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In the present study, the pyrolysis behaviour of Capsicum Annuum stem Waste (CAW) was studied applying thermogravimetric analysis to evaluate the intrinsic kinetic parameters to develop a pyrolysis reactor for utilizing CAW. The thermal decomposition of CAW was achieved between 373 – 1173 K under inert conditions at different heating rates of 10, 20, and 30 K min-1. Model-free kinetic methods like Kissinger-Akahira-Sunose (KAS), Ozawa Flynn Wall (OFW) and Coats Redfern methods were applied to work out the kinetic parameters. To identify the utility of CAW and its biochar, physio-chemical characteristics such as proximate and ultimate analysis, scanning electron microscopy, and Fourier Transform Infrared analysis are reported.

Plastic is a material that is used to a great extent. Nowadays Petroleum based plastics are widely used for the commercial purpose. This kind of plastics can take more than a century to degrade. Man made plastics have Polyurethane and Polyethylene polymers. It cannot be easily degraded by micro organisms as it don’t recognize as food. When burned plastics release cancer causing carcinogenic chemicals that are harmful to people and the environment. One eco friendly alternative the current commercial plastic is “Bio plastics”. Bio plastics are biodegradable plastics that have components derived directly from renewable raw plant materials. Seven type of bioplastics were made using various combinations of renewable raw materials and plasticizers. In this project, Cassava starch is used as a main biopolymer for the production of bio degradable bags.

Background: Acute phase insulin secretion is disturbed in individuals with prediabetes. We have previously shown a significant risk reduction in 4,417 prediabetes individuals through Yoga. The present study explores yoga’s mechanism of action. Method: This randomized control trial included 42 (20 yoga;22 walkings;5 days/ week, 3Months) prediabetes (fasting glucose of 100 to 125 mg/dl) adults between 23 to 50 years. Serum Insulin and C peptide were assessed at 0, 3, 5, 7, 10, 15, 30,60and 90th minutes after 75grams of oral glucose. Pre-post assessments included fasting Glucose, HOMA%B(derived), GLP-1, free Triiodothyronine, perceived stress, and Heart rate variability. Results: There was a significant increase in the 10th minute (acute phase) Insulin and c-peptide levels in the yoga as compared to the control group. Improvement in GLP-1 and HOMA%B was significantly better in yoga than walking group. Yoga group also showed positive change in perceived stress and HRV. Simple linear regression of post data using acute phase Insulin as the dependent variable indicated a significant association between all measures(R2 = 0.806; p = Conclusion: The shift to normoglycemia, due to an increase in acute phase insulin release, was associated with improved autonomic balance and reduced stress after yoga. More studies may confirm this unique first time observation. Disclosure R. Nagarathna: None. J. Kurian: None. V. Vijayakumar: None. H. R. Nagendra: None. R. N. Mavathur: None. Funding Ministry of AYUSH, Government of India (Z.28015-HPC (EMR)-AYUSH-B)

Chirality is a key factor in the safety and efficacy of many drug products and thus the production of single enantiomers of drug intermediates and drugs has become important and state of the art in the pharmaceutical industry. There has been an increasing awareness of the enormous potential of microorganisms and enzymes (biocatalysts) for the transformation of synthetic chemicals with high chemo-, regio- and enatioselectivities providing products in high yields and purity. In this article, biocatalytic processes are described for the synthesis of key chiral intermediates for development pharmaceuticals.

The production of single enantiomers of key chiral intermediates has become increasingly important for the development of drugs. Demand for chiral compounds continues to rise in the pharmaceutical industry due to federal regulations covering active pharmaceutical ingredients (APIs), and the recognition that enantiomers of a chiral compound could have dramatically different biological activities and toxicity. In this area, there are a number of advantages in using biocatalysis over chemical synthesis. Typically, enzyme-catalyzed reactions are highly enantioselective, regioselective and do not require protection and deprotection steps during synthesis. They can be carried out at ambient temperature, atmospheric pressure in aqueous systems, and are often regarded as green processes that avoid the use of more extreme conditions used in chemical reactions which could cause undesirable side reactions. Microbial cells and enzymes (biocatalysts) can be immobilized and reused for many cycles, and enzymes are generally over-expressed in suitable hosts such as Escherichia coli to make enzymatic processes economically efficient, safer and cheaper. The preparation of thermostable, pH stable, organic solvent tolerant and highly active biocatalysts towards substrates by random and site-directed mutagenesis by directed evolution has led to the production of novel biocatalysts to be used under desired process conditions. This chapter describes the synthesis of key intermediates for the synthesis of APIS by biocatalysis at Bristol-Myers Squibb to provide sustainable, energy efficient and economical processes.

