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Modern extraction methods and standardization of extracts
Published in C. P. Khare, Evidence-based Ayurveda, 2019
The third and more scientific method is bioactivity guided assay. These are relatively simple at a basic level or can be extended to clinical studies that define dose response, safety and efficacy. Extracts are then expressed in terms of the activity of mg/kilo body weight for the indication prescribed. As an example, the antioxidant activity can be determined by the Ferric Reducing Ability of Plasma (FRAP) assay and 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) assay methods.
Antioxidant assays
Published in Roger L. McMullen, Antioxidants and the Skin, 2018
Similar to the TRAP assay, the original FRAP assay was designed to measure the ferric reducing ability of plasma (FRAP).36 However, the assay has undergone revisions and is now commonly employed to measure the antioxidant capacity of substances other than plasma, especially extracts that can easily be dissolved in a suitable solvent.37–43 The assay is carried out using a complex of 2,4,6-tripyridyl-s-triazine (TPTZ) and the ferric (Fe3+) form of iron. Figure 6.12 contains the structure of the organic ligand, TPTZ, in which case two moles of TPTZ complex with one Fe3+ atom. At low pH, the ferric iron present in this complex may be reduced to ferrous iron (Fe2+) in the presence of antioxidants, as shown in Equation 6.5:
Nutrition and Nutraceutical Supplements for the Treatment of Hypertension
Published in Stephen T. Sinatra, Mark C. Houston, Nutritional and Integrative Strategies in Cardiovascular Medicine, 2015
Oxidative stress, with an imbalance between ROS and RNS and the antioxidant defense mechanisms, contributes to the etiology of hypertension in animals21 and humans.22,23 ROSs and RNSs are generated by multiple cellular sources, including NADPH oxidase, mitochondria, xanthine oxidase, uncoupled endothelium-derived NO synthase (U-eNOS), cyclo-oxygenase, and lipo-oxygenase.22 Superoxide anion is the predominant ROS produced by these tissues, which neutralizes NO and also leads to downstream production of other ROSs (Figure 9.3). Hypertensive patients have impaired endogenous and exogenous antioxidant defense mechanisms,24 an increased plasma oxidative stress, and an exaggerated oxidative stress response to various stimuli.24,25 Hypertensive subjects also have lower ferric reducing ability of plasma (FRAP), lower vitamin C levels, and increased plasma 8-isoprostanes correlate with both an increase in systolic blood pressure (SBP) and diastolic blood pressure (DBP). Various single-nucleotide polymorphisms (SNPs) in genes that codify for antioxidant enzymes are directly related to hypertension.26 These include NADPH oxidase, xanthine oxidase, superoxide dismutase (SOD) 3, catalase, GPx 1 (glutathione peroxidase), and thioredoxin. Antioxidant deficiency and excess free radical production have been implicated in human hypertension in numerous epidemiologic, observational, and interventional studies24,25,27 (Figure 9.7). ROSs directly damage endothelial cells; degrade NO; influence eicosanoid metabolism; oxidize low-density lipoprotein (LDL), lipids, proteins, carbohydrates, DNA, and organic molecules; increase catecholamines; damage the genetic machinery; and influence gene expression and transcription factors.1,22–25 The interrelations of neurohormonal systems, oxidative stress, and CVD are shown in Figure 9.8. The increased oxidative stress, inflammation, and autoimmune vascular dysfunction in human hypertension result from a combination of increased generation of ROSs and RNSs, an exacerbated response to ROSs and RNSs, and a decreased antioxidant reserve.24–29 Increased oxidative stress in the rostral ventrolateral medulla (RVLM) enhances glutamatergic excitatory inputs and attenuates γ-aminobutyric acid (GABA)-ergic inhibitory inputs to the RVLM, which contributes to increased sympathetic nervous system (SNS) activity from the paraventricular nucleus.30 Activation of AT1R in the RVLM increases NADPH oxidase and increases oxidative stress and superoxide anion and increases SNS outflow causing an imbalance of SNS/parasympathetic nervous system (PNS) activity with elevation of BP, increased heart rate, and alterations in heart rate variability and heart rate recovery time, which can be blocked by AT1R blockers.30,31
Evaluation of oxidative stress biomarkers and antioxidant parameters in allergic asthma patients with different level of asthma control
Published in Journal of Asthma, 2022
Behnaz Karadogan, Sengul Beyaz, Asli Gelincik, Suna Buyukozturk, Nazli Arda
Ferric reducing antioxidant power (FRAP) was determined usingthe method of Benzie and Strain (23), with a slight modification. FRAP assay is a colorimetric method that relies on the ferric-reducing ability of plasma at low pH. This reduction by antioxidants creates ferrous ions (Fe2+) with a blue color. This method was modified for the 96-well microplate reader. FRAP reagent was prepared by mixing sodium acetate buffer (300 mM, pH 3.6), TPTZ (2,4,6-tris(2-pyridyl)-s-triazine) (10 mM in 40 mMHCl), and FeCl3.6H2O (20 mM) in a volume ratio of 10:1:1, respectively. Ten microliters of plasma were mixed with FRAP reagent and incubated at 37 °C for 30 min. Absorbance was measured at 593 nm. A standard curve was generated using different concentrations (5–100 nmol/mL) of ferrous sulfate heptahydrate (FeSO4.7H2O) as a reference in order to evaluate total antioxidant status. The results were expressed in nmol/mL of plasma.
