基于非靶标代谢组学方法研究水稻种子的代谢物轮廓

建立了一种基于气相色谱-质谱联用的非靶标代谢组学方法,研究了不同品种水稻种子之间的代谢物差异。分别考察了3种提取方式及4种提取溶剂对水稻中代谢物提取效率的影响,发现涡旋提取方法及80%甲醇提取可获得较好的结果,该方法具有良好的稳定性和线性。将方法用于3种水稻种子(农大、稻花香、状元)的代谢轮廓分析,结果显示3种水稻的代谢特征不同。相对于农大种子,稻花香种子中脂肪酸(棕榈酸、油酸、亚油酸、硬脂酸)和糖类(葡萄糖、半乳糖、甘露醇)、天冬酰胺、柠檬酸的含量较高,此外,状元种子中乳酸含量也较高;稻花香种子中琥珀酸、果糖,状元种子中的苏氨酸含量低于农大种子。但相对于状元种子,稻花香中乳酸、琥珀酸、硬脂酸、果糖等物质含量较低。该方法可应用于不同品种、不同基因型或不同产地水稻种子的代谢表型分析,为改善水稻产量及质量等研究提供信息。     

小儿肺炎患者尿液的代谢组学初步研究

利用液相色谱-质谱联用法对小儿肺炎( Childhood pneumonia, CP)患者和健康儿童( Healthy control)的尿液进行分析,发现小儿肺炎患者尿液中的潜在标记物,为其发病机制及早期筛查提供科学依据。筛选10例小儿肺炎患者(age 47.72±2.35 months)及10例健康儿童(age 46.65±1.97 months)尿液样本,采用快速高分辨液相色谱四极杆-飞行时间质谱联用( RRLC-Q TOF/MS)技术对其尿液代谢物进行分析,通过主成分分析方法( PCA)对两组代谢物进行分类,并发现潜在生物标记物。 RRLC-Q TOF/MS检测表明,CP组和Healthy Control组尿液代谢物图谱能得到很好的区分,并鉴定了5种生物标记物,提示嘌呤代谢、氨基酸代谢可能在小儿肺炎发生发展中有重要作用。     

基于液质联用技术的高脂血症金黄地鼠肝脏非靶标代谢组学研究

采用液相色谱-飞行时间质谱联用技术(LC-TOF MS)对高脂血症金黄地鼠的肝匀浆液进行代谢谱分析,并利用模式识别挖掘潜在生物标志物以探索发病机制.采用反相色谱(RPLC)和亲水作用色谱(HILIC)两种模式分离肝匀浆液中的内源性代谢产物.采用偏最小二乘法判别分析(PLS-DA)进行数据分析,结合标准品与数据库比对、二...     

代谢组学法研究三聚氰胺对儿童尿液代谢的影响

运用代谢组学方法研究了三聚氰胺对儿童尿液代谢的影响.通过超高效液相色谱-飞行时间质谱(UPLC/TOF-MS)法分析儿童尿样的代谢指纹图谱,质谱数据采用MarkerLynx软件处理,然后使用主成分分析和偏最小二乘判别分析法分析病例组和正常对照组之间的代谢物谱差异,并通过变量重要性投影(VIP)选取潜在的生物标志物,结合质谱同位素分析和数据库检索对潜在的生物标志物进行鉴定.结果表明,三聚氰胺通过肾结石导致的物理性损伤干扰了柠檬酸代谢.代谢组学法能够应用于三聚氰胺导致的代谢异常的研究及三聚氰胺导致肾损伤的无创检测.     

基于肝脏代谢组分析研究低聚硒化氨基多糖对黑鲷的免疫调节作用

采用基于液相色谱-飞行时间质谱联用（LC-TOF-MS）技术的代谢组学方法，分析黑鲷肝脏内源性代谢物的变化，研究硒化氨基多糖增强黑鲷的免疫调节机制。采用XCMS^plus软件非靶向分析质谱采集数据，筛选潜在生物标志物，并通过MetaboAnalyst3.0网站分析相关代谢通路。结果表明，饲喂硒化氨基多糖组中的代谢物明显区分于空白组，发现并鉴定了32个有差异的生物标志物。代谢通路分析结果表明，硒化氨基多糖可通过氨基酰基-转运脱氧核糖核苷酸（tRNA）生物合成、氨基酸代谢、核苷酸代谢、氮代谢等代谢通路增强黑鲷自身的免疫机能。该研究为阐明硒化氨基多糖的免疫增强机制提供了科学依据。     

