Application of the general height equivalent to a theoretical plate equation to size exclusion chromatography. Study of the mass transfer of high-molecular-mass compounds in liquid chromatography

The mass transfer of nonretained, high-molecular-weight polystyrene standards in a chromatographic column packed with 4-microm-diameter silica particles for SEC (average pore size 80 A) and eluted with tetrahydrofuran was investigated. The HETPs of nine polystyrene standards (MW 560, 760, 1780, 3680, 6400, 13,200, 19,300, 31,600, and 90,000) and of ethylbenzene were measured by the method of moments. These HETPs were accounted for by a general HETP equation previously derived for the mass transfer of low-molecular-weight compounds in RPLC (Gritti, F.; Guiochon, G. Anal. Chem. 2006, 78, 5329.). The best fit between the experimental and the theoretical HETPs allowed the estimation of the internal restricted diffusion coefficients of the polymers. The internal diffusion coefficients of the polystyrene standards were also measured, using the peak parking method. Both methods give comparable results and show that the restriction diffusion coefficient gammapKD decreases linearly with increasing logarithm of the polymer mass, from 0.56 for Log MW = 2.0 to 0 for Log MW = 4.8. Interestingly, the C term or limit slope of the high-velocity branch of the HETP curve passes through a maximum (highest mass-transfer resistance) for a polymer mass of 10 kDa. This maximum is well accounted for by the theoretical expression of the trans-particle mass-transfer contribution.     

Second-harmonic-generation-polarimeter calibration by means of a quartz plate

A dual-rotating-retarder polarimeter was used to determine the six measurable observables of the first hyperpolarizability tensor. Calibration of such an instrument requires a reference sample dedicated to wavelength conversion. We calibrated our experimental setup by using a quartz-plate sample in a two step procedure: at first the first retarder then the second one. The retardance and ellipticity angle of both retarders were estimated by minimizing a chi(2) function. We estimated the standard deviation of each parameter from noise spreading and performed this calibration procedure for two experimental case studies, i.e., two angular positions of the quartz sample.     

Thermal compensation in GaPO4 beam resonators: experimental evidence for length extensional mode

Temperature effects in gallium orthophosphate (GaPO4) vibrating beams are reported. In addition to the well-known, thickness-shear AT-cut, temperature-compensated cuts exist in GaPO4 for length extensional modes. Experimental evidence of a temperature-compensated cut in GaPO4 rectangular beam resonator vibrating in length extension is given.     

Myelinosome Organelles in the Retina of R6/1 Huntington Disease (HD) Mice: Ubiquitous Distribution and Possible Role in Disease Spreading

Visual deficit is one of the complications of Huntington disease (HD), a fatal neurological disorder caused by CAG trinucleotide expansions in the Huntingtin gene, leading to the production of mutant Huntingtin (mHTT) protein. Transgenic HD R6/1 mice expressing human HTT exon1 with 115 CAG repeats recapitulate major features of the human pathology and exhibit a degeneration of the retina. Our aim was to gain insight into the ultrastructure of the pathological HD R6/1 retina by electron microscopy (EM). We show that the HD R6/1 retina is enriched with unusual organelles myelinosomes, produced by retinal neurons and glia. Myelinosomes are present in all nuclear and plexiform layers, in the synaptic terminals of photoreceptors, in the processes of retinal neurons and glial cells, and in the subretinal space. In vitro study shows that myelinosomes secreted by human retinal glial Müller MIO-M1 cells transfected with EGFP-mHTT-exon1 carry EGFP-mHTT-exon1 protein, as revealed by immuno-EM and Western-blotting. Myelinosomes loaded with mHTT-exon1 are incorporated by naive neuronal/neuroblastoma SH-SY5Y cells. This results in the emergence of mHTT-exon1 in recipient cells. This process is blocked by membrane fusion inhibitor MDL 28170. Conclusion: Incorporation of myelinosomes carrying mHTT-exon1 in recipient cells may contribute to HD spreading in the retina. Exploring ocular fluids for myelinosome presence could bring an additional biomarker for HD diagnostics.     

