Superlattices and Microstructures 81 (2015) 248–264

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Superlattices and Microstructures

journal homepage: www.elsevier.com/locate/superlattices

Modified structural and optical characteristics

of Au-NPs–MWCNTs nanohybrids
S. Mehmood a, A. Naeem b, S. Sabahat a, R. Ciancio c, E. Carlino c, M.F. Bhopal a,
A.S. Bhatti a,⇑
a
Center for Micro and Nano Devices, Department of Physics, COMSATS Institute of Information Technology, Islamabad, Pakistan
b
Department of Biosciences, COMSATS Institute of Information Technology, Islamabad, Pakistan
c
CNR-IOM TASC, Area Science Park, Basovizza S.S. 14 km 163.5, I-34149 Trieste, Italy

a r t i c l e i n f o a b s t r a c t

Article history: Nanohybrids formed by the multiwalled carbon nanotubes

Received 10 September 2014 (MWCNTs) and metallic nanoparticles (NPs) have gained immense
Received in revised form 16 December 2014 interest recently for their potential biological applications. In this
Accepted 23 January 2015
paper, we present a comprehensive study of the nanohybrid
Available online 3 February 2015
formed with varied concentrations of Au-NPs with MWCNTs. It is
demonstrated that the concentration of Au-NPs in the nanohybrid
Keywords:
MWCNTs
is crucial in defining ultimate characteristics of the nanohybrid.
Au-NPs The MWCNTs were functionalized with varied concentrations of
Raman spectroscopy Au-NPs (40–100 nano Molar (nM)) and characterized extensively
Biosensors by the X-ray diffraction, electron microscopy, Raman spectroscopy
and UV–Vis absorption spectroscopy. The process of purification
and acid treatment led to the activation of –COOH bond at the sur-
face of MWCNTs and functionalization with Au-NPs also induced
stresses as observed in the X-ray diffraction patterns. The fusion
of Au-NPs in the MWCNTs was clearly observed in the high resolu-
tion TEM images, which affected the D and G – Raman bands of the
MWCNTs significantly as studied by the line shape analysis. The
Au-NPs–MWCNTs nanohybrids were then used to study the effect
of various concentrations of E-coli using Raman spectroscopy and
absorption spectroscopy. Due to intrinsic negative charge present
on the E-coli, the local charge densities varied at the surface of
MWCNTs as soon as it was covered with E-coli, and resulted in
the shift of both G and D bands and increased intensity ratio of
the two bands. The variation in the charge density in Au-NPs due
to its binding with MWCNTs and adsorption of E-coli was reflected

⇑ Corresponding author.
E-mail address: asbhatti@comsats.edu.pk (A.S. Bhatti).

http://dx.doi.org/10.1016/j.spmi.2015.01.020
0749-6036/Ó 2015 Elsevier Ltd. All rights reserved.
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 249

in the blue shift of the surface plasmon modes. Finally, it is con-

cluded that ratio of Au-NPs and MWCNTs is crucial in forming
nanohybrid for applications.
Ó 2015 Elsevier Ltd. All rights reserved.

1. Introduction

Carbon nanotubes (CNTs), since their discovery have attracted exceptional interest as a new class of
advanced materials due to their unique properties [1–3]. CNTs can be either single-walled CNTs
(SWCNTs) or multi-walled CNTs (MWCNTs). MWCNTs are significantly easy to produce making them
pertinent for large-scale applications. A careful functionalization of CNTs [4,5] makes them biocom-
patible for their use as biosensors [6–8]. Several reports regarding the attachment of biomolecules
[9], nanoparticles (NPs) [10] and polystyrene nanofibers [11] to CNTs have been reported. Among
these, CNTs and metal-NPs hybrids are of particular interest, as hybrids exhibit properties of both
CNTs and metal-NPs [12]. However, the ratio of constituents in composites is quite crucial for
applications, which is the objective of present study.
Au-NPs are considered as the most stable biocompatible NPs that can be functionalized for chem-
istry and materials science applications [13]. Several methodologies proposed to bind Au-NPs to the
CNTs are through covalent or non covalent linkages [14–16]. The latter method is preferred because
it allows excellent electronic and mechanical properties and also preserves the unique properties of
CNTs [17,18].
Several papers have reported results of biosensors made up of MWCNTs and NPs hybrids demon-
strating the importance of these nanohybrids or composites [19–22]. For example, a non-enzymatic
glucose sensor was fabricated based on a composite of cubic Cu-NPs and arc-synthesized multiwalled
carbon nanotubes (MWCNTs) by using a substrate-enhanced electroless deposition (SEED) to detect
glucose in a real blood serum [19]. Oxidase-based amperometric glucose biosensor composed of mul-
tiwalled carbon nanotubes (MWCNTs), Au-NPs and glucose oxidase (GOD) was developed for the
specific detection of glucose which was used as an illustration for constructing various hybrid struc-
tures [20]. Controlled assembly of protein-protected Au-NPs on PANHS-functionalized MWCNTs has
also been demonstrated, where intermediate interactions of proteins took place with both MWCNTs
and Au-NPs [21]. Imprinted electrochemical sensor has also been developed based on chitosan-
Ag-NPs (CS-SNP)/grapheme – multiwalled carbon nanotubes (GR-MWCNTs) composites decorated
Au electrodes, which determined the neomycin in milk and honey [22].
Raman spectroscopy has been gaining recognition as a diagnostic tool for the identification of dif-
ferent cells and tissues, such as breast cancer [23], individual neoplastic and normal hematopoietic
cells [24], bladder cancer [25], skin cancer [26], and lung cancer [27]. Raman imaging of SWCNTs in
live cell as reported by Heller et al. [28] suggested that it was resistant to quenching and photobleach-
ing making them appropriate candidates for long time tracking and labeling. Chou et al. also suggested
the downshift of G band of SWCNTs was due to electron donating species of DNA molecules [29].
Wang et.al used the noble metal coated SWCNTs for enhanced Raman scattering and suggested that
water soluble noble NPs decoration can be used for labeling and fast Raman spectroscopic imaging
of biological samples for cancer detection [30]. Thus, the objective of the present study was to explore
the effect of formation of nanohybrid of MWCNTs and Au-NPs on its structural and biosensing
properties.
In the present study, different concentrations of Au-NPs were functionalized on the surface of com-
mercially available MWCNTs. Acid treatment (H2SO4/HNO3) was employed to create –COOH function-
al group at the surface of MWCNTs. MWCNTs and Au-NPs hybrids were then formed with various
concentrations of Au-NPs. The oxidized and Au-NPs functionalized MWCNTs were structurally char-
acterized using XRD, scanning electron microscopy (SEM), high resolution transmission electron
microscopy (HRTEM) and Raman spectroscopy. The hybrid was then characterized with different
250 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

