酶法合成中色氨酸的测定

建立了光度测定酶法合成中色氨酸含量的新体系。该检测体系不仅能准确地测定产物色氨酸的含量，且对底物吲哚能同时测定。有效地避免了其他色氨酸测定方法底物吲哚的干扰，通过标准加入法色氨酸及吲哚的回收率分别为９７．８％～１０３．３％，９７．６％～１０３％。实验证明该体系简单、实用，测定结果可靠。     

乙酰丙酮-甲醛为荧光衍生试剂同步荧光法测定胺类物质

在用乙酰丙酮-甲醛作荧光衍生试剂测定胺类物质时,由于试剂背景干扰,胺类物质无法准确测定。本文以盐酸普鲁卡因为例,研究了同步荧光光度法测定胺类物质消除背景干扰的条件。同法测定了苯丙氨酸、正丙氨、正丁氨、谷氨酸、色氨酸和亮氨酸,给出了上述物质的线性回归方程,检出限,回收率和分析条件。     

双铂电极电流滴定测定邻甲苯胺

邻甲苯胺是合成新型含氯有机农药杀虫脒及染料工业的主要原料。目前多采用NaNO_2—KI外指示剂容量法进行测定,但其滴定终点不易掌握,结果准确度较差。资料介绍了用双铂电极电流滴定测定乙基保棉磷。我们对用NaNO_2作滴定剂,以双铂电极电流滴定法(永停法)测定邻甲苯胺进行了研究。拟定了双铂电极电流滴定法测定邻甲苯胺的方法,结果尚满意。     

HPLC法测定D-色氨酸甲酯盐酸盐中的左旋异构体

建立了测定D-色氨酸甲酯盐酸盐中左旋异构体的手性高效液相色谱方法.采用EnantioPak Y5(4.6 mm×250 mm,5μm)色谱柱,以正己烷(含0.1％二乙胺)-乙醇(80:20)为流动相,检测波长280 nm.D-色氨酸甲酯盐酸盐与其左旋异构体之间分离度良好,L-色氨酸甲酯盐酸盐在0.9927～14.890...     

高效液相色谱法测定人血白蛋白制品中乙酰色氨酸的含量

目的应用高效液相色谱法测定人血白蛋白制品中乙酰色氨酸的含量。方法参照《中国药典》三部(2005版)附录ⅢD,采用凝胶色谱柱,以紫外吸收检测器在280nm波长处检测,按照《中国药典》三部(2005版)附录ⅢB中的公式计算人血白蛋白制品中乙酰色氨酸的含量。分析线性关系,计算加标回收率,并验证该方法的重复性。结果乙酰色氨酸含量在0.16～0.64mmol/L的范围内与峰面积有良好的线性关系,相关系数为0.99992;3个浓度梯度(0.16、0.32和0.48mmol/L)乙酰色氨酸的加标回收率分别为100.4%、103.9%和102.8%;连续5次检测人血白蛋白制品中乙酰色氨酸的含量,变异系数为0.46%。结论应用高效液相色谱法测定人血白蛋白制品中乙酰色氨酸的含量,结果准确可靠。     

D-色氨酸印迹中空纤维复合膜制备及其性能

以聚砜（PSF）中空纤维超滤膜为基膜，采用表面紫外光聚合方法制备了D-色氨酸印迹中空纤维复合膜(CMIHFCM)。由SEM分析表明经过表面聚合后的D-色氨酸印迹中空纤维复合膜表面具有层叠交联状复合层，其厚度约3 μm。实验结果表明D-色氨酸印迹中空纤维复合膜对模板分子D-色氨酸具有很好的识别作用，D,L-色氨酸的分离因子（α）最高可以达到5.0，大于非分子印迹复合膜的识别选择性。此外，引发剂的用量对D-色氨酸印迹中空纤维复合膜的识别性能有较大影响；当引发剂的浓度为1.0％，制得的D-色氨酸印迹中空纤维复合膜对D-色氨酸具有较好的识别性能，具有良好的应用前景。     

十二烷基磺酸钠所致外切葡聚糖纤维二糖水解酶底物专一性的变化

以低浓度的十二烷基磺酸钠(SDS)为微扰剂处理外切葡聚糖纤维二糖水解酶（CBHI）,由荧光、圆二和二阶导致话测定反应动力学常数。结果表明低浓度的SDS处理可显著增加CBHI的内切酶活力,推测是CBHI活性位点附近的色氨酸所处微环境发生了变化,影响了酶与纤维素的结合所致。对近年来在分子水平上研究纤维素酶废物专一性的结果作了评述。     

