Engineering 3D-Printed Bioresorbable Scaffold to Improve Non-Vascularized Fat Grafting: A Proof-of-Concept Study.

Autologous fat grafting is the gold standard for treatment in patients with soft-tissue defects. However, the technique has a major limitation of unpredictable fat resorption due to insufficient blood supply in the initial phase after transplantation. To overcome this problem, we investigated the capability of a medical-grade poly L-lactide-co-poly ε-caprolactone (PLCL) scaffold to support adipose tissue and vascular regeneration. Deploying FDM 3D-printing, we produced a bioresorbable porous scaffold with interconnected pore networks to facilitate nutrient and oxygen diffusion. The compressive modulus of printed scaffold mimicked the mechanical properties of native adipose tissue. In vitro assays demonstrated that PLCL scaffolds or their degradation products supported differentiation of preadipocytes into viable mature adipocytes under appropriate induction. Interestingly, the chorioallantoic membrane assay revealed vascular invasion inside the porous scaffold, which represented a guiding structure for ingrowing blood vessels. Then, lipoaspirate-seeded scaffolds were transplanted subcutaneously into the dorsal region of immunocompetent rats (n = 16) for 1 or 2 months. The volume of adipose tissue was maintained inside the scaffold over time. Histomorphometric evaluation discovered small- and normal-sized perilipin+ adipocytes (no hypertrophy) classically organized into lobular structures inside the scaffold. Adipose tissue was surrounded by discrete layers of fibrous connective tissue associated with CD68+ macrophage patches around the scaffold filaments. Adipocyte viability, assessed via TUNEL staining, was sustained by the presence of a high number of CD31-positive vessels inside the scaffold, confirming the CAM results. Overall, our study provides proof that 3D-printed PLCL scaffolds can be used to improve fat graft volume preservation and vascularization, paving the way for new therapeutic options for soft-tissue defects.

Metal-Free ATRP Catalyzed by Visible Light in Continuous Flow.

ATRP of methyl methacrylate catalyzed by Eosin Y, an inexpensive and an environmental benign dye, was performed in a continuous flow reactor made of FEP tubing and irradiated by visible light green LEDs. The reaction under flow conditions was significantly more rapid and controlled compared to that in batch giving 90% of polymerization after only 3 h of irradiation. The formed polymers in flow have M n measured by GPC and DOSY NMR in accordance with the theoretical values and show low dispersities (Ð < 1.5). The livingness of the polymers has been confirmed by LED on and LED off experiments and by the synthesis of block copolymers. The protocol described herein serves as a "proof of concept" of using Eosin Y as a photocatalyst for controlled polymerization and of using 1D and 2D NMR for polymer characterization. The protocol could be replicated in the future for other reversible-deactivation radical polymerizations.

Rationale for the design of 3D-printable bioresorbable tissue-engineering chambers to promote the growth of adipose tissue.

Tissue engineering chambers (TECs) bring great hope in regenerative medicine as they allow the growth of adipose tissue for soft tissue reconstruction. To date, a wide range of TEC prototypes are available with different conceptions and volumes. Here, we addressed the influence of TEC design on fat flap growth in vivo as well as the possibility of using bioresorbable polymers for optimum TEC conception. In rats, adipose tissue growth is quicker under perforated TEC printed in polylactic acid than non-perforated ones (growth difference 3 to 5 times greater within 90 days). Histological analysis reveals the presence of viable adipocytes under a moderate (less than 15% of the flap volume) fibrous capsule infiltrated with CD68+ inflammatory cells. CD31-positive vascular cells are more abundant at the peripheral zone than in the central part of the fat flap. Cells in the TEC exhibit a specific metabolic profile of functional adipocytes identified by 1H-NMR. Regardless of the percentage of TEC porosity, the presence of a flat base allowed the growth of a larger fat volume (p < 0.05) as evidenced by MRI images. In pigs, bioresorbable TEC in poly[1,4-dioxane-2,5-dione] (polyglycolic acid) PURASORB PGS allows fat flap growth up to 75 000 mm3 at day 90, (corresponding to more than a 140% volume increase) while at the same time the TEC is largely resorbed. No systemic inflammatory response was observed. Histologically, the expansion of adipose tissue resulted mainly from an increase in the number of adipocytes rather than cell hypertrophy. Adipose tissue is surrounded by perfused blood vessels and encased in a thin fibrous connective tissue containing patches of CD163+ inflammatory cells. Our large preclinical evaluation defined the appropriate design for 3D-printable bioresorbable TECs and thus opens perspectives for further clinical applications.

