Tocopherols were extracted from 100 mg frozen tissues in 6 mL hexane using the MagNALyser and centrifuged at 14,000× g for 15 min. Extracts were dried (CentriVap concentrator, Labconco, Kansas City, MO, USA) and the dried extract (CentriVap concentrator, Labconco, KS, USA) was resuspended in hexane, and tocopherols were separated and quantified by HPLC (Shimadzu, ‘s Hertogenbosch, The Netherlands) (normal phase conditions, Particil Pac 5 µm column material, length 250 mm, i.d. 4.6 mm). Dimethyl tocol (DMT) was used as the internal standard (5 ppm). Data were analysed with Shimadzu Class VP 6.14 software.
Class vp 6
Manufactured by Shimadzu
Sourced in Japan
The Class VP 6.14 software is a data processing and analysis software developed by Shimadzu for laboratory applications. It provides tools for acquiring, managing, and analyzing data from various Shimadzu instruments. The software's core function is to facilitate the collection, storage, and interpretation of data generated by Shimadzu's analytical equipment.
Automatically generated - may contain errors
Lab products found in correlation
High performance liquid chromatography (hplc), by Shimadzu (8 mentions)
Centrivap concentrator, by Labconco (6 mentions)
Magna lyser, by Roche (2 mentions)
Particil pac 5 m column material, by Shimadzu (1 mentions)
Centrivap, by Labconco (1 mentions)
Pac 5 m column material, by Shimadzu (1 mentions)
Lc 10at pump, by Shimadzu (1 mentions)
Sil 10a autosampler, by Shimadzu (1 mentions)
Spd m10a dad, by Shimadzu (1 mentions)
C18 column, by Shimadzu (1 mentions)
Lc 2010a, by Shimadzu (1 mentions)
Lc 10ad pump, by Shimadzu (1 mentions)
Sil 20ac prominence autosampler, by Shimadzu (1 mentions)
Gstf04700, by Merck Group (1 mentions)
13 protocols using class vp 6
1
Tocopherol Extraction and Quantification
2
Comprehensive Antioxidant Analysis in Plants
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TAC (ferric reducing antioxidant power, FRAP) was measured in ice-cold 80% ethanol with a MagNALyser (Roche, Vilvoorde, Belgium) and quantified using Trolox as a reference, as reported by Benzie and Strain (1999) (link). Plant samples were extracted in 80% ethanol and measured for ascorbate and glutathione using a MagNALyser (Roche, Vilvoorde, Belgium). Reduced ascorbate (ASC) and glutathione (GSH) were measured using HPLC. To extract phenols and flavonoids, fresh plant materials were homogenized in 80% ethanol before being centrifuged at 5000 rpm for 15 minutes. The clear extract was then used to measure the phenols and flavonoid concentrations using the Folin-Ciocalteu and aluminum chloride assays, respectively (AbdElgawad et al., 2023 (link)). Tocopherols were extracted with hexane (100 mg FW in 6 ml hexane) and centrifuged for 15 minutes at 14,000 g. The extracts were dried (Labconco, Kansas, USA) and resuspended in hexane. HPLC (Shimadzu, Hertogenbosch, the Netherlands) was used to separate and quantify tocopherols under normal phase conditions. Particil Pac 5 mm column material, length 250 mm, i.d. 4.6 mm, and internal standard dimethyl tocol (DMT) (5 ppm). The Shimadzu Class VP 6.14 software was used to examine the data.
3
Tocopherol Quantification by HPLC
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According to the procedures outlined by AbdElgawad et al. [36 (link)], tocopherols were extracted in n-hexane solvent and measured by HPLC (Shimadzu, Kyoto, Japan) under normal phase conditions (Partisil Pac 5 m column material, length 250 mm, i.d. 4.6 mm). As an internal standard, dimethyl tocol (DMT; 5 ppm) was also applied. The HPLC system’s included Shimadzu Class VP 6.14 software was used to analyze the data.
4
Quantification of Tocopherols in Plant Tissue
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One hundred milligram (FW) of frozen plant tissue was extracted in 6 ml of hexane, and centrifuged at 14,000 × g for 15 min. Extracts were dried (CentriVap concentrator, Labconco, Kansas City, MO, USA) and resuspended again in hexane. Tocopherols were separated and quantified by HPLC analysis (Shimadzu, Hertogenbosch, The Netherlands, normal phase conditions, Particil Pac 5 mm column material, length 250 mm, i.d. 4.6 mm). Dimethyl tocol (DMT) was used as internal standard (5 ppm). Data were analyzed with Shimadzu Class VP 6.14 software.
5
Polyphenol, Flavonoid and Tocopherol Analysis
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To extract polyphenols and flavonoids, a known weight of fine powdered plant tissues was homogenized in 80% ethanol. The slurry was centrifuged (5000 g for 15 min), and the clear extract was used to quantify total phenolics and flavonoids using Folin-Ciocalteu and AlCl3 methods, respectively. Additionally, tocopherols were extracted in hexane, followed by evaporation using CentriVap concentrator (Labconco, Kansas, USA), and the dry pellet was resuspended in hexane. At the end of the extraction, tocopherols were separated, and their levels were determined by HPLC (Shimadzu, ‘s Hertogenbosch, The Netherlands) coupled with a fluorometric detector (excitation at 290 nm and emission at 330 nm). Tocopherols were separated on normal phase conditions, Particil Pac 5 μm column material, length 250 mm, i.d. 4.6 mm. The mobile phase was applied at a flow rate of 0.45 ml min−1. Dimethyl tocol (DMT) was used as internal standard (5 ppm). Data were analyzed with Shimadzu Class VP 6.14 software.
