# Tricarbonylrhenium(I) complexes with aggregation-induced phosphorescence emission (AIPE) properties: Application to the selective detection of heparin and its main contaminant

Corinne Vanucci-Bacqué, Valentine Guilbaud, Maëlle Deleuzière, Charles-Louis Serpentini, Florence Bedos-Belval, Eric Benoist, Suzanne Fery-Forgues

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Corinne Vanucci-Bacqué, Valentine Guilbaud, Maëlle Deleuzière, Charles-Louis Serpentini, Florence Bedos-Belval, et al. Tricarbonylrhenium(I) complexes with aggregation-induced phosphorescence emission (AIPE) properties: Application to the selective detection of heparin and its main contaminant. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy [1994-..], 2025, 334, pp.125932. 10.1016/j.saa.2025.125932. hal-04993372

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Distributed under a Creative Commons Attribution 4.0 International License
Tricarbonylrhenium(I) complexes with aggregation-induced
phosphorescence emission (AIPE) properties: Application to the selective
detection of heparin and its main contaminant

Corinne Vanucci-Bacqué,ª Valentine Guilbaud,ª Maëlle Deleuzière,ª Charles-Louis Serpentini,ᵇ Florence Bedos-Belval,ª Eric Benoist a and Suzanne Fery-Forgues a*

a. SPCMIB, CNRS UMR 5068, Université de Toulouse, 118 route de Narbonne, 31062 Toulouse cedex 9,
France.

b. Laboratoire SOFTMAT, CNRS UMR 5623, Université de Toulouse, 118 route de Narbonne, 31062 Toulouse cedex 9, France

* Corresponding author.

E-mail: suzanne.fery-forgues@univ-tlse3.fr

† Electronic Supplementary Information (ESI) available: Experimental details including proton numbering for NMR, NMR spectra, emission spectra, fluorescence microscopy images.

**Abstract**

Phosphorescent tricarbonylrhenium(I) complexes that are more emissive in the aggregate state
than in solution could be very valuable probes for biological analyses, but their development is
delicate. The present work focuses on the synthesis and spectroscopic study of three new
complexes that differ by the nature of their positively charged substituent. The dissolved
complexes were very weakly emissive. Due to electrostatic interaction and aggregation, they
showed a strong aggregation-induced phosphorescence enhancement (AIPE) effect in the
presence of heparin, a polyanionic macromolecule of biological interest, and a weaker effect in
the presence of chondroitin sulfate, which contains fewer negative charges. The magnitude of
the AIPE effect depended on the nature of the complex. However, it was weaker than expected
from the behavior of the parent neutral complex studied in a conventional acetonitrile/water
system, which highlights the challenge of accurately predicting solid-state emission properties
for this class of molecules. This work introduces rhenium(I) complexes in the field of AIPE-
active probes for the detection of polyanionic biomolecules.

<figure><img src="image_2.png" /></figure>
**Keywords**

Rhenium; Photoluminescence; Aggregation-induced emission; Heparin

**Highlights**

Three new phosphorescent Re(I) complexes were tested for the detection of heparin

The double-charged one showed the best AIPE effect

A much weaker effect was obtained with chondroitin sulfate

Re(I) complexes are valuable AIPE probes, although photoluminescence properties are difficult to anticipate

# 1. Introduction

Transition metal complexes that exhibit solid-state luminescence enhancement (SLE) [1] and more specifically aggregation-induced phosphorescence emission (AIPE), i.e. the rare property of emitting phosphorescence much more intensely in aggregate form than in solution, have experienced a dramatic development over the past two decades [2–6]. Their diversity is remarkable and they are currently used in a very large number of applications ranging from materials to biology and theranostics. Compared to organic AIE probes, they stand out by their long luminescence lifetimes and photochemical stability, allowing unique detection techniques and prolonged use. However, the AIPE phenomenon is not only based on molecular stiffening. It also depends heavily on the intermolecular interactions that take place in the solid state and on the prevention of emission quenching by oxygen molecules. Original photophysical pathways may also vary between solution and the various solid phases. Controlling all this set of parameters can be very difficult. The challenging development of AIPE-gens based on transition metal complexes is probably the reason why they have been much less studied than their organic counterparts.

Tricarbonylrhenium(I) complexes have been little studied in this context, whereas their phosphorescence properties have long been exploited for conventional applications in biology [7]. Their AIPE properties were first reported in 2002 by the Rajagopal and Lu group on quadrinuclear Re(I) alkoxy-bridged molecular rectangles [8]. Next, dinuclear Re(I) alkoxy-bridged complexes and tetrarhenium metallacycles have been used to detect explosives [9–11], antibiotics [11], amyloid fibrils related to Alzheimer's disease [12] and insulin fibrillation [13]. A dinuclear species has been reported to show crystallization-induced emission, with the formation of two polymorphs that emit differently [14]. For mononuclear species, a variety of complexes with good SLE/AIPE properties have been developed without specific applications [15–21]. Gabr and Pigge took advantage of this effect to detect human serum albumin [22] and amyloid fibrils [23], using Re(I) complexes bound to tetraarylethylene, a well-known AIE-active motif.

Over the last few years, our group has studied tricarbonylrhenium(I) complexes with the aim of developing photoluminescent materials. These complexes incorporate an original ligand, made of a bidentate pyridyl-triazole moiety (pyta), which has excellent chelating capacity and
exists in various isomeric forms. The chemical modifications of the appended aromatic ring
allow to fine-tune the luminescence behavior. All complexes are poorly emissive in solution,
but an ingenious design has led to the emergence of remarkable SLE properties when in powder
form [24–32]. The AIPE properties of the best emitters were tested in acetonitrile/water
mixtures, with very promising results [28–32]. It is now important to show that our Re(I)
complexes can be used as AIPE-gens for the detection of biological molecules. The decision
was therefore made to choose a simple and easy-to-substitute framework, and to improve its
design for AIPE applications. We selected complex **Re-Phe** (Fig. 1), which is built with a 3-(2-
pyridyl)-1,2,4-triazole ligand and has a PL quantum yield of 0.42 in the form of microcrystalline
powder [25]. The target was the detection of heparin, a sulfonated polysaccharide commonly
used as anticoagulant for the prevention and treatment of thromboembolic disorders [33].
Heparin is considered to be one of the most negatively charged natural molecules, since it
contains one carboxyl group and 2–3 sulfonated groups per disaccharide unit. Its main
contaminant is chondroitin sulfate (CS), which contains a smaller number of sulfonated groups.
It is of major interest to distinguish between the two compounds because adulteration of heparin
can have an impact on its safety [33]. It has been shown in the literature that a variety of organic
AIE-gens [34–48] and arene-based probes that give highly luminescent excimers upon
aggregation [49–53] allow a very effective direct detection of heparin, with possible
discrimination from CS. Regarding transition metal probes, an osmium(II) complex has been
used, resulting in a phosphorescence emission multiplied by 2.3 in the presence of heparin, but
AIPE was not mentioned [54]. All these molecules bear at least one positive charge, so that
they interact with anionic polysaccharides by electrostatic interactions. Based on this model,
**Re-Phe** was provided with positively charged substituents, giving three distinct cationic
derivatives: complex **Re-Phe-MP** bears an *N*-methylpyrrolidinium group, while complexes **Re-
Phe-AM** and **Re-Phe-AM2** contain one and two ammonium groups, respectively (Fig. 1a).
These complexes are expected to combine with Heparin and CS, resulting in molecular
aggregation and enhanced photoluminescence emission (Fig. 1b). The comparison of the three
complexes should reveal whether the nature and number of cationic groups affect the
recognition process. It was assumed that the nature of the counter-anions associated to the
complexes does not play a significant role in the recognition process. The cationic complexes
were synthesized and their spectroscopic study was undertaken in solution and in the solid state.
Then, they were placed in the presence of increasing amounts of heparin or CS. The AIPE effect
obtained was discussed. A comparison was made with **Re-Phe** tested under conventional
acetonitrile/water conditions, to determine the degree of predictability of the AIPE effect. This
study highlights the interest of mononuclear Re(I) complexes for specific AIPE applications,
and underlines the limitations of the present system, in order to evolve towards an increasingly
efficient design.
<figure><img src="image_3.png" /></figure>

