Synthesis of Au(I) complex-based aqueous colloids for sensing of biothiols

Article abstract of DOI:10.1016/j.ica.2018.10.006

The present work represents new facile route for synthesis of luminescent hydrophilic core-shell colloids, where the luminescence results from Au(I) complex (AuCl)₂L with cyclic PNNP ligand (L). The synthesis is based on solvent mediated aggregation of (AuCl)₂L in aqueous organic solutions. Instability of (AuCl)₂L in aqueous organic solutions was minimized by specific hydrophilic shell deposition arisen from electrostatically driven adsorption of polyethylenimine onto (AuCl)₂L-based cores. Luminescence of the colloids is stable for week at least due to restricted degradation of the luminescent complex at the interface and high positive exterior charge of the colloids. The sensing properties of the colloids towards thiols results from their complex formation with Au⁺ ions. The thiol-induced stripping of Au⁺ ions from the colloids quenches the luminescence. The optimal conditions for fluorescent recognition of cysteine, homocysteine and glutathione with lower detection limits about 1 μM are highlighted in the work.

Full text of DOI:10.1016/j.ica.2018.10.006

Accepted Manuscript
Research paper
Synthesis of Au(I) complex-based aqueous colloids for sensing of biothiols
Julia Elistratova, Bulat Faizullin, Nataliya Shamsutdinova, Aidar Gubaidullin,
Igor Strelnik, Vasily Babaev, Kirill Kholin, Irek Nizameev, Elvira Musina, Rafil
Khairullin, Andrey Karasik, Asiya Mustafina
PII:
S0020-1693(18)31393-8
https://doi.org/10.1016/j.ica.2018.10.006
ICA 18552
DOI:
Reference:
To appear in:
Inorganica Chimica Acta
Received Date:
Revised Date:
Accepted Date:
5 September 2018
3 October 2018
3 October 2018
Please cite this article as: J. Elistratova, B. Faizullin, N. Shamsutdinova, A. Gubaidullin, I. Strelnik, V. Babaev, K.
Kholin, I. Nizameev, E. Musina, R. Khairullin, A. Karasik, A. Mustafina, Synthesis of Au(I) complex-based aqueous
colloids for sensing of biothiols, Inorganica Chimica Acta (2018), doi: https://doi.org/10.1016/j.ica.2018.10.006
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Synthesis of Au(I) complex-based aqueous colloids for sensing of
biothiols.
a*
b
a
Julia Elistratova , Bulat Faizullin , Nataliya Shamsutdinova , Aidar
a
a
a
a
a
Gubaidullin , Igor Strelnik , Vasily Babaev , Kirill Kholin , Irek Nizameev ,
a
b
a
a
Elvira Musina , Rafil Khairullin , Andrey Karasik , Asiya Mustafina
Arbuzov Institute of Organic and Physical Chemistry, FRC Kazan Scientific
a
Center of RAS, Arbuzov str., 8, 420088, Kazan, Russia
b
Kazan (Volga region) Federal University, Kremlyovskaya str., 18, 420008,
Kazan, Russia
*
Corresponding author: 969_969@bk.ru
Abstract
The present work represents new facile route for synthesis of luminescent
hydrophilic core-shell colloids, where the luminescence results from Au(I)
complex (AuCl)
solvent mediated aggregation of (AuCl)
Instability of (AuCl) L in aqueous organic solutions was minimized by specific
hydrophilic shell deposition arisen from electrostatically driven adsorption of
polyethylenimine onto (AuCl) L-based cores. Luminescence of the colloids is
2
L with cyclic PNNP ligand (L). The synthesis is based on
2
L in aqueous organic solutions.
2
2
stable for week at least due to restricted degradation of the luminescent complex
at the interface and high positive exterior charge of the colloids. The sensing
properties of the colloids towards thiols results from their complex formation with
+
+
Au ions. The thiol-induced stripping of Au ions from the colloids quenches the
luminescence. The optimal conditions for fluorescent recognition of cysteine,
homocysteine and glutathione with lower detection limits about 1M are
highlighted in the work.
Keywords: Sensor Biothiols Luminescence Au(I) complex 1,5,3,7-
diazadiphosphacyclooctane
1
.Introduction
Heavy-metal complexes with d electron configuration have gained great
1
0
attention during recent decades due to their unique photophysical characteristics,
which makes them a good alternative to organic molecules in design of
1

