Water-soluble polycarbodiimides and their cytotoxic and antifungal properties.

We have successfully synthesized water-soluble neutral and polyelectrolyte helical polycarbodiimides and studied their biological properties. These polymers were prepared by decorating carbodiimide backbones with nonionic, hydrophilic functional groups such as dimethylamine, piperazine, and morpholine. Additionally, the 3° amines present in these functional groups were quaternized using methyl iodide as the alkylating agent to produce their ionic analogs. Polycarbodiimides were chosen as the base polymer used because of their facile chemical modification, pH tolerance in terms of both their helical conformations and degradation behaviors, and tunable helical inversion barriers. Hydrophilic side groups, such as morpholine, dimethylamine, and piperazine, can be used to balance the amphiphilic architecture of the polycarbodiimides along with lipophilic groups, such as alkyl side chains. A chiral R or S BINOL Ti(IV) isopropoxide catalyst was used to control the handedness of the polycarbodiimide helices in these studies. These ionic and neutral polycarbodiimides were subsequently studied for potential antimicrobial and cytotoxic properties. Poly[N-methyl-N'-2-morpholinoethylcarbodiimide], as an example, exhibited significant antifungal properties against Candida albicans. Also, Poly[N-methyl-N'-2-morpholinoethylcarbodiimide] showed significant inhibition of biofilm formation. This suggests that the polymer is a promising candidate for antifungal biomedical applications. Measuring cytotoxicity against urinary bladder cancer cells, poly[N-[3-(dimethylamino)propyl)]-N'-[3-(morpholino)propyl]carbodiimide] (S-cat) and poly[N-[3-(dimethylamino)propyl)]-N'-[3-(morpholino)propyl]carbodiimide]-MeI (S-cat) showed significantly low IC50 values. The IC50 values of poly[N-[3-(dimethylamino)propyl)]-N'-[3-(morpholino)propyl]carbodiimide] (S-cat) and Poly[N-[3-(dimethylamino)propyl)]-N'-[3-(morpholino)propyl]carbodiimide]-MeI (S-cat) are 3.50 µM and 1.27 µM, respectively. The significantly low cancer cell growth inhibition concentration implies the highest cytotoxicity of the polymers, suggesting potential applications as cancer therapeutics. These results also showed that the functionalization and chirality of polycarbodiimides modulate their anticancer and antifungal activity.

Self-assembling Morphologies Obtained from Helical Polycarbodiimide Copolymers and Their Triazole Derivatives.

A facile method for the preparation of polycarbodiimide-based secondary structures (e.g., nano-rings, "craters," fibers, looped fibers, fibrous networks, ribbons, worm-like aggregates, toroidal structures, and spherical particles) is described. These aggregates are morphologically influenced by extensive hydrophobic side chain-side chain interactions of the singular polycarbodiimide strands, as inferred by atomic force microscopy (AFM) and scanning electron microscopy (SEM) techniques. Polycarbodiimide-g-polystyrene copolymers (PS-PCDs) were prepared by a combination of synthetic methods, including coordination-insertion polymerization, copper(I)-catalyzed azide alkyne cycloaddition (CuAAC) "click" chemistry, and atom transfer radical polymerization (ATRP). PS-PCDs were found to form specific toroidal architectures at low concentrations in CHCl3. To determine the influence of a more polar solvent medium (i.e., THF and THF/EtOH) on polymer aggregation behavior, a number of representative PS-PCD composites have been tested to show discrete concentration-dependent spherical particles. These fundamental studies are of practical interest to the development of experimental procedures for desirable architectures by directed self-assembly in thin film. These architectures may be exploited as drug carriers, whereas other morphological findings represent certain interest in the area of novel functional materials.

Utilization of a Meldrum's acid towards functionalized fluoropolymers possessing dual reactivity for thermal crosslinking and post-polymerization modification.

New thermally cross-linkable and/or post-functionalizable perfluorocyclobutyl (PFCB) polymers containing Meldrum's acid moieties have been successfully prepared via the thermal cyclopolymerization of a new Meldrum's acid functionalized aromatic trifluorovinyl ether (TFVE) monomer.

