Enantiomeric high-performance liquid chromatographic separation of β-substituted tryptophan analogues

Summary The high-performance liquid chromatographic enantioresolution of β-substituted tryptophan analogues by direct and indirect methods is reported. The direct separation was carried out on chiral crown ether-bonded, on the macrocyclic antibiotic teicoplanin-bonded or on different derivatized β-cyclodextrin-bonded phases. The indirect resolution was achieved by applying pre-column derivatization with the chiral reagents 1-fluoro-2,4-dinitrophenyl-5-L-alanine amide (FDAA) and 2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl isothiocyanate (GITC). In the separation of all four isomers of the analytes, the indirect methods seemed to be more efficient than the direct methods. In most cases, the FDAA derivatives of the stereoisomers could be separated with better selectivity in methanol-containing mobile phases. Presented at Balaton Symposium on High Performance Separation Methods, Siófok, Hungary, September 1–3, 1999     

Fluorimetric Analysis of the Structural-Dynamic State of Alkaline Phosphatase of Escherichia coli

By the methods of tryptophan fluorescence and phosphorescence at room temperature, the action on the conformational state and intramolecular dynamics (IMD) of alkaline phosphatase of Escherichia coli of the competitive inhibitor sodium pyrophosphate (1–10 mM) has been investigated. On the basis of the results of the fluorimetric investigations, it has been concluded that pyrophosphate is capable of changing the conformation and IMD of enzymatic protein towards an increase in the fast and slow mobility of the globule structural elements. The most probable cause of the shifts of the alkaline phosphatase structural-dynamic state observed by fluorimetric methods is the replacement of phosphate in the active center of one of the protein subunits by pyrophosphate.     

Chiral Brønsted acid catalyzed stereoselective addition of azlactones to 3-vinylindoles for facile access to enantioenriched tryptophan derivatives

Syn-gled out: The syn diastereo- and enantioselective addition of azlactones to 3-vinylindoles was accomplished by using a chiral, binapthol-derived, Br?nsted acid catalyst (see scheme). This method enables facile access to tryptophan derivatives with adjacent quaternary and tertiary stereogenic centers, which are potentially useful for the synthesis of peptidomimetics.     

高效液相色谱-程序波长紫外检测法同时测定血浆中的色氨酸及其代谢物

建立了一种高效液相色谱-程序波长紫外检测法同时测定血浆中色氨酸(Trp)及其主要代谢产物犬尿氨酸(Kyn)和5-羟色胺(5-HT)。以茶碱为内标(IS),采用BDS-Hypersil-C8柱(150 mm×4.6 mm, 5 μm)分离。流动相为10 mmol/L醋酸钠缓冲液(pH 4.5)-乙腈(94:6, v/v),流速为0.6 mL/min;柱温为25 ℃;紫外检测波长设定: Kyn和IS为360 nm, 5-HT为220 nm, Trp为302 nm。3种物质的平均回收率为87%～113%;线性范围分别为3.97～400 μmol/L(Trp), 0.421～20.2 μmol/L(Kyn), 4.36～980 nmol/L(5-HT);检出限分别为0.134 μmol/L(Trp), 0.0160 μmol/L(Kyn), 2.03 nmol/L(5-HT)。利用该方法对15例抑郁症患者和15例健康志愿者的血浆进行测定,结果表明两组间Trp的代谢存在显著的差异。     

阻抑速差催化光度法测定痕量色氨酸和酪氨酸

研究了色氨酸、酪氨酸在硫酸环境中抑制碘对亚砷酸与硫酸铈氧化还原反应的催化作用及动力学条件。在0．001mol／L Ce(SO4)2,0．00125mol／L As2O3,0．013mg／L KI,0．025g／L和0．25g／L NaCl溶液中测定色氨酸和酪氨酸,其线性范围分别为0．0～0．65mg／L,0．0～4．0mg／L和0．0～1．3mg／L。0．10～4．0mg／L,用联立方程分别求出各自含量。用此法对醇溶蛋白质和混合氨基酸溶液中色氨酸和酪氨酸进行测定,混合氨基酸的色氨酸、酪氨酸测量值为0．8037g／L,3．591g/L;蛋白质的色氨酸、酪氨酸测量值为1．8g/kg、3．43g/kg;回收率在88％～109％;色氨酸和酪氨酸的RSD分别为4．8％和5．2％,结果满意。     