nanosensors [1-3]. Unique electronic and spectral properties of Au(I) complexes
1
0
are prerequisite for their particular importance among other d -complexes in
sensing [4-8]. Solvent effect, interaction with substrates, phase transition or
mechanistic influence are highlighted in literature as the reasons for switching or
tuning of Au(I) complexes luminescence. Thus, unique sensing function of Au(I)
complexes has been widely exemplified by different sensing procedures for
recognition of vapors (vapochromism) and substrates in both organic and aqueous
solutions [1-5].
1
0
High complexing ability of d -ions towards biomolecules, including thiols
and biothiols can be considered as promising basis for their fluorescent
recognition procedures. Biothiols play crucial role in human physiology due to
their participation in biorelevant redox reactions. Cysteine and glutathione are
well-known antioxidants able to prevent damage to cellular components caused by
reactive oxygen species through redox transformations of the thiols to their
oxidized disulfide forms. A high level of homocysteine in the blood makes a
person more prone to endothelial cell injury, which leads to atherogenesis due to
inflammation in the blood vessels. Thus, recognition of low molecular weight
biothiols is very challenging task from biomedical point of view which has gained
growing attention during recent decades. The literature procedures are mostly
represented by organic dyes [6-11], gold or silver nanoparticles [12-19], which
tend to change their luminescence under interaction with thiols. The thiol-induced
luminescence response of both silver and gold nanoparticles commonly derives
from their aggregation or de-aggregation induced by interfacial coordination of
thiol molecules [12-19]. Since another biomolecules including peptides provide
the similar effect on aggregation of the nanoparticles [20], combination of the
nanoparticles with chemiluminescence-based assay was applied for more selective
recognition of biothiols [21]. Thus, development of new sensors on the basis of
Au(I) complexes is rather challenging task, although their wide application in
aqueous solutions is greatly restricted by both low solubility and/or stability of
such complexes in aqueous solutions. This problem can be solved by a synthesis
of water soluble Au(I) complexes with a sensing function. However, a conversion
of water insoluble complexes to hydrophilic nanosensors is worth noting as rather
promising facile alternate to synthesis of Au(I)-based water soluble sensors.
2

Thus, the main goal of the present work is the development of facile
synthetic route for conversion of the water insoluble Au(I) complex into selective
hydrophilic nanosensor for luminescent recognition of thiols and biothiols. The
recently represented Au(I) complex (AuCl) L with cyclic PNNP ligand (L) was
2
chosen as the subject of the present work due to its high stimuli responsive green
luminescence with both vapochromism in solid state and solvatochromism in
organic solutions [22, 23]. The report [23] highlights sensing properties of
(
AuCl) L in organic solutions resulted from the ability of the complex to bind
2
some alkylammonium cations or the solvent molecules in host-guest binding
mode.
The present report represents very facile one-pot synthetic procedure for
conversion of (AuCl)
efficient adsorption of PEI. The colloid stabilization is discussed in a correlation
with host-guest complex formation of (AuCl) L with ammonium groups of PEI.
2
L into aqueous hydrophilic colloids stabilized through an
2
Moreover, the performed measurements reveal PEI-stabilized colloids as efficient
nanosensors for low molecular weight thiols and biothiols versus thioethers.
2
. Experimental Section
2
.1 Materials
Sodium chloride NaCl, ascorbic acid, polyethylenimine (PEI branched,
MMaveraged = 25,000 Da) were purchased from Aldrich Chemistry, sodium
dihydrophosphate anhydrous (NaH PO , 99%) was purchased from ACROS
Organics, sodium ethylenediaminetetraacetate dihydrate (Na EDTA∙2H O, 99%),
2
4
2
2
L-cysteine, L-homocysteine, glutathione, DL-methionine, dithiothreitol,
Dulbecco’s Phosphate Buffered Saline (DPBS) and bovine serum albumin were
purchased from Sigma-Aldrich. Tris-(hydroxymethyl)-aminomethane extra pure
from Scharlau and MES hydrate 99% from ACROS Organics were also used as
the buffers.
Dimethyl sulfoxide (DMSO, 99.7%) purchased from Sigma-Aldrich was
used as solvent after the common purification by distillation.
Complex (AuCl)
,7-diphosphocyclooctane was synthesized according to the technique described
in [22].
2
L, where L is 1,5-bis (p-tolyl) -3,7-bis (pyridin-2-yl) -1,5-diaza-
3
3

