代谢组学方法研究水飞蓟宾对四氯化碳致小鼠肝损伤的保护作用

运用代谢组学方法研究四氯化碳(CCl4)致小鼠肝损及水飞蓟宾肝保护作用的机理. 通过气相色谱-质谱(GC-MS)技术分析CCl4及水飞蓟宾作用下的小鼠肝组织匀浆及血浆代谢物谱. 使用正交偏最小二乘判别分析法研究正常组与CCl4模型组之间的代谢物谱差异, 并通过变量重要性投影(VIP)选取肝组织与血浆中各12种标志性代谢物. 使用主成分分析法研究水飞蓟宾预防性及治疗性给药对CCl4肝损的干预效果. 通过比较选取的标志性代谢物的含量的差异, 探讨了水飞蓟宾对CCl4致小鼠肝损的保护机理. 结果表明, CCl4作用后, 小鼠机体能量代谢、氨基酸代谢及脂类代谢都受到不同程度的影响. 水飞蓟宾能有效地缓解CCl4所造成的体内线粒体功能及氨基酸代谢紊乱. 采用代谢组学方法能较全面地反应生物体的生理及代谢状态, 并可应用于物质毒性和药效评价研究.     

基于修饰组和探针分子的重要途径代谢物筛选和注释新方法北大核心CSCD

植物次生代谢物在抵御生物/非生物胁迫、生物间互作以及信息传递等方面发挥重要作用,次生代谢途径解析对植物分子育种、天然产物合成等方面具有重要意义。液相色谱-高分辨串联质谱(LC-HRMS/MS)为次生代谢物鉴定及途径表征提供了技术手段。非靶向LC-HRMS/MS方法可获得丰富的质谱信号,包括一级质谱和二级质谱(MS,MS/MS),但受质谱数据库规模以及次生代谢物复杂性的制约,次生代谢物注释十分困难。该研究以玉米叶片中苯丙烷途径代谢物为例,发展用于非靶向代谢组数据中重要途径代谢物的高效筛选和注释新方法。首先,利用公共代谢途径数据库及文献获取参与苯丙烷代谢途径的61种修饰反应类型,进而从非靶向实验数据中筛选出修饰代谢组。其次,获取开源串联质谱数据中的苯丙烷类化合物作为探针分子,构建探针分子质谱数据库。将探针分子与修饰代谢组共建分子网络,锁定目标途径代谢物并注释结构。该方法在正、负离子模式下分别筛选出玉米叶片中392个和417个苯丙烷途径候选代谢物,去冗余后共注释出129个代谢物,涉及苯丙烷代谢的主要分支途径,如黄酮途径的8个类黄酮、19个氧苷类黄酮和32个碳苷类黄酮,31个羟基肉桂酸途径代谢物以及22个木脂素途径代谢物;其中26个在PubChem和SciFinder数据库中未见收录。该研究利用探针分子结合修饰组可快速锁定途径代谢物,且有助于快速、准确的网络传播注释,可显著提高目标途径代谢物筛选与注释效率,为植物次生代谢途径的深入解析提供分析手段。     

硫酸铵提高裂殖壶菌不饱和脂肪酸积累的代谢组学研究

采用基于气相色谱-质谱(GC-MS)联用技术的代谢组学方法,通过分析裂殖壶菌胞内代谢物的变化,研究(NH4)2SO4提高裂殖壶菌不饱和脂肪酸积累的作用机制.利用GC-MS技术对添加0,1.5,2.5和3.5g/L (NH4)2SO4的发酵样品进行分析,结合K-Means算法对数据进行聚类分析,得到胞内代谢物的热图.主成分分析(PCA)和偏最小二乘判别分析(OPLS-DA)得分图可将不同添加条件下裂殖壶菌胞内物质明显区分.运用PLS-DA载荷图及模型的变量重要性因子(VIP)值,发现了7种代谢物可作为区别4种不同条件下的生物标志物.经NIST和Wiley谱库检索,它们分别为葡萄糖、肌醇、甘露醇、果糖、磷酸、乙醇胺、乙胺.代谢途径分析结果表明,(NH4)2SO4是通过弱化糖酵解途径和乙酸合成支路,提高苹果酸酶的活力,强化氨基酸代谢和NADPH的积累,从而提高不饱和脂肪酸积累.     

