J Plant Biotechnol 2022; 49(1): 99-105
Published online March 31, 2022
https://doi.org/10.5010/JPB.2022.49.1.099
© The Korean Society of Plant Biotechnology
Correspondence to : e-mail: taekyung7708@chungbuk.ac.kr
This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
This study assessed the antioxidant and anti-inflammatory activities of leaf extracts from grain amaranths (Amaranthus spp.). Among all the extracts, the ethanol extract of Amaranthus cruentus leaves (Ar) exhibited the highest antioxidant activity, including the DPPH free radical scavenging activity and ORAC. In addition, Ar strongly inhibited nitric oxide production by suppressing the MEK/ERK signaling pathway in lipopolysaccharide-stimulated RAW264 murine macrophages. HPLC analysis revealed 13 polyphenolic compounds in the leaf extracts of grain amaranth and indicated that Ar contained more rutin than the other extracts. Taken together, these results show the impact of species diversity on the phytochemical contents and bioactivities of plant extracts and suggest that the nonedible parts, such as leaves, of A. cruentus should be considered for use as crude drugs and dietary health supplements.
Keywords Grain amaranth, antioxidant activity, anti-inflammatory activity, rutin
Phytochemicals, such as sterols, polyphenols, alkaloids, and sulfur-containing compounds obtained from plants, have been extensively studied in terms of their pharmaceutical value or therapeutic potential by the cosmetic and pharmaceutical industries. The biosynthesis of specific phytochemicals is greatly affected by environmental and agronomic factors (Moniodis et al. 2018). Recent studies have revealed that the phytochemical levels of various plants significantly depend on their genetic background (Friedrich et al. 2017; Ju et al. 2021; Li et al. 2012), indicating that genetic factors are the primary factors that affect the production of phytochemicals.
Amaranth is regarded as one of the ancient crops worldwide. The genus
The leaves of grain amaranth (
In the present study, we assessed the antioxidant and anti-inflammatory effects of ethanol extracts obtained from nine different varieties of grain amaranth (six varieties of
Grain amaranth varieties (Table 1) were obtained from the National Agrobiodiversity Center of the Rural Development Administration, Republic of Korea, and were cultivated in the experimental field managed by Chungbuk National University to avoid environmental and agronomic effects on the composition and level of phytochemicals. Leaf materials were harvested 90 days after planting and lyophilized using a freeze dryer (FreeZone Freeze Dry System, Labconco, Kansas City, MO, USA). The ground materials were soaked in ethanol for 24 h and subjected to ultrasonication (15 min × thrice). Filtered ethanol extracts were evaporated and stored at -20°C until further use.
Table 1 Description of the samples used in this study
Sample | IT number | Species |
---|---|---|
Ah1 | IT197078 | |
Ah2 | IT235715 | |
Ah3 | IT238342 | |
Ah4 | IT262653 | |
Ah5 | IT262681 | |
Ah6 | IT288964 | |
Ar | IT199951 | |
Ac1 | IT197098 | |
Ac2 | IT218869 |
The free radical scavenging activity and oxygen radical antioxidant capacity (ORAC) of each extract were determined using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals and fluorescein, respectively, as described by Ju et al. (2021). The DPPH free radical scavenging activity was expressed as the concentration required to reduce half of the DPPH free radicals (RC50), and ORAC was expressed as µM of Trolox equivalents (µM TE).
RAW 264.7 murine macrophage cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM; GIBCO, Grand Island, NY, USA) supplemented with 100 U/mL of penicillin–streptomycin and 10% fetal bovine serum. The cells were plated at a density of 1.5 × 104 cells/mL in 96-well plates and incubated at 37°C for 24 h in a humidified incubator containing 5% CO2. Following this, the cells were treated with each concentration of extract and stimulated with lipopolysaccharide (LPS, 1 μg/mL). After incubation for 24 h, cell viability was determined using MTT solution, as described by Yoo et al. (2021). Moreover, NO production was assessed using the Griess reagent system (Promega Co., Ltd., Madison, USA), according to the manufacturer’s instructions. The results are representative of five independent experiments.
