Broadband Fourier transform rotational spectroscopy for structure determination: The water heptamer

Over the recent years chirped-pulse, Fourier-transform microwave (CP-FTMW) spectrometers have chan- ged the scope of rotational spectroscopy. The broad frequency and large dynamic range make possible structural determinations in molecular systems of increasingly larger size from measurements of heavy atom (13C, 15N, 18O) isotopes recorded in natural abundance in the same spectrum as that of the parent isotopic species. The design of a broadband spectrometer operating in the 2–8 GHz frequency range with further improvements in sensitivity is presented. The current CP-FTMW spectrometer performance is benchmarked in the analyses of the rotational spectrum of the water heptamer, (H2O)7, in both 2– 8 GHz and 6–18 GHz frequency ranges. Two isomers of the water heptamer have been observed in a pulsed supersonic molecular expansion. High level ab initio structural searches were performed to pro- vide plausible low-energy candidates which were directly compared with accurate structures provided from broadband rotational spectra. The full substitution structure of the most stable species has been obtained through the analysis of all possible singly-substituted isotopologues (H218O and HDO), and a least-squares rm(1) geometry of the oxygen framework determined from 16 different isotopic species compares with the calculated O–O equilibrium distances at the 0.01 Å level.

Broadband Fourier transform rotational spectroscopy for structure determination: The water heptamer

Over the recent years chirped-pulse, Fourier-transform microwave (CP-FTMW) spectrometers have changed the scope of rotational spectroscopy. The broad frequency and large dynamic range make possible structural determinations in molecular systems of increasingly larger size from measurements of heavy atom (C-13, N-15, O-18) isotopes recorded in natural abundance in the same spectrum as that of the parent isotopic species. The design of a broadband spectrometer operating in the 2-8 GHz frequency range with further improvements in sensitivity is presented. The current CP-FTMW spectrometer performance is benchmarked in the analyses of the rotational spectrum of the water heptamer, (H2O)(7), in both 2-8 GHz and 6-18 GHz frequency ranges. Two isomers of the water heptamer have been observed in a pulsed supersonic molecular expansion. High level ab initio structural searches were performed to provide plausible low-energy candidates which were directly compared with accurate structures provided from broadband rotational spectra. The full substitution structure of the most stable species has been obtained through the analysis of all possible singly-substituted isotopologues ((H2O)-O-18 and HDO), and a least-squares r(m)((1)) geometry of the oxygen framework determined from 16 different isotopic species compares with the calculated O-O equilibrium distances at the 0.01 angstrom level. (C) 2013 Elsevier B.V. All rights reserved.

Vibronic Spectra of Jet-Cooled 2-Aminopurine center dot H2O Clusters Studied by UV Resonant Two-Photon Ionization Spectroscopy and Quantum Chemical Calculations

For understanding the major- and minor-groove hydration patterns of DNAs and RNAs, it is important to understand the local solvation of individual nucleobases at the molecular level. We have investigated the 2-aminopurine center dot H2O. monohydrate by two-color resonant two-photon ionization and UV/UV hole-burning spectroscopies, which reveal two isomers, denoted A and B. The electronic spectral shift delta nu of the S-1 <- S-0 transition relative to bare 9H-2-aminopurine (9H-2AP) is small for isomer A (-70 cm(-1)), while that of isomer B is much larger (delta nu = 889 cm(-1)). B3LYP geometry optimizations with the TZVP basis set predict four cluster isomers, of which three are doubly H-bonded, with H2O acting as an acceptor to a N-H or -NH2 group and as a donor to either of the pyrimidine N sites. The "sugar-edge" isomer A is calculated to be the most stable form with binding energy D-e = 56.4 kJ/mol. Isomers B and C are H-bonded between the -NH2 group and pyrimidine moieties and are 2.5 and 6.9 kJ/mol less stable, respectively. Time-dependent (TD) B3LYP/TZVP calculations predict the adiabatic energies of the lowest (1)pi pi* states of A and B in excellent agreement with the observed 0(0)(0) bands; also, the relative intensities of the A and B origin bands agree well with the calculated S-0 state relative energies. This allows unequivocal identification of the isomers. The R2PI spectra of 9H-2AP and of isomer A exhibit intense low-frequency out-of-plane overtone and combination bands, which is interpreted as a coupling of the optically excited (1)pi pi* state to the lower-lying (1)n pi* dark state. In contrast, these overtone and combination bands are much weaker for isomer B, implying that the (1)pi pi* state of B is planar and decoupled from the (1)n pi* state. These observations agree with the calculations, which predict the (1)n pi* above the (1)pi pi* state for isomer B but below the (1)pi pi* for both 9H-2AP and isomer A.