高效液相色谱法测定发酵液中色氨酸和酪氨酸含量的研究

建立了高效液相色谱法测定发酵液中色氨酸和酪氨酸含量的方法。采用Agilent C18柱(250mm×4.6mm,5μm),以0.1mol/L醋酸钠(pH 4.0±0.05)-甲醇(90∶10)为流动相,流速为1.0mL·min-1,紫外检测器的检测波长为280nm,柱温30℃,结果显示,色氨酸的线性范围为50~500μg·mL-1,相关系数为r=0.9985,平均回收率为99.3%(n=4);酪氨酸的线性范围为20~2000μg·mL-1,相关系数为r=0.9991,平均回收率为100.2%(n=4)。该方法无需衍生,直接测定,简便,准确,用于发酵液中色氨酸和酪氨酸含量的测定,结果令人满意。     

超声波法提取花豆中的5-羟基色氨酸

5-羟基色氨酸是重要的神经类药物和保健类原料,有重要的利用价值。花豆产量大价格低,并且其5-羟基色氨酸含量很高。本实验以花豆为原料,以5-羟基色氨酸为研究对象,利用超声波法提取5-羟基色氨酸,提取效果好于传统有机溶剂萃取且过程绿色简洁。利用高效液相色谱法对试样进行了分析,得到色谱图与标准曲线。色谱条件为色谱柱Symmetry C_(18)(5μm,4.6 mm×150 mm);检测波长276 nm;流动相:甲醇∶水=5∶95(V/V);流速:1.0 mL/min;进样量:10μL;柱温:室温;灵敏度:0.01 AUFS。通过四因素三水平对比试验优化5-羟基色氨酸的最佳提取条件为:脱脂溶剂为石油醚,温度为50℃,脱脂时间为45 min,提取溶剂甲醇,超声波提取时间45 min;测定的5-羟基色氨酸含量为0.49%。     

色氨酸的合成及其应用

色氨酸(Tryptophane),又名β-吲哚-α-氨基丙酸,是Hopkins和Cole在1902年发现并分离得到的一种氨基酸。它是人类和动物必需的八种氨基酸之一,也是唯一具有多方面生理生化功能的氨基酸, 人们一直期望色氨酸能继赖氨酸、蛋氨酸之后,成为食品和饲料添体剂的第三大种氨基酸。但由于色氨酸生产难度高、价格昂贵,多年来只作医药用,其他领域未能大量推广应用。近年来,欧美各国应用色氨酸的营养补品已大量上市。新技术的开发应用则以日本活跃,日本产量从七十年代末的几十吨发展到上千吨,售价从七十年代的2万日元/公斤下降到5000日元/公斤(1988年11月)。本文将介绍色氨酸的合成、应用及其进展情况。     

The role of tryptophan in protein fibrillogenesis: relevance of Trp7 and Trp14 to the amyloidogenic properties of myoglobin

In order to understand the role of tryptophan in the mechanisms of fibrils formation, the ability of a series of analogs of the residue 7-18 span of myoglobin to form amyloid-like fibrils was investigated. Alternatively one or both tryptophans were substituted with alanine and leucine, to determine the contribution of hydrophobicity and aromaticity. The scale of aggregation propensity of the peptides determined indicates that tryptophan is crucial for the amyloidogenic process. Since the rare tryptophan residue is generally engaged in structural roles in proteins, or when exposed serves as binding sites, we surmise that its exposure in the amyloidogenic fragments allows for intermolecular clustering with residues from other molecules leading to the formation of amyloid aggregates.     

Characterization of a Tryptophan 6-Halogenase from Streptomyces albus and Its Regioselectivity Determinants

Tryptophan halogenases are found in diverse organisms and catalyze regiospecific halogenation. They play an important role in the biosynthesis of halogenated indole alkaloids, which are biologically active and of therapeutic importance. Here, a tryptophan 6-halogenase (SatH) from Streptomyces albus was characterized by using a whole-cell reaction system in Escherichia coli. SatH showed substrate specificity for chloride and bromide ions, leading to regiospecific halogenation at the C6-position of l -tryptophan. In addition, SatH exhibited higher performance in bromination than that of previously reported tryptophan halogenases in the whole-cell reaction system. Through structure-based protein mutagenesis, it has been revealed that two consecutive residues, A78/V79 in SatH and G77/I78 in PyrH, are key determinants in the regioselectivity difference between tryptophan 6- and 5-halogenases. Substituting the AV with GI residues switched the regioselectivity of SatH by moving the orientation of tryptophan. These data contribute to an understanding of the key residues that determine the regioselectivity of tryptophan halogenases.     

