No abstract

Recent developments in ionic liquid electrolytes for cellulose or biomass dissolution has also allowed for high-resolution H andC NMR on very high molecular weight cellulose. This permits the development of advanced liquid-state quantitative NMR methods for characterization of unsubstituted and low degree of substitution celluloses, for example, surface-modified nanocelluloses, which are insoluble in all molecular solvents. As such, we present the use of the tetrabutylphosphonium acetate ([P][OAc]):DMSO- d electrolyte in the 1D and 2D NMR characterization of poly(methyl methacrylate) (PMMA)-grafted cellulose nanocrystals (CNCs). PMMA- g-CNCs was chosen as a difficult model to study, to illustrate the potential of the technique. The chemical shift range of [P][OAc] is completely upfield of the cellulose backbone signals, avoiding signal overlap. In addition, application of diffusion-editing for H and HSQC was shown to be effective in the discrimination between PMMA polymer graft resonances and those from low molecular weight components arising from the solvent system. The bulk ratio of methyl methacrylate monomer to anhydroglucose unit was determined using a combination of HSQC and quantitativeC NMR. After detachment and recovery of the PMMA grafts, through methanolysis, DOSY NMR was used to determine the average self-diffusion coefficient and, hence, molecular weight of the grafts compared to self-diffusion coefficients for PMMA GPC standards. This finally led to a calculation of both graft length and graft density using liquid-state NMR techniques. In addition, it was possible to discriminate between triads and tetrads, associated with PMMA tacticity, of the PMMA still attached to the CNCs (before methanolysis). CNC reducing end and sulfate half ester resonances, from sulfuric acid hydrolysis, were also assignable. Furthermore, other biopolymers, such as hemicelluloses and proteins (silk and wool), were found to be soluble in the electrolyte media, allowing for wider application of this method beyond just cellulose analytics.

We characterize the dark-adapted photoresponses from mouse cones intact in the isolated retina, their virtually natural environment, by isolating pharmacologically the photoreceptor light responses from the electroretinogram (ERG). Due to the different photoresponse kinetics and sensitivity of rods and cones, the cone responses were readily attained by using a rod-saturating preflash. The stimulus wavelength (544 nm) was chosen to selectively stimulate the green sensitive ("M"-)pigment. Obtained responses were monophasic, showing fast kinetics (mean t(p)=51 ms) and low sensitivity (fractional single-photon response ca. 0.23%). The amplification coefficient of cones (4.6 s(-2)) was very close to that of rods (5.6 s(-2)), while the dominant time constant of recovery was clearly smaller for cones (33 ms) than for rods (160 ms).

Rod responses to brief pulses of light were recorded as electroretinogram (ERG) mass potentials across isolated, aspartate-superfused rat retinas at different temperatures and intensities of steady background light. The objective was to clarify to what extent differences in sensitivity, response kinetics and light adaptation between mammalian and amphibian rods can be explained by temperature and outer-segment size without assuming functional differences in the phototransduction molecules. Corresponding information for amphibian rods from the literature was supplemented by new recordings from toad retina. All light intensities were expressed as photoisomerizations per rod (Rh * ). In the rat retina, an estimated 34% of incident photons at the wavelength of peak sensitivity caused isomerizations in rods, as the (hexagonally packed) outer segments measured 1.7 µm × 22 µm and had specific absorbance of 0.016 µm −1 on average. Fractional sensitivity (S) in darkness increased with cooling in a similar manner in rat and toad rods, but the rat function as a whole was displaced to a ca 0.7 log unit higher sensitivity level. This difference can be fully explained by the smaller dimensions of rat rod outer segments, since the same rate of phosphodiesterase (PDE) activation by activated rhodopsin will produce a faster drop in cGMP concentration, hence a larger response in rat than in toad. In the range 15-25• C, the waveform and absolute time scale of dark-adapted dim-flash photoresponses at any given temperature were similar in rat and toad, although the overall temperature dependence of the time to peak (t p ) was somewhat steeper in rat (Q 10 ≈ 4 versus 2-3). Light adaptation was similar in rat and amphibian rods when measured at the same temperature. The mean background intensity that depressed S by 1 log unit at 12• C was in the range 20-50 Rh * s −1 in both, compared with ca 4500 Rh * s −1 in rat rods at 36 • C. We conclude that it is not necessary to assume major differences in the functional properties of the phototransduction molecules to account for the differences in response properties of mammalian and amphibian rods. The phototransduction cascade and its regulatory mechanisms are basically similar in all rod photoreceptors that have been studied (see . On the other hand, quantitative parameters of amplification, activation and deactivation kinetics, and light adaptation derived from the electrical responses to light differ so as to suggest important differences in the functioning of the phototransduction molecules in mammals and 'lower vertebrates' (commonly represented by amphibians). The rods of both classes can respond reliably to a single photon, but the initial amplification rate in mammalian rods is higher by two orders of magnitude and the response peaks at a much earlier time after photon absorption (Baylor et al. 1979b(Baylor et al. , 1984Matthews, 1991;Robinson et al. 1993;Kraft et al. 1993;Nikonov et al. 2000). Although mammalian rods, including those of humans, do have the capacity to light adapt, ...