基于气相色谱-飞行时间质谱技术的佐剂性关节炎大鼠尿液代谢组学的研究

采用弗氏完全佐剂(FCA)诱导佐剂性关节炎(AA)大鼠模型,观察大鼠足趾肿胀度和踝关节组织的病理学形态变化。应用气相色谱-飞行时间质谱(GC-TOF MS)技术检测AA大鼠尿液代谢物谱,并对数据进行主成分分析(PCA)、偏最小二乘法-判别分析(PLS-DA)及正交偏最小二乘法-判别分析(OPLS-DA),探讨可能的发病机制。通过变量重要性投影值(VIP>1)和P值(<0.05),筛选出尿液中的差异代谢物。在模型组大鼠的尿液中共发现异柠檬酸、α-酮戊二酸、柠康酸、肌酸、3-羟基丁酸等20种差异代谢物。推断AA代谢组学的发病机制可能与能量代谢、氨基酸代谢、脂肪酸代谢途径有关。     

蛋白质组学和代谢组学在微生物代谢工程中的应用

构建微生物细胞工厂是化学品、生物能源以及药物分子可持续生产的可行性策略。然而，微生物的代谢复杂、调控严谨，制约着目标产物高效合成。蛋白质组学和代谢组学可以从系统生物学角度分析酶和代谢物组分，从而理解复杂的生物系统，为微生物代谢工程改造提供重要线索。该文介绍了蛋白质组学和代谢组学在微生物代谢工程中的应用，包括基因组尺度代谢模型构建、菌株生物合成优化、指导菌株耐受性改造、限速步骤预测、植物次级代谢途径挖掘，从而为微生物合成天然产物提供新的基因或途径。在此基础上，该文还展望了生物大数据未来的发展方向。     

基于核磁共振技术的临床尿毒症的代谢组学研究

采用核磁共振技术与统计学分析方法,结合尿液的生化指标,进行临床尿毒症尿样的代谢组学研究。结果表明,尿毒症患者和健康对照人群尿样的代谢轮廓存在明显差异,通过核磁共振氢谱鉴定出70种代谢物,其中20种具有显著差异。与健康人群相比,尿毒症患者尿液中2-羟基异丁酸、3-羟基丁酸、丙酮、丁酸、谷氨酸、肌氨酸、肌酐、赖氨酸、N,N-二甲基组氨酸、柠檬酸、天冬酰胺、乙醇和乙醇胺的含量明显偏低,而支链氨基酸(包括亮氨酸、缬氨酸、异亮氨酸)、牛磺酸、乳酸、α-葡萄糖和β-葡萄糖的含量明显偏高。这些代谢物涉及氨基酸代谢、能量代谢和脂质代谢中的多条代谢途径,可作为尿毒症代谢影响的潜在的生物标志物,有助于理解尿毒症发病的生化机制。     

基于气相色谱-质谱的烟草根部代谢组学方法建立及应用

建立了一种基于气相色谱-质谱的烟草根部代谢组学研究方法,方法采用甲基叔丁基醚-甲醇-水和二氯甲烷分别作为极性和非(弱)极性代谢物的萃取溶剂,采用10 mg冻干烟草根粉末对应1.5 m L甲基叔丁基醚-甲醇-水和100 mg冻干烟草根粉末对应5 m L二氯甲烷分别作为极性和非极性代谢物的最佳萃取方案。方法建立了烟草根部代谢物定性信息库,信息库包含了100个极性代谢物和32个非极性代谢物的质谱谱图、保留指数、化学结构等信息。将所建立的方法应用于红大与TN90根部代谢组差异研究,发现缬氨酸等8种氨基酸、乙醇酸等7种有机酸、果糖等4种糖类物质等在两种烟草根中含量有显著性差异。     

