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A novel marine actinomycete strain designated ICN19T was isolated from the subtidal sediment of Chinnamuttam coast of Kanyakumari, India and subjected to polyphasic taxonomic analysis. Neighbour-joining tree based on 16S rRNA gene sequences of validly described type strains had revealed the strain ICN19T formed distinct cluster with Streptomyces wuyuanensis CGMCC 4.7042T, Streptomyces tirandamycinicus HNM0039T and Streptomyces spongiicola HNM0071T. Morphological, physiological and chemotaxonomic characteristics were consistent with those of members of the genus Streptomyces. The strain possessed ll-diaminopimelic acid as the diagnostic diamino acid. The predominant isoprenoid quinone was identified as MK-9(H8) (70%), MK-9(H6) (20%) and MK-9(H2) (2%), with the major cellular fatty acids (>10%) being anteiso-C15:0, C16:0 and iso-C16:0. The main polar lipids were found to be diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol mannosides and three unidentified phospholipids. The dendrogram generated on the basis of MALDI-TOF mass spectra supports the strain differentiated from its neigbours. The genome sequence of strain ICN19T was 9,010,366 bp in size with a total of 7420 protein-coding genes and 98 RNA genes. The genomic G+C content of the novel strain was 71.27 mol%. The DNA–DNA relatedness between strain ICN19T and the reference strains with S. wuyuanensis CGMCC 4.7042T, S. tirandamycinicus HNM0039T and S. spongiicola HNM0071T were 42.8%, 39.5% and 38%, respectively. Based on differences in physiological, biochemical, chemotaxonomic differences and whole-genome characteristics the isolated strain represents a novel species of the genus Streptomyces, for which the name Streptomyces marianii sp. nov. is proposed. Type strain is ICN19T (=MCC 3599T = KCTC 39749T).

Reversible addition fragmentation transfer (RAFT) polymerization can be used to produce highly branched polymer approximating the structure of dendrimers, but with less regularity and much less synthetic complexity. This can be accomplished by using a RAFT agent with a polymerizable double bond. Thus, the stoichiometry is fixed, with one branch point (RAFT agent double bond) per chain. Since each of these branch points will also branch, a highly regular highly branched material is formed. If this RAFT chemistry is applied to the monomer N-isopropyl-acrylamide (NIPAAm), the resulting water-soluble polymer has unique applications in drug delivery, due to the hyperbranched nature of the polymer, and also to the lower critical solution temperature exhibited by poly(NIPAAm). In this paper, a previous mathematical model of the RAFT-pNIPAAm system is extended, and validated against experimental polymerization data.

Branch lengths resulting from both backbiting and intermolecular chain transfer to polymer are examined for the solution polymerization of butyl acrylate, using a rate-Equation model and ordinary differential equations. Backbiting is allowed to generate branches of varying length, according to a cumulative distribution function obtained from a lattice kinetic Monte Carlo simulation. About 8% of the branches produced by backbiting are 10 mers or longer. In contrast to common assumptions about the origins of short-chain and long-chain branches, the model indicates that nearly all of the long-chain branches may be produced by backbiting, rather than intermolecular chain transfer to polymer.

(+) (1S,2S)-2-Amino-1-phenyl-1,3-propanediol reacts with ortho-esters to form 4-hydroxymethyl-5-phenyl-1,3-oxazolines. Subsequent reaction of their toluenesulfonyl derivatives with diphenylphosphinolithium yields the N,P-ligands, (4S,5S)-2-R-4-diphenylphosphinomethyl-5-phenyl-1,3-oxazoline (R = Me, Et, Ph). X-ray analyses of (4S,5S)-2-methyl-4-toluenesulfonylmethyl-5-phenyl-1,3-oxazoline and (4S,5S)-2,5-diphenyl-4-diphenylphosphinomethyl-1,3-oxazoline reveal retention of absolute configuration throughout the synthesis. The [Ir(COD)(N,P-oxazoline)]PF6 systems in CH2Cl2 effect catalytic hydrogenation of N-(1-phenylethylidene)benzylamine, PhCH2N = C(Me)Ph, to the corresponding amine with upto 63% e.e. with the R = Et ligand.

