Synthesis and Crystal Structure of N-[(2S)-4-bromo-2-(L-menthyloxy)-5-oxo-2,5-dihydro-3-furyl]-L-leucine

Starting from（5S）-（L-menthyloxy）-3,4-dibromo-5H-furan-2-one and L-leucine,the title compound N-[（2S）-4-bromo-2-（L-menthyloxy）-5-oxo-2,5-dihydro-3-furyl]-L-leucine 5（C20H32BrNO5,Mr = 446.37） was obtained in one-pot process via the tandem Michael addition-elimination reaction in the presence of potassium hydroxide.The chemical structure and absolute configuration of the title compound were confirmed via rotation,UV-Vis,FT-IR,1H NMR,13C NMR,MS and elemental analysis,especially by the X-ray single-crystal diffraction.The crystal crystallizes in an orthorhombic system,space group P212121 with a = 12.5249（16）,b = 19.005（3）,c = 19.719（3） ,V = 4693.7（10） 3,Z = 8,Dc = 1.263 g/m3,μ = 1.778 mm-1,F（000） = 1872,the final R = 0.0617 and wR = 0.1576 for 3967 observed reflections（I 2σ（I））.X-ray analysis reveals that the title compound has two independent molecules in the asymmetric part of the unit cell with the two five-membered furanones being almost planar.The essential part of the electron delocalization is concentrated in the N（1）,C（3）,C（1）,C（37） and O（7） region and N（2）,C（28）,C（27）,C（30） and O（4） region in the other molecule respectively,but does not take place at the expense of delocalization within the ester function.

STUDIES ON THE ASYMMETRIC DECOMPOSITION OF D,L-LEUCINE INDUCED BY ^(60)Co γ-IRRADIATION

Sublimed D-,L-,and DL-leucine under vacuum were subjected to ^(60)Co γ-irradiation with different doses.The enantiomeric compositions of samples were determined by chirasil- val capillary column gas chromatography,and the relative yields of radiolytic products(H_2, CO_2 and NH_3)were measured by packed column gas chromatography.The obtained data showed the peak area ratios D/L of DL-leucine after different absorbed doses of irradiation are all the same as that of samples with no irradiation,within the limit of error.So are the ratios for the separately irradiated D-and L-leuicne.This suggests that no asymmetric decom- position was found.The relative yields D/L of radiolytic products(H_2,CO_2 and NH_3)are very close to 1.00,indicating that the D-leucine cleavage rate was the same with that of the L-leucine.From the view of dissipative structure,an explanation for the result was presented.

Microwave-Assisted Facile and Rapid Esterification of Amino Acids I: Esterification of L-Leucine from Batch to Flow Processes and Scale-Up

We herein report our studies into the effects of microwave irradiation on the solvent-free esterification of L-leucine with alcohols. In the absence of solvent, microwave irradiation accelerated the reaction compared to conventional oil bath heating. Measurement of the dielectric properties under the reaction conditions revealed that the high dielectric loss factor of the reaction mixture containing L-leucine, n-butanol, and a p-toluene sulfonic acid catalyst could be attributed to the acceleration of the reaction. The depth of microwave penetration into the reaction mixture as derived from the in-situ measurement of the dielectric properties was ~13 mm, which suggested that a thinner reaction vessel was favorable for the esterification of L-leucine. In addition to the batch reaction using a desktop microwave reactor, two types of flow reaction were also performed using a desktop tubular reactor and a semi-bench-scale tubular reactor. These flow reactions also exhibited high performances, thus allowing the scale-up of this reaction system for industrial use.

Modifications of L-Glutamine and L-Leucine Transport in Proliferating Lymphocytes

Lymphocytes respond to mitogens that stimulate proliferation by increasing theirs metabolic activity. In this study we investigate L-Glutamine and L-Leucine uptake as markers of cell response to Concavalina A (ConcaA) stimulation, using a high-resolution flow technique. We found that lymphocytes induced to blast transformation enhanced rate and efficiency of amino acid uptake during cell proliferation. Considering that increases in transport is the first quantifiable response of cells during malignant transformation, amino acid uptake could also be useful as an early marker of malignancy.

