Elena Osipova

Ekaterina Gulyaeva

Nikolay Kireev

Sergey Kovalenko

Christian Bijani

Yves Canac

Dmitry A Valyaev

Oleg Filippov

Natalia Belkova

Elena Shubina

P Co

H Ph

Fac -to-mer isomerization triggers hydride transfer from Mn( i ) complex fac -[(dppm

Fac-to-mer isomerization triggers hydride transfer from Mn(I) complex fac-[(Ph

Since pioneering works of Beller 1 and Milstein 2 in 2016 the design of well-defined Mn(I) catalysts for (de)hydrogenationtype processes 3 became one of the most popular directions in homogeneous catalysis using Earth-abundant transition metals. While the majority of these catalytic systems are still based on pincer-type architectures, 4 it was also shown that octahedral complexes fac-[(L-L')Mn(CO)3Br] bearing less structurally elaborated NN-, 5 PN-, 6 CN-, 7 PP-, 8 CP- 9 and CC-type 10 bidentate ligands could be competitive and even sometimes superior in terms of catalytic activity and/or substrate scope. It was generally admitted that hydride species fac-[(L-L')Mn(CO)3H] typically generated in situ from the corresponding Mn(I) bromide precursors are important intermediates in hydrogenation processes. [5][6][7][8][9][10]11 In some cases the occurrence of a direct hydride attack across the polar C=X bonds with a prior formation of non-covalent adducts (Chart 1, path (a)) was confirmed by DFT calculations. 6d,8b,10,12 We report herein the first experimental evidence that the hydride transfer process for such catalytically relevant facial Mn(I) complexes could be facilitated by their transient isomerization into the elusive meridional species mer-[(L-L')Mn(CO)3H] (Chart 1, path (b)).

In a continuation of our investigations on transition metal hydrides reactivity relevant for (de)hydrogenation catalysis, 13 Chart 1. Representation of possible hydride transfer routes from octahedral Mn(I) complexes fac-[(L-L')Mn(CO)3H] bearing bidentate donor ligands we turned our attention to Mn(I) derivatives, the hydride transfer behavior for which remains virtually unexplored. Indeed, kinetic hydricity for two simplest Mn(I) prototypes [(CO)5MnH] and cis-[( PPh3)Mn(CO)4H] was experimentally determined by Bullock in 1998, 14 whereas more recently Suresh used DFT calculations to estimate relative thermodynamic hydricity in a series of model compounds [(PH3)nMn(CO)5-nH] (n = 0-5) and [(H2P(CH2)3PH2)nMn(CO)mH] (n = 1, m = 3; n = 2, m = 1). 15 We have initially selected for this study readily available complex fac-[(dppm)Mn(CO)3H] (fac-1) recently postulated as an active intermediate in Mn-catalyzed Guerbet's reaction. 8a IR spectroscopy monitoring of the reaction between fac-1 and [Ph3C](BAr4) (1 eq, Ar = C6F5) in CH2Cl2 in 180-300 K temperature range (Figure S1) revealed the gradual transformation of νCO bands of the starting complex into those of the cationic Mn(I) product fac- [3](BAr4) (Scheme 1) as a result of hydride transfer and coordination of CH2Cl2 to the initially formed 16-electron intermediate fac- [2] + , the last step being energetically favorable (ΔG190K ca. -1.5 kcal/mol) according to DFT calculations. The structure of fac- [3](BAr4) [START_REF]All known formally 16-e cationic Mn(I) complexes with noncoordinating anions 16 are stabilized by C-H agosPc interaction(s), inexistent for fac-[2] + according to DFT data[END_REF][START_REF] Fang | For the precedent of such structure in Mn(I) series[END_REF] was confirmed based on the presence of three νCO bands at 2044, 1978, 1957 cm -1 as well as two characteristic Mn-CO resonances at δC 219.7 and 214.8 ppm in its IR and 13 C NMR spectra, respectively. Additionally, a broad signal of the coordinated CD2Cl2 at δC 62.5 ppm displaying a characteristic downfield shift 18 relative to that of free deuterated solvent (δC 53.8) was observed in 13 C NMR spectrum at 193 K. Interestingly, according to 1 H and 31 P NMR spectra Toulouse, Cedex 4, France. E-mail: dmitry.valyaev@lcc-toulouse.fr. † These authors contributed equally. Electronic Supplementary Information (ESI) available: complete experimental and computational parts, copies of variable temperature IR and NMR spectra. Scheme 1. Initially proposed reaction pathway between hydride complex fac-1 and [Ph3C](BAr4) (top) and the unconventional route identified by low temperature NMR spectroscopy (down). BAr4 -counter anions are not shown for clarity.

fac- [3](BAr4) exists as 4:1 mixture of two isomers (Figures S13-14). DFT calculations revealed that these species differ only by the orientation of MnPCP metallacycle relative to the CH2Cl2 ligand being defined as fac,syn-and fac,anti- [3] + (Figure S41).