Interaction of the cis-[Rh(PR3)2(Solv)2]PF6 complexes (R = Ar or R3 = Ph2Me, Solv — solvent) under Ar with semicarbazones bearing a phenyl group on the imine-C atom gives the rhodium(iii)-hydrido-bis(phosphine)-orthometallated semicarbazone species [RhH(PR3)2{(o-C6H4(R")C=N—N(H)CONH2}]PF6 (R" = Me or Et), which are characterized generally by elemental analysis, 31P{1H} and 1H NMR spectroscopy, and mass-spectrometry. The PPh3-containing complex with R" = Me, structurally characterized by X-ray analysis, reveals coordination of the semicarbazone by the ortho-C atom, the imine-N atom, and the amide-carbonyl group. For a semicarbazone containing no Ph group, the rhodium(i) complex [Rh(PR3)2(Et(Me)C=N—N(H)CONH2)]PF6, containing the η2-semicarbazone bonded via the imine-N and carbonyl, is formed. Attempts to hydrogenate the C=N moiety in the complexes or to catalytically hydrogenate the semicarbazones were unsuccessful.

This contribution to the “special issue” of Applied Catalysis A: General entitled “Industrial catalytic processes” deals with the development of the methanol process during the last 10–15 years. Following a brief review of the history, the developments to improve methanol synthesis are presented along the lines of elements of catalyst system improvements and of reactor improvements.

The epoxidation of a number of cyclic and linear terminal alkenes by an O 2 –O 3 mixture at room temperatures catalysed by Fe(TMP)Cl is reported. It appears that the alkene forms a primary ozonide which rearranges to a secondary ozonide, and it is the latter which, when catalysed by Fe(TMP)Cl, affects the subsequent alkene epoxidation. A number of other metal complexes were also tested as catalysts, but Fe(TMP)Cl was the most effective.

Methods of polymer reaction engineering were used to develop a mechanistic model of nonenzymatic polynucleotide replication. The model is made possible by the rate constants developed by Rajamani et al. This model treats the ideas of stalling proposed by Rajamani in a direct kinetic mechanism. When one assumes that multiple incorrect insertions in a row are unlikely, the model returns estimates of the error threshold very much in agreement with the modified Eigen model (based on probability arguments) proposed by Rajamani. In addition, the current model gives average lengths of poly­nucleotides, categorized by ultimate nucleoside (type and correct/incorrect) as well as the average lengths of interior and terminal sequences of correct and incorrect insertions in a row. When the insights of Leu et al. (specifically, that incorrect additions following an incorrect addition are likely) are included into the model, the error threshold will drop to unrealistic levels. To compensate for this, a mechanism for the “recycle” of incorrect strands (via hydrolysis and repolymerization) was included. Calculations indicate that even with a “correction” mechanism (hydrolysis of chain containing errors), the error threshold drops to an unacceptable value.

Aldehyde-mediated olefin epoxidations appear to proceed by parallel peracid and radical addition pathways. For that portion of the reaction proceeding by radical addition, several lines of evidence favor an explanation in which the peroxyacyl-olefin adduct decomposes in a concerted manner to form alkyl radical, CO 2 , and epoxide.