concentration of E-coli using Raman spectroscopy and UV–Vis absorption spectroscopy. A thorough
line shape analysis of the Raman modes of CNTs was done to determine the effect of Au-NPs attach-
ment and adsorption of E-coli.

2. Experiment

XRD, SEM and HRTEM were performed to study the morphology, oxidation and nanostructural
arrangement of MWCNTS as well as related Au-NPs attachment to MWCNTs. Room temperature
Raman spectroscopy was done after each step of the synthesis of nanohybrids using LabRamIII by
Dongwoo Optron with an Ar+ ion laser exciting with 514 nm. All spectra were baseline corrected
for the detector response.

3. Sample preparations

3.1. Oxidation of pristine MWCNTs

MWCNTs were purchased from Beijing DK Nanotechnology, China. The average diameter of CNTs
was 45 ± 15 nm and length in the range of 2–10 lm. Pristine MWCNTs were added to 40 ml acid mix-
ture of H2SO4/HNO3 (with 3:1 volumetric ratios) and the mixture was ultrasonicated for 6 h to oxidize
MWCNTs [31]. The functionalized (–COOH) MWCNTs were then centrifuged for 60 min before wash-
ing for several minutes with de-ionized (DI) water to remove the acid contents. The MWCNTs were
then dried at 100 °C for 14 h.

3.2. Functionalization with Au-NPs

The oxidized MWCNTs divided into four equal parts of 20 mg each were dispersed in a 2 ml of DI
water to form a suspension solution. Au-NPs dissolved in a 2 ml volume of DI water with concentra-
tions of 40 nM, 60 nM, 80 nM and 100 nM were added to each part of the oxidized MWCNTs. Ultra-
sonication was performed for 4 h at room temperature to form nanohybrids. All of the Au-NPs
functionalized MWCNTs were then annealed at 150 °C for 4 h to strengthen the attachment of Au-
NPs with the MWCNTs.

3.3. Nanohybrids biosensing with different E-coli concentration

All samples of Au-NPs MWCNTs nanohybrids were dispersed in 2.5 ml of DI water by ultrasonica-
tion. Later, 5 sets of each sample of 0.5 ml dilution were separated for each concentration of nanohy-
brids (5.0 ± 0.2 mg) to form a total of 20 sets of dilutions. Each set of samples was then mixed with 102,
103, 104, 105/ml of E-coli concentration.

4. Results and discussion

4.1. Electron microscopy

Fig. 1(a) shows the representative SEM micrographs of the acid treated MWCNTs and (b) Au-NPs
decorated MWCNTs (80 nM) and (c) 100 nM concentrations. The average diameter of pure MWCNTs
as determined from the micrographs was 40 ± 5 nm with an average length of 3–5 microns. The aver-
age diameter of Au-NPs was 20 ± 5 nm. It is worth mentioning that the pristine MWCNTs were mostly
open ended (not clear in SEM but clearly observed in the HRTEM), however, after acid treatment most
of tubes became closed ended which is discussed in detail in the transmission electron microscopy
section. The acid treatment on the pristine MWCNTs created –COOH sites not only at the ends of
the MWCNTs but also on the walls. The SEM images also revealed that the large number of Au-NPs
was attached to the ends of the MWCNTs compared to the side walls. Highly curved hemispherical
fullerene like structure at the end caps of the acid treated MWCNTs caused this agglomeration of
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 251

Fig. 1. Representative SEM images of (a) pristine MWCNTs, (b) 80 nM Au-NPs–MWWCNTs, and (c) 100 nM Au-NPs–MWCNTs
nanohybrids. The images also show Au-NPs, and regions of their agglomeration.

