Assessment of in vitro antioxidant capacity and polyphenolic composition of selected medicinal herbs from Leguminosae family in Peninsular Malaysia

Determination of total phenolic (TPC), flavonoid and anthocyanin contents, and various antioxidant activities (2,2-diphenyl-1-picrylhydrazyl radical scavenging, ferric reducing power, ferrous ion chelating and lipid peroxidation inhibition) of leaves and flowers of Bauhinia kockiana, Caesalpinia pulcherrima and Cassia surattensis were performed in this study. The B. kockiana flower was found to possess the highest TPC (8280 ± 498 mg GAE/100 g), free radical scavenging activity (ascorbic acid equivalents 14,600 ± 2360 mg AA/100 g) and reducing ability (72.4 ± 8.7 mg GAE/g). Rutin and chlorogenic acid were detected in the plants, where the C. pulcherrima leaf contained the highest amount of rutin (669 ± 26 mg/100 g), while minute amounts of chlorogenic acid were detected in C. surattensis leaf (9.13 ± 0.44 mg/100 g). The C. pulcherrima leaf displayed the highest ferrous ion chelating and lipid peroxidation inhibition activities. Positive correlation was observed between TPC and various antioxidant activities.     

Polyphenol contents and in vitro antioxidant activities of lyophilised aqueous extract of kiwifruit (Actinidia deliciosa)

The aim of this study was to determine the antioxidant potency and total phenolic and flavonoid contents of kiwifruit (Actinidia deliciosa) in vitro by analysing the radical scavenging activity of lyophilised water extract from kiwifruit (LEK) for 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), 1,1-diphenyl-2-picryl-hydrazyl (DPPH), N,N-dimethyl-p-phenylenediamine (DMPD), and superoxide anion radical (O₂⁻) as well as the total reducing power by FRAP and CUPRAC assays and the metal chelating activities. LEK showed efficient radical scavenging activity with DPPH, ABTS, DMPD, and O₂⁻ radicals; ferric (Fe³⁺) and cupric (Cu²⁺) ion reducing power and metal chelating activities. Moreover, the amounts of phenolic compounds, such as caffeic acid, ferulic acid, syringic acid, ellagic acid, catechol, pyrogallol, p-hydroxy benzoic acid, vanillin, p-coumaric acid, gallic acid, quercetin, ??-tocopherol and ascorbic acid, in LEK were quantified by LC-MS-MS. The results show that pyrogallol (2070.0 mg/kg LEK) is the main phenolic compound responsible for the antioxidant and radical scavenging activities of LEK. Finally, total phenolic and flavonoid contents were determined as gallic acid (GAE) and quercetin equivalents (QE). The GAE and QE values in LEK were 16.67 ± 2.83 ??g GAE/mg and 12.95 ± 0.52 ??g QE/mg, respectively. The results suggest that consumption of kiwifruit (A. deliciosa) can be beneficial effects due to its antioxidant properties.     

Antioxidant activity of three edible seaweeds from two areas in South East Asia

Total phenolic content (TPC) and antioxidant activity (AOA) of 50% aqueous methanol extracts of the marine algae, Padina antillarum, Caulerpa racemosa and Kappaphycus alvarezzi were studied. TPC was measured using Folin-Ciocalteu method while 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, ferric reducing antioxidant power (FRAP), ferrous ion chelating (FIC) assay and beta carotene bleaching (BCB) assay were used to study their AOA. P. antillarum was found to have the highest TPC, 2430±208 mg gallic acid equivalents (GAE) per 100 g dried sample and ascorbic acid equivalent antioxidant capacity (AEAC), 1140±85 mg AA/100 g. C. racemosa and K. alvarezzi displayed lower TPC and AEAC. C. racemosa had 144±22 mg GAE/100 g dried sample of TPC and 14.3±2.0 mg AA/100 g of AEAC, while K. alvarezzi had 115±35 mg/100 g dried sample of TPC and 37.8±16.8 mg AA/100 g of AEAC. In addition, P. antillarum displayed the highest reducing power, 15.7±2.6 mg GAE/g and highest chelating ability. C. racemosa and K. alvarezzi exhibited lower reducing power, 0.737±0.423 mg GAE/g and 0.561±0.269 mg GAE/g, and lower chelating ability. However, the AOA of these three seaweeds as assessed by BCB assay were equally high.     