Iron(III)-Pivalate-Based Complexes with Tetranuclear {Fe-4(mu(3)-O)(2)}(8+) Cores and N-Donor Ligands: Formation of Cluster and Polymeric Architectures

Different synthetic routes have been used for the preparation of a new tetranuclear [Fe4O2(O2CCMe3)(8)(bpm)] cluster (1) and a one-dimensional coordination polymer [Fe4O2-(O2CCMe3)(8)(hmta)](n) (2) (bpm = 2,2'-bipyrimidine and hmta = hexamethylenetetramine). For cluster 1, two structural isomers, 1a and 1b center dot 3MeCN, have been found. X-ray crystallographic analysis showed that all complexes consist of a central {Fe-4(mu(3)-O)(2)}(8+) core. In 1a, metal ions in the core are additionally linked by six bridging pivalates as two other pivalates and a bpm ligand are chelated to Fe-III ions, whereas in cluster 1b, metal ions in the {Fe-4(mu(3)-O)(2)}(8+) core are linked by seven bridging pivalates and only one carboxylate as well as bpm are chelated to the iron centers. In coordination polymer 2, [Fe4O2(O2CCMe3)(8)] clusters are bridged by hmta ligands to form zigzag chains. Magnetic measurements have been carried out to characterize these complexes and revealed antiferromagnetic interactions between Fe-III ions with best-fit parameters of J(wb) = -72.2 (1a) and -88.7 cm(-1) (1b) for wing...body interactions.

Studying Electrophoretic Separations Using Cholate Micelles: Effects Of Ph, Temperature, And Composition

Micelle-forming bile salts have previously been shown to be effective pseudo-stationary phases for separating the chiral isomers of binaphthyl compounds with micellar electrokinetic capillary chromatography (MEKC). Here, cholate micelles are systematically investigated via electrophoretic separations and NMR using R, S-1, 1¿- binaphthyl- 2, 2¿-diylhydrogenphosphate (BNDHP) as a model chiral analyte. The pH, temperature, and concentration of BNDHP were systematically varied while monitoring the chiral resolution obtained with MEKC and the chemical shift of various protons in NMR. NMR data for each proton on BNDHP is monitored as a function of cholate concentration: as cholate monomers begin to aggregate and the analyte molecules begin to sample the micelle aggregate we observe changes in the cholate methyl and S-BNDHP proton chemical shifts. From such NMR data, the apparent CMC of cholate at pH 12 is found to be about 13-14 mM, but this value decreases at higher pH, suggesting that more extreme pHs may give rise to more effective separations. In general, CMCs increase with temperature indicating that one may be able to obtain better separations at lower temperatures. S-BNDHP concentrations ranging from 50 ¿M to 400 ¿M (pH 12.8) gave rise to apparent cholate CMC values from 10 mM to 8 mM, respectively, indicating that S-BNDHP, the chiral analyte molecule, may play an active role in stabilizing cholate aggregates. In all, these data show that NMR can be used to systematically investigate a complex multi-variable landscape of potential optimizations of chiral separations.