Au-NPs (Fig. 1(b) and (c)). The curvature of the CNTs caused a change in the hybridization due to spa-
tial overlap of p-orbitals. Local strain and the radical character in the p-bonding electron at the end of
nanotubes was also believed to be imposed with the curvature of the MWCNTs [32]. The effect of
stresses thus produced was also observed in the XRD and Raman spectroscopy results as discussed
later. The SEM images were also used to estimate the densities of Au-NPs attached to MWCNTs,
e.g., functionalized Au-NPs densities for 80 nM and 100 nM nanohybrids were (6.0 ± 0.5)  1012 m2
and (7.6 ± 0.5)  1012 m2, respectively. This confirmed an increase in the density and surface cover-
age with the increase in the Au-NPs concentration in the hybrid composites.
The SEM images also confirmed the immobilization of higher concentration of Au-NPs at ends and a
relatively small amount of Au-NPs on the side walls of MWCNTs. Advanced nanostructural charac-
terization was performed by HRTEM to determine the extent of defects created in the MWCNTs as well
as modifications in the size of Au-NPs and relative attachment to the MWCNTs.

4.2. High resolution transmission electron microscopy

Fig. 2(a) shows a representative TEM micrograph of pristine MWCNTs. Nanotubes are arranged in
bundles and appear to be mostly open ended and characterized by the presence of defects. In the
HRTEM of Fig. 2(b), obtained by focusing the beam at the dash-edged region shown in the inset, the
nature of the defects can be appreciated: nanotubes were bent and characterized by several internal
closed edges intercalating within their length. HRTEM also showed the multiwalled nature of nan-
otubes, characterized by ordered arrays of layers clearly visible in the HRTEM of Fig. 2(c) and in the
corresponding intensity profile (shown in Fig. 2(d)) taken in the selected region of the image, which
showed that the average number of layers of MWCNTs was 45 ± 3 layers with an interlayer spacing
of about 0.35 ± 0.05 nm. The dark contrast spots appearing on the background of the images are
remains of contamination attached during the growth/oxidation process.
The situation changed considerably once CNTs were acid treated as can be seen in a representative
HRTEM micrograph shown in Fig. 3(a). HRTEM image shows that in this case nanotubes were mostly
close ended, as emphasized at higher magnification in Fig. 3(b). Moreover, HRTEM on the individual
nanotubes highlighted an overall reduced number of layers within the walls which were in general
of smaller width compared to the pristine MWCNTs. More in detail, number of layers ranging between
22 and 35 was observed for the acid treated MWCNTs. In the HRTEM of Fig. 3(c) and in the correspond-
ing intensity profile of Fig. 3(d), the case of a multiwalled nanotube with 30 layers is provided. The
number of layers reduction was due to corrosion introduced by the acid treatment in the MWCNTs.
Russian doll type tubes were also observed as the acid treatment had led to the breaking up of the
tubes. This confirmed the modification of the tube was along the surface (i.e., the edge planes acces-
sible at the cylindrical layers of the tube) [33,34].
Fig. 4(a) shows a HRTEM micrograph of the MWCNTs functionalized with Au-NPs. Au-NPs appear as
randomly dispersed in the bundled nanotubes and mostly attached to the outer surface of the nanotube
252 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

Fig. 2. (a) TEM images of bundled MWCNTs, mostly open ended. (b) HRTEM image showing the defects of an individual open
ended nanotube (displayed in the inset). (c) HRTEM image and (d) intensity profile of the dashed edged region showing the case
of a 45 layers-nanotube’s wall.

Fig. 3. (a) TEM image of oxidized MWCNTs showing bundled MWCNTs were closed ended. (b) HRTEM image showing in higher
detail the closed termination of the oxidized nanotubes; (c) HRTEM image and (d) intensity profile of the enlightened region
showing the drop in number of layers (from 45 to 30).

structure. HRTEM images of Au-NPs sitting on the outer walls of the nanotubes are shown in Fig. 4(b) and
(c), giving evidence that Au-NPs had diffused or penetrated into the top layers of the MWCNTs’ surface.
This is a relevant aspect which is expected to play an important role in the modification of Raman modes
of MWCNTs and absorption due to plasmon modes of the Au-NPs. HRTEM investigations showed a
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 253

Fig. 4. (a) TEM image of Au-NPs and MWCNTs composites showing the Au-NPs were randomly functionalized on the surface of
MWCNTs. HRTEM images (b) and (c) showing that Au-NPs are mostly sitting outside the nanotubes’ walls. (d) Intensity profile
showing the drop in number of layers (from 30 to 22).