To efficiently utilize agricultural biomass waste, kinetic modeling of the pyrolysis of rice husk, including both physical (mainly heat transfer) and chemical (reactions) terms, was conducted at different heating rates from (10 to 20 K min−1) to develop a transport model. For chemical kinetics, the parameters were estimated using different kinetic models, namely the single- or parallel-reaction kinetic model with higher orders and the two-step consecutive reaction model. The two-step model could adequately explain the pyrolysis reaction of multiple reactions with different reaction orders i. e., first step is of the first order (m = 1) with respect to the mass of biomass, and the second step is of the second order (n = 2) with respect to the mass of the intermediate to char. The intrinsic kinetics at different heating rates in the absence of oxygen was derived through thermogravimetric analysis. The kinetics of the evolution of non-condensable gases was studied in a self-designed reactor, and an appropriate kinetic model of rice husk biomass pyrolysis that showed excellent agreement with experimental data was established.

An enzymatic reduction process was developed to convert the ketone 2-(6-oxo-2-phenyladamantan-2-yl)acetic acid to the chiral alcohol 2-((2S, 6S)-6-hydroxy-2-phenyladamantan-2-yl)acetic acid and to ...

Chirality is a key factor in the safety and efficacy of many drug products and thus the production of single enantiomers of drug intermediates and drugs has become increasingly important in the pharmaceutical industry. There has been an increasing awareness of the enormous potential of microorganisms and enzymes derived there from for the transformation of synthetic chemicals with high chemo-, regio- and enatioselectivities. In this article, biocatalytic processes are described for the synthesis of chiral alcohols and unntural aminoacids for pharmaceuticals.

Biocatalytic processes to selectively hydrolyze the N-carbobenzyloxy (CBz) group from CBz-protected d - or l -amino acids have been developed. The substrate specificities of the CBz-deprotecting enzymes from Burkholderia phenazinium SC 16530, and Sphingomonas paucimobilis expressed into Escherichia coli SC 16501 were evaluated on CBz-protected amino acids and structurally related compounds. Modifications of various structural components and their effects on enzyme activity and enantioselectivity provided a greater understanding of the two CBz-deprotecting enzymes.

Gasification of four Indian coals is carried out in a CO2 atmosphere, using a thermogravimetric analyzer (TGA) to determine the intrinsic kinetics over a temperature range of 800-1050 degrees C with different partial pressures of CO2. The applicability of three models, viz., the volumetric reaction model, the shrinking core model and the random pore model, is evaluated. Of these three models, the random pore model is found to be the most suitable for all the coals considered in the current study. The dependence of the reaction rate on the gas-phase partial pressures is explained by the Langmuir-Hinshelwood model, and the parameters for the inhibition due to CO and CO2 are determined by performing experiments at different partial pressures. In underground coal gasification, the reaction takes place on reasonably large sized coal particles, wherein diffusion effects are significant A one-dimensional reaction diffusion model is therefore developed in order to determine the diffusional resistance in the coal particle, and values of diffusivity are estimated.

Enzymes are widely used for peptide synthesis, both for coupling and for terminal activation and modification. Broad-specificity proteases of different mechanistic classes can be used for C-terminal activation and peptide coupling reactions, whereas enzymes that recognize specific sequences are increasingly used for selective modification and tagging. Protein engineering can tailor hydrolases for improved synthesis, better reaction medium tolerance, and desired substrate selectivity. This chapter highlights properties of natural and engineered peptidases, amidases, and peptide transferases for chemoenzymatic peptide synthesis and terminal modification, with an emphasis on basic principles and innovations leading to new applications.

The synthesis of optically active sulfoxides, compounds due to their unique properties, has been a main target for synthetic organic chemistry. Recent efforts in the field of biocatalysis have allowed the preparation of enantiopure sulfoxides starting from the corresponding sulfides while using relatively mild conditions. In fact, several different types of redox biocatalysts have been found that can catalyze enantio- and/or regioselective sulfoxidations. The most promising group of enzymes able to perform selective sulfoxidations is the flavin-containing monooxygenases (FMOs). Enzymes containing a flavin cofactor have already been widely studied and used in organic synthesis, especially in reduction and/or oxidation processes. This chapter highlights the recent efforts in the preparation of chiral sulfoxides catalyzed by different types of flavoenzymes, with special attention to the parameters that can improve their catalytic properties. Novel approaches that allow performing selective sulfoxidations in which modified flavin systems are used are also discussed.

Chirality is a key factor in the safety and efficacy of many drug products and thus producing single enantiomers of drug intermediates and drugs has become increasingly important in the pharmaceutical industry. There has been an increasing awareness of the enormous potential of microorganisms and enzymes derived from them for the transformation of synthetic chemicals with high chemo-, regio-, and enatioselectivities. In this article, biocatalytic processes are described for the synthesis of key chiral intermediates for development of pharmaceuticals and fine chemicals.