<figure><img src="image_4.png" /><figcaption>**Fig. 1** a) Chemical structure of the tricarbonylrhenium(I) complex **Re-Phe** and its ionic derivatives **Re-Phe-MP**, **Re-Phe-AM** and **Re-Phe-AM2**. b) Schematic interaction between heparin and one cationic complex, with generation of AIPE effect.</figcaption></figure>

**2. Experimental section**

**2.1. Materials and methods**

Heparin was sodic enoxaparin (Lovenox®) 4000 UI (40 mg)/0.4 mL. Chondroitin sulfate from shark cartilage, hyaluronic acid and dextran were purchased from Sigma-Aldrich. Hyaluronic acid was converted as its sodium salt by addition of dilute NaOH solution. Deionized water (18.2 MΩ.cm, pH 6) was produced by an Elga Pure Lab system from Veolia. For synthesis, all purchased chemicals were of the highest purity commercially available and used without further purification. [Re(CO)₅Cl] was from Thermo Scientific. Pyridine-oxadiazole and di-tert-butyl(azanediylbis(ethane-2,1-diyl))dicarbamate were prepared according to ref. [25] and [55], respectively. Analytical grade solvents were used as received. Experiments were carried out under a nitrogen atmosphere. Reactions were monitored by TLC on silica gel Alugram® Xtra SIL G/UV₂₅₄. Column chromatography was performed on Machery-Nagel silica gel.
NMR, mass and infrared spectra were obtained in the relevant 'Services communs de l'Institut de Chimie de Toulouse, Université de Toulouse III-Paul-Sabatier'. ¹H- and ¹³C-NMR spectra were recorded on a Bruker Avance 300 MHz spectrometer. Attributions of the signals were made using 2D NMR data (COSY, HSQC and HMBC). Signals are described as follow: s, singlet; brs, broad singlet; d, doublet; t, triplet; m, multiplet. HRMS data were recorded on a Xevo G2 QTOF (Waters) instrument. Infrared spectra were obtained on a Nexus Thermonicolet apparatus with DTGS as the detector. Microanalyses were performed in the 'Service d'analyse' of LCC using a Perkin Elmer 2400 series II analyzer.

## 2.2. Synthesis

### 4-(3-(Pyridin-2-yl)-4H-1,2,4-triazol-4-yl)phenethyl methanesulfonate (1)

To a solution of 2-(pyridin-2-yl)-1,2,3-oxadiazole (297 mg, 2.16 mmol) in xylenes (5.5 mL) were added 2-(4-aminophenyl)ethan-1-ol (318 mg, 2.16 mmol) and *p*-toluenesulfonic acid (*p*-TsOH) (41 mg, 0.16 mmol). The reaction was stirred under nitrogen atmosphere at 140°C for 19 h. After cooling to room temperature, $CH_2Cl_2$ was added and the organic layer was washed with 1M aqueous NaOH solution. The aqueous layer was extracted with $CH_2Cl_2$, the combined organic layers were washed with brine, dried over $Na_2SO_4$ and concentrated under reduced pressure. To the crude product dissolved in anhydrous $CH_2Cl_2$ (25 mL) and cooled to 0°C were successively added methanesulfonyl chloride (0.25 mL, 3.15 mmol) and $NEt_3$ (0.75 mL, 5.25 mmol). After stirring for 3 h at 0°C, the reaction was quenched with saturated aqueous $NH_4Cl$ solution. The aqueous layer was extracted with $CH_2Cl_2$, the combined organic layers were washed with brine, dried over $Na_2SO_4$ and concentrated under reduced pressure. Silica gel column chromatography (EtOAc/MeOH, 95:5 v/v) afforded the expected compound as a white solid (500 mg, 67%). $M_p$ = 134–135°C.

¹H NMR (300 MHz, $CDCl_3$) $δ$(ppm): 8.35-8.33 (m, 1H), 8.32 (s, 1H), 8.22-8.11 (m, 1H), 7.80 (td, *J* = 7.8, 1.8 Hz, 1H), 7.38-7.20 (m, 5H), 4.47 (t, *J* = 6.7 Hz, 2H), 3.13 (t, *J* = 6.7 Hz, 2H), 2.93 (s, 3H). ¹³C NMR (75 MHz, $CDCl_3$) $δ$(ppm): 152.1, 149.0, 146.5, 145.5, 137.5, 136.9, 134.2, 129.8, 126.1, 124.3, 124.0, 69.7, 37.4, 35.1. HRMS (ESI) *m/z* 345.1021 ([M + H]⁺ calculated for $C_{16}H_{17}N_4O_3S$: 345.1021).

### Complex [Re(CO)₃(L1)Cl], (Re-Phe-MP)

A mixture of **1** (110 mg, 0.32 mmol) and *N*-methylpyrrolidine (0.33 mL, 3.2 mmol) in $CH_3CN$ (3 mL) was stirred at 50°C for 17h. The reaction mixture was concentrated under reduced pressure to afford **L1**. The crude product was dissolved in MeOH (3 mL). [Re(CO)₅Cl] (116 mg, 0.32 mmol) was added and the mixture was stirred at 65°C for 23h. The resulting yellow solution was concentrated under reduced pressure giving the expected complex (232 mg, 98%).

¹H NMR (300 MHz, MeOD) $δ$(ppm): 9.11-9.09 (m, 1H), 8.88 (s, 1H), 7.99 (td, *J* = 7.9, 1.5 Hz, 1H), 7.80-7.62 (m, 5H), 7.17 (dt, *J* = 8.1, 1.1 Hz, 1H), 3.80-3.61 (m, 6H), 3.38 (dd, *J* = 8.0, 3.9 Hz, 2H), 3.25 (s, 3H), 2.69 (s, 3H), 2.35-2.27 (m, 4H). ¹³C NMR (75 MHz, MeOD) $δ$(ppm): 198.5, 197.3, 189.4, 156.3, 155.9, 149.0, 146.1, 141.8, 141.1, 132.9, 132.6, 129.3, 128.5, 124.4, 65.8, 65.7, 49.1, 39.6, 30.7, 22.7. HRMS (ESI) *m/z* 638.1098 ([M – OM]⁺ calculated for $C_{23}H_{24}N_5O_3Cl^{185}Re$: 638.1097). IR (KBr): $ν_{(CO)}$ = 2016, 1902, 1885 cm⁻¹.
### tert-Butyl (4-(3-(pyridin-2-yl)-4H-1,2,4-triazol-4-yl)phenethyl)carbamate (L2)

To a solution of 2-(pyridin-2-yl)-1,2,3-oxadiazole (125 mg, 0.85 mmol) in xylenes (3 mL) were added tert-butyl(4-aminophenethyl)carbamate (200 mg, 0.85 mmol) and p-TsOH (16 mg, 0.084 mmol). The reaction was stirred under nitrogen atmosphere at 140°C for 19 h. After cooling to room temperature, CH₂Cl₂ was added and the organic layer was washed with 1M aqueous NaOH solution. The aqueous layer was extracted with CH₂Cl₂, the combined organic layers were washed with brine, dried over Na₂SO₄ and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (EtOAc/MeOH from 100:0 to 97.5:2.5 v/v) to afford the expected product (144 mg, 47%) as a white solid.

