Phenoxazine-based supramolecular tetrahedron as biomimetic lectin for glucosamine recognition

Article abstract of DOI:10.1016/j.cclet.2020.07.028

The design and synthesis of a phenoxazine-based metal-organic tetrahedron (Zn₄L₄) as biomimetic lectin for selectively recognition of glucosamine (GlcN) was reported. Different from the free phenoxazine-based ligand (L), Zn₄L₄ displayed the highest fluorescent intensity enhancement efficiency toward GlcN over other related natural mono- and disaccharides. Fluorescence titration demonstrated a 1:1 stoichiometric host-guest complex was formed with an association constant about 4.03 × 10⁴ L/mol. ¹H NMR spectroscopic studies confirmed this selectivity resulted from the multiple hydrogen bonding interactions formed between GlcN and Zn₄L₄. The present results suggested that rational arrangement of recognition sites in the confined space of metal-organic cage is crucial for the selectivity toward target guests.

Full text of DOI:10.1016/j.cclet.2020.07.028

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Contents lists available at ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
Communication
Phenoxazine-based supramolecular tetrahedron as biomimetic lectin
for glucosamine recognition
*
*
Yuchao Li, Xuezhao Li , Lili Li, Bing Xiao, Jinguo Wu, Hechuan Li, Danyang Li, Cheng He
State Key Laboratory of Fine Chemicals, Dalian University of Technology, Dalian 116024, China
A R T I C L E I N F O
A B S T R A C T
Article history:
The design and synthesis of a phenoxazine-based metal-organic tetrahedron (Zn4L4) as biomimetic lectin
for selectively recognition of glucosamine (GlcN) was reported. Different from the free phenoxazine-
based ligand (L), Zn4L4 displayed the highest fluorescent intensity enhancement efficiency toward GlcN
over other related natural mono- and disaccharides. Fluorescence titration demonstrated a 1:1
stoichiometric host-guest complex was formed with an association constant about 4.03 ꢀ 104 L/mol. 1H
NMR spectroscopic studies confirmed this selectivity resulted from the multiple hydrogen bonding
interactions formed between GlcN and Zn4L4. The present results suggested that rational arrangement of
recognition sites in the confined space of metal-organic cage is crucial for the selectivity toward target
guests.
Received 22 June 2020
Received in revised form 2 July 2020
Accepted 14 July 2020
Available online xxx
Keywords:
Glucosamine
Phenoxazine
Metal-organic cage
Fluorescence
© 2020 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences.
Published by Elsevier B.V. All rights reserved.
Host-guest complex
Carbohydrates, as the carriers of energy in life, are essential
mediators of a wide range of biological processes including protein
folding, cell-cell recognition, infection by pathogens or tumor
metastasis [1,2]. The recognition of carbohydrates with high
specificity and sensitivity is biologically important but intrinsically
challenging, for both nature and host-guest chemists [3–5].
Saccharides are high hydrophilic, non-charged, subtle variation
of the three-dimensional structures, and the only one kind of the
recognition units are the hydrogen bonding groups. Even the most
common class of natural receptors, lectins, show notably low
affinities and modest selectivity [6,7]. Biomimetic synthetic
organic-box-like lectins equipped with multi-hydrogen bonding
groups in their inner cavities have been devoted to sensor
saccharides operating through non-covalent interactions [8–16].
However, these purely organic hosts [17–21] commonly required
lengthy and low-yielding synthetic process. Noted that complexa-
tion of saccharides with an organic host receptor was often
detected by the 1H NMR chemical-shift changes [22–28], but this
method showed a limited sensitivity. Alternatively, the using of
fluorescence spectroscopic method to recognize target carbohy-
drates is highly anticipant. This method possesses excellent
signaling properties to modulate the binding process and thus
might leave an improved detection sensitivity [29,30].
