Plant-Based Secondary Metabolites for Health Benefits
Hafiz Ansar Rasul Suleria, Megh R. Goyal, Masood Sadiq Butt in Phytochemicals from Medicinal Plants, 2019
Understanding the chemical constituents profile gives us an idea about the nature of the biological activity the plant. The pharmacological attributes of plant extracts can be studied using either in vitro or in vivo or using both kinds of studies.24,49,60,100 Since, plant extracts contain a vast number of biomolecules, it is not easy to obtain absolutely pure bioactive compound. Therefore, studying the therapeutic activity is more fruitful using bioassay-guided approach. In this way toxicity of the phytoconstituents can also be monitored. In vitro biochemical assays are very helpful in identifying different biological activities, for example, antioxidant activity, antidiabetic activity, etc. Apart from this, the in vivo studies are very crucial to give us a better insight into how the molecule or plant extracts work in the living system.9,44
Why Analyze Drugs in Biological Fluids?
Joseph Chamberlain in The Analysis of Drugs in Biological Fluids, 2018
Although classical bioassays are not extensively used in modern drug research as assay methods for drugs, being largely superseded by more reliable and consistent chemical assays, there has been a resurgence of assay methods depending on specific biochemical phenomena, exemplified by immunoassays and receptor assays (see Chapter 8). Such biochemical events may be controlled by single molecular interactions, so that, in theory, such assay methods could be extremely sensitive. However, they may be less specific than chemical assays, since the biochemical event may also be triggered by closely related compounds such as metabolites. In the case of receptor assays, one could argue that this is not such a bad thing because the assay would then measure total potential activity of the sample being measured. However, this argument is flawed for at least two reasons: (1) the typical receptor assay measures molecules that bind to the receptor and does not distinguish between agonists and antagonists, and (2) not all molecules in the plasma may be able to penetrate to the site of action to the same extent as the active compound. The receptor activity, then, is no more valid than using total immunoactivity of plasma samples — the immunoactivity of a compound being quite incidental to its pharmacological effect.
The Role of the Macrophage in Immunity
Richard C. Niemtzow in Transmembrane Potentials and Characteristics of Immune and Tumor Cell, 2020
There has been a longstanding question as to whether macrophage activating factor and MIF are truly separate entities or whether they are really a single molecular species to which separate functional terms have been applied. For the most part, a variety of purification techniques have not separated MAF activity from MIF activity. Recently, however, several investigators have reported that techniques of gel filtration and affinity chromatography can separate the two factors.48, 49 These findings should be regarded as being encouraging, since one would like to see that at least some of the activities reported as being separate lymphokines are indeed separate molecular species. However, as these investigators and others before have found, one must be extremely cautious when working with highly purified factors, because of their instability. The factors might be active at one point in time but not the next. Additionally, it is conceivable that the sensitivity of one assay may exceed that of another and a single factor might be present in a high enough concentration to just barely initiate one cellular effect but not another.
The future of Extracellular Vesicles as Theranostics – an ISEV meeting report
Published in Journal of Extracellular Vesicles, 2020
Carolina Soekmadji, Bo Li, Yiyao Huang, Haifang Wang, Taixue An, Chunchen Liu, Weilun Pan, Jing Chen, Lesley Cheung, Juan Manuel Falcon-Perez, Yong Song Gho, Harry B. Holthofer, Minh T. N. Le, Antonio Marcilla, Lorraine O’Driscoll, Faezeh Shekari, Tang Long Shen, Ana Claudia Torrecilhas, Xiaomei Yan, Fuquan Yang, Hang Yin, Yu Xiao, Zezhou Zhao, Xue Zou, Qian Wang, Lei Zheng
Omics analyses are widely used in the discovery of disease-specific EV biomarkers, in order to globally assess EV cargo of proteins, nucleic acids, metabolites and lipids. For instance, proteomic approach may be performed to find enriched proteins that are involved in metabolic processes or signal transduction. Moreover, biochemical assays may help to characterize the activity of enzymes. Measurement of Enzymatic activity may provide some quantifiable features to assess EV functions. The importance of this is exemplified by the fact that EVs secreted by hepatocytes carry enzymes induced by exposure to liver toxins and drugs in both in vitro cell lines and in mouse model of drug-induced liver damage (DILI) [33,34]. Utilizing advanced mass spectrometry to detect active enzymes in hepatocyte-secreted EVs further suggests that EVs are associated with altering metabolic profile as a response to drugs [33,35].
Oxidative stress tolerance and antioxidant capacity of lactic acid bacteria as probiotic: a systematic review
Published in Gut Microbes, 2020
Free radical detection is widely used to evaluate the antioxidant activity of various antioxidants.156 Most antioxidant evaluation methods based on reactive species can be applied to probiotics to evaluate their antioxidant capacity.69,162 Radical production and scavenging systems are the most straightforward methods, and most of them use biochemical methods without the requirement for eukaryotic cells. These biochemical assays are mostly based on fluorescence or chromophore reactions, and thus are straightforward and cheap. Radical production and scavenging systems are normally used to detect the antioxidant ability of probiotic strains in vitro, but they have also been used to evaluate radical production after probiotic treatment or supplementation in vivo.5,163 Further, these systems can be applied to intact cells as well as cell-free extracts and cell lysates or their metabolic products.72,75,164,165 Reactive species tests have also been applied to evaluate the antioxidant capacity of LAB-fermented foods, especially, milk.166 The methods used to assess the antioxidative capacity of probiotic LAB based on the detection of reactive species are summarized in Table 1. Most studies combine assays to evaluate the antioxidant capacity of probiotic strains.72,181
Hep G2 cell culture confluence measurement in phase-contrast micrographs – a user-friendly, open-source software-based approach
Published in Toxicology Mechanisms and Methods, 2020
This limitation can be overcome by confirming data with additional biochemical assays, but this increases the cost of experiments (Riss et al. 2004). Alternatively, digital micrographs of label-free cells can provide useful insights on the numbers of viable cells, without the need of costly or cytotoxic fluorophores and fixation. Proliferation data can be extracted by counting single cell numbers, but this process is time consuming. This is impossible in the case of some of the most established adherent cell lines as Hep G2 because single cells in the monolayer cannot be easily discerned. In such cases confluence, or the fraction of surface, taken up by cells, can be measured. However, the application of digital image analysis by life scientists is often limited by the price of specialized software or by insufficient knowledge on sophisticated analysis methods, reviewed elsewhere (Vicar et al. 2019).
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