Plant therapy in the Peruvian Amazon (Loreto) in case of infectious diseases and its antimicrobial evaluation.

ETHNOPHARMACOLOGICAL RELEVANCE: The plant species reported here are used in contemporary phytotherapies by native and neo-urban societies from the Iquitenian surroundings (district of Loreto, Peruvian Amazon) for ailments related to microbial infections. Inhabitants of various ethnic origins were interviewed and 81 selected extracts were evaluated for their antimicrobial properties against a panel of 36 sensitive and multi-resistant bacteria or yeast. Medicinal plant researches in the Peruvian Amazon are now significant, but none of them has focused on an exhaustive listing of identified species tested on so many microbes with standardized experiments (to obtain MIC value). AIM OF THE STUDY: The aim of the study was to inventory the plants used against infections in the Loreto, an Amazonian region of Peru. It led to the new identification of secondary metabolites in two plant species. MATERIALS AND METHODS: Ethnographic survey was carried out using "participant-observation" methodology and focus on bioprospecting of antimicrobial remedies. Selected plant extracts and antimicrobial drugs were tested in vitro with agar dilution method on 35 bacteria strains and 1 yeast to evaluate their Minimal Inhibitory Concentration (MIC). Microdilution methods using 96-well microtiter plates were used for the determination of MIC from isolated compounds, and cytotoxicity in HepG2 cells from some selected extracts were also evaluated. Activity-guided isolation and identification of compounds were performed by various chromatographic methods and structural elucidations were established using HRMS and NMR spectroscopy. RESULTS: This study outlined antimicrobial activities of 59 plant species from 33 families (72 single plant extracts and 2 fermented preparations), 7 mixtures, and one insect nest extract against 36 microorganisms. Of the 59 species analysed, 12 plants showed relevant antibacterial activity with MIC ≤0.15 mg/mL for one or several of the 36 micro-organisms (Aspidosperma excelsum, Brosimum acutifolium, Copaifera paupera, Erythrina amazonica, Hura crepitans, Myrciaria dubia, Ocotea aciphylla, Persea americana, Spondias mombin, Swartzia polyphylla, Virola pavonis, Vismia macrophylla). Examination by bioautography of E. amazonica, M. dubia and O. aciphylla extracts allowed the phytochemical characterization of antimicrobial fractions and compounds. CONCLUSION: This study suggested an a posteriori correlation of the plant extract antimicrobial activity with the chemosensory cues of the drugs and attested that those chemosensory cues may be correlated with the presence of antimicrobial compounds (alkaloids, tannins, saponosids, essential oil, oleoresin …). It also led to the first isolation and identification of three secondary metabolites from E. amazonica and M. dubia.

One- or Two-Step Synthesis of C-8 and N-9 Substituted Purines.

A novel and original strategy to obtain rapidly a large diversity of C-8 and N-9 substituted purines was developed. The present procedure describes annulation reactions in one or two steps starting from 5-aminoimidazole-4-carbonitriles 1-8 in moderate to good yields. 8,9-Disubstituted-6,9-dihydro-1H-purin-6-ones 9-14, 6-amino-8,9-disubstituted-3,9-dihydro-2H-purin-2-ones 15-20, 8,9-disubstituted-3,9-dihydro-2H-purin-2,6-diamines 21-24 and 6-imino-1-phenyl-8,9-disubstituted-6,9-dihydro-1H-purin-2-(3H)-ones 25-26 were synthesized in one step using formic acid, urea, guanidine carbonate, and phenylisocyanate, respectively, whereas 8,9-disubstituted-9H-purin-6-amines 27-31 and 6-imino-8,9-disubstituted-6,9-dihydro-1H-purin-1-amines 32-33 were obtained in two steps using formamide and hydrazine, respectively.