慢性心功能衰竭患者尿液的代谢组学研究

采用基于液相色谱-质谱联用的方法对慢性心力衰竭(Chronic heart failure, CHF)患者和正常对照(Control)人群的尿液进行分析, 筛选慢性心力衰竭患者尿液中的差异代谢物, 研究其发病机制, 并为临床治疗提供科学依据.选择15个慢性心力衰竭患者(年龄(62.27±3.14)岁)及15个正常人(年龄(65.41±4.63)岁), 采用高分辨度快速液相色谱-四极杆-飞行时间串联质谱(RRLC-QTOF/MS)技术对尿液代谢物进行分析, 采用主成分分析(PCA)对两组代谢物进行分类, 并筛选潜在生物标记物;运用偏最小二乘判别分析法(PLS-DA)建模, 考察生物标记物对疾病筛选的预测能力.研究结果表明, CHF组和Control组尿液代谢物谱能得到很好的区分, 发现并鉴定了2种潜在生物标记物尿苷及丙氨酰色氨酸, 提示嘧啶代谢和色氨酸代谢可能在心力衰竭发生发展中有重要作用.     

基于液相色谱-质谱技术的肾脏代谢组学分析方法研究

建立了一种基于高效液相色谱-超高分辨质谱技术的肾脏代谢组学分析方法。肾脏组织于液氮中研磨成粉后,采用甲基叔丁基醚/甲醇/水溶剂体系进行提取,分别得到强极性部分(下层)和弱极性部分(上层)提取物,依次采用HILIC亲水色谱柱和反相C₁₈色谱柱进行梯度洗脱分离后,进行高分辨质谱分析。采用电喷雾离子源(ESI),正、负离子模式检测,扫描方式为全扫描,扫描范围为100～1 000 Da,质量分辨率为120 000。结果表明,该方法灵敏度高,专属性强,稳定性良好,可同时获取肾脏组织中的强极性和弱极性代谢物信息,可为肾脏疾病和药物肾毒性生物标志物的发现提供一种新的方法。     

汞离子对细胞代谢通路影响的代谢组学

利用超高效液相色谱-四级杆飞行时间质谱（UPLC-Q-TOF-MS）技术对汞离子作用后细胞的代谢组学进行了研究,结合化学计量学方法,对代谢组学数据进行了多维统计分析.结果表明,与非汞离子作用组相比,汞离子作用组存在能量、磷脂、脂肪酸及氨基酸代谢异常,并发现16种有显著差异的生物标记物.进一步探讨了汞中毒的细胞代谢机理及细胞的应激保护.本研究建立的基于UPLC-Q-TOF-MS技术的细胞代谢组学快速分析方法可以对细胞在重金属作用后的代谢物变化进行轮廓分析.     

基于液相色谱-高分辨质谱的代谢组学技术用于麦卢卡蜂蜜的甄别

采用基于液相色谱-高分辨质谱的代谢组学技术对麦卢卡蜂蜜以及国内主要蜂蜜品种的代谢谱进行全面分析，实现了麦卢卡蜂蜜与其他蜂蜜的区分，建立了偏最小二乘法判别模型，对测试样品能够取得很好的鉴定效果。实验从麦卢卡蜂蜜中筛选出高度表达的3-苯乳酸、甲氧基苯乙酮、二氢麻醉椒苦素、芹菜素等黄酮和肉桂酸类等34种代谢标志物，所构建的标志物组合模型的受试者工作特征曲线下的面积达到了0.99。该文建立的代谢组学方法为麦卢卡蜂蜜的质量控制提供了新的思路。     

基于质谱技术的代谢组学研究及其在中国的发展

代谢组学是关于生物系统代谢物组成及变化规律的科学,是系统生物学的重要组成部分.质谱技术是目前代谢组学研究中最主要的分析手段之一,广泛应用于代谢组学各个领域.本文阐述了基于质谱技术的代谢组学方法及其应用,重点介绍和评论了近年来我国在该领域取得的进步和成果,并对基于质谱技术的代谢组学研究目前存在的问题及未来的发展进行了分析与展望.     

