Intracellular organelle targeting, especially of mitochondria, is of significant importance for drug delivery to improve therapeutic efficiency and overcome the endosomal trap. While mitochondria targeting has been achieved by mostly cationic systems, this communication reports a rare phenomenon of mitochondria targeting by water-soluble non-ionic polymers (representative substrate of interest), featuring an oligo-oxyethylene appended polymethyl-methacrylate backbone, and terminated with an amine-substituted single naphthalene-monoamide (NMI) derivative (a push-pull type chromophore). The push-pull type charge-neutral NMI chromophore facilitates both cellular uptake and crossing the mitochondrial membrane barrier, while its green emission allows for simultaneous fluorescence imaging. Considering the outstanding mitochondria targeting, cell compatibility, simultaneous imaging and wide-ranging covalent-conjugation possibilities, this simple yet versatile chromophore brings new opportunities in intracellular mitochondria targeted delivery.

This manuscript describes chain-folding-regulated hierarchical self-assembly of an amphiphilic polyurethane (P1), exhibiting superior surface functional group display and biological activity compared to an analogous amphiphilic block copolymer (P3) or a rigid polyurethane (P2) that is deprived of chain folding. In P1, intrachain hydrogen bonding directs the formation of a pleated structure that hierarchically assembles into hollow capsules. A glucose oxidase–peroxidase assay confirmed that this unique assembly of P1 allows displaying ∼50% glucose moieties on the outer surface, which significantly outperforms amphiphilic block copolymers. This renders outstanding multivalent binding with the lectin concanavalin A, as is evident by a very high association constant (∼105 M–1) and spontaneous glycocluster effect. In contrast, P2 or P3 fails to exhibit any notable glycocluster effect under identical conditions. Furthermore, P1, unlike P2 or P3, induced clustering of Staphylococcus aureus by targeting bacterial carbohydrate transporters, deactivated the bacteria, and prevented internalization into mammalian cells.

pH-responsive aqueous nanoassembly from a fluorescent neutral polyester selectively targets mitochondria in cancer cells over non-cancerous cells due to positive charge gain, while similar assemblies from cationic polyesters show no such selectivity.

The growing demand for biodegradable polymers capable of stimuli-responsive drug release is challenged by limitations in facile synthetic methods. In this study, two biotin-functionalized amphiphilic polyesters (P1 and P2) were synthesized through step-growth polymerization, aiming to achieve biotin receptor-mediated cancer cell selective uptake. In addition to polar biotin, P2 incorporates a hydrophobic fluorescent dye, which enabled intracellular fluorescence tracking. P2 self-assembled into highly biocompatible spherical nanoaggregates (∼120 nm) in water, which showed effective encapsulation of the hydrophobic anticancer drug doxorubicin (DOX). It displayed ∼85-90% internalization in biotin-overexpressed cancer cells (HeLa and MCF7) contrary to only ∼5-10% uptake in noncancerous cells (NIH 3T3), as determined by flow cytometry and fluorescence microscopy. Cell-selective DOX release was likely induced by the polyester degradation in the acidic cancer microenvironment and via endogenous esterases, evident from size exclusion chromatography (SEC) and dynamic light scattering (DLS) experiments. These findings highlight the potential of stimuli-responsive degradable polyester nanocarriers for targeted cancer treatment.

In biological systems, programmable supramolecular frameworks characterized by coordinated directional non-covalent interactions are widespread. However, only a small number of reports involve pure water-based dynamic supramolecular assembly of artificial π-amphiphiles, primarily due to the formidable challenge of counteracting the strong hydrophobic dominance of the π-surface in water, leading to undesired kinetic traps. This study reveals the pathway complexity in hydrogen-bonding-mediated supramolecular polymerization of an amide-functionalized naphthalene monoimide (NMI) building block with a hydrophilic oligo-oxyethylene (OE) wedge. O-NMI-2 initially produced entropically driven, collapsed spherical particles in water (Agg-1); however, over a span of 72 h, these metastable Agg-1 gradually transformed into two-dimensional (2D) nanosheets (Agg-2), favoured by both entropy and enthalpy contributions. The intricate self-assembly pathways in O-NMI-2 enable us to explore seed-induced living supramolecular polymerization (LSP) in water for controlled synthesis of monolayered 2D assemblies. Furthermore, we demonstrated the nonspecific surface adsorption of a model enzyme, serine protease α-Chymotrypsin (α-ChT), and consequently the enzyme activity, which could be regulated by controlling the morphological transformation of O-NMI-2 from Agg-1 to Agg-2. We delve into the thermodynamic aspects of such shape-dependent protein-surface interactions and unravel the impact of seed-induced LSP on temporally controlling the catalytic activity of α-ChT.

