Tunable NIR-II emitting silver chalcogenide quantum dots using thio/selenourea precursors: Preparation of an MRI/NIR-II multimodal imaging agent

Article abstract of DOI:10.1039/d0dt02974b

Aqueous-stable, Cd- and Pb-free colloidal quantum dots with fluorescence properties in the second near-infrared region (NIR-II, 1000-1400) are highly desirable for non-invasive deep-tissue optical imaging and biosensing. The low band-gap semiconductor, silver chalcogenide, offers a non-toxic and stable alternative to existing Pd, As, Hg and Cd-based NIR-II colloidal quantum dots (QDs). We report facile access to NIR-II emission windows with Ag2X (X = S, Se) QDs using easy-to-prepare thio/selenourea precursors and their analogues. The aqueous phase transfer of these QDs with a high conservation of fluorescence quantum yield (retention up to ～90%) and colloidal stability is demonstrated. A bimodal NIR-II/MRI contrast agent with a tunable fluorescence and high T1 relaxivity of 408 mM-1 s-1 per QD (size ～ 2.2 nm) and 990 mM-1 s-1 per QD (size ～ 4.2 nm) has been prepared by grafting 50 and 120 monoaqua Gd(iii) complexes respectively to two differently sized Ag2S QDs. The size of the nanocrystals is crucial for tuning the Gd payload and the relaxivity.

Full text of DOI:10.1039/d0dt02974b

View Article Online
View Journal
Dalton
Transactions
An international journal of inorganic chemistry
Accepted Manuscript
This article can be cited before page numbers have been issued, to do this please use: K. Bhardwaj, S.
Pradhan, S. Basel, M. Clarke, B. Brito, S. Thapa, P. Roy, S. Borthakur, L. Saikia, A. Shankar, G. J. Stasiuk, A.
Pariyar and S. Tamang, Dalton Trans., 2020, DOI: 10.1039/D0DT02974B.
Volume 46
Number 10
14 March 2017
Pages 3073-3412
This is an Accepted Manuscript, which has been through the
Royal Society of Chemistry peer review process and has been
accepted for publication.
Dalton
Transactions
An international journal of inorganic chemistry
Accepted Manuscripts are published online shortly after acceptance,
before technical editing, formatting and proof reading. Using this free
rsc.li/dalton
service, authors can make their results available to the community, in
citable form, before we publish the edited article. We will replace this
Accepted Manuscript with the edited and formatted Advance Article as
soon as it is available.
You can find more information about Accepted Manuscripts in the
Information for Authors.
Please note that technical editing may introduce minor changes to the
text and/or graphics, which may alter content. The journal’s standard
Terms & Conditions and the Ethical guidelines still apply. In no event
ISSN 0306-0012
COMMUNICATION
Douglas W. Stephen et al.
shall the Royal Society of Chemistry be held responsible for any errors
N-Heterocyclic carbene stabilized parent sulfenyl, selenenyl,
and tellurenyl cations (XH+, X = S, Se, Te)
or omissions in this Accepted Manuscript or any consequences arising
from the use of any information it contains.
rsc.li/dalton

Please do not adjust margins
Page 1 of 9
Dalton Transactions
View Article Online
DOI: 10.1039/D0DT02974B
ARTICLE
Tunable NIR-II Emitting Silver Chalcogenide Quantum Dots using
Thio/Selenourea Precursors: Preparation of MRI/NIR-II
Multimodal Imaging Agent
Received 00th January 20xx,
Accepted 00th January 20xx
Karishma Bhardwaj,a, ‡ Sajan Pradhan,a, ‡ Siddhant Basel,a Mitchell Clarke,b Beatriz Brito,b,c Surakcha
a
a
e
b,c,
d
d
DOI: 10.1039/x0xx00000x
Thapa, Pankaj Roy, Sukanya Borthakur, Lakshi Saikia, Amit Shankar, Graeme J. Stasiuk,
Anand Pariyar,a and Sudarsan Tamanga,*
*
Aqueous-stable, Cd- and Pb- free colloidal quantum dots with fluorescence properties in the second near-infrared region
(NIR-II, 1000-1400) are highly desirable for non-invasive deep-tissue optical imaging and biosensing. The low band-gap
semiconductor, silver chalcogenide offers a non-toxic and stable alternative to existing Pd, As, Hg and Cd-based NIR-II
colloidal quantum dots (QDs). We report facile access to NIR-II emission windows with Ag2X (X=S, Se) QDs using easy-to-
prepare thio/selenourea precursors and their analogues. Aqueous phase transfer of these QDs with a high conservation of
fluorescence quantum yield (retention up to ~90%) and colloidal stability is demonstrated. Bimodal NIR-II/MRI contrast
agent with tunable fluorescence and high T1 relaxivity 408 mM−1 s−1 per QD (size ~2.2 nm) and 990 mM−1 s−1 per QD (size ~
4.2 nm) have been prepared by grafting 50 and 120 monoaqua Gd (III) complexes respectively to two different sized Ag2S
QDs. The size of the nanocrystals is crucial for tuning Gd payload and the relaxivity.