Chirality is a key factor for the safety and efficacy of many drug products. The production of single enantiomers of drug intermediates has become increasingly important in the pharmaceutical industry. There has been an enormous potential of microorganisms and enzymes derived from there for the transformation of synthetic chemicals with high chemo-, regio-, and enatioselectivities. Recent development in the area of directed evolution has led screen mutants under process conditions to increase activity and selectivity of biocatalysts, thus making the enzymatic process highly efficient and economically feasible. In this review, chemoenzymatic processes are described for the synthesis of chiral intermediates for the development of pharmaceuticals.

Systematic laboratory scale experiments on coal blocks can provide significant insight into the underground coal gasification (UCG) process. Our earlier work has demonstrated the various features of the early UCG cavity shape and rate of growth through lab-scale experiments on coal combustion, wherein the feed gas is oxygen. In this paper, we study the feasibility of in situ gasification of coal in a similar laboratory scale reactor set-up, under conditions relevant for field practice of UCG, using an oxygen-steam mixture as the feed gas. By performing the gasification reaction in a cyclic manner, we have been able to obtain a product gas with hydrogen concentrations as high as 39% and a calorific value of 178 kJ/mol. The effect of various operating parameters such as feed temperature, feed steam to oxygen ratio, initial combustion time and so on, on the product gas composition is studied and the optimum operating conditions in order to achieve desired conversion to syngas, are determined. We also study the effect of various design and operating parameters on the evolution of the gasification cavity. Empirical correlations are proposed for the change in cavity volume and its dimensions in various directions. The results of the previous study on the combustion cavity evolution are compared with this gasification study. (C) 2011 Elsevier Ltd. All rights reserved.

Polypyrrole (PPy) was synthesized by chemical oxidative polymerization technique using monomer pyrrole and ammo-nium persulphate as an oxidant in a ratio of 1:1. Thin films of polypyrrole were prepared by dissolving polypyrrole in mcresol and cast using spin coating technique on glass substrates. Thin films of polypyrrole were characterized by X ray diffraction (XRD), Scanning electron microscopy (SEM), Fourier transform infra red spectroscopy (FTIR), UV visible spectroscopy and electrical resistivity by four probe method. The XRD spectra showed that the polypyrrole is amorphous in nature. SEM studies revealed a uniform granular structure of PPy. The FTIR spectra shows that the presence of all characteristics absorption peaks of polypyrrole that is, 890 cm?1 (=C–H out-of-plane vibration), 1040 cm?1 (=C–H in-plane vibration), 1170 cm?1 (N–C stretch bending) and 1410 cm?1 (pyrrole ring vibration). UV visible study shows PPy films exhibit absorption peak at 446 nm (2.77 eV). Room temperature electrical resistivity of PPy is in the critical regime of the metal –to –insulator (M-I) transition.

During underground coal gasification (UCG), a cavity is formed in the coal seam when coal is converted to gaseous products. This cavity grows three dimensionally in a nonlinear fashion as gasification proceeds. The cavity shape is determined by the flow field, which is a strong function of various parameters such as the position and orientation of the inlet nozzle and the temperature distribution and coal properties such as thermal conductivity. In addition to the complex flow patterns in the UCG cavity, several phenomena occur simultaneously. They include chemical reactions (both homogeneous and heterogeneous), water influx, thermomechanical failure of the coal, heat and mass transfer, and so on. Thus, enormous computational efforts are required to simulate the performance of UCG through a mathematical model. It is therefore necessary to simplify the modeling approach for relatively quick but reliable predictions for application in process design and optimization. The primary objective of this work is to understand the velocity distribution and quantify the nonideal flow patterns in a UCG cavity by performing residence time distribution (RTD) studies using computational fluid dynamics (CFD). The methodology of obtaining RTD by CFD is validated by means of of representative laboratory-scale tracer experiments. Based on the RTD studies, the actual UCG cavity at different times is modeled as a simplified network of ideal reactors, called compartments. The compartment model thus obtained could offer a computationally less expensive and easier option for determining UCG process performance at any given time, when used in a reactor-scale model including reactions. The network of ideal reactors can be easily simulated using a flowsheet simulator (e.g., Aspen Plus). We illustrate the proposed modeling approach by presenting selected simulation results for a single gas-phase second-order water gas shift reaction.

The (S)-amino acid, (S)-2-amino-3-(6-o-tolylpyridin-3-yl)propanoic acid (3), is a key intermediate needed for synthesis of an antidiabetic drug candidate. Three enzymatic routes to 3 were explored. (S)-Amino acid 3 could be prepared in 73% isolated yield with 99.9% ee from racemic amino acid 1 using (R)-amino acid oxidase from Trigonopsis variabilis expressed in Escherichia coli in combination with an (S)-aminotransferase using (S)-aspartate as amino donor. The (S)-aminotransferase was purified from a soil organism identified as Burkholderia sp. and cloned and expressed in E. coli. (S)-Amino acid 3 with 100% ee was also prepared in 68% solution yield and 54% isolated yield from 1 using recombinant (R)-amino acid oxidase from T. variabilis and an (S)-amino acid dehydrogenase from Sporosarcina ureae. The cofactor NADH required for the reductive amination reaction was regenerated using formate and formate dehydrogenase. The chemoenzymatic dynamic resolution of 1 by (R)-selective oxidation with Celite-immobil...