Conformational analysis of a potent SSTR3-selective somatostatin analogue by NMR in water solution

The three-dimensional structure of a potent SSTR3-selective analogue of somatostatin, cyclo(3-14)H-Cys(3)-Phe(6)-Tyr(7)-D-Agl(8)(N(beta) Me, 2-naphthoyl)-Lys(9)-Thr(10)-Phe(11)-Cys(14)-OH (des-AA(1, 2, 4, 5, 12, 13)[Tyr(7), D-Agl(8)(N(beta) Me, 2-naphthoyl)]-SRIF) (peptide 1) has been determined by (1)H NMR in water and molecular dynamics (MD) simulations. The peptide exists in two conformational isomers differing mainly by the cis/trans isomerization of the side chain in residue 8. The structure of 1 is compared with the consensus structural motifs of other somatostatin analogues that bind predominantly to SSTR1, SSTR2/SSTR5 and SSTR4 receptors, and to the 3D structure of a non-selective SRIF analogue, cyclo(3-14)H-Cys(3)-Phe(6)-Tyr(7)-D-2Nal(8)-Lys(9)-Thr(10)-Phe(11)-Cys(14)-OH (des-AA(1, 2, 4, 5, 12, 13)[Tyr(7), D-2Nal(8)]-SRIF) (peptide 2). The structural determinant factors that could explain selectivity of peptide 1 for SSTR3 receptors are discussed.

The stereochemistry of the amino acid side chain influences the inflammatory potential of muramyl dipeptide in experimental meningitis

Intrathecal injections of 50 to 100 micro g of (N-acetylmuramyl-L-alanyl-D-isoglutamine) muramyl dipeptide (MDP)/rabbit dose-dependently triggered tumor necrosis factor alpha (TNF-alpha) secretion (12 to 40,000 pg/ml) preceding the influx of leukocytes in the subarachnoid space of rabbits. Intrathecal instillation of heat-killed unencapsulated R6 pneumococci produced a comparable leukocyte influx but only a minimal level of preceding TNF-alpha secretion. The stereochemistry of the first amino acid (L-alanine) of the MDP played a crucial role with regard to its inflammatory potential. Isomers harboring D-alanine in first position did not induce TNF-alpha secretion and influx of leukocytes. This stereospecificity of MDPs was also confirmed by measuring TNF-alpha release from human peripheral mononuclear blood cells stimulated in vitro. These data show that the inflammatory potential of MDPs depends on the stereochemistry of the first amino acid of the peptide side chain and suggest that intact pneumococci and MDPs induce inflammation by different pathways.

Development of a high performance parallel computing platform and its use in the study of nanostructures : clusters, sheets and tubes .

Small clusters of gallium oxide, technologically important high temperature ceramic, together with interaction of nucleic acid bases with graphene and small-diameter carbon nanotube are focus of first principles calculations in this work. A high performance parallel computing platform is also developed to perform these calculations at Michigan Tech. First principles calculations are based on density functional theory employing either local density or gradient-corrected approximation together with plane wave and gaussian basis sets. The bulk Ga2O3 is known to be a very good candidate for fabricating electronic devices that operate at high temperatures. To explore the properties of Ga2O3 at nonoscale, we have performed a systematic theoretical study on the small polyatomic gallium oxide clusters. The calculated results find that all lowest energy isomers of GamOn clusters are dominated by the Ga-O bonds over the metal-metal or the oxygen-oxygen bonds. Analysis of atomic charges suggest the clusters to be highly ionic similar to the case of bulk Ga2O3. In the study of sequential oxidation of these slusters starting from Ga2O, it is found that the most stable isomers display up to four different backbones of constituent atoms. Furthermore, the predicted configuration of the ground state of Ga2O is recently confirmed by the experimental result of Neumark's group. Guided by the results of calculations the study of gallium oxide clusters, performance related challenge of computational simulations, of producing high performance computers/platforms, has been addressed. Several engineering aspects were thoroughly studied during the design, development and implementation of the high performance parallel computing platform, rama, at Michigan Tech. In an attempt to stay true to the principles of Beowulf revolutioni, the rama cluster was extensively customized to make it easy to understand, and use - for administrators as well as end-users. Following the results of benchmark calculations and to keep up with the complexity of systems under study, rama has been expanded to a total of sixty four processors. Interest in the non-covalent intereaction of DNA with carbon nanotubes has steadily increased during past several years. This hybrid system, at the junction of the biological regime and the nanomaterials world, possesses features which make it very attractive for a wide range of applicatioins. Using the in-house computational power available, we have studied details of the interaction between nucleic acid bases with graphene sheet as well as high-curvature small-diameter carbon nanotube. The calculated trend in the binding energies strongly suggests that the polarizability of the base molecules determines the interaction strength of the nucleic acid bases with graphene. When comparing the results obtained here for physisorption on the small diameter nanotube considered with those from the study on graphene, it is observed that the interaction strength of nucleic acid bases is smaller for the tube. Thus, these results show that the effect of introducing curvature is to reduce the binding energy. The binding energies for the two extreme cases of negligible curvature (i.e. flat graphene sheet) and of very high curvature (i.e. small diameter nanotube) may be considered as upper and lower bounds. This finding represents an important step towards a better understanding of experimentally observed sequence-dependent interaction of DNA with Carbon nanotubes.

