Hydrophilicity and Microsolvation of an Organic Molecule Resolved on the Sub-molecular Level by Scanning Tunneling Microscopy

Article abstract of DOI:10.1002/anie.201711062

Low-temperature scanning tunneling microscopy was used to follow the formation of a solvation shell around an adsorbed functionalized azo dye from the attachment of the first water molecule to a fully solvated molecule. Specific functional groups bind initially one water molecule each, which act as anchor points for additional water molecules. Further water attachment occurs in areas close to these functional groups even when the functional groups themselves are already saturated. In contrast, water molecules surround the hydrophobic parts of the molecule only when the two-dimensional solvation shell closes around them. This study thus traces hydrophilic and hydrophobic properties of an organic molecule down to a sub-molecular length scale.

Full text of DOI:10.1002/anie.201711062

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Accepted Article
Title: Hydrophilicity and microsolvation of an organic molecule resolved
on the sub-molecular level by scanning tunneling microscopy
Authors: Karsten Lucht, Dirk Loose, Maximilian Ruschmeier, Valerie
Strotkötter, Gerald Dyker, and Karina Morgenstern
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To be cited as: Angew. Chem. Int. Ed. 10.1002/anie.201711062
Angew. Chem. 10.1002/ange.201711062
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Hydrophilicity and microsolvation of an organic molecule
resolved on the sub-molecular level by scanning tunneling
microscopy
Karsten Lucht, Dirk Loose, Maximilian Ruschmeier, Valerie Strotkötter, Gerald Dyker, and Karina
Morgenstern*
Abstract: We used low-temperature scanning tunneling microscopy
to follow the formation of a solvation shell around an adsorbed
functionalized azo dye from the attachment of the first water
molecule to a fully solvated molecule. Specific functional groups bind
initially one water molecule each, which act as anchor points for
additional water molecules. Further water attachment occurs in
areas close to these functional groups even when the functional
groups themselves are already saturated. In contrast, water
molecules surround the hydrophobic parts of the molecule only,
when the two-dimensional solvation shell closes around them. Our
study thus traces hydrophilic and hydrophobic properties of an
organic molecule down to a sub-molecular length scale.
Solute-solvent interactions play an important role in chemistry.
Chemical reactions are highly sensitive towards changes in the
[
1,2]
solvation environment.
The most abundant solvent on our
Figure 1. NPAS molecules on Au(111): a) STM overview image, square
marks region shown in higher resolution in b). b) False color STM image of
single NPAS molecule. Light blue represents the Au(111) surface, the NPAS
molecule is bright pink. Contrast in lower right of image is enhanced; gas-
phase structure of NPAS molecule at same scale calculated semi-empirically
planet is water. Therefore, it is of utmost significance to
investigate the interactions between water and the substances it
solvates. This will undoubtedly bring progress to many scientific
fields and industrial applications. Corrosion science, drug
delivery or polymer synthesis might benefit from an in-depth
understanding of solvation. However, imaging single molecules
[3]
[
4]
[5]
[
29]
using the Parametric Method 3 (PM3)-parametrized MNDO Hamiltonian.
Dotted lines guide the eye. c) Line profile along solid line in b). Red lines
represent Gaussian fits to individual peaks. Blue curve: sum of the three
Gaussian fits. Numbers (1-3) in graph correspond to features in b). Dotted
lines: peak positions of Gaussian fits. d) Map of electrostatic potential of NPAS
with their solvation shells is
a challenging task. Current
experimental techniques cannot image individual solvated
molecules in their natural three-dimensional solvation
environment. Scanning probe microscopy techniques have the
required high resolution but need to support the molecules on a
surface. Adapting atomic force microscopy (AFM) to the liquid
environment offered the fascinating possibility to image the
[
29]
molecule at a contour value of 0.005 e/B³ calculated semi-empirically.
Charge is given in fractions of elementary charge.
[
13,14]
the spectroscopy of water doped helium nanodroplets.
However, the geometrical arrangement of the resulting solvation
[
6-8]
three-dimensional solvation structure of polar surfaces.
