Exploration of polyamide structure-property relationships by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

RATIONALE: Polyamides (PA) are among the most used classes of polymers because of their attractive properties. Depending on the nature and proportion of the co-monomers used for their synthesis, they can exhibit a very large range of melting temperatures (Tm ). This study aims at the correlation of data from mass spectrometry (MS) with differential scanning calorimetry (DSC) and X-ray diffraction analyses to relate molecular structure to physical properties such as melting temperature, enthalpy change and crystallinity rate. METHODS: Six different PA copolymers with molecular weights around 3500 g mol(-1) were synthesized with varying proportions of different co-monomers (amino-acid AB/di-amine AA/di-acid BB). Their melting temperature, enthalpy change and crystallinity rate were measured by DSC and X-ray diffraction. Their structural characterization was carried out by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Because of the poor solubility of PA, a solvent-free sample preparation strategy was used with 2,5-dihydroxybenzoic acid (2,5-DHB) as the matrix and sodium iodide as the cationizing agent. RESULTS: The different proportions of the repeating unit types led to the formation of PA with melting temperatures ranging from 115°C to 185°C. The structural characterization of these samples by MALDI-TOF-MS revealed a collection of different ion distributions with different sequences of repeating units (AA, BB; AB/AA, BB and AB) in different proportions according to the mixture of monomers used in the synthesis. The relative intensities of these ion distributions were related to sample complexity and structure. They were correlated to DSC and X-ray results, to explain the observed physical properties. CONCLUSIONS: The structural information obtained by MALDI-TOF-MS provided a better understanding of the variation of the PA melting temperature and established a structure-properties relationship. This work will allow future PA designs to be monitored.

Complete meiosis in rat prepubertal testicular tissue under in vitro sequential culture conditions.

BACKGROUND: Testicular tissue cryopreservation before gonadotoxic treatments allows fertility preservation in children suffering from cancer. Fertility restoration strategies, in particular in vitro maturation of prepubertal testicular tissue, are being developed mainly in animal models. The rat, widely used in biomedical research, including in reproductive biology, is a relevant model. OBJECTIVES: To determine whether sequential two-step culture protocols can improve the efficiency of rat in vitro spermatogenesis. MATERIALS AND METHODS: Rat prepubertal testicular tissues were cultured on agarose gels with either a one-step or two-step protocol with or without polydimethylsiloxane (PDMS) ceiling chips. The progression of spermatogenesis, germ/Sertoli cell ratio, cell proliferation, seminiferous tubule area, and intratubular cell density were assessed by histological and immunohistochemical analyses. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assays and Peanut Agglutinin (PNA) lectin labeling were performed to analyze the Deoxyribonucleic Acid (DNA) integrity and differentiation step of in vitro-produced spermatids. RESULTS: Sequential two-step protocols allowed the production of spermatids with a higher efficiency compared with the one-step culture protocol. However, the efficiency was low, as less than 1.5% of tubules contained spermatids. Most of the in vitro-produced spermatids contained unfragmented DNA and were at an early step of differentiation. Rare elongating spermatids could be detected in the cultured explants. Although complete in vitro spermatogenesis could not be obtained with PDMS ceiling chips, entry into meiosis was promoted in one-step organotypic cultures. DISCUSSION AND CONCLUSION: Complete in vitro meiosis and the beginning of the elongation phase of spermiogenesis were obtained in a rat model using sequential culture methods. Because of their low efficiency, further work will be necessary to identify the culture conditions allowing the completion of spermiogenesis. These optimizations could pave the way for future applications, including the development of an in vitro fertility restoration procedure for childhood cancer survivors, which is still far from being clinically available.

Solvent-based and solvent-free characterization of low solubility and low molecular weight polyamides by mass spectrometry: a complementary approach.