Antioxidant activity and proline content of leaf extracts from Dorystoechas hastata

Dorystoechas hastata (D. hastata) is a monotypic plant endemic to Antalya province of Turkey. D. hastata leaves are used to make a tea locally called “çalba tea”. Diethyl ether (E), ethanol (A), and water (W) were used for the sequential preparation of extracts from dried D. hastata leaves. A hot water extract (S) was also prepared by directly boiling the powdered plant in water. The antioxidant activities of the extracts were tested by ferric thiocyanate (FTC) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging methods. E extract exhibited the greatest antioxidant activity with FTC method, whereas S extract exhibited the lowest IC₅₀ value (6.17 ± 0.53 μg/ml) for DPPH radical scavenging activity. Total phenolic contents of the extracts were estimated by Folin–Ciocalteu method and S extract was found to contain the highest amount (554.17 ± 20.83 mg GAE/g extract) of phenolics. Extract A contained highest flavonoid content and there was a inverse linear correlation (R² = 0.926) between IC₅₀ values for DPPH radical scavenging activity and flavonoid contents of all extracts. Reducing power of extracts increased in a concentration-dependent manner. S extract was found to possess higher reducing power than equivalent amount of ascorbic acid at 20 and 25 μg/ml concentrations. Linear correlation between reducing power and concentration of E, A, and W extracts (R² > 0.95) was observed. A, W, and S extracts contained relatively high levels of proline. The results presented suggest that D. hastata may provide a natural source of antioxidants and proline.     

Antioxidant activity of compounds isolated from the pyroligneous acid, Rhizophora apiculata

The polyphenols (CPAE II) was isolated from the dichloromethane extract of the pyroligneous acid, Rhizophora apiculata by simultaneous acid base and solvent extraction method. Its qualitative and quantitative composition was studied by gas chromatography mass spectroscopy (GC/MS) and out of 57 peaks, 52 compounds were identified, representing 95.47% of the total polyphenols. The CPAE II was then fractionated to four fractions (F1–F4) by means of thin layer chromatography and silica gel column chromatography with dichloromethane, dichloromethane/chloroform/ethyl acetate mixture (8:1:1; 4:3:3, v/v/v), and ethyl acetate, respectively. The antioxidant properties of the CPAE II and the fractions were evaluated. Among the four fractions, fraction 1 (F1) was the most potent in DPPH radical scavenging activity and molybdenum (VI) reducing power. It was subjected to further purification by means of silica gel column chromatography with hexane, hexane/diethyl ether mixture (9:1, 6:1, 3:1, v/v), and diethyl ether, respectively. 2,6-Dimethoxyphenol (syringol) and dihydroxybenzenes (catechol and 3-methoxycatechol) were isolated and identified by GC/MS, ¹H NMR, ¹³C NMR spectral analyses, and confirmed by GC co-injection with authentic standards. Syringol, catechol and 3-methoxycatechol constitute 39.08, 4.21 and 1.10% of F1, respectively. Their antioxidant activities were evaluated by DPPH radical scavenging activity, ABTS radical cation scavenging activity, phosphomolybdenum and ferric reducing antioxidant power (FRAP). The trend in antioxidant capacity was similar in all the four assays, with dihydroxybenzenes > 2,6-dimethoxyphenols, although discrepancies in the ranking within the dihydroxybenzenes were present. These three compounds which showed significant antioxidant activities were isolated for the first time from the pyroligneous acid, R. apiculata.     