Fig. 5. (a) HRTEM showing an icosahedrically shaped Au particle attached onto a CNT wall, (b) diffractogram taken over the
upper facet of the nanoparticle (dashed area in (a)). The indexed spots are compatible with those of cubic Au in the [1 1 1] zone
axis. The spots of the [1 1 1] zone axis are highlighted by the dashed lines. The other visible spots are due to the contribution of
other domains of the icosahedron.

decrease in the average number of layers to 22 keeping an interlayer spacing of 0.35 nm (see HRTEM
image of Fig. 4(c) and the related intensity profile displayed in Fig. 4(d). HRTEM inspection of the
Au-NPs showed that they had mainly an icosahedral shape. The HRTEM image of the Fig. 5(a) and the
related diffractogram (as shown in Fig. 5(b)) taken in the highlighted selected area displayed the case
of an Au-NP with the upper facet having a crystal structure compatible with the cubic Au in the [1 1 1]
zone axis as marked by dashed lines. This was also in agreement with the X-ray diffraction patterns.
According to HRTEM and electron diffraction results, in the case of an interface formed by Au-NP –
MWCNTs, a plausible epitaxial relationship could be established with either the (02–2) Au planes of
the Au-NP which correspond to a d – spacing of 0.14 nm – or with the Au (22–4) planes - which
254 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

correspond to a d-spacing of 0.08 nm. The (02–2) Au planes correspond to an in-plane Au–Au rectangular
coordination with 0.288 nm and 0.407 nm lattice parameters along the [0–1–1] and [1 0 0]
crystallographic directions, respectively. Analogously, the (22–4) Au planes correspond to an in-plane
Au–Au rectangular coordination with 0.288 nm and 0.705 nm lattice parameters along the [1 1 0]
and [111] crystallographic directions, respectively. A rectangle-on-rectangle epitaxial relationship
between the CNT structure and the two mentioned Au planes can be established by identifying a rect-
angular sub-unit cell in the in-plane hexagonal CNT structure, with 0.145 nm and 0.251 nm lattice para-
meters along the [1 0 0] and [0 1 0] directions, respectively. The smallest length difference (e.g. minimum
lattice registry mismatch) between integer numbers of Au–Au and C–C distances occurs for 1 unit cell
along the [0,1,1]/[1,1,0]-directions (0.288 nm) and 2 unit cells along [1 0 0]-direction (0.290 nm),
respectively. Accordingly, for the (0,2,2) Au/(0 0 1) C interface, the smallest length difference along
the other two orthogonal in-plane directions occurs for 3 Au unit cells along the [1 0 0]-direction
(1.221 nm) and 5 unit cells of C along the [0 1 0]-direction (1.258 nm). Similarly, for the (2,2,4)Au/
(0 0 1)C interface, the minimum lattice mismatch occurs for 3 unit cells of Au along the [1,1,1]
direction (2.115 nm) and 8 unit cells of C along the [0 1 0]-direction (2.012 nm). By defining the strain
condition as aCNT–aAu/aCNT (where aCNT and aAu are the facing CNT/Au lattice lengths), a 3% (compressive)
and a 5% (tensile) strain are expected for the first and the second case, respectively.
The mechanical deformation of the MWCNT in the proximity of the site where the Au-NPs were
attached could be therefore tentatively explained as a consequence of induced strain relaxation by
the imperfect commensuration between the Au and C unit cells. It was quite interesting to note that
the effect was pronounced as the concentration of Au-NPs increased. This formed the basis of study of
the effect of functionalization on the Raman modes of MWCNTs. The HRTEM studies thus concluded
that the acid treatment resulted in the cleavage of MWCNTs and that the Au-NPs randomly function-
alized at the surface of MWCNTs.
The acid treatment and functionalization of Au-NPs, both have introduced the stresses on the
surface of MWCNTs which were further studied by the X-ray diffraction and Raman spectroscopy as
discussed in the following.

4.3. X-ray diffraction

Fig. 6(a–c) shows the XRD patterns of (a) the pristine MWCNTs, (b) oxidized MWCNTs, and (c)
Au-NPs functionalized MWCNTs. Fig. 6(a) shows a strong graphitic peak C (0 0 2) at 26° with an inter-
layer distance (d002) of 3.41 Å for the pristine MWCNTs. Also C (1 0 0) and C (1 0 1) peaks were
observed at 42.5° and 44.5°, respectively. In addition, a minute diffraction peak at 47.5° was also
observed due to Fe (catalyst) present in the MWCNTs. The C (0 0 2) peak was used to determine the
graphitic nature and also to explain the defects, which were produced during the oxidation and Au-
NPs functionalization processes. For ideal (0 0 2) reflection, d002 should be 3.35 Å [35]. Thus a residual
stress of 1.70 (±0.03)% was observed in the pristine MWCNTs attributed to the presence of amorphous
carbon and Fe in the structure. The X-ray diffraction pattern of the oxidized MWCNTs is shown in
Fig. 6(b), which showed a slight shift of (0 0 2) peak and gave an interlayer spacing d002 of 3.43 Å.
The activation of –COOH bond at the surface of MWCNTs increased the value of the stress to 2.10
(±0.03)%. The XRD pattern of the Au-NPs functionalized MWCNTs hybrid is shown in Fig. 6(c). The
C (0 0 2) peak was further down shifted to 25.92° (overall D2h = 0.20°). In addition, diffraction from
Au (1 1 1), Au (2 0 0), Au (2 2 0) were also observed at 38.5°, 44.4° and 64.8°, respectively. The XRD
analysis has further confirmed that oxidation and Au-NPs functionalization has created defect sites
which resulted in 2.10 ± 0.03% increase in the stress and resulted the peak shift by D2h = 0.20° of in
C (0 0 2).

