Polyaniline based nucleic acid sensor

Twenty-bases long NH2-modified DNA and PNA probes specific to a pathogen (Mycobacterium tuberculosis) were covalently immobilized onto a polyaniline (PANI)/Au electrode to detect nucleic acid hybridization with complementary, one-base mismatch and noncomplementary targets within 30 s using Methylene Blue. The PNA-PANI/Au electrode exhibits improved specificity (1000 times) and detection limit (0.125 x 10(-18) M) as compared to that of the DNA-PANI/Au electrode (2.5 x 10(-18) M). These PNA-PANI/Au electrodes can be utilized for detection of hybridization with the complementary sequence in 5 min sonicated M. tuberculosis genomic DNA within 1 min of hybridization time. These DNA-PANI/Au and PNA-PANI/Au electrodes can be used 6-7 and 13-15 times, respectively.     

Differentiation of the diastereomeric synthetic precursors of isofebrifugine and febrifugine by electrospray ionization tandem mass spectrometry

Febrifugine is an alkaloid with potent antimalarial activity isolated from Dichroa febrifuga and Hydrangea umbellate, and it exists naturally with its diastereomeric component, isofebrifugine. Here we report the differentiation of diastereomeric synthetic precursors of isofebrifugine (1, cis) and febrifugine (2, trans) and a structurally similar model diastereomeric pair without a halogen substituent (3 and 4) by electrospray ionization (ESI) tandem mass spectrometry. Compounds 1-4 contain a tert-butoxycarbonyl (BOC) substituent, and the collision-induced dissociation (CID) spectra of the [M+H](+), [M+Na](+) and [M+Li](+) ions of 1-4 include the expected product ions corresponding to the loss of C(4)H(8) (isobutene) and of C(5)H(8)O(2) (BOC-H). Loss of C(5)H(8)O(2) is dominant in cis isomers (1 and 3) and/or loss of C(4)H(8) ions is dominant in trans isomers (2 and 4). The decomposition of [M+H](+) ions shows stereoselectivity in the formation of the [M+H-(BOC-H)-C(3)H(5)OBr](+) and [M+H-(BOC-H)-C(6)H(5)CH(2)OH](+) ions. The [M+Cat](+) ions (where Cat = Na or Li) additionally show loss of NaBr and HBr from [M+Cat-(BOC-H)](+), and these product ions are constantly more abundant in cis isomers than in trans isomers. The stereoselectivity for the product ion corresponding to the loss of [(BOC-H)+C(3)H(5)OBr] from [M+H](+) ions differs from that from [M+Cat](+) ions.     

Halogen-substituted phenylalanines as enantioselective selectors for enantioselective discrimination of amino acids: effect of halogen

Halogen-substituted phenylalanines with a halogen X (X = F, Cl, Br or I) in the para position in the aromatic ring of L-phenylalanine are used as enantioselective selectors to explore the effect of the halogen substituent on the enantioselective discrimination of amino acids. Enantioselective discrimination is achieved by investigating the collision-induced dissociation spectra of the trimeric complex ion, [CuII(ref)2(A)-H]+, generated by electrospraying a solution of a mixture of D- or L-analyte amino acid (A), enantioselective reference ligand (ref) and CuCl2. The relative abundances of fragment ions resulting from the competitive loss of reference and analyte amino acids are considered for measuring the degree of enantioselective discrimination by applying the kinetic method. The enantioselectivity of the p-halogenated derivatives of L-Phe increases from fluorine to iodine for the studied amino acids (except for acidic amino acids). The validity of the present method has also been checked by cross enantioselective experiments using p-iodo-D-phenylalanine as the reference in place of p-iodo-L-phenylalanine. The enantioselectivity of fluoro-substituted L-phenylalanine is less than that obtained with L-phenylalanine. The high inductive effect of the fluorine atom decreases the strength of the pi-pi stacking interaction. The presence of halogen affects the enantioselectivity by inductive and steric effects.     

Nucleic acid sensor for M. tuberculosis detection based on surface plasmon resonance

Cysteine modified NH(2)-end peptide nucleic acid (PNA) (24-mer) probe and 5'-thiol end labeled deoxyribonucleic acid (DNA) probes specific to Mycobacterium tuberculosis have been immobilized onto BK-7 gold coated glass plates for the detection of complementary, one-base mismatch, non-complementary targets and complementary target sequence in genomic DNA of Mycobacterium tuberculosis using a surface plasmon resonance (SPR) technique. The DNA/Au and PNA/Au bio-electrodes have been characterized using contact angle, atomic force microscopy (AFM), electrochemical impedance spectroscopy (EIS) and cyclic voltammetric (CV) techniques, respectively. It is revealed that there is a 252 millidegrees SPR angle change in the case of PNA immobilization and 205 millidegrees for DNA immobilization, indicating increased amount of immobilized PNA molecules. Hybridization studies reveal that there is no binding of the non-complementary target to DNA/Au and PNA/Au electrode. Compared to the DNA/Au bioelectrode, PNA/Au electrode has been found to be more efficient for detection of one-base mismatch sequence. The PNA/Au bioelectrode shows better detection limit (1.0 ng ml(-1)) over the DNA-Au bioelectrode (3.0 ng ml(-1)). The values of the association (k(a)) and dissociation rate constant (k(d)) for the complementary sequence in case of the PNA/Au bioelectrode have been estimated as 8.5 x 10(4) m(-1) s(-1) and 3.6 x 10(-3) s(-1), respectively.     