Identification of the specific, shutter-like conformational reorientation in a chiroptical switching polycarbodiimide by VCD spectroscopy.

The specific conformational states responsible for the unique, reversible temperature- and solvent-driven chiroptical switching process experienced by poly(N-1-naphthyl-N'-octadecyl-carbodiimide) (PNOC) have been identified using VCD spectroscopy and DFT calculations. The distinct VCD spectra of PNOC corresponding to the two specific conformations were obtained for the polymer dissolved in DCM-d2 (state A) and CDCl3 (predominantly state B). To specifically assign the structures of both conformations, two simplified 7mer models were constructed and optimized using DFT calculations. The theoretical spectra associated with these model conformations show a high level of agreement when compared to the experimental VCD spectra. The two states consist of the naphthyl pendant groups aligned directionally opposing the helical rotation (model A) and aligned with the helicity of the backbone (model B). This pendant reorientation causes very large OR and ECD Cotton effect inversions upon modification to the temperature or solvent composition of dilute (+)-PNOC solutions in specific solvents. In addition, the pendant group equilibrium from state A to B causes a contraction of the helical pitch from the more expanded 5/1 pitch to the more contracted 7/2 pitch resulting in increased disorder of the solvation sphere surrounding the polymer chain.

Evidence of entropy-driven bistability through (15)N NMR analysis of a temperature- and solvent-induced, chiroptical switching polycarbodiimide.

The thermo- and solvo-driven chiroptical switching process observed in specific polycarbodiimides occurs in a concerted fashion with large deviations in specific optical rotation (OR) and CD Cotton effect as a consequence of varying populations of two distinct polymer conformations. These two conformations are clearly visible in the (15)N NMR and IR spectra of the (15)N-labeled poly((15)N-(1-naphthyl)-N'-octadecylcarbodiimide) (Poly-3) and poly((15)N-(1-naphthyl)-(15)N'-octadecylcarbodiimide) (Poly-5). Using van't Hoff analysis, the enthalpies and entropies of switching (ΔHswitching; ΔSswitching) were calculated for both Poly-3 and Poly-5 using the relative integrations of both peaks in the (15)N NMR spectra at different temperatures to measure the populations of each state. The chiroptical switching (i.e., transitioning from state A to state B) was found to be an endothermic process (positive ΔHswitching) for both Poly-3 and Poly-5 in all solvents studied, meaning the conformation correlating with the downfield chemical shift (ca. 148 ppm, state B) is the higher enthalpy state. The compensating factor behind this phenomenon has been determined to be the large increase in entropy in CHCl3 as a result of the switching. Herein, we propose that the increased entropy in the system is a direct consequence of increased disorder in the solvent as the switching occurs. Specifically, the chloroform solvent molecules are very ordered around the polymer chains due to favorable solvent-polymer interactions, but as the switching occurs, these interactions become less favorable and disorder results. The same level of solvent disorder is not achieved in toluene, causing the chiroptical switching process to occur at higher temperatures.

Synthesis of Guanidinium Functionalized Polycarbodiimides and Their Antibacterial Activities.

A family of guanidinium-side-chain functionalized polycarbodiimides has been synthesized by allowing an azido guanidinium salt to react with alkyne polycarbodiimides via the copper catalyzed [3 + 2] cycloaddition (Click) reaction. Poly-2(a-d) are cationic/amphiphilic polymers in which the global hydrophilic/hydrophobic balance has been tailored by local alteration of the length of alkyl side chain in the repeat unit of polymers prior to polymerization. The shorter alkyl chains yield water-soluble polymers, Poly-2c, -2d, and -2e. Antibacterial activities of these cationic polycarbodiimides have been investigated for Gram-positive and Gram-negative bacteria that include Staphylococcus aureus, methicillin-resistant Staphylococcus aureus (MRSA), Escherichia coli, and Acinetobacter baumannii. It was observed that the influence of hydrophobic-hydrophilic balance per repeat unit of these polymers have profound effects for both antimicrobial and hemolytic activities. In addition, these polycarbodiimide-guanidinium-triazole conjugates offered moderate to significant antibacterial activity and rapid interaction with red blood cells causing blood precipitation without significant hemolysis in case of Poly-2(b-e). This latter property has the potential to be exploited in the polymer coatings or wound protection.