Proteins were extracted using RIPA lysis buffer (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% Triton X-100, 0.1% sodium dodecyl sulfate, 0.5% sodium deoxycholate, 1 mM ethylenediaminetetraacetic acid, and 10 mM NaF) and quantified using the Pierce™ BCA Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA), according to the manufacturer’s protocol. Following this, 20 µg of protein was separated by SDS–PAGE and transferred to a PVDF membrane (Millipore, Billerica, MA, USA). The membrane was then blocked with 5% nonfat dried milk and incubated with antibodies. The signal was detected and visualized with the ECL reagent using an Azure c280 imaging system (Azure Biosystems, Inc., Dublin, CA, USA). The results are representative of three independent experiments.
Total RNA was extracted using TRI Reagent (Invitrogen, Carlsbad, CA, USA), and cDNA was synthesized using the ReverTra Ace® qPCR RT Master Mix with gDNA Remover (TOYOBO, Co., Ltd., Osaka, Japan). qRT-PCR was performed using the SYBR® Green Real-Time PCR Master Mix (TOYOBO, Co., Ltd, Osaka, Japan) on the CFX96™ Real-Time System (Bio-Rad, Hercules, CA, USA). The expression level of each gene was normalized to that of
Table 2 Sequences of the primers used in qRT-PCR analyses
Gene | Primer sequences (5ʹ-3ʹ) | Accession number |
---|---|---|
F-CCTCTGCGATGCTCTTCC | AF233596.1 | |
R-TCACACTTATACTGGTCAAA | ||
F-TCCTACACCACACCAAAC | AF427516.1 | |
R-CTCCAATCTCTGCCTATCC | ||
F-AGCACAGAAGCATGATCCG | AY423855.1 | |
R-CTGATGAGAGGGAGGCCATT | ||
F-CCACTTCACAAGGTCGGAGGCTTA | DQ788722.1 | |
R-GTGCATCATCGCTGTTCATACAATC | ||
F-CCCATCTCCTAAGAGGAGGATG | NM_007393.5 | |
R-AGGGAGACCAAAGCCTTCAT |
F, forward; R, reverse.
HPLC analysis was conducted using an Agilent Technologies 1260 series HPLC unit equipped with a diode array detector (Agilent Technologies, Waldbronn, Germany). Chromatographic separation was performed on a Poroshell 120 EC-c18 column (4.6 × 150 mm, 4 µm) using a mixture of solvent A (water) and solvent B (acetonitrile containing 0.1% formic acid) at a flow rate of 1.0 mL/min, as described by Ju et al. (2021). Polyphenolic compounds in the extracts were identified by comparing their retention times and UV spectral data with those of the standards.
All the experiments were conducted in three independent replicates. The data are expressed as the means ± standard errors (SEs). One-way analysis of variance (ANOVA) and Duncan’s test were performed using IBM SPSS software (version 25) in order to determine the significance of the data (
Reactive oxygen species (ROS) act as intracellular signaling molecules in the regulation of various biological processes; however, excessive ROS formation resulting from an imbalance between cellular production and antioxidative mechanisms causes oxidative damage to cellular components, such as DNA, membranes, and lipids, and various diseases, including vascular disorders, autoimmune diseases, neurodegenerative diseases, and respiratory diseases (Checa and Aran 2020). In the present study, to assess the antioxidant activities of different varieties of grain amaranth, ethanol extracts were prepared and their DPPH free radical scavenging activities were analyzed. As shown in Fig. 1A, Ar showed the highest radical scavenging activity (IC50 = 907.9 ± 60.3 µg/mL) among all the tested varieties of grain amaranth. In addition, compared with the other genotypes, Ar exhibited the highest ORAC (80.9 ± 16.8 μM TE) (Fig. 1B). DPPH free radical scavenging and ORAC assays indicated that genotypic variation within a species did not influence the antioxidant activity.
Various plant extracts have been found to potentially function as natural therapeutic agents against inflammation, which is characterized by the overproduction of inflammatory mediators, such as ROS and pro-inflammatory cytokines (Rodríguez-Yoldi 2021). Therefore, the anti-inflammatory effect of amaranth has previously been studied using the leaf extract of weedy amaranth (
In RAW 264.7 cells, LPS can induce an inflammatory response, at least in part, through its ability to increase the levels of pro-inflammatory mediators, including NO and prostaglandin E2 (Aldridge et al. 2008), which are mainly synthesized by inducible NOS (
The LPS-induced activation of mitogen-activated protein kinase (MAPK) cascades plays critical regulatory roles in the production of pro-inflammatory mediators and cytokines (Manzoor and Koh 2012). Therefore, the MAPK cascade is an important target for anti-inflammatory therapy. As shown in Fig. 4, LPS-induced MEK/ERK activation was inhibited by Ar. This indicates that the Ar-mediated anti-inflammatory effect can be attributed to the inactivation of the MEK/ERK cascade, resulting in the suppression of LPS-induced transcriptions of pro-inflammatory mediators.
