Comparison of a 3D-model of the classical α-scorpion toxin V from Leiurus quinquestriatus quinquestriatus with other scorpion toxins

In the present paper, a study of classical and insect alpha-scorpion toxins is described. A homology model of the classical alpha-toxin LqqV from Leiurus quinquestriatus quinquestriatus was developed. The model was compared to stable and energetically favourable conformations of AaHII from Androctonus australis Hector and LqhalphaIT from Leiurus quinquestriatus hebraeus, which are the most active alpha-toxins in mammals and insects. The conformations were retrieved from molecular dynamics simulations of known structures. The model of LqqV shows a C-terminal conformation similar to LqhalphaIT. This is mainly caused by electrostatic interactions between Lys10 /Lys60 and Glu59, which are comparable to the cation-pi interactions of Tyr10 and Arg64 in LqhalphaIT. During the simulations the structures of AaHII and LqqV were stabilised through electrostatic interactions between Glu32 and Lys50 and especially the loop adjacent to the alpha-helix is affected, which is in contrast to LqhalphaIT. When the molecular electrostatic potentials of the toxins were studied, a possibly important difference between the classical alpha-toxins and the insect alpha-toxin LqhalphaIT was found in the area around Lys30 and Arg56 of AaHII, where a positive potential is missing in LqhalphaIT. A large negative potential caused by Asp3, Glu15 and Asp19 in LqhalphaIT is also unique for this toxin. It is proposed that Arg18, which is important for activity of LqhalphaIT, restricts the negative potential in this area and is not essential for toxins where negatively charged residues in comparable positions are not present.     

Solvent gradient operation of simulated moving beds. I. Linear isotherms

The simulated moving bed (SMB) is a multi-column chromatographic separation process, which--with respect to the single-column preparative batch process--allows for a continuous separation with larger productivity and smaller solvent consumption at the same time. The benefits of this process have been shown for several different applications in fine chemistry, particularly for the separation of enantiomers. In general, SMBs are operated under isocratic conditions. However, separation performance can be further improved by applying some sort of gradient mode operation, in order to optimize the operating conditions of each individual section of the unit. This can be achieved by tuning the retention behavior of the solutes to be separated along the unit, namely by enforcing weak adsorption conditions in sections 1 and 2, and strong adsorption conditions in sections 3 and 4. This can be achieved by applying a temperature gradient (high temperature in section 1, and low temperature in section 4), a pressure gradient (e.g. in the supercritical SMB, when pressure is high in section 1, and low in section 4), or a solvent gradient, which is the aim of this work. In the solvent gradient mode the mobile phase consists of a mixture of two or more solvents. To different mobile phase compositions corresponds a different retention behavior of the solutes, i.e. different adsorption isotherms. In this work we study a closed loop SMB unit with solvent mixtures of two different compositions entering the unit at the feed and desorbent inlet ports, respectively. Thereby two different mobile phase compositions are established in sections 1 and 2, and sections 3 and 4, respectively. To optimize this process the equilibrium theory design criteria for non-linear SMBs are extended to describe this operation mode. It is shown how the region of separation is derived and how the optimal operating conditions can be found. Finally the solvent gradient mode is compared with the isocratic mode in terms of productivity and solvent consumption.     

A new type of intermediate, C+(BCH3)11- <--> C(BCH3)11, in a Grob fragmentation coupled with intramolecular hydride transfer. A nonclassical carbocation ylide or a carbenoid?

In solvolysis of alkyl halides Hal-(CH(2))(n)-C(BCH(3))(11)(-) (n = 2, 5, 6, but not 3, 4, or 7) and protonation of alkenes CH(2)=CH-(CH(2))(n)(-)(2)-C(BCH(3))(11)(-) (n = 3, 6, 7, but not 4 or 5) carrying the icosahedral electrofuge -C(BCH(3))(11)(-) attached through its cage carbon atom, generation of incipient positive charge on C(alpha) (as shown in Scheme 1 in the article) leads to simultaneous cleavage of the C(beta)-C(BCH(3))(11)(-) bond. The products are a C(alpha)=C(beta) alkene and a postulated intermediate C(+)(BCH(3))(11)(-) <--> C(BCH(3))(11), trapped as the adduct Nu-C(BCH(3))(11)(-) by one of the nucleophiles (Nu(-)) present. The reaction kinetics is E1, first order in the haloalkylcarborane and zero order in [Nu(-)], and the elimination appears to be concerted, as in the usual E2 mechanism. The process is best viewed as a Grob fragmentation. The loss of the longer chains involves intrachain hydride transfer from the C(alpha)-H bond to an incipient carbocation on C(delta)(') or C(epsilon)(') via a five- or six-membered cyclic transition state, respectively. The electronic structure of the postulated intermediate is believed to lie between those of a nonclassical carbonium ylide C(+)(BCH(3))(11)(-) and a carbenoid C(BCH(3))(11) whose electronic ground state resembles the S(2) state of ordinary carbenes.     

A new SCF procedure and its applications to ab initio calculations of the states of the fluorosulphate radical

Ab initio calculations using a small Gaussian basis set, including 3d orbitals on the sulphur atom, have been performed on the fluorosulphate radical and the related ions SO₃F⁺ and SO₃F^?. A new SCF procedure is described and applied to the open shell cases discussed here. The results are compared with recent CNDO calculations and with the experimental transition energies of the radical.     