In 80% ethanol extract, the total antioxidant capacity (TAC, FRAP) was determined. Centrifugation was done at 14000 g, 4°C, for 25 min, and then the FRAP test [TPTZ (0.01 mM) in HCl (0.04 mM), acetate buffer (0.3 M, pH3.6), and FeCl3.6H2O (0.02 M)] was performed by using Trolox (0 to 650 M), as already described (Benzie and Strain, 1996 (link)).
In 80% ethanol extract, the total antioxidant capacity (TAC, FRAP) was determined. Centrifugation was done at 14000 g, 4°C, for 25 min, and then the FRAP test [TPTZ (0.01 mM) in HCl (0.04 mM), acetate buffer (0.3 M, pH3.6), and FeCl3.6H2O (0.02 M)] was performed by using Trolox (0 to 650 M), as already described (Benzie and Strain, 1996 (link)).
6
Tocopherols Determination by HPLC
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Tocopherols were determined by HPLC analysis according to (AbdElgawad et al., 2015 (link)). Tocopherols were extracted with hexane using the MagNALyser (Roche, Vilvoorde, Belgium; 1 min, 7000 rpm). The dried extract (CentriVap concentrator, Labconco, KS, United States) was resuspended in hexane, and tocopherols were separated and quantified by HPLC (Shimadzu, ‘s Hertogenbosch, The Netherlands) (normal phase conditions, Particil Pac 5 μm column material, length 250 mm, i.d. 4.6 mm). Dimethyl tocol (DMT) was used as internal standard (5 ppm). Data were analyzed with Shimadzu Class VP 6.14 software.
7
Tocopherols and Antioxidant Capacity in Wheat
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Tocopherols were extracted in n-hexane solvent and quantified by HPLC (Shimadzu, Hertogenbosch, The Netherlands) using normal phase conditions (Particil Pac 5 μm column material, length 250 mm, i.d. 4.6 mm), based on the methods described by AbdElgawad et al. (2015) (link). Dimethyl tocol (DMT; 5 ppm) was also used as an internal standard. Data were analyzed with Shimadzu Class VP 6.14 software provided by the HPLC system.
The ferric reducing antioxidant power (FRAP) was measured to evaluate total antioxidant capacity in durum wheat grains, as fully described by AbdElgawad et al. (2021) . Briefly, the extraction was done by adding ethanol (80% v/v) and centrifuging at 14,000 for 20 min. For 30 minutes at room temperature, FRAP reagent (20 mM FeCl3 in 0.25 M acetate buffer, pH 3.6) was combined with a known volume of the produced extract. A multi-mode microplate reader was used to measure the absorbance at 517 nm.
The ferric reducing antioxidant power (FRAP) was measured to evaluate total antioxidant capacity in durum wheat grains, as fully described by AbdElgawad et al. (2021) . Briefly, the extraction was done by adding ethanol (80% v/v) and centrifuging at 14,000 for 20 min. For 30 minutes at room temperature, FRAP reagent (20 mM FeCl3 in 0.25 M acetate buffer, pH 3.6) was combined with a known volume of the produced extract. A multi-mode microplate reader was used to measure the absorbance at 517 nm.
8
Tocopherol Extraction and Quantification
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Tocopherols were extracted with hexane using the MagNALyser. The dried extract (CentriVap concentrator, Labconco, Kansas, USA) was resuspended in hexane and tocopherols were separated and quantified by HPLC (Shimadzu, 's Hertogenbosch, The Netherlands) (normal phase conditions, Particil Pac 5 µm column material, length 250 mm, id. 4.6 mm). Dimethyl tocol (DMT) was used as an internal standard (5 ppm). Data were analyzed with Shimadzu Class VP 6.14 software.
9
Ascorbate and Tocopherol Quantification
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Ascorbate content was determined in 100 mg dry plant tissue by reversed phase HPLC. Separation and detection processes by UV detector were performed following Potters et al. 34 protocol. Total ascorbate concentration was determined after reduction with DTT (0.04 M) for 10 min at room temperature. The redox status was calculated as the ratio of the reduced form to the total concentration. Tocopherols were extracted in hexane 35 . The extract was dried under vacuum conditions (CentriVap concentrator, Labconco, KS, USA) and was re-suspended in hexane.
Tocopherols were separated and quantified by HPLC (Shimadzu, Hertogenbosch, Netherlands) using normal phase conditions (Particil Pac 5 µm column material, length 250 mm, i.d. 4.6 mm). 5,7dimethyltocol (DMT; 5 ppm) was used as an internal standard. Data were analyzed with Shimadzu Class VP 6.14 software provided by the HPLC system (Shimadzu, Tokyo, Japan).
Tocopherols were separated and quantified by HPLC (Shimadzu, Hertogenbosch, Netherlands) using normal phase conditions (Particil Pac 5 µm column material, length 250 mm, i.d. 4.6 mm). 5,7dimethyltocol (DMT; 5 ppm) was used as an internal standard. Data were analyzed with Shimadzu Class VP 6.14 software provided by the HPLC system (Shimadzu, Tokyo, Japan).
10
Tocopherols Extraction and Quantification
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Tocopherols were extracted in hexane (100 mg FW in 6 ml hexane), and centrifuged at 14,000g for 15 min. Extracts were dried (CentriVap concentrator, Labconco, Kansas, USA) and resuspended in hexane. Tocopherols were separated and quantified by HPLC (Shimadzu, Hertogenbosch, The Netherlands), normal phase conditions, Particil Pac 5 mm column material, length 250 mm, i.d. 4.6 mm, with dimethyl tocol (DMT) as internal standard (5 ppm). Data were analyzed with Shimadzu Class VP 6.14 software.
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