Polyelectrolyte-stabilized (AuCl)
2
L-based colloids (PEI-(AuCl)
2
L) were
synthesized by drop-wise addition of (AuCl)
2
L (C=2mM) in DMSO solution (2
mL) by syringe pump (dropping speed of 0.35 mL/min) into aqueous solution (8
mL) of polyelectrolyte PEI (1 g/L) and NaCl (0.5 M) at pH 6.85 under the
vigorous stirring. These conditions favor the precipitation of (AuCl) L species and
2
their colloid stabilization by PEI adsorption. Obtained aqueous colloid was
subjected to ultrasonic dispersion for 30 minutes (the temperature of the water in
the ultrasonic bath, unless otherwise specified, was maintained at 22 ± 2° C).
Then colloid phase was separated from supernatant by the centrifugation (10,000
rpm for 15 minutes) in order to work off an excess of PEI and NaCl. After
separation of the supernatant deionized water was added to the precipitate to
adjust the concentration of (AuCl) L to 0.4 mM. This system was further
2
subjected to ultrasonic dispersion for 30 minutes.
For the luminescent titration of PEI-(AuCl)
2
L colloids the initial solutions
of substrates (ascorbic acid, L-cysteine, dithiothreitol, glutathione, DL-
methionine) (20 mM) were prepared.
2
.2 Methods
Dynamic light scattering (DLS) and electrokinetic potential
experiments were performed by means of the Malvern Mastersize 2000 particle
analyzer at 25 C. Electrokinetic potential values were calculated by the
Smoluchowski-Helmholtz equation [24]. Experimental autocorrelation functions
were analyzed with the Malvern DTS software and the second-order cumulant
expansion methods. The average error was ca. 4%. All samples were prepared in
deionized water filtered through PVDF membrane with a Syringe Filter (0.45
µm). All measurements were performed at least in triplicate at 25 °C.
pH-Measurements were carried out on a pH-meter of InoLab pH-720
with a SenTix Mic pH-electrode (WTW). The instrument was calibrated using
three standard buffer points (pH 4.01, 7.01 and 10.01).
Luminescence spectra. Luminescence spectra recorded on
a
spectrofluorometer FL3-221-NIR (Horiba Jobin Yvon) using 10 mm quartz
cuvettes. Excitation of samples was performed at 365 nm and emission was
detected at 400–650 nm, excitation and emission slits were 7 nm.
Transmission electron microscopy (TEM). TEM images were obtained
by use of Hitachi HT7700 (Japan) at an accelerating voltage of 100 kV.
4

Electrospray ionization mass spectrometry (ESI) was applied using an
AmaZon X ion trap mass spectrometer in positive mode.
X-ray powder diffraction (PXRD) measurements were performed on a Bruker
D8 Advance diffractometer equipped with a Vario attachment and Vantec linear
PSD, using Cu radiation (40 kV, 40 mA) monochromated by a curved Johansson
monochromator (λ Cu Kα1 1.5406 Å). Room-temperature data were collected in
the reflection mode with a flat-plate sample. Sample was applied in liquid form on
the surface of a standard zero diffraction silicon plate. After drying the layer, a
few more layers were applied on top of it to increase the total amount of the
sample. The sample was kept spinning (15 rpm) throughout the data collection.
o
o
o
Patterns were recorded in the 2Θ range between 3 and 105 , in 0.008 steps, with
a step time of 0.1–5.0s. Several diffraction patterns in various experimental modes
were collected for the samples. Processing of the data obtained was performed
using EVA [25] and TOPAS [26] software packages. The PDF-2 powder X-ray
diffraction database (ICDD PDF-2, Release 2011-2016) was used to identify the
crystalline phase.
Inductively Coupled Plasma Optical Emission Spectrometry (ICP-
OES). Au were identified in the colloids using simultaneous inductively coupled
plasma optical emission spectrometry (ICP-OES) model iCAP 6300 DUO by
Varian Thermo Scientific Company equipped with a CID detector. This
spectrometer enables the simultaneous measurement of peak heights within the
1
66 to 867 nm range. The optical resolution is less than 0.007 nm to 200 nm. The
working frequency is 27.12 MHz. Together, the radial and axial view
configurations enable optimal peak height measurements with suppressed spectral
noises. Concentration of Au ions was determined by the spectral line 242.795 nm.
3
. Results and discussion
3
.1. Synthesis of hydrophilic (AuCl) L-based colloids
2
Literature data introduce a plenty of synthetic routes for conversion of
water insoluble complexes into hydrophilic nanosized colloids. Commonly, they
are based on the well-known core-shell morphology, where water insoluble
5

