Biotransformation of <Emphasis Type="SmallCaps">l</Emphasis>-tyrosine to Dopamine by a Calcium Alginate Immobilized Mutant Strain of <Emphasis Type="Italic">Aspergillus oryzae</Emphasis>

The present research work is concerned with the biotransformation of l-tyrosine to dopamine (DA) by calcium alginate entrapped conidiospores of a mutant strain of Aspergillus oryzae. Different strains of A. oryzae were isolated from soil. Out of 13 isolated strains, isolate-2 (I-2) was found to be a better DA producer. The wild-type I-2 was chemically improved by treating it with different concentrations of ethyl methyl sulfonate (EMS). Among seven mutant variants, EMS-6 exhibiting maximal DA activity of 43 μg/ml was selected. The strain was further exposed with l-cysteine HCl to make it resistant against diversion and environmental stress. The conidiospores of selected mutant variant A. oryzae EMS-6 strain were entrapped in calcium alginate beads. Different parameters for immobilization were investigated. The activity was further improved from 44 to 62 μg/ml under optimized conditions (1.5 % sodium alginate, 2 ml inoculum, and 2 mm bead size). The best resistant mutant variable exhibited over threefold increase in DA activity (62 μg/ml) than did wild-type I-2 (21 μg/ml) in the reaction mixture. From the results presented in the study, it was observed that high titers of DA activity in vitro could effectively be achieved by the EMS-induced mutagenesis of filamentous fungus culture used.     

Enhancing <Emphasis Type="SmallCaps">l</Emphasis>-Lysine Production of Beet Molasses by Engineered <Emphasis Type="Italic">Escherichia coli</Emphasis> Using an In Situ Pretreatment Method

Reducing the viscosity of molasses environmentally and selectively removing the harmful ingredients for microbes are the keys to promoting the bioavailability of molasses. A simple and environmental in situ pretreatment method integrating surfactants and alkali was developed to reduce the viscosity of molasses prior to l-lysine production using Escherichia coli ZY0217. Adding activated carbon and modified orange peel based on the in situ pretreatment process effectively removed pigments and excessive zinc in the molasses and also significantly increased the cell growth and l-lysine yield from E. coli ZY0217. The experimental results showed that a mixture of secondary alkane sulfonate, an anionic surfactant, and HodagCB-6, a non-ionic surfactant, effectively reduced the viscosity of the molasses more so than any single surfactant. When the surfactant mixture was added at a concentration of 0.04 g/L to the molasses, the ω value was 0.4, and when ammonia was added at 0.6 %, the lowest viscosity of 705 mPa?·?s was obtained. Further, 91.5 % of the color and 86.68 % of the original levels of zinc were removed using an activated carbon and modified orange peel treatment on the molasses with the lowest viscosity, which further promoted cell growth and l-lysine production. In the fed-batch cultivation process, the l-lysine concentration achieved using a constant-speed feeding strategy was 45.89 g/L, with an l-lysine yield of 27.18 %, whereas the l-lysine yield from untreated molasses was only 10.13 %. The increase in l-lysine yield was related to the reduced viscosity and the detoxification of the molasses. Lastly, the pretreatment was found to significantly enhance the conversion of sugars in the molasses to l-lysine.     

The Study of Intermolecular Interactions for <Emphasis Type="SmallCaps">dl</Emphasis>-Alanine and Glycine in Aqueous <Emphasis Type="SmallCaps">l</Emphasis>(+)-Arabinose Solutions at Different Temperatures and Ambient Pressure,Using the Volumetric Approach