Comparison of Widely Targeted Metabolomics and Untargeted Metabolomics of Wild Ophiocordyceps sinensis

The authors of this paper conducted a comparative metabolomic analysis of Ophiocordyceps sinensis (OS), providing the metabolic profiles of the stroma (OSBSz) and sclerotia (OSBSh) of OS by widely targeted metabolomics and untargeted metabolomics. The results showed that 778 and 1449 metabolites were identified by the widely targeted metabolomics and untargeted metabolomics approaches, respectively. The metabolites in OSBSz and OSBSh are significantly differentiated; 71 and 96 differentially expressed metabolites were identified by the widely targeted metabolomics and untargeted metabolomics approaches, respectively. This suggests that these 71 metabolites (riboflavine, tripdiolide, bromocriptine, lumichrome, tetrahymanol, citrostadienol, etc.) and 96 metabolites (sancycline, vignatic acid B, pirbuterol, rubrophen, epalrestat, etc.) are potential biomarkers. 4-Hydroxybenzaldehyde, arginine, and lumichrome were common differentially expressed metabolites. Using the widely targeted metabolomics approach, the key pathways identified that are involved in creating the differentiation between OSBSz and OSBSh may be nicotinate and nicotinamide metabolism, thiamine metabolism, riboflavin metabolism, glycine, serine, and threonine metabolism, and arginine biosynthesis. The differentially expressed metabolites identified using the untargeted metabolomics approach were mainly involved in arginine biosynthesis, terpenoid backbone biosynthesis, porphyrin and chlorophyll metabolism, and cysteine and methionine metabolism. The purpose of this research was to provide support for the assessment of the differences between the stroma and sclerotia, to furnish a material basis for the evaluation of the physical effects of OS, and to provide a reference for the selection of detection methods for the metabolomics of OS.     

Untargeted Metabolomics Reveals the Potential Antidepressant Activity of a Novel Adenosine Receptor Antagonist

Depression is the most common mental illness, affecting approximately 4.4% of the global population. Despite many available treatments, some patients exhibit treatment-resistant depression. Thus, the need to develop new and alternative treatments cannot be overstated. Adenosine receptor antagonists have emerged as a promising new class of antidepressants. The current study investigates a novel dual A₁/A_2A adenosine receptor antagonist, namely 2-(3,4-dihydroxybenzylidene)-4-methoxy-2,3-dihydro-1H-inden-1-one (1a), for antidepressant capabilities by determining its metabolic profiles and comparing them to those of two reference compounds (imipramine and KW-6002). The metabolic profiles were obtained by treating male Sprague-Dawley rats with 1a and the reference compounds and subjecting them to the forced swim test. Serum and brain material was consequently collected from the animals following euthanasia, after which the metabolites were extracted and analyzed through untargeted metabolomics using both ¹H-NMR and GC-TOFMS. The current study provides insight into compound 1a’s metabolic profile. The metabolic profile of 1a was similar to those of the reference compounds. They potentially exhibit their antidepressive capabilities via downstream effects on amino acid and lipid metabolism.     

Development of a Work-Flow for High-Performance Thin-Layer Chromatography Data Processing for Untargeted Metabolomics

JPC – Journal of Planar Chromatography – Modern TLC - Although the different practical steps of a high-performance thin-layer chromatography (HPTLC) experiment are now all automated,...     

Untargeted Metabolomics Reveals New Markers of Food Processing for Strawberry and Apple Purees

In general, food processing and its conditions affect nutrients, bioactive compounds, and sensory characteristics of food products. This research aims to use a non-targeted metabolomics approach based on UPLC-ESI-QTOF-MS to determine how fruit processing can affect the metabolic profile of fruits and, through a comprehensive metabolic analysis, identify possible markers to assess their degree of processing. The present study uses a real case from the food industry to evaluate markers of the processing of strawberry and apple purees industrially elaborated with different processing techniques and conditions. The results from the multivariate analysis revealed that samples were grouped according to the type of processing, evidencing changes in their metabolic profiles and an apparent temperature-dependent effect. These metabolic profiles showed changes according to the relevance of thermal conditions but also according to the exclusively cold treatment, in the case of strawberry puree, and the pressure treatment, in the case of apple puree. After data analysis, seven metabolites were identified and proposed as processing markers: pyroglutamic acid, pteroyl-D-glutamic acid, 2-hydroxy-5-methoxy benzoic acid, and 2-hydroxybenzoic acid β-d-glucoside in strawberry and di-hydroxycinnamic acid glucuronide, caffeic acid and lysoPE(18:3(9Z,12Z,15Z)/0:0) in apple purees. The use of these markers may potentially help to objectively measure the degree of food processing and help to clarify the controversial narrative on ultra-processed foods.     