In-depth analysis of IR data revealed the formation at very low temperature of additional metal species with νCO 2063 and 2049 cm -1 converting at 220 K to the final product fac- [3](BAr4). The use of nBuCl as a solvent allowing to perform the reaction at 160K (Figure S2) led to a higher relative amount of this intermediate. Intrigued by the nature of this intermediate exhibiting even higher νCO values than those of cationic complex fac- [3](BAr4), we have performed a low-temperature NMR study in CD2Cl2 solution. 1 H and 31 P NMR spectra of 1:1 mixture of fac-1 and [Ph3C](BAr4) showed that hydride transfer was over within 10-15 min at 183K affording two cationic species in ca. 60:40 ratio, the major one being characterized by a singlet at δP 10.2 ppm and the minor one by two doublets at δP 10.6 and 7.1 ppm with a 2 JPP coupling constant of 44.4 Hz (Figure S16). While the main product was identified as fac- [3](BAr4) based on comparison with NMR data of authentic sample, the presence of two inequivalent 31 P nuclei and three distinct Mn-CO resonances at δC 218.9, 218.1, and 213.6 ppm led us to propose that the minor species is defined by a meridional geometry (Scheme 1). Strikingly, 1 H spectra of this product mer-[4](BAr4) revealed a CH2 moiety appearing as two doublets at δH 2.23 and 2.06 ppm with a geminal 2 JHH constant of 24.6 Hz as well as four others upfield CH resonances at δH 6.44, 5.78, 5.51 and 4.41 ppm (Figure S17) in favor of the dearomatization of a phenyl group further confirmed by 13 C NMR experiments (see Figures S22-23 for details). These results represent, to the best of our knowledge, the first evidence for the nucleophilic attack at the para-position in Ar3C + carbocations 19 due to steric contraints being conceptually similar to the reactivity of Ar3C • radicals. [START_REF] Tidwell | Stable radicals[END_REF] Upon warming to 203K mer-[4](BAr4) rapidly transformed into fac- [3](BAr4) (Scheme 1) by a sequence of alkene ligand dissociation and mer-to-fac isomerization being finished at 213 K. The concomitant formation of cyclic diene 5 isomerizing to Ph3CH above 213 K was also observed (Figure S25). Its structure was proposed based on the observation of three signals in 1 H NMR spectrum at δH 6.44, 5.88 and 3.07 ppm being similar to those reported in literature for structurally related products. [START_REF] Chen | [END_REF] The spectroscopic detection of cationic mer-[4](BAr4) as a kinetic reaction product may suggest the possible participation of meridional Mn-H derivaPves in hydride-ion transfer from fac-1. Only a few examples of Mn(I) halide complexes mer-[(L-L')Mn(CO)3X] (X = Cl, Br) bearing bidentate ligands were reported in literature 22 due to their lower thermodynamic stability compared to the corresponding facial isomers. In contrast, structurally similar Mn(I) mer-hydride species -[(L-L')Mn(CO)3H] have been never characterized and only recently proposed to be involved in syn-to-anti isomerization of bridge-substituted dppm complexes fac-[(dppm R )Mn(CO)3H] based on DFT calculations. 23 With the aim to characterize these elusive compounds, we decided to use weaker Lewis acid B(C6F5)3 (BAr3) 24 as a hydride abstractor. Gratifyingly, lowtemperature IR spectra of fac-1 in the presence of BAr3 (1.5 equiv.) in CH2Cl2 (Figure 1) evidenced not only the characteristic νCO bands at 2063 and 2003 cm -1 belonging to cationic mer- [3](BHAr3) complex, but also two new νCO bands rapidly disappearing upon warming to 220 K. Band deconvolution allowed us to resolve these two bands at 2049 and 1993 cm -1 (Figure S3). Taking into account relatively elevated νCO values, these bands were tentatively attributed to the non-covalent adduct of meridional hydride with BAr3 mer-1•••BAr3 (vide infra). Low-temperature IR data analysis allowed to estimate experimentally both thermodynamic (ΔH° = -3.3 ± 0.5 kcal/mol, ΔS° = 5.5 ± 0.1 cal/(mol•K) and kinetic (∆H ≠ = 9.4 ± 0.5 kcal/mol, ∆S ≠ = -26 ± 2 cal/(mol•K)) parameters for hydride transfer (see the ESI for details). Importantly, only traces of facial cation fac-[3](BHAr3) were observed at 180 K thus showing that hydride transfer proceeds mainly from meridional hydride derivatives. [START_REF]Similar phenomenon was observed by Bullock for Mo(II) hydride complexes with four-legged piano-stool geometry revealing faster hydride transfer for trans-[Cp(CO)2Mo(PR3)H] compared to cis[END_REF] Low-temperature NMR data in CD2Cl2 solution further corroborated the existence of non-covalent adducts in the reaction mixture. 1 H NMR spectra (Figure S26) contained a broad signal at δH -6.44 ppm with a characteristic shielding from the resonance of free hydride fac-1 (δH -5.64 ppm) 26 and not affected by 31 P and 11 B decoupling. In 31 P spectra the formation of two species with chemical shifts close to fac-1 (δP 32.1 ppm) was observed: a singlet at δP 29.1 ppm and an AB spin system appearing as two doublets at δP 30.7 and 23.2 ppm with a 2 JPP coupling constant of 77.5 Hz (Figure S29). These spectral data are consistent with the simultaneous observation of Lewis acid adducts of fac-and mer-isomers of Mn(I) hydride fac-1•••BAr3 and mer-1•••BAr3, respectively. In contrast to IR data (Figure 1) significant amount of the final hydride transfer product fac- [3](BHAr3) was detected by 31 P NMR even at 183 K, whereas no signals of cationic mer-species were observed under these conditions. This discrepancy may be rationalized by much higher concentration of the NMR sample (ca. 30 times) leading unavoidably to the acceleration of reaction kinetics and/or the occurence of dynamic phenomena (vide infra). However, from 233K two new broad doublets at δP 45.8 and 10.6 ppm with 2 JPP of ca. 40 Hz disappearing above 273 K were observed (Figure S30), which may be assigned to the meridional cationic intermediates mer- [3](BHAr3) or mer- [2](BHAr3). According to 1 H NMR data, at 233 K fac- [3](BHAr3) starts to dissociate a labile CH2Cl2 ligand to produce fac- [2](BHAr3) stabilized by B-H bond interaction with the cationic metal center (Figure S28). This species becomes predominant above 273 K (see the ESI).