leads to the formation of a multimodal contrast agent which
facilitates better visualization compared to a single imaging
technique.17–19 For example, MRI/NIR-II fluorescent imaging
probes enable both excellent molecular level sensitivity
(fluorescence) and anatomic resolution (MRI) due to the
combination of fluorescence and MRI techniques. In recent
years, the multimodal imaging agents have gained immense
significance in the domain of biomedical imaging.18–22
The majority of the existing line of QDs -based NIR-II
fluorophores contain highly toxic Cd2+, As3-, Pb2+ and Hg2+
ions23,24 posing a real concern and practical barrier for their
application in bio-imaging. In this respect, silver chalcogenides
have tremendous potential in the field of in vivo bioimaging and
sensing.2–4,25–27 Ag2S is non-toxic and has extremely low
solubility product (Ksp = 6.3 × 10–50) with minimum risk of
releasing Ag+ ions into the biological environment.3,28,29
Furthermore, its low band-gap (bulk: 1.1 eV) and possible
quantum confinement effect at size regime below 4 nm (exciton
Bohr radius: 2.2 nm) are ideal for tuning of their emission in the
Introduction
In biological imaging, the second near-infrared (NIR-II, 1000-
1400 nm) emitting fluorophores afford, in principle, a higher
signal-to-noise ratio compared to their visible (400-750 nm) and
the NIR-I (750-900 nm region) counterparts.1–8 A combination
of factors such as low absorbance of water and other
biomolecules and low scattering/autofluorescence by tissue
make the NIR-II window ideal for deep-tissue optical imaging
and sensing with a high spatial and molecular resolution.5–9
Compared to organic dyes, colloidal quantum dots (QDs) based
NIR-II fluorophores have superior quantum yield, size-tunable
fluorescence, broad absorption window and high molar
extinction coefficient.10–12 In addition, the surface of the QDs
can be grafted with a high payload of functional molecules such
as drugs, antibody, peptides, nucleic acids, cell-penetrating
peptides, MRI contrast agent etc.13–17 Specially, functionalizing
QDs with another imaging agent such as a MRI contrast agent
~
700-1400 nm range.13 Similarly, the bulk band-gap Ag2Se is
a.Department of Chemistry, School of Physical Sciences, Sikkim University, Sikkim
737102, India. E-mail: stamang@cus.ac.in
0.15 eV30,31 and their calculated exciton Bohr radius using bulk
parameters is 2.9 nm. Strong absorption and emission tunability
at sizes well below 4-5 nm are important for achieving low
hydrodynamic diameter and renal-clearance from the body.32
Studies have shown that Ag2S exhibit negligible cytotoxic or
genotoxic effects at lower to moderate doses, making it an ideal
candidate for practical use in in-vivo imaging.3,28,29,33,34 Despite
b.Department of Biomedical Sciences, University of Hull, Hull, HU6 7RX, UK.
c. Department of Imaging Chemistry and Biology, School of Biomedical Engineering
and Imaging, King’s College London, St Thomas’ Hospital, London, SE1 7EH,
United Kingdom. E-mail: graeme.stasiuk@kcl.ac.uk
d.Department of Material Science, North East Institute of Science and Technology
(NEIST), Assam 785006, India.
e. Department of Physics, Kurseong College, West Bengal, India 734203.
‡ These authors contributed equally to this work.
Electronic Supplementary Information (ESI) available: NMR spectra, FTIR spectra, tremendous potential in biology, the colloidal synthesis of Ag2X
and TGA spectra of precursors. Additional optical characterization, photographs and
XPS spectra of colloidal quantum dots solution. Optical constants of Ag2S and DFT
calculation. NMR, Mass and FTIR characterization of Gd complex and relaxivity
studies data. See DOI: 10.1039/x0xx00000x
(X=S, Se) is still less explored compared to II-IV, III-V and II-VI
semiconductor NCs. Typically, the control of the size of Ag2X
QDs has been accomplished mainly either by changing of
Please do not adjust margins

Please do not adjust margins
Dalton Transactions
Page 2 of 9
ARTICLE
Journal Name
View Article Online
DOI: 10.1039/D0DT02974B
(a)
(b)
(d)
(c)
5b, 5a, 5c, 7b, 7a, 7c, 9b, 9a
3a, 3b, 3c, 3d, 3e, 3f, 3g, 3h, 3i
1000
1250
1500
1000
1200
1400
Wavelength (nm)
Wavelength (nm)
Figure 1. (a) Synthesis of a wide range of thiourea, thiocarbamate, dithiocarbamates and selenourea derivatives. (b) General
scheme for the synthesis of Ag2S/Se colloidal quantum dots (QDs). (c) PL spectra with tunable emission wavelength of as-prepared
Ag2S QDs synthesized from thiourea precursors (3a-g) and (d) Ag2S/Se QDs synthesized from thiocarbamate (5a-c),
dithiocarbamates (7a-c) and selenourea derivatives (9a-b).
temperature35,36 and reaction time31,37 or both the
parameters.38,39 Both changing time and temperature can
lead to the tunability of average final size, however, the
methods undermine the competitive effects such as Ostwald
ripening and uncontrolled depletion of molecular precursor,
both leading to diminished control over final size-
distribution. Size-tunability of quantum dots is crucial in
many important areas of bio-imaging including-but not
limited to- (a) application in multiplexed identification of
several bio-molecules as spectrally distinct barcodes,40,41 (b)
controlling hydrodynamic diameter for efficient clearance of
the intravenously injected quantum dots from the body32
and (c) controlling functional molecule pay-load in quantum
dots-based multimodal contrast agents.16,17,42
prepare an MRI/NIR-II imaging agent. The details of the
finding are in the following.
Result and discussion
We prepared a range of sulfur (substituted thiourea,
thiocarbamate and dithiocarbamates) and selenium
precursors
with
the
direct
reaction
between
isothiocyanate/isoselenocyanate
with commercially
available phenols, thiophenols or aryl amines (Figure1a). The
reaction was carried out using off-the-shelf reagents in a
suitable solvent such as chloroform, ethanol, dioxane or
toluene at room temperature (cf. ESI; Experimental section),
depending upon the solubility of the reactants. The reaction
proceeds effortlessly yielding a good to excellent product
A recent study by Vela et al. on II-VI semiconductor NCs43,44
and Owen45,46 et al. on the synthesis of II-VI and IV-VI
semiconductor NCs have introduced a more rational
approach to control the photophysical properties of
nanocrystals (NCs) via the reactivity of precursor. Vela et al.
demonstrated the impact of phosphine-chalcogenide
precursor reactivity on the composition and morphology of
CdS/CdSe NCs.43,44 Similarly, the size-dependent optical
properties of PbS/PbSe was later reported by Owen et al. by
modulating the reactivity of thio/selenourea derivatives.45,46
However, the strategies have so far not been extended to
Ag2X (X=S, Se) NCs. Herein, we report a facile synthesis of
Ag2X (X=S, Se) NCs with environmentally benign and
biologically conducive attributes tunable in the crucial NIR
region (900-1400 nm) by controlling the reactivity of
substituted thio/selenourea and their analogues via
modification of substituent groups. These NCs have been
successfully phase transferred to the aqueous medium and
functionalized with a T1-weighed MRI contrast agent to
yield (~up to 91 %) within a short time (~5-30 min, Figure 1a).
