2 2 azinobis 3 ethylbenzothiazoline 6 sulfonic acid (abts)

Manufactured by Merck Group

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ABTS is a laboratory reagent used for the detection and quantification of peroxidase activity. It is a colorimetric substrate that undergoes a color change when oxidized by peroxidases, allowing for spectrophotometric or colorimetric analysis.

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Lab products found in correlation

2 2 diphenyl 1 picrylhydrazyl (dpph), by Merck Group (293 mentions) Trolox, by Merck Group (166 mentions) Gallic acid, by Merck Group (124 mentions) Potassium persulfate, by Merck Group (88 mentions) Quercetin, by Merck Group (68 mentions) Folin ciocalteu reagent, by Merck Group (67 mentions) Methanol, by Merck Group (51 mentions) Ascorbic acid, by Merck Group (50 mentions) Dimethyl sulfoxide (dmso), by Merck Group (33 mentions) Sodium hydroxide (naoh), by Merck Group (33 mentions) Sodium carbonate, by Merck Group (32 mentions) Catechin, by Merck Group (31 mentions) Rutin, by Merck Group (29 mentions) Ethanol, by Merck Group (28 mentions) Chlorogenic acid, by Merck Group (25 mentions) Ferulic acid, by Merck Group (23 mentions) Acetonitrile, by Merck Group (21 mentions) Acetic acid, by Merck Group (21 mentions) Formic acid, by Merck Group (20 mentions) Ferrozine, by Merck Group (19 mentions)

817 protocols using 2 2 azinobis 3 ethylbenzothiazoline 6 sulfonic acid (abts)

ABTS radical scavenging assay for PTX

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The free radical scavenging capacity of PTX was also studied using the ABTS radical cation de-colorization assay which is based on the reduction of ABTS^+• radicals by antioxidants [17 (link)].
ABTS (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in PBS to reach a final concentration of 7.4 mM. ABTS radical cation (ABTS^+•) was produced by a chemical reaction of ABTS with 2.6 mM potassium persulfate (Sigma-Aldrich St. Louis, MO, USA). The reaction was carried out overnight, in the dark, and at room temperature. The ABTS^+• solution was then diluted in methanol to reach an absorbance of 0.75 at 734 nm. A BioTek PowerWave HT, microplate spectrophotometer (BioTek Instruments, Inc., Winoosky, VT, USA) was used.
Different concentrations of PTX dissolved in acetone (Sigma-Aldrich St. Louis, MO, USA) (20 µL) were allowed to react with 200 µL of ABTS^+• solution into the wells of a 96-well microplate kept in the dark. Absorbance kinetics were measured at room temperature starting at 5 min after initial mixing. All solutions were used the same day of their preparation, and all determinations were carried out in triplicate.

Binatti E., Zoccatelli G., Zanoni F., Donà G., Mainente F, & Chignola R. (2021). Effects of Combination Treatments with Astaxanthin-Loaded Microparticles and Pentoxifylline on Intracellular ROS and Radiosensitivity of J774A.1 Macrophages. Molecules, 26(17), 5152.

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TEAC Assay for Antioxidant Capacity

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The TEAC assay was performed adapting the method described by Re et al. [100 (link)] to a microplate reader. 2,2′-Azinobis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS, Sigma-Aldrich, Oakville, ON, Canada) radical cations were prepared by mixing potassium persulfate 2.45 mM (final concentration) and an aqueous solution of ABTS 7 mM (final concentration) in the dark at room temperature for 12–16 h. The ABTS radical cation solution was diluted in PBS (pH 7.4) to an absorbance of 0.40 at 734 nm ± 0.02. A standard calibration curve of Trolox (0–16 μM) was constructed. A volume of 10 μL of Trolox or extracts diluted in PBS were added in the wells of a 96 well-plate (Costar, MERCK, Darmstadt, Germany) with 200 μL of diluted ABTS. Afterwards the absorbance reading at 734 nm was taken 6 min after initial mixing using an Infinite200 Pro plate reader (Tecan, Männedorf, Swizerland). Appropriate solvent blanks were run in each plate. The lipidic extract was assayed in at least three separate dilutions and in triplicate. The inhibition of absorbance at 734 nm of the lipidic extract was plotted as a function of concentration of Trolox and the TEAC value expressed as Trolox equivalent (in micromolar) per g of lipidic extract, using Magellan v 7.2 software (Tecan, Männedorf, Switzerland).