From 2005 to 2010, The Dow Chemical Company (formerly Rohm and Haas Company) was a member of the Department of Energy Center of Excellence on Chemical Hydrogen Storage, which conducted research to identify and develop chemical hydrogen storage materials having the potential to achieve DOE performance targets established for on-board vehicular application. In collaboration with Center co-leads Los Alamos National Laboratory (LANL) and Pacific Northwest National Laboratory (PNNL), and other Center partners, Dow's efforts were directed towards defining and evaluating novel chemistries for producing chemical hydrides and processes for spent fuel regeneration. In Phase 1 of this project, emphasis was placed on sodium borohydride (NaBH{sub 4}), long considered a strong candidate for hydrogen storage because of its high hydrogen storage capacity, well characterized hydrogen release chemistry, safety, and functionality. Various chemical pathways for regenerating NaBH{sub 4} from spent sodium borate solution were investigated, with the objective of meeting the 2010/2015 DOE targets of $2-3/gal gasoline equivalent at the pump ($$2-3/kg H{sub 2}) for on-board hydrogen storage systems and an overall 60% energy efficiency. With the September 2007 No-Go decision for NaBH{sub 4} as an on-board hydrogen storage medium, focus was shifted to ammonia borane (AB) for on-board hydrogen storage and delivery. However, NaBH{sub 4} is a key building block to most boron-based fuels, and the ability to produce NaBH{sub 4} in an energy-efficient, cost-effective, and environmentally sound manner is critical to the viability of AB, as well as many leading materials under consideration by the Metal Hydride Center of Excellence. Therefore, in Phase 2, research continued towards identifying and developing a single low-cost NaBH4 synthetic route for cost-efficient AB first fill, and conducting baseline cost estimates for first fill and regenerated AB using a variety of synthetic routes. This project utilized an engineering-guided R&D approach, which involved the rapid down-selection of a large number of options (chemical pathways to NaBH{sub 4}) to a smaller, more manageable number. The research began by conducting an extensive review of the technical and patent literature to identify all possible options. The down-selection was based on evaluation of the options against a set of metrics, and to a large extent occurred before experimentation was initiated. Given the vast amount of literature and patents that has evolved over the years, this approach helped to focus efforts and resources on the options with the highest technical and commercial probability of success. Additionally, a detailed engineering analysis methodology was developed for conducting the cost and energy-efficiency calculations. The methodology utilized a number of inputs and tools (Aspen PEA{trademark}, FCHTool, and H2A). The down-selection of chemical pathways to NaBH{sub 4} identified three options that were subsequently pursued experimentally. Metal reduction of borate was investigated in Dow's laboratories, research on electrochemical routes to NaBH{sub 4} was conducted at Pennsylvania State University, and Idaho National Laboratory researchers examined various carbothermal routes for producing NaBH{sub 4} from borate. The electrochemical and carbothermal studies did not yield sufficiently positive results. However, NaBH{sub 4} was produced in high yields and purities by an aluminum-based metal reduction pathway. Solid-solid reactive milling, slurry milling, and solution-phase approaches to metal reduction were investigated, and while both reactive milling and solution-phase routes point to fully recyclable processes, the scale-up of reactive milling processes to produce NaBH{sub 4} is expected to be difficult. Alternatively, a low-cost solution-phase approach to NaBH{sub 4} has been identified that is based on conventional process unit operations and should be amenable to scale-up. Numerous advances in AB synthesis have been made in recent years to improve AB yields and purities. Process analysis of several leading routes to AB (Purdue's formate-based metathesis route and PNNL's NH{sub 4}BH{sub 4}-based route) indicated the cost to produce first-fill AB to be on the order of $$9-10/kg AB, assuming a NaBH{sub 4} cost of $5/kg for a 10,000 metric tons/year sized AB plant. The analysis showed that the dominant cost component for producing first-fill AB is the cost of the NaBH4 raw material. At this AB cost and assuming 2.5 moles hydrogen released per mole of AB, it may be possible to meet DOE's 2010 storage system cost target, but the 2015 target will likely require lower cost AB and demonstrates the importance of having a low-cost route to NaBH{sub 4}. Substantial progress has also been made to define feasible pathways for the regeneration of spent ammonia borane fuel.

An account is given of the development of electrocatalysts for the phosphoric acid (phosphoric acid) fuel cell starting from unsupported platinum black and ending with carbon-supported platinum alloys. The incentive for this work was the need to reduce the cost of the phosphoric acid fuel cell by reducing the amount of platinum it contained while simultaneously raising its performance. Equally important was the concurrent effort aimed at optimizing the structure of the polytetrafluoroethylene (PTFE)-bonded, gas-diffusion electrodes so that the noble metal catalysts were used as effectively as possible. Methods to control the structure of the catalyst layer were devised based on studies of the way in which aqueous dispersions of the supported catalyst and PTFE interact. Procedures used to prepare electrodes in the laboratory were modified and scaled up for production purposes. A novel process for depositing catalyst layers on electrode substrates was developed and several different coating technologies were evaluated for applying PTFE-bonded, silicon carbide, electrolyte-retaining matrices to the electrodes. Some operating conditions that can have an adverse affect on the life of the electrodes are considered. Most of the work described here originated in the laboratories and workshops of International Fuel Cells — a unit of United Technologies Corporation — and culminated in the PC25 phosphoric acid fuel cell power plant. Keywords: phosphoric acid fuel cells; electrocatalysts; gas-diffusion electrodes; electrolyte retaining matrix; coating technologies

A novel process for producing dimethyl ether (DME) from synthesis gas in a single-step reaction sequence has been developed. The new process uses a slurry reactor in which methanol synthesis, methanol dehydration to DME, and water-gas shift reactions all proceed concurrently. All three reactions are exothermic and reversible. Operation with a back-mixed slurry reactor exploits synergisms of the three reactions, and moderates the reaction exotherm to permit higher conversion of the syngas to liquid products than could be achieved from the three reactions practiced separately. The process offers potential lower capital and operating costs than processes using individual shift, methanol, and DME reaction stages. Process development to-date has focused on the use of coal-derived synthesis gas that is rich in CO. Catalysts used in the process can be a physical mixture of methanol, shift, and dehydration catalysts. Selection of commercially available catalysts and the effect of their different ratios have been investigated. Some process variable results are presented. Commercial applications of the new process are illustrated. Further development of the process is currently underway. In the laboratory, the effects of temperature and feed composition are being studied in detail. Catalyst aging characteristics are also being defined. In addition, plans are being made to demonstrate the process in the Department of Energy's Alternative Fuels Development Unit at LaPorte, Texas.

