates that there are C, O, and Cu elements in the catalyst (Figure 1d). Figure 1e shows the EDS mapping image with all the elements of Cu 2 O@L, while Figure 1f-h shows the EDS mapping images of the C, O, and Cu elements, respectively. The SEM and EDS results showed that the Cu 2 O@L was a spherical structure containing the Cu element. Furthermore, the Cu concentration of the catalyst is confirmed to be 7.76% by AAS. Cu elements, respectively. The SEM and EDS results showed that the Cu2O@L was a spherical structure containing the Cu element. Furthermore, the Cu concentration of the catalyst is confirmed to be 7.76% by AAS. The morphology of the electrosprayed lignin nanospheres and Cu2O@L were further studied using LSCM. As shown in Figure 2a, the raw lignin has an irregular shape and good fluorescence characteristic, which is attributed to its molecular structure containing repeated units of phenylpropane. Interestingly, as compared with the electrosprayed lignin nanospheres (Figure 2b), Cu2O@L shows weaker fluorescence ( Figure 2c). Fluorescence spectrums of lignin, lignin nanospheres, and Cu2O@L were further investigated, which were in accordance with the results of LSCM. The lignin nanospheres show stronger fluorescence intensity than raw lignin. This is probably because the aggregation-induced emission appears among lignin molecules as the lignin is processed into nanospheres by electrospraying. The Cu2O nanoparticles can lead to fluorescent quenching of lignin (Figure 2d), and energy transfer from lignin to Cu2O nanoparticles occurs owing to some overlap between the emission wavelength range of lignin (420-700 nm) and the absorption wavelength of Cu2O (400-900 nm) ( Figure 2e). Furthermore, the interaction of phenolic hydroxyl groups and carbon-oxygen and cuprous bonds destroying conjugated structures or clusters of the carbonyl groups in lignin molecules [40] may result in the weakening of the characteristic fluorescence of the lignin and an obvious blue-shift of the luminescent spectrum of Cu2O@L. Aggregation-induced emission of lignin nanospheres that were prepared by electrospraying is observed (Figure 2f), similarly to the phenomenon of lignin derivatives reported by Qiu and coworkers [41].  The morphology of the electrosprayed lignin nanospheres and Cu 2 O@L were further studied using LSCM. As shown in Figure 2a, the raw lignin has an irregular shape and good fluorescence characteristic, which is attributed to its molecular structure containing repeated units of phenylpropane. Interestingly, as compared with the electrosprayed lignin nanospheres (Figure 2b), Cu 2 O@L shows weaker fluorescence ( Figure 2c). Fluorescence spectrums of lignin, lignin nanospheres, and Cu 2 O@L were further investigated, which were in accordance with the results of LSCM. The lignin nanospheres show stronger fluorescence intensity than raw lignin. This is probably because the aggregation-induced emission appears among lignin molecules as the lignin is processed into nanospheres by electrospraying. The Cu 2 O nanoparticles can lead to fluorescent quenching of lignin (Figure 2d), and energy transfer from lignin to Cu 2 O nanoparticles occurs owing to some overlap between the emission wavelength range of lignin (420-700 nm) and the absorption wavelength of Cu 2 O (400-900 nm) ( Figure 2e). Furthermore, the interaction of phenolic hydroxyl groups and carbon-oxygen and cuprous bonds destroying conjugated structures or clusters of the carbonyl groups in lignin molecules [40] may result in the weakening of the characteristic fluorescence of the lignin and an obvious blue-shift of the luminescent spectrum of Cu 2 O@L. Aggregation-induced emission of lignin nanospheres that were prepared by electrospraying is observed (Figure 2f), similarly to the phenomenon of lignin derivatives reported by Qiu and coworkers [41]. Figure 2 shows TEM and high-resolution TEM (HRTEM) images of Cu 2 O@L as well. The diameter of the electrosprayed lignin nanospheres is less than 1 µm for the most part and some nanoparticles are present on their surfaces, as illustrated in Figure 2g. Noting that the Cu 2 O are irregular in shape with the average size of 20 ± 2 nm ( Figure 2h). The HRTEM image (Figure 2i) shows that the lattice spacing of the nanoparticles is about 0.24 nm [42], which is in agreement with the crystal lattice theoretical value for the (111) planes of Cu 2 O. The result indicates that the nanoparticles on lignin containing the Cu element are Cu 2 O. All these results show that small Cu 2 O nanoparticles have been successfully loaded onto the surfaces of the lignin nanospheres.