To get a deeper insight into the mechanism of the hydride transfer process we have performed the theoretical study of the reaction between fac-1 and BAr3 in CH2Cl2 at ωB97XD/def2-TZVP level of calculations using SMD continuum solvation model. The results are summarized in Scheme 2 and calculated νCO frequencies for main stationary points are listed in Table 1.

While the meridional isomer of Mn(I) hydride complex 1 was found less thermodynamically stable than its facial one (Scheme 2) in agreement with literature data, 23 the opposite trend was observed for the BAr3 non-covalent adducts showing Scheme 2. The mechanism of hydride transfer from Mn(I) complex fac-1 to BAr3 based on DFT calculations (wB97XD/def2-TZVP level, the corresponding ∆G298К/∆G190К values in kcal•mol -1 are shown in blue/red).

Table 1. Calculated (wB97XD/def2-TZVP level) and experimental νCO frequencies and 31 P NMR data (when available) for both facial and meridional Mn(I) hydrides 1, their noncovalent adducts with BAr3 (1...BAr3) and cationic complexes [3](BHAr3).

that mer-1•••BAr3 is preferred in terms of ∆G vs. both fac-1•••BAr3 and starting hydride fac-1 both at RT and 190 K (Figures S43). This fac/mer stability pattern reverts for naked 16-electron cations [2] + (see the ESI) and corresponding CH2Cl2 complexes [3](BHAr3), in which facial forms become again strongly thermodynamically preferable (Figure S39 and S41). Though calculated νCO values for fac-1 and mer-1 are very close (Table 1) varying only in the relative band intensities (Figure S38), the difference of spectral properties for other fac-and mer-species is more notable making possible their identification in experimental IR spectra. In particular, the highest frequency νCO values in mer-1•••BAr3 and mer- [3](BHAr3) are greater by ca. 20 and 30 cm -1 than those of their facial analogues being therefore fully consistent with our original attribution of different Mn(I) intermediates in low-temperature IR spectra on Figure 1.

Based on combined spectroscopic and theoretical data we propose that the dominant pathway for the reaction between complex fac-1 and BAr3 goes via the initial formation of the noncovalent adduct with borane fac-1•••BAr3, which then transforms to more stable meridional intermediate mer-1•••BAr3. The alternative route involving initial fac-to-mer isomerization of complex 1 before the interaction with Lewis acid can also be possible, albeit to a minor extent considering the relatively small amount of the meridional hydride in the equilibrium mixture. It thus appears that, the Mn-H moiety in mer- [1] •••BAr3 has more hydridic character than in fac-1•••BAr3 facilitating hydride transfer to form mer- [3](BHAr3) complex as a kinetic product. Its transformation to a more stable fac- [3](BHAr3) seems to proceed with higher activation barrier than those for neutral hydride compounds 22 permitting the accumulation of cationic meridional species at very low temperature and concentration level. Finally, the absence of isotopic effect in the reaction between the deuteride derivative fac-1-d3 and BAr3 (kH/kD = 0.9 ± 0.1 see the ESI) shows that Mn-H bond cleavage is not likely to be involved in the ratedetermining step giving further arguments that fac-to-mer isomerization plays a crucial role in the hydride transfer process.

