Cu Nano particles immobilized on silk-fibroin as a green and biodegradable catalyst for copper catalyzed azide-terminal, internal alkynes cycloaddition

Article abstract of DOI:10.1002/aoc.6019

Copper immobilized on silk fibroin was successfully prepared and fully characterized using powder X-ray diffraction, scanning electron microscopy–energy-dispersive X-ray spectroscopy, Fourier transform-infrared, CHN elemental analysis, and inductively cou

Full text of DOI:10.1002/aoc.6019

Received: 6 February 2020
DOI: 10.1002/aoc.6019
Revised: 11 August 2020
Accepted: 25 August 2020
F U L L P A P E R
Cu Nano particles immobilized on silk-fibroin as a green
and biodegradable catalyst for copper catalyzed azide-
terminal, internal alkynes cycloaddition
Hakimeh Mirzaei
| Hossein Eshghi
| Seyed Mohammad Seyedi
Department of Chemistry, Faculty of
Science, Ferdowsi University of Mashhad,
Mashhad, Iran
Copper immobilized on silk fibroin was successfully prepared and fully charac-
terized using powder X-ray diffraction, scanning electron microscopy–energy-
dispersive X-ray spectroscopy, Fourier transform-infrared, CHN elemental
analysis, and inductively coupled plasma-atomic emission spectroscopy. Cata-
lytic activity of this catalyst was examined in the azide-alkyne cycloaddition
reaction with internal and terminal alkynes at room temperature under mild
conditions. The reusability of the heterogeneous supported Cu catalyst was
examined four times without significant loss of catalytic activity.
Correspondence
Hossein Eshghi, Department of
Chemistry, Faculty of Science, Ferdowsi
University of Mashhad, Mashhad, Iran.
Email: heshghi@um.ac.ir
Funding information
Research Council of Ferdowsi University
of Mashhad, Grant/Award Number:
3/44652
K E Y W O R D S
azide-alkyne cycloaddition reaction, click reaction, copper-fibroin catalyst, ligand free
1 | INTRODUCTION
Until now, various catalytic systems, including homo-
geneous and heterogeneous, have been proposed and
Over the past years, a variety of scientific and methodo-
logical development have been achieved, that encourage
chemists to use these tools to more easily perform synthe-
sis and related purification/separation processes.
Huisgen 1, 3-dipolar cycloaddition between organic
azides and alkynes is one among many synthetic tools
that has become quite well-known over the recent
decade, mainly due to its key improvement in term of
rate and regioselectivity.[1]
The prodigious synthetic potential of primary proto-
cols for Huisgen 1, 3-dipolar cycloaddition reaction
between organic azides and alkynes was limited by the
significant disadvantages such as heating requirement,
prolonged reaction time, and formation of structural iso-
mers due to the lack of selectivity.[2] Wonderful
Cu(I) catalyst modification introduced at the dawn of last
decade, allowed the cycloaddition to occur at room
temperature or with moderate heating leading to the
exclusive formation of 1,4-disubstituted triazole with
shorter workup and purification steps.[3,4] Now, this
method has engulfed almost every part of chemistry and
applied sciences.[5,6]
employed for copper(I) catalyzed azide-alkyne cycloaddi-
tion reactions with terminal and internal alkynes.[7–9]
Some of these catalytic systems are Cu(I) salt; CuSO4-
ascorbate
polymers,[11,12]
system[10]
zeolite,[13]
;
immobilized
and
Cu(I)
onto:
hydrotalcite,[7]
nanomagnetic supported thiourea–copper(I),[14] two
magnetic nanoparticle-supported copper(I)[15]; copper
nanoparticles[16]; and nanostructured copper oxide.[17]
In catalytic systems, it has always been important to
recover the catalyst from the reaction mixture and reuse
it. Especially in the case of CuACC reactions which are
commonly used for synthesis of substances that are
related to biological application,[18] the complete separa-
tion of the catalyst from the reaction mixture is a matter
of concern due to the toxicity of copper ions to cells and
organisms.[19] This illustrates the need to use heteroge-
neous catalytic systems in which the catalyst is
immobilized on a support. However, for homogeneous