¹H NMR (300 MHz, CDCl₃) δ(ppm): 8.36 (ddd, *J* = 4.9, 1.9, 1.0 Hz, 1H), 8.32 (s, 1H), 8.14 (dt, *J* = 7.9, 1.1 Hz, 1H), 7.79 (td, *J* = 7.8, 1.8 Hz, 1H), 7.35–7.09 (m, 6H), 4.60 (s, 1H), 3.41 (q, *J* = 6.9 Hz, 2H), 2.86 (t, *J* = 7.1 Hz, 2H), 1.44 (s, 9H). ¹³C NMR (75 MHz, CDCl₃) δ(ppm): 155.9, 152.1, 149.0, 146.6, 145.6, 140.2, 136.8, 133.5, 129.6, 125.8, 124.2, 124.0, 79.3, 41.6, 35.9, 28.4. HRMS (ESI) *m/z* 366.1936 ([$M^+$ + H]⁺ calcd for C₂₀H₂₄N₅O₂: 366.1930).

### Complex [Re(CO)₃(L2)Cl], (ReL2)

A mixture of ligand L2 (96 mg, 0.263 mmol) and [Re(CO)₅Cl] (96 mg, 0.265 mmol) in MeOH (4 mL) was heated at reflux temperature for 20 h. The reaction mixture was concentrated under vacuum and the residue was purified by silica gel column chromatography (CH₂Cl₂/MeOH: 98:2 v/v) to afford the expected complex (152 mg, 86%) as a yellow solid.

¹H NMR (300 MHz, CDCl₃) δ(ppm): 9.11 (d, *J* = 5.4 Hz, 1H), 8.31 (s, 1H), 7.82 (td, *J* = 7.9, 1.5 Hz, 1H), 7.64–7.41 (m, 5H), 7.08 (d, *J* = 8.1 Hz, 1H), 4.70 (s, 1H), 3.48 (q, *J* = 6.7 Hz, 2H), 2.98 (t, *J* = 6.9 Hz, 2H), 1.42 (s, 9H). ¹³C NMR (75 MHz, MeOD) δ(ppm): 198.5, 197.1, 189.5, 156.4, 155.8, 146.2, 145.5, 141.2, 132.3, 132.0, 129.2, 127.9, 124.5, 80.0, 42.6, 37.1, 28.8. HRMS (ESI) *m/z* 634.1239 ([$M^-$ – Cl]⁺ calcd for C₂₃H₂₃N₅O₅¹⁸⁵Re: 634.1229). IR (ATR): ν(CO) = 2020, 1916, 1883 cm⁻¹.

### Complex Re-Phe-AM

To complex ReL2 (120 mmol, 0.18 mmol) dissolved in CH₂Cl₂ (3 mL), TFA was added dropwise (0.9 mL). The reaction mixture was stirred at room temperature for 1 h and then concentrated under vacuum. The residue was dissolved in MeOH (2 mL) and 4M HCl solution in dioxane (0.5 mL) was added. The reaction mixture was stirred at room temperature for 45 min. Concentration under vacuum afforded the expected product as a yellow solid (109 mg, 100%).

¹H NMR (300 MHz, MeOD) δ(ppm): 9.09 (ddd, *J* = 5.5, 1.5, 0.8 Hz, 1H), 8.85 (s, 1H), 8.01 (td, *J* = 8.0, 1.5 Hz, 1H), 7.80–7.62 (m, 5H), 7.24 (td, *J* = 8.1, 1.1 Hz, 1H), 3.34 (m, 2H), 3.18 (dd, *J* = 8.6, 6.2 Hz, 2H). ¹³C NMR (75 MHz, MeOD) δ(ppm): 198.4, 197.1, 189.4, 156.3, 155.8, 148.4, 146.1, 142.4, 141.2, 132.8, 132.4, 129.3, 128.5, 124.5, 41.5, 34.2. HRMS (ESI) *m/z* 534.0702 ([$M^+$ – HCl – Cl]⁺ calcd for C₁₈H₁₅N₅O₃¹⁸⁵Re: 534.0704). IR (ATR): ν(CO) = 2025, 1911 and 1887 cm⁻¹.
**Di-tert-butyl (((4-(3-(pyridin-2-yl)-4H-1,2,4-triazol-4-yl)phenethyl)azanediyl)bis(ethane-2,1-diyl))dicarbamate (L3)**

A mixture of **1** (196 mg, 0.57 mmol) and di-tert-butyl(azanediylbis(ethane-2,1-diyl))dicarbamate (303 mg, 1 mmol) in anhydrous acetonitrile (8 mL) in the presence of diisopropylethylamine (DIPEA) (150 µL, 0.86 mmol) was heated at reflux temperature for 3 days. The reaction mixture was cooled to room temperature and diluted with CH₂Cl₂. The organic layer was washed with water and brine, dried over Na₂SO₄ and concentrated under vacuum. Silica gel column chromatography (EtOAc/MeOH from 100:0 to 95:5 v/v) afforded the expected compound (98 mg, 31%) as a white solid foam.

¹H NMR (300 MHz, CDCl₃) δ(ppm): 8.36 (ddd, *J* = 4.8, 1.8, 0.9 Hz, 1H), 8.33 (brs, 1H) 8.14 (dt, *J* = 7.9, 1.1 Hz, 1H), 7.79 (td, *J* = 7.8, 1.8 Hz, 1H), 7.33–7.21 (m, 3H), 7.27–7.17 (m, 2H), 4.78 (s, 2H), 3.10–3.05 (m, 4H), 2.76 (dt, *J* = 4.7, 2.6 Hz, 4H), 2.58 (t, *J* = 5.9 Hz, 4H). ¹³C NMR (75 MHz, CDCl₃) δ(ppm): 156.1, 152.2, 149.1, 146.8, 145.9, 141.9, 136.9, 133.5, 129.7, 126.0, 124.3, 124.1, 79.4, 55.8, 53.5, 38.5, 33.6, 28.5. HRMS (ESI) *m/z* 552.3321 ([M+H]⁺ calcd for C₂₉H₄₂N₇O₄: 552.3298).

**Complex [Re(CO)₃(L3)Cl], (ReL3)**

A mixture of ligand **L3** (97 mg, 0.176 mmol) and [Re(CO)₅Cl] (64 mg, 0.176 mmol) in MeOH (4 mL) was heated at reflux temperature for 19 h. The reaction mixture was concentrated under vacuum and the residue was purified by silica gel column chromatography (CH₂Cl₂/MeOH from 100/0 to 98:2 v/v) to afford the expected **ReL3** complex (105 mg, 70%) as a yellow solid.