Density measurements are used to calculate the apparent molar volumes V_φ, limiting apparent molar volumes \(V_{\varphi }^{0}\), limiting apparent molar volumes of transfer, \(\Delta_{\text{t}} V_{\varphi }^{0}\), limiting apparent molar expansibilities, \(E_{\varphi }^{0}\), and hydration numbers n_H, for dl-alanine and glycine in aqueous solutions of l(+)-arabinose at T?=?293.15 to 313.15 K. To obtain the limiting apparent molar volume, the V_φ values are extrapolated to zero molality using the linear form of the Redlich–Meyer equation. Also, the limiting apparent molar volumes of transfer, \(\Delta_{\text{t}} V_{\varphi }^{0}\), for the amino acids, from water to aqueous l(+)-arabinose solutions, are calculated from the \(V_{\varphi }^{0}\) values. The limiting apparent molar expansibility, \(E_{\varphi }^{0}\), values have been obtained from the first derivative of limiting apparent molar volumes with respect to temperature. Also the hydration number, n_H, for both amino acids in the ternary solutions are estimated. Possible solute–solvent interactions in the studied ternary systems are discussed.     

Study on the Interaction of Nicotinic Acid with <Emphasis Type="SmallCaps">l</Emphasis>-Phenylalanine in Buffer Solution by Heat Capacity Measurements at Various Temperatures

The interaction between nicotinic acid (NA) and l-phenylalanine (Phe) was studied in aqueous phosphate buffer solutions (pH = 7.35) by differential scanning calorimetry. Heat capacities of nicotinic acid–buffer, l-phenylalanine–buffer, and nicotinic acid–l-phenylalanine–buffer mixtures were determined at (283.15, 288.15, 293.15, 298.15, 303.15, 308.15, 313.15, 318.15 and 323.15) K using the microdifferential scanning calorimeter SCAL-1 (Pushchino, Russia). The apparent molar heat capacities, ^? C _p , of nicotinic acid in buffer solution and in buffer 0.0216 mol·kg^?1 amino acid solutions were evaluated. The concentration of NA was varied from (0.0106–0.0701) mol·kg^?1. The interaction of NA with Phe is accompanied by complex formation. The partial molar heat capacities of transfer of nicotinic acid from buffer to buffer amino acid solutions are positive. The results are discussed in terms of various interactions operating in this system.     

The chemo- enzymatic synthesis of labeled <Emphasis Type="SmallCaps">l</Emphasis>-amino acids and some of their derivatives

This review compiles the combined chemical and enzymatic synthesis of aromatic l-amino acids (l-phenylalanine, l-tyrosine, l-DOPA, l-tryptophan, and their derivatives and precursors) specifically labeled with carbon and hydrogen isotopes, which were elaborated in our research group by the past 20 years. These compounds could be then employed to characterize the mechanisms of enzymatic reactions via kinetic and solvent isotope effects methods.     

Establishing a Mathematical Equations and Improving the Production of <Emphasis Type="SmallCaps">l</Emphasis>-<Emphasis Type="Italic">tert</Emphasis>-Leucine by Uniform Design and Regression Analysis

l-tert-Leucine (l-Tle) and its derivatives are extensively used as crucial building blocks for chiral auxiliaries, pharmaceutically active ingredients, and ligands. Combining with formate dehydrogenase (FDH) for regenerating the expensive coenzyme NADH, leucine dehydrogenase (LeuDH) is continually used for synthesizing l-Tle from α-keto acid. A multilevel factorial experimental design was executed for research of this system. In this work, an efficient optimization method for improving the productivity of l-Tle was developed. And the mathematical model between different fermentation conditions and l-Tle yield was also determined in the form of the equation by using uniform design and regression analysis. The multivariate regression equation was conveniently implemented in water, with a space time yield of 505.9 g L^?1 day^?1 and an enantiomeric excess value of >99 %. These results demonstrated that this method might become an ideal protocol for industrial production of chiral compounds and unnatural amino acids such as chiral drug intermediates.     