Polyphenolic compounds are receiving increasing attention because of their health-promoting properties in many chronic disorders and diseases, including diabetes, cardiovascular diseases, inflammation, cancer, rheumatoid arthritis, and neurodegenerative diseases (Shakoor et al. 2021). In higher plants, genetic factors are considered to explain the intraspecific variability of phytochemicals, including polyphenolic compounds (Asensio et al. 2020). To further analyze the variations in phytochemicals among nine different varieties of grain amaranth, their polyphenolic compounds were determined using HPLC. All the tested samples were found to contain 4-hydroxybenzoic acid, syringic acid, and kaempferol 3-O-β-rutinoside; however, quercetin 3-β-D-glucoside was specific to Ar (Table 3). In
Table 3 Polyphenolic compounds in the leaf extracts from nine different varieties of grain amaranth
Compound number | Compound | µg/g of extract | ||||||||
---|---|---|---|---|---|---|---|---|---|---|
Ah1 | Ah2 | Ah3 | Ah4 | Ah5 | Ah6 | Ar | Ac1 | Ac2 | ||
1 | Gallic acid | NDa | 0.45±0.00a | NDa | 0.25±0.00a | 0.37±0.00a | NDa | 0.19±0.00a | NDa | NDa |
2 | 3,4-Dihydroxybenzoic acid | 116±1.53c | NDa | NDa | NDa | 51.6±12.0b | 198±39.3d | 26.5±6.41ab | NDa | NDa |
3 | 4-Hydroxybenzoic acid | 1.01±0.11b | 1.34±0.09b | 2.52±0.08c | 0.31±0.00a | 0.18±0.01a | 0.08±0.00a | 0.21±0.00a | 0.08±0.00a | 0.27±0.00a |
4 | 2, 4-Dihydroxybenzoic acid | 0.3±0.05a | NDa | 0.42±0.02a | 0.25±0.06a | NDa | NDa | 0.58±0.00a | 0.13±0.00a | NDa |
5 | Caffeic acid | NDa | NDa | NDa | NDa | 0.7±0.00a | NDa | 1.85±0.02b | NDa | NDa |
6 | Syringic acid | 256±49.4bc | 318±15.7c | 467±18.4d | 84.7±34.8a | 70±20.4a | 48.1±0.00a | 47.7±0.00a | 84±14.0a | 150±42.6ab |
7 | p-Coumaric acid | NDa | NDa | NDa | NDa | 0.29±0.09ab | 0.63±0.40b | 0.15±0.02a | 0.36±0.01ab | 0.09±0.00a |
8 | Ferulic acid | NDa | NDa | NDa | NDa | NDa | 1.51±0.50b | NDa | NDa | NDa |
9 | Sinapinic acid | NDa | NDa | NDa | NDa | NDa | 5.14±0.00a | NDa | NDa | NDa |
10 | Rutin | NDa | NDa | NDa | NDa | 2.51±0.08a | NDa | 46.8±1.51c | 5.88±0.57b | 1.44±0.25a |
11 | Quercetin 3-β-D-glucoside | NDa | NDa | NDa | NDa | NDa | NDa | 2.49±0.17b | NDa | NDa |
12 | Benzoic acid | NDa | NDa | NDa | NDa | 0.46±0.00a | 0.18±0.00a | 1.84±0.23b | 0.71±0.00a | NDa |
13 | Kaempferol 3-O-β-rutinoside | 6.67±0.03e | 9.5±0.04f | 24.1±0.01f | 5.33±0.01d | 0.68±0.04bc | 1.01±0.21c | 0.23±0.00a | 0.76±0.02bc | 0.64±0.04b |
Superscript letters indicate significant differences between groups (
The present results clearly indicate the variations in antioxidant and anti-inflammatory activities among nine different varieties of grain amaranth and suggest that species diversity plays an essential role in determining the polyphenol contents and biological activities. In addition, Ar was found to possess the highest antioxidant and anti-inflammatory activities among all the tested extracts, suggesting that Ar is a promising source as a crude drug and dietary health supplement.