In conclusion, we have demonstrated an intrinsic mechanistic complexity for the a priori trivial hydride abstraction from Mn(I) complex fac-[(dppm)Mn(CO)3H] and evidenced that the formation of elusive mer-hydride species having increased kinetic hydricity represents a key step of this process. Considering that the majority of organic substrates have lower hydride abstraction ability than Lewis acids used in this study, the possible implication of barely detectable but more reactive mer-hydride derivatives in hydrogenation-type reactions catalyzed by similar octahedral Mn(I) complexes fac-[(L-L')Mn(CO)3X] could indeed create alternative pathways with lower activation energies. In light of these results, this option has to be systematically checked in DFT investigations of the relevant catalytic cycles at least for those involving external sphere hydride transfer steps.

This work was supported by the Russian Science Foundation (grant no. 20-73-00311) and by the CNRS. Computational studies were performed using HPC resources from CALMIP (grant no. P18038). E. S. G. is grateful to French Embassy in Moscow for a joint PhD fellowship (Vernadski program). The Complex Calcd. νCO (cm -1 ) Exp. νCO (cm -1 ) δP (ppm) fac -1 1996, 1918, 1914 1993, 1911, 1902 32.1 mer-1 1999, 1918, 1914 -fac-1•••BAr3 2037, 1974, 1959 -29.1 mer-1•••BAr3 2056, 1975, 1968 2049, 1993 30.7, 23.2 fac- [3](BHAr3) 2044, 1984, 1979 (anti) 2043, 1982, 1974 (syn) 2040, 1973, 1935 11.2, 10.1 mer-[3](BHAr3) 2069, 1997, 1977 2063, 2003 45.8, 10.6 NMR data were in part collected using the equipment of the Center for Molecular Composition Studies of INEOS RAS with the support from the Ministry of Science and Higher Education of the Russian Federation.

2 PCH 2

 22 PPh 2 )Mn(CO) 3 H] Elena S. Osipova, a † Ekaterina S. Gulyaeva, a,b † Nikolay V. Kireev, a Sergey A. Kovalenko, a Christian Bijani, b Yves Canac, b Dmitry A. Valyaev,* b Oleg A. Filippov,* a Natalia V. Belkova a and Elena S. Shubina* a Low-temperature IR and NMR studies combined with DFT calculations revealed the mechanistic complexity of apparently simple reactions between Mn(I) complex fac-[(dppm)Mn(CO)3H] and Lewis acids (LA = Ph3C + , B(C6F5)3) involving the formation of sofar elusive meridional hydride species mer-[(dppm)Mn(CO)3H•••LA] and unusual dearomatization of Ph3C + cation upon hydride transfer.

Fig. 1 .

 1 Fig. 1. IR spectroscopy monitoring of the reaction of complex fac-1 with 1.5 equiv. of BAr3 in CH2Cl2 at 180-300 K temperature range (c = 0.003 M).

Conflicts of interest

There are no conflicts to declare.

Notes and references


Elizaveta Ermakova

Oleg Raitman

Alexander Shokurov

Maria Kalinina

Sofiya Selector

Aslan Tsivadze

Vladimir Arslanov
email: vladimir.arslanov@gmail.com.

Michel Meyer

A Frumkin

Alla Bessmertnykh-Lemeune

Roger Guilard

A metal-responsive interdigitated bilayer for selective quantification of mercury( ii

Intensive research activities are currently devoted to the development of sensitive and selective sensors for toxic metal ions owing to the essential impact of these species on the environment and human health. Among them, mercury is one of the main targets considering its rapid bioaccumulation and high neurotoxicity. Accordingly, the maximal allowed concentration in drinking water is 10 nM. 1 Although a plethora of colorimetric and fluorescent chemosensors for Hg 2+ are currently available, 2 efficient, costeffective, and rapidly-responding solid-state sensors operating at sub-micromolar concentration levels are still rare. 3 Sensors based on the surface plasmon resonance (SPR) effect are particularly attractive because of their ultra-high sensitivity with detection limits in the nanomolar range. The signal transduction mechanism based on the change of the refractive index is therefore one of the most effective for investigating the interactions between a receptor (ligand) deposited as a film on a metallic surface and an analyte (substrate) injected into the contact solution. 4 Besides, SPR detectors possess many other advantages, such as the absence of any label, the compatibility with aqueous media, the possibility to miniaturize the sensing device and to equip it with other analytical probes (e.g. fiber optics, 5 electrochemical setup, 6 or amplifying effect of gold nanoparticles). 7 Moreover, it is a quantitative method despite the extremely small amount of materials (sensitive layer and analyte) required for the analysis. It is also worth to mention that SPR devices can be fitted with a microflow cell. In the continuous flow mode, the concentration of the analyte during the measurement remains constant, which is of particular interest from both a practical and theoretical point of view.

