Christine Vauthier

Multimodal Dispersion of Nanoparticles: A Comprehensive Evaluation of Size Distribution with 9 Size Measurement Methods

ABSTRACTPurposeEvaluation of particle size distribution (PSD) of multimodal dispersion of nanoparticles is a difficult task due to inherent limitations of size measurement methods. The present work reports the evaluation of PSD of a dispersion of poly(isobutylcyanoacrylate) nanoparticles decorated with dextran known as multimodal and developed as nanomedecine.MethodsThe nine methods used were classified as batch particle i.e. Static Light Scattering (SLS) and Dynamic Light Scattering (DLS), single particle i.e. Electron Microscopy (EM), Atomic Force Microscopy (AFM), Tunable Resistive Pulse Sensing (TRPS) and Nanoparticle Tracking Analysis (NTA) and separative particle i.e. Asymmetrical Flow Field-Flow Fractionation coupled with DLS (AsFlFFF) size measurement methods.ResultsThe multimodal dispersion was identified using AFM, TRPS and NTA and results were consistent with those provided with the method based on a separation step prior to on-line size measurements. None of the light scattering batch methods could reveal the complexity of the PSD of the dispersion.ConclusionsDifference between PSD obtained from all size measurement methods tested suggested that study of the PSD of multimodal dispersion required to analyze samples by at least one of the single size particle measurement method or a method that uses a separation step prior PSD measurement.

Serial multiple crossed immunoelectrophoresis at a microscale: A stamp‐sized 2D immunoanalysis of protein C3 activation caused by nanoparticles

Crossed immunoelectrophoresis (C‐IE) is used to detect and quantify specific proteins. An application allowed the evaluation of complement system activation by nanomaterials. The work aimed to improve the C‐IE toward a higher throughput and less tedious method. A new concept was implemented to prepare and run agarose gels. The first and the second dimension of electrophoresis were performed on a single gel plate, prepared before the beginning of the analysis. Several samples were migrated simultaneously on the same migration line. Up to 35 analyses were run at once, providing stamp‐sized electrophoregrams (2.8 × 3 cm2) maintaining the performance of the original method performed on 5 × 7 cm2 gel slabs. Robustness and precision of the method were demonstrated through a validation approach using ANOVA. Handling, experimental duration, amount of reagents, and overall cost of one analysis were considerably reduced compared to the original method. With the same equipment, seven times more analyses can be performed in one run. C‐IE can be used to analyze many types of proteins. The new experimental modalities were suitable for the application developed in the present work that was to evaluate activation of protein C3 of the complement system triggered by nanomaterials.

Mucoadhesive properties of low molecular weight chitosan- or glycol chitosan- and corresponding thiomer-coated poly(isobutylcyanoacrylate) core-shell nanoparticles

Graphical abstract Figure. No caption available. ABSTRACT The aim of the present work was to evaluate the mucoadhesive properties of poly(isobutyl cyanoacrylate) (PIBCA) nanoparticles (NPs) coated with Low Molecular Weight (LMW) chitosan (CS)‐ and glycol chitosan (GCS)‐based thiomers as well as with the corresponding LMW unmodified polysaccharides. For this purpose, all the CS‐ and GCS‐based thiomers were prepared under simple and mild conditions starting from the LMW unmodified polymers CS and GCS. The resulting NPs were of spherical shape with diameters ranging from 400 to 600 nm and 187 to 309 nm, for CS‐ and GCS‐based NPs, respectively. The mucoadhesive characteristics of these core shell NPs were studied in Ussing chambers measuring the percentage of NPs stuck on the mucosal of fresh intestinal tissue after 2 h of incubation. Moreover, incubation of nanoparticle formulations with the intestinal tissue induced changes in transmucosal electrical resistance which were measured to gain information into the opening of tight junctions and to control the integrity of the mucosa. Thus, it was found that PIBCA NPs coated with the GCS–Glutathione conjugate (GCGPIBCA NPs) possessed the most favorable mucoadhesive performances. Moreover, both GCGPIBCA‐ and GCS‐N‐acetyl‐cysteine (GCNPIBCA)‐core‐shell NPs might induced an enlargement of the epithelial cell tight junctions. In conclusion, coating of PIBCA NPs with GCS‐based thiomers may be useful for improving the mucoadhesive and permeation properties of these nanocarriers.