On the other hand, self-assembled metal-organic cages (MOCs)
with unique hollow cavity environment and rich chemical/physical
properties enable them as promising receptors for various
compounds recognition [31–33]. The abundant structural config-
urations and fruitful functionalities can be successfully achieved by
rational selection of different metal centers and well-designed
ligands. Especially, when amide groups as multiple hydrogen
bonding triggers used in nature protein–carbohydrate complexes
were incorporated into MOCs, the novel synthetic hosts could be
used as efficient receptors for mimicking biological carbohydrate
recognition processes [34–38]. The key issues remain how to
subtly modulate the distribution of recognition sites, as well as to
form appropriate shape and size to encapsulate specific saccha-
rides. Furthermore, fluorescent properties are also desired by
integrating emissive ligands and/or coordination complexes into
the supramolecular architectures [39] to possess improved signal
output properties.
Herein, the design and synthesis of a metal-organic tetrahedron
(Zn4L4) equipped with four phenoxazinean-based signaling
components and a dozen of amide-based hydrogen binding sites
as biomimetic lectin for selectively recognition of glucosamine
(GlcN) over other related natural mono- and disaccharides was
reported (Scheme 1). By contrast, free phenoxazine-based ligand
(L) showed no obvious enhancement of the fluorescent intensity
towards GlcN. Fluorescence titration and ESI-MS studies demon-
strated that the newly obtained biomimetic lectin host could
encapsulate one GlcN in the cavity environment of Zn4L4 with high
association constant. 1H NMR spectroscopic studies confirmed that
* Corresponding authors.
E-mail addresses: xuezhaoli@dlut.edu.cn (X. Li), hecheng@dlut.edu.cn (C. He).
https://doi.org/10.1016/j.cclet.2020.07.028
1001-8417/© 2020 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences. Published by Elsevier B.V. All rights reserved.
Please cite this article in press as: Y. Li, et al., Phenoxazine-based supramolecular tetrahedron as biomimetic lectin for glucosamine recognition,
Chin. Chem. Lett. (2020), https://doi.org/10.1016/j.cclet.2020.07.028

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Scheme 1. (a) Coordination-driven self-assembly of metal-organic tetrahedron
Zn4L4; (b) Mono- and disaccharides used in the recognition experiments.
the selectivity resulted from the complementary multiple hydro-
gen bonding interactions between amine, hydroxy groups of GlcN
and the amide moieties of Zn4L4. The present results suggested
that rational arrangement of recognition sites in the confined space
of metal-organic cage is crucial for the selectivity toward target
guests, and the supramolecular host-guest species formation
between the cage containing hydrogen-bond recognition sites and
the encapsulated carbohydrates is responsible for the effective
detection process.
As shown in Fig. 1a, ligand L was obtained by connecting
phenoxazine-based core (1) and 2,20-bipyridine chelators through
amine-linkages in two steps [40–42]. Acylating of 1 with thionyl
chloride, and subsequently reacted with 2 to afford ligand L in 78%
yield. The successfully obtained ligand L was confirmed by 1H and
13C NMR spectra (Fig. 1b and Section S2 in Supporting informa-
tion). The reaction of ligand L (1.0 equiv.) and Zn(OTf)2 (1.0 equiv.)
(OTf = trifluoromethanesulfonate) in DMF at 80 ℃ for 12 h resulted
in the formation of Zn4L4 as the unique product. The 1H NMR
spectra of tetrahedron Zn4L4 displayed only one set of ligand
resonances in solution, suggesting the exclusive formation of a
discrete structure to be highly symmetrical (Fig. 1c). Diffusion-
ordered NMR spectroscopy (DOSY) of the product also showed that
the multiple aromatic signals possessed similar diffusion coef-
ficients (D = 6.31 ꢀ10ꢁ11 m2/s, Fig. S8 in Supporting information).
The formation of a discrete supramolecular species containing a
Fig. 1. (a) Schematic pathway for the synthesis of phenoxazine-containing ligand L,
which inserting three amide groups. Sections of the 1H NMR spectra (400 MHz in
DMSO-d6) of ligand L (b) and the tetrahedron Zn4L4 (c). (d) ESI-TOF MS spectrum of
Zn4L4 in DMF and the expanded MS peaks of Zn4L4 with charges +6 to +4. The
experimental results were shown in blue (bottom) and the calculated results were
shown in red (top).