Targeted and Untargeted Mass Spectrometry-Based Metabolomics for Chemical Profiling of Three Coffee Species

While coffee beans have been studied for many years, researchers are showing a growing interest in coffee leaves and by-products, but little information is currently available on coffee species other than Coffea arabica and Coffea canephora. The aim of this work was to perform a targeted and untargeted metabolomics study on Coffea arabica, Coffea canephora and Coffea anthonyi. The application of the recent high-resolution mass spectrometry-based metabolomics tools allowed us to gain a clear overview of the main differences among the coffee species. The results showed that the leaves and fruits of Coffea anthonyi had a different metabolite profile when compared to the two other species. In Coffea anthonyi, caffeine levels were found in lower concentrations while caffeoylquinic acid and mangiferin-related compounds were found in higher concentrations. A large number of specialized metabolites can be found in Coffea anthonyi tissues, making this species a valid candidate for innovative healthcare products made with coffee extracts.     

Untargeted Metabolomics Analysis Revealed the Major Metabolites in the Seeds of four Polygonatum Species

Most Polygonatum species are widely used in China as a source of medicine and food. In this study, a UPLC-QTOF-MS/MS system was used to conduct an untargeted metabolomics analysis and compare the classes and relative contents of metabolites in the seeds of four Polygonatum species: P. sibiricum (Ps), P. cyrtonema (Pc), P. kingianum (Pk), and P. macropodium (Pm). The objectives of this study were to clarify the metabolic profiles of these seeds and to verify their medicinal and nutritional value via comparative analyses. A total of 873 metabolites were identified, including 185 flavonoids, 127 lipids, 105 phenolic acids, and 36 steroids. The comparative analysis of metabolites among Polygonatum seed samples indicated that flavonoids, steroids, and terpenoids were the main differentially abundant compounds. The results of principal component analysis and hierarchical clustering were consistent indicating that the metabolites in Ps and Pm are similar, but differ greatly from Pc and Pk. The data generated in this study provide additional evidence of the utility of Polygonatum seeds for producing food and medicine.     

Uncovering the Mechanisms of Active Components from Toad Venom against Hepatocellular Carcinoma Using Untargeted Metabolomics

Toad venom, a dried product of secretion from Bufo bufo gargarizans Cantor or Bufo melanostictus Schneider, has had the therapeutic effects of hepatocellular carcinoma confirmed. Bufalin and cinobufagin were considered as the two most representative antitumor active components in toad venom. However, the underlying mechanisms of this antitumor effect have not been fully implemented, especially the changes in endogenous small molecules after treatment. Therefore, this study was designed to explore the intrinsic mechanism on hepatocellular carcinoma after the cotreatment of bufalin and cinobufagin based on untargeted tumor metabolomics. Ultraperformance liquid chromatography with tandem mass spectrometry (UHPLC-MS/MS) was performed to identify the absorbed components of toad venom in rat plasma. In vitro experiments were determined to evaluate the therapeutic effects of bufalin and cinobufagin and screen the optimal ratio between them. An in vivo HepG2 tumor-bearing nude mice model was established, and a series of pharmacodynamic indicators were determined, including the body weight of mice, tumor volume, tumor weight, and histopathological examination of tumor. Further, the entire metabolic alterations in tumor after treating with bufalin and cinobufagin were also profiled by UHPLC-MS/MS. Twenty-seven active components from toad venom were absorbed in rat plasma. We found that the cotreatment of bufalin and cinobufagin exerted significant antitumor effects both in vitro and in vivo, which were reflected in inhibiting proliferation and inducing apoptosis of HepG2 cells and thereby causing cell necrosis. After cotherapy of bufalin and cinobufagin for twenty days, compared with the normal group, fifty-six endogenous metabolites were obviously changed on HepG2 tumor-bearing nude mice. Meanwhile, the abundance of α-linolenic acid and phenethylamine after the bufalin and cinobufagin intervention was significantly upregulated, which involved phenylalanine metabolism and α-linolenic acid metabolism. Furthermore, we noticed that amino acid metabolites were also altered in HepG2 tumor after drug intervention, such as norvaline and Leu-Ala. Taken together, the cotreatment of bufalin and cinobufagin has significant antitumor effects on HepG2 tumor-bearing nude mice. Our work demonstrated that the in-depth mechanism of antitumor activity was mainly through the regulation of phenylalanine metabolism and α-Linolenic acid metabolism.     