This
[
15-17]
technique obtained atomic resolution in the liquid, delivered the
spatial distribution of water at interfaces, and generated three-
dimensional solvation maps showing individual layers for
shells for e.g. for the chloride ion is only known theoretically.
Solvation of organic molecules is even more complex because
their spatial extend as well as the varying charge density of
functional groups need to be considered. Consequently, they
offer more potential attachment sites, which complicate the
interpretation of spectra and increase the computational effort.
Scanning tunneling microscopy performed at UHV conditions
and low temperature is a valuable tool to address this issue.
Although these conditions are not ambient, they offer the unique
opportunity to observe individual solute-solvent interactions by
freezing out molecular mobility existing under ambient
[
9]
[10]
inorganic surfaces as CaF
2
, Mica
and organic surfaces as
[
11,12]
phospholipid bilayers.
Because of the large interaction of the
AFM tip during the 3D-imaging procedure and the intrinsic
mobility of individual organic molecules of low mass in liquid
environment, it seems unlikely that this method can be extended
to the solvation of individual organic molecules. Recently,
detailed insight into the microsolvation environment of ions and
small organic molecules was gained by ab-initio calculations and
[
18]
conditions. Organic molecules
molecules,
as well as individual water
[
19]
[20]
and large water structures
were imaged by
[*]
K. Lucht, M. Ruschmeier, V. Strotkötter, Prof. Dr. K. Morgenstern
Physikalische Chemie I
Ruhr-Universität Bochum
Universitätsstraße 150, D 44801 Bochum, Germany
E-mail: karina.morgenstern@rub.de
scanning tunneling microscopy with (sub-)molecular resolution.
While water structures as well as organic molecules were
characterized separately, a thorough investigation of structures
resulting from the combination of the two is still lacking. In a
recent study, we co-adsorbed 4,4‘-dihydroxy-azobenzene with
D. Loose, Prof. Dr. G. Dyker
Organische Chemie II
Ruhr-Universität Bochum
Universitätsstraße 150, D 44801 Bochum, Germany
[21]
small amounts of water. The water molecules were found to
attach exclusively to the hydroxyl groups, but as they are
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identical no conclusion about relative hydrophilicity of the
[
19]
endgroups could be drawn. Clusters of water
molecules
and organic
[
22-24]
are often dominated by intermolecular bonds and,
it was shown that these interactions are only marginally
influenced by the presence of the Au(111) surface. Therefore,
our approach towards a molecular-scale picture of solvation is to
adsorb solute (an azobenzene derivate) and solvent (water) onto
Au(111) at low temperature and to image the formed structures
using low-temperature scanning tunneling microscopy. In this
article, the functionalized azo dye 5-(4-Nitrophenylazo)salicylic
acid (hereafter designated NPAS) with groups of different water
affinity (carboxyl, hydroxyl, and nitro group) offers a diversified
binding environment to water molecules. Our study shows which
functional groups are favored over others, it reveals their relative
hydrophilicity and how different functional groups affect the
formation of a solvation shell on a single molecule basis. The
hydroxyl group is placed in ortho position with respect to the
carboxyl group, because of the resulting intramolecular
hydrogen bond. Theory predicted that this bond lowers the
propensity to hydrogen bond formation of water molecules
[
25]
towards the hydroxyl group which we confirm experimentally.
Because of the predominance of solute-solvent interaction on
the low-interacting gold surface, our results are relevant for the
microscopic understanding of solvation also in other
environments.
Figure 2. Single water molecules attached to NPAS. a), b) STM images and
[29]
semi-empirical gas-phase structures of the NPAS/water complexes to scale
with STM images. a) Water molecule attached to carboxyl group next to an
isolated water molecule. b) Water molecule attached to nitro group. c) and d):
Line profiles along solid lines in a) and b). Red and dark blue curves:
2
Gaussian fits to NPAS molecules and D O molecules, respectively. Light blue
Results and discussion
curve: sum of all fits. Inter-peak distances of Gaussian fits represented by
dashed lines.
