Significance of N source (urea vs. NH4NO3) and Ni supply for growth,urease activity and nitrogen metabolism of zucchini (Cucurbita pepo convar. giromontiina)

The effect of Ni supply on growth and N metabolism of zucchini plants grown with either NH4NO3 or urea as sole N source was investigated. Dry matter production of plants grown with NH4NO3 was not affected by the Ni status, while urea-based nutrition led to reduced growth, particularly when plants were grown without Ni supplementation. The activity of urease, which requires Ni for activation, was hardly detectable in leaves and roots of plants grown without supplementary Ni irrespective of N source. Low-Ni urea-grown plants were chlorotic, accumulated large amounts of urea and had lower amino acid contents indicating impaired usage of the N supplied. The lack of urease activation made these plants metabolically N deficient. The amino acid pools of plants grown with NH4NO3 was not markedly affected by the Ni supply, although these plants accumulated endogenous urea in their leaves when grown without supplementary Ni. In urea-grown plants the glutamine content was considerably increased by Ni supply, indicating that the efficient use of urea N is Ni (urease) dependant. Based on growth and urease activity, a critical Ni concentration in the leaves of around 100 µg kg-1 can be deduced. These results confirm the necessity of Ni for urease activation and thus for growth of plants on urea-based media, as well as for recycling endogenous urea.     

Stereospecific synthesis of l-2-amino-3-(7-methoxy-4-coumaryl)propionic acid, an alternative to tryptophan in quenched fluorescent substrates for peptidases

A high-yielding synthesis of the highly fluorescent amino acid l-2-amino-3-(7-methoxy-4-coumaryl)propionic acid (l-Amp) has been developed via (2R)-bornane-10,2-sultam glycinate. l-Amp facilitates the synthesis of sensitive fluorogenic proteinase substrates with N-terminal solubilising or reactive groups.     

Stereospecific synthesis of l-2-amino-3-(7-methoxy-4-coumaryl)propionic acid,an alternative to tryptophan in quenched fluorescent substrates for peptidases

Summary A ligh-yielding synthesis of the highly fluorescent amino acidl-2-amino-3-(7-methoxy-4-coumaryl)propionic acid (l-Amp) has been developed via (2R)-bormane-10,2-sultam glycinate.l-Amp facilitates the synthesis of sensitive fluorogenic proteinase substrates with N-terminal solubilising or reactive group.     

Synthesis of non-proteinogenic (D)- or (L)-amino acids by asymmetric hydrogenation

Summary Non-proteinogenic amino acids play an increasing role in oligopeptide chemistry. Their pharmacological and chemical properties, caused by D-configuration and unnatural residues, are more and more used for drug design. Different methods of asymmetric synthesis have been developed during the last decade to prepare unusual amino acids. One of them, the asymmetric hydrogenation of dehydroamino aids catalyzed by chiral rhodium (I) complexes, will be described. A series of examples, D- and L-configured, like naphthyl-, thienyl-, furyl-, and pyridylalanines, as well as phenylalanines substituted by chlorine, fluorine, p-nitro, p-methyl, p-trifluoromethyl, p-isopropyl, and p-tert-butyl have been prepared and characterized. Some analytical data like melting points and values of optical rotation are summarized in tables.Abbreviations (–)-DIOP (4R,5R)-4,5-Bis(diphenylphosphinomethyl)-2,2-dimethyl-1,3-dioxolane - (–)-BPPM (2S,4S)-N-tert-Butoxycarbonyl-4-diphenylphosphino-2-diphenylphosphinomethyl-pyrrolidine - Ph--glup Phenyl 4,6-O-(R)-benzylidene-2,3-O-bis(diphenylphosphino)--D-glucopyranoside - DuPHOS 1,2-bis-(phospholano)benzene - PROPRAPHOS 2,3-O,N-bis(diphenylphosphino)-1-(naphthoxy)-2-hydroxy-3-isopropylamino propane - PINDOPHOS 2,3-O,N-bis(diphenylphosphino)-1-(4-indolyloxy)-2-hydroxy-3-isopropylamino propane     

Preparation of (25R)- and (25S)-26-functionalized steroids as tools for biosynthetic studies of cholic acids

A new synthesis of both epimeric forms of 26-cholestanoic acids and 26-alcohols containing a 3beta-hydroxy-Delta(5)- or a Delta(4)-3-keto-functionality in ring A is described starting from stigmasterol or (20S)-3beta-acetoxy-pregn-5-en-20-carboxylic acid. The obtained compounds are useful as standards for studies of cholic acids. Construction of the side chain was achieved by linkage of steroidal 23-iodides to sulfones prepared from (2R)- and (2S)-3-hydroxy-2-methylpropanoates. Oxidation of intermediate 26-alcohols into the corresponding carboxylic acids ensuring preservation of stereochemistry at C-25 and functional groups in the cyclic part was achieved with sodium chlorite catalyzed by TEMPO and bleach.     