Resolving the Regioregularity of Poly(N-n-hexyl-N'-phenylcarbodiimide) via Nitrogen-15 Labeling.

Nitrogen-15 nuclear magnetic resonance (NMR) spectroscopy and infrared spectroscopy (IR) were performed on an isotope-enriched poly(N-n-hexyl-N'-phenylcarbodiimide) to determine directly the connectivity and regioregularity of a polymer. Up to this point, the imine, C═N, IR stretch at 1660-1620 cm-1 was thought to be a sufficient handle to elucidate the presence or absence of a regioregular microstructure; however, recent findings cast some uncertainties when expanded to all polycarbodiimides. Therefore, an enriched 15N NMR study was undertaken to unambiguously resolve that a N-n-hexyl-N'-phenylcarbodiimide, when polymerized with a 2,2,2-trifluoroethoxide trichlorotitanium(IV) catalyst in chloroform, will produce a completely regioregular polymer. The only regioisomer present is the one in which the phenyl pendant group is positioned on the imino-nitrogen. The study was expanded to a chiral, (R)-BINOL-Ti(IV)-diisopropoxide catalyst which revealed no change in the regioisomer or the degree of regioregularity. In addition to 15N NMR spectra, the IR imine stretch exhibited isotope shifts for poly(N-n-hexyl-N'-phenylcarbodiimide) when labeled on both imino- and amino-nitrogen or solely the imino-nitrogen of ∼11 cm-1; however, no shift was manifested when labeling was restricted to the amino-nitrogen.

Stable-isotope labeled hydrophobic hydrazide reagents for the relative quantification of N-linked glycans by electrospray ionization mass spectrometry.

This study presents the development of stable-isotope labeled hydrophobic, hydrazide reagents for the relative quantification of N-linked glycans. The P2GPN "light" ((12)C) and "heavy" ((13)C(6)) pair are used to differentially label two N-linked glycan samples. The samples are combined 1:1, separated using HILIC, and then mass differentiated and quantified using mass spectrometry. These reagents have several benefits: (1) impart hydrophobic character to the glycans affording an increase in electrospray ionization efficiency and MS detection; (2) indistinguishable chromatographic, MS, and MS/MS performance of the "light" and "heavy" reagents affording relative quantification; and (3) analytical variability is significantly reduced due to the two samples being mixed together after sample preparation. Obtaining these analytical benefits only requires ~4 h of sample preparation time. It is shown that these reagents are capable of quantifying changes in glycosylation in simple mixtures, and the analytical variability of the reagents in pooled plasma samples is shown to be less than ±30%. Additionally, the incorporation of an internal standard allows one to account for the difference in systematic error between the two samples due to the samples being processed in parallel and not mixed until after derivatization.

Inhibition of Acinetobacter baumannii biofilm formation on a methacrylate polymer containing a 2-aminoimidazole subunit.

A polymeric composite containing a 2-aminoimidazole derivative was synthesized. It was found that this polymer was resistant to biofilm colonization by Acinetobacter baumannii, no leaching of the 2-aminoimidazole derivative was observed after 2 weeks of treatment with deionized water, and the resulting polymer was not hemolytic.

Synthesis and characterization of random and block copolypeptides derived from gamma-methylglutamate and leucine N-carboxyanhydrides.

The synthesis of random and block copolypolyeptides derived from gamma-methylglutamate and leucine N-carboxyanhydrides using Al-Schiff's base complexes and allylamine as initiators is here reported. The copolymer structures were confirmed by (1)H and (13)C NMR. The calculation of the statistical average block lengths reveals the presence of longer methylglutamate units in the copolymer. The determination of the reactivity ratios indicated a slightly higher reactivity of gamma-methylglutamateNCA as compared to leucineNCA. Block copolypeptides containing glutamate and leucine units were obtained by sequential polymerization of the two NCAs using Al-Schiff's base complexes or allylamine in dioxane as solvent. Based on (13)C NMR spectra of copolymers exhibiting two signals corresponding to peptide linkages, we confirmed the block structure and concluded that the copolymerization proceeds by attack of an amino group present on a glutamate chain end onto a LeuNCA. The copolymerization with allylamine was also shown, from calculation of the average block lengths of sequences, to exhibit living behavior. Viscometry analysis further showed that molar masses of the copolypeptides obtained with Al-Schiff's base were quite close to those derived from allylamine, supporting the proposed mechanism of copolymerization.

