Aerobic synthesis of biocompatible copper nanoparticles: Promising antibacterial agent and catalyst for nitroaromatic reduction and C-N cross coupling reaction

Article abstract of DOI:10.1039/c4ra01126k

Herein, we report the synthesis of copper nanoparticles at ambient conditions using biopolymer, pectin, as a protecting agent and hydrazine as a reducing agent. The obtained nanoparticles catalyze the reduction of nitroaromatic compounds in aqueous solution and also catalyze the C-N cross coupling of amines with bromobenzene in good yields. This journal is the Partner Organisations 2014.

Full text of DOI:10.1039/c4ra01126k

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Aerobic synthesis of biocompatible copper
nanoparticles: promising antibacterial agent and
catalyst for nitroaromatic reduction and C–N cross
coupling reaction†
Cite this: RSC Adv., 2014, 4, 15003
Received 8th February 2014
Accepted 11th March 2014
Showmya Venkatakrishnan,a Ganapathy Veerappan,b Elangovan Elamparuthia
DOI: 10.1039/c4ra01126k
a
*
and Anbazhagan Veerappan
www.rsc.org/advances
Herein, we report the synthesis of copper nanoparticles at ambient robust, cost-effective and eco-friendly approach is still required
conditions using biopolymer, pectin, as a protecting agent and to synthesize stable CuNPs.
hydrazine as a reducing agent. The obtained nanoparticles catalyze
Of late, green chemistry methods utilizing the sources such
the reduction of nitroaromatic compounds in aqueous solution and as microbes, plant extract and biopolymer have received
also catalyze the C–N cross coupling of amines with bromobenzene in signicant attention in the synthesis of inorganic nano-
good yields.
particles.5 However, most of the green chemistry approaches of
nanoparticles synthesis have stopped at the level of synthesis
and basic characterization. In order to effectively utilize the
nanoparticles synthesized by green chemistry method, it is
imperative to develop its application.
Metal nanoparticles nd application in catalysis, bio-
diagnostics, drug delivery, nanomedicine, sensors, photonics
and electronics.1 Among many metal nanoparticles, group IB
metals such as silver, gold and copper have received consider-
able attention due to their attractive optical properties, arising
from localized surface plasmon resonance in the visible and the
near infrared region of the electromagnetic spectrum. In this
list copper was less popular, mainly because the fabrication of
chemically stable copper nanoparticles (CuNPs) was far more
complicated, because they are prone to fast oxidation.2 In-spite
of the difficulty, CuNPs have attracted considerable attention
because they are much more economical than the silver and
gold nanoparticles. Therefore, several methods have been
developed for the preparation of copper nanoparticles,
including chemical reduction, thermal reduction, pulsed
sonoelectrochemical reduction, radiation methods, metal
vapour synthesis, vacuum vapour deposition, microemulsion
techniques and laser ablation.3 Most of these methods are
either required sophisticated instrument or required biological
hazardous chemicals and also required oxygen free environ-
ment to synthesize CuNPs. There are very few reports exist on
CuNPs synthesis without inert atmosphere.4 Hence, a more
In this paper, we describe here the rst preparation of pectin
stabilized CuNPs at ambient conditions and report on their
excellent catalytic activity for the reduction of various nitro-
aromatic compounds. We have also demonstrated for the rst
time that pectin protected CuNPs has an ability to catalyze C–N
cross coupling of amines with bromobenzene. We have used
pectin, a polysaccharide isolated from citrus peel, consisting of
a linear backbone of (1–4) linked a-D-galacturonic acid residue
with varying degree of methylesteried carboxyl groups, as a
templating agent.6 It is widely used as gelling, thickening and
stabilizing agents in the food industry. The choice of pectin was
made because of its benign nature and the presence of oxygen
rich functional group and acid group allows CuNPs to be
very stable.
