Construction of Recombinant Escherichia coli Catalysts which Simultaneously Express an (S)‐Oxynitrilase and Different Nitrilase Variants for the Synthesis of (S)‐Mandelic Acid and (S)‐Mandelic Amide from Benzaldehyde and Cyanide

Recombinant Escherichia coli strains were constructed which simultaneously expressed the genes encoding the (S)‐oxynitrilase from cassava (Manihot esculenta) together with the wild‐type or a mutant variant of the arylacetonitrilase from Pseudomonas fluorescens EBC191 in a single organism under the control of a rhamnose‐inducible promoter. The whole cell catalysts obtained converted benzaldehyde and potassium cyanide in aqueous media at pH 5.2 mainly to (S)‐mandelic acid and/or (S)‐mandelic amide and synthesized only low amounts of the corresponding (R)‐enantiomers. The conversion of benzaldehyde and potassium cyanide (KCN) by a whole‐cell catalyst simultaneously expressing the (S)‐oxynitrilase and the wild‐type nitrilase resulted in a ratio of (S)‐mandelic acid to (S)‐mandelic amide of about 4:3. This could be explained by the strong nitrile hydratase activity of the wild‐type nitrilase with (S)‐mandelonitrile as substrate. The relative proportion of (S)‐mandelic amide formed in this system was significantly increased by coexpressing the (S)‐oxynitrilase with a carboxy‐terminally truncated variant of the nitrilase. This whole‐cell catalyst converted benzaldehyde and KCN to mandelic amide and mandelic acid in a ratio of about 9:1. The ee of the (S)‐mandelic amide formed was calculated to be >95%.     

Application of a Recombinant Escherichia coli Whole‐Cell Catalyst Expressing Hydroxynitrile Lyase and Nitrilase Activities in Ionic Liquids for the Production of (S)‐Mandelic Acid and (S)‐Mandeloamide

The conversion of benzaldehyde and cyanide into mandelic acid and mandeloamide by a recombinant Escherichia coli strain which simultaneously expressed an (S)‐hydroxynitrile lyase (oxynitrilase) from cassava (Manihot esculenta) and an arylacetonitrilase from Pseudomonas fluorescens EBC191 was studied. Benzaldehyde exhibited a pronounced inhibitory effect on the nitrilase activity in concentrations ≥25 mM. Therefore, it was tested if two‐phase systems consisting of a buffered aqueous phase and the ionic liquid 1‐butyl‐1‐pyrrolidinium bis(trifluoromethanesulfonyl)imide (BMpl NTf₂) or 1‐butyl‐3‐methylimidazolium hexafluorophosphate (BMim PF₆) could be used for the intended biotransformation. The distribution coefficients of the substrates, intermediates and products of the reaction were determined and it was found that BMpl NTf₂ and BMim PF₆ were highly efficient as substrate reservoirs for benzaldehyde. The recombinant E. coli strain was active in the presence of BMpl NTf₂ or BMim PF₆ phases and converted benzaldehyde and cyanide into mandelic acid and mandeloamide. The two‐phase systems allowed the conversion of benzaldehyde dissolved in the ionic liquids to a concentration of 700 mM with product yields (=sum of mandelic acid and mandeloamide) of 87–100%. The cells were slightly more effective in the presence of BMpl NTf₂ than in the presence of BMim PF₆. In both two‐phase systems benzaldehyde and cyanide were converted into (S)‐mandeloamide and (S)‐mandelic acid with enantiomeric excesses ≥94%. The recombinant E. coli cells formed, in the two‐phase systems with ionic liquids and increased substrate concentrations, higher relative amounts of mandeloamide than in a purely aqueous system with lower substrate concentrations.     

Characteristics of gold biosorption from cyanide solution

Gold adsorption from cyanide solution by bacterial (Bacillus subtilis), fungal (Penicillium chrysogenum) and seaweed (Sargassum fluitans) biomass was examined. At pH 2.0, these biomass types were capable of sequestering up to 8.0 µmol g⁻¹, 7.2 µmol g⁻¹ and 3.2 µmol g⁻¹, respectively. An adverse effect of increasing solution ionic strength (NaNO₃) on gold biosorption was observed. Gold‐loaded biomass could be eluted with 0.1 mol dm⁻³ NaOH with efficiencies higher than 90% at pH 5.0 at the Solid‐to‐Liquid ratio, S/L, = 4 (g dm⁻³). Cyanide mass balances for the adsorption, desorption as well as for the AVR process indicated the stability of the gold‐cyanide which did not dissociate either upon acidification or upon binding by biomass functional groups. Gold biosorption mainly involved anionic AuCN₂⁻ species bound by ionizable biomass functional groups carrying a positive charge when protonated. FTIR analyses indicated that the main biomass functional groups involved in gold biosorption are most probably nitrogen‐containing weak base groups. The present results confirmed that waste microbial biomaterials have some potential for removing and concentrating gold from solutions where it occurs as a gold‐cyanide complex. © 1999 Society of Chemical Industry     

