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Dynamic combinatorial libraries (DCLs) are excellent benchmark models to study the stimuli-responsiveness of chemical networks. However, increasingly complex systems are difficult to analyze with simple data analysis methods, because many variables and connections must be considered for their full understanding. Here we propose the use of multivariate data analysis methods to bisect the evolution of a complex synthetic dynamic library of pseudopeptidic macrocycles, containing side chains with charges of different sign. Several stimuli (ionic strength, pH and the presence of a biogenic polyamine) were applied to the same dynamic chemical mixture, and the adaptation of the whole system was characterized by HPLC and analyzed with principal component analysis (PCA) and multivariate curve resolution-alternating least squares (MCR-ALS) methods. Both multivariate data analysis chemometric approaches are an excellent combination to extract both qualitative and semi-quantitative information about the adaptive process of the library upon the action of each stimulus. The resolution of the system with these chemometric tools proved to be especially useful when two inter-connected stimuli were combined in the same dynamic system. Our results demonstrate the utility of these two approaches for the analysis of complex dynamic chemical systems and open the way toward the application of these powerful tools in the emergent field of systems chemistry., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A straightforward four-step synthesis leads to the preparation of C 2 -symmetric dithiols containing a central aromatic core and amino acid side chains. These building blocks allow the preparation of dynamic covalent libraries of pseudopeptidic macrocycles in aqueous media that cover a broad range of polarities, functional groups and bulkiness mirroring the diversity found in natural peptides. The versatility of the generated dynamic libraries has been illustrated by the amplification of two different members from the same library upon the action of two biologically relevant templates.

Pseudopeptidic compounds are chemical species with attractive applications in many fields of chemistry. The increasing complexity in pseudopeptidic molecules has allowed improvements of their properties, very often after an initial bio-inspiration. Two main types of complexity can be proposed: structural and interactional. These operational processes have recently been used for the generation of new elaborated pseudopetidic molecules and complex molecular systems. This feature article presents a brief discussion of the recent advances done following this rational and with these privileged molecules.

Dynamic combinatorial libraries are powerful systems for studying adaptive behaviors and relationships, as models of more complex molecular networks. With this aim, we set up a chemically diverse dynamic library of pseudopeptidic macrocycles containing amino-acid side chains with differently charged residues (negative, positive, and neutral). The responsive ability of this complex library upon the increase of the ionic strength has been thoroughly studied. The families of the macrocyclic members concentrating charges of the same sign showed a large increase in its proportion as the ionic strength increases, whereas those with residues of opposite charges showed the reverse behavior. This observation suggested an electrostatic shielding effect of the salt within the library of macrocycles. The top-down deconvolution of the library allowed us to obtain the fundamental thermodynamic information connecting the library members (exchange equilibrium constants), as well as to parameterize the adaptation to the external stimulus. We also visualized the physicochemical driving forces for the process by structural analysis using NMR spectroscopy and molecular modeling. This knowledge permitted the full understanding of the whole dynamic library and also the de novo design of dynamic chemical systems with tailored co-adaptive relationships, containing competing or cooperating species. This study highlights the utility of dynamic combinatorial libraries in the emerging field of systems chemistry., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The molecular recognition of short peptides is a challenge in supramolecular chemistry, and the use of peptide-like cage receptors represents a promising approach. Here we report the synthesis and characterization of a diverse family of pseudopeptidic macrobicycles, as well as their binding abilities toward N-protected dipeptides using a combination of different techniques (NMR, ESI-MS, and fluorescence spectroscopy). The cage hosts were assayed for dipeptide binding using competition ESI-MS experiments as high-throughput screening to obtain general trends for the recognition phenomena. Selected hosts were additionally studied by NMR spectroscopy ((1)H NMR titration and diffusion-ordered spectroscopy experiments) in different solvents. The results unambiguously demonstrated the formation of the [cage·dipeptide] supramolecular complexes and rendered quantitative information about the strength of the interaction (K(ass)). The structural variables within the pseudopeptidic cage framework that produced a stronger and more selective recognition were thus identified. The cages showed a remarkable selectivity for N-protected dipeptides with an aromatic amino acid at the carboxylic terminus, which prompted us to propose a mode of binding based on polar and nonpolar noncovalent interactions. Accordingly, we faced the molecular recognition of a target dipeptide (Ac-EY-OH) mimicking a biologically relevant sequence by NMR and fluorescence spectroscopy in highly competitive media.

The increase of the ionic strength amplifies the species bearing acidic side chains from a bio-inspired dynamic combinatorial library of macrocyclic pseudopeptides, in close resemblance to the evolution observed for the proteins of halophilic microorganisms.