基于气相色谱-质谱法的广泛靶向代谢组学方法开发

气相色谱-质谱法(GC-MS)的四极杆检测器具有扫描速率低、离子流失率高、浓度检测范围窄的特点，这些缺陷限制了该技术在代谢组学领域的广泛应用，因此亟需建立一种基于GC-MS的高覆盖率代谢组学分析方法。本文提出了一种基于GC-MS的广泛靶向代谢组学方法，广泛靶向代谢组学结合了靶向和非靶向的优点，可以实现对代谢物质的定性和半定量检测，该方法以The Fiehn library(FiehnLib)数据库中的代谢物质信息为基础，建立直链脂肪酸甲酯(FAMEs)的保留时间与FiehnLib数据库中的保留指数(RI)的关系，根据FiehnLib数据库中的保留指数计算数据库中代谢物质在具体实验条件下的保留时间；对比分析并确定保留时间的阈值为0.15 min,优化最佳扫描间隔为0.20 s;优化代谢物质的定量离子以避免出峰时间相近离子的干扰；最终构建了含有611种代谢物质的选择离子监测(SIM)方法表，这611种代谢物质覆盖了KEGG(Kyoto Encyclopedia of Genes and Genomes)中65%的代谢通路。与全扫描非靶向GC-MS方法相比，该广泛靶向GC-MS方法所检测的代...     

微生物代谢组学的样品前处理

微生物代谢组学是指全面分析(定性和定量)细胞生长或生产周期某一时刻细胞内和细胞周围的所有低分子量代谢物.微生物代谢组学研究需要可靠和重现地分析细胞内较宽动力学浓度范围(nmol～mmol)、化学功能各异的代谢产物,因此,需要对从生物量培养、灭活和代谢产物的提取到代谢物的定量分析这整个实验过程提出耐用、可重现和可靠的实验方案.本文介绍了灭活过程中细胞内代谢产物泄漏的处理方法,较为详细地论述了通过灭活草案捕获代谢反应活性方法和代谢物提取方法的优化,并对微生物代谢组学样品前处理的目前发展趋势提出初步见解.     

代谢组学应用于糖肾方治疗糖尿病肾病的疗效评价

建立了基于超高效液相色谱-飞行时间质谱(UPLC/TOF MS)分析技术的血浆代谢指纹谱,应用多元统计分析方法评价糖尿病肾病患者血浆代谢物变化差异及糖肾方的干预效果。通过研究糖肾方干预糖尿病肾病血浆内源性代谢物的变化,探索与该疾病密切相关的代谢途径,评价糖肾方的治疗效果。结果发现: 经糖肾方治疗后,血浆内源性代谢物发生了明显变化,磷脂代谢、脂肪酸代谢、氨基酸代谢、嘌呤嘧啶代谢、固醇类代谢等多个代谢途径得到纠正。本研究基于UPLC/TOF MS的代谢组学方法,能够从整体水平反映疾病治疗过程中代谢网络的变化趋势,证实糖肾方具有治疗糖尿病肾病的临床疗效并有助于阐释药物作用机理。     

Zingiber officinale var. rubrum: Red Ginger’s Medicinal Uses

Zingiber officinale var. rubrum (red ginger) is widely used in traditional medicine in Asia. Unlike other gingers, it is not used as a spice in cuisines. To date, a total of 169 chemical constituents have been reported from red ginger. The constituents include vanilloids, monoterpenes, sesquiterpenes, diterpenes, flavonoids, amino acids, etc. Red ginger has many therapeutic roles in various diseases, including inflammatory diseases, vomiting, rubella, atherosclerosis, tuberculosis, growth disorders, and cancer. Scientific evidence suggests that red ginger exhibits immunomodulatory, antihypertensive, antihyperlipidemic, antihyperuricemic, antimicrobial, and cytotoxic activities. These biological activities are the underlying causes of red ginger’s therapeutic benefits. In addition, there have been few reports on adverse side effects of red ginger. This review aims to provide insights in terms the bioactive constituents and their biosynthesis, biological activities, molecular mechanisms, pharmacokinetics, and qualitative and quantitative analysis of red ginger.     