Polymers 2018, 10, x FOR PEER REVIEW 6 of 15 that the lattice spacing of the nanoparticles is about 0.24 nm [42], which is in agreement with the crystal lattice theoretical value for the (111) planes of Cu2O. The result indicates that the nanoparticles on lignin containing the Cu element are Cu2O. All these results show that small Cu2O nanoparticles have been successfully loaded onto the surfaces of the lignin nanospheres. As shown in Figure 3, height map images of AFM demonstrate the spherical shape of the electrosprayed lignin nanospheres (Figure 3a,b) and Cu2O@L (Figure 3e,f). Figure 3c shows the defective surface of the lignin nanosphere, which may contribute to it being a favorable catalyst support for preparing the high-performance lignin-based catalyst. There are two phases of Cu2O@L (Figure 3g), indicating Cu2O being successfully loaded onto the lignin nanospheres. Besides, the 3D images of AFM (Figure 3d,h) also prove the spherical structures of the lignin nanospheres and Cu2O@L. The catalyst Cu2O@L exhibits a nanoscale particle size mainly ranging from 100 to 1000 nm, which appears as a large peak at a size of around 600 nm ( Figure S2). The particle-size analysis is in accordance with SEM, TEM, and the AFM results mentioned above.
The crystal structure of Cu2O@L was confirmed by XRD patterns. As shown in Figure 4a-1, the dispersion peak at 2θ = 30-40° is a typical XRD diffraction pattern of noncrystalline lignin. Because the overlap peak of the lignin diffraction peaks heighten at 29°, 36°, and 42° of Cu2O [43], all of the sharp diffraction peaks (2θ = 29°, 36°, 42°, 62°, and 74°) can be indexed to the face-centered cubic structure of Cu2O (JCPDS file: 65-3288) (Figure 4a-2). There are still XRD diffraction peaks of Cu2O from the reused Cu2O@L (Figure 4a-3). The XRD results are consistent with the results of HRTEM, indicating the successful loading of Cu2O onto the lignin nanospheres.  Figure 3c shows the defective surface of the lignin nanosphere, which may contribute to it being a favorable catalyst support for preparing the high-performance lignin-based catalyst. There are two phases of Cu 2 O@L (Figure 3g), indicating Cu 2 O being successfully loaded onto the lignin nanospheres. Besides, the 3D images of AFM (Figure 3d,h) also prove the spherical structures of the lignin nanospheres and Cu 2 O@L. The catalyst Cu 2 O@L exhibits a nanoscale particle size mainly ranging from 100 to 1000 nm, which appears as a large peak at a size of around 600 nm ( Figure S2). The particle-size analysis is in accordance with SEM, TEM, and the AFM results mentioned above.
The crystal structure of Cu 2 O@L was confirmed by XRD patterns. As shown in Figure 4a     XPS and Cu L3VV Auger line results for Cu 2 O@L were studied to investigate the components of the catalyst and the chemical state of the Cu element. The XPS complete spectra shows C 1s, O 1s, and Cu 2p peaks (Figure 4b). The result illustrates that there are C, O, and Cu elements in Cu 2 O@L, corresponding with the EDS result. The binding energies of the Cu 2p of Cu 2 O@L are located at 932.5 and 952.5 eV. In addition, there is a satellite peak at 944.6 eV in the XPS spectrum of the Cu 2p of the Cu 2 O@L (Figure 4c). Furthermore, it is found that the peak of the Cu L3VV signals is centered at 916.8 eV, indicating the primary existence of Cu(I) in the catalyst [44]. According to the HRTEM, XRD, and XPS results, the nanoparticles on electrosprayed lignin nanospheres are calculated to be Cu 2 O.
Moreover, the structural characteristics of Cu 2 O@L were investigated by FTIR spectrum. Figure 4d-1 shows the FTIR spectra of lignin. The characteristic FTIR absorption peaks of alkali lignin (from poplar) are 1327 cm −1 , 1269 cm −1 , and 1115 cm −1 that are related to guaiacyls, the stretching vibrations of carbon-oxygen bonds, and ether bonds, respectively. The peaks at 3390 cm −1 , 1216 cm −1 , 1515 cm −1 , and 557 cm −1 are attributed to phenolic hydroxyl groups, carbon double bonds of benzene, and acyl-oxygen bonds, respectively [45]. As shown in Figure 4d-2, the FTIR spectrum of the electrosprayed lignin nanospheres is similar to that of lignin. Figure 4d-3 shows the FTIR spectrum of Cu 2 O@L. The peak at 3390 cm −1 shifts to 3379 cm −1 , and the peak at 1216 cm −1 weakens, indicating interaction of phenolic hydroxyl and cuprous groups. As compared with lignin and lignin nanospheres, the peak at 1705 cm −1 disappears in the FTIR spectrum of Cu 2 O@L. This was probably due to the weakened π bond of carbon and oxygen resulting from interaction of cuprous and carbon-oxygen bonds. There are also some differences in the fingerprint region. The characteristic peak at 628 cm −1 may be attributed to Cu 2 O, which is caused by the overlap of the peak the 632 cm −1 from lignin and the peak of 624 cm −1 from Cu 2 O.