4.4. Raman spectroscopy

4.4.1. Pristine and oxidized MWCNTs

In the next step, the effect of oxidation and Au-NPs functionalization on the Raman modes of
MWCNTs was studied extensively. Fig. 7(a) shows the Raman spectrum of the pristine MWCNTs.
The G band was observed at 1601 cm1, which originated from the circumferential C–C vibrations
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 255

Au Nps & MWCNTs

Au(111)
(c)

Au(220)
Au(200)
(b) Oxidized MWCNTs
Intensity (arb. u)

MWCNTs
C(002)

(a) C(101)
C(100)

FeC3

20 30 40 50 60 70
2θ (degrees)
Fig. 6. XRD patterns of (a) pristine, (b) oxidized and (c) Au-NPs functionalized MWCNTs. Peaks are labeled for their diffraction
planes. Additional peaks Au (1 1 1), Au (2 0 0) and Au (2 2 0) observed in the Au-NPs functionalized MWCNTs are also labeled in
(c).

0
(TO phonon) or due to sp2 bonding [36]. A hump in the tail of G band was due to D band and a
Gaussian function was made to fit and find its contribution in the Raman scattering process.
0
The D and D bands were observed at 1366.8 cm1 and 1637.7 cm1, respectively. The presence of
the D band was a signature of intrinsic defects present in the rolled graphene sheets and curvature
0
at ends of MWCNTs. Intense D vibrational mode was observed probably due to intercalated graphite
compounds. It was enabled due to small vibrational domains in the direction perpendicular to
MWCNTs wall and appeared at the tail of the G band as expected [37]. The ratio of the two Raman
scattering intensities, i.e., D band to the G band (ID/IG) is a metric of the crystal quality of MWCNTs
and it was 1.02 for pristine MWCNTs. It has been reported that an increase in the ID/IG ratio represent-
ed the presence of more defective sites due to covering of MWCNTs with other materials or functional
0
groups [38]. Fig. 7(b) shows the Raman spectra of the oxidized MWCNTs, where G, D and D were
1 1 1
observed at shifted positions of 1602.9 cm , 1368.7 cm and 1639 cm , respectively. The oxidized
0
MWCNTs have shown nearly a 2 cm1 blue shift for the G, D and D bands. Also ID/IG increased consid-
erably to 1.27 from 1.02. The observed blue shift in the G band was possibly due to creation of circum-
ferential and axial stresses [39] as a result of oxidation. The formation of –COOH bonds at the surface
of the oxidized MWCNTs had become responsible for the enhanced disorder and created stresses to
result in the blue shift of the Raman bands and enhanced intensity ratio RD,G to 1.27.

4.4.2. Au-NPs functionalized MWCNTs

0
Fig. 8(a) shows the Raman spectra of the G and D bands fitted with two Gaussian functions for all
0
dried Au-NPs hybrid MWCNTs samples. The peak shift of D, G and D bands as a function of Au molar
concentration is plotted in Fig. 8(b). All samples showed identical behavior for the shift of Raman
modes with the Au-NPs functionalization. The Raman bands could shift for two reasons, one – due
256 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

1200 1300 1400 1500 1600 1700 1800

10000
Oxidized MWCNTS
8000

6000

4000
Raman Intensity (arb. Unit)

2000 (b)

0
1500 Pristine MWCNTS

G band
1000 D band

/
500 D band
(a)
0
1200 1300 1400 1500 1600 1700 1800
Raman Shift (cm-1)

Fig. 7. Raman spectra of (a) pristine MWCNTs, (b) oxidized MWCNTs. Major D and G bands were fitted with LORENTZIAN
0
functions to resolve the D band from the G band.