Inhibition of Very Long Chain Fatty Acids Synthesis Mediates PI3P Homeostasis at Endosomal Compartments

A main characteristic of sphingolipids is the presence of a very long chain fatty acid (VLCFA) whose function in cellular processes is not yet fully understood. VLCFAs of sphingolipids are involved in the intracellular traffic to the vacuole and the maturation of early endosomes into late endosomes is one of the major pathways for vacuolar traffic. Additionally, the anionic phospholipid phosphatidylinositol-3-phosphate (PtdIns (3)P or PI3P) is involved in protein sorting and recruitment of small GTPase effectors at late endosomes/multivesicular bodies (MVBs) during vacuolar trafficking. In contrast to animal cells, PI3P mainly localizes to late endosomes in plant cells and to a minor extent to a discrete sub-domain of the plant’s early endosome (EE)/trans-Golgi network (TGN) where the endosomal maturation occurs. However, the mechanisms that control the relative levels of PI3P between TGN and MVBs are unknown. Using metazachlor, an inhibitor of VLCFA synthesis, we found that VLCFAs are involved in the TGN/MVB distribution of PI3P. This effect is independent from either synthesis of PI3P by PI3-kinase or degradation of PI(3,5)P₂ into PI3P by the SUPPRESSOR OF ACTIN1 (SAC1) phosphatase. Using high-resolution live cell imaging microscopy, we detected transient associations between TGNs and MVBs but VLCFAs are not involved in those interactions. Nonetheless, our results suggest that PI3P might be transferable from TGN to MVBs and that VLCFAs act in this process.     

Preparation of 5‐Membered Rings via Radical Addition‐ Translocation‐Cyclization (RATC) Processes Mediated by Diethyl Thiophosphites

A practical method for the formation of thiophosphonates bearing functionalized monocyclic, fused bicyclic and spirocyclic residues is presented. The procedure requires the easily available terminal alkynes as starting materials as well as commercially and readily available reagents such as diethyl thiophosphite. The experimental procedure consists of a one‐pot process without any slow addition of one of the reagents.     

Self-assembly and characterization of layered double hydroxide/DNA hybrids

The purpose of this study was to control the fabrication of new labile supramolecular assemblies by formulating associations of DNA molecules with inorganic layered double hydroxides (LDHs). The results show that LDH/DNA hybrids synthesized by a coprecipitation route involving the in situ formation of LDHs around DNA molecules acting as templates were characterized by a lamellar organization, with DNA molecules sandwiched between hydroxide layers, exhibiting a regular spacing of 1.96 nm. Our results indicate that labile complexes resulting from the association of nucleic acids and inorganic materials can be obtained not only by anion exchange but also by a direct self-assembly route.     

Heterogeneity of the adsorption mechanism of low molecular weight compounds in reversed-phase liquid chromatography

The retention mechanism in RPLC mode was investigated based on the acquisition of adsorption isotherm data by frontal analysis measurements and their modeling. This work is a review of the results of four years of adsorption data measurements. The data were acquired on a wide variety of brands of C18-silica columns (from Akzo Nobel, Bishoff, Hypersil, Merck, Phenomenex, Supelco, Vydac, and Waters) with several low molecular weight compounds such as phenol (94 g/mol), caffeine (194 g/mol), tryptophan (204 g/mol), sodium 2-naphthalenesulfonate (235 g/mol), and propranololium chloride (295 g/mol). The mobile phase was a mixture of methanol and water at variable composition. The adsorption isotherms were all convex upward (langmuirian), and the degree of heterogeneity of the adsorption system was determined from the calculation of the adsorption energy distribution using the expectation-maximization method. The adsorption isotherm parameters (number of types of adsorption sites, surface concentration of each type of site, and difference between the adsorption energies E(i) - E(j) on sites i and j), obtained from the mathematical fit of the adsorption data to the appropriate multi-Langmuir adsorption isotherm model, were analyzed and compared. The results allow the drawing of general conclusions regarding the relationships between the size of the analyte and the adsorption properties (saturation capacities, adsorption energies) characterizing the retention mechanism in RPLC mode for neutral, anionic, and cationic compounds.     

Frequency-temperature behavior of flexural quartz resonators by means of Timoshenko's model

The frequency of a flexural resonator and its frequency-temperature behavior usually are computed by Bernoulli's classical approximation. This approach is valid for beams with a large length-over-thickness-ratio. For shorter beams, the effects of shear stress and rotary inertia may play a significant role for temperature-compensated resonators. These effects have been taken into account for isotropic beams. The aim of this paper is to discuss the extension of the shear coefficient in the case of an anisotropic material and to compute the frequency-temperature characteristic of an (XYt)theta cut resonator when the shear stress and the rotary inertia have been taken into account. Comparisons between the classical approximation and this treatment are given for quartz. Furthermore, the numerical predictions obtained by means of different sets of data available for thermal sensitivities of elastic coefficients are compared.