In some cases, for example, in the reaction of nitro aniline
with isothiocyanate in acetonitrile, heating to reflux
temperature and longer reaction time is required. Inspired
by the work of Owen and co-workers46 on the synthesis of
PbS, ZnS, SnS and CdS QDs using thiourea precursors, our
initial attempt to synthesize the Ag2S QDs by reacting Ag-
oleate with thiourea in 1-octadecene (a non-coordinating
solvent) at precursor decomposition temperature (~150 °C)
was unsuccessful. No photoluminescence (PL) emission was
detected and the colloidal dispersion was poor (Figure S2).
The colloidal stability and optical properties could not be
improved even when an amine (oleyl amine) was added
during the reaction as a co-capping ligand (Figure S3).
However, when 1-dodecanethiol (DDT) was used as a
capping ligand, stable Ag2S QDs was formed with excellent
optical properties. DDT as capping ligand was found to be
crucial for the successful synthesis of Ag2S QDs as the soft
acid Ag (I)ion prefers soft bases like thiol (-SH) or thiolate
more than amines or carboxylates. We propose that the
2 | J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 3 of 9
Dalton Transactions
Please do not adjust margins
Journal Name
ARTICLE
View Article Online
DOI: 10.1039/D0DT02974B
OMe (3c)
Me (3b)
H (3a)
QD1
(c)
(e)
(c)
(e)
QD2
(a)
I (3d)
NO (3e)
2
0.26 nm
(-1,2,1)
0.22 nm
(0,3,1)
20 nm
20 nm
800 1000 1200 1400 1600
Wavelength (nm)
(f)
(b)
QD1
(d)
QD2
OMe (3c)
Me (3b)
H (3a)
I (3d)
NO (3e)
2
-0.3 0.0 0.3 0.6 0.9
15 30 45 60 75
20
40
60
80
Hammet constant ()
2 (degrees)
2 (degrees)
Figure 2. (a) PL spectra of Ag2S QDs synthesized from thiourea precursors (3a-e) bearing various substituents at the para position
of phenyl ring (H, p-Me, p-OMe, p-I and p-NO2); (b) A linear correlation of PL emission wavelength ( max) obtained from (3a-e) with
corresponding Hammett constants (σ) for para substituents. (c) TEM micrograph (size 2.2 ± 0.52 nm) and (d) XRD pattern of
purified QD1 obtained from 3c. (e) TEM micrograph (size 4.2 ± 1.1 nm) of and (f) XRD pattern of purified QD2 obtained from 3e.

~
~
Insets in figure c and e show the lattice fringes corresponding to the monoclinic phase. In figure d and f, the XRD reflections of
monoclinic Ag2S (JCPDS 14-0072) are shown as bars for comparison.
termination of surface of QDs with thiol-containing capping
agents improve the colloidal stability and leads to
monodisperse QDs with characteristic optical properties.47–
49 Furthermore, the standard redox potential of thiol-bound
Ag1+ ion is lower than free Ag1+ ion which implies that the
thiol-bound Ag (I) is not easily reduced to the undesired
species.31,47,48 In a typical synthesis of Ag2S QDs, as prepared
substituted thiourea (3a-g) dissolved in DME, was quickly
injected to the hot solution of the silver precursor containing
a mixture of AgNO3 and DDT in ODE at elevated temperature
(120-150 °C) (Figure 1b). A distinct color change from red to
black after 15 min indicated the formation of Ag2S QDs and
the reaction was quenched by immediate cooling in an ice
bath. To further understand the role of DDT we attempted
to synthesize Ag2S QDs using only DDT at the reaction
temperatures (120-150 °C). No formation of Ag2S QDs was
noted in all the cases (Figure S4) possibly due to the high
decomposition temperature (>170 °C) of DDT.50 At elevated
infrared spectroscopy (FTIR) experiment performed on
purified Ag2S QDs confirmed the presence of surface-bound
DDT. The distinct shifting of the FTIR bands to 2916 cm-1 and
2847 cm-1 from 2922 cm-1 and 2851 cm-1 respectively
attributed to the asymmetric and symmetric C-H stretching
of DDT was observed (Figure S7).51 These results show that
DDT solely acts as the capping ligand for QDs in our synthetic
scheme. Similarly, the other precursors such as
thiocarbamates (5a-c) and dithiocarbamates (7a-c) as well as
selenourea derivatives (9a-b) were also effective as a
precursor under the optimized reaction condition to yield
highly fluorescent Ag2S/Se QDs. Figure 1c demonstrates the
photoluminescence (PL) emission spectra of Ag2S QDs in the
NIR region (1020-1210 nm) prepared using (3a-i). Similarly,
the NIR emission was also tunable in the range
1300 nm by simply changing the substituent groups in
thiocarbamates 5a-c), dithiocarbamates 7a-c and
~900 nm to
(
(
)
selenourea precursors (9a-b) under optimized hot injection
reaction condition (Figure 1d). In literature reports, the size
of Ag2X (X=S, Se) QDs has been controlled mainly by
changing the reaction temperature35,36 or the reaction
time31,37 or both the parameters.38,39 Our results offer the
opportunity to control the optical properties of Ag2X (X=S,
Se) QDs by controlling the precursor reactivity and
conversion rates. In addition to this, the change of reaction
temperature (Figure S8) and the reaction time (Figure S9)
also affected the tunability of emission properties as
temperature (~200 °C) DDT formed Ag2S QDs without the
use of additional S precursor, as detected by XRD and optical
studies (Figure S5). However, size distribution and optical
properties (emission) were poor. On the contrary, the lower
decomposition temperatures of as-prepared thiourea
precursor (<150 °C) compared to DDT is conducive for
controlled release of sulphur at the optimized reaction
temperature (120-150 °C, Figure S6) leading to improved size
distribution and optical properties. Fourier transform
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 3
Please do not adjust margins

Please do not adjust margins
Dalton Transactions
Page 4 of 9
ARTICLE
Journal Name
View Article Online
DOI: 10.1039/D0DT02974B
(a)
H2O
CHCl3
After exchange
Before exchange
(c)
Chloroform
Quantum Yield (%)
(b)
(QD1=6.9%);
(QD2=6.2%)
Water
-1
-1
2538 cm
(QD1=5.5%);
(QD2=5.6%)
1588 cm
-1
1572 cm
L-cys
L-cys capped Ag S
2
3150 2520 1890 1260
Wave number (cm )
900 1050 1200 1350 1500
-1
Wavelength (nm)
Figure 3. (a) Phase transfer of DDT-capped Ag2S QDs from organic phase (chloroform) to aqueous phase using L-cysteine at pH ~9.