to mechanical stresses produced by the Au-NPs, and two – due to coupling of Raman modes to the
surface charge present on Au-NPs. For 40 nM Au-NPs and MWCNTs composites, there was a blue shift
of 2 cm1 for all bands and then there was a red shift of nearly 10 cm1 of the G band for 100 nM Au-
NPs MWCNTs composites. For 40 nM Au-NPs, MWCNTs composites, the –COOH functional group cre-
ated the stone-wale (SW) defects, which were more reactive sites and provided nucleation sites for the
attachment of Au-NPs [40], and in this case, the effect of Au-NPs on electron screening was weak.
Thus, the mechanical stresses produced due to loading of Au-NPs at the surface of MWCNTS were
dominant to result in the blue shift of the G band. The gradual increase in the concentration of func-
tionalized Au-NPs to 100 nM Au-NPs in the MWCNTs composites resulted in the increased local charge
density and its coupling to the Raman active phonons resulted in the red shift of the G band [41–44].
0
While, the red shift in the D (D) Raman bands of nearly 10 (8) cm1 was also observed which clearly
showed the increased mechanical stresses introduced by the increased concentration of Au-NPs. The
ratio of ID to IG (RD,G) is plotted in Fig. 8(c), which decreased linearly from 1.27 for oxidized MWCNTs to
1.03 for 80 nM Au-NPs–MWCNTs composites and then increased back to 1.21 for 100 nM Au-NPs–
MWCNTs composites. This was due to the fact that for small concentrations of Au-NPs covered only
the defective sites that originated from pentagons, heptagons and dangling bonds, which initiated
the functionalization of Au-NPs to form NP islands on MWCNTs. In composites with 100 nM Au-
NPs, islands coalesced to increase the encapsulation of MWCNTs significantly. This resulted in an
increase of the RD,G and weakening of the Raman G feature relative to composites made with lower
Au-NPs concentrations [45]. From this discussion, it was concluded that Au-NPs functionalized the
MWCNTs at the SW defect sites, which decreased the RD,G to 1.03 and then attached at the remaining
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 257

1200 1300 1400 1500 1600 1700 0 20 40 60 80 100

5
/
D band
100 nM (a) (b)
0

-5

80 nM -10

-15
Intensity (arb. u.)

G band
0

Peak shift (cm )

-1
60 nM -5

-10
40 nM
-15
D band
0

Oxidized D band / -5
G band D band
MWCNTS
-10

-15
1200 1300 1400 1500 1600 1700 0 20 40 60 80 100
Raman Shift (cm-1) Au-NPs conc. (nM)

1.30
(c)
1.25

1.20
ID /I G

1.15

1.10

1.05

0 20 40 60 80 100
Au-NPs Conc. (nM)
Fig. 8. (a) Raman spectra of the oxidized MWCNTs (bottom), 40 nM (second from bottom), 60 nM (third from bottom), 80 nM
(second from top) and 100 nM (top spectrum) Au-NPs functionalized MWCNTs. The peak energy position, width and
0
corresponding intensity was obtained from the Lorentzian fits for D, G and D band as shown in all spectra. (b) The plot of the
0
peak energy shifts in the D, G and D bands as a function of Au-NPs molar concentration. (c) The plot of the ratio of the peak
intensities (ID–IG) as a function of Au-NPs molar concentration.

surface of MWCNTs which created additional stresses and increased RD,G to 1.21. The red shift of bands
0
was also attributed to the enhanced mechanical stresses (D and D) and charge screening (G band)
with the increase in Au-NPs. The average value of defects as a function of Au-NP concentrations
remained in the limit of 2.6–3 1011  cm2 (surface defects) as estimated from the Raman spectra.
This number was still higher than in the pristine MWCNTs.
258 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

0.15

G band
0.10
Raman Intensity
D band
D/ band
0.05

0.00

1200 1350 1500 1650 1800

Raman Shift (cm-1)

Fig. 9. Observed and Lorentzian fitted Raman spectra of the oxidized MWCNTs in water. A blue shift in the bands was observed
with respect to the dried oxidized MWCNTs.

4.4.3. Effect of E-coli on Raman modes

Fig. 9 shows the Raman spectra of the oxidized MWCNTs dispersed in DI water. The G band was
0 0
observed at 1605 cm1, while D and D were at 1371 cm1, 1648 cm1, respectively. The D and G
0
bands were again resolved by fitting two Gaussian functions. G, D and D bands showed an upward
shift of 4 cm1, 3 cm1, 9 cm1, respectively compared to the Raman shift observed in the dried oxi-
dized MWCNTs. This was attributed to the circumferential and axial stresses caused by DI water mole-
cules adsorbed on the surface. In the next step, the effect of attachment of E-coli on the Raman modes
of composites was studied in detail.
Fig. 10(a–d) shows the D and G bands Raman spectra of the nanohybrid formed for (a) 40 nM, (b)
60 nM, (c) 80 nM, and (d) 100 nM Au-NPs–MWCNTs for different E-coli concentrations, namely
102 ml1, 103 ml1, 104 ml1 and 105 ml1. The bottom two spectra were from the oxidized MWCNTs
and dried Au-NPs–MWCNTs composites as discussed in the last section. The spectra are placed verti-
cally for clarity. A number of observations were drawn by having a careful look at the spectra. For
small concentration of Au-NPs, the Raman spectra became very weak and broad as the concentration
of E-coli was increased (Fig. 10(a)). On the other hand, the Raman spectra remained quite sharp in the
case of 100 nM Au-NPs composites even for higher concentrations of E-coli (Fig. 10(d)). The clear dif-
ference between the qualities of the Raman signals for the two composites demonstrated that concen-
tration of Au-NPs in the nanohybrid composites could be a key parameter for its biological
applications. For low concentration of Au-NPs in the nanohybrid and higher concentration of E-coli,
it was expected that all Au-NPs surfaces were covered with E-coli and the additional E-coli were
attached to the bare surface of CNTs, which affected the Raman signal. The situation gradually
improved when higher concentration of Au-NPs was used as can be seen in Fig. 10 (from a to d), where
the Raman signal improved as obtained from 40 nM to 100 nM Au-NPs hybrid. This was attributed to
enhanced surface coverage of CNTs by Au-NPs leaving less surfaces for E-coli to interact with CNTs
directly. This was probably due to high affinity of E-coli to attach to Au surface than to CNTs. In addi-
tion, following observations were made in the Raman spectra for all composites as the concentration
of E-coli increased. For 40 nM Au-NPs–MWCNTs composites, with increase in E-coli concentration,
Raman signal became weak and broad. For 80 nM and 100 nM composites, G band was shifted towards
higher wave numbers, which was due to increase in charge transfer from E-coli. Fig. 11 shows the shift
in the G band for each concentration of Au-NP-MWCNTs hybrid when loaded with various concentra-
tions of E-coli. It is assumed that now the shift in the G band is due to charge transfer from the com-
posite. The experimental points were fitted with a straight line to determine the slope. It was observed
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 259