8+
Zn4L4 cation was confirmed by electrospray ionization time-of-
flight mass spectrometry (ESI-TOF MS). As showed in Fig. 1d, a
series of intense peaks at m/z = 458.0999, 544.8200, 660.4491,
822.3308,1065.1522 and 1469.8522 were observed. Comparison of
the experimental peaks with these simulated results obtained on
the basis of natural isotopic abundance, these peaks were safely
assigned to the species [Zn4(L)4]8+ (calcd. 458.1022), [Zn4(L)4+OTf]7
(calcd. 544.8243), [Zn4(L)4+2OTf]6+ (calcd. 660.4538),
+
[Zn4(L)4+3OTf]5+ (calcd. 822.3351), [Zn4(L)4+4OTf]4+ (calcd.
1065.1570) and [Zn4(L)4+5OTf]3+ (calcd. 1469.8602), respectively.
This result demonstrated the Zn-based tetrahedral architecture
was successfully assembled with substantially stability in solution.
To confirm the formation of a [4 + 4] metal-organic cage, UV–vis
Fig. 2. UV–vis titration of ligand L (20.0
24.0 mol/L) in CH3CN: DMF = 4:1 (v:v). Inset: Absorbance variation curves of L
upon the addition of Zn2+ at 310 nm and 432 nm.
mmol/L) with the addition of Zn(OTf)2 (0-
m
titration of ligand L (20.0 m
mol/L) upon the addition of Zn2+ was
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further well conducted. As showed in Fig. 2, the ligand L exhibited
two broad bands at =310 nm and 390 nm, which assignable to the
* and n–
* charge transfer bands. Upon the addition of Zn2+
ions, the band at =310 nm was decreased gradually, while the
intensity of new peaks appeared at = 324 and =432 nm were
and in the short edges was about 18.34(1) Å. The average dihedral
angle between the phenyl groups and phenoxazine planes in Zn4L4
was about 74.41(1)ꢂ. The structure possessed a confined cavity and
contains 12 amide groups, which provides a great possibility for
carbohydrate recognition.
l
p
–
p
p
l
l
l
increasing significantly. These bands remained constant after
The fluorescent nature of ligand L was preserved and
adding approximately 1.0 equiv. of Zn2+ ions. The presence of sharp
transferred to the final cage, which enable the cage Zn4L4 with
luminescent recognition process of relative guests (Fig. S10 in
Supporting information). Fluorescent emission spectrum of Zn4L4
exhibited a ligand-based broad emission band at lem =448 nm
isosbestic point at
l =322 nm indicated that only two species
coexisted in the equilibrium system. The almost linear relations
between the absorbance at both bands and the concentration of
Zn2+ added revealed that the formation of the cage complex was
quantitative and the complex exhibited 1:1 stoichiometry. This
result demonstrated that the M4L4 cage compound was the only
one complexation specie and the association constant of the
complexation specie is relatively high.
when excited at lex =396 nm (5.0 mmol/L in DMF). Subsequently,
the potential application of Zn4L4 as a fluorescent detector for
natural saccharides recognition was explored. Impressively, upon
the addition of GlcN (0–10 equiv.) into the DMF solution of Zn4L4
(5 mmol/L), the wavelength of the emission maximum at lem
Numerous efforts for crystallization of Zn4L4 to obtain suitable
crystals for the high-quality X-ray diffraction have been unsuc-
cessful. Therefore, in order to understand the three-dimensional
structure of Zn4L4 better, we use a MM2 energy-minimized model
to simulate the structure of Zn4L4 (Fig. 3). Through the optimized
structures, we can more intuitively insight that the Zn4L4 was a
twisted tetrahedron structure with a hollow cavity. The four metal
atoms occupied the vertices and each of them was coordinated
with three ligands. Each ligand was positioned on one of the four
faces of the tetrahedron which defined by four metal ions. The
Zn . . . Zn distance in the long edges of Zn4L4 was about 19.38(1) Å,
=448 nm did not change but the luminescence intensity enhanced
gradually with the increasing concentration of the guest. When
adding of 10.0 equiv. GlcN, the fluorescence intensity of Zn4L4 was
dramatically increased around 82%. The hill-plot profile of the
fluorescence titration curves at 448 nm demonstrated a 1:1
stoichiometric host-guest complex was formed with an associa-
tion constant about 4.03 ꢀ 104 L/mol (Fig. 4). The formation of
donor-type hydrogen bonds between the amide groups of Zn4L4
and the guest molecules could alter the electronic distribution of
the ligand backbone, which suppress the process of photo-induced
electron transfer (PET) process, thus leading to the significant
luminescence enhancement [36]. Interestingly, the glucose (Glu)
only induced negligible emission enhancement of Zn4L4 under the
same condition (Fig. 5). The only difference between these two
saccharides is that there is one amine group in the skeleton of GlcN.