¹H NMR (300 MHz, MeOD) δ(ppm): 9.10 (ddd, *J* = 5.5, 1.5, 0.8 Hz, 1H), 8.88 (s, 1H), 7.99 (td, *J* = 7.9, 1.6 Hz, 1H), 7.76–7.56 (m, 5H), 7.18 (dt, *J* = 8.1, 1.1 Hz, 1H), 3.11 (t, *J* = 6.5 Hz, 4H), 2.98–2.83 (m, 4H), 2.65 (t, *J* = 6.5 Hz, 4H), 1.43 (s, 18H). ¹³C NMR (75 MHz, MeOD) δ(ppm): 198.5, 197.2, 189.4, 158.4, 156.3, 155.8, 149.6, 146.6, 146.1, 141.1, 132.4, 131.6, 129.1, 127.8, 124.3, 80.1, 57.1, 54.8, 39.6, 34.5, 28.8. HRMS (ESI) *m/z* 856.2365 ([M + H]⁺ calcd for C₃₂H₄₂N₇O₇Cl¹⁸⁵Re: 856.2364). IR (ATR): *v*(CO) = 2021, 1914, 1887 cm⁻¹.

**Complex Re-Phe-AM2**

To complex **ReL3** (98 mmol, 0.114 mmol) dissolved in CH₂Cl₂ (1.5 mL), TFA was added dropwise (0.75 mL). The reaction mixture was stirred at room temperature for 45 min and then concentrated under vacuum. The residue was dissolved in MeOH (1 mL) and 4M HCl solution in dioxane (0.5 mL) was added. The reaction mixture was stirred at room temperature for 45 min. Concentration under vacuum afforded the expected product as a yellow solid (84 mg, 100%).

¹H NMR (300 MHz, MeOD) δ(ppm): 9.11 (d, *J* = 5.4 Hz, 1H), 8.87 (s, 1H), 8.01 (t, *J* = 7.9 Hz, 1H), 7.69 (d, *J* = 11.5 Hz, 5H), 7.21 (d, *J* = 8.0 Hz, 1H), 3.42–3.30 (m, 12H). ¹³C NMR (75 MHz, MeOD) δ(ppm): 29.5, 35.2, 51.9, 55.9, 124.6, 128.4, 129.2, 132.5, 132.6, 141.2, 146.1, 149.4, 155.9, 156.3, 189.4, 197.1, 198.4. HRMS (ESI) *m/z* 656.1321 ([neutral M – HCl – Cl]⁺ calcd for C₂₂H₂₆N₇O₃Cl¹⁸⁵Re: 656.1315). IR (ATR): *v*(CO) = 2026, 1910 and 1890 cm⁻¹.
## 2.3. Spectroscopy, photochemistry and particle characterization

For solutions and suspensions, spectroscopic measurements were conducted at 20°C in temperature-controlled cells. UV-visible absorption spectra and emission spectra were measured with a Xenius SAFAS spectrofluorometer using cells of 1 cm optical pathway. All emission spectra were corrected. The emission quantum yields in solution ($\Phi$) were determined using the classical formula:

$$ \Phi_x = (A_s \times I_x \times n_x^2 \times \Phi_s)/(A_x \times I_s \times n_s^2) \quad (1) $$

where A is the absorbance at the excitation wavelength, I the integrated emission intensity and n the refractive index. Subscripts s and x refer to the standard and to the sample of unknown quantum yield, respectively. Coumarin 153 ($\Phi_s$ = 0.53) in ethanol was used as the standard [56]. The absorbance of the solutions was equal or below 0.06 at the excitation wavelength. The error on the quantum yield values is estimated to be about 10 %.

The photochemical stability of the complexes was tested at 25°C using an Analytik Jena Specord S600 absorption spectrophotometer equipped with a 385 nm emitting diode (1.6 × 10¹⁶ photon s⁻¹) for irradiation.

For AIPE measurements on **Re-Phe**, a small volume (20 µL) of a concentrated solution of complex (9.6 × 10⁻³ M) in acetonitrile was injected in 2.98 mL of various acetonitrile/water mixtures. The complex concentration in all samples was 6.4 × 10⁻⁵ M. For AIPE measurements on cationic complexes in the presence of polysaccharides, stock solutions of heparin, CS, hyaluronic acid or dextran in water at 1 mg/mL were first prepared. Then, 160 µL of a concentrated solution of the cationic complexes in water (2 × 10⁻³ M) was injected into 2.84 mL water containing the polysaccharide at concentrations varying between 0 and 45 µg/mL. The complex concentration in all samples was 1 × 10⁻⁴ M. All samples were stirred for 3 h in the dark, and they were sonicated for 3 min before optical measurement. Each experiment was at least duplicated.

Regarding solid samples, emission spectra were recorded on a HORIBA Fluorolog 3-2iHR320 spectrofluorometer and were corrected. Absolute photoluminescence quantum yields ($\Phi_{PL}$) were measured on the Xenius SAFAS spectrofluorometer equipped with an integrating sphere. They were determined using a method based on the one developed by de Mello *et al* [57], as described elsewhere [31]. The error was estimated to be about 20%.

Emission decay curves were recorded on liquid samples, the absorbance of which was lower than 0.1 at $\lambda_{ex}$, and on solid samples deposited on a quartz holder. The time-correlated single photon counting (TCSPC) method was performed on the HORIBA Fluorolog 3-2iHR320 spectrofluorometer used at right-angle geometry and equipped with a Nanoled-370 ($\lambda_{ex}$ = 371 nm). Emitted photons were detected by means of a Hamamatsu R928 photomultiplier. Emission was recorded near the maximum with a bandpass of 4 nm. The instrumental response was recorded directly before each decay curve, using the scattering of the sample at 371 nm. All analyses were recorded using the Datastation v2.7 software from Horiba. The decay curves were analyzed with reconvolution and global non-linear least-squares minimization method using DAS6 v6.8 software.

The limit of detection (LOD) of heparin was measured from AIPE titrations ($\lambda_{ex}$ = 390 nm, $\lambda_{em}$ = 574 nm). The standard deviation ($\sigma$) of the blank was first calculated from ten
independent measurements of the phosphorescence intensity of **Re-Phe-AM2** (1 × 10⁻⁴ M) in water. Then, three independent measurements of the phosphorescence intensity of **Re-Phe-AM2** were made in the presence of various amounts of heparin, and the mean intensity was plotted as a function of heparin concentration. The slope (m) of the straight part of the curve was calculated. The LOD was finally determined using equation:

$$ LOD = 3\sigma/m \qquad (2) $$

Samples for transmission electron microscopy (TEM) were prepared by the drop casting method onto 3.05 mm copper grids with a 400-mesh density (supplied by Pelanne Instruments). These grids were pre-coated with a collodion film approximately 20-50 nm thick. TEM images were recorded on a MET JEOL JEM 1011 instrument. The hydrodynamic size was measured by dynamic light scattering (DLS), using a Zetasizer NanoZ device (Malvern instruments).

# 3. Results and discussion

## 3.1. Synthesis and characterization

Complex **Re-Phe** was synthesized as previously described [25]. The ionic complexes (**Re-Phe-MP**, **Re-Phe-AM** and **Re-Phe-AM2**) were obtained by effective synthesis starting from a common pyridine-oxadiazole scaffold [25] (Scheme 1). A condensation step with tert-butyl(4-aminophenethyl)carbamate [55] in refluxing xylenes with a catalytic amount of p-toluenesulfonic acid led to the corresponding pyridinyl-1,2,4-triazole (in 47% yield), which afforded the corresponding tricarbonylrhenium complex **ReL2** in 86% yield, after reflux in methanol in the presence of the commercially available precursor [Re(CO)₅Cl]. Boc protection was cleaved under classical TFA conditions to give the free amino complex. This latter was then reacted with HCl 4M in dioxane (metathesis reaction) to afford the expected complex **Re-Phe-AM** in quantitative yield.