Cavity formation is an important phenomenon in the underground coal gasification (UCG) process. In the early stages of cavity formation, only the combustion reaction is performed in order to stabilize the temperature field. In the present work, we study the formation of the combustion cavity and the effect of various design and operating parameters such as the distance between the wells, feed flow rate and operation time, on its evolution. This paper presents details of laboratory-scale experiments that demonstrate the shape and size of the combustion cavity, and their dependence on various parameters. Empirical correlations for the cavity volume and dimensions in various directions are developed, which indicate the strong effects of mass transport. Results from computational fluid dynamics (CFD), which map the velocity profiles in the cavity, support the experimental observations.

A synthetic gene encoding a Streptomyces l -proline-3-hydroxylase was constructed and used to produce the hydroxylase protein in recombinant Escherichia coli. A fermentation process for growth of this recombinant E. coli for enzyme production was scaled-up to 250 L. A biotransformation process was developed using cell suspensions of the recombinant E. coli and subsequently scaled-up to 10 L for conversion of l -proline to cis-3-hydroxy- l -proline. A reaction yield of 85 M% and d.e. of 99.9% was obtained for cis-3-hydroxy- l -proline.

The d-amino acid (R)-2-amino-3-(7-methyl-1 H-indazol-5-yl)propanoic acid (3) is a key intermediate needed for synthesis of a drug candidate compound. Enzymatic routes to 3 were explored. d-Amino acid 3 was prepared in 68% isolated yield with >99% ee from racemic amino acid 1 using l-amino acid deaminase from Proteus mirabilis expressed in Escherichia coli in combination with a commercially available d-transaminase using d-alanine as amino donor. The d-enantiomer was also prepared in 79% isolated yield with >99% ee from the corresponding keto acid 2 using the d-transaminase with racemic alanine as the amino donor. The rate and yield of this reaction could be accelerated by addition of lactate dehydrogenase (with NAD, formate and formate dehydrogenase to regenerate NADH) to remove the inhibitory pyruvate produced during the reaction. A d-transaminase was purified from a soil organism identified as Bacillus thuringiensis and cloned and expressed in E. coli. The d-transaminase was very effective for the prepa...

Screening was carried out to identify strains useful for the preparation of (R)-1-cyclopropylethylamine and (R)-sec-butylamine by resolution of the racemic amines with an (S)-specific transaminase. Several Bacillus megaterium strains from our culture collection as well as several soil isolates were found to have the desired activity for the resolution of the racemic amines to give the (R)-enantiomers. Using an extract of the best strain, Bacillus megaterium SC6394, the reaction was shown to be a transamination requiring pyruvate as amino acceptor and pyridoxal phosphate as a cofactor. Initial batches of both amines were produced using whole cells of Bacillus megaterium SC6394. The transaminase was purified to homogeneity to obtain N-terminal as well as internal amino acid sequences. The sequences were used to design polymerase chain reaction (PCR) primers to enable cloning and expression of the transaminase in E. coli SC16578. In contrast to the original B. megaterium process, pH control and aeration were not required for the resolution of sec-butylamine and an excess of pyruvate was not consumed by the recombinant cells. The resolution of sec-butylamine (0.68 M) using whole cells of E. coli SC16578 was scaled up to give (R)-sec-butylamine⋅1/2 H2SO4 in 46.6% isolated yield with 99.2% ee. An alternative isolation procedure was also used to isolate (R)-sec-butylamine as the free base.

Chiral intermediates were prepared by biocatalytic processes for the chemical synthesis of four pharmaceutical drug candidates. These include: (i) the microbial reduction of 3,5-dioxo-6-(benzyloxy) hexanoic ethyl ester to (3S,5R)-dihydroxy-6-(benzyloxy) hexanoic acid ethyl ester, an intermediate for a new anticholesterol drug; (ii) synthesis of (2R,3S)-(-)-N-benzoyl-3-phenyl isoserine ethyl ester, a taxol side-chain synthon; (iii) the microbial oxygenation of 2,2-dimethyl-2H-1-benzopyran-6-carbonitrile to the corresponding (3S,4S) epoxide and (3S,4R)-trans diol, intermediates for synthesis of potassium channel opener; (iv) the biotransformation of (exo,exo)-7-oxabicyclo [2.2.1] heptane-2,3-dimethanol to the corresponding chiral lactol and lactone, intermediates for thromboxane A2 antagonist.