Aggregation of amphiphilic polymers in block-selective solvents produces different nanostructures, which have been studied extensively for wide-ranging applications. Nevertheless, such immiscibility-driven aggregation does not endow them with the desired structural precision, predictability or surface functional group exposure, which significantly impact their functional applications. More recently, biomimetic folded structures of synthetic macromolecules (mostly oligomers) have come to the fore, but such studies have been limited to probe the secondary structures. In this article, we have collated hierarchical structures of foldamers, especially highlighting our recent contribution to the field of chain-folding regulated assembly of segmented polyurethanes (PUs) and their functional applications. A series of such PUs have been discussed, which contain a segmented hydrocarbon backbone and alternately placed pendant solvophilic groups. In either water or highly non-polar solvents (TCE, MCH), depending on the nature of the pendant group, they exhibit folded structures stabilized by intra-chain H-bonding. Hierarchical assembly of such folded chains by inter-chain H-bonding and/or π-stacking leads to the formation of well-defined nanostructures with functional applications ranging from organic optoelectronics to biomaterials. For example, a segmented PU with appended naphthalene-diimide (NDI) chromophores showed a pleated structure in MCH, which helped in organization of the NDI chromophores within π-stacking distance. Such folded polymer chains eventually produced nanotubular structures with excellent electron mobility. They also showed efficient intercalation of the pyrene (Py) donor by NDI-Py charge-transfer interaction and in this case the mixed nanotubular structure exhibited prominent room-temperature ferroelectricity. On the other hand, having cationic functionalities as the pendant groups such chain-folding regulated assembly produced unilamellar polymersomes with excellent antibacterial activity with very low minimum inhibitory concentrations (<10 μg mL-1). Replacing the pendant amine functionality with sulphate groups made these polyurethanes highly potent antiviral materials. In the absence of the alternating connectivity of the solvophobic and solvophilic segments or rigid hydrocarbon backbone, such folding propensity is destroyed, leading to structural collapse. While significant efforts have been made in correlating primary structures of wide-ranging polymers with their functional applications, this article demonstrates the direct correlation between the secondary structures of polymers and their functional properties.

Thermoresponsive polymers, exhibiting a lower critical solution temperature (LCST), are of diverse interest in designing stimuli-responsive materials. Most such systems are limited to poly(N-isopropylacrylamide) (PNIPAM)-derived macromolecules or others having a C-C backbone. This article reveals the synthesis of thermoresponsive alternating copolymers based on a segmented polyurethane (PU) scaffold. These polymers contain a hydrocarbon backbone, periodically grafted with a hydrophilic oligo-oxyethylene (OE) wedge. In water, they adopt a pleated conformation, driven by intrachain H-bonding (among the urethane groups), which by hierarchical assembly produces hollow capsules capable of sequestering hydrophilic guests. They exhibit a LCST with tunable cloud points in the range of similar to 19 to 52 degrees C, depending on the degree of polymerization (DP) or hydrophobic/hydrophilic balance. With the same hydrophobic/hydrophilic content, the cloud point decreases by similar to 10 degrees C with the increase in the DP from 7 to 16, which can be attributed to the difference in the free volumes caused by the polymer chains and solvent molecules. On the other hand, by decreasing the hydrophobic content (two -CH2 units in a repeating unit), the cloud point increases by similar to 30 degrees C. For a given candidate, the thermodynamics of nonspecific protein (BSA) adsorption on the surface of the capsule was correlated with the LCST by isothermal titration calorimetry studies. At T < LCST, protein adsorption was largely governed by favorable entropy contribution due to the freeing of the surface-bound water molecules. In contrast, the enthalpy contribution became more prominent above the LCST, suggesting a dominant role of the hydrophobic interaction with the already dehydrated OE chains. Considering excellent biocompatibility (> 85% cell viability with up to 500 mu g/mL polymer after 72 h incubation with HeLa cells), degradable backbone, container property, tunable LCST, and temperature-dependent stealth effect, this polymeric system appears promising as a delivery vehicle for biomedical applications.

