Highly hydrophilic surfaces from polyglycidol grafts with dual antifouling and specific protein recognition properties.

Homopolymer grafts from α-tert-butoxy-ω-vinylbenzyl-polyglycidol (PGL) were prepared on gold and stainless steel (SS) substrates modified by 4-benzoyl-phenyl (BP) moieties derived from the electroreduction of the parent salt 4-benzoyl benzene diazonium tetrafluoroborate. The grafted BP aryl groups efficiently served to surface-initiate photopolymerization (SIPP) of PGL. In similar conditions, SIPP of hydroxyethyl methacrylate (HEMA) permitted the production of PHEMA grafts as model surfaces. Water contact angles were found to be 66°, 15°, and 0° for SS-BP, SS-PHEMA, and SS-PPGL, respectively. The spontaneous spreading of water drops on SS-PPGL was invariably observed with 1.5 µL water drops. PPGL thus appears as a superhydrophilic polymer. Resistance to nonspecific adsorption of proteins of PPGL and PHEMA grafts on gold was evaluated by surface plasmon resonance (SPR) using antibovine serum albumin (anti-BSA). The results conclusively show that PPGL-grafts exhibit enhanced resistance to anti-BSA adsorption compared to the well-known hydrophilic PHEMA. PPGL grafts were further modified with BSA through the carbonyldiimidazole activation of the OH groups providing immunosensing surfaces. The so-prepared PPGL-grafted BSA hybrids specifically interacted with anti-BSA in PBS as compared to antimyoglobin. It is clear that the superhydrophilic character of PPGL grafts opens new avenues for biomedical applications where surfaces with dual functionality, namely, specific protein grafting together with resistance to biofouling, are required.

Capillary columns for reversed-phase CEC prepared via surface functionalization of polymer monolith with aromatic selectors.

Macroporous crosslinked organic polymers based on N-acryloxysuccinimide (NAS) and ethylene dimethacrylate (EDMA) were prepared in the confines of 75 µm id fused-silica capillaries by photoinitiated free radical copolymerization in the presence of 2-2'-azobisisobutyronitrile as initiator and toluene as porogen. Monoliths with good mechanical strength, large porosity as well as surface reactive sites (succinimide leaving groups) could be obtained. Nucleophilic aromatic derivatives, namely benzylamine, phenylbutylamine and naphthylamine were grafted on the monolith surface to introduce π-conjugated ligands to develop particular selectivity. Successful achievement of the post-copolymerization functionalization was ascertained on the basis of in situ chemical characterization by means of Raman spectroscopy. Electrochromatographic properties of π-functionalized poly(NAS-co-EDMA) regarding alkylbenzenes, polycyclic aromatic hydrocarbons, anilines and phenols were evaluated in terms of retention, selectivity and resolution. The as-designed monolithic columns exhibited π-π interaction in addition to hydrophobic interaction due to the aromatic and non-polar nature of the surface-grafted aromatic selectors. One of the major results of this study is that monolithic columns with mixed selectivity providing high potentiality for the separation of solutes with varied chemical structure variation can be obtained by the surface grafting of the appropriate selector. Herein, an example is given for the phenylbutylamine functionalized poly(NAS-co-EDMA) where the butyl and phenyl fragments afford enhanced hydrophobic and π-selectivity, respectively.

Zwitterionic polymeric monoliths for HILIC/RP mixed mode for CEC separation applications.

Polymer-based monoliths with zwitterionic surface character were synthesized in capillary columns following a two-step approach to provide versatile electrochromatographic stationary phases exhibiting potentiality of both hydrophilic interaction and RP separation modes. UV-initiated free radical copolymerization of N-acryloxysuccinimide and ethylene dimethacrylate was performed using azobisisobutyronitrile as initiator and toluene as porogen. One of the originalities of this approach relies on the dual role of the N-acryloxysuccinimide monomer that is successively used during the preparation protocol to first covalently graft chromatographic selectors on the monolith surface via simple nucleophilic substitution reaction and then to generate negative charges through hydrolysis of remaining N-hydroxysuccinimide units. In this respect, the grafting of hexyldiamine affords potential cationic surface charges. It is shown that it is possible to tune, controlling the pH of the mobile phase, the intensity and direction of the generated EOF. Moreover, the nature of the interfacial interaction process responsible for the observed separations is well governed by the composition of the mobile phase. Polymer backbone hydrophilization is proposed as an efficient way to improve the HILIC behavior of poly(N-acryloxysuccinimide-co-ethylene dimethacrylate) based monolithic CEC columns together with the grafting of an alkyldiamine incorporating a shorter aliphatic segment.

Biofunctionalized tilted Fiber Bragg Gratings for label-free immunosensing.

We present a study aimed at developing a label-free optical fiber biosensor for detection and quantification of biomolecules in real-time. The biosensor based on a Tilted Fiber Bragg Grating (TFBG) transduces a binding event between the probe and target molecules into a change in the refractive index of the medium surrounding the fiber. This work describes the experimental results obtained with three methods for immobilizing biomolecular probes on a TFBG silica cladding surface. Bovine serum albumin (BSA) and anti-BSA are used to assess the performances of the TFBG based biosensor in each configuration.