RATIONALE: Polyamides (PA) belong to the most used classes of polymers because of their attractive chemical and mechanical properties. In order to monitor original PA design, it is essential to develop analytical methods for the characterization of these compounds that are mostly insoluble in usual solvents. METHODS: A low molecular weight polyamide (PA11), synthesized with a chain limiter, has been used as a model compound and characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). In the solvent-based approach, specific solvents for PA, i.e. trifluoroacetic acid (TFA) and hexafluoroisopropanol (HFIP), were tested. Solvent-based sample preparation methods, dried-droplet and thin layer, were optimized through the choice of matrix and salt. Solvent-based (thin layer) and solvent-free methods were then compared for this low solubility polymer. Ultra-high-performance liquid chromatography/electrospray ionization (UHPLC/ESI)-TOF-MS analyses were then used to confirm elemental compositions through accurate mass measurement. RESULTS: Sodium iodide (NaI) and 2,5-dihydroxybenzoic acid (2,5-DHB) are, respectively, the best cationizing agent and matrix. The dried-droplet sample preparation method led to inhomogeneous deposits, but the thin-layer method could overcome this problem. Moreover, the solvent-free approach was the easiest and safest sample preparation method giving equivalent results to solvent-based methods. Linear as well as cyclic oligomers were observed. Although the PA molecular weights obtained by MALDI-TOF-MS were lower than those obtained by (1)H NMR and acido-basic titration, this technique allowed us to determine the presence of cyclic and linear species, not differentiated by the other techniques. TFA was shown to induce modification of linear oligomers that permitted cyclic and linear oligomers to be clearly highlighted in spectra. CONCLUSIONS: Optimal sample preparation conditions were determined for the MALDI-TOF-MS analysis of PA11, a model of polyamide analogues. The advantages of the solvent-free and solvent-based approaches were shown. Molecular weight determination using MALDI was discussed.

Use of telechelic cis-1,4-polyisoprene cationomers in the synthesis of antibacterial ionic polyurethanes and copolyurethanes bearing ammonium groups.

New crosslinked ionic polyurethanes and copolyurethanes were yielded by reaction of telechelic cis-1,4-oligoisoprenes, bearing a variable number of ammonium and hydroxy groups, with isocyanurate of isophorone diisocyanate (I-IPDI). Aiming for a comparative study, polyurethane elastomers based on non-ionic telechelic oligomers were also synthesized. Thermo-mechanical behavior and crosslinking density of these three families of materials were investigated by DMTA and swelling test, respectively. Surface properties were examined by static contact angle measurements and AFM imaging. The bactericidal activity of the polymers was investigated by enumerating living Pseudomonas aeruginosa on material surfaces and on water suspensions. The number of attached living bacteria was found to depend on the chemical structure of the material and on the contact time between the microorganisms and the surface. An exclusive bactericidal activity was obtained with the ionic copolyurethane family. Materials with weak crosslinking density were found to release bactericidal moieties. The abilities of the polymers to prevent bacterial growth were examined through zone of inhibition experiments against P. aeruginosa, which shown a bacteriostatical effect for each synthesized material. These experiments were not sufficiently sensitive to detect the leaching of bactericidal moieties from the materials with weak crosslinking density. When the zone of inhibition experiments was performed on more sensitive bacteria, namely Staphylococcus epidermidis, the leaching of bactericidal moieties as well as bacteriostatic effect was detected. This work demonstrates the potentiality for making functional biomaterials from natural rubber, a renewable resource.

Tandem mass spectrometry of low solubility polyamides.

The structural characterization of polyamides (PA) was achieved by tandem mass spectrometry (MS/MS) with a laser induced dissociation (LID) strategy. Because of interferences for precursor ions selection, two chemical modifications of the polymer end groups were proposed as derivatization strategies. The first approach, based on the addition of a trifluoroacetic acid (TFA) molecule, yields principally to complementary bn and yn product ions. This fragmentation types, analogous to those obtained with peptides or other PA, give only poor characterization of polymer end-groups [1]. A second approach, based on the addition of a basic diethylamine (DEA), permits to fix the charge and favorably direct the fragmentation. In this case, bn ions were not observed. The full characterization of ω end group structure was obtained, in addition to the expected yn and consecutive fragment ions.

Click grafting of seaweed polysaccharides onto PVC surfaces using an ionic liquid as solvent and catalyst.

Seaweed antibacterial polysaccharides were grafted onto poly(vinylchloride) (PVC) surfaces using an original click chemistry pathway. PVC isothiocyanate surfaces (PVC-NCS) were first prepared by nucleophilic substitution of the chloride groups by isothiocyanate groups in DMSO/water medium. Then, unmodified Ulvan, Fucan, Laminarin or Zosterin was directly grafted onto the PVC-NCS surface using 1-ethyl-3-methyl imidazolium phosphate, an ionic liquid, as solvent and catalyst. To attest the grafting effectiveness, the new PVC surfaces were well characterized by AFM, XPS and contact angle measurements.