Ionic conductivity and morphology of semi‐interpenetrating‐type polymer electrolyte entrapping poly(siloxane‐g‐allyl cyanide)

We prepared a semi‐IPN (interpenetrating network)‐type solid polymer electrolyte (SPE) using poly (ethylene glycol)dimethacrylate (PEGDMA) as a polymer matrix containing a monocomb‐type poly(siloxane‐g‐allyl cyanide) and poly(ethylene glycol)dimethylether (PEGDME) for the lithium secondary battery. The poly(siloxane‐g‐allyl cyanide)s were prepared by a hydrosilation reaction of poly (methyl hydrosiloxane) with allyl cyanide and characterized by ¹H NMR and FTIR. The semi‐IPN‐type electrolyte was prepared by thermal curing, and conductivities of samples were measured by impedance spectroscopy using an indium tin oxide (ITO) electrode. The ionic conductivity of the semi‐IPN‐polymer electrolyte was about 1.05 × 10^?5 S cm^?1 with 60 wt % of the poly(siloxane‐g‐allyl cyanide) and 6.96 × 10^?4 S cm^?1 with 50 wt % of the PEGDME and 10 wt % of the poly(siloxane‐g‐allyl cyanide) at 30°C. The SEM morphology of the cross section of the semi‐IPN‐polymer electrolyte film was changed from discontinuous network to continuous network as increasing the PEGDME content and decreasing the poly(siloxane‐g‐allyl cyanide) content. The mechanical stability was also enhanced when increasing the PEGDME content. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci, 2008     

Investigation of sensing effects of polystyrene‐graft‐polyaniline for cyanide compounds

Polyaniline and its derivatives have high antioxidative stability and are considered to belong to the most useful conductive polymers for practical application. In this work, we have linked 1,4‐phenylenediamine to poly(styrene‐co‐chloromethylstyrene) and prepared poly(styrene‐co‐p‐aminoanilinemethylstyrene), poly(S‐co‐PAAMS). Polystyrene‐graft‐polyaniline, (PS‐g‐PANi), has been synthesized by adding solution of ammonium persulfate and p‐toluenesufonic acid in water. The conductivity of these polymers was measured by the four‐point probe method. The copolymers produced exhibit electrical conductivity comparable to that of polyaniline. PS‐g‐PANi was also exposed to some cyanide compounds such as hydrocyanic acid (Hydrogen cyanide‐HC), ethanedinitrile (Cyanogen‐CY), cyanogen chloride (Chlorocyan‐CC), and cyanogen bromide (Bromocyan‐BC). The cyanide compounds are classified as blood agents groups, in terms of chemical warfare agents standards. First, we prepared different concentration of blood agents at 50, 100, and 150 ppm and exposed them on PS‐g‐PANi for 2 min. Second, we have drawing conductivity change curves versus time and produced optimum conductivity versus time taken. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 101: 3920–3926, 2006     

Probing the Outstanding Local Hydrophobicity Increases in Peptide Sequences Induced by Incorporation of Trifluoromethylated Amino Acids

In order to achieve accurate determination of the local hydrophobicity increases in peptide sequences produced by incorporation of trifluoromethylated amino acids (TfmAAs), the chromatographic hydrophobicity indexes (?₀) of three series of tripeptides containing three unnatural trifluoromethylated amino acids have been measured and compared with those of their non‐fluorinated analogues. The hydrophobic contribution of each fluorinated amino acid was quantified by varying the position and the protection of (R)‐ and (S)‐α‐trifluoromethylalanine (TfmAla), (R)‐trifluoromethylcysteine (TfmCys), and (S)‐trifluoromethionine (TFM) in a short peptide sequence. As a general trend, strong increases in hydrophobicity were precisely measured, even exceeding the high hydrophobic contribution of the natural amino acid isoleucine. This study validates the incorporation of trifluoromethylated amino acids into peptide sequences as a rational strategy for the fine‐tuning of hydrophobic peptide–protein interactions.     