(b) PL emission spectra of Ag2S QDs (QD1 and QD2) before (dispersed in CHCl3, black) and after ligand exchange process with L-
cysteine (dispersed in water, red). The spectra have been recorded at same absorbance (concentration) in both the phases. (c)
FTIR spectra of free L-cysteine (black) and L-cysteine-capped Ag2S QDs (red).
reported in the literature.
in sizes of QDs formed using 3c (QD1) and 3e (QD2). Ag2S
To understand the effect of thiourea conversion reactivity on
the final size, Ag2S QDs were synthesized using thiourea
precursors (3a-e) with different substituent group under the
same reaction condition (15 min, 150 °C). The final products
were monitored using photoluminescence (PL) emission
spectroscopy, X-ray diffraction (XRD) and transmission
electron microscope (TEM) (Figure 2). The thiourea with no
substituent group in the phenyl ring (3a) yielded Ag2S QDs
with a PL emission peak at 1094 nm (Figure 2a). On
increasing the electron-donating ability by introducing
methyl group as a para substituent in phenyl ring (p-Me, 3b),
the PL emission exhibited a distinct blue shift to 1071 nm.
Further increasing the electron-donating ability by using p-
OMe substituent in phenyl ring (3c) resulted in a blue shift in
QDs were fairly monodisperse with QD1 and QD2 of average
sizes 2.2 ± 0.52 nm and 4.2 ± 1.1 nm respectively (Figure 2c
and 2e). The XRD pattern for QD1 was broader compared to
that of QD2 (Figure 2d and 2f) respectively due to the finite
size effect and the diffraction patterns are consistent with
monoclinic α-Ag2S phase (JCPDS 14-0072).53,54 The strong
electron-withdrawing capacity of p-NO2 group decreased the
reactivity of the thiourea, leading to the formation of larger
sized QDs compared. On the other hand, the presence of
electron-donating group such as p-OMe led to enhanced
reaction rate and smaller QDs. It is interesting to note that,
this trend of size/emission tunability of Ag2S QDs with
respect to the reactivity of the substituted thiourea
precursor is contrary to that observed in the case of PbS, ZnS
or CdS QDs (Figure S10) in literature report46 indicating that
the underlining reaction pathway in our case is possibly
different. A detailed investigation is required to map out the
exact reaction mechanism, however, the above results can
be explained by assuming rate-limiting nucleophilic attack
on Ag+ by the precursor through C=S bond. Furthermore, the
electronics of other analogues of thiourea namely
thiocarbamate (5a-c) and dithiocarbamates (7a-c) revealed a
similar effect on the shift of PL of Ag2S QDs. The
the PL emission (λmax
p-I substituent (3d) having moderate electron-withdrawing
ability resulted in a red-shift of PL peak (λmax 1133 nm). A
significant red-shift (λmax 1209 nm) was observed in the
~1024 nm). In contrast, the presence of
~
~
presence of 3e bearing p-NO2 group indicating the formation
of larger sized Ag2S QDs. Interestingly, the PL emission
maxima linearly correlated with corresponding Hammett
constant (σ) of the individual substituents (3a-e; H, p-Me, p-
OMe, p-I and p-NO2) of thiourea precursor corroborating a
strong and predictable influence of electronic effects (Figure
2b).52 Consistent with the PL spectra, the TEM images (Figure
2c, 2e) and XRD spectra (Figure 2d, 2f) confirm the difference
thiocarbamate (5b) formed smaller Ag2S QDs (λmax~900 nm)
due to its higher reactivity compared to thiourea and
dithiocarbamates (Figure 1d). In the case of Ag2Se QDs
4 | J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 5 of 9
Dalton Transactions
Please do not adjust margins
Journal Name
ARTICLE
View Article Online
DOI: 10.1039/D0DT02974B
(a)
(b)
(c)
L-Cys capped QD
-1
bulk H2O
1572 cm
Gd-L
-1
1610 cm
H2O
Exchange
∆δ = 0.102
-1
1676 cm
Gd-QD
bound
H2O
-1
1647 cm
-1
1593 cm
3150
2520
1890
1260
-1
Wavenumber (cm )
Chemical shift (ppm)
Figure 4. (a) Schematic representation of Gd3+ complex-capped Ag2S QDs (Gd-QD) obtained by functionalization of L-cysteine-
capped Ag2S QDs with Gd3+ complex (Gd-L); (b) FTIR spectra of L-cysteine-capped Ag2S QDs (blue), Gd-L (red) and Gd-QD (black);
1
(c) HNMR spectrum (400 MHz) of Gd-QD1 demonstrating the exchange of water protons between bound and bulk water of
surrounding.
