Soft Corona Research Articles

Dynamics and Thermodynamics Study of the Adsorption of Lysozyme to 2D SnSe Nanoflake Surfaces with Associated Unfolding

AbstractIn this work, we explore the interaction as well as corona formation and interfacial phenomena of lysozyme (LYZ) and 2D SnSe nanoflake. The optical results indicate formation of ground state complex with an apparent association constant of 1.5 × 103 (mol)−1. The dynamics of the in situ interaction exhibited surface reorganization (t2 = 73.5 min) and unfolding, which were more prominent than binding (t1 = 3 min) for tryptophan residues of LYZ. The formations of soft and hard corona of the LYZ structure were clearly observed from HRTEM. The emission quenching exhibited tertiary deformation of the LYZ structure with positive cooperative binding (n > 1) and dynamics of quenching followed the sigmoidal Boltzmann nature of the energy transfer process. The ultrafast lifetime of the bioconjugate varied from 1.9–1.35 ns with a maximum 63% energy transfer efficiency within 300 min of the corona evolution. The decrement of α helix (58.87%–21.78%) and increment of β sheet (5.27%–23.98%) were observed after interaction, with the conversion of large amount of α helix into β sheet. The ITC results showed that the injection heat results followed two‐site binding model, and the exothermic binding was dominate over the interaction. The second phase binding is driven by larger positive entropy, where the first binding site has a stronger association.

Nanoplastic Size and Surface Chemistry Dictate Decoration by Human Saliva Proteins.

Environmental pollution with plastic polymers has become a global problem, leaving no continent and habitat unaffected. Plastic waste is broken down into smaller parts by environmental factors, which generate micro- and nanoplastic particles (MNPPs), ultimately ending up in the human food chain. Before entering the human body, MNPPs make their first contact with saliva in the human mouth. However, it is unknown what proteins attach to plastic particles and whether such protein corona formation is affected by the particle's biophysical properties. To this end, we employed polystyrene MNPPs of two different sizes and three different charges and incubated them individually with saliva donated by healthy human volunteers. Particle zeta potential and size analyses were performed using dynamic light scattering complemented by nanoliquid chromatography high-resolution mass spectrometry (nLC/HRMS) to qualitatively and quantitatively reveal the protein soft and hard corona for each particle type. Notably, protein profiles and relative quantities were dictated by plastic particle size and charge, which in turn affected their hydrodynamic size, polydispersity, and zeta potential. Strikingly, we provide evidence of the latter to be dynamic processes depending on exposure times. Smaller particles seemed to be more reactive with the surrounding proteins, and cultures of the particles with five different cell lines (HeLa, HEK293, A549, HepG2, and HaCaT) indicated protein corona effects on cellular metabolic activity and genotoxicity. In summary, our data suggest nanoplastic size and surface chemistry dictate the decoration by human saliva proteins, with important implications for MNPP uptake in humans.

Formation of a ‘protein corona’ on the Fe3O4@SiO2 nanoparticles surface

The patterns of interaction of the nanoparticles surface with protein components of biosystems have been studied on the example of the interaction of magnetic nanoparticles of the core–shell structure with albumin. Data on particle sizes and their spectra in the visible region in the presence of different quantity of protein have been obtained and analyzed by sorption, spectral and dynamic light scattering methods and albumin sorption isotherms in various media. The formation of the ordered protein monolayer (so-called ‘hard corona’) with subsequent forming of the multilayer of a different structure (‘soft corona’) has been experimentally shown; it demonstrates that it is possible to computationally distinguish between mono- and multi-layer using data from sorption experiments.

Ovalbumin interaction with polystyrene and polyethylene terephthalate microplastics alters its structural properties

Contaminating microplastics can interact with food proteins in the food matrix and during digestion. This study investigated adsorption of chicken egg protein ovalbumin to polystyrene (PS, 110 and 260 μm) and polyethylene terephthalate (PET, 140 μm) MPs in acidic and neutral conditions and alterations in ovalbumin structure. Ovalbumin adsorption affinity depended on MPs size (smaller > larger), type (PS > PET) and pH (pH 3 > pH 7). In bulk solution, MPs does not change ovalbumin secondary structure significantly, but induces loosening (at pH 3) and tightening (at pH 7) of tertiary structure. Formed soft corona exclusively consists of full length non-native ovalbumin, while in hard corona also shorter ovalbumin fragments were found. At pH 7 soft corona ovalbumin has rearranged but still preserved level of ordered secondary structure, resulting in preserved thermostability and proteolytic stability, but decreased ability to form fibrils upon heating. Secondary structure changes in soft corona resemble changes in native ovalbumin induced by heat treatment (80 °C). Ovalbumin is abundantly present in corona around microplastics also in the presence of other egg white proteins. These results imply that microplastics contaminating food may bind and change structure and functional properties of the main egg white protein.