Diffusion process of amino acids in polymer supports for solid‐phase peptide synthesis as studied by pulsed‐field‐gradient spin‐echo proton nuclear magnetic resonance

The diffusion coefficient (D) values of tert‐butyloxycarbonyl‐glycine, tert‐butyloxycarbonyl‐L ‐tryptophan, tert‐butyloxycarbonyl‐L ‐phenylalanine (Boc‐Phe), and 9‐fluorenylmethoxycarbonyl‐L ‐phenylalanine in Merrifield polystyrene (MPS) gels, poly(ethylene glycol)‐grafted polystyrene (PEG–PS) gels, and crosslinked ethoxylate acrylate (CLEAR) gels, as used in solid‐phase peptide synthesis, were determined by the pulsed‐field‐gradient spin‐echo ¹H‐NMR method. From these experimental results, it was found that the amino acids in MPS gels, PEG–PS gels, and CLEAR gels with N,N‐dimethylformamide‐d₇ (DMF‐d₇) as a solvent had multidiffusion components within a measurement timescale of 10 ms. The D value of Boc‐Phe in polystyrene gels (1% divinylbenzene crosslinked) with tetrahydrofuran‐d₈ was much larger than that in the same gels with DMF‐d₇. Furthermore, the required time in which an amino acid transferred from a reactive site to a reactive site was estimated, within which the solvents and amino acids in the polymer supports diffused in the swollen beads.© 2003 Wiley Periodicals, Inc. J Appl Polym Sci 89: 413–421, 2003     

Double metal cyanide complexes synthesized by solvent‐free grinding method for copolymerization of CO2 and propylene oxide

Two active double metal cyanide (DMC) complexes were successfully synthesized by solvent‐free grinding method. Their structures were characterized by FTIR spectrometer and X‐ray diffractometer. The results showed that Complex 1 (double metal cyanide complex with K₃Fe(CN)₆ and ZnCl₂) and Complex 2 (double metal cyanide complex with K₃Fe(CN)₆ and Zn(CH₃COO)₂) had the same structures, crystal forms, and lower crystallinity as both of them synthesized by conventional solvent‐based methods, respectively. Investigations on grinding conditions indicated that Complex 1 ground 14 min at a high grinding strength could achieve low crystallinity and showed substantially amorphous structures. Two speculated structures of DMC were given. The alternating copolymerization of CO₂ and propylene oxide with Complex 1 as catalyst obtained anticipated poly(propylene carbonate) (PPC) with very high catalytic activity. The PPC produced by optimized Complex 1 has molecular weight (M_n) up to 98,000 and narrow polydispersity of 1.93 with more than 90% carbonate linkages. Compared with Complex 1 , Complex 2 displayed low catalytic activity but high selectivity mainly due to the electron atmosphere and strong steric hindrance. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Enzymatic Synthesis of Chiral Phenylalanine Derivatives by a Dynamic Kinetic Resolution of Corresponding Amide and Nitrile Substrates with a Multi‐Enzyme System

Mutant α‐amino‐ε‐caprolactam (ACL) racemase (L19V/L78T) from Achromobacter obae with improved substrate specificity toward phenylalaninamide was obtained by directed evolution. The mutant ACL racemase and thermostable mutant D ‐amino acid amidase (DaaA) from Ochrobactrum anthropi SV3 co‐expressed in Escherichia coli (pACLmut/pDBFB40) were utilized for synthesis of (R)‐phenylalanine and non‐natural (R)‐phenylalanine derivatives (4‐OH, 4‐F, 3‐F, and 2‐F‐Phe) by dynamic kinetic resolution (DKR). Recombinant E. coli with DaaA and mutant ACL racemase genes catalyzed the synthesis of (R)‐phenylalanine with 84% yield and 99% ee from (RS)‐phenylalaninamide (400 mM) in 22 h. (R)‐Tyrosine and 4‐fluoro‐(R)‐phenylalanine were also efficiently synthesized from the corresponding amide compounds. We also co‐expresed two genes encoding mutant ACL racemase and L ‐amino acid amidase from Brevundimonas diminuta in E. coli and performed the efficient production of various (S)‐phenylalanine derivatives. Moreover, 2‐aminophenylpropionitrile was converted to (R)‐phenylalanine by DKR using a combination of the non‐stereoselective nitrile hydratase from recombinamt E. coli and mutant ACL racemase and DaaA from E. coli encoding mutant ACL racemase and DaaA genes.     