catalytic systems trace amounts of copper ions can be
removed from the reaction mixture and isolated products,
by washing with EDTA solution[20] but this procedure is
not always convenient, especially in parallel or
Appl Organomet Chem. 2020;e6019.
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MIRZAEI ET AL.
combinatorial systems. Immobilizing the copper catalyst
on a heterogeneous support is one of the main strategies
to circumvent the aforementioned problems. According
to the principles of green chemistry and the need for min-
imal use of chemicals and the production of Eco-friendly
catalysts, many researchers have focused studies on
applying natural biopolymers, such as silk,[21,22]
starch,[23] cellulose,[24] and chitosan,[8,25] as activated
supports for metal particles. Many of the electron-rich
functional groups in the protein bind with the metal
surface easily, leading to the bioconjugation. The main
characteristic of these biopolymers is their high amine
group content, leading to interesting chelating properties
for metal cations.[25–27] This situation eliminates the need
to use surface activator ligands. Sometimes, the sensitiv-
ity of these ligands to temperature, moisture and the
presence of oxygen, the tedious multistep synthesis,
hence the high cost of the ligands, the use of various
additives, and the need for an inert atmosphere during
the synthesis of the catalyst as well as during the
reaction, limits the use of these catalysts.[28–30] Also, uni-
form dispersion of amino groups leads to uniform
dispersion of the synthesized nanoparticles immobilized
on these supporting materials. In this paper, for the first
time a copper nanoparticle immobilized on silk fibroin
was prepared. The catalytic activity of this new heteroge-
neous catalyst investigated in one-pot synthesis of
1,2,3-triazoles by reaction of terminal and internal
alkynes and azides at room temperature under mild
conditions.
D4 X-ray diffractometer with Ni-filtered Cu KR radiation
(40 kV, 30 mA).
2.2 | Preparation of support
The raw silk fibers were processed three times (30 min
each times) in 0.5 wt.% Na2CO3 solution at 80ꢀC–90ꢀC to
remove sericin (degumming), rinsed with deionized
water and dried at room temperature.[32,33]
2.3 | Preparation of supported Cu0-
nanoparticles
The degummed fibroin (0.2 g) dissolved in deionized
water (10 ml) and sonicated for 10 min. Then a solution
of (0.1 M) copper(П) acetate (5 ml) was added to mixture
and was stirred for 6 h led to the absorption of Cu(П) on
the fibroin surface. Afterwards, copper(П) supported
fibroin was removed from the solution and finally 0.1 M
sodium borohydride solution (10 ml) was added dropwise
to this and was stirred for 24 h to provide a copper(0)-
fibroin catalyst. After completion of the reduction proce-
dure, catalyst was washed four times with water and
ethanol (50:50) mixture and dried at room temperature.
The Cu/Fib. catalyst structure is schematically shown in
Scheme 1.
2.4 | General procedure for AACR
reaction
2 | EXPERIMENTAL
2.1 | General
Aryl azides 1a-d (1 mmol), alkyne 2a-d (1.2 mmol),
tetrabutylammonium bromide, TBAB, (0.1 mmol), and
All chemicals were purchased from Merck or Fluka and
Aldrich Chemical Companies. Aryl azides 1a-d were
synthesized according to the procedure mentioned in
Pokhodylo et al.[31] All yields refer to isolated products.
The products were characterized by their spectral data.
1H NMR (300 MHz) and 13C NMR (75 MHz) spectra were
recorded as CDCl3 solutions with tetramethylsilane as an
internal standard using a Bruker Avance spectrometer.
Fourier transform infrared (FT-IR) spectra were recorded
as KBr discs with a Nicolet Avatar 370 FT-IR spectrome-
ter. Elemental compositions were determined through
energy-dispersive X-ray spectroscopy (EDS) analysis
(model 7353, Oxford Instruments UK), with a resolution
of 133 eV. Inductively coupled plasma atomic emission
spectroscopy (ICP-AES) analysis was carried out with a
Varian VISTAPRO CCD (Australia). Powder X-ray dif-
fraction (PXRD) patterns were obtained using a Bruker
SCHEME 1 Schematic representation of Cu/Fibroin catalyst