J Plant Biotechnol 2022; 49(1): 99-105
Published online March 31, 2022 https://doi.org/10.5010/JPB.2022.49.1.099
Copyright © The Korean Society of Plant Biotechnology.
Hyo Seong Ji ・Gayeon Kim ・Min-A Ahn ・Jong-Wook Chung ・Tae Kyung Hyun
Department of Industrial Plant Science and Technology, College of Agricultural, Life and Environmental Sciences, Chungbuk National University
Correspondence to:e-mail: taekyung7708@chungbuk.ac.kr
This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
This study assessed the antioxidant and anti-inflammatory activities of leaf extracts from grain amaranths (Amaranthus spp.). Among all the extracts, the ethanol extract of Amaranthus cruentus leaves (Ar) exhibited the highest antioxidant activity, including the DPPH free radical scavenging activity and ORAC. In addition, Ar strongly inhibited nitric oxide production by suppressing the MEK/ERK signaling pathway in lipopolysaccharide-stimulated RAW264 murine macrophages. HPLC analysis revealed 13 polyphenolic compounds in the leaf extracts of grain amaranth and indicated that Ar contained more rutin than the other extracts. Taken together, these results show the impact of species diversity on the phytochemical contents and bioactivities of plant extracts and suggest that the nonedible parts, such as leaves, of A. cruentus should be considered for use as crude drugs and dietary health supplements.
Keywords: Grain amaranth, antioxidant activity, anti-inflammatory activity, rutin
Phytochemicals, such as sterols, polyphenols, alkaloids, and sulfur-containing compounds obtained from plants, have been extensively studied in terms of their pharmaceutical value or therapeutic potential by the cosmetic and pharmaceutical industries. The biosynthesis of specific phytochemicals is greatly affected by environmental and agronomic factors (Moniodis et al. 2018). Recent studies have revealed that the phytochemical levels of various plants significantly depend on their genetic background (Friedrich et al. 2017; Ju et al. 2021; Li et al. 2012), indicating that genetic factors are the primary factors that affect the production of phytochemicals.
Amaranth is regarded as one of the ancient crops worldwide. The genus
The leaves of grain amaranth (
In the present study, we assessed the antioxidant and anti-inflammatory effects of ethanol extracts obtained from nine different varieties of grain amaranth (six varieties of
Grain amaranth varieties (Table 1) were obtained from the National Agrobiodiversity Center of the Rural Development Administration, Republic of Korea, and were cultivated in the experimental field managed by Chungbuk National University to avoid environmental and agronomic effects on the composition and level of phytochemicals. Leaf materials were harvested 90 days after planting and lyophilized using a freeze dryer (FreeZone Freeze Dry System, Labconco, Kansas City, MO, USA). The ground materials were soaked in ethanol for 24 h and subjected to ultrasonication (15 min × thrice). Filtered ethanol extracts were evaporated and stored at -20°C until further use.
Table 1 . Description of the samples used in this study.
Sample | IT number | Species |
---|---|---|
Ah1 | IT197078 | |
Ah2 | IT235715 | |
Ah3 | IT238342 | |
Ah4 | IT262653 | |
Ah5 | IT262681 | |
Ah6 | IT288964 | |
Ar | IT199951 | |
Ac1 | IT197098 | |
Ac2 | IT218869 |
The free radical scavenging activity and oxygen radical antioxidant capacity (ORAC) of each extract were determined using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals and fluorescein, respectively, as described by Ju et al. (2021). The DPPH free radical scavenging activity was expressed as the concentration required to reduce half of the DPPH free radicals (RC50), and ORAC was expressed as µM of Trolox equivalents (µM TE).
RAW 264.7 murine macrophage cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM; GIBCO, Grand Island, NY, USA) supplemented with 100 U/mL of penicillin–streptomycin and 10% fetal bovine serum. The cells were plated at a density of 1.5 × 104 cells/mL in 96-well plates and incubated at 37°C for 24 h in a humidified incubator containing 5% CO2. Following this, the cells were treated with each concentration of extract and stimulated with lipopolysaccharide (LPS, 1 μg/mL). After incubation for 24 h, cell viability was determined using MTT solution, as described by Yoo et al. (2021). Moreover, NO production was assessed using the Griess reagent system (Promega Co., Ltd., Madison, USA), according to the manufacturer’s instructions. The results are representative of five independent experiments.