Both rHis6-BoNT/E and rBoNT/E-His6 were produced from a codon optimized gene within Escherichia coli. Similar to native BoNT/E isolated from Clostridium botulinum, the recombinant proteins were isolated as single-chain molecules. The His6 affinity tag facilitated a two-step IMAC-IEX purification. While reasonable yields were obtained, single-chain rHis6-BoNT/E was produced less efficiently than was rBoNT/E-His6. Treatment with recombinant trypsin efficiently converted both purified rBoNT/E constructs to the activated dichain forms, with concomitant removal of the His6 tag, and the nicking profile for each rBoNT/E molecule matched that of native BoNT/E. Following nicking with trypsin, it was observed that the resultant rHis6-BoNT/E dichain displayed reduced in vivo potency (mouse potency assay) compared to activated native BoNT/E dichain. In contrast, activated rBoNT/E-His6 displayed a specific potency equivalent to that of trypsin-nicked native BoNT/E in the mouse potency assay. These results demonstrate that rBoNT/E, containing a C-terminal His6 affinity tag, can be expressed and purified in high yield, with the activated molecule displaying equivalent in vivo potency to activated native BoNT/E.

In the present study, a detailed activation time-course was examined for single chain native BoNT/E, enzymatically nicked in solution with trypsin. Through SDS-PAGE and Western blot analysis, the optimal nicking time was determined to be approximately 30 min. At 30 min, maximal di-chain BoNT/E and minimal secondary nicking (degradation) products were observed. The identity and integrity of the resulting light chain and heavy chain products were confirmed by N-terminal sequence analysis. Trypsin-activated BoNT/E samples covering a broad time-course were subsequently evaluated for potency and efficacy in pharmacological (DAS) and toxicological (im-LD50, ip-LD50) mouse models. In all three models, sample potencies varied with the degree of BoNT/E activation. In the DAS model, compared to single-chain BoNT/E, all samples displayed improved maximal efficacy and potency, except for 96 h-treated samples, which displayed lower potency. Both im- and ip-LD50 studies displayed similar dose–response profiles across the time-course. All three models yielded maximal potencies with samples activated for 30–100 min. These data demonstrate that purified, native BoNT/E can be activated in a controlled and reproducible manner with trypsin to near completion prior to the formation of any significant amounts of degradation products, with concomitant maximal in vivo potency.

Xeomin ® [Merz; 150 kDa Botulinum type A toxin (BoNT/A)] and BOTOX ® [Allergan; 900 kDa BoNT/A complex] were compared, to assess the degree of similarity between these products in terms of potency and antigenicity. Potency was evaluated in the Digit Abduction Score (DAS) mouse assay. Full-range dose–response profiles were achieved with 3 lots of each product, with similarity between lots for a given product. Between products, however, the mean potency of Xeomin ® was ~50% lower than that of BOTOX ® . Subsequently, studies were conducted to investigate whether rabbit-derived, BOTOX ® -neutralizing antibodies (nAbs) could also neutralize Xeomin ® . Equi-efficacious, near-maximal doses (ED94) were selected for each product lot from the DAS potency profiles. Each ED94 dose for each lot was combined 1:1 with either naive serum (control) or nAbs and then evaluated in the DAS assay. As expected, anti-BOTOX ® nAbs inhibited each of the BOTOX ® lots near-maximally (average DAS inhibition ~85% of matched control). Similarly, anti-BOTOX ® nAbs also inhibited each lot of Xeomin ® near-maximally (average DAS inhibition ∼90%). These results suggest that inhibition of the 150 kDa toxin is the common basis for neutralization for both products. With lower potency and similar antigenicity, Xeomin ® is not dose-equivalent to BOTOX ® and would not be expected to be effective in BoNT/A-resistant subjects.

The hydrated group 4 metal triflates, Hf(OTf)4 and Zr(OTf)4, were found to be excellent catalysts (10 mol%) for the mononitration of o-nitrotoluene using a single equivalent of concentrated (69%) nitric acid. The only side product is water and the catalysts are readily recycled from the aqueous phase and re-used.