complex species form the hard cores, which are coated by hydrophilic shell. A
choice of an optimal procedure for (AuCl) L is greatly affected by its
inconvenient stability in acidic and alkaline media. Thus, a hydrophilization of
AuCl) L via polymeric or silica coating is an inconvenient route due to the
2
(
2
complex degradation. The application of the solvent-mediated bottom-up
approach which results in formation of hard cores with their subsequent non-
covalent hydrophilic coating enables to obtain stable hydrophilic colloids without
any degradation of the complex. This procedure was previously successfully
applied for the efficient conversion of water insoluble lanthanide complexes into
hydrophilic colloids for sensing and imaging [27-30]. The adsorption of
polyelectrolytes onto charged hard cores provides very good opportunity for their
colloid stabilization.
As it has been above mentioned the water insolubility of the complex is the main
reason for its solvent-mediated conversion into hard templates for further
polyelectrolyte deposition. The previously reported synthetic route for the
conversion of water insoluble lanthanide complexes to polyelectrolyte-stabilized
nano-colloids is based on precipitation of water insoluble particles under a drop-
wise addition of the complex in organic solution to an aqueous solution of
polyelectrolyte [27-30]. Thus, the modification of the previously reported
procedure should be started from developing of convenient conditions for a
solvent-mediated precipitation of nanosized species of (AuCl)
2
L complex. The
drop-wise pouring of (AuCl) L complex in the organic (DMSO, DMF and
2
acetonitrile) solution to water results in the lack of precipitation. ESI
+
measurements reveal the appearance of [(Au
which reveals the dissociation of (AuCl)
2
Cl)L] in aqueous solution (Fig.1),
+
-
2
L to [(Au
2
Cl)L] and Cl (1) as the
reason for the dissolution of (AuCl)
organic solution.
2
L instead of its precipitation in aqueous
+
-
(
AuCl)
2
L= [(Au
2
Cl)L] + Cl
(1)
6

Fig. 1. ESI mass spectrum of the aqueous DMSO solution resulted from mixing
+
+
DMSO solution of (AuCl)
2
L with water: 1-[Au
2
ClL] , 2-[(AuCl) LNa] , 3-
2
+
[
(AuCl)
2
LK] .
-
Thus, excess amounts of Cl anions can shift the dissociation equilibrium (1)
towards water insoluble (AuCl) L. However, the peaks corresponding to
2
+
[
(AuCl)
2
LM] , where M=Na and K (Fig.1), point to efficiency of electrostatic
interactions of sodium and potassium ions with pre-organized P-Au-Cl moieties of
AuCl) L in accordance with Scheme 1. Therefore, host-guest complex formation
of (AuCl) L with sodium ions in NaCl solutions can provide additional
(
2
2
stabilization of the complex.
Scheme 1
Indeed, 0.5 M of NaCl is optimal to get detectable by a naked eye precipitation of
(
AuCl)
chloride. The optimized procedure (detailed description is in Exp. Section) is
based on the precipitation of (AuCl) L under mixing its DMSO solution with
2
L when its organic solution is mixed with aqueous solution of sodium
2
aqueous solution of sodium chloride and polyethylenimine (PEI) at pH 6.85,
where the latter provides a hydrophilic stabilization of the precipitated colloid
species. DMSO was chosen as organic solvent due to the significant solubility of
(
AuCl)
2
L in DMSO and miscibility of the latter with water. The use of acetonitrile
7

and DMF solutions in the synthetic procedure is also possible, although the use of
DMSO favors the stability of the hydrophilic colloids. The synthesized PEI-
stabilized colloids were separated from the supernatant in order to work off excess
amount of PEI and washed by water for several times. It is worth noting that the
synthesized hydrophilic colloids are easy to manipulate by centrifugation-assisted
phase separation for washing and controlling the colloids concentration.
Electrokinetic potential and dynamic light scattering measurements of the
synthesized colloids were performed after their redispersion in water. The results
provide clear indication of the positively charged exterior layer of PEI-stabilized
(
(
AuCl)
AuCl)
2
2
L-based colloids (Fig. 2), which will be further designated as PEI-
L. The measured electrokinetic potential of precipitated (AuCl) L-based
2
colloid species in the saline aqueous DMSO solution is negative (Fig. 2).
Fig. 2. Size distribution by volume in saline aqueous DMSO solutions of
AuCl) L (1) and in PEI-(AuCl) L measured by DLS along with electrokinetic
potential values (, mV) designated at the top of the peaks.
(
2
2
The ability of (AuCl)
interfacial adsorption of PEI onto the precipitated species, where ammonium
groups of PEI can be bound with Au-P-Cl moieties at the interface of (AuCl) L-
L and
2
L molecule to bind cations explains the efficiency of the
2
based cores. Scheme 1 illustrates the binding mode between (AuCl)
ammonium groups of PEI.
2
8