Proteins were extracted using RIPA lysis buffer (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% Triton X-100, 0.1% sodium dodecyl sulfate, 0.5% sodium deoxycholate, 1 mM ethylenediaminetetraacetic acid, and 10 mM NaF) and quantified using the Pierce™ BCA Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA), according to the manufacturer’s protocol. Following this, 20 µg of protein was separated by SDS–PAGE and transferred to a PVDF membrane (Millipore, Billerica, MA, USA). The membrane was then blocked with 5% nonfat dried milk and incubated with antibodies. The signal was detected and visualized with the ECL reagent using an Azure c280 imaging system (Azure Biosystems, Inc., Dublin, CA, USA). The results are representative of three independent experiments.
Total RNA was extracted using TRI Reagent (Invitrogen, Carlsbad, CA, USA), and cDNA was synthesized using the ReverTra Ace® qPCR RT Master Mix with gDNA Remover (TOYOBO, Co., Ltd., Osaka, Japan). qRT-PCR was performed using the SYBR® Green Real-Time PCR Master Mix (TOYOBO, Co., Ltd, Osaka, Japan) on the CFX96™ Real-Time System (Bio-Rad, Hercules, CA, USA). The expression level of each gene was normalized to that of
Table 2 . Sequences of the primers used in qRT-PCR analyses.
Gene | Primer sequences (5ʹ-3ʹ) | Accession number |
---|---|---|
F-CCTCTGCGATGCTCTTCC | AF233596.1 | |
R-TCACACTTATACTGGTCAAA | ||
F-TCCTACACCACACCAAAC | AF427516.1 | |
R-CTCCAATCTCTGCCTATCC | ||
F-AGCACAGAAGCATGATCCG | AY423855.1 | |
R-CTGATGAGAGGGAGGCCATT | ||
F-CCACTTCACAAGGTCGGAGGCTTA | DQ788722.1 | |
R-GTGCATCATCGCTGTTCATACAATC | ||
F-CCCATCTCCTAAGAGGAGGATG | NM_007393.5 | |
R-AGGGAGACCAAAGCCTTCAT |
F, forward; R, reverse..
HPLC analysis was conducted using an Agilent Technologies 1260 series HPLC unit equipped with a diode array detector (Agilent Technologies, Waldbronn, Germany). Chromatographic separation was performed on a Poroshell 120 EC-c18 column (4.6 × 150 mm, 4 µm) using a mixture of solvent A (water) and solvent B (acetonitrile containing 0.1% formic acid) at a flow rate of 1.0 mL/min, as described by Ju et al. (2021). Polyphenolic compounds in the extracts were identified by comparing their retention times and UV spectral data with those of the standards.
All the experiments were conducted in three independent replicates. The data are expressed as the means ± standard errors (SEs). One-way analysis of variance (ANOVA) and Duncan’s test were performed using IBM SPSS software (version 25) in order to determine the significance of the data (
Reactive oxygen species (ROS) act as intracellular signaling molecules in the regulation of various biological processes; however, excessive ROS formation resulting from an imbalance between cellular production and antioxidative mechanisms causes oxidative damage to cellular components, such as DNA, membranes, and lipids, and various diseases, including vascular disorders, autoimmune diseases, neurodegenerative diseases, and respiratory diseases (Checa and Aran 2020). In the present study, to assess the antioxidant activities of different varieties of grain amaranth, ethanol extracts were prepared and their DPPH free radical scavenging activities were analyzed. As shown in Fig. 1A, Ar showed the highest radical scavenging activity (IC50 = 907.9 ± 60.3 µg/mL) among all the tested varieties of grain amaranth. In addition, compared with the other genotypes, Ar exhibited the highest ORAC (80.9 ± 16.8 μM TE) (Fig. 1B). DPPH free radical scavenging and ORAC assays indicated that genotypic variation within a species did not influence the antioxidant activity.
Various plant extracts have been found to potentially function as natural therapeutic agents against inflammation, which is characterized by the overproduction of inflammatory mediators, such as ROS and pro-inflammatory cytokines (Rodríguez-Yoldi 2021). Therefore, the anti-inflammatory effect of amaranth has previously been studied using the leaf extract of weedy amaranth (
In RAW 264.7 cells, LPS can induce an inflammatory response, at least in part, through its ability to increase the levels of pro-inflammatory mediators, including NO and prostaglandin E2 (Aldridge et al. 2008), which are mainly synthesized by inducible NOS (
The LPS-induced activation of mitogen-activated protein kinase (MAPK) cascades plays critical regulatory roles in the production of pro-inflammatory mediators and cytokines (Manzoor and Koh 2012). Therefore, the MAPK cascade is an important target for anti-inflammatory therapy. As shown in Fig. 4, LPS-induced MEK/ERK activation was inhibited by Ar. This indicates that the Ar-mediated anti-inflammatory effect can be attributed to the inactivation of the MEK/ERK cascade, resulting in the suppression of LPS-induced transcriptions of pro-inflammatory mediators.