Phenolic content and antioxidant capacity of Philippine sweet potato (Ipomoea batatas) varieties

The study established baseline data on the total phenolic content and antioxidant activities of five sweet potato (Ipomoea batatas) varieties grown in the Philippines including Dakol, Emelda, Haponita, PSBSP and Violet. Phenolic content ranged from 192.7 to 1159.0 mg gallic acid equivalent (GAE) /100 g dry sample. Antioxidant activities were highest for Dakol, with an EC₅₀ value of 0.7 ± 0.2 mg/mL for DPPH radical scavenging activity, 2.5 ± 0.5 mg/mL for reducing power, and 2.4 ± 0.3 mg/mL for iron-chelating ability, on a dry basis. However, Haponita had the best inhibitory action on linoleic acid oxidation at 99.4 ± 0.9%. Methanolic sweet potato extracts had higher radical scavenging activity, reducing power and oxidation inhibition than α-tocopherol and higher iron-chelating capacity than ethylenediamine tetraacetic acid (EDTA). Significant (∗P < 0.05) negative correlation was observed between total phenolic content and the EC₅₀ for DPPH radical scavenging activity (R = −0.826), reducing power (R = −0.876) and iron-chelating capacity (R = -0.800).     

Antioxidant activity of microwave-assisted extract of Buddleia officinalis and its major active component

Buddleia officinalis Maxim, commonly used as rice dye for festivals, was extracted with ethanol using microwave-assisted extraction and Soxhlet extraction. The antioxidant activities of microwave-assisted extract of B. officialis (MEB) and Soxhlet extract of B. officianils (SEB) at the optimum extraction conditions were evaluated and compared with synthetic antioxidant butylated hydroxytoluene (BHT) employing DPPH free radical assay, ABTS assay, total antioxidant activity and reducing power. MEB and SEB had stronger antioxidant activities than BHT in all assays except reducing power, and the effects decreased as follows: MEB > SEB > BHT. The total phenolic contents of MEB and SEB reached 113.56 mg/g and 100.94 mg/g dry weight of extract, respectively, expressed as pyrocatechol equivalents, while the total flavonoids contents were 75.33 mg/g and 62.56 mg/g dry weight of extract, respectively, expressed as catechin equivalents (P < 0.05). Higher phenolic and flavonoids compounds may be major contributors to their higher antioxidant activities. Following activity-oriented separation, luteolin was isolated as an active principle, which exhibited excellent free radical scavenging activities with DPPH IC₅₀ 3.09 μg/ml and ABTS IC₅₀ 2.20 μg/ml.     

Free radical scavenging activities and bioactive substances of Jerusalem artichoke (Helianthus tuberosus L.) leaves

The total phenolic content and radical scavenging activities of Jerusalem artichoke (Helianthus tuberosus L.) leaves were investigated. Results indicated that the ethyl acetate fraction contained the highest total phenolic content (266.69 ± 2.51 mg GAE/g dry extract) accompanied with strongest free radical scavenging abilities. Following an in vitro radical scavenging activity-guide fractionation procedure, six phenolic compounds which strongly quenched free radicals were separated from ethyl acetate fraction. Among them, 3-O-caffeoylquinic acid and 1,5-dicaffeoylquinic acid played a dominant role due to their strong free radical scavenging abilities and their high contents. The content of 3-O-caffeoylquinic acid in n-butanol fraction was 74.58 ± 1.05 mg/g, while 1,5-dicaffeoylquinic acid in ethyl acetate fraction was 104.51 ± 2.86 mg/g. The results imply that the leaves of Jerusalem artichoke might be a potential source of natural antioxidants.     