This manuscript reports the effect of hydrogen-bonding functionality on the supramolecular assembly of naphthalene-diimide (NDI)-derived amphiphilic building blocks in water. All the molecules contain a central NDI chromophore, functionalized with a hydrophilic oligo-oxyethylene (OE) wedge in one arm and a phenyl group on the opposite arm. They differ by a single H-bonding functionality, which links the NDI chromophore and the phenyl moiety. The H-bonding functionalities are amide, thioamide, urea, and urethane in NDI-A, NDI-TA, NDI-U, and NDI-UT, respectively. All of these molecules exhibit π-stacking in water, as evident from their distinct UV/vis absorption spectra when compared to that of the monomeric dye in THF. However, among these four, only NDI-A and NDI-TA show hydrogelation, while the other two precipitate out of the medium. The NDI-A hydrogel also exhibits transient stability and leads to a crystalline precipitate within ∼5 h. Only NDI-TA produces stable transparent hydrogel with the entangled fibrillar morphology that is typical for gelators. Both NDI-A and NDI-TA showed a thermoresponsive property with a lower critical solution temperature of about 41-42 °C. Powder XRD studies show a parallel orientation for NDI-A and an antiparallel orientation for NDI-TA. Computational studies support this experimental observation and indicate that the NDI-A assembly is highly stabilized by strong H-bonding among the amide groups and π-stacking interaction in the parallel orientation. On the other hand, due to weak H-bonding among the thioamide groups, the binding energy of the parallelly oriented NDI-TA was significantly lower and the optimized structure was disordered. Instead, its antiparallel orientation was more stable, with criss-cross aligned H-bonding interactions and π-π interactions between adjacent aromatic rings. The NDI-TA hydrogel with less ordered OE chains on the surface showed prominent adsorption of serum protein BSA. In sharp contrast, NDI-A did not exhibit any notable interaction with BSA, as evident from the ITC studies.

CONSPECTUS: Bioinspired self-assembly has been explored with diverse synthetic scaffolds, among which amphiphiles are perhaps the most extensively studied systems. Classical surfactants or amphiphilic block copolymers, depending on the hydrophobic-hydrophilic balance, produce distinct nanostructures, which hold promise for applications ranging from biology to materials sciences. Nevertheless, their immiscibility-driven aggregation does not provide the opportunity to precisely regulate the internal order, morphology, or functional group display, which is highly desirable, especially in the context of biological applications. A new class of amphiphiles have emerged in the recent past in which the hydrophilic segment(s) is appended with a hydrophobic supramolecular-structure-directing-unit (SSDU), consisting of a pi-conjugated chromophore and a H-bonding group. Self-recognition of the SSDU by attractive directional interactions governs the supramolecular assembly, which is fundamentally different than the repulsive solvent-immiscibility driven aggregation of traditional amphiphiles. Such SSDU-appended hydrophilic polymers exhibit entropy-driven highly stable self-assembly producing distinct nanostructures depending on the H-bonding functional group. For example, polymers with the hydrazide-functionalized SSDU attached form a polymersome, while in a sharp contrast, the same polymers when connected to an amide containing SSDU produce a cylindrical micelle via a spherical-micelle intermediate. This relationship holds true for a series of SSDU-attached hydrophilic polymers irrespective of the hydrophobic/hydrophilic balance or chemical structure, indicating that the supramolecular-assembly is primarily controlled by the specific molecular-recognition motif of the SSDU, instead of the packing parameter-based norms. Beyond synthetic polymers, SSDU-attached proteins also exhibit similar molecular-recognition driven self-assembly as well as coassembly with SSDU-attached polymers or hydrophilic wedges, producing multi-stimuli-responsive nanostructures in which the protein gains remarkable protection from thermal denaturation or enzymatic hydrolysis and exhibits redox-responsive enzymatic activity. Furthermore, SSDU-derived bola-shape pi-amphiphiles have been recognized as a useful scaffold for the synthesis of unsymmetric polymersomes, rarely reported in the literature. The building block consists of a hydrophobic naphthalene-diimide (NDI) pi-system attached to a hydrophilic functional group (ionic or nonionic) and a nonionic wedge on its two opposite arms. Extended H-bonding among the hydrazide groups, placed only on one side of the central chromophore by design, ensures stacking of the NDIs with parallel orientation and induces a preferred direction of curvature so that the H-bonded chain and consequently the functional groups attached to the same side remain at the inner-wall of the supramolecular polymersome. Automatically, the functional groups, located on the other side, are displayed at the outer surface. This design works for different amphiphiles, which by virtue of efficient and predictable functional group display, strongly influences the multivalent binding with different biological targets resulting in efficient enzyme inhibition, glycocluster effect, or antibacterial activity, depending on the nature of the functional group. By taking advantage of the electron accepting nature of the NDI, electron rich pyrene-containing amphiphiles can be costacked in alternating sequence, producing temperature and redox-responsive supramolecular polymers with NDI/pyrene stoichiometry-dependent morphology, lower critical solution temperature (LCST), functional group display, and antibacterial activity.

