| Chromatographic Analysis for Targeted Metabolomics of Antioxidant and Flavor-Related Metabolites in Tomato
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Author:
Date:
2021-03-05
[Abstract] Targeted metabolomics is a useful approach to evaluate crop breeding studies. Antioxidant and flavor-related traits are of increasing interest and are considered quality traits in tomato breeding. The present study presents chromatographic methods to study antioxidants (carotenoids, vitamin C, vitamin E, phenolic compounds, and glutathione) and flavor-related characters (sugars and organic acids) in tomato. Two different extraction methods (for polar and apolar entities) were applied to isolate the targeted compounds. The extraction methods developed in this work were time and cost-effective since no further purification was needed. Carotenoids, vitamin C, glutathione, and phenolic acids were analyzed by HPLC-PDA using a RP C18 column at an appropriate wavelength for each compound. ...
[摘要] [摘要]靶向代谢麦克风是评估作物育种研究的有用方法。抗氧化剂和与风味有关的性状受到越来越多的关注，被认为是番茄育种中的品质性状。本研究提供了色谱方法来研究番茄中的抗氧化剂（类胡萝卜素，维生素C，维生素E，酚类化合物和谷胱甘肽）和与风味相关的特征（糖和有机酸）。应用了两种不同的提取方法（针对极性和非极性实体）来分离目标化合物。由于不需要进一步纯化，因此这项工作中开发的提取方法既省时又经济。使用RP C18色谱柱，通过RP-C18色谱柱在适当波长下对每种化合物进行类胡萝卜素，维生素C，谷胱甘肽和酚酸的分析。用RP C18和NH2色谱柱通过HPLC分析维生素E和糖，并分别通过FLD和RI检测器进行检测。此外，MSTFA衍生化后，使用Rtx 5DA色谱柱使用GC-FID分析有机酸。结果，开发了定量重要植物代谢物的灵敏分析方法，并在本文中进行了描述。这些方法不仅适用于番茄，但都还表征其他物种的香味相关的和抗氧化的化合物。因此，这些协议可用于指导作物育种的选择。
[背景]代谢组学是其受到关注的其潜在的援助作物育种研究施加生化途径。在番茄育种中，由于消费者对提高作物品质的偏好，抗氧化剂和与风味有关的代谢产物都受到了越来越多的关注。人类无法自己合成抗氧化剂分子; ...
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| Measurement of Ascorbic Acid and Glutathione Content in Cyanobacterium Synechocystis sp. PCC 6803
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Author:
Date:
2020-10-20
[Abstract] Ascorbic acid (AsA) and gluthathione (GSH) are two key components of the antioxidant machinery of eukaryotic and prokaryotic cells. The cyanobacterium Synechocystis sp. PCC 6803 presents both compounds in different concentrations (AsA, 20-100 μM and GSH, 2-5 mM). Therefore, it is important to have precise and sensitive methods to determine the redox status in the cell and to detect variations in this antioxidants. In this protocol, we describe an improved method to estimate the content of both antioxidants (in their reduced and oxidized forms) from the same sample obtained from liquid cultures of Synechocystis sp. PCC 6803.
[摘要] [摘要] 抗坏血酸（AsA）和明胶硫（GSH）是真核细胞和原核细胞抗氧化机制的两个重要组成部分。蓝藻Synechocystis sp.pcc6803以不同浓度（AsA，20-100μM和GSH，2-5mm）呈现这两种化合物。因此，有精确和敏感的方法来确定细胞中的氧化还原状态和检测这种抗氧化剂的变化是很重要的。在该方案中，我们描述了一种改进的方法，用以从Synechocystis sp.pcc6803的液体培养中获得的同一样品中估算两种抗氧化剂（以还原形式和氧化形式）的含量。
[背景] 细胞中的氧化还原状态可以被多种因素改变，产生氧化应激。我们使用该方案来量化暴露于50℃高温下的Synechocystis sp. PCC 6803中的GSH和AsA含量。如拟南芥所述，热胁迫导致抗氧化剂含量下降，并通过铁作用引起细胞死亡(Distefano et al.， 2017;Aguilera等人，2019年预印本)。虽然本方案是针对Synechocystis sp. PCC 6803制定的，但也可用于测定其他蓝藻中GSH和AsA的含量。蓝藻中AsA含量很低，正常条件下uM含量在一定范围内，某些处理下pM含量在一定范围内。因此，我们提出了这个敏感的方法与一个改进的细胞裂解程序。
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| Fluorometric Estimation of Glutathione in Cultured Microglial Cell Lysate
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Author:
Date:
2017-06-05
[Abstract] Glutathione is one of the major antioxidant defense components present in cells. It is predominantly present as reduced glutathione (GSH) and converted into oxidized glutathione (GSSG) while reducing the free radicals like hydroxyl ions (OH-). For the measurement of GSH and GSSG, o-phthalaldehyde (OPT) has been used as a fluorescent reagent. O-phthalaldehyde has an ability to react specifically with GSH at pH 8 and GSSG at pH 12 respectively. N-ethylmaleimide (NEM) has been used to prevent auto-oxidation of GSH during measurement of GSSG in the present protocol. The original protocol by Hissin and Hilf was developed for glutathione estimation in Rat liver tissue. The present protocol has been standardized following Hissin and Hilf (1976) for the estimation of ...
[摘要] 谷胱甘肽是细胞中存在的主要抗氧化防御成分之一。它主要作为还原型谷胱甘肽（GSH）存在并转化为氧化型谷胱甘肽（GSSG），同时还可以减少自由基，如羟基离子（OH）。对于GSH和GSSG的测定，邻苯二甲醛（OPT）已被用作荧光试剂。邻苯二甲醛具有与pH8的GSH和pH12的GSSG特异性反应的能力。已经使用N-甲基马来酰亚胺（NEM）来在本方案中测量GSSG期间防止GSH的自动氧化。 Hissin和Hilf的原始方案被开发用于大鼠肝组织中的谷胱甘肽估计。目前的方案已经在Hissin和Hilf（1976）之后被标准化，用于估计培养的小胶质细胞裂解物中的谷胱甘肽，但它也可以用于其它哺乳动物细胞裂解物。在我们的实验中，相同的方案已被用于估计小鼠神经母细胞瘤细胞N2a的全细胞裂解物中的谷胱甘肽。
背景 该方法于1976年由Hissin和Hilf在分析生物化学方面出版（Hissin和Hilf，1976）。然而，有可用于准确检测GSH的方法;由于GSH容易氧化转化为GSSG，所以大多数方法给出过高的GSSG。 ...
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