The preparation of CuNPs using biopolymer, pectin as a
capping agent and hydrazine as a reducing agent was illustrated
in Fig. 1A. In a typical experiment, 50 mM of CuCl2 was mixed
with 50 mL aqueous suspension of pectin (1 mg mLꢀ1) through
vigorous stirring. The greenish blue gel acidic solution brought
to basic by adding 200 mL of concentrated ammonia solution.
The colour changes from greenish blue to blue due to the
formation of copper–ammonia complex. Aer 5 minutes of
gentle mixing, 200 mL of hydrazine hydrate was added as a mild
reducing agent. Aer stirring the solution for 5 minutes, the
container was le undisturbed in open air for 3 hours. The
formation of copper nanoparticles was easily noticeable due to
the change in the color of the solution to reddish brown – the
characteristic color of the copper nanoparticles (Fig. 1A).
aDepartment of Chemistry, School of Chemical and Biotechnology, SASTRA University,
Thanjavur – 613 401, Tamil Nadu, India. E-mail: anbazhagan@scbt.sastra.edu.in;
Fax: +91-4362-26120; Tel: +91-4362-264101-3689
bSKKU Advanced Institute of Nano Technology, Sungkyunkwan University, Suwon,
South Korea
† Electronic supplementary information (ESI) available: Description of
experimental procedure. UV-Vis spectra, kinetics data, 1H-NMR spectra and
antimicrobial activity plates. See DOI: 10.1039/c4ra01126k
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XRD displays high crystallinity. The thin lm of CuNPs showed
diffraction peak at 43.70ꢁ, 50.78ꢁ and 74.45ꢁ, corresponding to
the crystal facets of (1 1 1), (2 0 0) and (2 2 0), respectively
(Fig. 1B) and compared with the standard diffraction pattern of
metallic copper (JCPDS card no. 4-0836). The pattern was very
clean, with no indication of impurities such as copper oxides
(CuO, Cu2O). The TEM images, as shown in Fig. 1C, suggested
that the CuNPs are spherical in nature with size is about 5 nm.
The catalytic activity of the synthesized CuNPs was measured
by sodium borohydride (NaBH4) mediated reduction of p-
nitrophenol (p-NP) to p-aminophenol (p-AP). The chosen reac-
tion was easy to follow by UV-Vis absorption spectroscopy,
therefore, this reaction have been used widely to evaluate the
catalytic activity of various metallic NPs including gold, silver,
nickel, palladium and platinum.7 Addition of NaBH4 to the
aqueous solution of p-NP was accompanied by a colour change
from light yellow to yellowish-green (Fig. 3A). The absorption
maximum of p-NP shis from 317 nm to 400 nm (Fig. 3B),
suggested the formation of p-nitrophenolate anion. The addi-
tion of CuNPs was accompanied by color change from
yellowish-green to colorless suggested the reduction of p-NP to
Fig. 1 (A) Schematic preparation of pectin-stabilized copper nano-
particles from 50 mM CuCl2, (B) X-ray diffraction pattern of pectin
stabilized CuNPs (C) TEM image of the synthesized CuNPs. The inset is
the high resolution TEM micrograph.
The UV-visible spectra of the CuNPs solution exhibit a
characteristic surface plasmon resonance peak at 580 nm
(Fig. 2A). Time dependent absorption studies of CuNPs showed
that the NPs formation was accompanied by three stages
(Fig. 2B). Stage I, initiation process, which occurs between 0 to
65 minutes. The conversions of copper complex to CuNPs (Stage
II) occur between 65 minutes to 120 minutes. Aer 120 minutes,
the absorption intensity does not change, indicating that the
NPs formation was completed (Stage III). Analyzing the time
dependent formation of NPs yielded the half time (95.06
minutes), the time required to convert half of the copper
complex to CuNPs. The dispersion of the CuNPs in water was
very clear and stable for more than 30 days without settling
down. The initial reddish brown color of CuNPs remains stable,
even exposing the CuNPs to open air for 10 days. The stability of
CuNPs was also monitored by measuring their intensity at 580
nm over a period of 30 days and no signicant change in the
intensity was observed. The synthesized CuNPs examined by
Fig. 3 (A) Catalytic reduction of p-NP to p-AP. (B) UV visible spectra of
p-nitrophenol (a), p-nitrophenolate ion (b) and its reduced product, p-
Fig. 2 Formation of pectin stabilized CuNPs (A) time dependent UV aminophenol (c). (C) Plot of ln(Ct/C0) versus reaction time. Ct/C0 is
studies of CuNPs formation, (B) time dependent changes in the calculated based on the absorbance of p-NP at 400 nm. The
absorption maximum at 580 nm.