Ruthenium‐Catalyzed Remote Electronic Activation of Aromatic C?F Bonds

The tandem isomerization and nucleophilic aromatic substitution of allylic fluoro‐substituted benzylic alcohols is described for the first time. In the presence of the ruthenium complex Ru(PPh₃)₃(CO)(H)₂, 1‐(4‐fluorophenyl)prop‐2‐en‐1‐ol is converted into the corresponding para‐amino ketone or para‐phenolic substituted ketone.     

Synthesis of 20‐O‐linked 20(S)‐Camptothecin Glycoconjugates: Impact of the Side Chain of the Ester‐linked Amino Acid on Epimerization During the Acylation Reaction and on Hydrolytic Stability of the Final Glycoconjugates

To improve solubility and tumor selectivity of 20(S)‐camptothecin the synthesis of 20‐O‐linked glycoconjugates 11A — G is described. Particular focus of the paper is the utilization of N‐tert‐butoxycarbonyl protected amino acid N‐carboxy anhydrides (UNCAs) 2a — f for an efficient acylation of the sterically hindered and deactivated tertiary 20‐hydroxy group of 20(S)‐camptothecin 1 . Depending on the solvent and on the side chain of the amino acid different extents of epimerization of the amino acids during the coupling reaction are observed; however, the epimers can easily be separated after removal of the tert‐butoxycarbonyl protecting group and camptothecin amino acid conjugates 4B — E with L‐ and D‐ configured amino acids, respectively, are obtained. Particularly, bulky and β‐branched amino acids can be attached to camptothecin in high yields and with low epimerization. Starting from the camptothecin amino acid conjugates 4B — E the synthesis of the glycoconjugates 11A — G is straightforward following standard procedures. The glycoconjugate hydrochlorides 11A — G show good water solubility (> 5mg‐ ml) and hydrolytic stability of the ester bond which again depends on the side chain of the amino acid residue linked to camptothecin. Particularly, glycoconjugates 11B — E with a bulky and β‐branched amino acid at the ester linkage show high hydrolytic stability in aqueous solutions with less than 2.5% of 20(S)‐camptothecin cleaved within 24 h. These results provide a basis for the selection of appropriate spacer groups for delivery systems of 20(S)‐camptothecin for therapeutic use.     

Novel 7‐maleimido‐2(1H)‐quinolones as potential fluorescent sensors for the detection of sulphydryl groups

Novel compounds, based on the 2(1H)‐quinolone skeleton, were synthesised and characterised using ¹H‐nuclear magnetic resonance (NMR) spectroscopy and chemical ionisation mass spectrometry (CI MS). Their spectroscopic properties, such as absorption and emission spectra and fluorescence quantum yield, were also examined. 7‐maleimido‐4‐methyl‐2(1H)‐quinolone, 7‐maleimido‐3,4‐dimethyl‐2(1H)‐quinolone, 7‐maleimido‐4‐propyl‐2(1H)‐quinolone and 7‐maleimido‐4‐phenyl‐2(1H)‐quinolone were evaluated as potential sensors for the detection of sulphydryl amino acids groups. These new compounds demonstrate a high ‘turn‐on’ fluorescence response and selectivity towards l ‐cysteine in the presence of other amino acids and metal cations.     

Mono‐, Di‐ and Tetra‐Cationic Surfactants as Carbon Steel Corrosion Inhibitors

Corrosion inhibition of three new synthesized cationic surfactants, N‐(2‐(((Z)‐4‐(pyridin‐4‐yl)but‐3‐en‐1‐yl)amino)ethyl)‐N‐(2‐((E)‐(pyridin‐4‐ylmethylene)amino)ethyl)dodecan‐1‐aminium bromide I(4N), N¹,N²‐didodecyl‐N¹‐((Z)‐4‐(pyridin‐4‐yl)but‐3‐en‐1‐yl)‐N²‐(2‐((E)‐(pyridin‐4‐ylmethylene)amino)ethyl)ethane‐1,2‐diaminium bromide II(4N) and 1‐dodecyl‐4‐((E)‐((2‐(dodecyl(2‐(dodecyl((Z)‐4‐(1‐dodecylpyridin‐1‐ium‐4‐yl)but‐3‐en‐1‐yl)ammonio)ethyl)ammonio)ethyl)imino)methyl)pyridin‐1‐ium bromide IV(4N) on carbon steel was investigated by weight loss, electrochemical impedance spectroscopy and polarization measurements. Results show that the synthesized cationic surfactants inhibit corrosion of carbon steel in 1 M HCl. The inhibitive action occurs by virtue of adsorption on the metal surface following a Langmuir adsorption isotherm model. Polarization curves reveal that the investigated cationic surfactants can be classified as mixed inhibitor types. The variations in the corrosion inhibition efficiency between three cationic surfactants are correlated with their chemical structures, with more hydrophobic surfactants yielding higher inhibition efficiency.     