The preparation of the ligands required for the synthesis of complexes **Re-Phe-MP** and **Re-Phe-AM2** began with the condensation of 2-(4-aminophenyl)ethanol with pyridineoxadiazole under the conditions described above. The newly synthesized pyridinyl-1,2,4-triazole-phenylethanol was reacted with mesyl chloride to give compound **1** by converting the -OH group to the corresponding mesyl (–OMs) substituent. For the preparation of **Re-Phe-MP**, a subsequent substitution of the mesyl group with N-methylpyrrolidine yielded the expected ligand **L1**, which was used in the ultimate complexation step to afford the corresponding complex with 98% yield over the two steps. The preparation of **Re-Phe-AM2** required two additional steps. First of all, di-tert-butyl(azanediylbis(ethane-2,1-diyl))dicarbamate prepared from diethylenetriamine using a published protocole [58] was reacted with mesyl compound **1** to afford to the corresponding di-Boc-amino ligand **L3** (31% yield). The latter was reacted with [Re(CO)₅Cl] in refluxing methanol to give the corresponding rhenium(I) complex (70% yield). Boc-cleavage followed by HCl treatment as described for **Re-Phe-AM** afforded the expected **Re-Phe-AM2** complex in quantitative yield over the two steps. Detailed schemes of the synthesis are given in the ESI (Schemes S1–3†). The products were characterized by NMR and FT-IR spectroscopy, and HRMS (Fig. S1–25†). Unfortunately, it
was not possible to grow single crystals of sufficient quality for X-ray analysis despite a large number of attempts.

<figure><img src="image_5.png" /></figure>

**Scheme 1.** Synthesis of tricarbonylrhenium(I) complexes **Re-Phe-AM**, **Re-Phe-MP** and **Re-Phe-AM2**. Conditions and reagents: (a) *p*-Toluenesulfonic acid, xylenes, 140°C, 19 h; (b) Methanesulfonyl chloride, NEt₃, CH₂Cl₂, 0°C, 3 h; (c) *N*-Methylpyrrolidine, CH₃CN, 50°C, 17 h; (d) [Re(CO)₅Cl], MeOH, reflux, 19–23 h; (e) Trifluorocetic acid, CH₂Cl₂, rt, 45 min; (f) HCl (4M in dioxane), MeOH, rt, 45 min; (g) Di-*tert*-butyl(azanediylbis(ethane-2,1-diyl))dicarbamate, diisopropylethylamine, CH₃CN, reflux, 3 days.

### 3.2. Spectroscopic properties of the ionic complexes

For the sake of comparison, the spectroscopic properties of **Re-Phe** in acetonitrile were characterized (Table 1). The three ionic complexes were studied in water (Fig. S26† and Table 1). Since the addition of a carbon chain on the **Re-Phe** framework does not alter the conjugated electron system, the spectroscopic properties of the ionic complexes must be close to those of **Re-Phe**, which were previously analyzed in CH₂Cl₂ based on DFT calculations [25]. For instance, the intense high-energy band of the absorption spectra may be attributed to a combination of intra-ligand charge transfer (ILCT) and metal-to-ligand charge transfer (MLCT) transitions, while the low-energy band of moderate intensity, peaking at 340 nm, has strong MLCT character. Small variations in absorptivity may be explained by differences in solubility and aggregate formation at the concentrations used.

The three complexes emitted weak red light when illuminated by a UV lamp. The emission spectra, attributed to phosphorescence, showed a single unresolved band centered at 610 nm (Fig. S26†), with quantum yields around 6.0 × 10⁻³. The emission decays were
biexponential (Fig. S27†). The very widely predominant lifetime, attributed to dissolved complexes, was 14 ns. A lifetime of more than 100 ns confirmed the presence of a few aggregates. Bubbling the complex solutions with argon for 4 min hardly changed the emission intensity, showing that dissolved oxygen had no significant quenching effect, as can be expected for compounds with a relatively short lifetime. In comparison, the ionic complexes in water emitted ten times less than **Re-Phe** in acetonitrile, and had a shorter lifetime. This effect may result from quenching by water molecules [59].

While the complexes showed exactly the same emission in water, their microcrystalline powders emitted light ranging from yellow-green (**Re-Phe-MP**) to yellow-orange (**Re-Phe-AM** and **Re-Phe-AM2**), probably due to crystal packing effects. The emission spectra showed a single band peaking between 538 and 578 nm (Fig. 2 and Table 1). The quantum yields were rather low, between 0.12 and 0.06, and decreased with the red shift of emission spectra, in agreement with the energy gap law [60]. The SLE effect was clear in all cases. All decays were found to be multiexponential (Fig. S28†). The predominant contribution was associated to lifetimes longer than 700 ns, and attributed to bulk molecules. The second significant contribution, with lifetimes shorter than 200 nanoseconds, can be attributed to molecules experiencing a different environment, for example on the surface or near crystal defects. The molecules are therefore well stabilized in the solid relative to the solutions. However, compared to **Re-Phe**, the much lower quantum yields of ionic complexes suggest that the molecular arrangement driven by the electric charges induces intermolecular interactions detrimental to emission.

<figure><img src="image_6.png" /><figcaption>**Fig. 2** Normalized photoluminescence spectra of the microcrystalline powders of complexes **Re-Phe-MP**, **Re-Phe-AM** and **Re-Phe-AM2**. $\lambda_{ex} = 380$ nm.</figcaption></figure>
<table><thead><tr><th rowspan="2">Compound</th><th colspan="5">Solutions <sup>a</sup></th><th colspan="3">Microcrystalline powders</th><th colspan="3">Suspensions <sup>b</sup></th></tr><tr><th>λ<sub>abs</sub><br>(nm)</th><th>ε<br>(M<sup>-1</sup>cm<sup>-1</sup>)</th><th>λ<sub>P</sub><br>(nm)</th><th>Φ<sub>P</sub></th><th>τ<sup>c</sup><br>(ns), (f), [χ<sup>2</sup>]</th><th>λ<sub>PL</sub><br>(nm)</th><th>Φ<sub>PL</sub></th><th>τ<sub>PL</sub><sup>d</sup><br>(ns), (f), [χ<sup>2</sup>]</th><th>λ<sub>PL</sub><br>(nm)</th><th>Φ<sub>PL</sub></th><th>τ<sub>PL</sub><sup>e</sup><br>(ns), (f), [χ<sup>2</sup>]</th></tr></thead><tbody><tr><td rowspan="3">Re-Phe-MP</td><td>240</td><td>13000</td><td rowspan="3">608</td><td rowspan="3">0.006</td><td>14 (90.8)</td><td rowspan="3">538</td><td rowspan="3">0.12</td><td>9 (0.6)</td><td rowspan="3">586</td><td rowspan="3">0.02</td><td>15 (13.9)</td></tr><tr><td>280</td><td>9800</td><td>153 (9.2)</td><td>174 (13.3)</td><td>76 (30.4)</td></tr><tr><td>340</td><td>4600</td><td>[1.17]</td><td>775 (86.1)</td><td>295 (55.7)</td></tr><tr><td rowspan="3">Re-Phe-AM</td><td>240</td><td>12000</td><td rowspan="3">610</td><td rowspan="3">0.005</td><td>14 (97.5)</td><td rowspan="3">570</td><td rowspan="3">0.09</td><td>11 (1.1)</td><td rowspan="3">578</td><td rowspan="3">0.03</td><td>16 (12.2)</td></tr><tr><td>280</td><td>9400</td><td>104 (2.5)</td><td>193 (24.1)</td><td>84 (27.9)</td></tr><tr><td>340</td><td>4400</td><td>[1.16]</td><td>760 (74.8)</td><td>291 (59.9)</td></tr><tr><td rowspan="3">Re-Phe-AM2</td><td>238</td><td>14600</td><td rowspan="3">610</td><td rowspan="3">0.006</td><td>14 (94.8)</td><td rowspan="3">578</td><td rowspan="3">0.06</td><td>15 (1.4)</td><td rowspan="3">574</td><td rowspan="3">0.06</td><td>9 (6.5)</td></tr><tr><td>280</td><td>10600</td><td>165 (5.2)</td><td>190 (24.1)</td><td>86 (18.5)</td></tr><tr><td>340</td><td>5100</td><td>[1.12]</td><td>781 (74.5)</td><td>367 (74.9)</td></tr><tr><td rowspan="3">Re-Phe<sup>f</sup></td><td>234</td><td>13800</td><td rowspan="3">626</td><td rowspan="3">0.052</td><td>54</td><td rowspan="3">542<sup>g</sup></td><td rowspan="3">0.42</td><td>1 (0.2)</td><td rowspan="3">562</td><td rowspan="3">0.50</td><td>16 (1.3)</td></tr><tr><td>278</td><td>9800</td><td>[1.43]</td><td>60 (2.3)</td><td>363 (98.7)</td></tr><tr><td>366</td><td>3600</td><td></td><td></td><td>639 (97.5)</td><td>[1.32]</td></tr></tbody></table>