Identification and quantification by 1H nuclear magnetic resonance spectroscopy of seven plasticizers in PVC medical devices.

Medical devices are generally made of polyvinyl chloride plasticized by six authorized plasticizers as alternatives to di-(2-ethylhexyl)-phthalate (DEHP) classified as reprotoxic class 1b. These are acetyl tri-n-butyl citrate (ATBC), di-(2-ethylhexy) adipate (DEHA), di-(2-ethylhexyl) terephthalate (DEHT), di-isononyl cyclohexane-1,2-dicarboxylate (DINCH), di-isononyl phthalate (DINP), and tri-octyl trimellitate (TOTM). The main objective of this study was to propose a new method using 1H NMR spectroscopy to determine and quantify these seven plasticizers in PVC sheets, standard infusion tubings, and commercially available medical devices. Two techniques were compared: dissolution in deuterated tetrahydrofuran and extraction by deuterated chloroform. Plasticizer 1H NMR spectra were very similar in both deuterated solvents; dissolution and extraction provided similar results. The sensitivity of this method enabled us to detect and quantify the presence of minor plasticizers in PVC. In nine commercially available samples, the major plasticizer was identified and quantified by 1H NMR. In six samples, one, two, or three minor plasticizers were identified and also quantified. DEHP was detected in only one tubing. NMR is therefore very convenient for studying plasticizers contained in medical devices. Only small quantities of solvents and sample are required. It is not necessary to dilute samples to enter a quantification range, and it is sufficiently sensitive to detect contaminants.

Evaluation of three neutral capillary coatings for the determination of analyte-cyclodextrin binding constants by affinity capillary electrophoresis. Application to N,N'-disubstituted piperazine derivatives.

The performances of three neutral static coatings (hydroxypropyl cellulose, polyethylene oxide and poly(N,N-dimethylacrylamide) have been evaluated in order to determine the binding constants of the complexes formed between four polycationic compounds (piperazine derivatives) and four cyclodextrins of pharmaceutical interest (ß-CD, HP-ß-CD, Me-ß-CD and sulfobutyl ether-ß-CD) by affinity capillary electrophoresis. The physically-adsorbed poly(N,N-dimethylacrylamide) coating proves to be the more efficient to mask the silanol groups of the capillary wall since the lowest electroosmotic flow was measured for this coating. Moreover, it drastically reduces the adsorption of the compounds since it allows a correct repeatability of their migration time, higher efficiencies of the peaks and no baseline shift. Then, it was verified for four complexes that this coating allows a correct determination of the binding constants avoiding the CD adsorption which is responsible of an undervaluation of binding constants. The highest binding constants are obtained using the anionic sulfobutyl ether-ß-CD (SBE-ß-CD). The structure of the complex formed between the tacrine derivative and the SBE-ß-CD was further investigated through 2D ROESY NMR experiments and structure-binding constant relationships. Results suggest that the inclusion in the SBE-ß-CD cavity occurs through the aliphatic ring portion of the tacrine moiety.

NMR investigation of the complexation and chiral discrimination of pyrazole sulfonamide derivatives with cyclodextrins.

The complexes formed between six original chiral diaryl-pyrazole sulfonamide derivatives, displaying poor solubility, and various CDs (native α-, ß- and γ-CDs, hydroxypropylated HP-ß-CD, methylated Me-ß-CD or amino NH2-ß-CD) were studied by 1D and 2D (1)H NMR at physiological pH in order to determine their apparent binding constant, stoichiometry and structure of the supramolecular assembly. For some complexes, the spectra obtained for free racemic compound and for racemic compound in presence of CD indicate a splitting of signal(s). Additional experiments with pure enantiomer and enriched enantiomer allow us to attribute this behavior to chiral discrimination. The complexing ability of the native ß-CD towards our compounds appears the most promising since binding values around 7×10(2)M(-1) are obtained. The two-dimensional ROESY ((1)H-(1)H) experiments prove the inclusion of the aliphatic part of the compound in the CD cavity. It is noteworthy that this inclusion occurs via the smaller opening of the cavity.