Molecular modeling of the carbohydrate corona formation on a polyvinyl chloride nanoparticle and its impact on the adhesion to lipid bilayers.

The pervasive presence of nanoplastics (NPs) in the environment has gained increasing attention due to their accumulation in living organisms. These emerging contaminants inevitably interact with extracellular polymeric substances along respiratory or gastrointestinal tracts, and diverse organic coating on the surface of NPs, known as bio- or eco-corona, is formed. Although its impact on altering the NP properties and potential cell internalization has been extensively examined, studies on its role in NP partitioning in the cell membrane are elusive yet. In this work, molecular dynamics is used to investigate the formation of chitosan (CT) corona centered on a polyvinyl chloride (PVC) nanoparticle and the uptake of the resulting complex onto lipid membranes. Coarse-grained models compatible with the newly developed Martini 3.0 force field are implemented for the two polymers employing the atomistic properties as targets in the parameterization. The reliability of the coarse-grained polymer models is demonstrated by reproducing the structural properties of the PVC melt and of solvated CT strands, as well as by determining the conformation adopted by the latter at the NP surface. Results show that the spontaneous binding of CT chains of high and intermediate protonation degrees led to the formation of soft and hard corona that modulates the interaction of PVC core with model membranes. The structural changes of the corona adsorbed at the lipid-water interface enable a subsequent transfer of the NP to the center of the saturated lipid membranes and a complete or partial transition to a snorkel conformation depending on the hydrophilic/hydrophobic balance in the CT-PVC complex. Overall, the computational investigation of the coarse-grained model system provides implications for understanding how the eco-corona development influences the uptake and implicit toxicology of NPs.

The In Vivo Biological Fate of Protein Corona: A Comparative PET Study of the Fate of Soft and Hard Protein Corona in Healthy Animal Models.

Radiolabeling and nuclear imaging techniques are used to investigate the biodistribution patterns of the soft and hard protein corona around poly (lactic-co-glycolic acid) nanoparticles (PLGA NPs) after administration to healthy mice. Soft and hard protein coronas of 131I-labeled BSA or 131I-labeled serum are formed on PLGA NPs functionalized with either polyehtylenimine (PEI) or bovine serum albumin (BSA). The exchangeability of hard and soft corona is assessed in vitro by gamma counting exposing PLGA NPs with corona to non-labeled BSA, serum, or simulated body fluid. PEI PLGA NPs form larger and more stable coronas than BSA PLGA NPs. Soft coronas are more exchangeable than hard ones.The in vivo fate of PEI PLGA NPs coated with preformed 18F-labeled BSA hard and soft coronas is assessed by positron emission tomography (PET)following intravenous administration. While the soft corona shows a biodistribution similar to free 18F BSA with high activity in blood and kidney, the hard corona follows patterns characteristic of nanoparticles, accumulating in the lungs, liver, and spleen. These results show that in vivo fates of soft and hard corona are different, and that soft corona is more easily exchanged with proteins from the body, while hard corona is largely retained on the nanoparticle surface.

Areas of anomalous properties as function of shape of potential

By molecular dynamics and Monte Carlo simulations we have determined regions and hierarchy of anomalies in the purely repulsive core-softened system. The particles are repelling each other through an isotropic core-softened potential with two characteristics lengths. The first is a hard core with one diameter and the second a soft corona with at larger distance. The thermodynamics and structure was assessed and anomalous regions determined. We have shown that in two dimensional systems the size of 100 particles is enough to get correct details by comparing these results to data from simulations 1000 and even 10000 particles. The model has anomaly of density, diffusivity and structure and their hierarchy is water-like no matter what parameters are used. We have shown that in two dimensional systems the size of 100 particles is enough to get correct details by comparing these results to data from simulations of 1000 and even 10000 particles.

Multivariate Analysis of Protein-Nanoparticle Binding Data Reveals a Selective Effect of Nanoparticle Material on the Formation of Soft Corona.