synthesis, a similar trend in the shift of PL emission as a
function of the para substituent (9a and 9b) in selenourea
derivative (Figure 1c and 1d) was observed. In some cases,
L-cysteine in basic pH (Figure 3a). We chose L-cysteine due
to its small size and zwitterionic nature, which are
considered crucial for achieving compact size and low non-
specific binding in the biological set-up.61,62 The DDT-capped
such as 3i, 5a and 5b (Figure 1c and 1d), small additional peak
is observed in addition to the main PL emission, similar to
that reported for silver chalcogenides and are attributed to
the presence of surface defects.30 Ag2S and Ag2Se NCs
formed were further confirmed using X-ray photoelectron
spectroscopy (XPS) (Figure S11 and S13). The C 1s signal was
calibrated at 284.8 eV, prior to the assigning of other
observed peaks (Figure S9a and S9b). Consistent with the
literature report,55,56 the presence of two prominent peaks
at 368.9 eV and 374.9 eV in the Ag 3d core-level spectra of
Ag2S QDs (Figure S9c) are ascribed to Ag 3d5/2 and3d3/2
respectively. The spectrum of S 2p (Figure S11d) could be
deconvoluted to three individual components corresponding
to S-H (162.9 and 164.1 eV), S-C (161.9 and 163.1 eV) and S-
Ag (161.3 and 162.5 eV) as shown in the figure S12.57 The
individual peaks S2p1/2 and S2p3/2 corresponding to each
component are separated by a spin–orbit splitting of ~1.2
eV58 and have FWHM 1-1.2. The presence of S-C and S-H
binding energies, in addition to S-Ag peak explain the
observed atomic ratio (S/Ag) of 1.6: 1, which is on the higher
side. This is attributed to the presence of the DDT on the
surface of Ag2S QDs which is also confirmed by FTIR results
(Figure S7). Similar observation is also reported in the
literature.55
Ag2S QDs of two different sizes QD1 (size
~2.2 nm) and QD2
(size 4.2 nm) dispersed in chloroform showed a quantum
~
yield (QY) of about 6.9 % and 6.2 % respectively (measured
with respect to IR-140, the NIR-II standard dye as a
reference, Figure 3b). The purified DDT-capped Ag2S QDs
dispersed in chloroform was subjected to ligand exchange
process in the presence of L-cysteine dissolved in water at a
pH~
9 (cf. ESI; Experimental section).62 The slightly elevated
pH ensures deprotonation of thiol to thiolate. The pH of the
phase transfer reaction is important because the metal-
thiolate binding energy is higher than the metal-thiols.62 The
phase transfer was successful within 2 h yielding the L-
cysteine-capped Ag2S QDs in excellent yields (yield~93% and
90.0 % for QD1 and QD2 respectively) with a high
conservation of the colloidal stability and the quantum yield
(QY~5.5% and 5.6% for QD1 and QD2 respectively, Figure
3b). The FTIR spectrum of L-cysteine-capped Ag2S QDs is
depicted in Figure 3c. The FTIR spectrum of pure L-cysteine
shows a characteristic S-H stretching vibration signal at 2538
cm-1 (Figure 3c). The absence of this peak in aqueous Ag2S
QDs (Figure 3c) confirmed the absence of unbound L-
cysteine in solution. A prominent peak at 1572 cm-1 is
ascribed to the asymmetric stretching vibration of
carboxylate (COO–) group of surface-bound L-cysteine. For
free L-cysteine, this peak is observed at a slightly higher
The XPS spectra for Ag2Se QDs are shown in Figure S13.
These QDs have excellent colloidal stability in non-polar
solvents such as hexane and toluene. However, the practical
application of QDs for biomedical applications relies mostly
on their colloidal stability in the aqueous system with the
conservation of the optical properties.59–62 In the following,
we demonstrate the phase transfer and surface
functionalization of the as-prepared QDs. We substituted
hydrophobic surface ligands (DDT) with a hydrophilic ligand,
frequency (~
1588 cm-1).
Ag2S QDs offer NIR-II emission with improved signal-to-noise
ratio and functionalizing it with a T1-weighed MRI contrast
agent further improves its imaging efficiency and accuracy.
We synthesized the Gd3+ based complex (Gd-L, Figure S14a)
based on our earlier report16 (cf. ESI; Experimental section)
and subjected to the functionalization of the surface of L-
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 5
Please do not adjust margins

Please do not adjust margins
Dalton Transactions
Page 6 of 9
ARTICLE
Journal Name
cysteine capped Ag2S QDs to obtain an MRI/NIR-II
fluorescence contrast agent, Gd-QD (figure 4a). We
functionalized the surface of Ag2S QDs of two different sizes
values of Gd-QD1
/
Gd-QD2 with other MRI contrast agents
View Article Online
DOI: 10.1039/D0DT02974B
are listed in Table 1. The high longitudinal relaxivity values
for both Gd-QD1 and Gd-QD2 in comparison to other
nanoprobes suggest that Gd-QD1 and Gd-QD2 have a
promising potential as a bright MRI contrast agent in
bioimaging applications.
QD1 (Size~2.2 nm) and QD2 (Size~4.2 nm) with Gd-L to
obtain Gd-QD1 and Gd-QD2 respectively (cf. ESI;
Experimental section). The Gd3+ based complex is known for
its ability to lower the longitudinal relaxation (T1) of bound
water protons for improving localized signal intensity,63
resulting in the enhancement of the image contrast.64–73
Table 1. Longitudinal relaxivity values of Gd-QD1/Gd-QD1
and other nanoprobes
nanoprobe
Gd-QD1
Gd-QD2
longitudinal relaxivity (r)
The FTIR spectrum of Gd-L complex (Figure 4b) showed
peaks at 1676 cm-1 and 1610 cm-1 owing to the asymmetric
stretching vibration of COO– group and bending vibration of
N-H group respectively. The successful grafting of Gd-L on to
the Ag2S QDs surface, thereby forming Gd-QD is confirmed
by observation of asymmetric stretching vibration of COO–
r1 = 7.75 mM-1s-1
r1 = 8.27 mM-1s-1
r1 = 5.36 mM-1s-1
r1 = 4.34 mM-1s-1
r1 = 5.5–6.4 mM-1s-1
r1 = 6.9 mM-1s-1
Gd-L
Gadovist66
Gd-doped QDs67,74,78
Gd2O3 nanoparticles79
group at a lower frequency (~
1593 cm-1). The broad
absorption peaks at the range of 3000-3500 cm-1 are due to
O-H and N-H stretching vibrations.74 The formation of Gd-QD
conjugate is further confirmed from nuclear magnetic
resonance (NMR, 400 MHz) spectroscopy by monitoring the
change in chemical shift of water due to the presence of the
paramagnetic center. The difference in the chemical shifts of
bulk and bound water (∆δ) for pure ligand, Gd-L is 0.3917
ppm (Figure S14b). After functionalization with QD1 and
QD2, the distinct change in ∆δ was observed for both Gd-
QD1 (∆δ = 0.102, Figure 4c) and Gd-QD2 (∆δ = 0.0301, Figure
S14c) respectively. This result, taken together with FTIR
result where no free Gd-L was detected, unanimously
supports the successful grafting of Ag2S with Gd-L. Notably,
the smaller sized Gd-QD1 exhibited a smaller longitudinal
relaxivity (r1) of 7.78 mM-1s-1 per Gd-L and vice versa was
shown by larger sized Gd-QD2 (r1=8.27 mM-1s-1 per Gd-L).