Fig. 3. Emission spectra of PEI-(AuCl) L (0.4 mM) colloids measured: (a) exactly
2
after their preparation (1) and under their storage within 2 (2), 4 (3) and 9 (4)
days; (b) before (1) and after addition of various amounts of L-cysteine, ex=365
nm.
Taking into account that PEI solutions are highly alkaline the adjusting of pH to
neutral range (5.6-6.0) is the important prerequisite of the successful synthetic
procedure. Analysis of the supernatant solutions reveal very small concentration
of Au (for more details see Suppl. Mat.). The losses of the complex during its
conversion to the colloids is about 1%, which confirms the efficiency of (AuCl)
precipitation in the applied synthetic conditions. The emission spectrum of the
colloids similar with that of (AuCl) L in DMSO solutions possesses one widened
2
L
2
band with the maximum wavelength at about 505 nm under the excitation at 365
nm (Fig. 3 and Fig.S1 in Suppl. Mat.). The emission band arises from the Au(I)-
disturbed ligand-centered luminescence [23]. Dynamic light scattering
measurements reveal the averaged size at 380 nm with rather wide size
distribution in PEI-(AuCl)
2
L colloids, while TEM images of the dried samples
L. Such
(
Fig.4) reveal ultra-small sizes of the hard templates for PEI-(AuCl)
2
deviation between the size values revealed by DLS and TEM methods derives
from the difference of the applied techniques. In particular, TEM images of the
dried samples reveal the size of the hard (AuCl) L-based templates, while DLS
2
data indicate the size of the hydrated polyelectrolyte layer. The aggregation
behavior of the colloids is worth noting as another factor influencing the size
values revealed from the DLS measurements. This tendency is rather anticipated
for the colloids synthesized in such saline conditions (0.5 M of NaCl), because the
aggregation behavior of polyelectrolyte-stabilized colloids is greatly assisted by
an exterior charge neutralization due to a counter-ion binding [31].
9

Fig. 4. TEM image of dried PEI-(AuCl)
2
L colloids.
0
3+
The disproportionation of (AuCl)
organic solutions of (AuCl)
results in the degradation of (AuCl)
2
L to Au and Au was previously revealed for
2
L under their irradiation by sunlight [23], which
L emission within few hours in organic
2
solutions. It is worth noting that the luminescence of the synthesized aqueous
colloids remains unchanged for nine days at least (Fig.3a). Thus, conversion of
DMSO-based solutions of (AuCl)
complex stability.
2
L to PEI-(AuCl) L colloids increases the
2
PXRD data of the dried samples of PEI-(AuCl) L reveal a crystallinity of the hard
2
cores inside the polyelectrolyte coating, although the crystal structure of the
colloids differs from that in the crystals obtained from DMSO solutions of
(
AuCl)
nucleophilic binding site of (AuCl)
moieties as the main reason for formation of solvato-complexes in DMSO
solutions of (AuCl) L [23]. The bound DMSO molecules can be easily substituted
by sodium ions or alkylammonium groups of PEI molecule in the aqueous PEI-
AuCl) L colloids (Scheme 1). The previously reported X-ray results [22, 23]
indicate that the host-guest binding of (AuCl) L with solvent molecules
significantly affects the crystal packing in (AuCl) L-based solvates. Fig. S2 in
Suppl. Mat. illustrates the similarity in the PXRD patterns for the isostructural
crystal solvates of (AuCl) L with DMSO and acetone.
2
L (Figure 5). Our previous report is worth noting for highlighting the
2
L derived from two pre-organized P-Au-Cl
2
(
2
2
2
2
1
0