Chiroptical switching polyguanidine synthesized by helix-sense-selective polymerization using [(R)-3,3'-dibromo-2,2'-binaphthoxy](di-tert-butoxy)titanium(IV) catalyst.

A series of chiral binaphthyl titanium alkoxide complexes were synthesized. Among them, chiral titanium complex [(R)-3,3'-dibromo-2,2'-binaphthoxy](di-tert-butoxy)titanium(IV) (R-3) exists as a crystallographic C2 dimer in the solid state but a monomer in solution at room temperature. Application of R-3 in the helix-sense-selective polymerization of achiral carbodiimide, N-(1-anthryl)-N'-octadecylcarbodiimide (1), yielded a well-defined regioregular, stereoregular poly[N-(1-anthryl)-N'-octadecylguanidine] (poly-1b) with a relatively narrow polymer dispersity index of 2.7. Full racemization of poly-1b at +80 degrees C in toluene requires more than 100 h. Interestingly, poly-1b was found to undergo fast reversible chiroptical switching at +38.5 degrees C in toluene. Furthermore, at room temperature, poly-1b shows a positively signed Cotton effect in toluene, but negative ones in THF and chloroform, respectively. The chiroptical switching takes place around the toluene content of 90% (vol) in the mixed toluene/THF solvents. This is the first example of chiroptical switching phenomenon occurring in a helical polymer possessing no chiral moieties in the polymer chains. We believe this reversible chiroptical switching phenomenon occurs by reorientation of anthracene rings relative to the chain director.

Helix-sense selective polymerization of carbodiimides: building permanently optically active polymers from achiral monomers.

The helix-sense selective polymerization of achiral monomers by homochiral catalysts was investigated. Polymerization of chiral carbodiimides (N-(R)-2,6-(dimethylheptyl)-N'-phenylcarbodiimide) by achiral catalysts yields polymers that undergo mutorotation at elevated temperatures, thus illustrating that these chains are formed under kinetic rather than thermodynamic control. Building on this observation, the polymerization of achiral carbodiimides by (S-BINOL)Ti(OiPr)2, I, was studied. Monomers (N-hexyl-N'-(X)carbodiimide, where X = isopropyl (3), hexyl (4) or phenyl (5)), N-methyl-N'-(2-methyl-6-isopropylphenyl)carbodiimide, 6, and N-dodecyl-N'-(1-naphthyl)carbodiimide, 7, were all polymerized with I in good yields (86-95%), and all showed varying degrees of asymmetric induction. Poly-3, -4, and -5 racemized upon heating at elevated temperatures, but poly-6 and poly-7, bearing nonsymmetric phenyl groups, yielded optically active polymers that could not be racemized even at elevated temperatures. Thin films of poly-7 were found to be highly opalescent.

Stable helical polyguanidines: poly[N-(1-anthryl)-N'-[(R)- and/or (S)-3,7-dimethyloctyl]guanidines].

Using chiral catalysts of (R)- and/or (S)-BINOL-Ti, the asymmetrical polymerization of achiral monomer, N-(1-anthryl)-N'-n-octadecylcarbodiimide, yielded soluble nonregioregular polyguanidines of Poly-R1 and Poly-S1. A racemization process occurred when the toluene solution of Poly-R1 was annealed at elevated temperatures (70-80 degrees C). Kinetic studies reveal this to be a slow process with an activation energy of ca. 36 kcal/mol. On the other hand, using titanium(IV) trifluoroethoxide catalyst, the polymerization of N-(1-anthryl)-N'-[(R)- and/or (S)-3,7-dimethyloctyl]carbodiimides afforded highly regioregular polyguanidines of Poly-R2 and Poly-S2. These polymers adopt stable helices in various solvents and elevated temperatures, whose kinetically controlled conformations and thermodynamically controlled conformations are essentially the same.