When nanoparticles are introduced into the bloodstream, plasma proteins accumulate at their surface, forming a protein corona. This corona affects the properties of intravenously administered nanomedicines. The firmly bound layer of plasma proteins in direct contact with the nanomaterial is called the "hard corona". There is also a "soft corona" of loosely associated proteins. While the hard corona has been extensively studied, the soft corona is less understood due to its inaccessibility to analytical techniques. Our study used dynamic light scattering to determine the dissociation constant and thickness of the protein corona formed in solutions of silica or gold nanoparticles mixed with serum albumin, transferrin or prothrombin. Multivariate analysis showed that the nanoparticle material had a greater impact on binding properties than the protein type. Serum albumin had a distinct binding pattern compared to the other proteins tested. This pilot study provides a blueprint for future investigations into the complexity of the soft protein corona, which is key to developing nanomedicines.

Blood Compatibility of Drug-Inorganic Hybrid in Human Blood: Red Blood Cell Hitchhiking and Soft Protein Corona.

A drug-delivery system consisting of an inorganic host-layered double hydroxide (LDH)-and an anticancer drug-methotrexate (MTX)-was prepared via the intercalation route (MTX-LDH), and its hematocompatibility was investigated. Hemolysis, a red blood cell counting assay, and optical microscopy revealed that the MTX-LDH had no harmful toxic effect on blood cells. Both scanning electron microscopy and atomic force microscopy exhibited that the MTX-LDH particles softly landed on the concave part inred blood cells without serious morphological changes of the cells. The time-dependent change in the surface charge and hydrodynamic radius of MTX-LDH in the plasma condition demonstrated that the proteins can be gently adsorbed on the MTX-LDH particles, possibly through protein corona, giving rise to good colloidal stability. The fluorescence quenching assay was carried out to monitor the interaction between MTX-LDH and plasma protein, and the result showed that the MTX-LDH had less dynamic interaction with protein compared with MTX alone, due to the capsule moiety of the LDH host. It was verified by a quartz crystal microbalance assay that the surface interaction between MTX-LDH and protein was reversible and reproducible, and the type of protein corona was a soft one, having flexibility toward the biological environment.

Protein corona investigations of polyplexes with varying hydrophobicity – From method development to in vitro studies

In the field of non-viral drug delivery, polyplexes (PXs) represent an advanced investigated and highly promising tool for the delivery of nucleic acids. Upon encountering physiological fluids, they adsorb biological molecules to form a protein corona (PC), that influence PXs biodistribution, transfection efficiencies and targeting abilities. In an effort to understand protein – PX interactions and the effect of PX material on corona composition, we utilized cationic branched 10 kDa polyethyleneimine (b-PEI) and a hydrophobically modified nylon-3 polymer (NM0.2/CP0.8) within this study to develop appropriate methods for PC investigations. A centrifugation procedure for isolating hard corona – PX complexes (PCPXs) from soft corona proteins after incubating the PXs in fetal bovine serum (FBS) for PC formation was successfully optimized and the identification of proteins by a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method clearly demonstrated that the PC composition is affected by the underlying PXs material. With regard to especially interesting functional proteins, which might be able to induce active targeting effects, several candidates could be detected on b-PEI and NM0.2/CP0.8 PXs. These results are of high interest to better understand how the design of PXs impacts the PC composition and subsequently PCPXs-cell interactions to enable precise adjustment of PXs for targeted drug delivery.

Protein Adsorption on Carboxy-Functionalized Microparticles Revealed by Zeta Potential and Absorption Spectroscopy Measurements

Abstract Addition of micro/nanoparticles to a protein solution leads to the formation of a protein layer on the particle surface, called a protein corona. We investigate here the adsorption behavior of myoglobin, hemoglobin, cytochrome-c, and lysozyme on carboxy-functionalized polystyrene microparticles using zeta potential and absorption spectroscopy measurements. The observed adsorption behavior differs according to the method of detection: monolayer for zeta potential and multilayer for absorption spectroscopy. Langmuir-type monolayer adsorption is observed with zeta potential measurements, because zeta potential (ζ) responds only to the charge density at the outermost protein layer. Multilayer adsorption is observed by absorption spectroscopy. Spectroscopic results were analyzed by the Guggenheim-Anderson-de Boer (GAB) model, which comprises a hard corona formed by strong interaction between the protein and the particle surface and a soft corona formed by weak interaction between adsorbed and bulk-solution proteins. The extent of hard and soft corona formation depends on pH. When a protein monolayer is prepared by covalent modification of the particle surface, the number of layers in the protein corona decreases relative to the case of protein adsorption on bare particles. This result demonstrates that electrostatic interactions between the protein and particle surface play a key role in the formation of a protein corona.