Fig. 5. Experimental diffraction pattern of the dried PEI-[(AuCl) L] colloids (1),
2
and calculated diffraction patterns of ligand L (2) and the complex grown from
DMSO (3). Blue vertical lines show the position of the interference peaks
corresponding to a crystalline NaCl, Ref.№ 01-070-2509 in the PDF-2 database.
Thus, the host-guest binding of cations or cationic moieties of polyelectrolyte with
(
AuCl)
specific crystal packing of the complex within nanosized colloid species. For this
reason, the diffraction pattern of dried PEI-(AuCl) L was examined and compared
with that of (AuCl) L∙2DMSO, which is the crystal solvate of (AuCl) L with
DMSO (Figure 5). The presence of the peaks deriving from crystalline NaCl in
the dried sample of PEI-(AuCl) L is rather anticipated due to the saline synthetic
conditions. These peaks do not interfere with the diffraction pattern of (AuCl) L,
which is quite different from the pattern of (AuCl) L∙2DMSO calculated from the
2
L molecules at the interface of the colloid species can be the reason for
2
2
2
2
2
2
single crystal X-ray data. This result indirectly confirms the substitution of DMSO
+
molecule from the nucleophilic binding site of (AuCl)
2
L by cations (Na or
ammonium groups of PEI). The substitution is schematically illustrated by
Scheme 1.
3
.2. Substrate-induced luminescence response
1
1

Fig. 6. I/I
0
values at 505 nm (λ =365 nm) of PEI-(AuCl) L colloids (0.4 mM)
2
ex
versus concentration of substrates: G-SH (1); DTT (2); Cys (3); Hcy (4) where I
0
and I refers to intensity of the colloids before and after the substrate addition
correspondingly. The data in wide concentration range without adjusting pH value
(
a); the same data in the narrow concentration range (b); the data, when pH was
adjusted by MES to 5.6 (c) and 6.8 (d), where standard deviations are shown as
standard bars.
The discussion of thiol-mediated luminescence response of PEI-(AuCl) L colloids
2
should be preceded by the data revealing another factors, such as pH, buffer or
some well-known chelating anions influencing the luminescence of the colloids.
The obtained results indicate that the luminescence of PEI-(AuCl)
unchanged within pH 5.0-7.5 (Fig.S3, Suppl. Mat.) and in wide concentration
range of Na EDTA (Fig.S4, Suppl. Mat.). These data indicate the lack of the
2
L remains
2
stripping effect of ethylenediaminetetraacetate anions. Moreover, the presence of
ascorbate anions which are well-known reducing agents in synthesis of gold
nanoparticles results in rather small quenching effect on the emission of PEI-
(
AuCl) L instead of their whole degradation (Fig.S4, Suppl. Mat.).
2
Stability of the luminescence of PEI-(AuCl)
2
L colloids in blood serum is
important prerequisite for their bioanalytical application. Buffer (DPBS) solutions
1
2

of bovine serum albumin (BSA) are commonly applied as a simplest model of
blood plasma. It is worth noting that the luminescence of PEI-(AuCl) L remains
2
unchanged in DPBS-based solution of BSA (Fig.S5, Suppl. Mat.). Cysteine
residues are worth noting as common component of the proteins, although they
are regarded as the least exposed residue in proteins due to great extent of burial
and clustering of the cysteine residues within proteins [32]. The lack of the
luminescent response on BSA is in good agreement with the structure of BSA.
Moreover, this result argues for the applicability of PEI-(AuCl) L in bioanalysis.
2
Both reducing and complexing ability of thiols should induce a stripping of Au(I)
ions from PEI-(AuCl) L nanoparticles. Indeed, the increase in the concentration of
2
thiols exemplified by cysteine (Cys), homocysteine (Hcy), glutathione (G-SH)
and dithiothreitol (DTT) result in the concentration-dependent quenching of the
luminescence in PEI-(AuCl)
2
L colloids (Fig. 6 a,b). The plotting of the quenching
, where I and I are the colloids luminescence
data as relative intensity (I/I
0
0
intensity before and after addition of thiols) versus thiol concentration (Fig.6a)
indicates that the quenching extent dramatically depends on the structure of thiol.
In particular, the thiol-induced quenching of PEI-(AuCl) L luminescence tends to
2
increase in the following sequence G-SH  DTT  Cys (Fig.6a). The
measurements represented in Figs. 6 a,b were done in nonbuffered conditions in
order to reveal the acidification of the solution resulted from the thiol-induced
quenching. Indeed, pH is about 5.6 in the initial colloid solution, while tends to
continuously decrease to 3.6-4.0 at 200 M of the thiols (for more details see
Table S1, Suppl. Mat.). Thus, the quenching derives from the process, which can
be schematically represented by equilibrium (2), followed by further
+
-
transformation of AuSR due to high redox activity of Au and RS . The
0
equilibrium (3) producing Au and the disulfide can be assumed as the result of
the transformation.
(
AuCl)
2
L + 2 RSH = 2AuSR + L + 2HCl
(2)
(3)
0
2
AuSR = 2 Au +RS-SR
The appearance of emission at 420 nm as the additional shoulder at the wide
emission band of (AuCl) L in PEI-(AuCl) L colloids after the addition of the
thiols (Fig. S6, Suppl. Mat.) can be explained by surface plasmon resonance
2
2
1
3