Stabili L., Acquaviva M.I., Angilè F., Cavallo R.A., Cecere E., Del Coco L., Fanizzi F.P., Gerardi C., Narracci M, & Petrocelli A. (2019). Screening of Chaetomorpha linum Lipidic Extract as a New Potential Source of Bioactive Compounds. Marine Drugs, 17(6), 313.

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ABTS Radical Scavenging Assay

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2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS, Sigma-Aldrich, ST Louis, MO, USA) was dissolved in water to make a final 7 mM concentration. ABTS radical cation was produced a reacting ABTS stock solution with 2.45 mM potassium persulfate standing on the dark at room temperature for 24 h. The ABTS solution was diluted with ethanol at absorbance of 0.7 nm. And then mixed by adding 0.1 mL of the antioxidant solutions (resveratrol (trans-3,4,5-trihydroxystilbene), Vitamin E and trolox, Sigma-Aldrich, USA) to make the reaction mixtures in a final volume of 1 mL. The reaction mixtures were shaken vigorously and kept in the dark for 6 min at 37°C. The absorbance of the remaining ABTS radical cation was determined at 734 nm by spectrophotometer (Genesys10-S, Thermo, Inc., Hudson, NH, USA). trolox equivalent antioxidant capacity (TEAC, μM trolox equivalent to each antioxidant) values were calculated by the equation (difference of sample absorbance [Δsample abs]/difference of trolox absorbance [Δtrolox abs]).

Kim J.H., Park E.Y., Ha H.K., Jo C.M., Lee W.J., Lee S.S, & Kim J.W. (2015). Resveratrol-loaded Nanoparticles Induce Antioxidant Activity against Oxidative Stress. Asian-Australasian Journal of Animal Sciences, 29(2), 288-298.

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ABTS Radical Scavenging Activity Assay

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The ABTS radical scavenging activity of the sample was investigated by using a modified method from a previous study [16 (link)]. Briefly, the ABTS• stock solution (7 mM) was prepared by reacting ABTS (Sigma-Aldrich, Singapore) with 2.45 mM potassium persulfate (Sigma-Aldrich, Singapore) solution at room temperature under dark conditions for 12 h. A working ABTS• solution was then prepared by diluting the stock solution with distilled water. The working reagent was adjusted to obtain an absorbance of 0.80 ± 0.05 at 734 nm. Then, 200 μL of sample or Trolox (Sigma-Aldrich, Singapore) were reacted with 1.8 mL of working ABTS• solution for 30 min before measuring at 734 nm. The ABTS radical scavenging activity was calculated and compared with the Trolox standard curve. The result was expressed as mg Trolox equivalent/g sample.

Rodboon T., Okada S, & Suwannalert P. (2020). Germinated Riceberry Rice Enhanced Protocatechuic Acid and Vanillic Acid to Suppress Melanogenesis through Cellular Oxidant-Related Tyrosinase Activity in B16 Cells. Antioxidants, 9(3), 247.

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Antioxidant Capacity Assay with ABTS

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ABTS (Sigma-Aldrich, Inc., St. Louis, MO, USA) was dissolved in phosphate-buffered saline (PBS, pH 7.4) to 7 mM. Then, the ABTS solution was mixed with potassium persulfate solution that was dissolved in PBS to 2.45 mM. The mixture was stored in the dark at room temperature for 24 h. After the ABTS^•+ radical was formed, the ABTS solution was diluted with PBS to an absorbance of 0.70 (±0.02) at a wavelength of 734 nm. Then, 990 µL of ABTS radical solution and 10 µL of fungal extract (10 mg/mL in DMSO) were reacted in a cuvette. The absorbance was measured at 734 nm after 6 min using a spectrophotometer (Sunrise^TM, Tecan, Männedorf, Switzerland).