The design of selective SPR devices relies on the adequate functionalization of a gold layer, a task that can be easily achieved by attaching specific receptors endowed with a high affinity and selectivity for the targeted analyte. As host-guest recognition processes are ubiquitous in life-sciences (base-pairing, antibodyantigen interactions etc.), it is not wondering that SPR sensors have been applied overwhelmingly for monitoring the interactions between biological substrates (proteins, enzymes, nucleic acids etc.). 4a,8 Works on the detection of metal ions in aqueous media are more seldom and challenging. These SPR sensors often obtained by covering the gold surface with a layer of polymer incorporating molecular receptors (ligand or biomolecules) adapted to the target. 13 Likewise, gold-coated electrodes are commonly used in the most popular electrochemical detection of mercury. 91011 However, the efficiency of both types of detectors is limited by the slow diffusion of the analyte into the polymeric matrix and the restricted accessibility of the recognition sites. Additional issues might be low film adhesion, high film resistivity, and low chemical stability especially in case of biomolecules.

In that respect, deposition of self-assembled monolayers (SAM) is emerging as a promising alternative. 13b,14 Covalently-bonded SAMs incorporating molecular receptors or appropriate biomolecules located close to the interface and in proximity of the gold layer give rise to stable and selective responses upon analyte binding. However, the design of such films is challenging and major improvements are still sought to overcome the limitations intrinsic to SPR technology. Among them, the most important ones are (i) the low chemical stability of self-assembled monolayers (SAM) (cf. oxidation of thiol groups) and (ii) the non-selective interaction with impurities or analytes diffusing directly to the gold substrate as a consequence of the incomplete coverage of the surface 15 that leads to the presence of defects (pinholes) in the SAM. 14f,16 In addition, thiolated chelators inevitably contain, unlike alkylthiols, bulky functional groups which might perturb the alignment of the molecular chains, a prerequisite for achieving a highly-ordered coverage of the surface. 16a,17 Beside SAMs, Langmuir-Blodgett (LB) films can also be used as SPR sensors. However, multilayer LB films of voluminous or branched amphiphilic chelators exhibit usually a lower stability as they are washed away from the gold surface in the course of the analysis. 18 LB monolayers can be used instead, provided that they remain firmly fixed onto the solid support and that the ligand bears hydrophilic binding sites pointing away from the surface in direction to the analyzed solution. However, stable LB monolayers can only be obtained by the "bottom up" methodology, which supposes that some functional groups from the chelator are oriented towards the surface interacting with the gold atoms and are no longer available for metal binding, thus lowering the uptake efficiency.

Herein, an alternative approach for constructing the sensing film on the gold surface is described. We demonstrate that the immobilization of the chemosensor by combining both SAM and LB techniques 19 affords sensitive and selective metal-responsive SPR chips with enhanced stability and uniform orientation of the binding units towards the solution. This bilayer methodology is applied for the direct quantification of Hg 2+ cations at the trace level (ppt to ppb). To that aim, an interdigitated bilayer is constructed stepwise on a gold substrate by self-assembling first octanethiol into a rarefied monolayer. 19 Next, a second monolayer consisting of the amphiphilic and highly mercury-selective ligand L (Fig. 1A,B), 20 is deposited on the top of the SAM by means of the LB technique. The resulting interpenetrated architecture of the SAM/LB bilayer film differs from that of lipid membrane bilayers or Y-type multilayers (LB films and SAM/LB-films). Indeed, in the three latter cases the films consist of closely-packed hydrocarbon chains, but hydrophobic interactions between adjacent monolayers cannot provide the same high level of interlayer adhesion as the second LB-layer tends to be anchored on the first one by hydrogen bonds or electrostatic interactions. 21 In our approach, the weak van-der-Waals interactions between the hydrocarbon chains of the adjacent layers compete with the Au-S bonds. As a consequence, the alkanethiol molecules move laterally over the surface, 3b producing a beneficial reduction of pinholes in the film.