The D-amino acid oxidase-carbon nanotubes: evaluation of cytotoxicity and biocompatibility of a potential anticancer nanosystem

The ‘enzyme prodrug therapy’ represents a promising strategy to overcome limitations of current cancer treatments by the systemic administration of prodrugs, converted by a foreign enzyme into an active anticancer compound directly in tumor sites. One example is D-amino acid oxidase (DAAO), a dimeric flavoenzyme able to catalyze the oxidative deamination of D-amino acids with production of hydrogen peroxide, a reactive oxygen species (ROS), able to favor cancer cells death. A DAAO variant containing five aminoacidic substitutions (mDAAO) was demonstrated to possess a better therapeutic efficacy under low O2 concentration than wild-type DAAO (wtDAAO). Recently, aiming to design promising nanocarriers for DAAO, multi-walled carbon nanotubes (MWCNTs) were functionalized with polyethylene glycol (PEG) to reduce their tendency to aggregation and to improve their biocompatibility. Here, wtDAAO and mDAAO were adsorbed on PEGylated MWCNTs and their activity and cytotoxicity were tested. While PEG-MWCNTs-DAAOs have shown a higher activity than pristine MWCNTs-DAAO (independently on the DAAO variant used), PEG-MWCNTs-mDAAO showed a higher cytotoxicity than PEG-MWCNTs-wtDAAO at low O2 concentration. In order to evaluate the nanocarriers’ biocompatibility, PEG-MWCNTs-DAAOs were incubated in human serum and the composition of protein corona was investigated via nLC-MS/MS, aiming to characterize both soft and hard coronas. The mDAAO variant has influenced the bio-corona composition in both number of proteins and presence of opsonins and dysopsonins: notably, the soft corona of PEG-MWCNTs-mDAAO contained less proteins and was more enriched in proteins able to inhibit the immune response than PEG-MWCNTs-wtDAAO. Considering the obtained results, the PEGylated MWCNTs conjugated with the mDAAO variant seems a promising candidate for a selective antitumor oxidative therapy: under anoxic-like conditions, this novel drug delivery system showed a remarkable cytotoxic effect controlled by the substrate addition, against different tumor cell lines, and a bio-corona composition devoted to prolong its blood circulation time, thus improving the drug’s biodistribution.

Identification and characterization of soft protein corona absorbed on iron oxide nanoparticles

Nanomedicines have recently garnered significant attention due to their great potential to improve the efficiency of disease diagnosis and treatment. Nonetheless, a wide gap still exists between innovative research and clinical application because of limited understanding of the protein corona formed on the nanocarrier's surface. When NPs are introduced to living systems, the protein corona rather than the originally expected surface of the nanoparticle is initially recognized, further affecting immune clearance and tumor-specific delivery. This study found that after incubation with human plasma, the protein corona formed on the iron oxide nanoparticle's (IONP) surface significantly increased the uptake of IONPs in the leukemic cell line MOLM13. The formation and thicknesses of the protein coronas were measured by dynamic light scattering, nanoparticle tracking analysis, and flow cytometry, and the hard corona was measured by transmission electron microscopy. The size distribution of IONPs before and after incubation with plasma was measured, and the change in particle size was calculated to derive the thickness of the soft corona (7.58±3.94 nm) by subtracting the thickness of the hard corona from that of the full protein corona. The results can provide an approximation of the soft corona area after plasma incubation and contribute to the understanding of the features of the protein corona of nanoparticles with different physical and chemical properties.