Stereoselective pharmacokinetics of ketamine and norketamine after constant rate infusion of a subanesthetic dose of racemic ketamine or S-ketamine in Shetland ponies

OBJECTIVE: To evaluate pharmacokinetics of ketamine and norketamine enantiomers after constant rate infusion (CRI) of a subanesthetic dose of racemic ketamine or S-ketamine in ponies. ANIMALS: Five 6-year-old Shetland pony geldings that weighed between 101 and 152 kg. PROCEDURES: In a crossover study, each pony received a CRI of racemic ketamine (loading dose, 0.6 mg/kg; CRI, 0.02 mg/kg/min) and S-ketamine (loading dose, 0.3 mg/kg; CRI, 0.01 mg/kg/min), with a 1-month interval between treatments. Arterial blood samples were collected before and at 5, 15, 30, 45, and 60 minutes during drug administration and at 5, 10, 30, and 60 minutes after discontinuing the CRI. Plasma ketamine and norketamine enantiomers were quantified by use of capillary electrophoresis. Individual R-ketamine and S-ketamine concentration-versus-time curves were analyzed by use of a monocompartmental model. Plasma disposition curves for R-norketamine and S-norketamine were described by estimating the area under the concentration-versus-time curve (AUC), maximum concentration (Cmax), and time until Cmax. RESULTS: Plasma concentrations of S-ketamine decreased and biodegradation products increased more rapidly after S-ketamine CRI, compared with results after racemic ketamine CRI. The R-norketamine was eliminated faster than was the S-norketamine. Significant differences between treatments were found for the AUC of S-ketamine and within the racemic ketamine CRI for the AUC and Cmax of norketamine isomers. CONCLUSIONS AND CLINICAL RELEVANCE: CRI of S-ketamine may be preferable over CRI of racemic ketamine in standing equids because the S-enantiomer was eliminated faster when infused alone instead of as part of a racemic mixture.

Synthesis of bicyclo-DNA nucleosides with additional functionalization in the carbocyclic ring

Two novel bicyclo nucleoside isomers carrying the base thymine in the furanose ring and an ester substituent in the carbocyclic ring were synthesized from a common bicyclic sugar precursor via a cyclopropanation/fragmentation pathway in nine steps. The relative configuration of the ester substituent in both isomers as well as the anomeric configuration in one nucleoside was determined by 1H-NMR difference NOE spectroscopy.

Synthesis and Biological Activity of 7,8,9-Trideoxy- and 7 R DesTHP-Peloruside A

The stereoselective syntheses of 7,8,9-trideoxypeloruside A (4) and a monocyclic peloruside A analogue lacking the entire tetrahydropyran moiety (3) are described. The syntheses proceeded through the PMB-ether of an ω-hydroxy β-keto aldehyde as a common intermediate which was elaborated into a pair of diastereomeric 1,3-syn and -anti diols by stereoselective Duthaler–Hafner allylations and subsequent 1,3-syn or anti reduction. One of these isomers was further converted into a tetrahydropyran derivative in a high-yielding Prins reaction, to provide the precursor for bicyclic analogue 4. Downstream steps for both syntheses included the substrate-controlled addition of a vinyl lithium intermediate to an aldehyde, thus connecting the peloruside side chain to C15 (C13) of the macrocyclic core structure in a fully stereoselective fashion. In the case of monocyclic 3 macrocyclization was based on ring-closing olefin metathesis (RCM), while bicyclic 4 was cyclized through Yamaguchi-type macrolactonization. The macrolactonization step was surprisingly difficult and was accompanied by extensive cyclic dimer formation. Peloruside A analogues 3 and 4 inhibited the proliferation of human cancer cell lines in vitro with micromolar and sub-micromolar IC50 values, respectively. The higher potency of 4 highlights the importance of the bicyclic core structure of peloruside A for nM biological activity.