Chirality is a key factor in the safety and efficacy of many drug products and, thus, the production of single enantiomers of drug intermediates has become increasingly important in the pharmaceutical industry. There has been an increasing awareness of the enormous potential of microorganisms and enzymes derived therefrom for the transformation of synthetic chemicals with high chemo-, regio- and enantioselectivities. In this article, some selected enzymatic processes are described for the synthesis of chiral intermediates for pharmaceuticals.

Protection against a high dose of a toxicant by prior exposure to another toxicant is called heteroprotection. Our objective was to establish a heteroprotection model in RBCs. Female Sprague Dawley rats treated with an LD90 dose of 2-butoxyethanol (BE, 1500 mg/kg in water, 5 ml/kg po) 14 days after priming with 0.9% NaCl suffered 90% mortality by 15 days, whereas all rats receiving the LD90 dose of BE 14 days after priming with phenylhydrazine (PHZ, 125 mg/kg in 0.9% NaCl, 3 ml/kg po) survived. Hematocrit decreased from normal 45% to 24% by day 3 after PHZ priming and improved thereafter. Increasing the time interval between the priming and LD90 dose to 21 days abolished the heteroprotection. RBCs obtained on days 7 and 14 after PHZ priming unlike those on day 21 were resilient to the hemotoxic metabolite of BE, butoxyacetic acid (BAA). Unaltered hepatic alcohol and aldehyde dehydrogenase activities upon PHZ priming suggested that bioactivation of BE to BAA was unaffected. Lower renal (6 and 12 h) and hepatic (12 h) BAA levels and 3 fold higher excretion of BAA in PHZ-primed rat urine suggested a protective role of toxicokinetics. Higher erythropoietin, reticulocytes, and resiliency of PHZ-primed rat RBCs indicated that newly formed RBCs are resilient to hemolytic BAA. The antioxidant levels in the PHZ-primed rat RBCs did not indicate a protective role in heteroprotection. In conclusion, the resistance of PHZ-primed rats against BE-induced hemotoxicity and lethality is mediated by a combination of altered toxicokinetics, robust erythropoiesis, and resiliency of new RBCs.

An efficient biocatalytic process has been developed for the resolution of methyl (1 RS )- N - t Boc-6-hydroxy-3,4-dihydro-1 H -isoquinoline-1-carboxylate rac - 1 by means of Seaprose S-mediated enantioselective hydrolysis to afford (1 S )- 2 and (1 R )- 1 in 87% and 93% isolated yield, 101% and 96% potency, and ee >99.8% and >99.5%, respectively.

The non-proteinogenic amino acid 2-(3-hydroxy-1-adamantyl)-(2S)-aminoethanoic acid [2, (S)-3-hydroxyadamantylglycine], is a key intermediate required for the synthesis of Saxagliptin, a dipeptidyl peptidase IV inhibitor under development for treatment of type 2 diabetes mellitus. Keto acid 2-(3-hydroxy-1-adamantyl)-2-oxoethanoic acid (1) was converted to (S)-3-hydroxyadamantylglycine by reductive amination using a phenylalanine dehydrogenase from Thermoactinomyces intermedius expressed in a modified form in Pichia pastoris or Escherichia coli. NAD (nicotinamide adenine dinucleotide) produced during the reaction was recycled to NADH (reduced form of nicotinamide adenine dinucleotide) using formate dehydrogenase. Pichia pastoris produces an endogenous formate dehydrogenase when grown on methanol, and the corresponding gene was cloned and expressed in E. coli. The modified phenylalanine dehydrogenase contains two amino acid changes at the C-terminus and a 12-amino acid extension of the C-terminus. The modified enzyme is more effective with keto acid 1 than the wild-type enzyme, but less effective with the natural substrate, phenylpyruvate. Production of multi-kg batches was originally carried out with extracts of Pichia pastoris expressing the modified phenylalanine dehydrogenase from Thermoactinomyces intermedius and endogenous formate dehydrogenase, and further scaled up using a preparation of the two enzymes expressed in E. coli.

An efficient biocatalytic process has been developed for the resolution of 1-(3′-bromophenyl)ethylamine ( RS )- 1 by way of enantioselective lipase-mediated ( R )-selective acylation with ethyl 2-methoxyacetate to afford ( S )-amine ( S )- 1 and ( R )-2″-methoxyacetamide (( R )- 2 ) in 91–95% and 90–92% isolated yield, respectively, and both with >99% ee.