Nonspecific adsorption of proteins on the surface of nanocarriers plays a critical role in their cellular uptake and other biological functions. This article reports vesicular assemblies of two π-amphiphiles (NDI-1 and NDI-2) and thermodynamic aspects of their interaction with bovine serum albumin (BSA). Both contain a hydrophobic naphthalene-diimide (NDI) core and two oligo-oxyethylene (OE) wedges but differ by the presence of the hydrazide group in NDI-1. NDI-2 exhibits a constricted π-stacking and enthalpy-driven adsorption of BSA. In contrast, NDI-1 exhibits a stronger interaction due to enhanced entropy contribution. It is postulated that a tight packing of NDI chromophores in NDI-2 results in an inadequate space in the corona, leading to the dehydration of OE chains, which contributes to the observed enthalpy-driven binding. On the other hand, due to H-bonding along the direction of π-stacking in NDI-1, an enhanced interchromophoric distance provides more space in the shell, resulting in less dehydration of the OE chains, which results in an entropy gain from the BSA binding-induced release of water from the OE chains. Intercalation of an electron-rich pyrene in the electron-deficient NDI-1 stack further reduces the grafting density of the OE chains, resulting in negligible BSA adsorption, similar to a stealth polymer. A correlation can be seen between the thermodynamic landscape of the protein adsorption and the trend of their lower critical solution temperature (LCST), which follows the order NDI-1 + Py < NDI-1 < NDI-2.

Cellular uptake is an important event in drug delivery and other biomedical applications. Amphiphilic polymers produce aggregates of different size and shape depending on the intrinsic structural differences and the packing parameter. Although they have been explored for various biomedical applications with immense interest, the relationship between the shape of the aggregate and cellular uptake has been studied only in limited examples. This work reports two polymers (P1 and P2), both of which contain a hydrophobic supramolecular structure-directing unit (SSDU) at the chain-end of a fluorescence dye-labeled hydrophilic polymer. Depending on the difference in the structure of the single H-bonding functional group (hydrazide or amide) of the SSDU, P1 and P2 produce polymersomes (NS1) and spherical micelles (NS2), respectively. An aged solution of P2 produces cylindrical micelles (NS3). Confocal microscopy studies reveal that the uptake of these nanostructures in HeLa cells greatly depends on the shape of the aggregate. Spherical NS1 and NS2 show appreciable uptake at 1 or 4 h of incubation, whereas NS3 shows negligible uptake. Temperature-dependent cellular uptake studies reveal an energy-dependent endocytosis pathway. Kinetic studies show gradual increase in the cellular uptake with time, and at 24 h the relative uptake ratio (NS1:NS2:NS3) is 1.0:0.2:<0.1, implying the polymersome morphology (NS1) is most efficient for cellular uptake compared to the spherical or cylindrical micelles. The same trend was also noticed for MDA-MB 231 cells. Confocal microscopy studies further reveal cellular internalization and intracellular location of NS1, which showed maximum cellular uptake. As the intrinsic difference in the chemical structure of the two polymers is negligible, the observed difference can be explicitly assigned to their difference in shape.

Synthesis, aqueous aggregation, hydrophobic guest encapsulation, non-covalent encapsulation stability and glutathione responsive degradation of amphiphilic hyperbranched polydisulfides have been reported.