MIRZAEI ET AL.
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Cu/Fibroin catalyst (0.02 mol%) were added to water
(5 ml), and the reaction mixture was stirred at room
temperature for appropriation time. After completion
of the reaction (monitored by TLC), the reaction
mixture was diluted with chloroform (20 ml) and
filtered to remove catalyst. The organic layer was
washed with water (40 ml × 2), dried and concen-
trated under reduce pressure to obtain crude residue.
The residue was purified by washing with cold metha-
nol to obtained pure products 3a-d in good to high
yields (Scheme 2).
and 13C NMR spectroscopy and mass spectrometry are
described below (Supporting information).
2.5.1 | Product 3i
N,N-dimethyl-1-(1-(3-nitrophenyl)-1H-1,2,3-triazol-4-yl)
1
methanamine, mass: m/z = 247 (M+), HNMR (CDC13,
300 MHz): δ = 2.38 (s, 6H, N-(Me)2), 3.75 (s, 2H, CH2),
7.78 (t, 1H, J = 8.1, meta-Ph), 8.10 (s,1H, triazole), 8.23
(ddd, 1H, J 1 = 8.1, J 2 = 2.1, J 3 = 0.9, ortho-Ph), 8.33
(ddd, 1H, J 1 = 8.1, J 2 = 2.1, J 3 = 0.9, para-Ph), 8.64 (t,
1H, J = 2.1, ortho-Ph). 13CNMR (CDC13, 300 MHz):
δ = 45.2 (dimethyl amino), 54.2 (CH2), 115–150, 8C (Ar).
2.5 | Representative spectral data for the
selected products
The known products (3a–h) were reported previously so
these products were identified by HNMR and compari-
2.5.2 | Product 3j
1
son of their physical data with those of authentic samples
(Supporting information). But compounds 3i and 3j are
first synthesized so their analytical and spectral data (1H
N,N-dimethyl-1-(1-(2-methyl-4-nitrophenyl)-1H-
1,2,3-triazol-4-yl)methanamine, m/z = 261 (M+), 1HNMR
(CDC13, 300 MHz): δ = 2.39 (s, 6H, N-(Me)2), 2.45 (s, 2H,
SCHEME 2 General procedure for the
synthesis of triazole compounds
FIGURE 1 Fourier-transform infrared (FT-
IR) spectra of fibroin fibers: (a) without copper
and (b) with copper NPs

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MIRZAEI ET AL.
CH3), 3.77 (s, 2H, CH2), 7.61 (d, 1H, J = 9, ortho-ph),
7.82 (s, 1H, Triazole), 8.25 (dd, 1H, J 1 = 9, J 2 = 3, meta-
ph), 8.32 (d, 1H, j = 3, meta-ph). 13CNMR (CDC13,
300 MHz): δ = 18.6 (Me), 45.2 (dimethyl amino), 54.2
(CH2), 122–147, 8C, (Ar).
vibrations of carboxylate group and 1,339 cm−1 indi-
cate the presence of methylic groups of alanine in the
silk chain. The peak at 3,423 cm−1 is a symptom of
the hydroxyl ( OH) stretching vibration from the phe-
nolic acids of Tyr molecule of SF chain. Infrared
(IR) analysis demonstrated that complexation of copper
by the fibroin fiber had occurred, with the prominent
2.6 | Recycling and reusing of the catalyst
C N stretching vibration of the imine (1,247 cm−1
)
being shifted to a lower frequency (1,230 cm−1) by
Due to the fact that the catalyst is insoluble in water, it
can be recycled by simple filtration. The separated cata-
lyst was washed with ethanol, dried at room temperature,
and reused in the model reaction for four times.
17 cm−1, indicative of metal–fibroin interaction.[34]
3.1.2 | X-ray diffraction
Powder XRD analysis was applied to identify the crystal-
line structure of Cu NPs (Cu/Fib.) into the surface of
fibroin. As shown in Figure 2, three characteristic diffrac-
tion peaks at 2θ values 43.7ꢀ, 51. 1ꢀ, and 74.6ꢀ,
3 | RESULTS AND DISCUSSION
3.1 | Characterization results of the
Cu/Fib. catalyst
3.1.1 | Fourier transform-infrared spectra
In the FT-IR spectrum of Pure Silk Fibroin
(Figure 1a), the appearance of absorption bands at
1,647 cm and 1,545 cm−1, which are related to the
amide—I (C O stretching) and amide—II (N H
bending), respectively, confirm the Silk II structural
conformation (β-sheet). Another absorption band
observed at 1,247 cm−1 corresponded to amide—III
(C N stretching), which is typically silk I conforma-
tion (α-helix or random coil structure).[34] Peaks were
observed at 1,411 cm−1 is due to symmetric stretching
FIGURE 2 X-ray diffraction (XRD) patterns of Cu/Fibroin
catalyst
FIGURE 3 Scanning electron microscopy (SEM) images of (a) pure fibroin, (b–e) Cu/Fib.; and (f) cutting section of fibroin fiber