5
(a) 40 nM (b) 60 nM
10 /ml

4
10 /ml

3
10 /ml

2
10 /ml
Normalized intensity

Au bonded

Oxidized

(c) 80 nM (d) 100 nM

1200 1350 1500 1650 1800 1200 1350 1500 1650 1800
-1
Raman shift (cm )
Fig. 10. (a–d): Raman spectra of various concentrations of Au-NPs–MWCNTs composites reacted with E-coli bacteria for (a)
40 nM, (b) 60 nM, (c) 80 nM, and (d) 100 nM of Au-NPs concentrations.

that composite formed with 80 nM Au-NP-MWCNTs displayed most drastic change with the E-coli
concentration. The shifting of G band is an indication of transfer of charge from the CNTs to the bac-
teria [29,41], which is strongest for the 80 nM Au-NP-MWCNTs composite. It is interesting to note that
at this concentration, the ratio ID/IG was the lowest, which meant a large of already existing defects.
This may also lead to conclude that the presence of defects help transfer of charge from the composite,
thus making it more sensitive for detection or killing of bacteria. The difference between the G and D
bands also increased with the increase in E-coli concentration (5 cm1 increase for 80 nM and 4 cm1
increase for 100 nM), which indicated that for higher concentrations of Au-NPs, the shift was attribut-
ed to more charge transfer than mechanical stresses.
The densities of Au-NPs covering the MWCNTs determined for each concentration were
3.0  1012 m3, 4.5  1012 m3, 6.0  1012 m3 and 7.6  1012 m3 for 40 nM, 60 nM, 80 nM and
100 nM, respectively. It was expected that for small concentrations of E-coli from 102 ml to 104 ml,
not all Au-NPs were covered by E-coli. As soon as E-coli got attached to CNTs, there would be transfer
of charge from E-coli to CNTs as E-coli surface has a negative charge. This affected the Raman modes,
which was quite evident in the Raman spectra for high concentrations of E-coli in Fig. 10(a). However,
for increased Au-NPs in composites, the Raman signal for higher concentrations also improved as
E-coli had more Au-NPs to attach. Increase in the free carrier densities due to electron transfer to
Au-NPs and MWCNTs composites were also observed by UV–Vis spectroscopy as discussed in the
following section.
260 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

Fig. 11. Plot of shift in G band of each series of Au-MWCNTs hybrid as a function of E-coli concentration. The lines are the linear
fits to the experimental points. The slopes of the lines are 0.996 ± 0.168, 0.483 ± 0.047, 1.797 ± 0.231, and 1.088 ± 0.077,
for 40 nM, 60 nM, 80 nM and 100 nM, respectively.

4.4.4. UV–Vis spectroscopy

It was interesting to perform absorption spectroscopy of Au-NPs in suspension solution and com-
posites with MWCNTs. A peak at 526 ± 2 nm in the absorption spectrum was clearly observed due to
surface plasmons of Au-NPs as shown in Fig. 11 for all concentrations, i.e., from 40 nM to 100 nM. All
Au-NPs concentrations showed the absorption peak at the same position with more or less identical
width (52 ± 2 nm) as clearly shown in Fig. 11(a–d). This reflected the uniformity in the size of the syn-
thesized Au-NPs. However, their number densities were quite different as have already been deter-
mined from the SEM images.
This picture modified drastically for the Au-NPs–MWCNTs composites, the absorption peak
became weak, and blue shifted. The absorption spectra of the composites after subtracting the back-
ground along with a reference absorption spectra of MWCNTs only (bottom spectrum) is shown in
Fig. 12(a) for 40 nM and (b) 80 nM Au-NPs–MWCNTs composites (2nd spectra from bottom) and then
mixed with varied concentrations of E-coli (upper 4 spectra). It can be seen in Fig. 12 that there was no
absorption in MWCNTs in the region of interest (bottom reference spectrum). However, when Au-NPs
were attached to MWCNTs, the absorption peak appeared at 497 nm and was less broad and asym-
metric compared to the one obtained for pure Au-NPs. The average width of peaks was also reduced
to 34 ± 4 nm. The variation in the absorption peak position and its width as a function of E-coli con-
centration is plotted in Fig. 13(a and b) for 40 nM (red) and 80 nM (black) Au-NPs-MWCNT nanohy-
brids. It was observed that 40 nM Au-NPs–MWCNTs composites were more sensitive to the E-coli
concentration compared to 80 nM Au-NPs MWCNTs composites as the absorption peak shifted for
higher concentrations of E-coli. However, the widths of the absorption peaks showed identical behav-
ior (Gaussian).
The shift in the plasmon resonance as shown in Fig. 14(a) in the Au-NPs- MWCNTs composites
could be ascribed to two factors; (i) a change in the medium’s refractive index [46], and (ii) to a change
in the free carrier concentration [47,48]. In the present case, the binding of Au-NPs to the surface of
MWCNTs was believed to change the free carrier density in Au-NPs, which caused a shift in the surface
plasmon resonance. The latter was due to Au-NPs merged into first few layers of MWCNTs as clearly
seen in the TEM images. This reduced the effective radii of Au-NPs by about 20%, which modified the
local charge densities in the Au-NPs sphere and caused a blue shift in the absorption [49]. The reduc-
tion in size also affected the mean free path of the scattering events, which reduced the width of the
absorption peaks (see Fig. 14(b)).
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 261