This article reports the facile synthesis of a cylindrical micelle from mostly hydrophilic polymers by a specific molecular interaction among a hydrophobic single supramolecular structure-directing unit (SSDU), appended at the chain terminal. The SSDU contains a naphthalene-diimide (NDI) chromophore, an amide group, and a hydrophobic wedge. H bonding, pi stacking, and hydrophobic interaction among the SSDU lead to the formation of entropy-driven aggregates for appended hydrophilic polymers including one dendronized polymer. In water, the freshly prepared solution exhibits a spherical morphology, which gradually transforms into the cylindrical micelle for only one tested polymer having oligo-oxyethylene pendant chains. But no such transformation was noticed for the other two polymers having pendant hydroxyl groups. In the presence of 10% good solvent such as tetrahydrofuran (THF), the rate of such morphology transformation could be significantly enhanced for all of the tested polymers producing long (>5 mu m) cylindrical micelles. The rate was inversely proportional to the sample concentration, indicating that the initially formed spherical species was not an intermediate but an off-pathway aggregate. In the presence of a good solvent, the mole fraction of the off-pathway aggregate reduced marginally and increased unimer population, which facilitated the nucleation for the thermodynamic product (cylindrical micelle). Isothermal titration calorimetry studies revealed the disassembly of the initially formed micelle upon dilution, but no such effect was noticed for the cylindrical micelle in the tested concentration window, suggesting enhanced stability. Noncovalent encapsulation stability, probed by fluorescence resonance energy transfer (FRET) studies, revealed fast chain exchange by the expulsion/insertion mechanism for the off-pathway spherical aggregate. In contrast, the average lifetime increased significantly for the cylindrical micelle with predominant splitting/merging of the micelle mechanism for chain exchange, similar to amphiphilic block copolymers, although in the present system, the hydrophobic SSDU was merely 5-8 wt %.

Ionic liquids (ILs) have been extensively used for stabilization and long-term DNA storage. However, molecular level understanding of the role of the hydrogen bond of DNA with ILs in its stabilization is still inadequate. Two ILs, namely, 1,1,3,3-tetramethylguanidinium acetate (TMG) and 2,2-diethyl-1,1,3,3-tetramethylguanidinium acetate (DETMG), have been synthesized, of which TMG has a hydrogen bonding N-H group whereas DETMG does not contain any hydrogen bonding site. It has been found that both TMG and DETMG cations interact in the groove region of DNA; however, their mode of interaction is distinctly different, which causes the stabilization of DNA in the presence of TMG, whereas the effect is opposite in the case of DETMG. It is apparent from the data that only the accommodation of ILs in the groove region is not enough for the stabilization of DNA. MD simulation and spectroscopic studies combinedly indicate that the hydrogen bonding capability of the TMG cation enhances the hydrogen bonding between the Watson-Crick base pairs of DNA, resulting in its stabilization. In contrast, the bigger size as well as the absence of the hydrogen bonding site of the DETMG cation perturbs the minor groove width and base pair step parameters of DNA during its intrusion into the minor groove, which decreases the hydrogen bond between the Watson-Crick base pairs of DNA, leading to destabilization.

This article reports molecular interaction driven aqueous assembly of supramolecularly engineered amphiphilic macromolecules to cylindrical structure. Each polymer contains a single hydrophobic trialkoxybenzamide-linked naphthalene diimide (NDI) chromophore at the chain terminal as the supramolecular structure directing unit (SSDU). Irrespective of the structure of the appended hydrophilic polymer, H-bonding promoted J-aggregation among the NDI chromophore leads to the formation of thermally stable spherical micelle (critical aggregation concentration: 0.01-0.03 mM) which reorganizes to cylindrical micelle after a few hours. The reorganization time can be regulated by pH in the case of the anionic polymer as it affects the dynamics. Isothermal titration calorimetric (ITC) studies reveal positive Delta S values for assembly of all the polymers, reflecting the self-assembly process is favored by the entropy factor similar to the elegant examples in the biological domain. In contrast, a small molecule analogue of these polymers, having a short hydrophilic wedge (instead of a water-soluble polymer), shows a reverse trend, typically expected in a process of supramolecular organization. This can be attributed to the tightly packed J-aggregation of the NDI chromophore of the SSDU that compels a close packing of the hydrophilic polymer chains in the corona, leading to the release of the surrounding water molecules and causing entropy enhancement.

This Review Article highlights the utility of the fluorescence resonance energy transfer (FRET) to probe the dynamics and related issues in amphiphilic polymeric aggregates and supramolecular polymers. Amphiphilic polymers are more attractive compared to their small molecule analogues because they exhibit significantly lower critical aggregation concentration, relatively larger particle size (suitable for the enhanced permeation and retention effect), and a much slower dynamics of exchange between the unimer and the aggregate. Representative examples of exchange dynamics in amphiphilic polymer aggregates and their noncovalent encapsulation stability as a function of the structure of the macromolecule, cross-linking, environmental parameters, and biological conditions, as probed by FRET studies, have been included in this article. Further, related observations on the utility of FRET in studying the exchange dynamics in supramolecular polymers, particularly in aqueous medium, have been discussed at length, revealing a strong impact of chirality, side chain polarity, and other parameters. Overall, this Review Article brings out the strength of this technique to probe dynamics of aggregates and assembled systems, mostly in water medium, which has a paramount importance in designing future biomaterials.