Furthermore, thermal stabilities of the lignin nanospheres and Cu 2 O@L were displayed in Figure S3. The lignin nanospheres and the catalyst have similar thermal properties, in that both of them show decomposition temperatures above 150 • C. Because the Huisgen [3+2] cycloaddition reactions in this study were carried out at room temperature, the thermal properties of Cu 2 O@L could fully meet the temperature requirement of the reaction.

Catalytic Performance of Lignin Nanosphere-Supported Catalyst in the Huisgen "Click" Reaction
Solvent-free conditions confer to the reaction process simple operation, easy separation, and reduced environmental hazards [46]. With the aim to develop a "green" method for "click" chemistry, we attempted to synthetize triazoles under solvent-free conditions. The result is inspiring. As shown in Table S1, the reaction yield is almost 99% under solvent-free conditions without adding any base (with 1.2 mol % catalyst). The reason for this is that Cu 2 O@L may achieve better contact with reaction reagents in such a situation. The product is able to crystallize out from the reactants during the reaction process, and the whole reaction can be monitored visually under the solvent-free condition. Moreover, different solvents, such as water, alcohol and, methanol have also been adopted as solvents for the reaction of benzyl azide and phenylacetylene. As compared with the solvent-free condition, the reaction yields are slightly lower than those of the other solvents using Cu 2 O@L.
The classic cycloaddition reaction of benzyl azide and phenylacetylene was used to explore the optimum condition of the reaction. The results were encouraging, noting that the reaction between benzyl azide and phenylacetylene could be completed within 3 h under the solvent-free condition at ambient temperature. The effect of the Cu 2 O@L quantity on the reaction was investigated. The reaction almost did not take place under the solvent-free condition at room temperature even after 24 h of stirring without the catalyst. When 0.6 mol % Cu 2 O@L was used, the productivity reached 93%. The reaction could be completed in 3 h and provided a quantitative yield of the desired product with Cu 2 O@L. The optimized result (99% yield) was obtained within 3 h under the solvent-free condition at room temperature with 1.2 mol % Cu 2 O@L, while the yield of the reaction was only 27% and 25% using equivalent Cu 2 O nanoparticles as the catalyst under the solvent-free condition or using H 2 O as the solvent, respectively. Besides, the reaction did not take place when the lignin nanospheres were used alone.
After establishment of the optimum condition for the cycloaddition of benzyl azide and phenyl acetylene, a series of azides and terminal alkynes were further subjected to the Huisgen "click" reactions using Cu 2 O@L at room temperature under the solvent-free condition to further evaluate the efficiency of this methodology. The products obtained have been tabulated in Table 1. All the reactions could be completed within 3 h to give high yields of the corresponding 1,4-triazoles. The substituents of phenyl acetylene (electron-withdrawing, electron-rich, and heterocyclic) had no effect on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules  on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules on the reaction, and tolerance for variations in the azide component was also excellent. In particular, the catalyst was very effective for aliphatic alkynes as well, and we could isolate 97% of the product between 4-nitrobenzyl azide and 1-hexyne. The possible mechanism for the catalytic cycle is proposed in Scheme 1, in which the whole reaction process is related to two copper atoms. In our work, Cu(I) is also considered to be the most active species. Cu(I) coordinates with strongly polar phenolic hydroxyl groups in lignin molecules and can form Cu 2 O@L coordination compounds. Then, there is the formation of Cu 2 O@L-acetylidine composites by initial coordination with alkynes. The Cu 2 O@L-acetylidine composites are added to azide groups to form the active cycloaddition composites. Finally, the cycloaddition composites give 1,2,3-triazole.
Polymers 2018, 10, x FOR PEER REVIEW 11 of 15 Scheme 1. The proposed catalytic mechanism for the "click" reaction catalyzed by Cu2O@L (red colors are from reactants and green colors are from catalyst).