a: Water for the ionic complexes, acetonitrile for **Re-Phe**. Concentrations around 7 × 10⁻⁵ M for absorption and 1–2.5 × 10⁻⁵ M for emission. b: Heparin 45 µg/mL in water for the ionic complexes (1 × 10⁻⁴ M); water/acetonitrile 90:10 v/v for **Re-Phe** (6.4 × 10⁻⁵ M). c: Decays are given in Fig. S27†. d: Decays of ionic complexes are given in Fig. S28†. e: Decays are given in Fig. S34†. f: The Φ<sub>PL</sub> and τ<sub>PL</sub> values of the microcrystalline powder are from ref. [25]. g: This work.

**Table 1.** Spectroscopic data of complexes in solution, as suspensions and in the solid state. Maximum absorption wavelength (λ<sub>abs</sub>), molar extinction coefficient (ε), maximum wavelength of emission (λ), emission quantum yield (Φ), lifetime (τ) with relative amplitude (f) in % and chi square (χ²) values. Subscripts P and PL refer to phosphorescence in solution and photoluminescence in suspensions and in the solid state.
### 3.3. Photochemical stability

Acetonitrile and water are coordinating solvents that could replace one of the carbonyl ligands in rhenium complexes, especially after photochemical reaction [61, 62]. **Re-Phe** in acetonitrile and **Re-Phe-AM** in water were tested for stability by irradiating dilute solutions with a 385 nm emitting diode for 8 h. In both cases, no changes in the UV-vis spectra were detected, showing that both compounds are very stable chemically and photochemically in these media (Fig. S29†).

### 3.4. AIPE behavior of Re-Phe in the acetonitrile/water mixture

The AIPE behavior of **Re-Phe** was studied by increasing the proportion of water in acetonitrile solution, while maintaining the complex concentration constant. The corresponding absorption spectra were drastically modified. For a complex concentration of 6.4 × 10⁻⁵ M, they showed a blue shift of the MLCT band, with formation of a quasi-isosbestic point around 352 nm, as well as a decrease in absorbance and a broadening of the high energy bands. This evolution can be attributed both to a solvatochromic effect and to the formation of aggregates (Fig. 3a). It should be noted that a high complex concentration (1.3 × 10⁻⁴ M) is required for the absorptivity to collapse at high water proportions with a deviation of the baseline (Fig. S30†), due to the precipitation of microcrystalline agglomerates clearly visible under a fluorescence microscope. Samples examined under UV excitation showed that the light emitted gradually changed from faint red to bright yellow-green as the water proportion increased (Fig. 3b). The corresponding emission spectra were recorded by setting the excitation at 352 nm, so as to minimize variations in absorbance. They showed that the weak emission band initially centered at 626 nm became a strong emission band with a maximum at 562 nm (Fig. 3c). The intensity of the PL signal at this wavelength was multiplied by 20 (Fig. S31†). Bubbling the suspension formed in 90% water with argon for 4 min resulted in a 35% increase in PL intensity. This shows that the presence of dissolved oxygen has a clear influence on the PL efficiency of the particles, which have a long lifetime. The AIPE phenomenon can be explained by a combination of many factors. In all cases, the non-radiative deactivation processes, which are predominant in solution, are blocked during the transition to the solid state. In fact, in solution, the excitation energy of tricarbonylrhenium(I) complexes is mainly released by the small amplitude vibrations of carbonyl ligands and the bidentate ligand-metal framework. Vibrations of the surrounding solvent molecules also play an important role in deactivating excited states [63–65]. The rotation of the phenyl group around the single bond could also contribute to energy dispersion [66]. In aggregate and microcrystalline states, rigidity prevents the vibrations and intramolecular motions necessary to reach the numerous conical intersections between excited and ground states [67]. The access of dissolved oxygen molecules, which can quench the emission, is also restricted. Finally, the crystal packing mode that leads to very little π-π stacking between neighbouring molecules is very favourable to a good phosphorescence
emission. The resulting strong AIPE effect justifies the choice of the **Re-Phe** scaffold for
subsequent applications.

<figure><img src="image_7.png" /><figcaption>**Fig. 3** a) Absorption spectra of **Re-Phe** at 6.4 × 10⁻⁵ M in acetonitrile solutions containing from 0 to 90% water; b) Corresponding samples (from left to right) illuminated at 365 nm; c) Evolution of the emission spectra for the same samples (λem = 352 nm, same color code as above).</figcaption></figure>