L-leucine stimulates glutamate dehydrogenase activity and glutamate synthesis by regulating mTORC1/SIRT4 pathway in pig liver

The liver is the most essential organ for the metabolism of ammonia, in where most of ammonia is removed by urea and glutamine synthesis. Regulated by leucine, glutamate dehydrogenase(GDH) catalyzes the reversible inter-conversion of glutamate to ammonia. To determine the mechanism of leucine regulating GDH, pigs weighing 20 ± 1 kg were infused for 80 min with ammonium chloride or alanine in the presence or absence of leucine. Primary pig hepatocytes were incubated with or without leucine. In the in vivo experiments with either ammonium or alanine as the nitrogen source, addition of leucine significantly inhibited ureagenesis and promoted the production of glutamate and glutamine in the perfused pig liver(P < 0.05). Similarly, leucine stimulated GDH activity and inhibited sirtuin4(SIRT4)gene expression(P < 0.01). Leucine could also activate mammalian target of rapamycin complex 1(m TORC1) signaling(P < 0.05), as evidenced by the increased phosphorylation levels of ribosomal protein S6 kinase 1(S6 K1) and ribosomal protein S6(S6). Interestingly, the leucine-induced m TORC1 pathway activation suitably correlated with increased GDH activity and decreased expression of SIRT4.Similar results were observed in primary cultured hepatocytes. Notably, leucine exerted no significant change in GDH activity in SIRT4-deficient hepatocytes(P > 0.05), while m TORC1 signaling was activated.Leucine exerted no significant changes in both GDH activity and SIRT4 gene expression in rapamycin treated hepatocytes(P > 0.05). In conclusion, L-leucine increases GDH activity and stimulates glutamate synthesis from different nitrogen sources by regulating m TORC1/SIRT4 pathway in the liver of pigs.

Effects of “Bioactive” amino acids leucine, glutamate, arginine and tryptophan on feed intake and mRNA expression of relative neuropeptides in broiler chicks

Feed intake control is vital to ensuring optimal nutrition and achieving full potential for growth and development in poultry. The aim of the present study was to investigate the effects of L-leucine, L-glutamate, L-tryptophan and L-arginine on feed intake and the mRNA expression levels of hypothalamic Neuropeptide involved in feed intake regulation in broiler chicks. Leucine, glutamate, tryptophan or arginine was intra-cerebroventricularly （ICV） administrated to 4d-old broiler chicks respectively and the feed intake were recorded at various time points. Quantitative PCR was performed to determine the hypothalamic mRNA expression levels of Neuropeptide Y （NPY）, agouti related protein （AgRP）, pro-opiomelanocortin （POMC）, melanocortin receptor 4 （MC4R） and corticotrophin releasing factor （CRF）. Our results showed that ICV administration of L-leucine （0.15 or 1.5 μmol） significantly （P〈0.05） increased feed intake up to 2 h post-administration period and elevated both hypothalamic NPY and AgRP mRNA expression levels. In contrast, ICV administration of L-glutamate （1.6 μmol） significantly （P 〈 0.05） decreased feed intake 0.25, 0.5 and 2 h post-injection, and increased hypothalamic CRF and MC4R mRNA expression levels. Meanwhile, both L-tryptophan （10 or 100 μg） and L-arginine （20 or 200 μg） had no significant effect on feed intake. These findings suggested that L-leucine and L-glutamate could act within the hypothalamus to influence food intake, and that both orexigenic and anorexigenic Neuropeptide genes might contribute directly to these effects.

Relief of osteoarthritis with a herbal-amino acid supplement: A randomized double-blind placebo controlled trial

A redox active medicinal plant and L-leucine mixture (HLM) was investigated in subjects with established osteoarthritis of the knee in a multi-center, rando- m-ized, placebo-controlled, double-blind clinical trial. A total of 96 subjects with osteoarthritis were enrolled and randomized to either placebo (n = 38) or HLM treatment group (n = 38). The HLM group re- ceived a combination of Uncaria tomentosa (300 mg), Boswellia serrata (200 mg), Lepidium meyenii (1000 mg) and L-Leucine (700 mg) given as 3 capsules once a day. The placebo group received matching capsules with carboxymethylcellulose. The treatment period was 8 weeks, with assessments made at days 7, 14, 28 and 56. The primary outcome was reduction in total WOMAC score. VAS pain, tolerability, investigator assessments, use of rescue medication (acetominophen), and safety assessments of vital signs and laboratory assessments were included. Subject randomization was effective for age, gender and disease severity. In the placebo group 32/38 subjects completed the trial and for HLM 35/38. WOMAC scores (pain, stiffness, physical performance and total) steadily declined over the course of the 8 week study in both groups, but the magnitude was significantly greater for HLM (P < 0.05). Total WOMAC was reduced 46.5% for HLM and 25.4 % for placebo. VAS pain was reduced 21.8% in the placebo group (p < 0.002) but the changes were significantly greater (37.8% p < 0.03) with HLM treatment. Investigator’s global assessment rating of good-excellent was 24/35 (69%) for HLM and 14/32 (44%) for placebo (P = 0.05). Rescue medication consumption and tolerability were comparable for HLM and placebo. No safety issues were evident with either group. As expected a placebo effect was observed, nevertheless HLM was clearly more effective in relieving the symptoms of osteoarthritis. This HLM represents a safe and effective new approach to the management of osteoarthritis symptoms.