Comparative study of the complex forming ability and enantioselectivity of cyclodextrin polymers by CE and 1H NMR.

The interactions between nine drugs (baclofen, bupivacaine, chlorpheniramine, ketoconazole, paliperidone, promethazine, propranolol, risperidone and verapamil) and six cyclodextrins (α-CD, ß-CD, γ-CD, HP-ß-CD, HP-γ-CD and Me-ß-CD) or six polymers of cyclodextrins (polyα-CD, polyß-CD, polyγ-CD, polyHP-ß-CD, polyHP-γ-CD and polyMe-ß-CD) were studied by affinity capillary electrophoresis and/or (1)H NMR at pH 2.5. An exhaustive qualitative study was performed through the determination of the retardation factor. Then, four compounds and both ß-CD and polyß-CD were selected for the quantitative study of the interactions at pH 2.5 and 7.0. By comparing the results obtained with the ß-CD and polyß-CD, it appears that the apparent binding constants are up to five times higher with the polymer. The 2D-NMR results seem to indicate that the structure of the polymeric network favours the inclusion of the guest in the hydrophobic cavity of the CD units. Moreover, the poly-CDs have shown very high enantioselective abilities at both pH.

Complexation of triptolide and its succinate derivative with cyclodextrins: affinity capillary electrophoresis, isothermal titration calorimetry and 1H NMR studies.

The complexation of the triptolide PG490 and its succinate derivative PG490-88Na with various cyclodextrins was studied using three complementary techniques: affinity capillary electrophoresis (ACE), isothermal titration calorimetry (ITC) and nuclear magnetic resonance (NMR). The apparent binding constants of the complexes formed between the drugs and 8 CDs (α-CD, ß-CD, γ-CD, HP-α-CD, HP-ß-CD, HP-γ-CD, CM-ß-CD and amino-ß-CD) were determined by ACE through linear Scott's plots. The apparent and averaged binding constants of the complexes formed between PG490-88 and ß-CD, γ-CD, HP-α-CD, HP-ß-CD or HP-γ-CD are contained in the narrow range 135-167 M(-1). For the anionic CM-ß-CD and cationic amino-ß-CD, these constants are 38 and 278 M(-1), respectively, which is in accordance with electrostatic repulsions or attractions with the succinate moiety. ITC and NMR investigations for the binding constants determinations were performed for 2 CDs allowing high complexation: HP-ß-CD and amino-ß-CD. The three techniques provided similar results. ITC and NMR, in contrast to ACE, allowed to study the complexes formed between the neutral compound PG490 and neutral cyclodextrins. A more advanced characterization of the PG 490-88Na/amino-ß-CD complex, which displays the highest apparent binding constant, was undertaken using NMR spectroscopy. The 1:1 stoichiometry of the complex was established by (1)H NMR 1D and selective 1D TOCSY experiments using the continuous variation method. Moreover, the 1D and 2D ROESY experiments revealed the inclusion of the isopropyl moiety of the triptolide derivative in the hydrophobic CD cavity. Altogether, the data provide strong evidences that the two triptolide compounds can be efficiently complexed with CD.

Experimental study on infusion devices containing polyvinyl chloride: to what extent are they di(2-ethylhexyl)phthalate-free?