0
emission band arisen from the formation of Au nanoparticles in the thiol-
containing solutions. The solution acidification confirms the protons deliverance
due to equilibrium (2) and shifts the equilibrium (2) to the left. Thus, pH of the
solutions of PEI-(AuCl) L was adjusted to various levels in order to affect the
2
quenching effect of the thiols, since the pH is the reason for shifting of the
equilibrium (2). The shifting, in turn, is the reason for influence on the
luminescent response to the biothiols.
Indeed, the quenching effect of the thiols increased when the measurements were
performed in the buffer solutions at pH 5.6 and 6.8 (Figs. 6 c,d). pH 6.8 provides
optimal conditions for the greatest deviation between concentration dependencies
of I/I for G-SH, Cys and DTT, while the quenching effects induced by Cys and
0
Hcy are very similar. The greatest quenching effect of DTT (Fig.6 d) is in good
agreement with its enhanced complexing ability versus Cys and G-SH due to the
presence of two SH groups [33-36]. The insignificant effect of methionine
possessing C-S-CH moiety instead of SH group on the colloids emission (Fig.S4,
3
Suppl. Mat.) points to the impact of the latter on the quenching effect of the thiols.
This result along with the small quenching effect of ascorbate anions (Fig. S4 in
+
Suppl. Mat.) confirms that the complex formation of the thiols with Au ions due
to equilibrium (2) is the reason for their stripping from the colloids. It is also
worth noting that Cys and Hcy mediate more enhanced quenching than G-SH.
The core-shell nano-architecture of PEI-(AuCl)
based exterior layer, is worth noting as the factor affecting the fluorescence
response of (AuCl) L on the tiols. It is in particular well-known that a charge of
exterior layer commonly plays a crucial role in a luminescent response of core-
shell colloids [27-29]. Thus, PEI-based adlayer of PEI-(AuCl) L is worth noting
2
L, in particular the PEI-
2
2
as a factor favoring the luminescence response on anions versus cations, although
the heterogeneous nature of the colloids can be a reason for a time-dependent
luminescent response. The luminescence response of PEI-(AuCl) L colloids was
2
measured at different time duration after the sample preparation. The obtained
data (Fig.S7a,b in Suppl. Mat.) indicate that the quenching effect of Cys and G-
SH tend to increase within ten minutes. Nevertheless, the immediate (within one
minute) measurements provide better linearity of I/I
0
versus concentration of Cys,
Hcy and G-SH at pH 6.8 (Fig.6d) than the similar dependence on the basis of the
1
4

measurements performed within ten minutes after the sample preparation (Fig.6,
Fig.S7c in Suppl. Mat.).
The represented data are enough to reveal the origin and regularities of the thiol-
mediated luminescence response of PEI-(AuCl)
2
L, while the detection conditions
should be optimized for the lower detection level of the thiols.
1
0
0
0
0
0
,0
,8
,6
,4
,2
,0
2
2
R (1)=0.998
2
1
3
R (2)=0.978
2
R (3)=0.982
0
5
10
15
,M
20
C
Substrate
Fig. 7. Linear relationships between I/I
0
at 505 nm (λ =365 nm) of PEI-
ex
(
(
AuCl)
2
L colloids (0.04 mM) and thiols concentration: Cys (1); G-SH (2); Hcy
2
3). Coefficient of determination (R ) is presented in the plot, slope (b) and
intercept (a) values are presented in Table S2 in Suppl. Mat.
Figure 7 represents the luminescence response of PEI-(AuCl)
2
L on Cys, Hcy and
G-SH in the narrow concentration range 0-20 M of the thiols. The results (Fig.7)
indicate no significant difference in the quenching effect of Cys, Hcy and G-SH
within 0-5 M of the thiols. Deviation between the I/I values plotted versus Cys
0
and G-SH concentration tends to increase within 5-20 M, which results in better
linearity of the plot for Cys versus G-SH (Fig.7, Table 1). Taking into account
that relative standard deviation of I/I values lies within 1-1.5 % the lower
0
detection limit of Cys, Hcy and G-SH is about 1 M (Table 1). Table 1 represents
the lower detection (LD) and linear range (LR) values both estimated in the
present work and taken from the literature data. The obtained results (Table 1)
highlight PEI-(AuCl) L colloids as good (not the best) sensors on the biothiols
2
without significant discrimination between them within the 0-5 M concentration
range.
1
5