DNA immobilisation procedures for surface plasmon resonance imaging (SPRI) based microarray systems.

Two different surface chemistries have been studied for the development of surface plasmon resonance imaging (SPRI) based DNA microarray affinity sensors: (1) 11-mercaptoundecanoic acid-poly(ethylenimine) (MUA-PEI) and (2) dextran procedures. The MUA-PEI method consists of assembling a multilayer on the basis of electrostatic interactions formed with: 11-mercaptoundecanoic acid (MUA), poly(ethylenimine) (PEI) and extravidin layers. The dextran procedure involves assembling a multilayer formed with 11-mercaptoundecanol, dextran and streptavidin layers, which are linked by covalent bonds. The oligonucleotide probes are immobilised onto the sensor surface as spots forming a matrix 14x14, which is spotted by a robot, while the target sequences are free in solution. The system allows the interaction (hybridisation) monitoring, in real-time and in parallel, of unlabeled oligonucleotide solution targets to oligonucleotide probes immobilised on a 196 spots matrix. Using oligonucleotides as probes and targets, both functionalised surfaces have been evaluated in view of their application to the diagnosis of gene mutations involved in human diseases. In particular, we demonstrate the ability to detect, in parallel, several mutations causing human cystic fibrosis (CF), which lie within exon 10 of the human cystic fibrosis transmembrane conductance regulator (CFTR) gene. The immobilised probes were complementary to sequences corresponding the mutant or wild type alleles. Two deletions of three bases (DeltaF508 and DeltaI507) and four single nucleotide polymorphisms (M470V, Q493X, V520F and 1716 G>A) were investigated. In both functionalised surfaces, the system showed the capacity to discriminate normal and mutant sequences differing by a single base.

In situ functionalization of N-acryloxysuccinimide-based monolith for reversed-phase electrochromatography.

Capillary electrochromatography (CEC) monolithic columns were prepared following a two-step synthetic pathway based on (i) UV-induced in situ radical polymerization of N-acryloxysuccinimide (NAS) and ethylene dimethacrylate (EDMA) and (ii) in situ functionalization of the NAS-containing monolithic matrix with various alkylamines. The first synthetic step was performed using toluene as a porogenic solvent. The successful grafting of the alkylamines onto the reactive matrix was confirmed on the basis of qualitative analysis of Raman spectra recorded before and after the chemical modification step. All the electrochromatographic results indicate a strong dependence of the retention, efficiency and selectivity of the monolithic columns on small variations of mobile phase composition and nature of the grafted aliphatic selector in agreement with the typical reversed-phase behaviour. Van Deemter plots for a series of alkylbenzene homologues injected on a column bearing hexyl-segments as side chains are also presented.

Supramolecular interactions between beta-cyclodextrin and hydrophobically end-capped poly(ethylene glycol)s: a quartz crystal microbalance study.

In this study, the supramolecular interactions occurring between beta-cyclodextrin-based surfaces and macromolecular chains modified at one end with naphthyl, adamantyl, or phenyladamantyl hydrophobic groups were investigated by means of a quartz crystal microbalance. beta-Cyclodextrin-functionalized gold electrodes were obtained through the amide-coupling reaction between mono-6-deoxy-6-amino-beta-cyclodextrin and 11-mercaptoundecanoic acid self-assembled monolayer allowing the reproducible preparation of densely grafted surfaces with host properties. The interaction data obtained for the three different modified poly(ethylene glycol)s are in good agreement with our previous studies performed by high performance liquid chromatography and surface plasmon resonance. This evidences that the driving force for the supramolecular interaction is based on the inclusion of the hydrophobic terminal group of the chains within the cyclodextrin cavities. The reversibility of the inclusion process was proven through the regeneration of the original host properties of the sensing surfaces using sodium dodecylsulfate as a competitor for the desorption of the poly(ethylene glycol) chains.

Retention properties of hydrophobically end-capped poly(ethylene glycol)s on a beta-cyclodextrin support.

High-performance liquid chromatography (HPLC) was used to examine the retention behavior of monomethoxypoly(ethylene glycol)s bearing one hydrophobic naphthyl end group (Nap-MPEG) on beta-cyclodextrin polymer (poly-beta-CD) immobilized on a silica support, under isocratic elution conditions and using water as mobile phase. Studies of retentions and theoretical plate heights H were conducted at infinite dilution by comparing the behavior of Nap-MPEGs having different molecular weight (750, 1000 and 5000 g/mol). The larger is its molecular size, the lower is the retention of the polymer. The linear increase of H with mobile phase velocity reveals slow mass-transfer kinetics arising from the restricted diffusion into the pores of the support. The complexation constants between the Nap-MPEGs and beta-CD in solution (around 500M(-1)) were determined from the decrease of retention observed by adding increasing concentrations of hydroxypropyl beta-CD into the eluent. The peak profiles in mass-overload conditions were studied by fitting a model based upon bi-Langmuir kinetics which assumes a non-uniform support having two types of binding sites and apparent adsorption rate constants are used to describe mass-transfer kinetics. A three-parameter adsorption equilibrium isotherm was sufficient to account for the modifications of peak shapes observed when increasing amounts of polymer were injected. This result indicates an interaction with a heterogeneous poly-beta-CD support mainly composed of low affinity groups, non-saturable in the range of polymer concentration studied. An upper limit was estimated for the equilibrium constant (<1000 M(-1)) characterizing the affinity of Nap-MPEG for the non-saturable sites of the poly-beta-CD support. Large affinity constants (8-9 x 10(4)M(-1)) were found for the interaction of Nap-MPEGs with a small percentage of active sites.