This is an increase from Gd-L which on its own exhibits an r1
= 5.36 mM-1s (Figure S15 and S16). The higher relaxivity of
Gd-QD compared to Gd-L is due to the presence of quantum
dots which restricts free rotation in solution, therefore
increasing r1.16 The concentration of Ag2S QDs were
Conclusions
A new method for the synthesis of bio-compatible silver
chalcogenide QDs using easy-to-prepare substituted
thiourea, thiocarbamate, dithiocarbamates and selenourea
as S or Se precursors and DDT as capping agent is developed.
The emission wavelength can be tuned in the NIR-II region
by tuning reactivity of the precursor. The electronic effect of
substituents groups and atoms on the size is evident from
the blue or red shift in PL emission of the QDs consistent with
electron-withdrawing or donating abilities of the substituent
group. In the case of substituted phenyl containing
precursors, the final size of QDs fits linearly with Hammett
equation. The phase transfer of Ag2S QDs from chloroform
to aqueous phase was achieved by surface ligand exchange
of DDT with L-cysteine and the aqueous QDs were
subsequently functionalized with Gd3+ complex to form a
dual-modal contrast agent with high NIR-II quantum yield
(~
5-6 %) and MRI T1 water relaxivity (408-990 mM−1 s−1 per
QD) depending upon the size of the QDs.
determined at high photon energy (
extinction coefficient is independent of the size of QDs 75,76
We used molar extinction coefficient of 1.73
105 M-1 cm-1
~450 nm), where molar
.
Conflicts of interest
×
calculated using Ricard equation77 at 450 nm (The optical
constants used for calculation of molar extinction coefficient
were determined using DFT, cf. ESI; Determination of Optical
constants). At 400 MHz, the relaxivity per-QD for Gd-QD1
and Gd-QD2 was found to be 407.8 mM-1s-1 and 990.3 mM-
1s-1 respectively (cf. ESI; Relaxivity measurements). Thus, the
observed increase in relaxivity in Gd-QD2 compared to Gd-
QD1 can be attributed to both the increase in the size of the
nanocrystals which results in more restricted rotation in
solution and increase in Gd payload due to larger surface
area. In our study, we have demonstrated functionalization
of the size-tunable Ag2S QDs exhibiting fluorescence in
biologically important NIR-II diagnostic window. The
obtained values for relaxivity are much higher than the
smaller complexes and the commercially available MRI
contrast agents.66 The comparison of longitudinal relaxivity
The authors declare no competing financial interest.
Acknowledgements
ST, SP and KB acknowledge SERB-DST, Government of India
for
research
funding
(EEQ/2016/000751
and
EMR/2016/002505). SB would like to thank Department of
Science and Technology, Government of India
(DST/INSPIRE/03/2016/001207) [IF160689] for financial
support under DST-INSPIRE Scheme. AP acknowledges SERB-
DST
(EEQ/2016/000685)
and
DST-Inspire
(DST/
INSPIRE/04/2015/002674), for financial assistance. ST and
GJS thank the Royal Society (Grant IE160227) for finacial
assistance. GJS would like to thank the Medical Reseasrch
Council for funding (MR/T002573/1). AS acknowledges DST-
6 | J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 7 of 9
Dalton Transactions
Please do not adjust margins
Journal Name
ARTICLE
SERB (EEQ/2017/000319) for research funding. The authors
thank Debesh Sharma, Khanindram Baruah and Sangeeta
Saikia for initial help.
2013, 1, 6485–6494.
P. Zrazhevskiy, M. Sena and X. GDaOo,I:C1h0e.1m03.9S/oDc0.DRTe0v2.9, 74B
2010, 39, 4326–4354.
View Article Online
24
25
26
27
28
29
30
P. Jiang, C. N. Zhu, Z. L. Zhang, Z. Q. Tian and D. W.
Pang, Biomaterials, 2012, 33, 5130–5135.
G. Chen, F. Tian, Y. Zhang, Y. Zhang, C. Li and Q. Wang,
Adv. Funct. Mater., 2014, 24, 2481–2488.
B. Dong, C. Li, G. Chen, Y. Zhang, Y. Zhang, M. Deng and
Q. Wang, Chem. Mater., 2013, 25, 2503–2509.
D. He, S. Garg, Z. Wang, L. Li, H. Rong, X. Ma, G. Li, T. An
and T. D. Waite, Environ. Sci. Nano, 2019, 6, 1674–1687.
R. Gui, J. Sun, D. Liu, Y. Wang and H. Jin, Dalt. Trans.,
2014, 43, 16690–16697.
M. Yarema, S. Pichler, M. Sytnyk, R. Seyrkammer, R. T.
Lechner, G. Fritz-Popovski, D. Jarzab, K. Szendrei, R.
Resel, O. Korovyanko, M. A. Loi, O. Paris, G. Hesser and
W. Heiss, ACS Nano, 2011, 5, 3758–3765.
C. N. Zhu, P. Jiang, Z. L. Zhang, D. L. Zhu, Z. Q. Tian and
D. W. Pang, ACS Appl. Mater. Interfaces, 2013, 5, 1186–
1189.
H. Soo Choi, W. Liu, P. Misra, E. Tanaka, J. P. Zimmer, B.
Itty Ipe, M. G. Bawendi and J. V. Frangioni, Nat.
Biotechnol., 2007, 25, 1165–1170.
C. Lu, G. Chen, B. Yu and H. Cong, Adv. Eng. Mater.,
2018, 20, 1700940.
P. Reiss, M. Carrière, C. Lincheneau, L. Vaure and S.
Tamang, Chem. Rev., 2016, 116, 10731–10819.
I. Hocaoglu, M. N. Çizmeciyan, R. Erdem, C. Ozen, A.
Kurt, A. Sennaroglu and H. Y. Acar, J. Mater. Chem.,
2012, 22, 14674–14681.
Y. Zhang, Y. Liu, C. Li, X. Chen and Q. Wang, J. Phys.
Chem. C, 2014, 118, 4918–4923.