This article describes self-assembly of supramolecularly engineered naphthalene-diimide (NDI)-derived amphiphiles NDI-1 and NDI-2. They have the same hydrophobic/hydrophilic balance but merely differ by a single functional group, amide or ester. They exhibit distinct self-assembly in water; NDI-1 forms hydrogel, which upon aging forms crystals, whereas NDI-2 forms micelles as revealed by in-depth structural analysis using cryo-TEM, dynamic light scattering, and small-angle X-ray scattering studies. These results suggest that the H-bonding among the amide groups fully regulates the self-assembly by overruling the packing parameters. Further, the present study elucidates sharp lower critical solution temperature exhibited by these π-amphiphiles, which has been extensively studied for many important applications of water-soluble polymers but hardly known in the literature of small-molecule surfactants. Control experiments with the same water-soluble hydrophilic wedge did not show such a property, confirming this to be a consequence of the supramolecular polymerization by extended amide-amide H-bonding and not inherent to the structure of the hydrophilic wedge containing oligo-oxyethylene chains.

This chapter collates several examples of self-assembled nanofibres, gels and other mesoscopic materials from a naphthalenediimide (NDI) chromophore driven by different weak interactions including π-stacking, H-bonding, charge-transfer (CT) interactions and solvophobic forces. Ease of synthesis, possibility of versatile imide substitution, efficient π-stacking and the ability to form an alternating stack with electron-rich donor chromophores by CT interactions make NDI an extremely versatile building block for exploring supramolecular chemistry. Furthermore, recent developments in the self-assembly of ring substituted NDIs with diverse photophysical and redox properties have been discussed. Aqueous self-assembly of suitably derivatized NDIs and their implications in the biological domain have also been included.

Amphiphilic polymers have emerged as an important class of materials owing to their ability to produce a diverse range of self-assembled structures with container properties that can be used to address growing challenges in biomedical applications. Thus, in-depth understanding on their aggregation properties is important fundamentally as well as from an application viewpoint. In this article we describe utilization of fluorescence resonance energy transfer (FRET) as a powerful tool to elucidate various physical properties of amphiphilic block copolymer aggregates at very low polymer concentration (∼10–7 M) which otherwise are difficult to achieve using other commonly used techniques such as microscopy, scattering, or external probe based spectroscopic techniques. We synthesized a prepolymer based on PEO-b-PMMA-co-PHEMA and subsequently utilized the hydroxy groups of the HEMA units to covalently attach with either a green (D) or a red (A) fluorescent dye (D–A pair suitable for FRET) to get D- and A-labeled...

Supramolecular architectures with the synchronized combination of various directional noncovalent forces are ubiquitous in biological systems. However, reports of such abiotic synthetic systems involving H-bonding in aqueous medium are rare due to the challenge faced in the formation of such structures by overcoming the competition from the water molecules. In this paper we have studied self-assembly of two structurally related naphthalene-diimide (NDI) conjugated bola-amphiphiles (NDI-1 and NDI-2) in water with an aim to realize the specific role of H-bonding among the hydrazide units present in one of the two building blocks (NDI-2) on the self-assembly. Both chromophores showed vesicular assembly in aqueous solution driven primarily by π-stacking among the NDI chromophores, which could be probed by UV-vis absorption spectra. Contrary to common belief, the lack of an H-bonding group in NDI-1 was found to be a boon in disguise in terms of the stability of the aggregates. Whereas NDI-2 aggregates showed LCST around 65-70 °C owing to the breaking of the H-bonds with increased temperature, the NDI-1 aggregates were found to be structurally intact until 90 °C, which may be attributed to the increased hydrophobicity introduced by the absence of the polar hydrazide group. Further concentration- and solvent-dependent UV-vis studies showed that NDI-1 formed assembled structure at greatly dilute solution and also in a solvent such as THF, confirming greater propensity for its self-assembly. As both bola-amphiphiles contain an electron-deficient NDI chromophore, interaction of their vesicles was studied with an externally added electron-rich pyrene derivative. Surprisingly, NDI-1 did not show any charge-transfer interaction with the donor, whereas NDI-2 could effectively intercalate, leading to a functional membrane with tunable surface functionalities. This was attributed to the additional stability of the intercalated state by H-bonding among the hydrazide units.