Ex vivo Application of δ-Aminolevulinic Acid Induces High and Specific Porphyrin Levels in Human Skin Tumors: Possible Basis for Selective Photodynamic Therapy

Abstract— In photodynamic therapy with topically applied δ-aminolevulinic acid porphyrins are acting as photosensitiz-ers. The profile of porphyrin metabolites in normal or in neoplastic skin after administration of δ-aminolevulinic acid has not been determined in detail yet. Thus, to study porphyrin biosynthesis in human skin an organ culture model was developed. Explant pieces of normal skin, ker-atoacanthoma, and basal cell carcinoma were incubated with 1 niM δ-aminolevulinic acid for 36 h. Levels of δ-aminolevulinic acid, porphyrins and porphyrin metabolites were measured in tissues and supernatants. After incubation with δ-aminolevulinic acid, higher porphyrin levels were demonstrated in tumors as compared to normal skin. In supernatants, most of formed porphyrins, preferentially highly carboxylated porphyrin metabolites, were measured. The pattern of synthesized porphyrins differed between normal and neoplastic skin explants. In tissues of basal cell carcinomas protoporphyrin was preferentially shown and tissues of keratoacanthomas were characterized by a predominance of coproporphyrin as compared to normal skin. The results show that explant cultures offer an easy approach to examine the porphyrin biosynthesis of various tissues. The tumor-specific δ-aminolevulinic acid metabolism indicates additional porphyrin metabolites such as coproporphyrin apart from protoporphyrin as effective photosensitizers and may offer a novel approach to tumor-selective photodynamic damage.     

Rapid Identification of 3,6′-Disinapoyl Sucrose Metabolites in Alzheimer’s Disease Model Mice Using UHPLC–Orbitrap Mass Spectrometry

Alzheimer’s disease (AD) is a degenerative disease of the central nervous system characterized by the progressive impairment of neural activity. Studies have shown that 3,6′-disinapoyl sucrose (DISS) can alleviate the pathological symptoms of AD through the activation of the cAMP/CREB/BDNF signaling pathway. However, the exact biochemical mechanisms of action of DISS are not clear. This study explores metabolism of DISS in an AD mouse model, induced by the microinjection of a lentiviral expression plasmid of the APPswe₆₉₅ gene into CA1 of the hippocampus. After gavage administration of DISS (200 mg/kg), the kidneys, livers, brains, plasma, urine, and feces were collected for UHPLC–Orbitrap mass spectrometry analysis. Twenty metabolites, including the prototype drug of DISS, were positively or tentatively identified based on accurate mass measurements, characteristic fragmentation behaviors, and retention times. Thus, the metabolic pathways of DISS in AD mice were preliminarily elucidated through the identification of metabolites, such as ester bond cleavage, demethoxylation, demethylation, and sinapic acid-related products. Furthermore, differences in the in vivo distribution of several metabolites were observed between the model and sham control groups. These findings can provide a valuable reference for the pharmacological mechanisms and biosafety of DISS.     

Metabolic alterations in triptolide‐induced acute hepatotoxicity

Triptolide, a major active constitute of Tripterygium wilfordii Hook. F, is prescribed for the treatment of autoimmune diseases in China. One of its most severe adverse effects observed in the clinical use is hepatotoxicity, but the mechanism is still unknown. Therefore, the present study applied an LC/MS‐based metabolomic analysis to characterize the metabolomic changes in serum and liver induced by triptolide in mice. Mice were administered triptolide by gavage to establish the acute liver injury model, and serum biochemical and liver histological analyses were applied to assess the degree of toxicity. Multivariate data analyses were performed to investigate the metabolic alterations. Potential metabolites were identified using variable importance in the projection values and Student's t‐test. A total of 30 metabolites were observed that were significantly changed by triptolide treatment and the abundance of 29 metabolites was correlated with the severity of toxicity. Pathway analysis indicated that the mechanism of triptolide‐induced hepatotoxicity was related to alterations in multiple metabolic pathways, including glutathione metabolism, tricarboxylic acid cycle, purine metabolism, glycerophospholipid metabolism, taurine and hypotaurine metabolism, pantothenate and CoA biosynthesis, pyrimidine metabolism and amino acid metabolism. The current study provides new mechanistic insights into the metabolic alterations that lead to triptolide‐induced hepatotoxicity.     