MIRZAEI ET AL.
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FIGURE 4 Energy-dispersive X-ray spectroscopy (EDS) report of fibroin fibers: (a) before; and (b) after copper loading
corresponding to the (1 1 1), (2 0 0), (2 2 0) indices of
FCC lattice of metallic Copper nanoparticles (JCPDS card
no. 98-410-5041). Because copper nanoparticles are sensi-
tive to oxygen in the air, they can be oxidized to copper(I)
oxide (Cuprite). Thus, there are some extra peaks in the
XRD pattern attributed to Cu(I) (Cuprite) that are posi-
tioned at 2θ = 36.7, 42.3, and 73.5, corresponding to the
(200) (220) and (311) planes of copper oxide (JCPDS card
no. 96-900-7498). The average crystallite sizes of Cu
nanoparticles, calculated using the Debye–Scherrer equa-
tion (d = Kλ/βcosθ), is about 30 nm.
TABLE 1 CHN component of fibroin fibers, before and after
Copper loading
Element (% w)
Nitrogen
Pure fibroin
16.431
Cu/Fibroin
15.500
3.1.3 | Scanning electron microscopy
Carbon
45.240
42.830
The morphological features were studied by scanning
electron microscopy (SEM) technique. SEM images of
Hydrogen
5.596
5.288
TABLE 2 Synthesis of compound 3e in the presence of Cu/Fibroin as catalyst under different reaction conditiona
Entry
Catalyst (mol% Cu)
Solvent
Methanol
Temp. (ꢀC)
Time (h)
Yield (%)b
1
0.05
0.05
0.05
0.05
0.05
0.05
0.02
0.01
-
50
50
50
50
80
r.t
r.t
r.t
r.t
50
50
4
4
4
3
3
3
3
3
7
4
4
80
2
H20
60
3
H20/tButOH (1:2)
H20/TBABc (10:1)
H20/TBAB (10:1)
H20/TBAB (10:1)
H20/TBAB (10:1)
H20/TBAB (10:1)
H20/TBAB (10:1)
n-hexane
81
4
83
5
71
6
90
7
90
8
80
9
-
10
11
0.05
0.05
Trace
Trace
Toluene
aOther conditions: 3-nitroazide (1 mmol), phenylacetylene (1. 5 mmol), H2O/TBAB (10:1) as a solvent at r.t.
bIsolated yield of the pure product based on 3-nitroazide.
cTetra-n-butylammonium bromide.

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MIRZAEI ET AL.
TABLE 3 Synthesis of 1,2,3-triazoles 3a-j using Cu/Fibroin as catalysta
Entry
Aryl azide
Alkyne
Productb
Time (h)
Yieldc (%)
1
2a
2
95
1a
3a
2
3
2b
2b
3
90
93
1b
3b
2:40
1c
3c
4
2c
2
95
1d
3d
5
6
2a
2a
2
3
90
92
1b
3e
1c
3f
7
8
2a
2
3
97
93
1d
3g
2b
1a
1b
3h
9
2:20
2
98
94
2d
3i
3j
10
2d
1d
aReaction conditions: Aryl azide (1 mmol), Alkyne (1.2 mmol), 0.02 mol% Cu/Fibroin and H2O/TBAB (10:1) as a solvent at r.t. temperature.
bAll the products were characterized by 1H-NMR.
cIsolated yield.