A highly efficient synthesis of small pseudopeptidic cages from simple precursors has been achieved by the triple S(N)2 reaction between tripodal tris(amido amines) and several 1,3,5-tris(bromomethyl)benzene electrophiles. The success of the macrobicyclization strongly depends on the central triamine scaffold, which dictates the correct preorganization of the intermediates. The chloride binding properties of the protonated pseudopeptidic cages have been studied in the solid state (by X-ray diffraction) as well as in solution (by NMR spectroscopy and ESI-MS) and in the gas phase (by collision-induced dissociation (CID)-MS). The crystal structure of the HCl salts of several cages show a chloride partially or completely caged within the cavity of the macrobicycle. Both the amino acid side chain and the substitution at the aromatic tripodal ring have an effect on the chloride binding ability. The cages derived from the 1,3,5-benzene moiety show low affinity, whereas the triple substitution in the ring (either with Me or Et) increases the chloride binding by one order of magnitude. Besides, the cages derived from aliphatic amino acids display a stronger interaction than those derived from phenylalanine. The basis for the different mode of binding depending on the receptor structure is proposed according to the structural data (X-ray and NMR spectroscopy). Finally, the transport of the chloride anion through lipid bilayers has been studied for selected cages. Despite the important differences in the chloride binding, the transport properties are better correlated with the lipophilicity of the molecules. Therefore, the pseudopeptidic cages sharing the same binding motif for chloride rendered very different interaction and transport properties depending on the peripheral substitution., (Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

An anionic dicarboxylate is able to template the formation of geometrically disfavoured macrocycles from a dynamic covalent mixture of open chain oligoimines.

The molecular recognition properties of pseudopeptidic macrocycles have been studied by ESI-MS and NMR spectroscopy, as highly complementary experimental techniques in solution and in the gas phase. We used ESI-MS competition experiments for the high throughput screening of the supramolecular interaction between four macrocyclic receptors and different peptide-like substrates in solution, rendering the best-fitted host-guest pairs. Further insights on the non-covalent recognition process in the gas-phase were obtained through collision induced dissociation (CID) experiments. Solution studies using NMR spectroscopy ((1)H NMR titrations, NOESY and DOSY) were carried out to prove the validity of ESI-MS as a high-throughput screening method for studying the molecular recognition of the investigated pseudopeptidic macrocycles. A clear selectivity for N-protected dipeptides over N-protected amino acids, and a slight preference for dipeptides bearing aromatic side chains were observed. On the basis of the results obtained from this approach, a mode of binding has been proposed.

The self-assembling abilities of several pseudopeptidic macrocycles have been thoroughly studied both in the solid (SEM, TEM, FTIR) and in solution (NMR, UV, CD, FTIR) states. Detailed microscopy revealed large differences in the morphology of the self-assembling micro/nanostructures depending on the macrocyclic chemical structures. Self-assembly was triggered by the presence of additional methylene groups or by changing from para to meta geometry of the aromatic phenylene backbone moiety. More interestingly, the nature of the side chain also plays a fundamental role in some of the obtained nanostructures, thus producing structures from long fibers to hollow spheres. These nanostructures were obtained in different solvents and on different surfaces, thus implying that the chemical information for the self-assembly is contained in the molecular structure. Dilution NMR studies (chemical shift and self-diffusion rates) suggest the formation of incipient aggregates in solution by a combination of hydrogen-bonding and pi-pi interactions, thus implicating amide and aryl groups, respectively. Electronic spectroscopy further supports the pi-pi interactions because the compounds that lead to fibers show large hypochromic shifts in the UV spectra. Moreover, the fiber-forming macrocycles also showed a more intense CD signature. The hydrogen-bonding interactions within the nanostructures were also characterized by attenuated total-reflectance FTIR spectroscopy, which allowed us to monitor the complete transition from the solution to the dried nanostructure. Overall, we concluded that the self-assembly of this family of pseudopeptidic macrocycles is dictated by a synergic action of hydrogen-bonding and pi-pi interactions. The feasibility and geometrical disposition of these interactions finally render a hierarchical organization, which has been rationalized with a proposal of a model. The understanding of the process at the molecular level has allowed us to prepare hybrid soft materials.

The binding between a pseudopeptidic macrocyclic naphthalenophane and different N-protected amino acid derivatives has been thoroughly studied by ESI-MS, NMR, fluorescence, and molecular modeling. Careful NMR titration experiments led to the characterization of the intermolecular noncovalent interactions, reflecting a slight side chain and l-stereoselectivity of the host-guest complexes. The data suggest the formation of an intimate ionic pair after the proton transfer from the carboxylic substrate to the amino macrocycle. Additional intermolecular interactions like H-bonding and pi-pi contacts are also important. This receptor shows a stronger interaction with substrates bearing aromatic rings, either in the side chain or in the N-protecting group. Besides, for N-Z-Phe-OH, a moderate enantioselectivity has been observed. Mass spectrometry suggests the formation of supramolecular complexes with stoichiometries higher than 1:1. The dual nature of the fluorescence emission of the macrocyclic receptor allowed determining binding constants and pertinent thermodynamic parameters. On the basis of the experimental data (NMR titrations, intermolecular ROESY, VT-NMR) and with the help of molecular modeling, a reasonable structure for the supramolecular complexes can be proposed, in which the interactions with the naphthyl ring of the receptor play a fundamental role in the strength and selectivity of the molecular recognition event.