Identification of epidermal L‐tryptophan and its oxidation products by in vivo FT‐Raman Spectroscopy further supports oxidative stress in patients with vitiligo

In the past, non‐invasive in vivo FT‐Raman spectroscopy has been used to detect H₂O₂‐mediated oxidation of methionine to methionine sulfoxide and methionine sulfone, as well as cysteine to cysteic acid, in the sequence of proteins in the epidermis of patients with vitiligo. L ‐tryptophan (Trp) is another potential target for this oxidation. Owing to the presence of 10⁻³M epidermal albumin which contains one Trp residue, it was tempting to follow the oxidation of this amino acid. Using in vivo and in vitro FT‐Raman spectroscopy, we show for the first time that epidermal Trp is oxidised in patients with vitiligo, yielding 5‐OH‐Trp at 930 cm⁻¹ and other oxidation products (i.e. N‐formyl kynurenine and kynurenine) from indole ring oxidation peaking at 1050 cm⁻¹. On the basis of detailed in vitro results, we could conclude that 5‐OH‐Trp as well as formyl kynurenine and kynurenine are formed via H₂O₂‐mediated Fenton chemistry. These results once again bring out the strength of non‐invasive in vivo FT‐Raman Spectroscopy in dermatology to follow the effect of oxidative stress in the skin of patients with vitiligo. Copyright © 2008 John Wiley & Sons, Ltd.     

Oxidation Mechanism of 1-Methyl-tryptophan and Tryptophan on Glassy Carbon Electrode: A Comparative Study

The anodic behaviour of 1-methyl-tryptophan (1-mTrp) in aqueous electrolytes was investigated on a glassy carbon electrode (GCE), using voltammetric techniques. The oxidation of 1-mTrp was associated with an electrochemical-chemical (EC) mechanism: one electron and one proton were removed of C2 to form an intermediate radical, 1-mTrp⋅. This was followed by a two-way reaction, producing a 1-mTrp dimer and/or reaction with water to form a final hydroxylated product. The oxidation mechanism of 1-mTrp proposed was also compared with the anodic oxidation Trp on GCE. Differential pulse voltammetry was also explored for quantification of Trp and 1-mTrp in neutral medium with low detection limits, on an anodically pre-treated GCE.     

Simultaneous determination of serum tryptophan metabolites in an older Chinese population

Tryptophan (TRP) and its metabolites exhibit significant biological effects and are strongly associated with age-related disease and mortality. However, reports on quantitatively analyzing these metabolites in older individuals are not available. We used ultra-high-performance liquid chromatography–tandem mass spectrometry to optimize and validate a method for isotope dilution analysis of TRP metabolites in older individuals. The targeted analytes are TRP, serotonin or 5-hydroxytryptamine, kynurenine, kynurenic acid, xanthurenic acid, indole-3-acetic acid, indole-3-propionic acid, and tryptamine. The serum sample was purified using solid-phase extraction and was separated on a Waters HSS T3 column (100 mm × 2.1 mm, 1.8 μm). The analytes were detected in the multiple reaction monitoring mode under positive ionization. TRP was confirmed and measured after being diluted 100 times. This method exhibited satisfactory linearity (r > 0.99). The intrabatch and interbatch accuracies (85.7–114%) and precisions (<15%) were acceptable. The standard-normalized matrix effects ranged from 51.6 to 145%. This method was successfully applied to a cohort of 1021 older Chinese individuals, and this study may enable further understanding of the metabolic phenotypes associated with TRP in other populations.