I. Hocaoglu, F. Demir, O. Birer, A. Kiraz, C. Sevrin, C.
Grandfils and H. Yagci Acar, Nanoscale, 2014, 6, 11921–
11931.
P. Jiang, Z. Q. Tian, C. N. Zhu, Z. L. Zhang and D. W.
Pang, Chem. Mater., 2012, 24, 3–5.
A. Sahu, L. Qi, M. S. Kang, D. Deng and D. J. Norris, J.
Am. Chem. Soc., 2011, 133, 6509–6512.
M. Han, X. Gao, J. Z. Su and S. Nie, Nat. Biotechnol.,
2001, 19, 631–635.
References
1
A. Zebibula, N. Alifu, L. Xia, C. Sun, X. Yu, D. Xue, L. Liu,
G. Li and J. Qian, Adv. Funct. Mater., 2018, 28, 1703451.
G. Hong, J. T. Robinson, Y. Zhang, S. Diao, A. L. Antaris,
Q. Wang and H. Dai, Angew. Chem. Int. Ed., 2012, 51,
9818–9821.
2
3
4
5
6
7
Y. Zhang, G. Hong, Y. Zhang, G. Chen, F. Li, H. Dai and Q.
Wang, ACS Nano, 2012, 6, 3695–3702.
Y. P. Gu, R. Cui, Z. L. Zhang, Z. X. Xie and D. W. Pang, J.
Am. Chem. Soc., 2012, 134, 79–82.
K. Welsher, S. P. Sherlock and H. Dai, Proc. Natl. Acad.
Sci. U. S. A., 2011, 108, 8943–8948.
J. T. Robinson, G. Hong, Y. Liang, B. Zhang, O. K. Yaghi
and H. Dai, J. Am. Chem. Soc., 2012, 134, 10664–10669.
G. Hong, J. C. Lee, J. T. Robinson, U. Raaz, L. Xie, N. F.
Huang, J. P. Cooke and H. Dai, Nat. Med., 2012, 18,
1841–1846.
31
32
8
Y. T. Lim, S. Kim, A. Nakayama, N. E. Stott, M. G.
Bawendi and J. V. Frangioni, Mol. Imaging, 2003, 2, 50–
64.
33
34
35
9
A. M. Smith, M. C. Mancini and S. Nie, Nat.
Nanotechnol., 2009, 4, 710–711.
10
S. Kim, Y. T. Lim, E. G. Soltesz, A. M. De Grand, J. Lee, A.
Nakayama, J. A. Parker, T. Mihaljevic, R. G. Laurence, D.
M. Dor, L. H. Cohn, M. G. Bawendi and J. V. Frangioni,
Nat. Biotechnol., 2004, 22, 93–97.
36
37
11
12
J. Li and J. J. Zhu, Analyst, 2013, 138, 2506–2515.
U. Resch-Genger, M. Grabolle, S. Cavaliere-Jaricot, R.
Nitschke and T. Nann, Nat. Methods, 2008, 5, 763–775.
S. J. Rosenthal, J. C. Chang, O. Kovtun, J. R. McBride and
I. D. Tomlinson, Chem. Biol., 2011, 18, 10–24.
J. Zhou, Y. Yang and C. Y. Zhang, Chem. Rev., 2015, 115,
11669–11717.
G. Wang, Z. Li and N. Ma, ACS Chem. Biol., 2018, 13,
1705–1713.
G. J. Stasiuk, S. Tamang, D. Imbert, C. Poillot, M.
Giardiello, C. Tisseyre, E. L. Barbier, P. H. Fries, M. De
Waard, P. Reiss and M. Mazzanti, ACS Nano, 2011, 5,
8193–8201.
13
14
15
16
38
39
40
41
42
P. Zrazhevskiy, L. D. True and X. Gao, Nat. Protoc., 2013,
8, 1852–1869.
W. J. M. Mulder, R. Koole, R. J. Brandwijk, G. Storm, P.
T. K. Chin, G. J. Strijkers, C. De Mello Donegá, K. Nicolay
and A. W. Griffioen, Nano Lett., 2006, 6, 1–6.
Y. Guo, S. R. Alvarado, J. D. Barclay and J. Vela, ACS
Nano, 2013, 7, 3616–3626.
T. P. A. Ruberu, H. R. Albright, B. Callis, B. Ward, J.
Cisneros, H. J. Fan and J. Vela, ACS Nano, 2012, 6, 5348–
5359.
M. P. Campos, M. P. Hendricks, A. N. Beecher, W.
Walravens, R. A. Swain, G. T. Cleveland, Z. Hens, M. Y.
Sfeir and J. S. Owen, J. Am. Chem. Soc., 2017, 139,
2296–2305.
M. P. Hendricks, M. P. Campos, G. T. Cleveland, I. J.-L.
Plante and J. S. Owen, Science, 2015, 348, 1226–1230.
17
18
19
H. Yang, S. Santra, G. A. Walter and P. H. Holloway, Adv.
Mater., 2006, 18, 2890–2894.
L. E. Jennings and N. J. Long, Chem. Commun., 2009,
3511–3524.
X. Michalet, F. F. Pinaud, L. A. Bentolila, J. M. Tsay, S.
Doose, J. J. Li, G. Sundaresan, A. M. Wu, S. S. Gambhir
and S. Weiss, Science, 2005, 307, 538–544.
S. Lee and X. Chen, Mol. Imaging, 2009, 8, 87–100.
Y. Jiao, Y. Sun, X. Tang, Q. Ren and W. Yang, Small,
2015, 11, 1962–1974.
43
44
45
46
20
21
22
23
X. Gao, Y. Cui, R. M. Levenson, L. W. K. Chung and S.
Nie, Nat. Biotechnol., 2004, 22, 969–976.
H. Sun, F. Zhang, H. Wei and B. Yang, J. Mater. Chem. B,
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 7
Please do not adjust margins

Please do not adjust margins
Dalton Transactions
Page 8 of 9
ARTICLE
Journal Name
47
48
49
P. Jiang and Z. Chen, New J. Chem., 2017, 41, 5707–
5712.
Stasiuk, J. Gallo and N. J. Long, Nanoscale,V2ie0w1A5rt,ic7le, Online
16119–16128.
C. Rivas, G. J. Stasiuk, M. Sae-Heng and N. J. Long, Dalt.
Trans., 2015, 44, 4976–4985.