Analysis of bioactive components in Ghost chili (Capsicum chinense) for antioxidant, genotoxic, and apoptotic effects in mice
Published in Drug and Chemical Toxicology, 2020
Sarpras M, Sushil Satish Chhapekar, Ilyas Ahmad, Suresh K. Abraham, Nirala Ramchiary
The 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity was determined by using the previously reported method (Abraham and Khandelwal 2013). For obtaining the calibration curve, five concentrations of ascorbic acid (100 µg–6.25 µg) was used. FRAP (ferric reducing ability of plasma) assay was performed by adopting the method of Benzie and Strain (1996) with slight modification. Test samples (10 µl) were added in to 310 µl freshly prepared FRAP reagent and 31 µl of distilled water. After 5 min incubation at 37 °C the absorbance was measured at 593 nm. Plasmid DNA nicking assay method was adopted from Lee et al. (2002). This assay was performed by using pBR322 supercoiled DNA plasmid. Quercetin 50 mM solution was used as a control.
Sour Cherries but Not Apples Added to the Regular Diet Decrease Resting and fMLP-Stimulated Chemiluminescence of Fasting Whole Blood in Healthy Subjects
Published in Journal of the American College of Nutrition, 2018
Piotr Bialasiewicz, Anna Prymont-Przyminska, Anna Zwolinska, Agata Sarniak, Anna Wlodarczyk, Maciej Krol, Jaroslaw Markowski, Krzysztof P. Rutkowski, Dariusz Nowak
Concentrations of selected polyphenols and their metabolites (dihydrocaffeic acid, vanillic acid, caffeic acid, homovanillic acid, hippuric acid, 4-hydroxyhippuric acid, 3-hydroxyhippuric acid, chlorogenic acid, 3,4-dihydroxybenzoic acid, 3-hydroxyphenylacetic acid, urolithin A) were determined in plasma and urine with a combination of solid-phase extraction technique with high-performance liquid chromatography with electrochemical (HPLC-ECD) or ultraviolet-visible detection (HPLC-UV/VIS), as previously described (19,25). Total phenolic compound concentrations were measured by the Folin-Ciocalteau method (26) and expressed as catechin equivalent in millimoles per liter or millimoles per gram of creatinine for plasma or spot morning urine, respectively. Plasma uric acid and urinary creatinine were determined with an Alpha Diagnostics Creatinine kit and Uric Acid kit (Alpha Diagnostics, Warsaw, Poland). Blood cell count was measured with a Micros Analyzer OT 45 (ABX, Montpellier, France). Plasma 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity (DPPH test) was measured according to our previously described procedure (27). Measurement of ferric-reducing ability of plasma (FRAP) was conducted following the procedure originally described by Benzie and Strain (28) with some modifications (26). Both FRAP and DPPH tests were performed with native plasma samples and those deprived of uric acid by incubation with uricase and catalase (nonurate plasma) (25). Apple and cherry phenolics were determined using HPLC according to previously described protocols (29–31). Fruit glucose, fructose, and saccharose were determined with a combination of solid-phase extraction and HPLC (32).