固定化色氨酸合成酶基因工程菌合成L-色氨酸的研究

以海藻酸钠作为包埋剂、戊二醛作为交联剂和氯化钙作为填充剂对色氨酸合成酶基因工程菌进行固定化,同时探究三种物质和菌体负载量对固定化菌影响,响应面法优化色氨酸合成酶基因工程菌合成L-色氨酸。固定化色氨酸合成酶基因工程菌最优制备条件为：海藻酸钠28.92 g/l、戊二醛0.95%、氯化钙19.82 g/l、菌体负载量25 g/l。底物L-丝氨酸浓度为1%、固定化菌8g, L-色氨酸转化率为28.16%。固定化菌可连续使用15批次。     

Cholesterol interaction with recombinant human sterol carrier protein-2

The interaction of human recombinant sterol carrier protein-2 (SCP-2) with sterols was examined. Two independent ligand binding methods, Lipidex 1000 binding of [³H]cholesterol and a fluorescent dehydroergosterol binding assay, were used to determine the affinity of SCP-2 for sterols. Binding analysis indicated SCP-2 bound [³H]cholesterol and dehydroergosterol with aK _d of 0.3 and 1.7 μM, respectively, and suggested the presence of a single binding site. Phase fluorometry and circular dichroism were used to characterize the SCP-2 sterol binding site. Alterations in dehydroergosterol lifetime, SCP-2 tryptophan lifetime, and SCP-2 tryptophan quenching by acrylamide upon cholesterol binding demonstrated a shielding of the SCP-2 tryptophan from the aqueous solvent by bound sterol. Differential polarized phase fluorometry revealed decreased SCP-2 tryptophan rotational correlation time upon cholesterol binding. Circular dichroism of SCP-2 indicated that cholesterol elicited a small decrease in SCP-2 alpha helical content. The data suggest that SCP-2 binds sterols with affinity consistent with a lipid transfer protein that may act either as an aqueous carrier or at a membrane surface to enhance sterol desorption.     

Non-Invasive Brain Stimulation Effects on Biomarkers of Tryptophan Metabolism: A Scoping Review and Meta-Analysis

Abnormal activation of the kynurenine and serotonin pathways of tryptophan metabolism is linked to a host of neuropsychiatric disorders. Concurrently, noninvasive brain stimulation (NIBS) techniques demonstrate high therapeutic efficacy across neuropsychiatric disorders, with indications for modulated neuroplasticity underlying such effects. We therefore conducted a scoping review with meta-analysis of eligible studies, conforming with the PRISMA statement, by searching the PubMed and Web of Science databases for clinical and preclinical studies that report the effects of NIBS on biomarkers of tryptophan metabolism. NIBS techniques reviewed were electroconvulsive therapy (ECT), transcranial magnetic stimulation (TMS), and transcranial direct current stimulation (tDCS). Of the 564 search results, 65 studies were included with publications dating back to 1971 until 2022. The Robust Bayesian Meta-Analysis on clinical studies and qualitative analysis identified general null effects by NIBS on biomarkers of tryptophan metabolism, but moderate evidence for TMS effects on elevating serum serotonin levels. We cannot interpret this as evidence for or against the effects of NIBS on these biomarkers, as there exists several confounding methodological differences in this literature. Future controlled studies are needed to elucidate the effects of NIBS on biomarkers of tryptophan metabolism, an under-investigated question with substantial implications to clinical research and practice.     

UHPLC-ESI-MS/MS Quantification of Relevant Substrates and Metabolites of the Kynurenine Pathway Present in Serum and Peritoneal Fluid from Gastric Cancer Patients—Method Development and Validation

Metabolites and enzymes involved in the kynurenine pathway (KP) are highly promising targets for cancer treatment, including gastrointestinal tract diseases. Thus, accurate quantification of these compounds in body fluids becomes increasingly important. The aim of this study was the development and validation of the UHPLC-ESI-MS/MS methods for targeted quantification of biologically important KP substrates (tryptophan and nicotinamide) and metabolites(kynurenines) in samples of serum and peritoneal fluid from gastric cancer patients. The serum samples were simply pretreated with trichloroacetic acid to precipitate proteins. The peritoneal fluid was purified by solid-phase extraction before analysis. Validation was carried out for both matrices independently. Analysis of the samples from gastric cancer patients showed different accumulations of tryptophan and its metabolites in different biofluids of the same patient. The protocols will be used for the evaluation of tryptophan and kynurenines in blood and peritoneal fluid to determine correlation with the clinicopathological status of gastric cancer or the disease’s prognosis.     