G. J. Stasiuk and N. J. Long, Chem. Commun., 2013, 49,
2732–2746.
DOI: 10.1039/D0DT02974B
L. Jing, S. V. Kershaw, Y. Li, X. Huang, Y. Li, A. L. Rogach
and M. Gao, Chem. Rev., 2016, 116, 10623–10730.
A. Heuer-Jungemann, N. Feliu, I. Bakaimi, M. Hamaly, A.
Alkilany, I. Chakraborty, A. Masood, M. F. Casula, A.
Kostopoulou, E. Oh, K. Susumu, M. H. Stewart, I. L.
Medintz, E. Stratakis, W. J. Parak and A. G. Kanaras,
Chem. Rev., 2019, 119, 4819–4880.
L. Li and P. Reiss, J. Am. Chem. Soc., 2008, 130, 11588–
11589.
J. Jayabharathi, G. A. Sundari, V. Thanikachalam, P.
Jeeva and S. Panimozhi, RSC Adv., 2017, 7, 38923–
38934.
70
71
72
N. G. Chabloz, M. N. Wenzel, H. L. Perry, I. C. Yoon, S.
Molisso, G. J. Stasiuk, D. S. Elson, A. E. G. Cass and J. D.
E. T. Wilton-Ely, Chem. Eur. J., 2019, 25, 10895–10906.
G. J. Stasiuk, S. Tamang, D. Imbert, C. Gateau, P. Reiss,
P. Fries and M. Mazzanti, Dalt. Trans., 2013, 42, 8197–
8200.
H. Chen, L. Wang, H. Fu, Z. Wang, Y. Xie, Z. Zhang and Y.
Tang, J. Mater. Chem. B, 2016, 4, 7472–7480.
S. Tamang, S. Lee, H. Choi and S. Jeong, Chem. Mater.,
2016, 28, 8119–8122.
P. Yu, M. C. Beard, R. J. Ellingson, S. Fernere, C. Curtis, J.
Drexler, F. Luiszer and A. J. Nozik, J. Phys. Chem. B,
2005, 109, 7084–7087.
D. Ricard, M. Ghanassi and M. C. Schanne-Klein, Opt.
Commun., 1994, 108, 311–318.
H. Chen, G. D. Wang, W. Tang, T. Todd, Z. Zhen, C.
Tsang, K. Hekmatyar, T. Cowger, R. B. Hubbard, W.
Zhang, J. Stickney, B. Shen and J. Xie, Adv. Mater, 2014,
26, 6761–6766.
M. Ahrén, L. Selegård, F. Söderlind, M. Linares, J.
Kauczor, P. Norman, P. Käll and K. Uvdal, J. nanoparticle
Res., 2012, 14, 1–34.
50
51
73
74
75
76
52
53
54
E. J. Choi and S. B. Park, Org. Biomol. Chem., 2015, 13,
5202–5208.
Y. Du, B. Xu, T. Fu, M. Cai, F. Li, Y. Zhang and Q. Wang, J.
Am. Chem. Soc., 2010, 132, 1470–1471.
S. I. Sadovnikov, A. I. Gusev, A. V. Chukin and A. A.
Rempel, Phys. Chem. Chem. Phys., 2016, 18, 4617–
4626.
J. Xiang, H. Cao, Q. Wu, S. Zhang, X. Zhang and A. A. R.
Watt, J. Phys. Chem. C, 2008, 112, 3580–3584.
Q. Wu, M. Zhou, J. Shi, Q. Li, M. Yang and Z. Zhang,
Anal. Chem., 2017, 89, 6616–6623.
Q. Wu, M. Zhou, J. Shi, Q. Li, M. Yang and Z. Zhang,
Anal. Chem., 2017, 89, 6616–6623.
J. Van Embden and Y. Tachibana, J. Mater. Chem., 2012,
22, 11466–11469.
77
78
55
56
57
58
59
60
79
L. Tan, A. Wan, T. Zhao, R. Huang and H. Li, ACS Appl.
Mater. Interfaces, 2014, 6, 6217−6222.
B. Dubertret, P. Skourides, D. J. Norris, V. Noireaux, A.
H. Brivanlou and A. Libchaber, Science, 2012, 298,
1759–1762.
61
W. Liu, H. S. Choi, J. P. Zimmer, E. Tanaka, J. V Frangioni
and M. Bawendi, J. Am. Chem. Soc., 2007, 129, 14530–
14531.
62
63
64
S. Tamang, G. Beaune, I. Texier and P. Reiss, ACS Nano,
2011, 5, 9392–9402.
R. G. Aswathy, Y. Yoshida, T. Maekawa and D. S. Kumar,
Anal. Bioanal. Chem., 2010, 397, 1417–1435.
S. G. McAdams, D. J. Lewis, P. D. McNaughter, E. A.
Lewis, S. J. Haigh, P. O’Brien and F. Tuna, Chem.
Commun., 2017, 53, 10500–10503.
65
V. S. Marangoni, O. Neumann, L. Henderson, C. C.
Kaffes, H. Zhang, R. Zhang, S. Bishnoi, C. Ayala-Orozco,
V. Zucolotto, J. A. Bankson, P. Nordlander and N. J.
Halas, Proc. Natl. Acad. Sci., 2017, 114, 6960–6965.
P. Caravan, J. J. Ellison, T. J. McMurry and R. B. Lauffer,
Chem. Rev., 1999, 99, 2293–2352.
X. Ren, L. Liu, Y. Li, Q. Dai, M. Zhang and X. Jing, J.
Mater. Chem. B, 2014, 2, 5541–5549.
G. J. Stasiuk, F. Minuzzi, M. Sae-Heng, C. Rivas, H. P.
Juretschke, L. Piemonti, P. R. Allegrini, D. Laurent, A. R.
Duckworth, A. Beeby, G. A. Rutter and N. J. Long, Chem.
Eur. J., 2015, 21, 5023–5033.
66
67
68
69
N. A. Keasberry, M. Bañobre-López, C. Wood, G. J.
8 | J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 9 of 9
Dalton Transactions
View Article Online
DOI: 10.1039/D0DT02974B
84x52mm (300 x 300 DPI)