As a proof of concept, we report hereafter on the elaboration of a highly-sensitive and selective, but also reusable, mercury(II) sensor with a quantification limit close to 10 -11 M in aqueous media and a linear response spanning up to 5 orders of magnitude. Our rational design of the SPR chip rests on the use of the amphiphilic chemosensor L derived from 1,8-diamino-9,10-anthraquinone (Fig. 1A) as the solution-exposed component of the rarefied SAM/LB bilayer architecture. The choice of the chelator is based on the following criteria. We have recently shown that polyamino-9,10anthraquinones endowed with diethoxyphosphoryl-substituted carbamoyl fragments allow for cheap, fast, sensitive, and highly selective colorimetric detection of various metal ions in aqueous solutions at the ppm levels. 20,22 High affinity and selectivity towards Hg 2+ was found for chemosensor L and its water-soluble analogue L1 that lacks the lipophilic alkoxy substituents (Fig. 1A). Spectrophotometric, pH-metric, and NMR studies evidenced both the formation of mono-and dinuclear [Hg(L1)] (log K11 = 5.3) and [Hg2(L1)] complexes (log K21 = 4.0) and the involvement of the aminoanthraquinoyl moiety in the Hg 2+ binding scheme. 20 Highresolution ESI-MS measurements in methanol/water mixtures confirmed the [Hg(L1)] and [Hg2(L1)] stoichiometries deduced from UV-vis titrations. Structural information on these species was tentatively gained by 1 H and 31 P NMR spectroscopy in D2O. 20 Accordingly, the tertiary amines, one anthraquinone oxygen atom, the carbamoyl, and phosphoryl groups are most likely involved in coordination, in such a way that each polyamine branch binds a single metal ion. However, in the absence of a crystal structure, the exact binding mode of the amide moieties (Hg-O or Hg-N coordination) cannot be definitively ascertained. Ligation of the deprotonated amidic nitrogen atoms, as described for mercury complexes formed with cyclic amides, 23 would result in stable 5membered cycles and seems the most probable coordination mode in agreement with the pH-metric studies.

Noteworthy, chemosensor L retains its high affinity and selectivity towards Hg 2+ once incorporated in a Langmuir monolayer or LB multilayers. 24 Moreover, the strongly light-absorbing anthraquinone scaffold of L might also contribute to increase the sensitivity of the SPR system. 25 Bilayer films were prepared on freshly cleaned gold-coated glass slides by formation of an octanethiol SAM followed by the deposition of a monolayer of L by the vertical top-down LB method (ESI †). After each deposition step, the films were characterized by 2A. Upon covering the pure gold surface by a SAM of octanethiol, a significant decrease of the current together with a concomitant increase of the anodic and cathodic peak separation point out the existence of an array of pinhole defects in the microelectrode assembly. 26 For the Au/SAM/LB(L) electrode, only the non-faradic current is observed, indicating the formation of a compact and robust interdigitated bilayer (Fig. 2A, Fig. S2,ESI †). After its exposure to an aqueous 10 -11 M and 10 -6 M Hg(ClO4)2 solution, an additional decrease of the current intensity was observed, reflecting an even more regular and dense coverage of the metallic conductor by the insulating bilayer. It can be concluded that the lower current is a direct consequence of the structural changes undergone by the flexible receptor upon Hg 2+ binding. Becoming more rigid, the metal-bound ligands give rise to a denser packing, thus affording a more organized and protective layer. Noteworthy, the voltammograms for Au/SAM/LB(C) (C = Hg 2+ complex of L) were insensitive to exposure times ranging from 2 min to 1 h, revealing the high stability of the Au/SAM/LB(L) bilayer in water towards leaching.

To gain a deeper insight into the structure of Au/SAM and Au/SAM/LB(L) systems, they were also examined by electrochemical impedance spectroscopy (EIS) under the experimental conditions used for the CV studies. The Nyquist diagrams for bare and modified Au/SAM electrodes are shown in Fig. 2B and compared to those recorded for an Au/SAM/LB(L) chip before and after contact with an aqueous 10 -11 M and 10 -6 M Hg(ClO4)2 solution. They were analyzed according to the Randles equivalent circuit approach (Fig. S3, ESI †), [START_REF] Lvovich | Impedance spectroscopy: applications to electrochemical and dielectric phenomena[END_REF] the fitted parameters being reported in Table 1. It should be stressed that in case of naked gold, the frequency window was limited by the too high electron-transfer rate. As expected, the film thickness exerts a strong influence: the double-layer capacitance (Cdl) decreases whereas the charge-transfer resistance (Rct) increases, as evidenced by the larger left-sided Nyquist half-circles. Concomitantly, the Warburg impedance (W), which describes the diffusional impedance to/from a flat electrode, rises as the coating becomes thicker. The higher Rct value obtained for the Au/SAM/LB(L) bilayer supports the idea that interdigitation of both monolayers is accompanied with a defect-healing process that restricts the accessibility of the [Fe(CN)6] 3-/[Fe(CN)6] 4-anions to the surface. Moreover, exposure of Au/SAM/LB(L) to mercury(II) produces a two-fold increase of Rct, reflecting the robustness and rather high level of organization of the Au/SAM/LB(C) bilayer, as already evidenced by CV measurements. Assuming that the electron transfer between the redox couple and gold occurs only at pinholes, the electrode coverage () could be estimated (Eq. S1, ESI †). Data reported in Table 1 evidence a significantly higher coverage level of the surface upon formation of the bilayer, which almost totally insulates the gold electrode.