Determination of tyrosine using <Emphasis Type="Italic">p</Emphasis>-sulfonated calix[<Emphasis Type="Italic">n</Emphasis>]arene modified electrode

Cyclic voltammetry was used to detect p-sulfonated calix[n]arenas (SCnA) how to immobilize on gold surface. P-sulfonated calix[n]arenes not only increased the specific surface area of the modified electrode, but also improved the enrichment ability of tyrosine. This led to a significant increase in peak current, and improved the sensitivity of tyrosine determination on the p-sulfonated calix[n]arenes-modified electrode. The modified electrode showed good catalytic ability of l-tyrosine oxidation reaction. The peak current of l-tyrosine increased and the oxidation peak potential shifted negatively with cavity size of the SCnA-modified electrode, which indicated that the catalytic ability of the modified electrode to l-tyrosine oxidation reaction was also enhanced. For the tyrosine guest molecule, the order of electrochemical activity and magnitude of catalytic ability of the oxidation reaction of the three modified gold electrodes was as follows: SC4A?>?SC6A?>?SC8A. The electrode had high selectivity and stability for the determination of tyrosine, with a wide linear range, low detection limit and high sensitivity at different concentrations under different pH values. This resulted in an accurate, fast, sensitive electrochemical method for the determination of tyrosine. The sensor was used for the determination of tyrosine in human urine with satisfactory results.     

A Comprehensive Review on <Emphasis Type="SmallCaps">l</Emphasis>-Asparaginase and Its Applications

l-asparaginase (LA) catalyzes the degradation of asparagine, an essential amino acid for leukemic cells, into ammonia and aspartate. Owing to its ability to inhibit protein biosynthesis in lymphoblasts, LA is used to treat acute lymphoblastic leukemia (ALL). Different isozymes of this enzyme have been isolated from a wide range of organisms, including plants and terrestrial and marine microorganisms. Pieces of information about the three-dimensional structure of l-asparaginase from Escherichia coli and Erwinia sp. have identified residues that are essential for catalytic activity. This review catalogues the major sources of l-asparaginase, the methods of its production through the solid state (SSF) and submerged (SmF) fermentation, purification, and characterization as well as its biological roles. In the same breath, this article explores both the past and present applications of this important enzyme and discusses its future prospects.     

Bioactive compounds from <Emphasis Type="Italic">Smilax excelsa</Emphasis> L.

Phytochemical investigation of EtOAc extract of Smilax excelsa has led to isolation and structure elucidation of five compounds. The structures of these compounds are established by different spectroscopic techniques including 1D and 2D-NMR, HRMS and ECD spectroscopy. The compounds were: solanesol (1), violasterol A (2), trans-resveratrol (3), 5-O-caffeoylshikimic acid (4) and 6-O-caffeoyl-β-d-fructofuranosyl-(2-1)-α-d-glucopyranoside (5). The configuration of compound 2 was established by electronic circular dichroism (ECD) spectroscopy. Meanwhile the cytotoxicity and antibacterial activity of the compounds were evaluated by MTT and MIC assays. Compounds 1 and 2 showed promising inhibition on MCF-7 cell line with IC₅₀ of 161.6 and 190.0 µM, respectively. Also compounds 2 and 3 illustrated activity against Staphylococcus aureus with MIC values of 142.5 and 136.9 µM, respectively.     

Thermodynamics of the Protonation of an Analogue of Glyphosate, <Emphasis Type="Italic">N</Emphasis>-(phosphonomethyl)-<Emphasis Type="SmallCaps">l</Emphasis>-proline,in Aqueous Solutions

N-(phosphonomethyl)-l-proline is an analogue of glyphosate. The protonation for N-(phosphonomethyl)-l-proline was studied by potentiometry, calorimetry, ³¹P NMR spectroscopy and quantum chemical calculations to further understand the protonation process of glyphosate. The results confirmed that the order of successive protonation sites of totally deprotonated N-(phosphonomethyl)-l-proline are a phosphonate oxygen, amino nitrogen, and finally the carboxylate oxygen. The results can improve the understanding of the biological activity of these types of molecules in solution.     