However, the very different recognition ability of Zn4L4 toward
them suggested that the amine might played an important role.
Under the same condition, when 10.0 equiv. of cyclohexanamine
and aniline were added, the fluorescent emission intensity of Zn4L4
was increased around 25% and 29%, respectively (Fig. S13 in
Supporting information). Noted that other mono-saccharides
including mannose (Man), ribose (Rib) and xylose (Xyl) showed
negligible emission enhancement less than 10%. Furthermore, the
addition of excess disaccharides including the sucrose (Suc),
lactose (Lac), melibiose (Mel), maltose (Mal), and trehalose (Tre)
did not cause any obvious spectroscopic changes of Zn4L4 as well
(Fig. 5). Combined with the result that free ligand L showed no
recognition ability to GlcN, these results suggested that the specific
recognition process mainly occurs in the cavity of the cage, and the
larger disaccharide could not be encapsulated into the cavity.
The excellent selectivity of Zn4L4 toward GlcN encouraged us to
further investigate the interaction detail of the host–guest
Fig. 3. The optimized structure of Zn4L4. (a) Ball-and-stick model (Zn atoms are
shown as green spheres); (b) Space-filling model. Disorder atoms and solvent
molecules of crystallization are omitted for clarity. Color code: C = gray; N = blue;
O = red; H = white; Zn = green.
Fig. 4. Family of luminescence spectra of Zn4L4 (5 mmol/L) upon the addition of
different concentrations GlcN (0-10.0 equiv.). Inset: Hill-plot titration curve
showing the 1:1 host–guest behavior.
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In summary, we reported a supramolecular tetrahedron as
biomimetic lectin for GlcN recognition with high specificity and
sensitivity over other related natural mono- and disaccharides. The
Zn4L4 displayed phenoxazine-based broad fluorescent emission
property, and equipment of 12 amide groups in the skeleton. The
formation of a 1:1 host-guest complexes has been characterized by
fluorescence titration spectroscopy and ESI-MS. 1H NMR spectro-
scopic titration studies and NOESY spectra confirmed that the
hydrogen bonding interactions between GlcN and receptor Zn4L4,
resulting a stronger host-guest binding affinity. The present results
demonstrated that the rational distribution of amide groups and
luminescent ligands into the 3D metal-organic cage structure not
only provides a recognizable hydrogen-bonding functional site for
the guest molecule, which facilitates the formation of the host-
guest complex, but also provides a way to convert identification
information into effective communicator for optical signals.
Accordingly, changing the topology and the arrangement of
hydrogen binding sites in the confined space of host, the selectivity
toward other target guests could be expected.
Fig. 5. The fluorescent emission enhancement efficiency of Zn4L4 upon addition of
different carbohydrates. Inset: different fluorescent emission enhancement
efficiency of Zn4L4 and L toward GlcN.
Declaration of competing interest
The authors declare that they have no known competing
financial interests or personal relationships that could have
appeared to influence the work reported in this paper.
Acknowledgments
We appreciate the National Natural Science Foundation of
China (Nos. 21701019 and 21861132004) and Doctoral Scientific
Research Launching Fund Project of Liaoning province (No. 2019-
BS-050).
Appendix A. Supplementary data
Supplementarymaterialrelatedtothisarticlecanbefound, inthe
online version, at doi:https://doi.org/10.1016/j.cclet.2020.07.028.
Fig. 6. 1H NMR titration (400 MHz in DMSO-d6) of Zn4L4 (10 mmol/L) upon addition
of GlcN (0-10.0 equiv.).
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Chin. Chem. Lett. (2020), https://doi.org/10.1016/j.cclet.2020.07.028