Many of the familiar relations in the theory of linear viscoelasticity which relate the relaxation spectrum H(τ) to the experimentally measured quantities were extended to embrace the concept of shear dependent relaxation spectrum H(τ, ). The concept is based on the relation H(τ, ) = H(τ)h(θ)g(θ)3/2, where θ = c τ and c = 0.5. Here is the shear rate and τ is the relaxation time. The replacement of H(τ) by H(τ',) in the integral relations of the linear theory produced the corresponding nonlinear parameters, e.g., the shear rate dependent viscosity η(), the shear stress relaxation following cessation of steady-state flow σ(,t) and the dynamic parameters under parallel superimposed rotation η'(ω) and G'(ω, ). The concept was used successfully for several high-density polyethylene melts in interconversion between η(), c(,t) and the dynamic viscosity η'(ω) without involving coordinate shifts or empirical parameters. The failure of this approach for interconversion between η(ω) and η() for the highly branched polyethylenes was ascribed to the level of long-chain branching. For the high-density resins, the agreement between measured η'(ω,) and G'(ω,) and those calculated using H(τ,) was not quantitative since the parallel superimposed experiments require an additional term in ∂H(τ,)/∂ inside the integral.

The enantioselective microbial reduction of 6-oxo-8-[4-[4-(2-pyrimidinyl)-1-piperazinyl]butyl]-8-azaspiro[4.5]decane-7,9-dione (1) to either of the corresponding (S)- and (R)-6-hydroxy-8-[4-[4-(2-pyrimidinyl)-1-piperazinyl]butyl]-8-azaspiro[4.5]decane-7,9-diones (2 and 3, respectively) is described. The NADP+-dependent (R)-reductase (RHBR) which catalyzes the reduction of 6-ketobuspirone (1) to (R)-6-hydroxybuspirone (3) was purified to homogeneity from cell extracts of Hansenula polymorpha SC 13845. The subunit molecular weight of the enzyme is 35,000 kDa based on sodium dodecyl sulfate gel electrophoresis and the molecular weight of the enzyme is 37,000 kDa as estimated by gel filtration chromatography. (R)-reductase from H. polymorpha was cloned and expressed in Escherichia coli. To regenerate the cofactor NADPH required for reduction we have cloned and expressed the glucose-6-phosphate dehydrogenase gene from Saccharomyces cerevisiae in E. coli. The NAD+-dependent (S)-reductase (SHBR) which catalyzes the reduction of 6-ketobuspirone (1) to (S)-6-hydroxybuspirone (2) was purified to homogeneity from cell extracts of Pseudomonas putida SC 16269. The subunit molecular weight of the enzyme is 25,000 kDa based on sodium dodecyl sulfate gel electrophoresis. The (S)-reductase from P. putida was cloned and expressed in E. coli. To regenerate the cofactor NADH required for reduction we have cloned and expressed the formate dehydrogenase gene from Pichia pastoris in E. coli. Recombinant E. coli expressing (S)-reductase and (R)-reductase catalyzed the reduction of 1 to (S)-6-hyroxybuspirone (2) and (R)-6-hyroxybuspirone (3), respectively, in >98% yield and >99.9% e.e.

Microbial reduction of ethyl 1-benzyl-3-oxo-piperidine-4-carboxylate by the majority of evaluated microorganisms gave the ethyl cis-(3R,4R)-1-benzyl-3R-hydroxy-piperidine-4R-carboxylate as the major product in high diastereo- and enantioselectivities. The 3R,4R-hydroxy ester was produced in 97.4% diastereomeric excess (de) and 99.8% enantiomeric excess (ee) by Candida parapsilosis SC16347, while 99.5% de and 98.2% ee were obtained from reduction by Pichia methanolica SC16415. A few of the evaluated microorganisms gave 10–40% of the ethyl trans-(3R,4S)-1-benzyl-3R-hydroxy-piperidine-4S-carboxylate as the minor product.

Previously we have demonstrated the reduction of ethyl diketoester 4 to the corresponding dihydroxy ester 6a by Acinetobacter sp. SC13874. Recently we screened more than 100 cultures for microbial reduction of both the ethyl and t-butyl diketoesters 4 and 5. Most yeast cultures showed a preference for reduction at the C-3 with low enantioselectivity. Among the three Acinetobacter strains screened, Acinetobacter sp. SC13874 reduced both compounds 4 and 5 to the corresponding (3R)- and (5S)-monohydroxy compounds. Monohydroxy compounds were isolated and their absolute configurations determined. (3R)- and (5S)-Monohydroxy compounds were reduced further to the corresponding dihydroxy esters 6a and 8a to provide alternate routes for the synthesis of compounds 14a and 16a, potential intermediates for the synthesis of HMG-CoA reductase inhibitors. Cell suspensions of Acinetobacter sp. SC13874 reduced the ethyl diketoester 4 to a mixture of desired syn and undesired anti diastereomers. The desired syn-(3R,5S)-dihydroxy ester 6a was obtained with an enantiomeric excess (ee) of 99% and a diastereomeric excess (de) of 63%. Cell suspensions reduced the t-butyl diketoester 5 to a mixture of mono- and dihydroxy esters with the dihydroxy ester showing an ee of 87% and de of 51% for the desired syn-(3R,5S)-dihydroxy ester 8a. Three different ketoreductases were purified to homogeneity, and their biochemical properties compared. Reductase I only catalyzes the reduction of ethyl diketoester 4 to its monohydroxy products 10 and 11, whereas reductase II catalyzes the formation of dihydroxy products 6 and 7 from monohydroxy substrates 10 and 11. A third reductase (III) was identified, which catalyzes the reduction of diketoester 4 to syn-(3R,5S)-dihydroxy ester 6a.