concentration of p-NP in presence of NaBH4 are shown in blue dots.
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p-AP (Fig. 3A). The absorption maximum at 400 nm in presence formation encouraged us to extend the application of CuNPs to
of CuNPs decreases with time and a new peak due to the synthesis various C–N cross coupling product with aryl amines,
formation p-AP appears at 298 nm (Fig. 3B). As shown in Fig. 3C, alkyl amine and N-heterocyclic amines. 3-Chloro aniline, tyra-
the concentration of p-NP at 400 nm remained unaltered in mine, dopamine, glycine, pyrrole and indole underwent reac-
presence of NaBH4, indicating that the NaBH4 does not reduce tion with bromobenzene to afford desired product in moderate
p-NP in the absence of CuNPs.
to good yield. The obtained products was puried by ash
To determine the reaction rate, we monitored time depen- chromatography and characterized by thin layer chromatog-
dent changes in the absorption peak of p-NP at 400 nm in the raphy and 1H-NMR (details in ESI†). The results of these
presence of CuNPs. Before the catalytic reduction of p-NP, an experiments are summarized in Table 2. The reusability of the
induction time up to 120 s was observed (Fig. 3C). This induc- catalyst was examined by reacting aniline with bromobenzene.
tion time is typical for a heterogeneous catalytic process and Aer completion of the reaction, the catalyst was recovered by
commonly related to activation or restructuring of the metal centrifugation at 14 000 rpm and reused for the fresh reaction
surface by nitrophenol before the reaction could start.8 Aer an of aniline with bromobenzene and observed no loss of activity
induction time t0, the reaction starts and completed within even aer 3 cycles.
250 s. In this catalytic reaction, the concentration of NaBH4 was
The gradual increases in the identication of new multi-drug
much higher than that of p-NP, thus, the reduction kinetics can resistant bacteria strain have forced the research community to
be described as pseudo rst-order with respect to p-NP alone. discover new antibacterial agent. Copper and its complex have
Fig. 3C show the plots of ln(Ct/C0) versus reaction time for the been well known for the bactericidal effect.10 In order to exploit
reduction of p-NP, where Ct and C0 are the concentration of the pectin stabilized CuNPs against pathogenic bacteria we
p-NP at time t and 0, respectively. The linear correlation between tested the bactericidal effect of CuNPs by disk diffusion method
ln(Ct/C0) and the reaction time indicate that the reduction against Gram positive and Gram negative bacteria (Fig. S12 and
reaction follows pseudo rst order kinetics. The rate constant, k S13†). As shown in Table 3, the diameter of inhibition zones
(1.08 ꢂ 10ꢀ2 sꢀ1) was obtained directly from the slope of the around the disk containing CuNPs varies for different bacterial
linear part of the kinetic trace.
species. This difference in zone of inhibition among the bacterial
The scope of the above described procedure has been species can be attributed to different cell wall composition. It is
examined with various nitroaromatic compounds (Fig. S1–S4†) clear from Table 3 that the zone of inhibition of the CuNPs was
and the results are summarized in Table 1. As noted from comparable to the standard antibiotic, ooxacin and kanamycin.