Cytoprotective and antioxidant activity of free,conjugated and insoluble-bound phenolic acids from swallow root (Decalepis hamiltonii)

Swallow root (Decalepis hamiltonii) was extracted for free (SRFP), conjugated (SRCP) and insoluble-bound phenolic acids (SRIBP), and evaluated for cytoprotectivity, 1,1,diphenyl-2-picrylhydrazyl (DPPH) scavenging ability, reducing power and protection to DNA damage. In addition, the constituent phenolic acids in the extracts were also analysed. Results indicated a total phenol content of 20.72, 7.97 and 11.52 mg gallic acid equivalents (GAE)/g for SRFP, SRCP and SRIBP extracts, respectively. At 0.12 μg/mL concentration SRCP showed 87% cytoprotection (on NIH 3T3 cells) compared to SRFP (47%) and SRIBP (65%). DPPH radical scavenging activity indicated an IC₅₀ of 0.046, 0.06 and 0.128 μg/mL for SRCP, SRIBP and SRFP, respectively. Also, SRCP showed higher reducing power and DNA protectivity (80%). HPLC analysis of phenolic acid extracts showed the presence of hydroxybenzoate and cinnamate derivatives. Among the phenolics identified gallic, gentisic, protocatechuic and p-coumaric acids were the major contributors to antioxidant activity.     

Determination of antioxidative potential of lichen Usnea ghattensis in vitro

The study was aimed at evaluating the antioxidant activities of extract of Usnea ghattensis. The antioxidant activity, reducing power, superoxide anion radical scavenging and free radical scavenging activities were studied. The antioxidant activity increased with the increasing amount of extracts (2-20 mg/ml) added to the linoleic acid emulsion. Lipid peroxidation upto 73.3% was inhibited by the extract of 20 mg/ml and 39.2% by α-tocopherol at the same concentration. These effects were statistically significant (r²=0.876,P<0.01) when compared with control. However, the extract had no significant effect on superoxide anion scavenging by the PMS/NADH-NBT method. Like antioxidant activity, the reducing power and free radical scavenging activity of extract depends on concentration and increasing with increased amount of sample. The reducing power and DPPH radical-scavenging activity of U. ghattensis extract were found greater than the BHA and BHT. The results obtained in the present study indicate that U. ghattensis is a potential source of natural antioxidant.     

Standardised Mangifera indica extract is an ideal antioxidant

The standardised ethanolic and aqueous extracts of Mangifera indica leaf were prepared and analysed for their free radicals scavenging activity. The IC₅₀ values using the DPPH assay were 0.17 ± 0.02 and 0.49 ± 0.4 mg/ml, respectively. Standardised ethanolic extracts of the M. indica leaf had a solid content of 9.1 ± 0.7%, mangiferin concentration of 73 ± 0.17 mg/g of dry weight of the extract, free radical scavenging activity (IC₅₀) of 0.17 ± 0.02 mg/ml and total phenolic content of 590 ± 48 mg/g of extract. The protection exhibited by these extracts against lipid peroxidation was superior to butylated hydroxytoluene (BHT) and commercial grape seed extract. These extracts at higher concentration did not exhibit pro-oxidant activities when compared to vitamin C. Our findings also show that the aqueous and ethanolic extracts of M. indica leaf protect NIH/3T3 cells from oxidant-induced cell death.     

Antioxidative and anti-inflammatory properties of Citrus sulcata extracts

Citrus sulcata was subjected to ultrasound, high-pressure, and Soxhlet extractions. The antioxidant and anti-inflammatory activities of the extracts were evaluated qualitatively and quantitatively. The antioxidant content of the peel extract was twice that of the fruit extract. The quantitative analysis showed that the narirutin and hesperidin contents in the peel extracts were 8.8 and 7.5 mg/100 g, respectively. These extracts had a total phenolic content of 112.22 ± 2.89 gallic acid equivalent (GAE) mg/100 g, a total flavonoid content of 54.09 ± 1.01 rutin equivalent (RE) mg/100 g, DPPH radical scavenging activity of 46%, and antioxidant activity of 213.25 ± 2.82 μM of Trolox equivalents (TEAC). C. sulcata extracts could prevent hydrogen peroxide (H₂O₂)-induced oxidative stress, reduce expression of the inflammatory markers nuclear Factor kappa B (NFκB) and phosphorylated IκBα (p-IκBα) in A549 human lung carcinoma cells, and inhibit Lipopolysaccharide (LPS)-induced THP-1 monocyte differentiation to an extent of 85%.     