Synthesis and structure of a vesicular assembly of dialkylammonium-type amphiphile carrying L-3-(3-N-ethylcarbazolyl)alanine

Summary A novel unnatural amino acid, L-3-(3-N-ethylcarbazolyl)alanine, was synthesized and used for synthesis of a chiral dialkylammonium-type amphiphile. The synthetic amphiphile formed a vesicular assembly in aqueous dispersion showing gel-liquid crystalline phase transition around 25 °C. The bilayer membrane composed of the amphiphile was nearly free from excimer as evidenced by fluorescence spectroscopy.     

1-(t-Butyldimethylsilyloxy) benzotriazole (TBDMS-OBt): A new and novel reagent for the synthesis of peptides

Synthesis and use of 1-(t-butyldimethylsilyloxy)benzotriazole (TBDMS-OBt) in the coupling of Fmoc-amino acid chlorides to amino free amino acid esters in homogeneous solution phase is described. The coupling required no addition of base and was fast and racemization free. Work up and isolation of products were easy. Yield, purity and ¹H NMR analysis of peptides, synthesised by this method, were satisfactory.     

1-(t-Butyldimethylsilyloxy) benzotriazole (TBDMS-OBt): A new and novel reagent for the synthesis of peptides

Summary Synthesis and use of 1-(t-butyldimethylsilyloxy)benzotriazole (TBDMS-OBt) in the coupling of Fmoc-amino acid chlorides to amino free amino acid esters in homogeneous solution phase is described. The coupling required no addition of base and was fast and racemization free. Work up and isolation of products were easy. Yield, purity and¹H NMR analysis of peptides, synthesised by this method, were satisfactory.     

The extent of N epsilon-(carboxymethyl)lysine formation in lens proteins and polylysine by the autoxidation products of ascorbic acid.

The autoxidation of ascorbic acid (ASA) leads to the formation of compounds which are capable of glycating and crosslinking proteins in vitro. When the soluble crystallins from bovine lens were incubated with ASA in the presence of sodium cyanoborohydride, a single major adduct was observed, whose appearance correlated with the loss of lysine. When polylysine was reacted with equivalent amounts of ASA under the same conditions, this product represented half of the total lysine content after four weeks of incubation at 37 degrees C. This adduct was isolated and identified as N epsilon-(carboxymethyl)lysine (CML) by TLC, GC/MS and amino acid analysis. Several oxidation products of ASA were each reacted with polylysine in the presence of sodium cyanoborohydride to identify the reactive species. CML was the major adduct formed with either ASA and dehydroascorbic acid (DHA). Markedly diminished amounts were seen with L-2,3-diketogulonic acid (DKG), and L-threose, while no CML was formed with L-threo-pentos-2-ulose (L-xylosone). In the absence of sodium cyanoborohydride the yield of CML was similar with each of the ASA autoxidation products and required oxygen. Reactions with [1-14C]ASA gave rise to [14C]CML, but only with NaCNBH3 present. At least two routes of CML formation appear to be operating depending upon whether NaCNBH3 is present to reduce the putative Schiff base formed between lysine and DHA.     

l-3-(3′-Carboxy-4′-hydroxyphenyl)alanine (3-carboxytyrosine) in seeds of Neonotonia wightii (Glycine wightii) and its possible systematic significance

l-3-(3′-Carboxy-4′-hydroxyphenyl)alanine (3-carboxytyrosine) constitutes 3% of the seeds of Neonotonia wightii (Glycine wightii); it has also been detected in the seeds of 3 species belonging to 2 other genera of the Glycineae. The systematic significance of these findings is discussed.     