Geometrical characterization of solute and solvent
To set the stage, we characterize STM images of individual
NPAS molecules. The NPAS molecule is a functionalized
azobenzene composed of salicylic acid and nitro benzene
subunits that are linked via a diazo bridge (Fig. 1b, ball-and-stick
model). At the adsorption temperature of 30 K NPAS molecules
are mobile and agglomerate at the elbows of the herringbone
reconstruction (Fig. 1a). In order to ascertain that intermolecular
interactions do not influence our results, only single molecules in
between elbows are considered in this study, as marked in
Fig 1a. The STM image of a single NPAS molecule consists of
two protrusions (marked by 1 and 2) and a shallow depression
range influence of the other end-group.
First steps of NPAS solvation
Having identified the solute we now co-adsorb NPAS and water
molecules to reveal the preferred coordination sites of water
molecules at NPAS molecules. The overall appearance of the
NPAS molecules is not altered by the attachment of single water
molecules, which add a coordination site dependent feature (Fig.
2a,b). Water molecules attached to the carboxyl group, cause a
further elongation of the already ellipsoidal protrusion (Fig. 2a).
Coordination to the nitro group adds a round protrusion along
the molecular axis (Fig. 2b). These two binding motifs make up
(
marked by 3). Distances between them are determined through
the analysis of line profiles along the molecular axis (solid line in
Fig. 1b), which are fitted by Gaussian profiles (Fig. 1c). The
90 % of all measured NPAS/water complexes at small
D
2
O:NPAS ratios (Ntot = 261). Coordination to the carboxyl group
distance between protrusion
1
and
2
amounts to
occurs in 53 % of imaged NPAS/water complexes, while
coordination to the nitro group makes up 37 %. The lower affinity
of the nitro group towards water compared to the carboxyl group
may be attributed to the rather hydroneutral properties of the
(
0.80 ± 0.03) nm. It is evident that protrusion 1 is slightly ellipsoid
and oriented at an angle from the main molecular axis. In order
to assign this protrusion to a specific part of the molecule, we
compare the image to an ESP map, in which regions of large
negative charge density correspond to large local-density of
[27]
nitro group. The spreading of the negative partial charge over
the two oxygen atoms of the nitro group prevents the formation
of strong hydrogen bonds. However, it is still more hydrophilic
than the phenyl rings or the azo group, which did not bind single
water molecules in any of the imaged NPAS/water complexes.
In the following we characterize the two predominant binding
motifs geometrically.
Compared to Fig. 1b the ellipsoidal appearance of the carboxyl
group is clearly more prominent (Fig. 2a). A line profile over the
long axis of the ellipsoidal protrusion yields two Gaussians with
a separation of (0.66 ± 0.05) nm (Fig. 2c). The Gaussian fit of
the smaller protrusion closely matches the one obtained from
[
26]
states in an STM image.
ESP) map exhibits an elongation in the region of the carboxyl
group, which is caused by an extension of the benzene ring’s
-system (Fig. 1d). Hence, we conclude that the carboxyl group
Indeed, the electrostatic potential
(

of the salicylic acid subunit causes the observed bulge of
electron density. Therefore, the shallow depression of 20 pm (at
5
0 mV bias voltage) at the other end of the molecule is caused
by the nitro group. Its shallow depth at a large distance of
0.6 ± 0.06) nm from protrusion 2 suggests that the molecule
(
changes the electronic structure of the neighboring substrate
atoms (see dotted lines in Fig. 1b). Surprisingly, such a
depression was not observed in previous studies of nitro
isolated water molecule, corroborating that
molecule is attached to the NPAS molecule. Interestingly, we
2
a single D O
[
26]
functionalized azo dyes or phenyl rings and indicates a long-
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never observed a single water molecule that binds to the
hydroxyl group before the carboxyl group is occupied. This can
be understood in view of a recent DFT study on the solvation of
salicylic acid by water-modified supercritical carbon dioxide that
showed that the hydroxyl group forms an intramolecular
hydrogen bond to the carbonyl oxygen atom of the carboxyl
[
25]
group.