Antioxidant and hepatoprotective activity of ethanolic extract of leaves of Solidago microglossa containing polyphenolic compounds

The antioxidative and hepatoprotective potential of Solidago microglossa D.C, a widely used medicinal plant from Brazil was investigated. The leaf extract showed inhibition against thiobarbituric acid reactive species (TBARS) induced by different prooxidants (10 μM FeSO₄ and 5 μM sodium nitroprusside SNP) in rat liver, brain and phospholipid homogenates from egg yolk. Moreover, the free radical scavenging activities of the extract was evaluated by the scavenging of 2,2-diphenyl-1-picrylhydrazyl (DPPH) (IC_50, 3.8 ± 0.5 μg/ml) and hydroxyl radical on benzoic acid hydroxylation (IC_50, 32.3 ± 1.3 μg/ml) and deoxyribose (IC_50, 39.1 ± 2.4 μg/ml) assays. The ethanolic extract showed significant hepatoprotective activity against paracetamol (250 mg/kg) induced liver damage in mice in a dose dependent manner. The phenolic composition and their quantification by high performance liquid chromatography (HPLC) resulted in the identification of gallic acid and flavonoids: quercetrin (quercetin-3-O-rhamnoside), rutin (quercetin-3-O-rutinoside) and quercetin.     

Phenolic composition,antioxidant and acetylcholinesterase inhibitory activities of Sclerocarya birrea and Harpephyllum caffrum (Anacardiaceae) extracts

Total phenolic content, proanthocyanidins, gallotannins, flavonoids, and antioxidant activities of Sclerocarya birrea and Harpephyllum caffrum methanolic extracts were evaluated using in vitro assays. S. birrea young stem extract contained the highest levels of total phenolic content (14.15 ± 0.03 mg GAE/g), flavonoids (1.21 ± 0.01 mg CE/g) and gallotannins (0.24 ± 0.00 mg GAE/g). H. caffrum stem bark extract had the highest content of proanthocyanidins (1.47%). The EC₅₀ values of the extracts in the DPPH free radical scavenging assay ranged from 4.26 to 6.92 μg/ml, compared to 6.86 μg/ml for ascorbic acid. A dose-dependent linear curve was obtained for all extracts in the ferric-reducing power assay. Dichloromethane and methanol extracts exhibited dose-dependent acetylcholinesterase inhibitory activity. Similarly, all extracts exhibited high antioxidant activity comparable to butylated hydroxytoluene based on the rate of β-carotene bleaching (84.1–93.9%). The two Anacardiaceae species provide a source of natural antioxidants and acetylcholinesterase inhibitors, and may be beneficial to the health of consumers.     

Antioxidant capacities of Gundelia tournefortii L. extracts and inhibition on glutathione-S-transferase activity

Gundelia tournefortii L. is an important food source and a well-known medicinal plant in Eastern Anatolia. Therapeutic effects of medicinal plants are known to be closely related to their antioxidant capacities. Antioxidant activities of G. tournefortii, both for the aerial parts and seeds, were investigated by using both 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and lipid peroxidation inhibition methods. The seeds were found to have higher antioxidant potential than the aerial, with IC₅₀ values of 0.073 mg/mL for DPPH scavenging and 0.146 mg/mL for lipid peroxidation inhibition capacities. In addition, total phenolic contents of the Gundelia tournefortii L. extracts, especially the seed extracts correlates to its high antioxidant activity with 105.1 ± 8.7 μg gallic acid equivalents (GAEs) per mg of seed extract. Plant extracts with high phenolics content are known to have important effects on various enzymes, as well as glutathione-S-transferases, which are important detoxification enzymes in phase II systems with an important role in developing multi-drug resistance to chemotherapy in tumour cells. Consequently, the effects of G. tournefortii extracts on crude cytosolic glutathione-S-transferase was also studied and the seed extracts have shown effective inhibition of cytosolic GST activity, with an IC₅₀ of 97.51 μg/mL.     

