Expression of Avian Influenza Virus (H5N1) Hemagglutinin and Matrix Protein 1 in Pichia pastoris and Evaluation of their Immunogenicity in Mice

The conventional avian influenza vaccines rely on development of neutralizing antibodies against the HA and NA antigens. However, these antigens are highly variable, and hence there is a need for better vaccine candidates which would offer broader protection in animals. The M1 of avian influenza is another major structural protein that has conserved epitopes that are reported to induce CD8+ T cells and can contribute to protection against morbidity and mortality from influenza. Hence in an effort to study the immune response of rM1 either alone or in combination with rHA, the hemagglutinin (HA) and matrix protein (M1) of A/Hatay/2004/H5N1 strain of avian influenza were expressed in Pichia pastoris as his-tagged proteins and purified through Ni-NTA chromatography. The His-tag was removed using TEV protease cleavage site and the immunogenicity of purified rHA and rM1 either alone or in combination was determined in mice. One group of mice was immunized with 5 μg of purified rHA, the other group was immunized with rM1, and a third group of mice were immunized with 5 μg of rHA and rM1. All the animals were boosted twice, once on 28 days postimmunization (dpi) and the second on 42 dpi. The immune response was evaluated by enzyme-linked immunosorbent assay (ELISA) and hemagglutination inhibition (HI) assay. The group of mice immunized with rHA and rM1 together showed significantly higher immune response against rHA and rM1 than mice immunized with either HA or M1 antigens. The addition of rM1 with rHA resulted in increased HI titer in animals immunized with both the antigens. These results suggest that the HA and M1 expressed in P. pastoris can be utilized in combination for the development of faster and cost-effective vaccines for circulating and newer strains of avian influenza and would aid in combating the disease in a pandemic situation, in which production time matters greatly.     

Regulation of vincamine biosynthesis and associated growth promoting effects through abiotic elicitation,cyclooxygenase inhibition,and precursor feeding of bioreactor grown Vinca minor hairy roots

Hydroxylase/acetyltransferase elicitors and cyclooxygenase inhibitor along with various precursors from primary shikimate and secoiridoid pools have been fortified to vincamine less hairy root clone of Vinca minor to determine the regulatory factors associated with vincamine biosynthesis. Growth kinetic studies revealed that acetyltransferase elicitor acetic anhydride and terpenoid precursor loganin significantly reduce the growth either supplemented alone or in combination (GI?=?140.6?±?18.5 to 246.7?±?24.3), while shikimate and tryptophan trigger biomass accumulation (GI?=?440.2?±?31.5 to 540.5?±?40.3). Loganin also downregulates total alkaloid biosynthesis. Maximum flux towards vincamine production (0.017?±?0.001 % dry wt.) was obtained when 20-day-old hairy roots were fortified with secologanin (10 mg/l) along with tryptophan (100 mg/l), naproxen (8.4 mg/l), hydrogen peroxide (20 μg/l), and acetic anhydride (32.4 mg/l). This was supported by RT PCR (qPCR) analysis where 2- and 3-fold increase in tryptophan decarboxylase (TDC; RQ?=?2.0?±?0.09) and strictosidine synthase (STR; RQ?=?3.3?±?0.36) activity, respectively, was recorded. The analysis of variance (ANOVA) for growth kinetics, total alkaloid content, and gene expression studies favored highly significant data (P?<?0.05–0.01). Above treated hairy roots were also up-scaled in a 5-l stirred-tank bioreactor where a 40-day cycle yielded 8-fold increase in fresh root mass.     

Poly(3,4‐ethylenedioxythiophene) (PEDOT)‐Coated MWCNTs Tethered to Conducting Substrates: Facile Electrochemistry and Enhanced Coloring Efficiency

Composite films of poly(3,4‐ethylenedioxythiophene) (PEDOT)‐coated over functionalized multiwalled coiled and linear carbon nanotubes (CNTs) have been fabricated by a simple oxidative electropolymerization route. The nanotubular morphology of the polymer–CNT composite is responsible for the lower charge transfer impedance, lower internal resistance, and superior capacitive response in comparison to that shown by the control PEDOT film doped by trifluoromethanesulfonate ions. This facile electrochemistry exhibited by the PEDOT–CNT composite film ensues in a remarkably high coloration efficiency of 367 cm² · C⁻¹ at 550 nm, hitherto unrealized for PEDOT; thus demonstrating the huge potential the PEDOT–CNT composite film has as cathode for the entire spectrum of electrochromic devices.

     

Chiral ionic liquid-catalyzed Biginelli reaction: stereoselective synthesis of polyfunctionalized perhydropyrimidines

A chiral ionic liquid-catalyzed, efficient and unprecedented version of the Biginelli reaction using new variants of its active methylene component, viz. 2-phenyl-1,3-oxazol-5-one/2-methyl-2-phenyl-1,3-oxathiolan-5-one, with aromatic aldehydes and urea/thiourea enantio- and diastereoselectively, yields 5-amino-/mercaptoperhydropyrimidines. This three-component domino cyclocondensation reaction is effected via ring transformation of an isolable intermediate in a one-pot procedure.     

Ionic liquid [Hmim]HSO4-promoted one-pot oxidative conjugate addition of sulfur-centred nucleophiles to Baylis-Hillman adducts

The first example of the one-pot oxidative conjugate addition of sulfur-centred nucleophiles to Baylis-Hillman adducts is reported. The reaction involves oxidation of Baylis-Hillman adducts with NaNO₃ in the Brønsted acidic ionic liquid [Hmim]HSO₄ to give [E]-α-cyanocinnamaldehydes followed by conjugate hydrothiocyanation/hydrosulfenylation with NH₄SCN/PhSH to afford the corresponding β-thiocyanato (or β-phenylsulfenyl)-α-cyanohydrocinnamaldehydes diastereoselectively in 76-89% yields in a one-pot procedure. After isolation of the product, the ionic liquid [Hmim]HSO₄ could be easily recycled for further use.     