Discussion
Cu(I) has always been considered to be one of the most active species in "click" chemistry. In this work, the catalytic active center of Cu2O@L is monovalent copper. Furthermore, the lignin nanosphere support has improved the catalytic activity of the Cu2O nanoparticles for the yield of Huisgen "click" reactions, attributed to the formation of Cu2O@L composites ( Figure S4). Therefore, the lignin nanosphere support can further provide sufficient catalytic active sites, which favors the continuation of the azide-alkyne cycloaddition reaction. These may be the reasons why all the yields of the products are impressive under the solvent-free condition at room temperature using Cu2O@L. A comparison of some selected protocols in the literature and our method are listed in Table 2. The catalyst system was also characterized by electrochemistry ( Figure S5).
Cu2O@L could be reused for four times without obvious change in its activity. As shown in Figure S6a, it can be observed that with the increasing number of cycles of the reaction, the catalytic activity of the catalyst does not decrease significantly. The Cu2O@L works extremely well even up to four subsequent cycles with 95% yield. These observations suggest that after few cycles, the catalytic efficiency can be preserved, which is attributed to the stable structure of Cu2O@L. Metal leaching was studied by AAS analysis of the catalyst before and after four reaction cycles. The Cu element concentration is found to be 7.76% before the reaction and 7.62% after the reaction. The XRD result shows that there are still diffraction peaks in the reused catalyst (Figure 4a-3). The morphology of Cu2O@L after four cycles was investigated as well, as shown in Figure S6b,c. It is clearly shown that the Cu2O nanoparticles still remain irregular in shape and dispersed uniformly on the surface of the lignin nanospheres. The XRD and TEM results indicate that the lignin nanosphere is a favorable support for cuprous oxide to catalyze Huisgen [3+2] cycloaddition reactions.

Discussion
Cu(I) has always been considered to be one of the most active species in "click" chemistry. In this work, the catalytic active center of Cu 2 O@L is monovalent copper. Furthermore, the lignin nanosphere support has improved the catalytic activity of the Cu 2 O nanoparticles for the yield of Huisgen "click" reactions, attributed to the formation of Cu 2 O@L composites ( Figure S4). Therefore, the lignin nanosphere support can further provide sufficient catalytic active sites, which favors the continuation of the azide-alkyne cycloaddition reaction. These may be the reasons why all the yields of the products are impressive under the solvent-free condition at room temperature using Cu 2 O@L. A comparison of some selected protocols in the literature and our method are listed in Table 2.
The catalyst system was also characterized by electrochemistry ( Figure S5). Cu 2 O@L could be reused for four times without obvious change in its activity. As shown in Figure S6a, it can be observed that with the increasing number of cycles of the reaction, the catalytic activity of the catalyst does not decrease significantly. The Cu 2 O@L works extremely well even up to four subsequent cycles with 95% yield. These observations suggest that after few cycles, the catalytic efficiency can be preserved, which is attributed to the stable structure of Cu 2 O@L. Metal leaching was studied by AAS analysis of the catalyst before and after four reaction cycles. The Cu element concentration is found to be 7.76% before the reaction and 7.62% after the reaction. The XRD result shows that there are still diffraction peaks in the reused catalyst (Figure 4a-3). The morphology of Cu 2 O@L after four cycles was investigated as well, as shown in Figure S6b,c. It is clearly shown that the Cu 2 O nanoparticles still remain irregular in shape and dispersed uniformly on the surface of the lignin nanospheres. The XRD and TEM results indicate that the lignin nanosphere is a favorable support for cuprous oxide to catalyze Huisgen [3+2] cycloaddition reactions.

Conclusions
In summary, we developed a facile and powerful method to prepare Cu 2 O@L using widely available and sustainable lignin by electrospraying technology. Alkali lignin was successfully processed into uniform nanospheres when electrospraying lignin precursor solution at an appropriate concentration under a determined voltage condition. The lignin nanosphere was further exploited as part of a high-performance cuprous oxide heterogeneous catalyst, and the catalyst exhibited an excellent catalytic activity for the Huisgen "click" reaction under the solvent-free and mild conditions. As compared with other copper-mediated "click" reactions, reactions adopting the Cu 2 O@L showed higher yield (up to 99%) and required shorter reaction time and relatively mild conditions to synthesize 1,2,3-triazoles. The preparation process of the catalyst is economical, facile, and can be implemented on a large scale. In general, this work provided a "green" method for "click" chemistry and an efficient approach for application of the biomass resource lignin.  Table S1. Effect of polarity of solvent on Huisgen "click" reaction of azides and alkynes using Cu2O@L in different solvents; Spectroscopic data, 1H and 13C NMR Spectra of products.