Polyphenolic compounds are receiving increasing attention because of their health-promoting properties in many chronic disorders and diseases, including diabetes, cardiovascular diseases, inflammation, cancer, rheumatoid arthritis, and neurodegenerative diseases (Shakoor et al. 2021). In higher plants, genetic factors are considered to explain the intraspecific variability of phytochemicals, including polyphenolic compounds (Asensio et al. 2020). To further analyze the variations in phytochemicals among nine different varieties of grain amaranth, their polyphenolic compounds were determined using HPLC. All the tested samples were found to contain 4-hydroxybenzoic acid, syringic acid, and kaempferol 3-O-β-rutinoside; however, quercetin 3-β-D-glucoside was specific to Ar (Table 3). In
Table 3 . Polyphenolic compounds in the leaf extracts from nine different varieties of grain amaranth.
Compound number | Compound | µg/g of extract | ||||||||
---|---|---|---|---|---|---|---|---|---|---|
Ah1 | Ah2 | Ah3 | Ah4 | Ah5 | Ah6 | Ar | Ac1 | Ac2 | ||
1 | Gallic acid | NDa | 0.45±0.00a | NDa | 0.25±0.00a | 0.37±0.00a | NDa | 0.19±0.00a | NDa | NDa |
2 | 3,4-Dihydroxybenzoic acid | 116±1.53c | NDa | NDa | NDa | 51.6±12.0b | 198±39.3d | 26.5±6.41ab | NDa | NDa |
3 | 4-Hydroxybenzoic acid | 1.01±0.11b | 1.34±0.09b | 2.52±0.08c | 0.31±0.00a | 0.18±0.01a | 0.08±0.00a | 0.21±0.00a | 0.08±0.00a | 0.27±0.00a |
4 | 2, 4-Dihydroxybenzoic acid | 0.3±0.05a | NDa | 0.42±0.02a | 0.25±0.06a | NDa | NDa | 0.58±0.00a | 0.13±0.00a | NDa |
5 | Caffeic acid | NDa | NDa | NDa | NDa | 0.7±0.00a | NDa | 1.85±0.02b | NDa | NDa |
6 | Syringic acid | 256±49.4bc | 318±15.7c | 467±18.4d | 84.7±34.8a | 70±20.4a | 48.1±0.00a | 47.7±0.00a | 84±14.0a | 150±42.6ab |
7 | p-Coumaric acid | NDa | NDa | NDa | NDa | 0.29±0.09ab | 0.63±0.40b | 0.15±0.02a | 0.36±0.01ab | 0.09±0.00a |
8 | Ferulic acid | NDa | NDa | NDa | NDa | NDa | 1.51±0.50b | NDa | NDa | NDa |
9 | Sinapinic acid | NDa | NDa | NDa | NDa | NDa | 5.14±0.00a | NDa | NDa | NDa |
10 | Rutin | NDa | NDa | NDa | NDa | 2.51±0.08a | NDa | 46.8±1.51c | 5.88±0.57b | 1.44±0.25a |
11 | Quercetin 3-β-D-glucoside | NDa | NDa | NDa | NDa | NDa | NDa | 2.49±0.17b | NDa | NDa |
12 | Benzoic acid | NDa | NDa | NDa | NDa | 0.46±0.00a | 0.18±0.00a | 1.84±0.23b | 0.71±0.00a | NDa |
13 | Kaempferol 3-O-β-rutinoside | 6.67±0.03e | 9.5±0.04f | 24.1±0.01f | 5.33±0.01d | 0.68±0.04bc | 1.01±0.21c | 0.23±0.00a | 0.76±0.02bc | 0.64±0.04b |
Superscript letters indicate significant differences between groups (
The present results clearly indicate the variations in antioxidant and anti-inflammatory activities among nine different varieties of grain amaranth and suggest that species diversity plays an essential role in determining the polyphenol contents and biological activities. In addition, Ar was found to possess the highest antioxidant and anti-inflammatory activities among all the tested extracts, suggesting that Ar is a promising source as a crude drug and dietary health supplement.