This study suggests that this intramolecular hydrogen
bond causes the hydroxyl group to show little propensity to
engage in hydrogen bonding of water molecules and water
molecules bind almost exclusively via the hydrogen atom of the
carboxyl group. We here experimentally confirm that the bond
between a water molecule and the carboxyl group is stronger
than the one between a water molecule and the hydroxyl group.
A possible explanation is the presence of stronger positive and
negative partial charges at the oxygen and hydrogen atoms of
the respective groups as is evident from the ESP map in Fig. 1d.
In the second binding motif, the water molecule is bound to the
nitro group and located in line with the molecular axis (Fig. 2b).
Fitting of a line profile along the molecular axis results in a
separation of the two NPAS protrusions of (0.80 ± 0.03) nm,
while the distance between the NPAS and water protrusion
amounts to (0.74 ± 0.05) nm (Fig. 2d).
Figure 3. NPAS hydration. a) False color STM image of pristine NPAS
molecule with calculated gas-phase structure of NPAS superimposed. (b - d):
2
STM images of NPAS/water complexes for three preparations with D O/NPAS
Because of the overwhelming majority of these two binding
motifs we conclude that even at temperatures below 30 K the
water molecules are mobile enough to diffuse to preferred
positions at the NPAS molecule. This implies that the carboxyl
and nitro group are energetically favored over other functional
groups like the diazo bridge or the hydroxyl group.
ratios of: a) 2:1, b) 18:1, c) 70:1 with calculated gas-phase structure of NPAS
superimposed. Features differing from a) are caused by the presence of water.
(
e – g): Stacked semi-transparent STM images for the three preparations in
b – d). Number of stacked STM images is given by N. Diagonal lines in the
(
images are stacking artifacts.
water at any given position around the NPAS molecule at that
particular ratio. Evidently, the carboxyl group and the nitro group
are the nucleation centers for the observed solvation shell
Formation of a complete solvation shell
By gradually increasing the NPAS:water ratio we now follow the
formation of the two-dimensional solvation shell around the
NPAS molecules. The pink dumbbell shape in the centers of all
images in Fig. 3 represents the NPAS molecule (compare Fig.
(
Fig. 3e). In addition, as already seen for the competition
between carboxyl and nitro group, the carboxyl group attracts
slightly more water. This is indicative of a stronger influence of
the carboxyl group than the nitro group on the water molecule.
With increasing NPAS:water ratio the solvation shell extends
further, but neither the phenyl rings nor the diazo bridge get
solvated (Fig. 3f). At this intermediate NPAS:water ratio, we thus
image the hydrophilic and hydrophobic properties of an organic
molecule on the molecular scale. Further increase of
NPAS:water ratio eventually leads to fully solvated NPAS
molecules (Fig. 3g). Note, that at larger ratios the hydroxyl group
influences the orientation of solvation shell, though it does not
show a distinct propensity for initial water coordination. This is
especially evident from Fig. 3g, where the two bulges above the
NPAS molecule are of comparable size and are equally
positioned at carboxyl and hydroxyl group.
3a and Fig. 1b). All features in the images Fig. 3b – 3g that differ
from Fig. 3a are caused by the presence of water. At low ratio,
the water molecules coordinate to the carboxyl and nitro group
exclusively. Hydroxyl group, diazo bridge and the phenyl rings
do not attract water molecules at all (Fig. 3b). At intermediate
NPAS:water ratio a water network grows around the initial
coordination sites. Water molecules added to the saturated nitro
group do not coordinate to the phenyl rings, but build a solvation
shell surrounding the first coordinated water molecule. The
situation at the carboxyl group appears to be more complex.