The use of medical devices containing highly criticized phthalates including di(2-ethylhexyl) phthalate (DEHP) has been challenged by European directive 2007/47/CE, put into effect in March 2010. New plasticizers are now being used to soften PVC in medical devices: trioctyltrimellitate (TOTM), di-isononyl-cyclohexan-1,2-dicarboxilate (DINCH) and di(2-ethylhexyl) terephthalate (DEHT). To quantify DEHP in nine DEHP-free medical devices made of PVC softened by alternative plasticizers, high performance liquid chromatography analysis with ultraviolet detection at 220 nm wavelength was achieved. An NMR spectroscopy was performed to confirm DEHP presence. Only two medical devices out of the nine tested were truly without DEHP. One of them showed traces of DEHP exceeding the threshold contamination of 0.1% in plastic mass set by REACH regulations. TOTM plasticizer is still incriminated when polyvinyl-chloride (PVC) is contaminated with DEHP. Manufacturers must verify the purity of their raw material, not only on PVC, but also on other soft plastics entering into the composition of medical infusion devices. The clinical consequences of exposure to certain levels of DEHP have not been evaluated. A solution could be to use alternative PVC-free materials.

Unusual inversion phenomenon of ß-cyclodextrin dimers in water.

A conformational analysis of three triazole-containing bridged bis-ß- cyclodextrins (CD) has been carried out to evaluate their recognition ability. NMR spectroscopy and ITC measurements clearly demonstrate that one of the CD glucopyranose units undergoes a 360° rotation in water so that the spacer linking the two CDs is deeply included into one of the CD cavities. The amplitude of this inversion phenomenon depends on the nature of the spacer and results in a limited accessibility to the CD cavities in line with previous catalytic results.

New phosphane based on a beta-cyclodextrin, exhibiting a solvent-tunable conformation, and its catalytic properties.

A new diphenylphosphane based on a beta-cyclodextrin skeleton that exhibits a dual solubility in water and in organic solvent was synthesised. Interestingly, a solvent-dependent conformation change was evidenced by NMR spectroscopy studies; the self-inclusion of a phenyl group of the phosphane moiety into cyclodextrin cavity observed in water disappeared in organic solvents due to a change in conformation. Hydrogenation or hydroformylation reactions performed in water and in organic solvents showed that this ligand was able to stabilise catalytically active rhodium species in solution. In the case of the hydroformylation reaction, it was demonstrated that regioselectivity was influenced by the solvent-dependent conformation of the ligand.

Molecular interactions between zoledronic acid and bone: An in vitro Raman microspectroscopic study.

The aim of this study was to investigate molecular interactions between a bisphosphonate (BP), zoledronic acid, and bone tissue by the use of Raman microspectroscopy. In this way, samples of hydroxyapatite (HA), as a bone model, and Wistar rat femurs were soaking in zoledronic acid solutions. Sample surfaces were studied by Environmental Scanning Electron Microscopy and Raman spectroscopy. The amount of zoledronic acid incorporated onto the samples and the inorganic phosphate released in solution were determined by (31)P NMR spectroscopy. Total carbonate content in solution was evaluated by inorganic carbon analyser. After impregnation new Raman bands with frequencies close to characteristic peaks of zoledronic acid (in particular phosphate moieties and imidazole ring of the R2 side-chain) were observed on both types of samples. Physico-chemical parameters of the bone were also significantly modified (P<0.0001). The mineral to organic ratio and the carbonate to phosphate ratio decreased and the crystallinity increased. Released inorganic phosphate and carbonate were detected in the solutions. The Raman shift of the bands corresponding to the phosphate groups and the imidazole ring of the BP highlight their implication in the binding to the mineral. The detection of released inorganic phosphate and carbonate in solution, the modifications of the mineral to phosphate ratio and the carbonate to phosphate ratio reveal that BP decrease the amount of inorganic phosphate and limit the dissolution of bone mineral. The increase of the crystallinity after BP binding shows a re-organisation of the lattice with a higher symmetry. Thus, it seems that zoledronic acid has an important contribution on the increase of crystallinity. The use of Raman spectrometry brings new and complementary information on the impact of zoledronic acid on bone composition at molecular level. Raman spectrometry could help to understand by which way BPs improve bone strength and decrease fracture risk.

Stepwise aggregation of dimethyl-di-n-octylammonium chloride in aqueous solutions: from dimers to vesicles.