Serum metabolomic profiling in patients with Alzheimer disease and amnestic mild cognitive impairment by GC/MS

The aim of this study was to characterize the serum metabolic profiles of patients with Alzheimer’s disease (AD) and amnestic mild cognitive impairment (AMCI) using metabolomics based on gas chromatography–mass spectrometry (GC/MS). Serum samples were collected from patients with AD (n = 30) and AMCI (n = 32), and normal healthy controls (NOR, n = 40). Metabolite profiles were performed with GC/MS in conjunction with multivariate statistical analysis, and possible biomarker metabolites were identified. Thirty-one kinds of endogenous metabolites could be identified simultaneously. Eleven components were chosen as biomarker metabolites between AD and NOR groups, and these metabolites were closely related to seven biological pathways: arginine and proline metabolism, phenylalanine metabolism, β-alanine metabolism, primary bile acid synthesis, glutathione metabolism, starch and sucrose metabolism, and steroid hormone biosynthesis. Meanwhile, 10 components were chosen as biomarker metabolites between AMCI and NOR groups and seven biological pathways were closely related: arginine and proline metabolism, phenylalanine metabolism, citrate cycle, alanine, aspartate and glutamate metabolism, taurine and hypotaurine metabolism, starch and sucrose metabolism, and steroid hormone biosynthesis. Our study distinguished serum metabotypes between AD, AMCI and NOR patients successfully. The implementation of this metabolomic strategy may help to develop biochemical insight into the metabolic alterations in AD/AMCI and will be helpful for the further understanding of pathogenesis.     

Metabolome-Based Discrimination Analysis of Shallot Landraces and Bulb Onion Cultivars Associated with Differences in the Amino Acid and Flavonoid Profiles

Shallot landraces and varieties are considered an important genetic resource for Allium breeding due to their high contents of several functional metabolites. Aiming to provide new genetic materials for the development of a novel bulb onion cultivar derived from intraspecific hybrids with useful agronomic traits from shallots, the metabolic profiles in the bulbs of 8 Indonesian shallot landraces and 7 short-day and 3 long-day bulb onion cultivars were established using LC–Q-TOF-MS/MS. Principal component analysis, partial least squares discriminant analysis, and dendrogram clustering analysis showed two major groups; group I contained all shallot landraces and group II contained all bulb onion cultivars, indicating that shallots exhibited a distinct metabolic profile in comparison with bulb onions. Variable importance in the projection and Spearman’s rank correlation indicated that free and conjugated amino acids, flavonoids (especially metabolites having flavonol aglycone), and anthocyanins, as well as organic acids, were among the top metabolite variables that were highly associated with shallot landraces. The absolute quantification of 21 amino acids using conventional HPLC analysis showed high contents in shallots rather than in bulb onions. The present study indicated that shallots reprogrammed their metabolism toward a high accumulation of amino acids and flavonoids as an adaptive mechanism in extremely hot tropical environments.     

Effect of Calcium Compound Type and Dosage on the Properties of Acid Rennet Goat’s Milk Gels