3.5. *Detection of heparin and chondroitin sulfate by the ionic complexes*

Heparin (Hep) or chondroitin sulfate (CS) were added till a concentration of 45 mg/mL to aqueous solutions of the three cationic complexes (1 × 10⁻⁴ M). The observations were made after stirring for 3 h at 20°C. The concentrated samples of **Re-Phe-AM2** became cloudy, those of the other two analogues were slightly opalescent. Fluorescence microscopy did not allow to distinguish the shape of luminescent particles in suspensions. The samples with the highest
amount of Hep were examined by transmission electron microscopy (TEM), which clearly indicated the presence of many compact nanoparticles, 20-30 nm in diameter for **Re-Phe-AM2** more difficult to measure with precision for the other two complexes. (Fig. 4 and S32†). All particles tended to agglomerate. Dynamic light scattering revealed objects measuring 226 nm (**Re-Phe-AM2**), 308 nm (**Re-Phe-AM**) and 344 nm (**Re-Phe-MP**), which probably correspond to nanoparticle agglomerates (Table S1†). In the absence of Hep, no signal correlates with the quality criteria required for an analysis. In absorption spectroscopy, the shape of the spectra was unchanged when the polysaccharide concentration increased, but a progressive deviation of the baseline confirmed the presence of aggregates (Fig. S33†). It was decided to excite at 390 nm, so that the absorbance was low enough to allow reliable emission measurements. The PL of all samples was more intense and shifted towards blue as the concentration of sulfonated polysaccharides was increased (Fig. 5a–g). The main lifetimes were much longer, typical of solid-state emissions (Fig. S34†). A clear AIPE effect was therefore observed for the three complexes in the presence of Hep and CS. The most spectacular effect was observed using double-charged **Re-Phe-AM2** and Hep. In this case, the blue shift was 36 nm and the intensity of the PL signal at 574 nm was multiplied by ~13. The bicationic complex was also the most effective for CS detection. All complexes showed a stronger AIPE effect for Hep than for CS, whose charge density is lower (Fig. 5h). The three cationic complexes were also tested in the presence of hyaluronic acid sodium salt, a polysaccharide that only contains carboxylate anions, and dextran that is neutral. The variation in emission intensity was one third of that observed with Hep in the first case, and not significant in the second one. All these observations show that the AIPE effect obtained with Hep and CS is indeed due to strong electrostatic interaction. In addition, the ammonium-substituted complex **Re-Phe-AM** provided better discrimination between heparin and CS than its *N*-methylpyrrolidinium counterpart **Re-Phe-MP**, which shows that the nature of the charged group also plays a role. Samples containing Hep and CS continued to evolve over time, resulting in more turbid and emissive suspensions, without any chemical or photochemical degradation. For instance, after three weeks, the emission intensity at maximum wavelength was multiplied by 6, 13 and 15 for **Re-Phe-MP**, **Re-Phe-AM** and **Re-Phe-AM2**, respectively, in the presence of 45 mg/mL Hep. The full growth of aggregates is therefore a slow process, as already reported in the literature for Re(I) complexes [9]. Finally, no emission change was detected by bubbling argon in a sample of **Re-Phe-AM2** with 45 mg/mL Hep, suggesting that the aggregates formed are not very sensitive to oxygen.

Compared to **Re-Phe** in the acetonitrile/water mixture, the maximum blue shift of emission of **Re-Phe-AM2** in the presence of Hep was halved (36 nm instead of 64 nm). The overall amplitude of the AIPE effect was quite close, but the quantum yield of the **Re-Phe-AM2** suspensions with heparin remained ten times lower than that of the **Re-Phe** particles formed in 90% water. Remarkably, the spectroscopic properties of the aggregates formed in AIPE experiments reflect those of the corresponding microcrystalline powders (Table S1†). Except for **Re-Phe-MP** that interacts weakly with substrates, emission maxima and quantum yields are comparable. These observations suggest that the molecular arrangement could be quite close between the aggregates formed in water and the microcrystals. In contrast, the average lifetime of species generated in water was systematically equal to half that of
microcrystals, indicating that excited states are less stabilized in the first case. This can be
attributed to structural defects or to a less rigid environment. Remarkably, for this series of
compounds, the emission efficiency of microcrystalline powders is a good indicator of the
amplitude of the AIPE effect that can be expected when interaction with the substrate is
sufficient.

<figure><img src="image_8.png" /><figcaption>**Fig. 4.** TEM images of the aqueous suspension of **Re-Phe-AM2** (1 × 10⁻⁴ M) in the presence of 45 µg/mL heparin.</figcaption></figure>