While also in this case additional water molecules extend the
water network, they may now attach to the hydroxyl group once
the preferred carboxyl group is saturated (Fig. 3c). At even
larger NPAS:water ratio we find NPAS molecules, which are
completely surrounded by water molecules. However, as the
shell extends, the influence of carboxyl and nitro group
decreases and the most distant water molecules form structures
that resemble those of pure water structures on Au(111) grown
Conclusion
On a specifically functionalized azo dye, we demonstrate how
single water molecules may serve to probe the hydrophilic and
hydrophobic properties of adsorbed molecules. This technique
reveals solute/solvent interactions on a sub-molecular scale and
allows pinpointing key positions for solvent coordination on
solute molecules. In principle, it is applicable to any type of
molecule of appropriate size or super-molecular assembly on a
surface and extendable to other solvent molecules as well. By
varying the functional groups, it is conceivable to create a
sequence of solvent affinity for different functional groups. In
order to transfer results gained by this method to three-
dimensional solvation at ambient conditions, cooperative effects
[
28]
at 20 K (Fig. 3d).
As expected from the flexibility of water
networks none of the resulting NPAS/water complexes shows
the same configuration. We derive an average structure of the
solvation shell by stacking semi-transparent STM images from
solvation shells at the same NPAS:water ratios on top of each
other. Using this procedure, we are able to identify the
barycenters of water molecule presence around the NPAS
molecule at different NPAS:water ratios. The map may be
viewed as a lateral probability distribution for the presence of
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for hydrogen bonding, as observed here in two-dimensions need
to be taken into account also for the third dimension, because
they influence the formation of hydrogen bond networks.
Nevertheless, our work offers unique insights into the
interactions between water molecules and functional groups on
the sub-molecular length scale. By revealing the lateral extend
and structure of the solvation shell of a single molecule our work
complements the work by liquid-phase AFM. While the latter
gives a three-dimensional picture of water distribution dominated
by the arrangement of the ions and molecules that compose a
surface, our results shed light on the influence of individual
functional groups on water distribution. The fundamental
understanding of solvation gained in our model study is an
important step forward in the ongoing quest to understand
solvation on the single molecule level.
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STM measurements are performed with a low-temperature
[
[
[
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STM
p < 2 · 10 mbar). The Au(111) surface is cleaned by repeated
cycles of sputtering and annealing. The sputtering gas used is
under
ultrahigh
vacuum
(UHV)
conditions
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−
10
(
+
−5
Ne (3 · 10 mbar, 1−2 μA, 1.3 keV, 30 min). Annealing is at
00 K for 30 min. Details for the preparation of NPAS can be
9
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found in the supporting information. NPAS Molecule deposition:
For deposition on the surface, the molecules are held at 413 K
which yields a pressure of 5 · 10 mbar in a sealed off molecule
deposition chamber. NPAS molecules are deposited onto the
surface held at 23 to 30 K. The deposition time of 45 s is
[
[
[
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controlled via a shutter in front of the effusion cell. The D
milli-q quality is further purified in vacuum through freeze-pump-
thaw cycles. For D O deposition a desired D O pressure is set
within the sealed off molecule deposition chamber. In
dependence on the desired D O coverage it ranges between 1 ·
mbar and 1 · 10 mbar. The procedure is similar and details
are given in the supporting information. Note that the local D
2
O of
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8
2
−
8
−7
10
2
O
and NPAS pressure at the sample surface is much lower than
the one set in the molecule deposition chamber. After exposure,
the sample is transferred to the STM that is operated at a
temperature of 6 K, in constant current mode. All images are
recorded at a bias voltage of 50 mV and a current of 8 pA.
Gas phase structures are calculated semi-empirically using the
Parametric
Hamiltonian.
Method
3
(PM3)-parametrized
MNDO
[
29]
The accuracy of this method is better than the
pixel resolution of our STM images (~ 20 pm/pixel).
Keywords: Scanning probe microscopy • Surface chemistry •
Solvation • Single-molecule studies
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[
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Direct imaging of the hydration
of a substituted azo benzene
by low-temperature scanning
tunnelling microscopy. From
single water molecules to a
complete solvation shell.
Karsten Lucht, Dirk Loose,
Maximilian Ruschmeier], Valerie
Strotkötter, Prof. Gerald Dyker
and Prof. Karina Morgenstern*
Page No. – Page No.
Title
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