It is also worth noting that high affinity of G-SH to PEI-(AuCl) L makes the latter
2
very promising basis for affecting the so-called ROS level in living cells due to
the well-known impact of G-SH on the maintaining optimal ROS level [37]. This
task lies out of the present work scope but will be studied in a nearest future and
represented elsewhere.
Table 1. Comparison of various sensors for biothiols detection using fluorescence
technique: LD and LR values calculated from data obtained in the present work
and taken from literature data.
Materials used
Fe quantum
Thiols
LR
LD
Ref.
clusters incorporated into Hb/Zn(II)
complex
Cys
1-20M
0.25M
[38]
Triethylenetetramine modified
CDs/Cu(II) complex
G-SH
0.2-175M
0-11M
0.11M
[39]
[40]
Cys
Hcy
86nM
86nM
N,S- doped CDs loaded with Cu(II)
G-SH
G-SH
Cys
86nM
30nM
N-doped CDs
0.2-1000M
0.1-0.6M
0.1-0.5M
[41]
[42]
1.59nM
2.88nM
Single-stranded DNA stabilized Ag
2
+
NCs/Hg system
G-SH
HSA-stabilized Au/Ag
core/shell NCs
Cys and Hcy
(total)
0
.1-5M
15nM
[43]
[44]
Graphene QDs coupled to Au NPs
Cys
Cys
1-4M
0.02-80M
2-70M
0.32nM
5.87nM
1.01M
0.51nM
Au@Ag nanoclusters
[16]
[45]
[46]
[47]
G-SH
Au NPs decorated reduced graphene
oxide nanohybrid
Cys
Cys
0-3nM
+
GSH-Au NCs/ Ag system
0-2.71M
0.01-5M
16.54nM
3.6nM
N,S- doped CDs/AuNPs
G-SH
Cys
Hcy
0-20M
0-20M
0-20M
1M
1M
1M
This
work
2
PEI-[(AuCl) L] colloids
G-SH
Conclusions
The present work for the first time highlights facile synthetic approach for
conversion of luminescent Au(I) complex (AuCl) L with poor stability in
solutions into stable in time hydrophilic colloids as the nanosensors for biothiols.
2
1
6

Dissociation of (AuCl)
deposition of PEI on (AuCl)
an ionic background are the factors restricting the dissociation of (AuCl)
is the reason for significant stability of (AuCl) L in the synthesized core-shell
colloids. In particular, the luminescence of the colloids arisen from the emission
of (AuCl) L remains unchanged within one week at least. No changes in the
2
L is one of the reasons for its instability in solutions. Both
L-based nanoparticles and the presence of NaCl as
L. This
2
2
2
2
luminescence is observed in the solutions of ethylenediaminetetraacetate and
ascorbate anions, as well as in saline buffer solutions of bovine serum albumin.
The concentration- and pH-dependent quenching of PEI-(AuCl)
revealed in the solutions of thiols and biothiols. The stripping effect of SH groups
plays a key role in the luminescence response of PEI-(AuCl) L. The present work
introduces the recognition of cysteine, homocysteine and glutathione versus
methionine by luminescent response of PEI-(AuCl) L which highlights the
2
L emission is
2
2
synthesized colloids as promising for bioanalytical and biomedical purposes.
Compliance with ethical standards
The author(s) declare that they have no competing interests.
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[
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Synthesis of aqueous core-shell luminescent colloids for sensing of thiols
Au(I) complex as basis for hard cores coated by soft polyelectrolyte shell
Specific interactions at core/shell interface provide stability of colloids
pH-dependent quenching effects of thiols on colloids luminescence
+
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