Tuning complement activation and pathway through controlled molecular architecture of dextran chains in nanoparticle corona

The understanding of complement activation by nanomaterials is a key to a rational design of safe and efficient nanomedicines. This work proposed a systematic study investigating how molecular design of nanoparticle coronas made of dextran impacts on mechanisms that trigger complement activation. The nanoparticles used for this work consisted of dextran-coated poly(isobutylcyanoacrylate) (PIBCA) nanoparticles have already been thoroughly characterized. Their different capacity to trigger complement activation established on the cleavage of the protein C3 was also already described making these nanoparticles good models to investigate the relation between the molecular feature of their corona and the mechanism by which they triggered complement activation. Results of this new study show that complement activation pathways can be selected by distinct architectures formed by dextran chains composing the nanoparticle corona. Assumptions that explain the relation between complement activation mechanisms triggered by the nanoparticles and the nanoparticle corona molecular feature were proposed. These results are of interest to better understand how the design of dextran-coated nanomaterials will impact interactions with the complement system. It can open perspectives with regard to the selection of a preferential complement activation pathway or prevent the nanoparticles to activate the complement system, based on a rational choice of the corona configuration.

Characterization of nanomedicines: A reflection on a field under construction needed for clinical translation success

&NA; The nanotechnology revolution offers many expectations for the improvement of medicine treatments. At present, nanomedicine (NM) development is hampered by methodological barriers for a better characterization and a wider understanding of their in vivo behavior. While regulatory agencies setup guidelines to support NM translation from bench to bedside, the gap is still hardly overcome by main nanomedicines. One lever for filling this gap is a better characterization, thus increasing the global knowledge about the NM itself but also validate the confidence in terms of batch to batch reproducibility of such complex nano‐objects. Here, we review the current methodologies routinely used for clinical release of nanomedicine batches in compliance with official guidelines. We confront them to the extreme sharpness of biological systems and finally discuss future possible orientations for a better characterization of NMs, needed to bridge the gap between physicochemical properties and biological fate. Graphical abstract Figure. No Caption available. Abbreviations: AFM: Atomic force microscopy; API: Active pharmaceutical ingredient; BSAI: Biological Surface Adsorption Index; CARPA: Complement activation related pseudo allergy; CRO: Contract research organization; DCS: Differential centrifugal sedimentation; DLS: Dynamic light scattering; ELS: Electrophoretic light scattering; EM: Electron microscopy; EMA: European medicine agency; EPR: Enhanced permeation and retention; EUNCL: European Nanomedicine Characterization Laboratory; FDA: Food and drug administration; FFF: Field flow fractionation; GC‐MS: Gas chromatography ‐ mass spectrometry; ISO: International Standard Organization; LC‐MS: Liquid chromatography‐mass spectrometry; MALS: Multi‐angle light scattering; NC: Nanocarrier; NCL: Nanotechnology Characterization Laboratory; NIH: National Institute of Health; N. meningitidis: Neisseria meningitidis; NM: Nanomedicine; NMR: Nuclear magnetic resonance; NP: Nanoparticle; OECD: Organization for Economic Co‐operation and Development; PD: Pharmacodynamics; PEG: Polyethylene glycol; PK: Pharmacokinetics; PTA: Particle tracking analysis; SAXS: Small angle X‐ray scattering; SEC: Size exclusion chromatography; SEM: Scanning electron microscopy; SIMS: Secondary ion mass spectrometry; SPE: Solid phase extraction; STORM: Stochastic optical reconstruction microscopy; SUSTU: SUrface proteo‐ mics, Safety, Targeting, Uptake; TEM: Transmission electron microscopy; TRPS: Tunable resistive pulse sensing; WAXS: Wide angle X‐ray scattering.