Configuration interaction calculations of the valence ionization energies of the water dimer

The eight vertical valence ionization energies of the water dimer are calculated by the ΔCI method. Excellent agreement with measurements of the first and second ionization energies is found. Calculations of the remaining six ionization energies is found. Calculations of the remaining six ionization energies are sufficiently accurate to be of value in the identification and assignment of the dimer photoelectron spectrum.     

Mechanisms of elimination reactions—XXVI: The α'-β mechanism in elimination reactions of sulforium salts

Tracer studies with β-deuterated sulfonium salts have shown that the α'-β, or ylid mechanism can be the major path of elimination in the reaction of sulfonium salts with t-butoxide in t-butyl alcohol, while the E2 reaction is dominant with hydroxide in water or n-butoxide in n-butyl alcohol. The structure of the sulfonium salt also affects the propensity toward α'-β elimination, with 3-pentyl > 3-propyl and eyclopentyl > 3-pentyl > cyclohexyl. The S-methyl protons of the sulfonium salt exchange at a rate much faster than that of the elimination reaction. A strongly basic medium and a syn-periplanar arrangement of the α-C-S and β-C-H bonds seem to be the two most important factors favoring the α'-β mechanism.     

Small-angle neutron scattering study of temperature-induced emulsion gelation: the role of sticky microgel particles

In this work, small-angle neutron scattering (SANS) is used to probe the structural transformations that accompany temperature-induced gelation of emulsions stabilized by a temperature-responsive polymer. The latter is poly(NIPAM-co-PEGMa) (N-isopropylacrylamide and poly(ethyleneglycol) methacrylate) and contains 86 mol% NIPAM. Turbidity measurements revealed that poly(NIPAM-co-PEGMa) has a lower critical solution temperature (T(LCST)) of 36.5 degrees C in D(2)O. Aqueous polymer solutions were used to prepare perfluorodecalin-in-water emulsions (average droplet size of 6.9 mum). These emulsions formed gels at 50 degrees C. SANS measurements were performed on the poly(NIPAM-co-PEGMa) solutions and emulsions as a function of temperature. The emulsion was also prepared using a D2O/H2O mixture containing 72 vol% D2O in order to make scattering from the droplets negligible (on-contrast). The SANS data were analyzed using a combination of Porod and Ornstein-Zernike form factors. The results showed that the correlation length (xi) of the polymer scaled as xi approximately phi(p)(-0.68) at 32 degrees C, where phi(p) is the polymer volume fraction. The xi value increased for all systems as the temperature increased, which was attributed to a spinodal transition. At temperatures greater than T(LCST), the polymer solution changed to a polymer dispersion of poly(NIPAM-co-PEGMa) aggregates. The aggregates have features that are similar to microgel particles. The average size of these particles was estimated as 160-170 nm. The particles are "sticky" and are gel-forming. The on-contrast experiments performed using the emulsion indicated that the interfacial polymer chains condensed to give a relatively thick polymer layer at the perfluorodecalin-water interface at 50 degrees C. The gelled emulsions appear to consist of perfluorodecalin droplets with an encapsulating layer of collapsed polymer to which sticky microgel particles are adsorbed. The latter act as a "glue" between coated droplets in the emulsion gel.     

Method development and validation for the analysis of didanosine using micellar electrokinetic capillary chromatography

A selective MEKC method was developed for the analysis of didanosine in bulk samples. Successful separation of didanosine from 13 of its potential impurities, derived from the various synthetic preparation procedures, was achieved. As CZE gave poor separation selectivity, MEKC was preferable. The use of EKC allowed achievement of the separation in a significantly shorter time than conventional HPLC. An anionic long-chain surfactant, lithium dodecyl sulfate (LiDS), was used as the pseudostationary phase and sodium tetraborate buffer as the aqueous phase. In order to obtain the optimal conditions and to test the method robustness, a central composite response surface modeling experiment was performed. The optimized electrophoretic conditions include the use of an uncoated fused-silica capillary with a total length of 40 cm and an ID of 50 microm, a BGE containing 40 mM sodium tetraborate and 110 mM LiDS at pH 8.0, an applied voltage of 18.0 kV, and the capillary temperature maintained at 15 degrees C. The method was found to be robust. The parameters for validation such as linearity, precision, and sensitivity are also reported. Three commercial bulk samples were analyzed with this system.     

Linking protein fractionation with multidimensional monolithic reversed-phase peptide chromatography/mass spectrometry enhances protein identification from complex mixtures even in the presence of abundant proteins

Recently, multidimensional shotgun proteomics has proven to be an alternative technology able to identify hundreds of proteins from single samples. Two major limitations of the technology are the presence of high abundance proteins (e.g. RUBISCO in plant leaf tissue) and the enormous number of co-eluting peptides that overstrain the loading and resolving capacity of conventional particle-packed columns as well as the capacity of electrospray ionisation due to ion suppression. Here, the coupling of fast performance liquid chromatography (FPLC) pre-fractionation of an Arabidopsis leaf protein extract and subsequent two-dimensional liquid chromatography/mass spectrometry with improved resolution using a monolithic silica C18 capillary column allowed the identification of 1032 unique proteins in a single 4 mg total protein plant leaf tissue sample. The reassignment of peptide IDs to distinct FPLC protein fractions enhances the identification procedure, especially in the case of present protein isoforms. The proposed strategy is useful to detect proteins otherwise not seen in conventional multidimensional chromatography/mass spectrometry approaches.