牛血清白蛋白为分离剂超滤拆分色氨酸的机理和模型(Ⅰ)

以牛血清白蛋白（BSA）作为手性选择剂,利用膜超滤法拆分色氨酸（Trp）可以达到良好的手性分离效果。系统地研究了溶液pH值、温度、色氨酸和BSA的初始浓度、超滤压力等因素对分离效果的影响,指出除pH值外,温度对分离效果同样具有显著的影响。并依据pH值和温度对BSA空间结构和色氨酸带电性质的影响规律,定性分析了BSA拆分色氨酸对映体的机理。pH值主要通过影响BSA的空间构象和色氨酸的带电性质影响分离效果,温度则主要通过影响BSA的空间结构影响分离效果。     

Developmental Programming and Reprogramming of Hypertension and Kidney Disease: Impact of Tryptophan Metabolism

The concept that hypertension and chronic kidney disease (CKD) originate in early life has emerged recently. During pregnancy, tryptophan is crucial for maternal protein synthesis and fetal development. On one hand, impaired tryptophan metabolic pathway in pregnancy impacts fetal programming, resulting in the developmental programming of hypertension and kidney disease in adult offspring. On the other hand, tryptophan-related interventions might serve as reprogramming strategies to prevent a disease from occurring. In the present review, we aim to summarize (1) the three major tryptophan metabolic pathways, (2) the impact of tryptophan metabolism in pregnancy, (3) the interplay occurring between tryptophan metabolites and gut microbiota on the production of uremic toxins, (4) the role of tryptophan-derived metabolites-induced hypertension and CKD of developmental origin, (5) the therapeutic options in pregnancy that could aid in reprogramming adverse effects to protect offspring against hypertension and CKD, and (6) possible mechanisms linking tryptophan metabolism to developmental programming of hypertension and kidney disease.     

An Emerging Cross-Species Marker for Organismal Health: Tryptophan-Kynurenine Pathway

Tryptophan (TRP) is an essential dietary amino acid that, unless otherwise committed to protein synthesis, undergoes metabolism via the Tryptophan-Kynurenine (TRP-KYN) pathway in vertebrate organisms. TRP and its metabolites have key roles in diverse physiological processes including cell growth and maintenance, immunity, disease states and the coordination of adaptive responses to environmental and dietary cues. Changes in TRP metabolism can alter the availability of TRP for protein and serotonin biosynthesis as well as alter levels of the immune-active KYN pathway metabolites. There is now considerable evidence which has shown that the TRP-KYN pathway can be influenced by various stressors including glucocorticoids (marker of chronic stress), infection, inflammation and oxidative stress, and environmental toxicants. While there is little known regarding the role of TRP metabolism following exposure to environmental contaminants, there is evidence of linkages between chemically induced metabolic perturbations and altered TRP enzymes and KYN metabolites. Moreover, the TRP-KYN pathway is conserved across vertebrate species and can be influenced by exposure to xenobiotics, therefore, understanding how this pathway is regulated may have broader implications for environmental and wildlife toxicology. The goal of this narrative review is to (1) identify key pathways affecting Trp-Kyn metabolism in vertebrates and (2) highlight consequences of altered tryptophan metabolism in mammals, birds, amphibians, and fish. We discuss current literature available across species, highlight gaps in the current state of knowledge, and further postulate that the kynurenine to tryptophan ratio can be used as a novel biomarker for assessing organismal and, more broadly, ecosystem health.     

Tryptophan Metabolism via Kynurenine Pathway: Role in Solid Organ Transplantation

Solid organ transplantation is a gold standard treatment for patients suffering from an end-stage organ disease. Patient and graft survival have vastly improved during the last couple of decades; however, the field of transplantation still encounters several unique challenges, such as a shortage of transplantable organs and increasing pool of extended criteria donor (ECD) organs, which are extremely prone to ischemia-reperfusion injury (IRI), risk of graft rejection and challenges in immune regulation. Moreover, accurate and specific biomarkers, which can timely predict allograft dysfunction and/or rejection, are lacking. The essential amino acid tryptophan and, especially, its metabolites via the kynurenine pathway has been widely studied as a contributor and a therapeutic target in various diseases, such as neuropsychiatric, autoimmune disorders, allergies, infections and malignancies. The tryptophan-kynurenine pathway has also gained interest in solid organ transplantation and a variety of experimental studies investigating its role both in IRI and immune regulation after allograft implantation was first published. In this review, the current evidence regarding the role of tryptophan and its metabolites in solid organ transplantation is presented, giving insights into molecular mechanisms and into therapeutic and diagnostic/prognostic possibilities.