Size of monodispersed nanomaterials evaluated by dynamic light scattering: Protocol validated for measurements of 60 and 203 nm diameter nanomaterials is now extended to 100 and 400 nm

In vivo fate of nanomaterials is influenced by the particle size among other parameters. Thus, Health Agencies have identified the size of nanomaterial as an essential physicochemical property to characterize. This parameter can be explored by dynamic light scattering (DLS) that is described in the ISO standard 22412:2008(E) and is one of the methods recognized by Health Agencies. However, no protocol of DLS size measurement has been validated over a large range of size so far. In this work, we propose an extension of validation of a protocol of size measurement by DLS previously validated with certified reference materials (CRM) at 60 and 203nm. The present work reports robustness, precision and trueness of this protocol that were investigated using CRM at 100 and 400nm. The protocol was robust, accurate and consistent with the ISO standard over the whole range of size that were considered. Expanded uncertainties were 4.4 and 3.6% for CRM at 100 and 400nm respectively indicating the reliability of the protocol. The range of application of the protocol previously applied to the size measurement of liposomes and polymer nanoparticles was extended to inorganic nanomaterial including silica nanoparticles.

towards quality assessed characterization of nanomaterial: Transfer of validated protocols for size measurement by dynamic light scattering and evaluation of zeta potential by electrophoretic light scattering

Quality control analysis of nanomaterials has been identified as a major issue to pursue their development in different industrial fields including nanomedicine. One difficulty is the lack of standardized and validated protocols suitable to achieve their characterization. In a previous work, we have developed standardized protocols for the evaluation of the size and zeta potential of nanomaterials based on methods described in the ISO standard and have performed validation of each one. The present work was aimed to transfer these protocols in three independent receiving laboratories. No official guideline was described in the literature to achieve such a transfer. A comparative study for receiving laboratories equipped with the same instrument as the sending laboratory was designed based on the Code of Federal Regulation edited by the Food and Drug Administration. For the receiving laboratory equipped with an instrument working at a different wavelength, a new validation was designed and applied. Corresponding statistical methods were used for the analysis of the results. A successful transfer of the protocols in all receiving laboratories was achieved. All laboratories recorded consistent results applying in blind the protocol of size measurements on two samples of nanomaterials from which included one reference.

Assessment of Complement Activation by Nanoparticles: Development of a SPR Based Method and Comparison with Current High Throughput Methods

PurposeA Surface Plasmon Resonance chip (SPR) was developed to study the activation of complement system triggered by nanomaterials in contact with human serum, which is an important concern today to warrant safety of nanomedicines.MethodsThe developed chip was tested for its specificity in complex medium and its longevity of use. It was then employed to assess the release of complement fragments upon incubation of nanoparticles in serum. A comparison was made with other current methods assessing complement activation (μC-IE, ELISA).ResultsThe SPR chip was found to give a consistent response for C3a release upon activation by nanoparticles. Results were similar to those obtained by μC-IE. However, ELISA detection of iC3b fragments showed an explained high non-specific background. The impact of sample preparation preceding the analysis was assessed with the newly develop SPR method. The removal of nanoparticles before analysis showed an important modification in the obtained response, possibly leading to false negative results.ConclusionThe SPR chip developed in this work allows for an automated assessment of complement activation triggered by nanoparticles with possibility of multiplexed analysis. The design of the chip proved to give consistent results of complement activation by nanoparticles.

Development of a Gas Chromatography Method for the Analysis of Copaiba Oil

A rapid, simple, precise and economic method for the quantification of main compounds of copaiba resin and essential oils (Copaifera langsdorffii Desf.) by gas chromatography (GC) has been developed and validated. Copaiba essential oil was extracted by hydrodistillation from the copaiba resin. Resin derivatization allowed the identification of diterpenes compounds. A gas chromatography-mass spectroscopy (GC/MS) method was developed to identify compounds composing the copaiba resin and essential oil. Then the GC/MS method was transposed to be used with a flame ionization detector (FID) and validated as a quantitative method. A good correlation between GC/MS and GC/FID was obtained favoring method transposition. The method showed satisfactory sensitivity, specificity, linearity, precision, accuracy, limit of detection and limit of quantitation for β-caryophyllene, α-humulene and caryophyllene oxide analyses in copaiba resin and essential oils. The main compounds identified in copaiba essential oil were β-bisabolene (23.6%), β-caryophyllene (21.7%) and α-bergamotene (20.5%). Copalic acid methyl ester (15.6%), β-bisabolene (12.3%), β-caryophyllene (7.9%), α-bergamotene (7.1%) and labd-8(20)-ene-15,18-dioic acid methyl ester (6.7%) were diterpenes identified from the derivatized copaiba resin. The proposed method is suitable for a reliable separation, identification and quantification of compounds present in copaiba resin and essential oil. It could be proposed as an analytical method for the analysis of copaiba oil fraction in raw and essential oil parent extracts and after they have been incorporate in pharmaceutical formulations.