Antioxidant and free radical scavenging potential of Achillea santolina extracts

The antioxidative activities of hydroalcoholic extract of Achillea santolina were investigated employing various established in vitro systems including total antioxidant activity in linoleic acid emulsion system, 1,1-diphenyl-2-picrylhydrazyl (DPPH), superoxide and hydroxyl radicals scavenging, reducing power, and inhibitory effect on protein oxidation as well as the inhibition of Fe²⁺/ascorbate induced lipid peroxidation in rat liver homogenate. Total phenolic and flavonoid content of A. santolina extract (ASE) was also determined by a colorimetric method. The results revealed that ASE has notable inhibitory activity on peroxides formation in linoleic acid emulsion system along with concentration-dependent quenching of DPPH and superoxide radicals. Furthermore, the extract showed both nonsite-specific (Fe²⁺ + H₂O₂ + EDTA) and site-specific (Fe²⁺ + H₂O₂) hydroxyl radical scavenging suggesting potent hydroxyl radical scavenging and chelating ability for iron ions in deoxyribose degradation model. A linear correlation between ASE and the reducing power was also observed (r² = 0.9981). ASE prevents thiobarbituric acid reactive substances formation in Fe²⁺/ascorbate induced lipid peroxidation in rat liver tissue in a dose-dependent manner. Moreover, free radical induced protein oxidation was suppressed significantly by the addition of ASE over a range of concentration. These results clearly demonstrated that A. santolina extract possess a marked antioxidant activity.     

Antioxidant and antibacterial activities of Nephelium lappaceum L. extracts

Ether, methanolic and aqueous extracts of lyophilized rambutan (Nephelium lappaceum L.) peels and seeds were evaluated for phenolic contents, antioxidant and antibacterial activities. High amounts of phenolic compounds were found in the peel extracts and the highest content was in the methanolic fraction (542.2 mg/g dry extract). Several potential antioxidant activities, including reducing power, β-carotene bleaching, linoleic peroxidation and free radical scavenging activity, were evaluated. The peel extracts exhibited higher antioxidant activity than the seed extracts in all methods determined (P < 0.05). The methanolic fraction was found to be the most active antioxidant as shown by their 50% DPPH inhibition concentration, 4.94 μg/mL. The results indicated this fraction exhibited greater DPPH radical scavenging activity than BHT and ascorbic acid (0.32 g dry extract/g BHT or ascorbic acid). Antibacterial activity against eight bacterial strains was assessed by disc diffusion and broth macrodilution methods. All peel extracts exhibited antibacterial activity against five pathogenic bacteria. The most sensitive strain, Staphylococcus epidermidis, was inhibited by the methanolic extract (MIC 2.0 mg/mL).     

Targeted phenolic analysis in Hericium erinaceum and its antioxidant activities

The ABTS⁺-radical-cation scavenging activity, DPPH radical scavenging activity, ferric reducing/antioxidant power, and nitrite scavenging activity of the methanol extract from Hericium erinaceum and its subfractions were assessed. Among the methanol extract subfractions tested, the chloroform subfraction exhibited the strongest antioxidant activity in the most experiments, except for the ferric reducing/antioxidant power. The Trolox equivalent antioxidant capacity (TEAC) of the chloroform subfraction was 378.89 μmol/g of sample. This subfraction also scavenged 35.80% of DPPH radicals at 500 μg/mL. The highest ferric reducing/ antioxidant power was found in the n-hexane subfraction (174.82 μmol FeSO₄·7H₂O/g). The chloroform, n-hexane, and n-butanol subfractions had high total phenolic compound content, with ferulic acid equivalents of 35.18, 19.08, and 11.23 mg/g, respectively. Flavonoids were found mostly in the chloroform subfraction, and the 4 phenolic compounds were identified in the same fraction as 4-hydroxybenzoic acid, syringic acid, 4-coumaric acid, and ferulic acid by electrospray ionization (ESI) LC-MS/MS analysis.