How Dimensionality Affects the Structural Anomaly in a Core-Softened Colloid

The interaction between hard core–soft shell colloids are characterized by having two characteristic distances: one associated with the penetrable, soft corona and another one corresponding to the impenetrable core. Isotropic core-softened potentials with two characteristic length scales have long been applied to understand the properties of such colloids. Those potentials usually show water-like anomalies, and recent findings have indicated the existence of multiple anomalous regions in the 2D limit under compression, while in 3D, only one anomalous region is observed. In this direction, we perform molecular dynamics simulations to unveil the details about the structural behavior in the quasi-2D limit of a core-softened colloid. The fluid was confined between highly repulsive solvophobic walls, and the behavior at distinct wall separations and colloid densities was analyzed. Our results indicated a straight relation between the 2D- or 3D-like behavior and layer separation. We can relate that if the system behaves as independent 2D-layers, it will have a 2D-like behavior. However, for some separations, the layers are connected, with colloids hopping from one layer to another, thus having a 3D-like structural behavior. These findings fill the gap in the depiction of the anomalous behavior from 2D to 3D.

Peptide Self-Assembly into Amyloid Fibrils at Hard and Soft Interfaces-From Corona Formation to Membrane Activity.

Peptides and proteins are exposed to a variety of interfaces in a physiological environment, such as cell membranes, protein nanoparticles (NPs), or viruses. These interfaces have a significant impact on the interaction, self-assembly, and aggregation mechanisms of biomolecular systems. Peptide self-assembly, particularly amyloid fibril formation, is associated with a wide range of functions; however, there is a link with neurodegenerative diseases, such as Alzheimer's disease. This review highlights how interfaces affect peptide structure and the kinetics of aggregation leading to fibril formation. In nature, many surfaces are nanostructures, such as liposomes, viruses, or synthetic NPs. Once exposed to a biological medium, nanostructures are coated with a corona, which then determines their activity. Both accelerating and inhibiting effects on peptide self-assembly have been observed. When amyloid peptides adsorb to a surface, they typically concentrate locally, which promotes aggregation into insoluble fibrils. Starting from a combined experimental and theoretical approach, models that allow for a better understanding of peptide self-assembly near hard and soft matter interfaces are introduced and reviewed. Research results from recent years are presented and relationships between biological interfaces, such as membranes and viruses, and amyloid fibril formation are proposed.

In situ characterization of nanoparticle protein corona using real-time 3D single-particle tracking.

Nanoparticles (NPs) adsorb proteins when they are exposed to biological fluids, forming a dynamic protein corona that consists of tightly bound “hard” corona and rapidly exchanged “soft” corona. This protein layer alters the NPs’ surface identity and affects their fate in vivo. However, current methods fail to directly investigate single NP-protein interactions in the solution phase due to the fast NP diffusion and protein exchange dynamics. To address this gap, we introduce a real-time lock-on protein corona spectroscopy based on previously developed 3D single molecule active real-time tracking (3D-SMART), primarily using polystyrene NP (PSNP) and bovine serum albumin (BSA) as a model system. 3D-SMART uses real-time photon arrival information to estimate the PSNP's position within a rapid 3D laser scanning pattern. The position estimate is then used to move a piezoelectric stage to counteract the PSNP's diffusion, keeping the NP-protein complex locked in the center of detection volume for spectral interrogation. Consequently, the 3D trajectory of the NP-protein complex and the fluorescence traces of the associated proteins can be synchronously measured for size measurement and hard corona quantification. Moreover, we applied a lock-in filtering-based algorithm to the protein intensity trace to extract the full protein corona signal in situ at signal-to-background ratios of 1%. It was discovered that the full corona consists of a sub-monolayer of BSA on PSNP, which contains twice the quantity of proteins compared to hard corona measurements, indicating that half of the full protein corona can be classified as soft corona. To expand this method to smaller NP-protein systems, we have further implemented a Galvo mirror as control actuator with response time five times faster than the piezoelectric stage, opening the possibility of studying a wider range of transient NP-protein interaction in vivo.

The Effect of Mesogenic Coronas on the Type and Anisotropy of Gold Nanoparticle Superlattices: When Can the Tail Wag the Dog?

The correlation between the size of nanoparticles, the structure and shape of mesogenic ligands and the ensuing assembly behaviour is not really understood. Closer inspection shows very surprising features. Here, 2- and 4-nm gold nanoparticles (NPs) were synthesized, and grafted with a forked ligand containing two rod-like mesogens in its branches: one cholesterol, the other with azobenzene. The 4-nm NPs also contained n-hexylthiol as co-ligand. They were found to form a FCC cubic superlattice, whereas the 2-nm NPs form hexagonal HCP with weak birefringence, hence with partially oriented ligands. The structures were compared with those of related systems containing a range of different azobenzene-to-cholesterol ratios, all giving body-centred tetragonal superlattices with various degrees of anisotropy. Geometric analysis is presented in terms of the asphericity of the NPs' surroundings, requirement for space-filling and structural anisotropy. Some general rules are derived to help design the soft corona around the NPs in order to obtain superlattices with the desired structure and anisotropy.