Contact angle measurements clearly pointed out the higher hydrophobicity of Au/SAM ( = 71°) and Au/SAM/LB(L) ( = 74°) in comparison to bare gold ( = 57°). Interestingly, Hg 2+ uptake by the Au/SAM/LB(L) bilayer does not affect the wetting properties of the film ( = 74.2° and 75°), in accordance with the formation of neutral complexes at the surface and the reorientation of the binding units upon complexation, as previously observed at the air/water interface. 24,[START_REF] Ranyuk | [END_REF] Analytical performances of Au/SAM/LB(L) chips were studied by using a commercial SPR spectrophotometer equipped with a microflow cell. In preliminary experiments, the stability of Au/SAM and Au/SAM/LB(L) was ascertained by observing a constant SPR signal when deionized water was pumped through the cell for up to 40 min at a constant flow rate of 100 L/min. A second control experiment involving an Au and Au/SAM chip ruled out Hg/Au amalgam formation on the gold surface, as well as unspecific adsorption of either Hg 2+ or ClO4 -ions when an aqueous Hg(ClO4)2 solution (c = 10 -11 -10 -6 M) flowed through the cell (Fig. S4-S5, ESI †). A typical SPR sensogram recorded for a Au/SAM/LB(L) chip for mercury(II) concentrations in the range 10 -11 -10 -6 M is reproduced in Fig. S6 (ESI †). It shows a regular signal growth with increasing mercury concentration. The sensor is sensitive enough to reliably determine mercury concentrations as low as 10 -11 M (2 ppt) in spiked solutions. The semi-log calibration function displayed in Fig. 3A evidences a linearity range spanning 5 decades.

The Hg 2+ binding kinetics by the Au/SAM/LB(L) chip was investigated by analyzing the sensogram data (Fig. S6, ESI †) with the classical pseudo-first-order Langmuir adsorption kinetic model (FLM) (Eq. S2, ESI †) that commonly applies to various types of analytes. 14f However, the nonlinear semi-log plot displayed in Fig. S8 (ESI †) definitively rules out this model. This mismatch can be related to one or several of the following factors: diffusion limitations, presence of more than one type of binding centers, or steric hindrance between immobilized receptors. 14c,29 In contrast, the pseudo-first-order Langmuir adsorption kinetic model with diffusion limitations (DLM) (Eq. S4, ESI †) fits properly the experimental data (Fig. S9, ESI †). To the best of our knowledge, this is the first time that a metal ion adsorption process on a thin film built in a microflow cell is found to follow the DLM model, although the adsorption of various alkanedithiols on gold were rationalized by this rate law too. 29b However, the reported value of the apparent rate constant (km) is about one order of magnitude higher (km = 0.13 s -1/2 at c = 5  10 -6 М) 29b than the one found herein (km = 0.018 s -1/2 at c = 10 -6 M). Interestingly, the km values summarized in Table S2 (ESI †), which approximately double when the Hg 2+ concentration increases from 10 -11 М to 10 -6 М, evidence no direct proportionality with the total metal concentration. This non-linear dependence of km vs. c has already been observed at several instances, for example in the case of various thiols adsorption. 30 As a matter of facts, the mercury uptake kinetics by the immobilized ligand L can be adequately described by the model that takes into account both adsorption and diffusion processes. Experimentally, the stationary regime is reached after some tens of minutes (Fig. S6,ESI †). Considering the diffusion coefficient of Hg 2+ in aqueous solution (D = 9.1  10 -6 cm 2 s -1 ), the mass-transfer should only take a few seconds and should not strongly depend on the metal-ion concentration. 31 Accordingly, the diffusion of Hg 2+ in the receptor sublayer with a thickness of about 1 nm cannot account on its own for the observed time-dependence of the SPR signal. Hence, the mass transfer is likely limited by the binding process, although it can also be argued that Hg 2+ is a very labile cation which was found to react extremely rapidly with a watersoluble analog of L that lacks both aliphatic chains. 20 However, socalled "surface effects" related to local changes in viscosity, permittivity, and desolvation rates at the interface, as well as molecular rearrangements within the bilayer may significantly contribute to slowdown the metal-ion uptake kinetics by the film. It turns out that the mercury sorption process is significantly slower than the adsorption of alkanethiols under similar experimental conditions. 29b To evaluate the reusability of the sensor and check for possible losses in sensitivity, regeneration tests were carried out in two steps after exposing the Au/SAM/LB(L) bilayer to a 10 -6 M Hg(ClO4)2 aqueous solution for 25 min. The chip was first washed by pumping for 25 min deionized water while monitoring the SPR signal. Then, the microflow cell was flushed for 5 min with 10 -2 M HCl. As shown in Fig. 3B, about 70% of the bound metal is washed away by the water flow with a pseudo-first-order dissociation rate constant of kdis = 8.2  10 -4 s -1 (Eq. S7, ESI †). This value, which is about eleven times lower than the one obtained for the dissociation of Cd 2+ /metallothionein surface complexes in similar SPR-experiments (kdis = 10 -2 s -1 ), 29d reflects the multidentate character of the receptor and the structural reorganization undergone by the top sublayer upon decomplexation. The partial elution of mercury by water, while complete desorption requires a wash with a dilute acid stream, suggests the occurrence of both weak and strong binding sites within the bilayer. Accordingly, the structural parameters of immobilized complexes are different from those observed in solution studies of ligand L1. It means that a significant fraction of Hg 2+ cations is only loosely bound, most likely to carbamoyl and/or phosphoryl oxygen atoms. A second fraction is more tightly entrapped by the polydentate N,O-receptor, as found in solution. 20 The acid-promoted dissociation of the chelate enables to fully recover the uptake capacity of the sensor and its sensitivity towards Hg 2+ . Furthermore, Fig. 3B highlights the excellent chemical stability and resistance towards leaching, as the analytical performances remain unaltered even after five adsorption/desorption cycles.