Production of 6-<Emphasis Type="Italic">O</Emphasis>-<Emphasis Type="SmallCaps">l</Emphasis>-Ascorbyl Palmitate by Immobilized <Emphasis Type="Italic">Candida antarctica</Emphasis> Lipase B

l-ascorbyl palmitate (ASP) is an oil-soluble derivative of ascorbic acid which is used extensively in food, cosmetics industry, and medical hygiene. Enzymatic synthesis of ascorbyl palmitate in tert-butyl alcohol was carried out using indigenously immobilized lipase preparation PyCal with ascorbic acid and palmitic acid as starting material. The developed batch process under optimized reaction conditions resulted in conversion of 90% with relatively shorter reaction time of 6 h. Continuous process in packed bed reactor gave conversion of 50% with space time yield of 15.46 g/L/h which was found to be higher than the reported literature on enzymatic synthesis of ascorbyl palmitate. The immobilized lipase used in the present work showed good reusability. Characterization of formed ascorbyl palmitate was carried out by FTIR, MS/MS, H¹-NMR, and C¹³-NMR. The enzymatic process resulted in selective synthesis of 6-O-l-ascorbyl palmitate with purity of 98.6% and no side product formation. The use of underivatized starting materials, high space time yield of 15.46 g L^?1 h^?1, high recyclability of catalyst, and no by-product formation make the overall process highly efficient and clean in terms of energy consumption and waste generation, respectively. The optimized reaction parameters for ascorbyl palmitate synthesis in the present study can be used as a useful reference for industrial synthesis of fatty acid esters of ascorbic acid by enzymatic route.     

Enhancing the Production of <Emphasis Type="SmallCaps">d</Emphasis>-Mannitol by an Artificial Mutant of <Emphasis Type="Italic">Penicillium</Emphasis> sp. T2-M10

d-Mannitol belongs to a linear polyol with six-carbon and has indispensable usage in medicine and industry. In order to obtain more efficient d-mannitol producer, this study has screened out a stable mutant Penicillium sp. T2-M10 that was isolated from the initial d-mannitol-produced strain Penicillium sp.T2-8 via UV irradiation as well as nitrosoguanidine (NTG) induction. The mutant had a considerable enhancement in yield of d-mannitol based on optimizing fermentation. The production condition was optimized as the PDB medium with 24 g/L glucose for 9 days. The results showed that the production of d-mannitol from the mutant strain T2-M10 increased 125% in contrast with the parental strain. Meanwhile, the fact that d-mannitol is the main product in the mutant simplified the process of purification. Our finding revealed the potential value of the mutant strain Penicillium sp. T2-M10 to be a d-mannitol-producing strain.     

Concise synthesis of two natural steroidal glycosides isolated from <Emphasis Type="Italic">Allium schoenoprasum</Emphasis>

Two natural steroidal glycosides, diosgenin 3-O-α-l-rhamnopyranosyl-(1→2)-[β-d-glucopyranosyl-(1→4)]-β-d-glucopyranoside (1) and laxogenin 3-O-α-l-rhamnopyranosyl-(1→2)-[β-d-glucopyranosyl-(1→4)]-β-d-glucopyranoside (2) with important cytotoxic activity against the HCT 116 and HT-29 human colon cancer cell lines have been efficiently synthesized via straightforward sequential glycosylation reaction with the combined use of N-phenyltrifluoroacetimidates and trichloroacetimidates donors at room temperature. All structures of the synthesized new compounds were identified by ¹H NMR, ¹³C NMR and HRMS spectra.     

Asymmetric polymerisation of substituted phenylacetylene using chiral Rh(2,5-norbornadiene)(<Emphasis Type="SmallCaps">l</Emphasis>-proline) catalyst

The Rh(nbd)(l-proline) (nbd = 2,5-norbornadiene) catalyst was synthesised with l-proline as ligand. The achiral monomer phenylacetylene, having two hydroxyl groups and a dodecyl group (DoDHPA), was polymerised for the first time using an isolated chiral Rh(nbd)(l-proline) as catalyst to afford polymers of M_r of 28.5 × 10⁴ and 36.2 × 10⁴. The resulting polymers exhibited the Cotton effect at wavelengths assignable to the main chain, indicating that the polymers adopted one-handed helical conformation. These findings suggest that the rhodium complex with chiral amine may be the true active species for helix-sense-selective polymerisation (HSSP) of DoDHPA.     