The aim of this study was to determine the effect of adding calcium compounds to processed goat’s milk, and on the properties of acid rennet goat’s milk gels, which are a middle product obtained in the manufacture of acid rennet cheese. The properties of the gels directly affect the quality of acid rennet cheeses. The analysis of raw goat’s milk was carried out, then acid rennet gels were produced with the addition of six different calcium compounds (chloride, citrate, bisglycinate, gluconate, lactate, and carbonate). The dynamics of milk fermentation were performed by monitoring the pH value of milk during acidification. The pH, syneresis, color, and texture profile were determined in the formulated acid rennet gels. An organoleptic evaluation was also performed. The study demonstrated that, not only calcium chloride, but also calcium citrate, gluconate, lactate, bisglycinate, and calcium carbonate could be used in the production of goat’s milk acid rennet gels, or the middle product in the manufacture of acid rennet curd cheese from goat’s milk. Notably, the addition of citrate, bisglycinate, and calcium carbonate in doses of 20 mg Ca 100 g⁻¹ most effectively reduced syneresis compared to the control sample by 4.76% (citrate), 7.85% (bisglycinate), and 10.28% (carbonate). The hardness of the control gels ranged from 2.35 N to 2.99 N. The addition of chloride, citrate, gluconate, lactate, and calcium carbonate to the milk improved the acid rennet gel’s hardness. The addition of 20 mg Ca 100 g⁻¹ as gluconate increased the hardness the most (3.61 N). When increasing the calcium dosage in the form of all compounds, there was a tendency to increase the gel’s springiness. The addition of chloride, citrate, and bisglycinate to milk did not result in a darkening of the gel’s color. The addition of calcium compounds mostly reduced the intensity of goatish taste and odor. Calcium gluconate, in particular, reduced the goatish taste the most, a taste which is not always acceptable by the consumers.     

In Silico Analysis of Metabolites from Peruvian Native Plants as Potential Therapeutics against Alzheimer’s Disease

Background: Despite research on the molecular bases of Alzheimer’s disease (AD), effective therapies against its progression are still needed. Recent studies have shown direct links between AD progression and neurovascular dysfunction, highlighting it as a potential target for new therapeutics development. In this work, we screened and evaluated the inhibitory effect of natural compounds from native Peruvian plants against tau protein, amyloid beta, and angiotensin II type 1 receptor (AT1R) pathologic AD markers. Methods: We applied in silico analysis, such as virtual screening, molecular docking, molecular dynamics simulation (MD), and MM/GBSA estimation, to identify metabolites from Peruvian plants with inhibitory properties, and compared them to nicotinamide, telmisartan, and grapeseed extract drugs in clinical trials. Results: Our results demonstrated the increased bioactivity of three plants’ metabolites against tau protein, amyloid beta, and AT1R. The MD simulations indicated the stability of the AT1R:floribundic acid, amyloid beta:rutin, and tau:brassicasterol systems. A polypharmaceutical potential was observed for rutin due to its high affinity to AT1R, amyloid beta, and tau. The metabolite floribundic acid showed bioactivity against the AT1R and tau, and the metabolite brassicasterol showed bioactivity against the amyloid beta and tau. Conclusions: This study has identified molecules from native Peruvian plants that have the potential to bind three pathologic markers of AD.     

Sensory Quality Evaluation of Korla Pear from Different Orchards and Analysis of Their Primary and Volatile Metabolites

Metabolites play vital roles in shaping the quality of fresh fruit. In this study, Korla pear fruit harvested from twelve orchards in South Xinjiang, China, were ranked in sensory quality by fuzzy logic sensory evaluation for two consecutive seasons. Then, gas chromatography-mass spectrometry (GC-MS) was applied to determine the primary metabolites and volatile compounds. Sensory evaluation results showed that the panelists were more concerned about ‘mouth feel’ and ‘aroma’ than about ‘fruit size’, ‘fruit shape’ and ‘peel color’. In total, 20 primary metabolites and 100 volatiles were detected in the pear fruit. Hexanal, (E)-2-hexenal, nonanal, d-limonene, (Z)-3-hexen-1-yl acetate and hexyl acetate were identified as the major volatile compounds. Correlation analysis revealed that l-(+)-tartaric acid, hexanoic acid, trans-limonene oxide and 2,2,4-trimethyl-1,3-pentanediol diisobutyrate were negatively correlated with sensory scores. Furthermore, OPLS-DA results indicated that the fruit from three orchards with lower ranks in quality could be distinguished from other samples based on the contents of l-(+)-tartaric acid and other eight metabolites, which were all associated with ‘mouth feel’ and ‘aroma’. This study reveals the metabolites that might be closely associated with the sensory quality attributes of Korla pear, which may provide some clues for promoting the fruit quality in actual production.