3.6 Limit of detection and comparison with literature data

The limit of detection (LOD) of **Re-Phe-AM2** for heparin was determined to compare the
efficacy of our complex with that of other probes reported in the literature. As shown in Fig.
S35†, the emission intensity at 574 nm was increased linearly when the Hep concentration
ranged from 5 to 45 µg/mL with a correlation coefficient of 0.9915. The LOD measured by
taking into account this linear relationship and a noise level of three times the standard deviation
of a blank solution was found to be 5.25 µg/mL (from 0.65 to 2.62 µM since the MW of
enoxaparin may vary between 2000 and 8000 D). An overview of the recently reported results
on heparin detection using aggregation-induced emission procedures is presented in Table 2.
The comparison is difficult because LOD values are often given in molar concentration while
the type of heparin used is not specified, and molecular weight can vary greatly from batch to
batch. Many authors use high molecular weight heparin (HMWH). However, Wang *et al*. have
shown that HMWH readily forms aggregates in the presence of cationic probes, and leads to a
much better AIE response than low molecular weight heparin (LMWH) such as enoxaparin,
which was used in the present work [37]. Lee *et al.*, working with LMWH, found a LOD of
138.0 pM in aqueous solutions and 2.6 nM in serum sample. Obviously, the organic sensor used
by these authors has a detection threshold much lower than our complex. The recommended
therapeutic dose for Hep in cardiovascular surgery is 17–67 µM, and 1.7–10 µM for post-
operative and long-term care. Complex **Re-Phe-AM2** could be used in the first case for the
detection of Hep in clinical samples, assuming that the AIPE response is not too altered in
biological medium
<figure><img src="image_9.png" /><figcaption>**Fig. 5** a-f) Variation of the emission spectra ($\lambda_{ex} = 390$ nm) of the cationic complexes ($1 \times 10^{-4}$ M) in the presence of increasing amounts (0, 5, 10, 15, 20, 25, 30, 35, 40, and 45 µg/mL) of anionic polysaccharides: **Re-Phe-MP** with Hep (a) and CS (b), **Re-Phe-AM** with Hep (c) and CS (d), **Re-Phe-AM2** with Hep (e) and CS (f); g) Samples of **Re-Phe-AM2** with Hep under illumination at 365 nm; Comparison of the amplitude of the AIPE effect for the six samples.</figcaption></figure>
<table><caption>**Table 2.** Comparison with the results reported in the literature for the detection of heparin based on aggregation-induced emission. *ns*: not specified, R = Ratiometric. Some enhancement factors, not given in publications, were estimated.</caption><thead><tr><th>Sensor</th><th>Heparin</th><th>λ (nm)</th><th>Enhancement</th><th>LOD</th><th>Medium</th><th>Dynamic range</th><th>Ref.</th></tr></thead><tbody><tr><td>TPE/bis(butan-1-ammonium) dichloride</td><td>Sodium salt</td><td>480</td><td>x~15</td><td>1.53 ng/mL</td><td>10 mM Tris-HCl, pH = 6.0</td><td>36–180 ng/mL</td><td>34</td></tr><tr><td>TPE/tetra(ethane-1-amine)</td><td>Sodium salt</td><td>475</td><td>x22</td><td>35.89 ng/mL</td><td>10 mM Tris-HCl, pH = 6.0</td><td>0–2.0 µg/mL</td><td>35</td></tr><tr><td>TPE/pentane-1-trimethylammonium bromide</td><td>ns</td><td>421/497</td><td>x~8 (R)</td><td>20 nM</td><td>5 mM HEPES, pH = 7.4</td><td>0–16 µM</td><td>36</td></tr><tr><td>TPE/tetra(ethane-1-trimethylammonium) tetrabromide</td><td>Sodium salt from porcine intestine LMWH, enoxaparin sodium salt</td><td>475</td><td>x10<br>x4</td><td>0.75 µg/mL</td><td>5 mM HEPES, pH = 7.4</td><td>0–1.0 µg/mL</td><td>37</td></tr><tr><td>TPE/Polyfluorene/bis(propane-1-trimethylammonium) dibromide</td><td>ns, MW = 20 kD</td><td>563</td><td>x3.2</td><td>30 nM</td><td>2 mM HEPES, pH = 7.4</td><td>0–0.7 µM</td><td>38</td></tr><tr><td>TPE/Arg-Lys-Arg peptide</td><td>Sodium salt from porcine intestine</td><td>475</td><td>x100</td><td>138 pM<br>2.6 nm</td><td>Water<br>Serum</td><td>0–700 pM</td><td>39</td></tr><tr><td>TPE/AG73 peptide</td><td>Unfractionated from Hog intestine MW = 644.2 g/mol</td><td>470</td><td>x31</td><td>38 ng/ML</td><td>HEPES/DMSO (95:5 v/v)</td><td>0.064–0.32 µg/ML</td><td>40</td></tr><tr><td>TPE/pyridinium Iodide</td><td>ns</td><td>575</td><td>x36</td><td>4 nM</td><td>10 mM HEPES, pH= 7.0</td><td>0–4 µM</td><td>41</td></tr><tr><td>TPE/tetra(pyridinium) tetrabromide</td><td>Sodium salt</td><td>565</td><td>x~47</td><td>6.9 nM</td><td>Water, pH = 7</td><td>0–22 µM</td><td>42</td></tr><tr><td>TPE/triazolium (BF<sub>4</sub><sup>-</sup> salt)</td><td>ns</td><td>486</td><td>x~100</td><td>30 ng/mL</td><td>10 mM Tris-HCl, pH = 6.0</td><td>0–1200 ng/mL</td><td>43</td></tr><tr><td>N,N'-bis[4-[[3-(trimethylammonio) ethyl]oxy]salicylidene]ethylenediamine dibromide (BTASE)</td><td>Sodium salt 150 U/mg</td><td>530</td><td>x40</td><td>57.6 ng/ML</td><td>10 mM Tris-HCl, pH = 7.0</td><td>0.2–14 µg/mL</td><td>44</td></tr><tr><td>Tetraphenylsilole/ propane-1-trimethylammonium Iodide</td><td>ns</td><td>480</td><td>x92</td><td>23 nM</td><td>5.0 mM HEPES, pH = 7.4</td><td>0–10 µM</td><td>45</td></tr><tr><td>4-[3-Methyl-2(3H)-benzoxazolylidene) methyl]-1-[3-(trimethylammonio) propyl]-quinolinium diiodide (YO-PRO-1)</td><td>Sodium salt</td><td>580</td><td>x55</td><td>0.48 nM</td><td>Water</td><td>0–1600 nM</td><td>46</td></tr><tr><td>Tetraphenyl imidazole/ 2,3-dimethyl-benzo[d]thiazol-3-ium iodide (TIBI)</td><td>ns</td><td>650</td><td>x~60</td><td>0.08 µM</td><td>10 mM PBS, pH = 7.4</td><td>0–10 µM</td><td>47</td></tr><tr><td>Thioflavin T chloride salt</td><td>Sodium salt from porcine intestine</td><td>560</td><td>x45</td><td>18 nM</td><td>Water, pH = 4.5-9</td><td>0–15 µM</td><td>48</td></tr><tr><td>Pyrene/(Arg)<sub>3</sub>NH<sub>2</sub></td><td>Sodium salt from porcine intestine</td><td>484/376</td><td>x5947 (R)</td><td>36 pM<br>299 pM</td><td>10 mM HEPES, pH = 7.4<br>+ 5% human serum</td><td>0–900 pM<br>0–2400 pM</td><td>49</td></tr><tr><td>Pyrene/Arg-Lys-Arg peptide</td><td>Sodium salt from porcine intestine</td><td>480/396</td><td>x1600 (R)</td><td>2.46 nM<br>3.43 nM</td><td>10 mM HEPES, pH 7.4 + 1% DMSO<br>+5% serum sample</td><td>ns<br>0–36 nM</td><td>50</td></tr><tr><td>Pyrene/benzimidazolium dibromide (PYNN)</td><td>ns</td><td>580</td><td>x8</td><td>6.1 ng/mL</td><td>10 mM HEPES, pH = 7</td><td>0–6 µg/mL</td><td>51</td></tr><tr><td>Pyrene/bis(methylimidazolium) dibromide</td><td>Sodium salt from hog intestine</td><td>511/385</td><td>x~12 (R)</td><td>8.5 nM (153 ng/mL)</td><td>10 mM HEPES, pH = 7.4</td><td>10–100 nM</td><td>52</td></tr><tr><td>Pyrene/bis(imidazolium) dibromide</td><td>Sodium salt from hog intestine</td><td>485/396</td><td>x28.6 (R)</td><td>9.5 nM (6.3 ng/mL)</td><td>5 mM Tris-HCl, pH = 7.4</td><td>0–1 µM</td><td>53</td></tr><tr><td>[Os(1,10-phenanthroline)<sub>2</sub>CO(4-phenyl-pyridine)](PF<sub>6</sub>)<sub>2</sub></td><td>Sodium salt >180 units/mg</td><td>560</td><td>x2.3</td><td>~1.0 µg/mL</td><td>Water</td><td>1.0–14.5 µg/mL</td><td>54</td></tr><tr><td>Rhenium(I)complex Re-Phe-AM2</td><td>Enoxaparin sodium salt</td><td>574 nm</td><td>x~13</td><td>5.25 µg/mL</td><td>Water</td><td>5 to 45 µg/mL</td><td>This work</td></tr></tbody></table>
## 4. CONCLUSIONS

This study showed that our ionic rhenium(I) complexes generate a marked AIPE effect for the detection of heparin. Discrimination against chondroitin sulfate is an asset. However, these complexes are less efficient than some organic AIE-probes. Conceptually, the substituted analogues (**Re-Phe-MP**, **Re-Phe-AM** and **Re-Phe-AM2**) do not show the good solid-state emission properties of their parent complex **Re-Phe**, as our group has already noted for other complexes [31]. Indeed, for this type of compounds, photoluminescence properties are highly dependent on intermolecular interactions, which vary with the slightest structural change and are difficult to predict. The reduction intramolecular motions is not sufficient to ensure a good enhancement in emission, as may be the case for organic probes. However, the PL quantum yields of our aggregated complexes are one to two orders of magnitude higher than those reported in the literature for Re(I) complexes used in AIPE applications [8–10]. The excellent photochemical stability of our compounds allows for prolonged observation and their simple design lends itself well to various chemical modifications, in order to play on recognition groups and solubility. This study validated their use as AIPE-probes and highlighted important structural criteria. Further developments would be needed to obtain highly effective complexes that deserve being tested under biological conditions.

### Acknowledgements

We thank Dr. Alexandre Poirot and Dr. Nadine Leygue for the synthesis of **Re-Phe**, and ANR (#ANR-22-CE51-0023-04) for financial support.

### Author contributions: CRediT

C. Vanucci-Bacqué: Investigation, Writing - original manuscript; V. Guilbaud: Investigation, Reviewing and editing; C.-L. Serpentini: Investigation; F. Bedos-Belval: Writing - original draft, Reviewing and editing; E. Benoist: Reviewing and editing; S. Fery-Forgues: Conceptualization, Methodology, Investigation, Writing - original draft, Funding acquisition.

### Funding sources

Agence Nationale de la Recherche (ANR) (#ANR-22-CE51-0023-04)

## Appendix A. Supplementary data

Supplementary data to this article can be found online at...

### Conflicts of interest

There are no conflicts to declare.
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