Determination of binding constants of hydrophobically end-capped poly(ethylene glycol)s with beta-cyclodextrin by affinity capillary electrophoresis.

The formation of inclusion complexes between methoxypoly(ethylene glycol)s (MPEG)s bearing one hydrophobic group (phenyladamantyl) per chain and beta-cyclodextrin (beta-CD) was studied by capillary electrophoresis (CE). The effect of highly sulphated beta-CD (HS-beta-CD) on the migration behaviour of the phenyladamantyl-modified MPEG (MPEG-PhAd) analyte was investigated. It was established that the interaction between the modified PEG and beta-CD involved a 1:1 stoichiometry. Non-linear regression and three usual linearization methods (y-reciprocal, x-reciprocal and double reciprocal) were employed to estimate the binding constants. It was demonstrated that the binding constants were similar (around 400 M(-1)) for two MPEG-PhAd having different chain lengths (2000 and 5000 g/mol).

Immobilization of a (dextran-adamantane-COOH) polymer onto beta-cyclodextrin-modified silica.

Adamantane-modified compounds are known to form stable complexes with beta-cyclodextrins (beta-CD) by host-guest interactions. In this study, the inclusion complex formed between beta-CD cavities and the adamantane group was evaluated for the elaboration of a cation-exchange support. The synthesis of the chromatographic supports involved three steps: (i) a polymer of beta-CD was grafted to diol-modified silica, (ii) a dextran polymer was modified by both adamantane groups and ionizable COOH functions, (iii) the dextran derivative (Ad-Dex-COOH) was bound to the chromatographic support by complexation between the adamantane groups of the dextran and beta-CD cavities of the support. The polymer immobilization on the beta-CD support was successful as the resulting support exhibited weak cation-exchange properties. The stationary phase was easy to prepare under mild conditions (aqueous media, room temperature) and was quite stable when using aqueous mobile phases. The chromatographic behaviour of model proteins was studied in isocratic elution by examining the effect of salt concentration in the buffer on retention. A mixed retention mode was found for lysozyme, revealing both electrostatic and hydrophobic interactions with the stationary phase.

Functionalization of macroporous organic polymer monolith based on succinimide ester reactivity for chiral capillary chromatography: a cyclodextrin click approach.

Macroporous cross-linked organic polymer based on N-acryloxysuccinimide (NAS) and ethylene dimethacrylate (EDMA) was prepared inside 75 µm id fused silica capillary as a functionalizable monolithic stationary phase for chromatographic applications. Succinimide groups on the monolith surface provide reactive sites able to react readily through standard electrophile-nucleophile chemistry. Propargylamine was used to prepare alkyne functionalized poly(NAS-co-EDMA). Onto this azido-reactive polymer surface was grafted ß-cyclodextrin (CD) via a triazole ring utilizing the copper(I)-catalyzed 1,3-dipolar cyclo-addition reaction. Chemical characterization was performed in situ after each synthetic step by means of Raman spectroscopy. Good enantioseparations of flavanone enantiomers, chosen as test chiral compound, were achieved under reversed phase conditions by both capillary electrochromatography and nano-liquid chromatography (nano-LC) techniques. These results demonstrate the potentiality and usefulness of click chemistry in the preparation of ß-CD containing chiral organic polymer monolith.

CEC separation of aromatic compounds and proteins on hexylamine-functionalized N-acryloxysuccinimide monoliths.

Reactive organic polymer monoliths were prepared in fused-silica capillaries by UV-initiated free radical polymerization of N-acryloxysuccinimide (NAS) as reactive monomer, ethylene dimethacrylate as crosslinker, azobisisobutyronitrile as initiator, and toluene as porogen. In a second synthetic step, chemical derivatization of the activated-ester moieties was performed in situ through alkylation reaction with alkylamines to afford monolithic stationary phases with potential reversed-phase properties. A correlation between the synthesis conditions--composition of the reactive solution--chemical characteristics of the reactive polymer monoliths--nitrogen/NAS content--and the reversed-phase separation properties of the functionalized monolithic columns--selectivity towards homologous series of akylbenzenes--was clearly established. This finding offers the possibility of adjusting the experimental conditions with respect to the target applications. The monolithic stationary phases with optimized chemical and porous structures were used for the CEC separation of alkylbenzenes, phenols, anilines, organic acids, amino acids, and proteins. The data indicate that depending on the nature of the analytes (charge, hydrophobic/hydrophilic balance, size) reversed-phase or mixed modes may account for the observed separation.