Lanthanide(III) triflates catalyse (1–10 mol%) the nitration of a range of simple aromatic compounds in good to excellent yield using stoichiometric quantities of 69% nitric acid; the only by-product is water and the catalyst can be readily recycled by simple evaporation.

Bovine follicular fluid was used as a source for the isolation of gonadal inhibin, the activity of which was monitored by the dose dependent suppression of the FSH content of cultured pituitary cells. The procedures presented result in over 3000-fold purification of the starting material and the purified inhibin has an apparent molecular weight of 56000. The purified inhibin can be dissociated under reducing conditions into two subunits with molecular weights of 44000 and 14000 daltons.

The primary amino acid structures of the 43-kDa (A) and 15-kDa (B) subunits of the 58-kDa form of the hormone inhibin have been elucidated by cloning and analysis of cDNA species derived from bovine granulosa cell mRNA. The A subunit (Mr = 32,298) is a protein of 300 amino acids with two potential N-glycosylation sites and two potential proteolytic processing sites and has a pre-pro region of 60 amino acids. The mature B subunit (Mr = 12,977) is a protein of 116 amino acids synthesized from a separate mRNA. These data establish that a 31-kDa form of inhibin also isolated from bovine follicular fluid, with subunits of 20 kDa (Ac) and 15 kDa (B), is derived from the 58-kDa form by proteolytic processing of the A subunit.

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Type I and Type II adrenal steroid receptors from rat renal and hippocampal cytosols were studied by the technique of Fast Protein Liquid Chromatography. Type I receptors were labelled with [ 3 H]aldosterone plus excess RU26988, and Type II receptors with [ 3 H]dexamethasone. On a Mono Q anion exchange column the molybdate-stabilized renal and hippocampal Type I receptors both eluted as single symmetrical peaks at 0.27 M NaCl, with a recovery of ~90% and 60-fold purification (renal) and 10–15-fold (hippocampal). Molybdate-stabilized Type II binding sites from both hippocampal and renal cytosols co-eluted with the Type I sites. On Superose gel nitration renal Type I receptor-steroid complexes consistently eluted two fractions later than hippocampal Type I complexes, suggesting that the renal complexes are smaller; Type II receptor-steroid complexes from both cytosols co-eluted, consistently one fraction behind hippocampal Type I sites. Sequential gel filtration and anion exchange Chromatography achieved a 1000-fold purification of renal Type I binding sites, with an overall recovery of 10%.

This report describes the application of high performance liquid chromatography (HPLC) and associated techniques to the isolation of inhibin from ovine follicular fluid (oFF). Two modes of HPLC have been assessed. High speed gel permeation chromatography on a Waters I-125 support showed that at pH 7.0 inhibin bioactivity was associated with an approx. 80000 mol. wt protein fraction which co-eluted with the bulk of the protein in oFF and gave a yield of 30–44% of the applied activity. Reversed-phase HPLC (RP-HPLC) of a peptide-rich fraction extracted from oFF was also investigated. Using analytical liquid chromatography on octadecylsilylsilica (ODS-silica), two distinct inhibin bioactive fractions were obtained, with a resultant purification of 23-fold and 3-fold respectively and an overall recovery of 8.5%. These results demonstrate that RP-HPLC may be employed as a direct and rapid technique for the isolation of inhibin from oFF.

Two forms of inhibin with molecular weights of 65 000 and 30 000 (65 and 30 kD) were isolated from ovine follicular fluid using a combination of gel permeation chromatography, reversed-phase high-performance liquid chromatography and preparative polyacrylamide gel electrophoresis. The 65 kD form was partially purified approximately 315-fold whilst the 30 kD form was isolated as two isoforms (29 and 30 kD) of similar biological activity and in >95% purity (1210-fold purification and 4·2% recoveries). On reduction the 30 kD form resolved into four components of 36, 31, 20–21 and 16 kD of which the 20–21 and 16 kD components were similar to the corresponding inhibin subunits isolated from porcine and bovine follicular fluid. The 36 kD component was established as a non-reducible inhibin-like material, based on its binding to antiserum raised against bovine 58 kD inhibin. The nature of the remaining non-reducible 31 kD component is unknown. Two NH2-terminal amino acid sequences (first 13 amino acids) identified in purified 30 kD inhibin were identical to the corresponding subunit amino acid sequences of bovine 31 kD inhibin. J. Endocr. (1987) 113, 213–221