A cross-metathesis approach for the synthesis of tedanolide and 13-deoxytedanolide: stereoselective synthesis of the C3-C16 segment

A unified synthetic strategy has been devised for the synthesis of both tedanolide and 13-deoxytedanolide, which involves a cross-metathesis reaction as the key step. We report herein the stereoselective synthesis of the C3-C16 segment (+)-5b and subsequent manipulation of the C12-C13 double bond leading to the preparation of both tedanolides.     

Chiral discrimination of D- and L-amino acids using iodinated tyrosines as chiral references: Effect of iodine substituent

L-Tyrosine and iodinated L-tyrosines, i.e., 3-iodo-L-tyrosine and 3,5-diiodo-L-tyrosine, are successfully used as chiral references for the chiral discrimination of aliphatic, acidic, and aromatic amino acids. Chiral discrimination is achieved by investigating the collision-induced dissociation spectra of the trimeric complex [Cu(II)(ref)(2)(A) - H](+) ion generated by electro spraying the mixture of D- or L-analyte amino acid (A), chiral reference ligand (ref) and M(II)Cl(2) (M = Ni and Cu). The relative abundances of fragment ions resulted by the competitive loss of reference and analyte amino acids are considered for measuring the degree of chiral discrimination by applying the kinetic method. The chiral discrimination ability increases as the number of iodine atom increases on the aromatic ring of the reference and the discrimination is better with Cu when compared with Ni. A large chiral discrimination is obtained for aliphatic and aromatic amino acids using iodinated L-tyrosine as the reference. Computational studies on the different stabilities of the diastereomeric complexes also support the observed differences measured by the kinetic method. The suitability of the method in the measurement of enantiomeric excess over the range of 2% to 100% ee with relative error 0.28% to 1.6% is also demonstrated.     

Chiral discrimination of alpha-amino acids by the DNA triplet GCA using amino acids as a co-selector

The DNA triplet GCA is successfully used as a chiral selector for the chiral discrimination of amino acids using amino acids themselves as a co-selector. Chiral discrimination was achieved by investigating the collision-induced dissociation spectra of the [X(A) + X(R) + 2Y - 2H](2-) ion generated by electrospraying a mixture of analyte amino acid (X(A)), reference amino acid (X(R)) and GCA (Y). The relative abundances of fragment ions resulting from the competitive loss of reference and X(A)'s are considered for measuring the degree of chiral discrimination. GCA successfully shows D-selectivity for all the amino acids, except Tyr and Lys. The success of the method lies in the selection of a suitable 10(R) that has closer GCA binding affinity to that of analyte. The degree of discrimination by GCA is improved in the presence of the reference, and the chirality of the reference does not change the selectivity of GCA. The suitability of the method for the measurement of optical purity is also demonstrated.     

Formation of left-handed helices in hybrid peptide oligomers with cis beta-sugar amino acid and L-Ala as building blocks

Residue based control of specific helical folding is explored in hybrid peptide oligomers consisting of alternating L-Ala and cis-beta-furanoid sugar amino acid (FSAA) residues as building blocks; two series of these hybrid oligomers are designed, synthesized and extensively characterized by using NMR, CD, FT-IR and MD simulation studies; results show the co-existence of left-handed 11- and 14/15-helical conformations in these short oligomers of Boc-(alpha/beta) and Boc-(beta/alpha) series.     

Umbrella sampling for nonequilibrium processes

The authors introduce an algorithm for determining the steady-state probability distribution of an ergodic system arbitrarily far from equilibrium. By enforcing equal sampling of different regions of phase space, as in umbrella sampling simulations of systems at equilibrium, low probability regions are explored to a much greater extent than in physically weighted simulations. The algorithm can be used to accumulate joint statistics for an arbitrary number of order parameters for a system governed by any stochastic dynamics. They demonstrate the efficiency of the algorithm by applying it to a model of a genetic toggle switch which evolves irreversibly according to a continuous time Monte Carlo procedure.     

Mass spectral study of substituted allyl aryl and allyl alkyl selenides and some analogous sulfides

The electron impact (EI) mass spectra of allyl aryl selenides showed abundant molecular ions and many fragment ions containing the selenium atom. alpha-Cleavage is the dominant process in the fragmentation of selenides, and cleavage product ions are characteristic of the substituents. In the case of 3-methyl allyl and related aryl selenides, characteristic delta-hydrogen migration to the selenium atom is observed. A McLafferty-type rearrangement is found in benzyl allyl selenides and substituted alkyl allyl selenides. The charge on the rearrangement products preferably remains on the fragments containing the phenyl group. The [M - SeH](+), [M - CH(3)](+) and [M - C(2)H(4)](+) ions are found only in the EI mass spectrum of allyl phenyl selenide, and are attributed to a Claisen rearrangement in the source of the mass spectrometer. All structurally informative fragmentation processes are supported by collision induced dissociation spectra of molecular ions. The fragmentation patterns found in methane chemical ionization (CI) spectra of the selenides were significantly different from those observed in EI. The EI and CI mass spectra of analogous sulfides showed similar behaviour to that observed in the corresponding selenides. Copyright 1999 John Wiley & Sons, Ltd.