Table 1, the nitro group present at para position undergoes The zone of inhibition in the present study was comparatively
faster reduction with lesser induction time. When the nitro
group is in ortho or meta position, the rate of reduction
decreases considerably with higher induction time. This
suggest that the activation or restructuring of the metal surface
by the ortho or meta substituted nitro compound is delayed R ꢀ NH2 & R ꢀNH
because of the steric hindrance and yielded lesser rate (Table 1).
However, irrespective of the substituted position, all the nitro
compounds are converted efficiently to their corresponding
amino compounds in less than 5 minutes.
Table 2 C–N cross coupling of amines with bromobenzene
Cu NPs; PhꢀBr
-
R ꢀ HN ꢀ Ph & R ꢀN ꢀ Ph
ƒƒƒƒƒƒƒƒƒ!
2
2
DMSO; 110 ꢁ C; 2 h
Entry
Substrate
Product
Yield
1
1a, 85%
To further expand the scope of pectin stabilized CuNPs, we
next explored the C–N cross coupling of amines with aryl halide.
The formation of C–N bonds by cross coupling reaction repre-
sents a powerful method to prepare biological and pharma-
ceutical compounds.9 The reaction of aniline with
bromobenzene was rst studied as standard substrate with
CuNPs act as a catalyst. The reaction occurred to produce
diphenylamine in 85% yield when it was stirred at 110 ꢁC in the
presence of 2 mol% of CuNPs and 1 mol% of NaOH in DMSO
under aerobic conditions. The success of diphenylamine
2
3
4
2a, 82%
3a, 75%
4a, 69%
Table
1 Induction time (t0) and rate constants (k) of catalyzed
5
6
5a, 71%
6a, 70%
reductions of chosen nitroaromatic compounds
Substrate
t0 (s)
(k) (sꢀ1
)
p-Nitrophenol
120.7
79.8
160.2
169.8
139.8
1.08 ꢂ 10ꢀ2
1.32 ꢂ 10ꢀ2
0.21 ꢂ 10ꢀ2
0.68 ꢂ 10ꢀ2
0.75 ꢂ 10ꢀ2
p-Nitrobenzamine
m-Nitrobenzamine
o-Nitrobenzamine
p-Methyl-o-nitrobenzamine
7
7a, 75%
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Table 3 Antibacterial activity estimated by zone of inhibition
2008, 37, 1896; (c) B. C. Ranu, K. Chattopadhyay, L. Adak,
A. Saha, S. Bhadra, R. Dey and D. Saha, Pure Appl. Chem.,
2009, 81, 2337.
Zone of inhibition (mm)
2 K. K. R. Datta, C. Kulkarni and M. Eswaramoorthy, Chem.
Commun., 2010, 46, 616.
Bacteria
CuNPs
Ooxacin
Kanamycin
Bacillus thuringiensis
Staphylococcus aureus
Klebsiella pneumonia
Escherichia coli
Salmonella typhi
Pseudomonas aeruginosa
Shigella exneri
16
19
17
17
12
16
16
26
16
19
19
15
16
14
15
20
14
16
18
11
12
17
11
20
3 (a) D. Kim, S. Jeong and J. Moon, Nanotechnology, 2006, 17,
4019; (b) J. Hambrock, R. Becker, A. Birkner, J. Weiß and
R. A. Fischer, Chem. Commun., 2002, 68; (c) I. Haas,
S. Shanmugam and A. Gedanken, J. Phys. Chem. B, 2006,
110, 16947; (d) A. A. Ponce and K. J. Klabunde, J. Mol.
Catal. A: Chem., 2005, 225, 1.
4 (a) A. K. Chatterjee, R. K. Sarkar, A. P. Chattopadhyay,
P. Aich, R. Chakraborty and T. Basu, Nanotechnology, 2012,
23, 85103; (b) M. Vaseem, K. M. Lee, D. Y. Kim and
Y.-B. Hahn, Mater. Chem. Phys., 2011, 125, 334; (c) Y. J. Lee,
K. J. Lee, N. E. Stott and D. Kim, Nanotechnology, 2008, 19,
415604.