The self-aggregation of dimethyl-di-n-octylammonium chloride, in diluted aqueous solutions, was studied with various experimental and theoretical techniques: zetametry, conductimetry, dimethyl-di-n-octylammonium and chloride-selective electrodes, tensiometry, NMR spectroscopy ((1)H and DOSY), and molecular modeling (PM3 and molecular dynamic). The combination of the data obtained by these techniques led us to propose a stepwise aggregation process with increasing concentration: dimers (0.2-10 mM), bilayers (10-30 mM), and finally vesicles (>30 mM).

Self-association of short-chain nonionic amphiphiles in binary and ternary systems: comparison between the cleavable ethylene glycol monobutyrate and its ether counterparts.

In the context of environmental concerns for the production of surface active species, the introduction of a carbonyl function into the skeleton of ethyleneglycol-derived solvo-surfactants is a way to access cleavable compounds with presumed enhanced biodegradability. Ethylene glycol monobutyrate (C(3)COE(1)) was synthesized and compared to its ether counterparts, ethylene glycol monopropyl (C(3)E(1)) and monobutyl ethers (C(4)E(1)), to assess the effect of the insertion of a carbonyl function in the skeleton of short-chain ethoxylated amphiphilic compounds. In aqueous solutions, the ester has intermediate behavior between that of the two ethers with regard to surface tension, solubilization of Me-naphtalene in water, and self-diffusion by PGSE NMR. In ternary systems, C(3)COE(1) and C(3)E(1) have the same optimal oil (EACN = 2.8), which is much more polar than that of C(4)E(1) (EACN = 8.5). With regard to the ability to form structured systems, the behavior in water does not differ significantly for the three compounds, and the transition between nonassociating solvents and amphiphilic solvents, sometimes called solvo-surfactants, is gradual. In ternary systems, however, only C(4)E(1) and C(3)COE(1) form a third phase near the optimal formulation, which tends to show that C(3)COE(1) possesses the minimum amphiphilicity to get a structuration. Self-diffusion NMR studies of the one-phase domains do not, however, allow us to distinguish between different degrees of organization in the three systems.

Characterization and in vitro evaluation of the formoterol/cyclodextrin complex for pulmonary administration by nebulization.

The aim of this work is to investigate the effect of cyclodextrin complexation on the pulmonary deposition of formoterol, a drug with a very poor aqueous solubility, after jet nebulization. Two types of cyclodextrins, a hydroxypropyl beta cyclodextrin (Kleptose HP) and a polydispersed methyl beta cyclodextrin (Crysmeb) were used. The interactions of formoterol with the cyclodextrins were studied by NMR. The aqueous cyclodextrin solutions containing formoterol were defined by their physicochemical properties in relation to nebulization capacity: density, surface tension and viscosity. Nebulization efficiency was evaluated by measuring droplet size, nebulization rate, quantity nebulized and nebulization time. The NMR ROESY spectra suggest that formoterol or a part of it is included inside the cyclodextrins. Densities and viscosities of the solutions tested are close to those of water; the lower surface tensions compared to water (53.7 and 56.7 vs 70 mN/m) favour the formation of small droplets. The aqueous solutions of cyclodextrins and formoterol studied can generate aerosols with a particle size that is compatible with pulmonary deposition. Respirable fraction values between 57.5% and 88 % were obtained when nebulizing the solutions with four nebulizers that differ geometrically. Nebulization rates varied from 0.19 to 0.47 g/min. Large quantities of drug nebulized over acceptable delivery times were observed. beta-cyclodextrin derivatives can be used to formulate nebulizable solutions of formoterol. It is indispensable to define the appropriate nebulizers and operating conditions associated with the solutions to obtain adapted and reproducible activity.

Study of the complexation of risperidone and 9-hydroxyrisperidone with cyclodextrin hosts using affinity capillary electrophoresis and (1)H NMR spectroscopy.