MIRZAEI ET AL.
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Cu/Fibroin (Figure 3b–e) proved that these copper
nanoparticles are almost spherical and narrowly distrib-
uted on the smooth surface of the Silk Fibroin
(Figure 3a). This homogeneous dispersion was attributed
to an interaction between the Cu(OAc)2 and amino acids
of the Silk Fibroin, increasing the resistance to the
growth of the Cu clusters. The SEM image of cutting
section of a fibroin fiber in Figure 3f proves that the seri-
cin was completely removed,[20] and according to
Figure 3d, the average particle size is about
50 nanometers.
Comprising between the elemental analysis (EDS)
spectrum of pure fibroin and Cu/Fibroin catalyst shows
the successful immobilization of Cu nanoparticles on
fibroin fibers. As can be seen in Figure 4, no additional
peak related to other impurities was appeared in the
spectrum.
3.2 | Catalytic activity of the Cu/Fibroin
nanocatalyst in the synthesis of
1,2,3-triazoles
The reaction of 3-nitro-phenylazide (1 mmol),
phenylacetylene (1. 5 mmol) for the synthesis of com-
pound 3e was selected as a model, to optimize the reac-
tion conditions Such as different solvents, amount of
catalyst and required optimum temperature of the reac-
tion, and the results are summarized in Table 2.
Initially, we used methanol as solvent in the presence
of 0.05 mol% Cu/Fibroin catalyst at 50ꢀC, which resulted
80% yield in 4 h (Table 2, entry 1). The water was then
replaced with water as a green solvent in the same condi-
tion However, the reaction yield decreased as the solubil-
ity of the reactants in the water solvent decreased
(Table 2, entry 2). Then t-Butanole was added to increase
the solubility of the reactants in water (1:2) which
increased the reaction yield to 81%.
3.1.4 | Determining of Cu content of
Cu/Fibroin catalyst
Tetra-n-butylammonium bromide was also evaluated
as a phase transfer catalyst in the water (10:1) and was
able to produce product 3e with high yield (Table 2, entry
4). The highest yield was obtained when the reaction was
done in mixture of H2O and TBAB with the ratio 10:1 at
room temperature in the presence of 0.02 mol%
Cu/Fibroin (entry 7). Normal hexane and toluene as non-
polar solvent were also evaluated, which very small
amount of product resulted in 4 h (Table 2, entries
10 and 11). Having these optimized reaction conditions
in hand, the scope of reaction was extended to various
azides and different terminal and internal alkynes.[8]
According to the results shown in Table 3, all phenyl
azides carrying either electron-donating or electron-
withdrawing groups reacted successfully whit terminal
The Cu content of Cu/Fibroin catalyst was determined
using the inductively coupled plasma-atomic emission
spectrometry (ICP-AES) technique. This resulted in a
value of about 1.45% w/w.
Due to confirmation of the copper content resulting
from the ICP-AES method, the CHN content was
investigated before and after copper loading. The
results are shown in Table 1, which shows that the
CHN content (% w) decreased from 67.26 to 63.61% w
after Copper loading, which is approximately in accor-
dance with the copper content resulting from the ICP-
AES method.
FIGURE 5 Investigation of reusability of
the catalyst in the model reaction condition

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MIRZAEI ET AL.
FIGURE 6 Characterization of the Cu/Fibroin catalyst after the third recovery and reuse process. (a) Fourier-transform infrared (FT-
IR) spectra; (b) X-ray diffraction (XRD) pattern
and symmetric internal alkynes and gave the products in
high yields within short reaction time. In the case of ter-
minal alkynes, the reaction showed good regioselectivity
and produced a high yield of 1,4-disubstituted products.
To determine the applicability of catalyst recovery,
the same model reaction was again studied under the
optimized conditions. The catalyst was recovered
according to the procedure mentioned in Section 2 and
reused in a similar reaction, for three more times. The
catalyst showed high stability after it was reused four
times with only a slight reduction of its activity. Results
have been shown in Figure 5. FT-IR and powder XRD,
were carried out to characterize the catalyst after the
third recovery and reuse process, which does not show
any considerable change in the morphology. However,
an increase in the intensity of diffraction peaks related to
Cu2O (cuprite) was observed in the recovered catalyst
(Figure 6) compared to the fresh catalyst (Figure 2). This
is due to the oxidation of copper nanoparticles by air dur-
ing catalyst recovery.
In order to compare the efficiency of the synthe-
sized catalyst and other previous catalytic systems used
in the CuACC reactions, studies were performed, and
the results are given in Table 4. As can be seen in
Table 4, these comparative results establish the advan-
tage of the Cu/Fib. catalyst over the previously
reported methods (based on catalyst loading and reac-
tion times and yield).
TABLE 4 Comparative results between our study and other previously reported catalytic systems in the synthesis of some triazole
compounds
Entry
Product
Catalyst/condition
Yield (%)
Ref.
1
Cu/Fibroin (0.02 mol%), H2O/TBAB, rt, 2 h
95
89
This work
Velasco et al.[37]
Dialkyldithiocarbamic acid copper complexes,
(0.005 mmol), acetonitrile, rt, 11 h
(3a)
Cu (PPh3)2NO3 (0.5 mol%), rt, solvent free, 24 h
Cu/Fibroin (0.02 mol%), H2O/TBAB, rt, 2 h
Cu (PPh3)2NO3 (0.5 mol%), rt, solvent free, 5 h
45
90
81
92
Wang et al.[38]
This work
Wang et al.[38]
Murthy et al.[39]
2
(3e)
Aryl halide, NaN3, CuI/D-lucosamine KOH,
1:1(DMF + H2O), 80ꢀC–90ꢀC, ethanol,
phenylacetylene, Sod.ascorbate, rt, 16 h
C32H32Cu2N10(2+) * 2F6P(1−) (0.3 mol%), sodium
99
Han et al.[40]
L-ascorbate (3 mol%), methanol, rt, N2, 16 h
3
Cu/Fibroin (0.02 mol%), H2O/TBAB, rt, 3 h
Catalyst free, chloroform, 30ꢀC–70ꢀC, 15 days
93
44
This work
Kauer et al.[41]
(3h)