Development of an Amperometric Biosensor Platform for the Combined Determination of <Emphasis Type="SmallCaps">l</Emphasis>-Malic,Fumaric, and <Emphasis Type="SmallCaps">l</Emphasis>-Aspartic Acid

Three amperometric biosensors have been developed for the detection of l-malic acid, fumaric acid, and l -aspartic acid, all based on the combination of a malate-specific dehydrogenase (MDH, EC 1.1.1.37) and diaphorase (DIA, EC 1.8.1.4). The stepwise expansion of the malate platform with the enzymes fumarate hydratase (FH, EC 4.2.1.2) and aspartate ammonia-lyase (ASPA, EC 4.3.1.1) resulted in multi-enzyme reaction cascades and, thus, augmentation of the substrate spectrum of the sensors. Electrochemical measurements were carried out in presence of the cofactor β-nicotinamide adenine dinucleotide (NAD⁺) and the redox mediator hexacyanoferrate (III) (HCFIII). The amperometric detection is mediated by oxidation of hexacyanoferrate (II) (HCFII) at an applied potential of + 0.3 V vs. Ag/AgCl. For each biosensor, optimum working conditions were defined by adjustment of cofactor concentrations, buffer pH, and immobilization procedure. Under these improved conditions, amperometric responses were linear up to 3.0 mM for l-malate and fumarate, respectively, with a corresponding sensitivity of 0.7 μA mM^?1 (l-malate biosensor) and 0.4 μA mM^?1 (fumarate biosensor). The l-aspartate detection system displayed a linear range of 1.0–10.0 mM with a sensitivity of 0.09 μA mM^?1. The sensor characteristics suggest that the developed platform provides a promising method for the detection and differentiation of the three substrates.     

Molecular and crystal structure of 15-acetylsongorine and songoramine isolated from <Emphasis Type="Italic">Aconitum szechenyianum</Emphasis> Gay

Two napelline skeletal diterpenoid alkaloids 15-acetylsongorine, C₂₄H₃₃NO₄ I, and songoramine, C₂₂H₂₉NO₃ II, were first isolated from the roots of Aconitum Szechenyianum Gay. The crystal structures were determined by X-ray single-crystal diffraction analysis. The crystal I is the triclinic system with space group P1 having unit cell parameters of a = 9.360(8) Å, b = 11.593(9) Å, c = 11.830(16) Å, α = 113.223(15)°, β = 105.950(16)°, γ = 101.296(12)°, and Z = 2. Hydrogen bonds O–H···O and O–H···N joint the molecules into dimer. The crystal II belongs to the orthorhombic system with space group P2₁2₁2₁ having unit cell parameters of a = 8.950(2) Å, b = 13.272(3) Å, c = 15.454(4) Å and Z = 4. The O–H···O hydrogen bonding interaction links the molecule into linear chains. The distortion of rings of compound I and II were evaluated by calculation of the Cremer and Pople puckering parameters. The presence of the C–O–C bond in the compound II results in the changes of ring conformations compared with that of the compound I.     

Capillary Electrophoresis-Based Enzyme Assay for Nicotinamide <Emphasis Type="Italic">N</Emphasis>-Methyltransferase

A capillary electrophoresis method was developed for the determination of the activity of nicotinamide N-methyltransferase. Separation of the substrate nicotinamide and the product N-methylnicotinamide as well as the cofactor S-(5′-adenosyl)-l-methionine and its metabolite S-(5′-adenosyl)-l-homocysteine were achieved in a 50/60.5 cm fused-silica capillary using 100 mM sodium phosphate buffer, pH 3.0 as electrolyte and an applied voltage of 25 kV. Analyte detection was carried out at 193 nm. Using lidocaine as internal standard the method was validated for nicotinamide and N-methylnicotinamide with regard to range, limit of detection, limit of quantitation, repeatability, intermediate precision and accuracy. The assay was applied to the determination of the Michaelis–Menten kinetic parameters of the enzyme.     

