Frozen leaves (2 g) were homogenized with 10 mL of 0.05 M sodium phosphate buffer solution (pH 7.8) with 1% polyvivylpyrrolidone and was centrifuged at 12,000× g at 4 °C for 15 min. The supernatant was collected for malondialdehyde (MDA) and enzyme analysis. MDA content was measured using spectrophotometric assay based on the thiobarbituric acid reaction [101 (link)]. Superoxide dismutase activity was analyzed using spectrophotometric method of Giannopolitis-Ries with measurements of the inhibition levels of nitroblue tetrazolium reduction at 560 nm [102 (link)]. Catalase activity was measured using spectrophotometric method of Abassi et al. which include the measuring of H2O2 extinction decline at 240 nm [103 (link)].
The total antioxidant capacity of D. palmatum extracts was determined using the phosphomolybdic acid method [104 (link)]; the DPPH• radical scavenging activity and ABTS•+ radical scavenging activity was assessed as described early [105 (link)]; the determination of superoxide anion scavenging activity (O2•−-SA) was measured in phenazine methosulfate-nicotinamide adenine dinucleotide-nitroblue tetrazolium systems using the method of Ozen et al. [106 (link)]; the Br• radical scavenging activity (Br•-SA) was determined using culometric method [105 (link)] with electrogenerated bromine radicals; the potential to inactivate NO was assessed by the sodium nitropusside technique [107 (link)]; the ability to inactivate H2O2 was determined using the technique developed by Badami and Channabasavaraj [108 (link)]; to evaluate the chelating activity for Fe2+ ions the o-phenanthroline technique was applied [59 (link)].
For preparation of the extract, accurately-weighed dried sample of D. palmatum herb (10 g) was transferred in a conical flask for preparation of the extracts. After that, 150 mL of solvent (60% methanol solutions) was added with stirring and put in an ultrasonic bath. The extraction conditions were 60 min at 45 °C, ultrasound power of 100 W, the frequency 35 kHz. The extraction was repeated twice. The obtained extract were filtered through a cellulose filter and combined. The filtrates were evaporated in vacuo until dryness with the use of a rotary evaporator.
The total antioxidant capacity of D. palmatum extracts was determined using the phosphomolybdic acid method [104 (link)]; the DPPH• radical scavenging activity and ABTS•+ radical scavenging activity was assessed as described early [105 (link)]; the determination of superoxide anion scavenging activity (O2•−-SA) was measured in phenazine methosulfate-nicotinamide adenine dinucleotide-nitroblue tetrazolium systems using the method of Ozen et al. [106 (link)]; the Br• radical scavenging activity (Br•-SA) was determined using culometric method [105 (link)] with electrogenerated bromine radicals; the potential to inactivate NO was assessed by the sodium nitropusside technique [107 (link)]; the ability to inactivate H2O2 was determined using the technique developed by Badami and Channabasavaraj [108 (link)]; to evaluate the chelating activity for Fe2+ ions the o-phenanthroline technique was applied [59 (link)].
For preparation of the extract, accurately-weighed dried sample of D. palmatum herb (10 g) was transferred in a conical flask for preparation of the extracts. After that, 150 mL of solvent (60% methanol solutions) was added with stirring and put in an ultrasonic bath. The extraction conditions were 60 min at 45 °C, ultrasound power of 100 W, the frequency 35 kHz. The extraction was repeated twice. The obtained extract were filtered through a cellulose filter and combined. The filtrates were evaporated in vacuo until dryness with the use of a rotary evaporator.
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