Ruthenium‐Catalyzed Asymmetric Hydrogenation of β‐Keto‐ enamines: An Efficient Approach to Chiral γ‐Amino Alcohols

A highly efficient and enantioselective hydrogenation of unprotected β‐ketoenamines catalyzed with ruthenium(II) dichloro{(S)‐(−)‐2,2′‐bis[di(3,5‐xylyl)phosphino]‐1,1′‐binaphthyl}[(2S)‐(+)‐1,1‐bis(4‐methoxyphenyl)‐3‐methyl‐1,2‐butanediamine] {Ru[(S)‐xylbinap][(S)‐daipen]Cl₂} has been successfully developed. This methodology provides a straightforward access to free γ‐secondary amino alcohols, which are key building blocks for a variety of pharmaceuticals and natural products, with high yields (>99%) and excellent enantioselectivities (up to 99% ee) in all cases.     

Enantioselective Cyanosilylation of Ketones with Amino Acid/BINAP/Ruthenium(II)‐Lithium Phenoxide Catalyst Systems

Enantioselective reactions of simple ketones, α,α‐ and β,β‐dialkoxy ketones, and α‐alkoxy ketones with trimethylsilyl cyanide catalyzed by the bimetallic systems of amino acid/BINAP/ruthenium(II) complexes and lithium phenoxide have been studied [BINAP=2,2′‐bis(diphenylphosphino)‐1,1′‐binaphthyl]. The Ru(PhGly)₂(BINAP)‐lithium phenoxide system showed high enantioselectivity for the reaction of acetophenone derivatives to afford the cyanated products in up to 90% ee [PhGly=phenylglycinate]. For the cyanosilylation of dialkoxy ketones and α‐alkoxy ketones, the Ru(t‐Leu)₂(BINAP)‐lithium phenoxide system exhibited the best catalyst performance to produce the cyanohydrin derivatives in up to 99% ee and 98% ee, respectively [t‐Leu=tert‐leucinate]. The excellent catalytic activity resulted in complete conversion in the reaction with a substrate‐to‐catalyst molar ratio (S/C) of 10,000 in the best cases.     

Solvent extraction of gold(I) from alkaline cyanide solution by dibutylcarbitol (DBC) with n‐octanol

BACKGROUND: Currently, cyanidisation is preferred for the extraction of gold because it has a number of advantages over other methods. Gold(I) can be extracted with various extractants, but there are no reports on the extraction of gold(I) from cyanide solution by dibutylcarbitol (DBC). In this work the extraction of gold(I) from alkaline cyanide solution using DBC with n‐octanol was studied. Several factors affecting the percentage extraction of gold(I), including DBC concentration, diluent concentration, equilibrium time, phase ratio, pH and gold concentration in aqueous phase, were investigated. RESULTS: The results showed that 96.2% of gold(I) could be extracted using an organic phase composed of 40% (v/v) DBC, 50% (v/v) n‐octanol and 10% (v/v) odourless kerosene. The extraction was quite fast and equilibrium could be established within 2 min. Stripping of the gold‐laden organic phase was carried out using sodium sulfite (Na₂SO₃) and sodium thiosulfate (Na₂S₂O₃) solutions, with Na₂S₂O₃ proving better than Na₂SO₃. The percentage stripping of gold(I) was 96.5% when the Na₂S₂O₃ concentration was 4% (w/w), and the stripping capacity of gold(I) exceeded 311.3 mg L^?1 when the phase ratio (A/O) was equal to 0.2. CONCLUSION: Gold(I) can be extracted from aqueous cyanide solution by DBC in the presence of n‐octanol and efficiently stripped by aqueous Na₂S₂O₃ solution. This method has the potential for practical application in the extraction and separation of gold. Copyright © 2008 Society of Chemical Industry     