Differences in protein distribution, conformation, and dynamics in hard and soft coronas: dependence on protein and particle electrostatics.

We perform all-atom molecular dynamics simulations of a 9 nm-thick protein layer, which consists of serum albumin (SA) or a mixture of SA and immunoglobulin gamma-1, formed on 10 nm-sized cationic, anionic, and neutral polystyrene particles. More than half of the proteins are densely concentrated within a distance of ∼3 nm from the particle surface, while fewer proteins are broadly distributed in the range of 3-9 nm from the particle. This compares favorably with the experimental observations of a hard corona as the first layer adjacent to the particle and a soft corona as a loose protein-network. The conformation and diffusivity of the proteins vary in different positions of the layer, and are to an extent dependent on the protein and particle electrostatics. These, combined with free energy calculations, show that the protein and particle charges do not significantly modify the strength of protein-particle binding but do influence the distribution of proteins in the layer. In particular, a free protein more strongly binds to the complex of a protein and particle than to either one, showing the synergistic effect of already adsorbed proteins and a particle. This helps explain the experimental observation regarding the formation of a denser protein layer and the stronger protein-protein interaction in the hard corona than the soft corona.

Quantitative comparison of the protein corona of nanoparticles with different matrices

Nanoparticles (NPs) are paving the way for improved treatments for difficult to treat diseases diseases; however, much is unknown about their fate in the body. One important factor is the interaction between NPs and blood proteins leading to the formation known as the “protein corona” (PC). The PC, consisting of the Hard (HC) and Soft Corona (SC), varies greatly based on the NP composition, size, and surface properties. This highlights the need for specific studies to differentiate the PC formation for each individual NP system. This work focused on comparing the HC and SC of three NPs with different matrix compositions: a) polymeric NPs based on poly(lactic-co-glycolic) acid (PLGA), b) hybrid NPs consisting of PLGA and Cholesterol, and c) lipidic NPs made only of Cholesterol. NPs were formulated and characterized for their physico-chemical characteristics and composition, and then were incubated in human plasma. In-depth purification, identification, and statistical analysis were then performed to identify the HC and SC components. Finally, similar investigations demonstrated whether the presence of a targeting ligand on the NP surface would affect the PC makeup. These results highlighted the different PC fingerprints of these NPs, which will be critical to better understand the biological influences of the PC and improve future NP designs.

Phase Control of Colloid-like Block Copolymer Micelles by Tuning Size Distribution via Thermal Processing

The formation of a Frank–Kasper (FK) phase in a one-component block copolymer (bcp) has been attributed to the sufficiently large conformational asymmetry parameter that leads to the deformation of the micellar core into the polyhedral shape templated by the Voronoi cells for relieving the packing frustration of the coronal blocks. Here we present the insight into the evolution of body-centered cubic (BCC) and Laves C14 phases from the corresponding metastable liquidlike packing (LLP) phases in a conformationally symmetric poly(2-vinylpyridine)-block-poly(dimethylsiloxane) (P2VP-b-PDMS) forming the colloid-like micelle with a hard P2VP core and a soft PDMS corona at the temperature of micelle ordering (Tgcorona < Ta < Tgcore). Different thermal processing conditions were applied to produce four distinct types LLP phase characterized by different average micelle size and size dispersity. The LLP phase was found to transform to BCC, C14 and a reorganized LLP phase with increasing polydispersity of particle size, showing the existence of a proper range of size dispersity for the formation of the Laves phase. The real-space analysis of the local environments of the particles revealed that the C14 phase can accommodate the particles with a broader range of interparticle distance and a less uniform local environment, such that the PDMS coronal blocks of the micelles with greater size dispersity suffered a lower degree of packing frustration when they organized into a C14 lattice. In view of the colloid-like nature of the micelle, a size fractionation process found in the crystallization of polydisperse colloidal particles was proposed as the mechanism for directing the effective development of C14 phase from the LLP phase. Due to the metastable nature of the ordered phases, their order–disorder transition temperatures were found to depend strongly on the structures of the LLP phases from which they developed and a strong memory effect underlying the phase transitions was identified.