Synergistic extraction and spectrophotometric determination of palladium(II), iron(III), and tellurium(IV) at trace level by newly synthesized p-[4-(3,5-dimethylisoxazolyl)azophenylazo]calix(4)arene

A spectrophotometric method for the determination of palladium, iron and tellurium from nitric acid media after extraction of their p-[4-(3,5-dimethylisoxazolyl)azophenylazo]calix(4)arene [DMIAPAC] complexes has been developed and possible synergistic effects have been investigated. Chloroform, carbon tetrachloride, cyclohexane, 1,2-dichloroethane, toluene and xylene were used as the diluents. The maximum enhancement was obtained in the presence of 30% 1,2-dichloroethane. The trace amounts of metals were determined spectrophotometrically. Beer’s law obeyed in the concentration range of 5.0–95.0 μg, 8.0–120.0 μg and 10.0–140.0 μg/10 mL of the final solution of palladium, iron and tellurium, respectively. The molar absorptivities (l mol^?1cm^?1) and Sandell’s sensitivities (μg cm ^?1) were calculated: Pd(II) = 1.73 × 10⁴ and 0.0061; Fe(III) = 1.08 × 10⁴ and 0.0052; Te(IV) = 1.67 × 10⁴ and 0.0077. Ten replicate analyses containing 20 μg of Pd(II), 12.5 μg of Fe(III) and 32 μg of Te(IV) gave mean absorbance of 0.326, 0.242 and 0.418 with relative standard deviation of 0.36, 0.65 and 0.82% for Pd(II), Fe(III) and Te(IV), respectively. The interference of various ions was studied and optimum conditions were developed for the determination of these metals in certain alloys and synthetic mixtures.     

Determination of plutonium isotopes in urine samples from radiation workers using <Superscript>236</Superscript>Pu tracer,anion exchange resin and alpha spectrometry

Bioassay technique is used for the estimation of actinides present in the body based on their excretion rate through body fluids. For occupational radiation workers urine assay is the preferred method for monitoring of chronic internal exposure. Determination of low concentrations of actinides such as plutonium, americium and uranium at low level of mBq in urine by alpha spectrometry requires pre-concentration of large volumes of urine. This paper deals with standardization of analytical method for the determination of Pu-isotopes in urine samples using anion exchange resin and ²³⁶Pu tracer for radiochemical recovery. The method involves oxidation of urine followed by co-precipitation of plutonium along with calcium phosphate. Separation of Pu was carried out by Amberlite, IRA-400, anion exchange resin. Pu-fraction was electrodeposited and activity estimated using tracer recovery by alpha spectrometer. Twenty routine urine samples of radiation workers were analyzed and consistent radiochemical tracer recovery was obtained in the range 74–96% with a mean and standard deviation of 85 and 6% respectively.     

Phytochemical investigation and antifeedant activity of Premna latifolia leaves

The essential oil of Premna latifolia Roxb. was obtained by hydrodistillation of fresh leaves of the plant having an oil yield of 0.05%, both non-polar and essential oil were analysed by GC and GC-MS. Hexane fraction of the leaves of P. latifolia was transesterified and analysed by GC and GC-MS, 40 non-polar components were identified comprising 89.3%. The most abundant fatty acid constituents were hexadecanoic acid (25.04%), 8,11,14-docosatrienoic acid (13.62%), octadecanoic acid (6.82%), 9,12-octadecadienoic acid (4.19%) and 29 components were investigated in the essential oil which comprises 78.1%. The most abundant oil constituents were 1-octen-3-ol (35.69%), terpendiol II (7.19%), δ-guaiene (7.49%) 2-undecanone (4.80%) and α-pinene (3.27%). Different extracts were also tested against polyphagous crop pest Spodoptera litura for antifeedant activity. Essential oil showed maximum growth reduction of 56.83% followed by chloroform extract of 43.93%.     

Miscellaneous applications of ferrocene‐based peptides/amides

This article provides a critical review of the different applications of ferrocene‐based peptides/amides in biological as well as in non‐biological systems. Ferrocene‐based peptides/amides find many applications in different fields such as materials science, medicine, organic synthesis, bio‐organometallic and biological chemistry, asymmetric catalysis, nonlinear optics, in polymer science as redox active polymers and dendrimers, in molecular recognition as biosensors and in electrochemistry). Extensive research is being done on ferrocene‐based peptides/amides but we will highlight the various applications of ferrocene‐based peptides/amides for the period 2006–2010. The main factors that govern the potential biological and non‐biological applications are an electroactive core, a conjugated linker that can act as a chromophore and lower the oxidation potential of the ferrocene part, an amino acid or peptide derivative that can interact with other molecules via hydrogen bonding or any secondary bonding, and symmetric and asymmetric substitution on the ferrocene moiety. Copyright © 2011 John Wiley & Sons, Ltd.     

The Mechanism of Thionyl Chloride Reaction with Dialkyl Alkylphosphonothionate Using 31 P NMR

Based on 31 P NMR studies of thionyl chloride reaction with dialkyl alkylphosphonothionates, a method for preparation of alkylphosphonic dichloride has been investigated. A mechanism via intermediacy of ester chloride is suggested.