Amino‐functional electrospun nanofibrous membrane for detecting nitroaromatic compounds

A novel amino‐functionalized polystyrene copolymer (PS‐NH₂) was designed and synthesized with styrene and 4‐vinylbenzyl amine. Additionally, an amino modified glass (G‐NH₂) was obtained as a carrier. (PS‐NH₂/pyrene)/G‐NH₂ fluorescent nanofibrous membrane [named (PS‐NH₂/pyrene)/G‐NH₂] was designed and prepared via electrospinning technique to detect representative saturated nitroaromatic (NAC) explosive vapor. The (PS‐NH₂/pyrene)/G‐NH₂ showed highly fluorescence stability in ambient condition and further displayed a high quenching efficiency of 70.9% toward trinitrotoluene (TNT) vapor (～10 ppb) with an exposure time of 150 s at room temperature. The abundance of amino groups could effectively adsorb NACs and the binding of electron‐deficient NACs to the amino groups on the (PS‐NH₂/pyrene)/G‐NH₂ surface led to the formation of charge‐transfer complexes. The quenching constant (K_SV) to TNT was obtained to be 1.07 × 10¹¹ mL/g in gaseous phase with a limit of detection up to 2.76 × 10^?13g/mL. Importantly, the (PS‐NH₂/pyrene)/G‐NH₂ showed notable selectivity toward TNT and 2,4‐dinitrotoluene vapors. Straightforwardly, the colorimetric sensing performance can be visualized by naked eye with a color change for detecting of different vapor phase NACs explosives. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2018 , 135, 46708     

Electrolytic oxidation of cuprocyanide electroplating waste waters under different pH conditions

The main purpose of this work was to investigate the electrolytic oxidation of cuprocyanide solution with various total cyanide to copper molar ratios ranging from 2.8 to 20 and under different pH conditions. In strong alkaline solution (pH12), cuprocyanide ions Cu(CN) _n ^/(n–1)– , wheren=2, 3 or 4, are directly electroxidized, and copper oxide precipitates on the anode. Cyanate ions, as well as nitrogen gas, were detected as the products and 0.30–0.43 g mol of total cyanide was destroyed per Faraday. For less alkaline solutions (pH<12), cuprocyanide ions first dissociated to free cyanide ions and then electroxidized. At a pH of 10.5–11.7, cyanate ion and brown azulmin polymer were produced in the anolyte. In the neutral solution (pH=7.0–8.6), carbonate and ammonium ions and azulmin were formed and 0.52–0.56 g mol of total cyanide was destroyed per Faraday. In weak acidic solution (pH=5.2–6.8), oxalate and ammonium ions and white oxamide were produced and 1.01–1.18 g mol of total cyanide were destroyed per Faraday.Nomenclature C _CN molar concentration of total cyanide (kmol m^–3) - C _Cu molar concentration of total copper (kmol m^–3) - C _d equivalent concentration of cyanide destroyed due to the formation of cupric oxide (kg m^–3) - C _f concentration of cyanide destroyed by dissociation of complex ion to free cyanide ion and then electroxidized (kg m^–3) - C _i initial concentration of total cyanide (kg m^–3) - C _t change of total cyanide concentration during electrolysis (kg m^–3) - F Faraday constant (96 487 C mol^–1) - K ₁,K ₂,K ₃ formation constant of dicyanocuprate, tricyanocuprate and tetracyanocuprate ions - R molar ratio of total cyanide concentration to total copper concentration (i.e.C _CN/C _Cu) - W weight of precipitates on electrodes or in anolyte (kg) - angle of incidence     

A One‐Pot Synthesis of [1,2,4]Triazino[4,3‐b]‐[1,2,4,5]tetrazines

Reactions of hydrazonoyl halides 6 with either 4‐amino‐2,3‐dihydro‐6‐substituted‐3‐thioxo‐[1,2,4]‐triazin‐5(4H)ones 1 ( 2 ) or 4‐amino‐3‐methylthio‐6‐substituted‐[1,2,4]‐triazin‐5(4H)ones 3 ( 4 ) gave [1,2,4]‐triazino‐[4,3‐b][1,2,4,5]tetrazine derivatives 9 ( 10 ), respectively. The mechanism of the reactions studied is discussed.     

Preparation of (R)‐Amines from Racemic Amines with an (S)‐Amine Transaminase from Bacillus megaterium

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.