Heo Y.M., Kim K., Ryu S.M., Kwon S.L., Park M.Y., Kang J.E., Hong J.H., Lim Y.W., Kim C., Kim B.S., Lee D, & Kim J.J. (2018). Diversity and Ecology of Marine Algicolous Arthrinium Species as a Source of Bioactive Natural Products. Marine Drugs, 16(12), 508.

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ABTS Radical Cation Assay for Antioxidant Capacity

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The DPPH assay was performed as previously described in [36 (link)]. In detail, the 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonate) radical cation (ABTS^•+) solution was prepared by the reaction of 7 mM ABTS^•+ (Sigma-Aldrich, St. Louis, MO, USA) and 2.45 mM potassium persulphate (Merck KGaA, Darmstadt, Germany) after incubation at room temperature in the dark for 12–16 h. The (ABTS^•+) solution was thus diluted with ethanol to obtain an absorbance of 0.700 ± 0.020 at 734 nm. After the extension of 3.0 mL of diluted (ABTS^•+) solution (A734 nm = 0.700 ± 0.020) to 30 μL of ethanolic extracts, the absorbance reading at 30 °C, t = 6 min after initial mixing (A sample) was acquired. A calibration curve was obtained using the Trolox (Aldrich, Steinheim, Germany) standard ethanolic solution, and the results were expressed in terms of Trolox Equivalent Antioxidant Capacity (TEAC) as mg Trolox equivalents per mL. All determinations were carried out in triplicates.

Christofi A., Margariti G., Salapatas A., Papageorgiou G., Zervas P., Karampiperis P., Koukourikos A., Tarantilis P.A., Kaparakou E.H., Misiakos K, & Makarona E. (2023). Determining the Nutrient Content of Hydroponically-Cultivated Microgreens with Immersible Silicon Photonic Sensors: A Preliminary Feasibility Study. Sensors (Basel, Switzerland), 23(13), 5937.

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Antioxidant Activity Assessment of Formulation

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An anti-oxidant activity of both the selected formulation and the extract was performed by DPPH and ABTS scavenging assay (Meda et al., 2005 (link); Thaipong et al., 2006 (link)). For DPPH scavenging assay, the tested formulation and the extract were dissolved in methanol and mixed with 1.5 mL of 80 µM DPPH solution in methanol. After 15 min of incubation in the dark, the absorbance of the reaction was determined at 517 nm UV–V is spectrophotometer.
For ABTS scavenging assay, an aliquot solution of 7 mM ABTS (Merck, Germany) was oxidized with potassium persulfate for 16 h in the dark at room temperature. The ABTS+solution was diluted with phosphate buffer saline (PBS; Merck, Darmstadt, Germany) pH 7.4 to initial an absorbance of 0.70 ± 0.02 at 734 nm. An aliquot (20 µL) of each formulation was mixed with 2 mL ABTS+solution for 6 min in the dark. The absorbance was read at 734 nm using UV-visible spectrophotometer. Trolox was used as a positive control for both experiments. The radical scavenging activity was calculated according to the following formula;

% I n h i b i t i o n = [(b l a n k a b s o r b a n c e - s a m p l e a b s o r b a n c e) ∕ b l a n k a b s o r b a n c e] \times 100

Chokpaisarn J., Chusri S., Amnuaikit T., Udomuksorn W, & Voravuthikunchai S.P. (2017). Potential wound healing activity of Quercus infectoria formulation in diabetic rats. PeerJ, 5, e3608.

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Preparation and Use of ABTS and BPO Solutions

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The chemical reagents used throughout this study were analytical grade and utilized without any further purification. The deionized water was purified by Milli-Q, Millipore apparatus. ABTS was obtained from Sigma-Aldrich (Sigma-Aldrich, Oakville, Canada). BPO was purchased from Panreac (Panreac, Barcelona, Spain). Ethanol 96% came from Merck (Merck, Darmstadt, Germany).
The stock solution of chromogenic reagent as ABTS 1000 mg L⁻¹ was prepared by dissolving 0.1 g of ABTS in 100 mL of deionized water.
1000 mg L⁻¹ BPO stock solution was generated by dissolving BPO 0.1 g in 100 mL of 96% ethanol. The working solution of BPO was prepared fresh daily by diluting appropriate volume of stock BPO in 96% ethanol.