The anion-templated synthesis of different pseudopeptidic macrocycles has been studied in detail by using a multidisciplinary approach. The reaction between an open-chain pseudopeptidic diamine and the appropriate dialdehyde is highly affected by the presence of the best fitting anionic template. The formation of the corresponding macrocyclic tetraimino-template supramolecular complex is demonstrated by NMR (ROESY and PGSE) and mass spectrometry (ESI-TOF). These supramolecular complexes can be easily reduced to the corresponding more stable tetraamine macrocycles. Accordingly, we have used this reaction to efficiently synthesize a family of new pseudopeptidic macrocycles in a one-pot two-steps anion-templated reductive amination reaction, which comprises a multicomponent macrocyclization through the selective formation of four chemical bonds to yield a unique macrocyclic structure. Different variables like the aliphatic spacer between amino acidic moieties, geometry of the dialdehyde, and structure of the amino acid side chains were thoroughly studied, and their effect in the formation and stability of the supramolecular complexes discussed. The conformational preorganization induced by the template has been monitored by circular dichroism, reflecting the differences observed in the isolated yields, as well as by NMR spectroscopy. This effect has been also supported by molecular modeling. All the experimental and theoretical techniques were strongly consistent and reflected the same trends by comparing the different structural variables introduced in the system.

The interaction of a synthetic enantiopure azamacrocyclic receptor (L) with biologically important chiral dicarboxylates (A, 1-7) has been studied by means of potentiometric titrations in 0.15 M NaCl aqueous solution in a wide pH range. This macrocycle forms strong complexes of the type [HnLA](n-2) (with n = 0-5). As a general trend, the binding is much tighter at basic or neutral pH than in acidic medium. Interestingly, nonprotected excitatory amino acids (Asp and Glu) are strongly bound even at acidic pH. Regarding selectivity, the receptor showed stereoselective binding toward those substrates bearing an H-bonding donor at Calpha, being S-selective in most of the cases, except for glutamic acid. Thus, L displayed an excellent enantioselectivity for (S)-malate dianion (KS/KR = 11.50 at pH 10.0 and KS/KR = 6.86 at pH 7.0) and exhibited moderate enantiopreference for (S,S)-tartrate (KSS/KRR = 3.01 at pH 10 and KSS/KRR = 1.70 at pH 7.0). For this last anion, a very good diastereopreference was also observed (KSS/KRS = 8.46 at pH 10 and KSS/KRS = 4.99 at pH 7.0). On the contrary, L is smoothly R-selective toward (R)-Glu (KR/KS = 3.22 at pH 10 and KR/KS = 2.05 at pH 7.0) due to its longer and more flexible molecular structure. The stereoselectivity of the corresponding complexes decreased when decreasing pH values. For the hydroxy derivatives, mass spectrometry also reflected the trends observed by potentiometry and confirmed the receptor:dicarboxylate 1:1 stoichiometry of the supramolecular complexes. Additional experimental techniques were used to study the most stereoselective example. Solution studies by NMR suggested a good geometrical complementarity between the malate dianion and the receptor, which showed a predominant helical conformation in solution. Besides, self-diffusion rates (PGSE) of the diastereomeric complexes with malate also agree with binding data. Circular dichroism was also used in this case at different pH values, showing a very good correlation between the helical content of the receptor and the stereoselectivity of the molecular recognition process.

Cadmium promoted diastereoselective amplification of a single member from a dynamic combinatorial library of stereoisomeric oligoimines of different sizes allows the efficient preparation of a new macrocyclic polyamine.

The conformational behavior of 16- to 18-membered ring peptidomimetic p-cyclophanes 1a,b-3a,b has been studied by NMR. The cycles bearing 16 and 17 atoms showed a dynamic process within the NMR time scale, produced by the rotation of the aromatic p-diphenylene moiety with respect to the macrocyclic main plane. The temperature dependence of 1H NMR spectra has been studied in order to get activation parameters of the energetic barrier for the process (VT-NMR and line shape analysis). The rate of the movement clearly depends on the macrocyclic ring size and the nature of the peptidomimetic side chain. Entropic and enthalpic contributions to the free energy of activation are discussed. The rotation of the aromatic ring is closely related to the intramolecular hydrogen bonding pattern, as suggested by temperature factors of NH chemical shifts (DeltadeltaNH/DeltaT) and molecular modeling. The interconnected roles of the solvation and the intramolecular H-bonds have been established by measurements (VT-NMR and DeltadeltaNH/DeltaT) in environments of different polarities and H-bonding abilities. We concluded that the conformational stability of the systems directly depends on the stability of the intramolecular H-bonding pattern. We finally showed how one of these peptidomimetics behaves as a methanol-dependent artificial molecular rotor. In this simple molecular device, the well-defined molecular rotation is tuned by the competition between intramolecular hydrogen bonds and interactions with the solvent.