Table 1 . Description of the samples used in this study.
Sample | IT number | Species |
---|---|---|
Ah1 | IT197078 | |
Ah2 | IT235715 | |
Ah3 | IT238342 | |
Ah4 | IT262653 | |
Ah5 | IT262681 | |
Ah6 | IT288964 | |
Ar | IT199951 | |
Ac1 | IT197098 | |
Ac2 | IT218869 |
Table 2 . Sequences of the primers used in qRT-PCR analyses.
Gene | Primer sequences (5ʹ-3ʹ) | Accession number |
---|---|---|
F-CCTCTGCGATGCTCTTCC | AF233596.1 | |
R-TCACACTTATACTGGTCAAA | ||
F-TCCTACACCACACCAAAC | AF427516.1 | |
R-CTCCAATCTCTGCCTATCC | ||
F-AGCACAGAAGCATGATCCG | AY423855.1 | |
R-CTGATGAGAGGGAGGCCATT | ||
F-CCACTTCACAAGGTCGGAGGCTTA | DQ788722.1 | |
R-GTGCATCATCGCTGTTCATACAATC | ||
F-CCCATCTCCTAAGAGGAGGATG | NM_007393.5 | |
R-AGGGAGACCAAAGCCTTCAT |
F, forward; R, reverse..
Table 3 . Polyphenolic compounds in the leaf extracts from nine different varieties of grain amaranth.
Compound number | Compound | µg/g of extract | ||||||||
---|---|---|---|---|---|---|---|---|---|---|
Ah1 | Ah2 | Ah3 | Ah4 | Ah5 | Ah6 | Ar | Ac1 | Ac2 | ||
1 | Gallic acid | NDa | 0.45±0.00a | NDa | 0.25±0.00a | 0.37±0.00a | NDa | 0.19±0.00a | NDa | NDa |
2 | 3,4-Dihydroxybenzoic acid | 116±1.53c | NDa | NDa | NDa | 51.6±12.0b | 198±39.3d | 26.5±6.41ab | NDa | NDa |
3 | 4-Hydroxybenzoic acid | 1.01±0.11b | 1.34±0.09b | 2.52±0.08c | 0.31±0.00a | 0.18±0.01a | 0.08±0.00a | 0.21±0.00a | 0.08±0.00a | 0.27±0.00a |
4 | 2, 4-Dihydroxybenzoic acid | 0.3±0.05a | NDa | 0.42±0.02a | 0.25±0.06a | NDa | NDa | 0.58±0.00a | 0.13±0.00a | NDa |
5 | Caffeic acid | NDa | NDa | NDa | NDa | 0.7±0.00a | NDa | 1.85±0.02b | NDa | NDa |
6 | Syringic acid | 256±49.4bc | 318±15.7c | 467±18.4d | 84.7±34.8a | 70±20.4a | 48.1±0.00a | 47.7±0.00a | 84±14.0a | 150±42.6ab |
7 | p-Coumaric acid | NDa | NDa | NDa | NDa | 0.29±0.09ab | 0.63±0.40b | 0.15±0.02a | 0.36±0.01ab | 0.09±0.00a |
8 | Ferulic acid | NDa | NDa | NDa | NDa | NDa | 1.51±0.50b | NDa | NDa | NDa |
9 | Sinapinic acid | NDa | NDa | NDa | NDa | NDa | 5.14±0.00a | NDa | NDa | NDa |
10 | Rutin | NDa | NDa | NDa | NDa | 2.51±0.08a | NDa | 46.8±1.51c | 5.88±0.57b | 1.44±0.25a |
11 | Quercetin 3-β-D-glucoside | NDa | NDa | NDa | NDa | NDa | NDa | 2.49±0.17b | NDa | NDa |
12 | Benzoic acid | NDa | NDa | NDa | NDa | 0.46±0.00a | 0.18±0.00a | 1.84±0.23b | 0.71±0.00a | NDa |
13 | Kaempferol 3-O-β-rutinoside | 6.67±0.03e | 9.5±0.04f | 24.1±0.01f | 5.33±0.01d | 0.68±0.04bc | 1.01±0.21c | 0.23±0.00a | 0.76±0.02bc | 0.64±0.04b |
Superscript letters indicate significant differences between groups (
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