MIRZAEI ET AL.
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SCHEME 3 Proposed CuAAC reaction mechanism for (a) terminal and (b) internal alkynes
3.3 | Proposed reaction mechanism
ACKNOWLEDGMENT
The authors are gratefully acknowledged for financial
support of this study (grant no: 3/44652) by Research
Council of Ferdowsi University of Mashhad.
Under our conditions, the presence of CuNPs on the sur-
face of fibroin fibers probably acts as the catalyst. A tenta-
tive reaction mechanism for the synthesis of 1H-
1,2,3-triazoles catalyzed by CuNPs under heterogeneous
conditions is proposed in Scheme 3. It is proposed that,
in the first step, a terminal alkyne is coordinated to
CuNPs supported on fibroin, which activates the C H
bond. As a result, the corresponding copper–alkylidine
complex is formed on the surface of the fibroin fibers.
This is a very favorable step because Cu metal is well
known to exhibit high alkynophilicity for terminal
alkynes. In the second step, synthesized aryl azides attack
the copper-alkylidine complex followed by intramolecu-
lar cyclization, finally leading to formation of a five-
membered ring of triazole as a product. In the case of
internal alkynes, π-coordination between alkyne and
copper(I) spices leading to activation of alkyne and can
acts as driving force for this reaction.[35,36]
AUTHOR CONTRIBUTIONS
Hakimeh Sadat Mirzaei Hesari: Conceptualization;
data curation; formal analysis; investigation; methodol-
ogy; project administration; validation; visualization.
Hossein Eshghi: Conceptualization; data curation; for-
mal analysis; funding acquisition; investigation; method-
ology; project administration; resources; supervision;
validation; visualization. Seyed Mohammad Seyedi:
Conceptualization; methodology; project administration;
supervision.
DATA AVAILABILITY STATEMENT
The data that support the findings of this study are avail-
able from the corresponding author upon reasonable
request.
ORCID
Hakimeh Mirzaei https://orcid.org/0000-0002-4556-
0832
4 | CONCLUSION
Hossein Eshghi https://orcid.org/0000-0002-8417-220X
Seyed Mohammad Seyedi https://orcid.org/0000-0001-
5542-5880
In conclusion, novel copper nanoparticles immobilized
on silk fibroin (Cu/Fibroin) were successfully prepared
and fully characterized using FT-IR, SEM, EDS, XRD,
ICP, and CHN techniques. We have developed a simple
new catalytic method for the synthesis of 1,2,3-triazoles
from aryl azides and terminal and internal alkynes in the
presence of Cu/Fibroin NPs as ligand-free, efficient, reus-
able, and heterogeneous catalysts. Some attractive fea-
tures of this protocol are high yields, simple procedure to
produce catalyst, easy work-up, high catalytic activity,
good regioselectivity with terminal alkynes, reactivity
toward internal alkynes and reusability of the catalyst.
The catalyst can be used at least four times.
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How to cite this article: Mirzaei H, Eshghi H,
Seyedi SM. Cu Nano particles immobilized on silk-
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copper catalyzed azide-terminal, internal alkynes
cycloaddition. Appl Organomet Chem. 2020;e6019.
https://doi.org/10.1002/aoc.6019
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