Antide B,an antagonist of LHRH with cis-3-(4-pyrazinylcarbonylaminocyclohexyl)alanine in position 5

Summary Several LHRH antagonists with trans-3-(4-pyrazinylcarbonylaminocyclohexyl)alanine (trans-PzACAla) in the position 5 were synthesized and their antiovulatory activity was compared with the activity of the analogs containing cis-PzACAla in this position. In all cases cis-isomer produced more potent analogs. Introduction of cis-PzACAla in the position 5 of Antide gave Antide B which completely inhibits ovulation at a dose of 0.5µg/rat. Antide B releases negligible histamine (ED₅₀ = 104µg/mL), and has excellent solubility in water. Also, an improved synthesis of cis-PzACAla is reported, involving the hydrogenation of 4-aminophenylalanine on a rhodium catalyst to give the desired cis-isomer with a 53% yield.Abbreviations Cpa 3-(4-chlorophenyl)alanine - ILys N-isopropyllysine - Nal 3-(2-naphthyl)alanine - NicLys N-nicotinoyllysine - Pal 3-(3-pyridyl)alanine - PicLys N-picolinoyllysine - PzACAla 3-(4-pyrazinylcarbonylaminocyclohexyl)alanine - Qal 3-(3-quinolyl)alanine     

Inhibition of COX-1 activity and COX-2 expression by 3-(4'-geranyloxy-3'-methoxyphenyl)-2-trans propenoic acid and its semi-synthetic derivatives

Oxyprenylated naturally occurring cinnamic acids displayed efficient and promising biological activities. Aim of this study was to characterize the effects of 3-(4'-geranyl-3'-methoxy)phenyl-2-trans propenoic acid and its selected semi-synthetic analogues, on COX-2 expression and activity, and on COX-1 activity, in purified systems or in whole cell systems. The anti-inflammatory activity of title compounds (1) was tested as inhibition of COX-2 on isolated monocytes stimulated with LPS (10 μg/ml). COX-2 expression was completely suppressed when monocytes were incubated with 100 μM of 3-(4'-geranyl-3'-methoxy)phenyl-2-trans propenoic acid (1) or 3-(4'-isopentenyloxy)phenyl-2-trans propenoic acid (4). Moreover compounds (1) and (4) inhibit dose-dependently LPS-induced COX-2 expression.     

Enzymatic synthesis and characterization of N5-(carboxymethyl)-L-ornithine and N6-(carboxymethyl)-L-lysine

Summary This report describes the enzyme-catalyzed synthesis, characterization, and chromatographic separation of N⁶-(carboxymethyl)-L-lysine and N⁵-(carboxymethyl)-L-ornithine. The two N -(carboxyalkyl)amino acids are formed via a reductive condensation between glyoxylate and the- or-amino groups of lysine and ornithine, respectively. Both reactions are catalyzed by the NADPH-dependent enzyme, N⁵-(carboxyethyl)ornithine synthase [EC 1.5.1.24], found in some strains of the lactic acid bacteriumLactococcus lactis subsp.lactis.     

Copper(II) complexes of N2-alkyl-(S)-amino acid amides as chiral selectors for dynamically coated chiral stationary phases in RP-HPLC

Copper(II) complexes of N²-octyl-(S)-phenylalaninamide (Noc-Phe-NH₂), N²-dodecyl-(S)-phenylalaninamide (Ndo-Phe-NH₂), and N²-octyl-(S)-norleucinamide (Noc-NLeu-NH₂), dynamically adsorbed on a reversed-phase C₁₈ column, were able to perform the direct enantiomeric separation of unmodified amino acids, amino acid amides and esters, hydroxy acids, and dipeptides by elution with aqueous or mixed aqueous-organic solutions containing copper(II) sulphate or acetate. The role played by several parameters in the separation procedure was examined with the copper(II) complex of Noc-Phe-NH₂ [concentration of the copper(II) ion in the eluent, pH and eluent polarity, amount of adsorbed selector]. The separation was shown to occur entirely on the stationary phase. The mechanism of chiral discrimination is discussed in terms of the chromatographic parameters and of the structure of the copper(II) complexes in solution and in the solid state. The chiral stationary phase maintained its separation ability for about 3 months. However, the column could be easily restored by recovering the selector with methanol and repeating the loading procedure. © 1996 Wiley-Liss, Inc.     

Synthesis,biological evaluations and computational studies of N-(3-(-2-(7-Chloroquinolin-2-yl)vinyl) benzylidene)anilines as fungal biofilm inhibitors

In the present investigation, new chloroquinoline derivatives bearing vinyl benzylidene aniline substituents at 2nd position were synthesized and screed for biofilm inhibitory, antifungal and antibacterial activity. The result of biofilm inhibition of C. albicans suggested that compounds 5j (IC₅₀ value?=?51.2?μM) and 5a (IC₅₀ value?=?66.2?μM) possess promising antibiofilm inhibition when compared with the standard antifungal drug fluconazole (IC₅₀?=?40.0?μM). Two compounds 5a (MIC?=?94.2?μg/mL) and 5f (MIC?=?98.8?μg/mL) also exhibited good antifungal activity comparable to standard drug fluconazole (MIC?=?50.0?μg/mL). The antibacterial screening against four strains of bacteria viz. E. coli, P. aeruginosa, B. subtilis, and S. aureus suggested their potential antibacterial activity and especially all the compounds except 5g were found more active than the standard drug ciprofloxacin against B. subtilis. To further gain insights into the possible mechanism of these compounds in biofilm inhibition through the agglutinin like protein (Als), molecular docking and molecular dynamics simulation studies were carried out. Molecular modeling studies suggested the clear role in inhibition of this protein and the resulting biofilm inhibitory activity.     