Objective of the present study was to test the importance of tissue repair in the final outcome of S-(1,2-dichlorovinyl)-L-cysteine (DCVC)-induced nephrotoxicity using colchicine (CLC) intervention. Male Swiss Webster (SW) mice were administered a normally nonlethal dose of DCVC (30 mg/kg, i.p.) on day 0 and CLC (2 mg/kg, i.p.) at 42 and 66 h after administration of DCVC. The mice were observed for mortality and various renal injury and repair parameters were studied during a time course of 0-14 days. Administration of 30 mg DCVC/kg led to loss of renal architecture by day 1, which sustained until day 5, and regressed thereafter to reach normal architecture by day 10 resulting in 100% survival. Renal dysfunction as assessed by increases in plasma BUN and creatinine levels was concordant during this time course. Urinary volume increased significantly between days 10 and 14 with significant increases in urinary glucose concentrations on days 1-4. Calpain leakage increased from day 1 and remained so until day 5 before declining at later time points. In contrast, CLC intervention led to marked inhibition of S-phase DNA synthesis and 100% mortality by 120 h. H&E sections of kidneys revealed loss of renal architecture on day 1 which progressively worsened from day 2 to 4. Polyuria and glycosuria were evident during the first 2 and 3 days, respectively. Calpain immunohistochemistry revealed progressive leakage of calpain in the extracellular space during 2-4 days which lead to increased renal injury as evident from significant increases in calpain specific breakdown products (CSBPs) of alpha-fodrin during the same period of time. The group of mice receiving 2 mg CLC/kg alone showed a significant increase in urinary creatinine concentration on day 5. Neither the expression nor localization of aquaporin 1 was altered in any of the treatment groups. These results show that antimitotic intervention after DCVC-initiated renal injury leads to expansion and progression of that injury, which appears to be due to proteolytic destruction of neighboring cells mediated by calpain leaking out of necrosed renal tubular epithelial cells.

An efficient biocatalytic method has been developed for the conversion of (5S)-4,5-dihydro-1H-pyrrole-1,5-dicarboxylic acid, 1-(1,1-dimethylethyl)-5-ethyl ester (1) into the corresponding amide (5S)-5-aminocarbonyl-4,5-dihydro-1H-pyrrole-1-carboxylic acid, 1-(1,1-dimethylethyl)ester (2), which is a critical intermediate in the synthesis of the dipeptidyl peptidase IV (DPP4) inhibitor Saxagliptin (3). Candida antartica lipase B mediates ammonolysis of the ester with ammonium carbamate as ammonia donor to yield up to 71% of the amide. The inclusion of Ascarite and calcium chloride as adsorbents for carbon dioxide and ethanol byproducts, respectively, increases the yield to 98%, thereby offering an efficient and practical alternative to chemical routes which yield 57-64%.

Enantioselective asymmetric hydrolysis of (1α,2β,3α)-2-(benzyloxymethyl)-cyclopent-4-ene-1,3-diol diacetate 1 to the corresponding (+)-monoacetate 2 was carried out using lipase PS-30 from Pseudomonas cepacia or pancreatin. A reaction yield of 85 M % with an enantiomeric excess (ee) of 98% was obtained. Using pancreatin, a reaction yield of 75 M % with an ee of 98.5% was obtained. Asymmetric acetylation of (1α,2β,3α)-2-(benzyloxymethyl)-cyclopent-4-ene-1,3-diol 3 to the corresponding (−)-monoacetate 4 was carried out using lipase PS-30 with isopropenyl acetate as the acylating agent. A reaction yield of 80 M % with an ee of 98% was obtained for (−)-monoacetate 4 .

Electrospray ionization (ESI) quadrupole ion trap mass spectrometry (QIT-MS) and collisionally activated dissociation (CAD) were used to evaluate the rare-earth binding properties of two hydrophobic carbamoylmethylphosphine oxide (CMPO) ligands, the normal bidentate variety, (t-BuC6H4)2P(O)CH2C(O)N(i-Bu)2 (A), a new potentially tridentate extractant, (t-BuC6H4)2P(O)CH[CH2C(O)N(i-Bu)2]C(O)N(i-Bu)2 (B), and tributyl phosphate. The mass spectral results obtained from analysis of 1% HNO3/methanol solution containing the ligands and dissolved lanthanide salts reveal that the favorable stoichiometries of the ligand/metal/nitrate complexes are 2:1:2 for the bidentate ligand A, 1:1:2 for the tridentate ligand B, and 3:1:2 for the monodentate tributyl phosphate. These observed stoichiometries correlate with the number of available binding sites on each ligand as well as with potential steric effects. Energy-variable collisionally activated dissociation experiments showed that for the 2:1:2 complexes involving ligand A or B, as the ionic radius of the bound metal decreased, the removal of nitric acid required less energy and resulted in less extensive spontaneous solvent coordination. This experimental trend suggests that, as the ionic radius of the lanthanide ion decreases, a pair of the carbamoylmethylphosphine ligands is able to more completely solvate the bound metal ion thereby weakening the nitrate-metal interaction.