Ponhong K., Supharoek S.A., Siriangkhawut W, & Grudpan K. (2015). A rapid and sensitive spectrophotometric method for the determination of benzoyl peroxide in wheat flour samples. Journal of Food and Drug Analysis, 23(4), 652-659.

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Antioxidant Potential Evaluation by ABTS Assay

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The ABTS (2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid) assay was used to determine the radical scavenging (antioxidant) effect of TCT using vitamin C (Sigma-Aldrich, St. Louis, MO, USA) as a positive control. ABTS is a chromogenic free radical generated overnight by combining it with potassium persulfate (7.4 mM ABTS (Merck Millipore, Burlington, MA, USA) + 10% 24.5 mM K-persulfate). Antioxidant properties of the compounds were determined by the decolorization of the intense green ABTS radical solution. The ABTS solution was prepared as described [51 (link)]. The ABTS solution was diluted with 50 mM Gly-HCl buffer to reach a concentration where the absorbance of the solution was A = 1 (1 cm light path at 405 nm) in a Spark photometer (Tecan Spark, Männedorf, Switzerland). Three-fold serial dilutions of TCT and vitamin C were pipetted into 96-well plates containing the assay solution and after 30 min incubation time, absorbance was measured at 405 nm.

Nagy-Pénzes M., Hajnády Z., Regdon Z., Demény M.Á., Kovács K., El-Hamoly T., Maléth J., Hegyi P., Hegedűs C, & Virág L. (2022). Tricetin Reduces Inflammation and Acinar Cell Injury in Cerulein-Induced Acute Pancreatitis: The Role of Oxidative Stress-Induced DNA Damage Signaling. Biomedicines, 10(6), 1371.

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Antioxidant Activity of Camel Milk Yogurt

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The radical scavenging assay was performed using 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) (Sigma-Aldrich, St. Louis, MO, USA) stock solution, then 14.8 mM of ABTS solution was incorporated into potassium persulfate (1:1, v/v) (5 mM) solution. This solution was incubated at 25 °C in the dark for 16 h. Later, the solution was added to distilled water until obtaining an absorbance of 734 nm. Then, 20 μL digested camel milk yogurt was diluted to 180 μL ABTS+ solution and incubated at 25 °C in the dark for 15 min. A solution of 180 μL ABTS+ solution and 20 μL distilled water was considered a control [39 (link),49 (link)]. ABTS+ scavenging activity was then calculated as follows (Equation (2)):

ABTS radical scavenging activity (%) = (\frac{A b s c o n t r o l - A b s s a m p l e}{A b s c o n t r o l}) \times 100

The DPPH radical scavenging assay was performed using 2,2-difenil-1-picrilhidracilo (DPPH) (Sigma-Aldrich, St. Louis, MO, USA)) stock solution. We mixed 20 μL of yogurt extract with 180 μL 0.1 mM DPPH solution and incubated this at 25 °C in the dark for 30 min. The absorbance was recorded at 515 nm [50 (link),51 (link)]. A solution of 180 μL DPPH+ solution and 20 μL ethanol was considered a control. The DPPH scavenging activity was calculated as shown in Equation (3):

DPPH Inhibition (%) = (\frac{A b s c o n t r o l - A b s s a m p l e}{A b s c o n t r o l}) \times 100

Marcia J.A., Aleman R.S., Kazemzadeh S., Manrique Fernández V., Martín Vertedor D., Kayanush A, & Montero Fernández I. (2023). Isolated Fraction of Gastric-Digested Camel Milk Yogurt with Carao (Cassia grandis) Pulp Fortification Enhances the Anti-Inflammatory Properties of HT-29 Human Intestinal Epithelial Cells. Pharmaceuticals, 16(7), 1032.

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