非靶向尿液代谢组学方法研究五味子治疗精尿病并发症作用机制

采用超高效液相色谱-串联四级杆飞行时间质谱联用技术(Ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry,UPLC/Q-TOF-MS),结合主成分分析技术(Principal component analysis,PCA),进行了五味子治疗糖尿病并发症大鼠尿液代谢组学研究,对五味子治疗糖尿病并发症大鼠后尿液生物标志物变化进行了考察.采用UPLC/Q-TOF-MS方法分析了健康组、糖尿病并发症模型组和五味子给药组的大鼠尿液,采用偏最小二乘法-判别分析(Partial least squares discriminant analysis,PLS-DA)和正交偏最小二乘法-判别分析(Orthogonal partial least squares discriminant analysis,OPLS-DA)载荷图筛选,通过分析对各组分离贡献较大(VIP>1,P°0.05)化合物的串联质谱数据,再经Human Metabolome Data-base(HMDB)等数据库检索,进行质谱信息匹配,尿液中共鉴定出28种潜在生物标记物,其中正谱13种,负谱15种.这些生物标记物主要影响戊糖和葡糖醛酸相互转化通路、核黄素代谢、泛酸和CoA合成、精氨酸和脯氨酸代谢、肠内菌代谢、嘌吟代谢、Vc代谢胆酸合成、色氨酸代谢等通路,五味子从能量代谢、氨基酸代谢、肠内菌代谢、脂类代谢等各个角度发挥治疗糖尿病并发症的作用.从各个通路的相关生物功能分析,五味子治疗糖尿病肾病的作用较强,此外还具有降脂、抗氧化的功效.     

基于药代动力学和代谢组学研究咖啡因对异丙嗪镇静作用的影响

采用药代动力学和代谢组学方法研究了咖啡因对异丙嗪镇静作用的影响. 首先, 采用航天警戒作业模拟系统和斯坦福睡眠量表, 比较了异丙嗪单独给药和联合咖啡因给药对志愿者认知功能的影响; 然后, 通过Kinetica软件计算异丙嗪在咖啡因联用前后的药代动力学参数变化, 并讨论可能的影响因素; 最后, 使用代谢组学方法对潜在生物标志物进行了鉴定. 实验结果表明, 咖啡因可以对抗异丙嗪的镇静作用, 并改善志愿者的认知功能. 这可能与联合给药后异丙嗪在体内代谢加快, 多巴胺, 去甲肾上腺素和肾上腺素相关通路发生变化有关. 该方法为异丙嗪和咖啡因联合用药研究提供了新思路.     

Untargeted Metabolomics Combined with Bioassay Reveals the Change in Critical Bioactive Compounds during the Processing of Qingzhuan Tea

Qingzhuan tea (QZT) is a typical Chinese dark tea that has a long-time manufacturing process. In the present study, liquid chromatography coupled with tandem mass spectrometry was used to study the chemical changes of tea samples during QZT processing. Untargeted metabolomics analysis revealed that the pile-fermentation and turnover (post-fermentation, FT) was the crucial stage in transforming the main compounds of QZT, whose contents of flavan-3-ols and flavonoids glycosides were decreased significantly. The bioactivities, including the antioxidant capacities and inhibitory effects on α-amylase and α-glucosidase, were also reduced after the FT process. It was suggested that although the QZT sensory properties improved following pile-fermentation and aging, the bioactivities remained restrained. Correlation analysis indicated that the main galloylated catechins and flavonoid glycosides were highly related to their antioxidant capacity and inhibitory effects on α-amylase and α-glucosidase.