Synthesis,crystal structure,and thermodynamics of a high-nitrogen copper complex with <Emphasis Type="Italic">N</Emphasis>, <Emphasis Type="Italic">N</Emphasis>-bis-(1(2)H-tetrazol-5-yl) amine

A new high-nitrogen complex [Cu(Hbta)₂]·4H₂O (H₂bta = N,N-bis-(1(2)H-tetrazol-5-yl) amine) was synthesized and characterized by elemental analysis, single crystal X-ray diffraction and thermogravimetric analyses. X-ray structural analyses revealed that the crystal was monoclinic, space group P2(1)/c with lattice parameters a = 14.695(3) Å, b = 6.975(2) Å, c = 18.807(3) Å, β = 126.603(1)°, Z = 4, D _c = 1.888 g cm^?3, and F(000) = 892. The complex exhibits a 3D supermolecular structure which is built up from 1D zigzag chains. The enthalpy change of the reaction of formation for the complex was determined by an RD496–III microcalorimeter at 25 °C with the value of ?47.905 ± 0.021 kJ mol^?1. In addition, the thermodynamics of the reaction of formation of the complex was investigated and the fundamental parameters k, E, n, \( \Updelta S_{ \ne }^{{{\uptheta}}} \), \( \Updelta H_{ \ne }^{{{\uptheta}}} \), and \( \Updelta G_{ \ne }^{{{\uptheta}}} \) were obtained. The effects of the complex on the thermal decomposition behaviors of the main component of solid propellant (HMX and RDX) indicated that the complex possessed good performance for HMX and RDX.     

Effect of Amino Acids and Sodium Chloride on <Emphasis Type="SmallCaps">d</Emphasis>-Sorbitol in Aqueous Solutions at Different Temperatures: Volumetric and Acoustic Approach

Apparent molar volumes and apparent molar compressibilities for d-sorbitol in (0.05, 0.1, 0.2 and 0.3) mol·kg^?1 aqueous solutions of l-alanine, l-cysteine and l-histidine and NaCl have been determined from measurements of solution density at T?=?(288.15, 298.15, 308.15 and 318.15) K and sound velocity at T?=?298.15 K, as a function of the concentration of the sugar alcohol. The data were used to obtain the limiting apparent molar volumes, limiting apparent molar compressibilities and the corresponding transfer parameters. Limiting apparent molar expansibilities and their second order derivatives and volume interaction coefficients were also estimated. These parameters are discussed in terms of d-sorbitol and co-solute (amino acid or sodium chloride) interactions in aqueous solutions.     

A sensitive electrochemical sensor for lead based on gold nanoparticles/nitrogen-doped graphene composites functionalized with <Emphasis Type="SmallCaps">l</Emphasis>-cysteine-modified electrode

Glassy carbon electrodes (GCEs) modified with l-cysteine (l-cys)/gold nanoparticles (AuNPs)/nitrogen-doped graphene (NG) composite were prepared to fabricate a novel electrochemical sensor for lead. AuNPs were uniformly dispersed into NG and l-cys was successfully decorated on AuNPs through the S–Au bond. The l-cys/AuNPs/NG exhibited a well-distributed nanostructure and high responsivity toward Pb(II). The results indicated that l-cys/AuNPs/NG/GCE exhibited the highest peak current, reflecting that the l-cys/AuNPs/NG composites showed the best response signal toward Pb²⁺. Under optimized conditions, a linear relationship between the current intensity and Pb²⁺ concentration was obtained in a range of 0.5–80 μg L^?1 with a detection limit of 0.056 μg L^?1 (S/N = 3). The analytical interference procedure and practical application were investigated using the prepared electrode, which exhibited an acceptable result.