Finally, interference studies involving eight environmentally relevant cations, including toxic metals, were carried out (Fig. 3C). In these cross-selectivity experiments, equimolar amounts of perchlorate salts (c = 10 -5 M) were injected into the analyzed solution in the following sequence: Na + , Ca 2+ , Zn 2+ , Ni 2+ , Pb 2+ , Cd 2+ , Co 2+ , Cu 2+ , and Hg 2+ . Among them, only Cu 2+ produced a noticeable SPR signal. However, when Hg 2+ was spiked at the same concentration into the cocktail of the 8 other tested cations, a reliable response could be recorded, demonstrating an excellent selectivity of the Au/SAM/LB(L) chips.

In conclusion, a novel approach for preparing metal-responsive SPR chips is proposed, which takes advantage of the chemical diversity and stability provided by the interdigital arrangement of a SAM/LB bilayer deposited on a gold surface. This strategy has several advantages over the classical Au/SAM or Au/LB approaches, including the less-demanding synthesis of molecular receptors (i.e. no need of anchoring SH groups) as well as the formation of a denser, and thus more stable, film. The bilayer technology enabled to selectively quantify mercury(II) in aqueous solutions at the subnanomolar level (0.01 nM or 2 ppt) thanks to the outstanding coordination properties of the amphiphilic chemosensor L. The performances of this chip withstand the comparison with those of ICP-MS and are superior to any other optical sensor. Processing of the time-dependent SPR signals allows one to solve not only a purely analytical problem, but provides also some insights into the molecular interactions occurring in planar organized systems. It can also be emphasized that the efficient fabrication method described herein for obtaining SPR sensors might be also useful for elaboration of ion-selective electrodes, as evidenced by our impedance spectroscopic data.

Figure 1 .Figure 2 .

 12 Figure 1. Structure of chemosensors L and L1 based on the 1,8diaminoanthraquinone chromophore (A). Schematic representation of the interdigitated Au/SAM/LB(L) bilayer architecture (B).

Figure 3 .

 3 Figure 3. Semi-log calibration curve of the SPR response recorded for the Au/SAM/LB(L) chip in the presence of Hg 2+ in deionized water (A). Timedependent SPR signal change after the sequential exposure of the Au/SAM/LB(L) sensor to a 10 -5 M Hg(ClO4)2 solution in deionized water (rising curve), pure water (first stage of decline), and 0.01 M HCl (second stage of the decline) (B). Time-dependent SPR signal changes upon sequential addition of 8 metal ions and Hg 2+ (c = 10 -5 M) (C).

Table 1

 1 Parameters obtained for different film systems on a gold electrode

	from impedance plots
	Electrode	Hg 2+ (M) Rs () Rct (/cm 2 ) Cdl (F/cm 2 ) W ()  (%)
	Au		4.58	7.25	21.16	72.75	0
	Au/SAM		6.55	131.19	11.02	107.58 94.5
	Au/SAM/LB(L)		7.63	578.43	8.63	129.48 98.7
	Au/SAM/LB(С)	10 -11	6.78	707.40	8.43	408.80 99.0
	Au/SAM/LB(С)	10 -6	6.58 1761.10	6.58	799.50 99.4

† This work was carried out in the frame of the International Associated French-Russian Laboratory of Macrocycle Systems and Related Materials (LAMREM) of the Centre National de la Recherche Scientifique (CNRS) and the Russian Academy of Sciences (RAS). It was financially supported by the Russian Foundation for Basic Research (Project 12-03-93105), the RAS Presidium Program (P39), the CNRS, and the Conseil Régional de Bourgogne (Program PARI II CDEA). Electronic Supplementary Information (ESI) available: Experimental section and further characterization of the SAM monolayer and the SPR mercury(II) recognition process. See DOI: 10.1039/