The enantioselective microbial reduction of 6-oxo-8-[4-[4-(2-pyrimidinyl)-1-piperazinyl]butyl]-8-azaspiro[4.5]decane-7,9-dione 1 to either of the corresponding (R)- or (S)-6-hydroxy-8-[4-[4-(2-pyrimidinyl)-1-piperazinyl]butyl]-8-azaspiro[4.5]decane-7,9-diones 2 and 3 is described.

6-Hydroxybuspirone is an active metabolite of the antianxiety drug buspirone. The (R)- and (S)-enantiomers of 6-hydroxybuspirone were prepared using an enzymatic resolution process. l -Amino acid acylase from Aspergillus melleus (Amano Acylase 30000) was used to hydrolyze racemic 6-acetoxybuspirone to (S)-6-hydroxybuspirone in 95% ee after 45% conversion. The remaining (R)-6-acetoxybuspirone with 88% ee was converted to (R)-6-hydroxybuspirone by acid hydrolysis. The ee of both enantiomers could be improved to 99% by crystallization as a metastable polymorph. (S)-6-Hydroxybuspirone was also obtained in 88% ee and 14.5% yield by hydroxylation of buspirone using Streptomyces antibioticus ATCC 14890.

The synthesis of the leading candidate compound in an anticancer program required (S)-2-chloro-1-(3-chlorophenyl)-ethanol as an intermediate. Other possible candidate compounds used analogues of the S-alcohol. Of 119 microbial cultures screened for reduction of the corresponding ketone to the S-alcohol, Hansenula polymorpha ATCC 58401 (73.8% ee) and Rhodococcus globerulus ATCC 21505 (71.8% ee) had the highest enantioselectivity for producing the desired alcohol. A ketoreductase from Hansenula polymorpha, after purification to homogeneity, gave the S-alcohol with 100% ee. Amino acid sequences from the purified enzyme were used to design PCR primers for cloning the ketoreductase. The cloned ketoreductase required NADP(H), had a subunit molecular weight of 29,220 and a native molecular weight of 88,000. The cloned ketoreductase was expressed in E. coli together with a cloned glucose 6-phosphate dehydrogenase from Saccharomyces cerevisiae to allow regeneration of the NADPH required by the ketoreductase. An extract of E. coli containing the two recombinant enzymes was used to reduce 2-chloro-1-(3-chloro-4-fluorophenyl)-ethanone and two related ketones to the corresponding S-alcohols. Intact E. coli cells provided with glucose were used to prepare (S)-2-chloro-1-(3-chloro-4-fluorophenyl)-ethanol in 89% yield with 100% ee.

Resolution of (±)- sec -butylamine by Candida antarctica lipase provided a very low enantiomeric excess of the residual amine when either ethyl or vinyl butyrate was used as the acylating agent. The enantiomeric excess was increased by using ethyl esters of long chain fatty acids. The rate of the reaction was increased by using methyl t -butyl ether as a solvent. ( S )- sec -Butylamine of very high enantiomeric excess was obtained by C. antarctica lipase catalyzed acylation with ethyl decanoate in methyl t -butyl ether.

The synthesis of a new series of arylmethyltin chlorides is reported, ArnMeSnCl3−n (Ar = RC6H4, R = H, ethyl, i-propyl, t-butyl; hexyl, octyl). The synthesis involves initial formation of triarylmethyltin compounds, Ar3MeSn, via Grignard techniques followed by HCl–Et2O aryl group cleavage, preferably in a stepwise manner. Preliminary biological activity against Staphylococcus aureus illustrates the importance of the para-alkyl substituents and reinforces that an optimal hydrophobic character is needed for maximum efficacy. Copyright © 2005 John Wiley & Sons, Ltd.

The single-crystal structure of the hydrophobic actinide extractant di(p-tert-butylphenyl)-N,N-di-(iso-butyl)carbamoylmethylphosphine oxide (CMPO, 1) has been determined, as has that of its 1:1 chelate complex with uranyl nitrate (2), its 1:1 monodentate complexes with Ph3SnCl (3) and Me2SnCl2 (4) bonding via the PO group. A 1:1 bidentate complex with Ph2SnCl2 bonding via both PO and CO functionalities (5) was obtained which in solution exhibits a CO Sn bond dissociation to exist as the monodentate isomer coordinating only through the PO group. Copyright © 2005 John Wiley & Sons, Ltd.