Watson–Crick and Sugar-Edge Base Pairing of Cytosine in the Gas Phase: UV and Infrared Spectra of Cytosine·2-Pyridone

While keto-amino cytosine is the dominant species in aqueous solution, spectroscopic studies in molecular beams and in noble gas matrices show that other cytosine tautomers prevail in apolar environments. Each of these offers two or three H-bonding sites (Watson–Crick, wobble, sugar-edge). The mass- and isomer-specific S1 ← S0 vibronic spectra of cytosine·2-pyridone (Cyt·2PY) and 1-methylcytosine·2PY are measured using UV laser resonant two-photon ionization (R2PI), UV/UV depletion, and IR depletion spectroscopy. The UV spectra of the Watson–Crick and sugar-edge isomers of Cyt·2PY are separated using UV/UV spectral hole-burning. Five different isomers of Cyt·2PY are observed in a supersonic beam. We show that the Watson–Crick and sugar-edge dimers of keto-amino cytosine with 2PY are the most abundant in the beam, although keto-amino-cytosine is only the third most abundant tautomer in the gas phase. We identify the different isomers by combining three different diagnostic tools: (1) methylation of the cytosine N1–H group prevents formation of both the sugar-edge and wobble isomers and gives the Watson–Crick isomer exclusively. (2) The calculated ground state binding and dissociation energies, relative gas-phase abundances, excitation and the ionization energies are in agreement with the assignment of the dominant Cyt·2PY isomers to the Watson–Crick and sugar-edge complexes of keto-amino cytosine. (3) The comparison of calculated ground state vibrational frequencies to the experimental IR spectra in the carbonyl stretch and NH/OH/CH stretch ranges strengthen this identification.

Denaturation and unfolding of human anaphylatoxin C3a: an unusually low covalent stability of its native disulfide bonds.

The complement C3a anaphylatoxin is a major molecular mediator of innate immunity. It is a potent activator of mast cells, basophils and eosinophils and causes smooth muscle contraction. Structurally, C3a is a relatively small protein (77 amino acids) comprising a N-terminal domain connected by 3 native disulfide bonds and a helical C-terminal segment. The structural stability of C3a has been investigated here using three different methods: Disulfide scrambling; Differential CD spectroscopy; and Reductive unfolding. Two uncommon features regarding the stability of C3a and the structure of denatured C3a have been observed in this study. (a) There is an unusual disconnection between the conformational stability of C3a and the covalent stability of its three native disulfide bonds that is not seen with other disulfide proteins. As measured by both methods of disulfide scrambling and differential CD spectroscopy, the native C3a exhibits a global conformational stability that is comparable to numerous proteins with similar size and disulfide content, all with mid-point denaturation of [GdmCl](1/2) at 3.4-5M. These proteins include hirudin, tick anticoagulant protein and leech carboxypeptidase inhibitor. However, the native disulfide bonds of C3a is 150-1000 fold less stable than those proteins as evaluated by the method of reductive unfolding. The 3 native disulfide bonds of C3a can be collectively and quantitatively reduced with as low as 1mM of dithiothreitol within 5 min. The fragility of the native disulfide bonds of C3a has not yet been observed with other native disulfide proteins. (b) Using the method of disulfide scrambling, denatured C3a was shown to consist of diverse isomers adopting varied extent of unfolding. Among them, the most extensively unfolded isomer of denatured C3a is found to assume beads-form disulfide pattern, comprising Cys(36)-Cys(49) and two disulfide bonds formed by two pair of consecutive cysteines, Cys(22)-Cys(23) and Cys(56)-Cys(57), a unique disulfide structure of polypeptide that has not been documented previously.