5 (a) L. Sintubin, W. Verstraete and N. Boon, Biotechnol.
Bioeng., 2012, 109, 2422; (b) S. Iravani, Green Chem., 2011,
13, 2638.
6 (a) H. V. Scheller, J. K. Jensen, S. O. Sørensen, J. Harholt and
N. Geshi, Plant Physiol., 2007, 129, 283; (b) B. L. Ridley,
M. A. O'Neill and D. Mohnen, Phytochemistry, 2001, 57,
929.
Proteus mirabilis
higher than the chitosan stabilized CuNPs and alginate stabi-
lized CuNPs.11 While considering the mucoadhesive nature of
pectin, the reported antibacterial activity of the newly synthe-
sized pectin stabilized CuNPs is promising and required detail
investigation for future application.
Conclusions
In conclusion, we presented a facile route to synthesize copper
nanoparticles at room temperature. The materials used in this
synthesis are biocompatible and the obtained product is air
stable. The synthesized CuNPs catalyze the reduction of nitro-
aromatic compounds in aqueous solution and also catalyze the
C–N cross coupling of amines with bromobenzene in good
yields. They also display promising antibacterial activity, as
good as ooxacin and kanamycin, towards the tested bacteria.
Further investigation on the catalytic property and antibacterial
activity will enable the use of this nanomaterial in the eld of
nanomedicine and catalysis.
7 (a) B. Liu, D. W. Zhang, J. C. Wang, C. Chen and X. L. Yang,
J. Phys. Chem. C, 2013, 117, 6363; (b) J. M. Zhang, D. H. Han,
H. J. Zhang, Y. Zhao and D. L. Ma, Chem. Commun., 2012, 48,
11510; (c) Z. M. Zhu, X. H. Guo, S. Wu, R. Zhang, J. Wang and
L. Li, Ind. Eng. Chem. Res., 2011, 50, 13848; (d) X. Zhang and
Z. H. Su, Adv. Mater., 2012, 24, 4574.
´
´
´
8 P. Herves, M. Perez-Lorenzo, L. M. Liz-Marzan, J. Dzubiella,
Y. Lu and M. Ballauff, Chem. Soc. Rev., 2012, 41, 5577.
9 (a) M. Negwar, in Organic-Chemical Drugs and their Synonyms:
(An International Survey), Akademie, Berlin, Germany, 1994,
7th edn; (b) J. B. Buckingham, in Dictionary of Natural
Products, Chapman and Hall, London, 1994, vol. 1.
Acknowledgements
The Department of Science and Technology, Government of
India is acknowledged for nancial support (SB/FT/LS-217/
2012). We thank Central Research Facility, SASTRA University
for providing the necessary infrastructure.
10 (a) B. Jia, Y. Mei, L. Cheng, J. Zhou and L. Zhang, ACS Appl.
Mater. Interfaces, 2012, 4, 2897; (b) F. Gao, H. Pang, S. Xu and
Q. Lu, Chem. Commun., 2009, 3571; (c) G. Borkow and
J. Gabbay, FASEB J., 2004, 18, 1728.
11 (a) M. S. Usman, M. E. El Zowalaty, K. Shameli,
N. Zainuddin, M. Salama and N. A. Ibrahim, Int. J.
Notes and references
´
Nanomed., 2013, 8, 4467; (b) J. Dıaz-Visurraga, C. Daza,
1 (a) P. K. Jain, X. Huang, I. H. El-Sayed and M. A. El-Sayed, Acc.
Chem. Res., 2008, 411, 1578; (b) R. A. Sperling, P. Rivera-gil,
F. Zhang, M. Zanella and W. J. Parak, Chem. Soc. Rev.,
´
C. Pozo, A. Becerra, C. von Plessing and A. Garcıa, Int. J.
Nanomed., 2012, 7, 3597–3612.
15006 | RSC Adv., 2014, 4, 15003–15006
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