Determination of N7-(2-hydroxyethyl)guanine by HPLC with electrochemical detection

A method has been developed for the determination of N7-(2-hydroxyethyl)guanine (N7-EtOHGua) via HPLC with electrochemical detection (EC). N7-EtOHGua is the major base adduct formed in DNA upon exposure to ethylene oxide. N7-EtOHGua, released from DNA, was separated from the unmodified nucleobases by chromatography on a reversed-phase column. For electrochemical detection, an amperometric detector cell was used with a glassy carbon working electrode, set at 1.35 V relative to an Ag/AgCl reference electrode. With purified N7-EtOHGua a linear dose-response relation was observed in the range between 0.11 and 13 pmol. The signal-to-noise ratio during analysis of 0.11 pmol N7-EtOHGua was about 8 to 1. Determination of adducts in a series of DNA samples treated with 0.16–10 mM ethylene oxide showed a linear dose-dependent increase in the level of N7-modifications. For DNA samples, the detection limit of this HPLC-EC analysis is 1 N7-EtOHGua per 6 × 10⁶ nucleotides.     

Irreversible Inhibition of High-Affinity [3H]Kainate Binding by a Novel Photoactivatable Analogue: (2'S,3'S,4'R)-2'-Carboxy-4'-(2-Diazo-1-Oxo-3,3,3-Trifluoropropyl)-3'-Pyrrolidinyl Acetate

Abstract: A photolabile trifluoromethyldiazoketone derivative of kainate (KA), (2' S ,3' S ,4' R )-2'-carboxy-4'-(2-diazo-1-oxo-3,3,3-trifluoropropyl)-3'-pyrrolidinyl acetate (DZKA), was synthesized and evaluated as an irreversible inhibitor of the high-affinity KA site on rat forebrain synaptic plasma membranes (SPMs). In the absence of UV irradiation, DZKA preferentially blocked [³H]KA binding with an IC₅₀ of 0.63 µ M , a concentration that produced little or no inhibition at AMPA or NMDA sites. At 100 µ M , however, DZKA inhibited [³H]AMPA and l -[³H]glutamate binding by ∼50%. When examined electrophysiologically in HEK293 cells expressing human KA (GluR6) or AMPA (GluR1) subtypes, DZKA acted preferentially at KA receptors as a weak agonist. DZKA also exhibited little or no excitotoxic activity in mixed rat cortical cultures. Irreversible inhibition was assessed by pretreating SPMs with DZKA (50 µ M ) in the presence of UV irradiation, removing unbound DZKA, and then assaying the reisolated SPMs for radioligand binding. This protocol produced a selective and irreversible loss of ∼50% of the [³H]KA sites. The binding was recoverable in SPMs pretreated with DZKA or UV alone. Coincubation with l -glutamate prevented the loss in [³H]KA binding, suggesting that the inactivation occurred at or near the ligand binding site. These results are consistent with the action of DZKA as a photoaffinity ligand for the KA site and identify the analogue as a valuable probe for future investigations of receptor structure and function.     

Simultaneous determination of N2-(3-aminopropyl)biopterin (oncopterin), biopterin and neopterin by high-performance liquid chromatography with fluorescence detection

A high-performance liquid chromatographic method is described for the simultaneous determination of N²-(3-aminopropyl)biopterin (oncopterin, a newly found natural pteridine in urine from cancer patients), biopterin, and neopterin in urine. For the detection and quantification of the compounds, fluorometry was used. Using Develosil ODS K-5 and Develosil ODS HG-5 reversed-phase columns and a Nucleosil 100-5SA strong cation-exchange column, oncopterin, biopterin, and neopterin in urine were completely separated and assayed simultaneously by fluorescence detection. Similar values of oncopterin were obtained using each of the three columns, and the Develosil ODS K-5 reversed-phase column gave the most satisfactory separation. The sensitivity was high enough to measure 1 pmol of each pteridine. The HPLC method was highly reproducible. Our preliminary results indicate that oncopterin could be a most sensitive marker for cancer.