The complexation of risperidone (Risp) and 9-hydroxyrisperidone (9-OH-Risp), atypical antipsychotics, with seven cyclodextrins (CDs) of pharmaceutical interest (native and hydroxypropylated (HP) alpha-, beta-, gamma-CDs and methyl (Me)-beta-CD) was studied by affinity capillary electrophoresis (ACE) and nuclear magnetic resonance spectroscopy (NMR) for acidic pH 2.5 and physiological pH 7.4. The 1:1 stoichiometry of the complexes was established by (1)H NMR spectroscopy using the continuous variation method developed by Job. The apparent binding constants of the 14 complexes at both pH were determined by ACE through the linear Scott's plots. The NMR spectroscopy investigation of the binding constants was achieved for the two CDs allowing the highest complexation: the beta-CD and Me-beta-CD. Both ACE and NMR spectroscopy studies provide similar conclusions by considering the influence of the 9-hydroxylation, the influence of the CD substitution and the influence of the pH. Moreover, the NMR spectroscopy results have allowed to suppose a pH-dependent inclusion mechanism. A thermodynamic study was then performed by ACE at both pH for the Risp.Me-beta-CD and 9-OH-Risp.Me-beta-CD complexes: the opposite signs of the entropic change (DeltaS degrees <0 at pH 2.5 and DeltaS degrees >0 at pH 7.4) confirms the influence of the pH on the complexation mechanism and the possible difference in the depth of the analyte inclusion in the hydrophobic cavity of the CD. Last, the two-dimensional ROESY (rotating-frame Overhauser spectroscopy) ((1)H-(1)H) and HOESY (heteronuclear Overhauser effect spectroscopy) ((19)F-(1)H) experiments have proved the inclusion of the aromatic part of the Risp and 9-OH-Risp in the hydrophobic CD cavity and lead us to propose a model of complexation.

Quantitative determination of glyphosate in human serum by 1H NMR spectroscopy.

The determination and quantification of glyphosate in serum using (1)H NMR spectroscopy is reported. This method permitted serum samples to be analysed without derivatization or any other sample pre-treatment, using 3-trimethylsilyl 2,2',3,3'-tetradeuteropropionic acid (TSP-d(4)) as a qualitative and quantitative standard. Characterization of the herbicide N-(phosphonomethyl)glycine was performed by analysing chemical shifts and coupling constant patterns. Quantification was performed by relative integration of CH(2)-P protons to the TSP-d(4) resonance peak. The method was tested for repeatability (n=5) and yielded coefficients of variation of 1% and 3%, respectively: detection and quantification limits were also determined and were 0.03 and 0.1mmol/L, respectively. The method was applied to the quantification of glyphosate in a case of acute poisoning.

Aqueous phase behavior of tetraethylene glycol decanoyl ester (C9COE4) and Ether (C10E4) investigated by nuclear magnetic resonance spectroscopic techniques.

The binary phase diagram of tetraethylene glycol decanoyl ester (C9COE4) was investigated in the micellar region by PGSE-NMR (pulse field gradient spin echo nuclear magnetic resonance) and in the lamellar liquid crystalline state by 2H NMR. Its behavior was compared to the ether counterpart, tetraethylene glycol decanoyl ether (C10E4), whose phase diagram is well-described. The determination of the self-diffusion coefficient as a function of concentration permitted not only a determination of the critical micellar concentration (cmc) values but also the determination of the size and shape of micelles formed by both compounds. The evolution of the self-diffusion coefficients in the vicinity of the cloud point was also studied, showing no micellar growth with increasing temperature. 2H NMR analyses at the border of and within the liquid crystalline region gave an insight into the lamellar phase structure. We investigated in detail the lamellar phase of both compounds by comparing the values of quadrupolar splittings (Deltanu) measured under the same conditions. Lower Deltanu values were found for the ester compared to the ether: since the ester probably binds more water than the ether, these lower Deltanu values would indicate a lower order parameter in the liquid crystal phase. NMR techniques, either PGSE-NMR or 2H NMR, were confirmed as useful tools to characterize aqueous phase behavior of surfactants, providing supplementary information to the classical techniques such as visual observations, polarized optical microscopy (POM), and surface tension measurements. They also provide a unique insight into the molecular organization in the different phases formed.