Presence of three pesticides in rural drinking water sources in Kenya

A cross-sectional study on the use of three pesticides and their presence in drinking water sources was conducted in Githunguri/Kiaria community between January 1994-March 1995. The main objective of the study was to determine the extent to which some of the pesticides used by the Githunguri/Kiaria agricultural community were polluting their drinking water sources. Due to monetary and physical limitations, only DDT, its isomers and metabolites, carbofuran and carbaryl pesticides were identified and used as surrogates of pollution for the other pesticides.^ The study area was divided into high and low lying geographic surface areas. Thirty-four and 38 water sampling sites were randomly selected respectively. During wet and dry seasons, a total of 144 water samples were collected and analyzed at the Kenya Bureau of Standards Laboratory in Nairobi. Gas chromatography was used to analyze samples for possible presence of DDT, its isomers and metabolites, while high pressure liquid chromatography was used to analyze samples for carbofuran and carbaryl pesticides.^ Six sites testing positively for DDT, its isomers and metabolites represented 19.4% of the total sampled sites, with a mean concentration of 0.00310 ppb in the dry season and 0.0130 ppb in the wet season. All the six sites testing positively for the same pesticide exceeded the European maximum contaminant limit (MCL) in the wet season, and only one site exceeded the European MCL in the dry season.^ Those sites testing positively for carbofuran and carbaryl represented 5.6% of the total sampled sites. The mean concentration for the carbofuran at the sites was 2.500 ppb and 1.590 ppb in the dry and wet seasons respectively. Similarly, the mean concentration for carbaryl at the sites was 0.281 ppb in the dry season and 0.326 ppb in the wet season.^ One site testing positively for carbofuran exceeded the European MCL and WHO set limit in the wet season, while one site testing positively for the same pesticide exceeded the USA, Canada, European and WHO MCLs in the dry season. Similarly, one site which tested positively for carbaryl pesticide exceeded the European MCL in both seasons.^ Out of the 2,587 community members in the study area, 333 (13%) were exposed through their drinking water sources to the three pesticides investigated by this study. As a public health measure, integrated pest management approaches (IPM), protection of the wells and education of the community is necessary to minimize the pollution of the environment and safeguard the drinking water sources from pollution by the pesticides. ^

THE ISOLATION AND IDENTIFICATION OF O-ACETYLATED SIALIC ACIDS ON HUMAN CELLS (NEURAMINIC ACIDS, PAPER CHROMATOGRAPHY, HUMAN MALIGNANCY, BIOSYNTHETIC LABELING)

Unlike most carbohydrates, sialic acids have a restricted distribution in nature, being present in higher animals and in certain bacteriae. Unfortunately, most studies have not taken into account the fact that the parent sialic acid molecules, N-acetyl(or N-glycolyl)-neuraminic acid can be O-substituted at the 4, 7, 8 and 9 positions, generating many compounds and isomers. The approach and results of this research study demonstrates that proportions of non-, mono-, di-, and tri-O-acetylated sialic acids can be identified and quantitated on normal and malignant human cells. This was accomplished using a paper chromatographic technique to isolate and resolve individual species of non and O-substituted sialic acids. The chemical nature of these O-substituents, as an acetyl ester, was determined on the basis of chemical degradation, enzymatic and fast atom bombardment-mass spectrometry analysis.^ The working hypothesis of this study, that O-acetylated sialic acids are expressed in a restricted manner on normal and malignant cells, was confirmed using the above experimental approach; which identified mono-, di-, and tri-O-acetylated sialic acids on a variety of normal and malignant human cells. These O-acetylated sialic acids were expressed in restricted manner on subpopulations and subcellular fractions of PHL melanoma cells. Aberrant expression of O-acetylated sialic acids was associated with adenocarcinoma of the colon, leading to a nearly complete loss of di- and tri-O-acetylated sialic acids.^ Thus, the ability to isolate and identify biosynthetically radiolabeled O-acetylated sialic acids offers an efficient method of monitoring the expression of O-acetylated sialic acids in biochemical and cellular interactions. Furthermore, the ability to identify abnormal ratios of O-acetylated sialic acids in the human colon, represents a possible diagnostic tool to evaluate and identify patients who may be genetically or culturally predisposed to the development of adenocarcinoma of the colon. ^