高效液相(HPLC)检测5种氨基酸在酮酸制备过程中的应用

过去氨基酸分析必须应用价格昂贵的专一的氨基酸分析仪,近几年来越来越多的人应用高效液相进行氨基酸定量分析。建立一种测定酮酸制备过程中,5种氨基酸原料L-亮氨酸、L-异亮氨酸、L-苯丙氨酸、L-缬氨酸、L-蛋氨酸的液相分析方法。以乙腈-磷酸氢二胺水溶液为流动相,采用HPLC对氨基酸进行含量测定。当乙腈-50Mm磷酸氢二胺水溶液(pH=2.6)=73:27,最适检测波长205nm。经验证,本方法的线性关系良好,其中L-亮氨酸R^(2)=1、L-缬氨酸R^(2)=1、L-异亮氨酸R^(2)=1、L-苯丙氨酸R^(2)=0.9997、L-蛋氨酸R^(2)=0.9996。本方法具有操作便捷、结果准确的特点,且重复性好,适用于氨基酸的快速定量测定。

Synthesis of optically active polymers bearing amino acid moieties by atom transfer radical polymerization

We report here an approach toward the synthesis of optically active polyacrylamide bearing amino acid moieties, poly[Nmethacryloyl L-leucine methyl ester] （PMALM）, with controlled average number molecular weight （Mn） and relatively narrow polydispersity index （PDI, Mw/Mn 〈 1.3） by atom transfer radical polymerization （ATRP） using initiating system methyl 2-bromopropionate/CuBr/tris（2-dimethylaminoethyl） amine. The optical properties of the resulting polymers were evaluated from specific optical rotation value and CD spectra.

Synthesis,Characterization,and DNA Binding Properties of Dinuclear Copper(Ⅱ) Complex [Cu_2(TATP)_2(L-Leu)_2(ClO_4)_2]_2·2H_2O

A dinuclear copper（Ⅱ） complex[Cu2（TATP）2（L-Leu）2（CIO4）2]2·2H2Owas synthesized and characterized, where, TATP=1,4,8,9-tetraazatriphenylene, and L-Leu=L-leucinate. The complex was crystallized in the triclinic space group P1, with two independent molecules in a unit cell. Two Cu（Ⅱ） ions in each complex [Cu2（TATP）2（L-Leu）2（CIO4）2] molecule were found to be in different coordination geometries, i.e., Cu2 or Cu4 of a distorted square-pyramidal geometry coordinated with two nitrogens of TATP, the amino nitrogen and one carboxylate oxygen of L-Leu and one oxygen of perchlorate, and Cul or Cu3 with an octahedral geometry coordinated with the above stated similar coordinated atoms, and another carboxylate oxygen of L-Leu coordinating to Cu2 or Cu4. The complex can interact with CT-DNA by an intercalative mode and cleave pBR322 DNA in the presence of ascorbate.

Modification of the substrate specificity of leucine dehydrogenase by site‑directed mutagenesis based on biocomputing strategies

Synthesis of chiral amines by amine dehydrogenase has the advantages of environmental friendliness,high stereoselectivity,and mild reaction conditions.However,amine dehydrogenase has low catalytic activity,which greatly limits its application in the large-scale synthesis of chiral amines.In this study,a novel amine dehydrogenase was obtained by modifying the substrate specificity of leucine dehydrogenase via computer-aided protein engineering strategy.Furthermore,conservation analysis,homology modeling and molecular docking analysis were carried out via biocomputing strategy to select the mutation sites,and the mutants L52S and T143C were obtained.The enzyme activities of the two mutants to 2-pentanone were 1.55 U/mg and 2.06 U/mg,respectively.The enzyme activity of the latter was 188%and the Tm value was 2.55°Chigher than those of the original mutant,which laid a foundation for the efficient preparation of chiral amines by using this novel enzyme.

Synthesis of depsipeptides from L-amino acids and lactones

By using the corresponding L-amino acid sodium as initiator,ε-caprolactone-depsipeptides CL-Ala and CL-Leu were prepared by the reactions ofε-caprolactone(CL)with L-alanine and L-leucine,respec-tively,and p-dioxanone-depsipeptide(PDO-Leu)was prepared by the reaction of p-dioxanone(PDO)with L-leucine.Two poly(ε-caprolactone)oligomers(PCL-Ala and PCL-Leu)of different molecular weights with depsipeptide unit were synthesized by controlling the feed ratio of L-amino acid sodium and CL.The presence of the depsipeptide structure in these obtained products was confirmed by 1H NMR spectra and the molecular weight of the poly(ε-caprolactone)oligomers was measured by gel permeation chromatography(GPC).These products con-tain a hydroxyl group and a carboxyl group in one molecule,which means they could act as bifunctional monomers for further polymerization to prepare high molecular weight polymers.By this way,the depsipeptide unit could be introduced into the polymers and the biodegradation rates of the novel polymers could be well controlled in vivo by the tailored molecular structures.