(d) 100 nM

(c) 80 nM
Normalized Intensity

(b) 60 nM

(a) 40 nM

425 450 475 500 525 550 575 600 625

Energy (nm)
Fig. 12. Normalized absorption spectra of Au-NPs prepared for different concentrations from 40 nM (bottom) to 100 nM (top)
as mentioned in figure.

Finally, absorption spectroscopy confirmed that conjugate or nanohybrid formed by Au-NPs and
MWCNTs modify the carrier density in Au-NPs. In addition, extent of penetration of Au-NPs in
MWCNTs varied to result in asymmetrical absorption peak as the mean free path is reduced.

5. Conclusion

The present study demonstrated the effect of concentration of Au-NPs in the Au-NPs–MWCNTs
nanohybrids on the structural characteristics of MWCNTs. It also demonstrated that the composition
of constituent components of the composite is crucial to avoid the modifications in the system prop-
erties. Such composites are important where CNTs role is of a substrate or platform for NPs for biolo-
gical applications. It was observed that MWCNTs were eroded on oxidation as number of layers
262 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264

(a) 40 nM (b) 80 nM
5
10 /ml

4
10 /ml
Normalized Intensity

3
10 /ml

2
10 /ml

Au+MWCNTs

MWCNTs

470 480 490 500 510 470 480 490 500 510 520
Energy (nm)
Fig. 13. Normalized absorption spectra of Au-NPs–MWCNTs composites mixed with various concentrations of E-coli as
mentioned in figure for Au-NPs concentration (a) 40 nM, and (b) 80 nM. The bottom spectra in both (a) and (b) are from
MWCNTs only showing flat response in the region of interest, i.e., plasmon response.

decreased after acid treatment and created stresses and nucleation sites for Au-NPs to attach on the
surface of MWCNTs. The formation of defects was confirmed by the XRD and Raman spectroscopy.
Electron microscopy has shown that immobilized Au-NPs penetrated a few layers into the MWCNTs.
Strong variation in the Raman modes of MWCNTs successfully proved the crucial role of structural
modifications in the composites with the increase of Au-NPs and increased concentrations of E-coli.
It was found that Raman modes remained unchanged when E-coli was attached to Au-NPs. However,
for increased E-coli concentration, it also covered MWCNTs, which affected Raman bands adversely,
which clearly indicated that nature of CNTs was modified. The effect of Au-NPs penetration in the
MWCNTs was responsible for the blue shift of absorption peak and reduced peak width. The modified
properties of MWCNTs and Au-NPs were attributed to the change in the total charge present at the
surface of MWCNTs and stresses produced as a result of Au-NPs attachment. The present study suc-
cessfully demonstrated that the concentration of Au-NPs in CNTs composites is crucial for its biologi-
cal applications.
 S. Mehmood et al. / Superlattices and Microstructures 81 (2015) 248–264 263

500
(a) 80 nM
40 nM
499

Peak position (nm)

498

497

496

495
(b)
40
Width (nm)

38

36

34

32

30
102
Au + CNTs

103 104 105

Au + CNTs + E-coli (/ml)

Fig. 14. Variation in the (a) Plasmon absorption peak, and (b) absorption peak widths as a function of E-coli concentration for
40 nM (red) and 80 nM (black) Au-NPs–MWCNTs composites. The solid lines are the fits to the experimental values showing the
trend. (For interpretation of the references to color in this figure. 14 legend, the reader is referred to the web version of this
article.)

Acknowledgements

The research work was funded by the HEC NRPU grant numbers 261 and 1770. One of authors SM
is thankful to the Higher Education Commission for PhD scholarship. Authors are thankful to ICTP and
Center for Electron Microscopy of CNR-IOM, TASC laboratories, Trieste Italy for TEM experiments.

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