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. 2024 Sep 30;29(3):311-320.
doi: 10.3746/pnf.2024.29.3.311.

Antioxidant and Longevity-Related Properties of the Ethyl Acetate Fraction of Cnidium officinale Makino in Caenorhabditis elegans

Hyeon-Ji Kim  1 , Ji-Su Mun  1 , Suk-Heung Oh  1 , Jun-Hyeong Kim  1
Affiliations

Antioxidant and Longevity-Related Properties of the Ethyl Acetate Fraction of Cnidium officinale Makino in Caenorhabditis elegans

Hyeon-Ji Kim et al. Prev Nutr Food Sci. .

Abstract

Reactive oxygen species (ROS) are produced from energy metabolism and may cause diseases or cell death. Antioxidation refers to the suppression of ROS production and is considered beneficial in preventing diseases. This study aimed to examine the antioxidative effects of Cnidium officinale Makino (COM) extracts and fractions using Caenorhabditis elegans as an experimental model. The COM ethanol extract was fractionated according to polarity. The results showed that the ethyl acetate fraction of COM showed powerful radical scavenging activities and increased the activities of superoxide dismutase (SOD) and catalase in C. elegans in a concentration-dependent manner. Moreover, the ethyl acetate fraction reduced the ROS production rate in C. elegans and increased the cell survival rate, suggesting oxidative and thermal stress resistance. In addition, the SOD-3::green fluorescent protein (GFP) expression level in the transformed cells of C. elegans (CF1553) increased, suggesting oxidative stress resistance. Similarly, the HSP-16.2::GFP expression level increased, suggesting thermal stress resistance. In conclusion, the ethyl acetate fraction of COM demonstrated the strongest antioxidative effects, indicating that it may help extend longevity.

Keywords: Caenorhabditis elegans; Cnidium officinale Makino; antioxidant; reactive oxygen species.

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Conflict of interest statement

AUTHOR DISCLOSURE STATEMENT The authors declare no conflict of interest.

Figures

Fig. 1
Fig. 1
(A) DPPH radical scavenging effects of the ethanol extract, and its fractions from the Cnidium officinale Makino. (B) ABTS radical scavenging effects of the ethanol extract, and its fractions from the Cnidium officinale Makino. Different letters (a-d) represent significant differences at P<0.05, as determined by Duncan’s multiple range test. DPPH, 2,2-diphenyl-1-picrylhydrazyl; ABTS, 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt; Vit. C, vitamin C; EtOH, ethanol; n-Hex, n-hexane; MC, methylene chloride; EA, ethyl acetate; n-BuOH, n-butanol.
Fig. 2
Fig. 2
Effects of the ethyl acetate fraction of Cnidium officinale Makino on the antioxidant enzyme activity of wild-type N2 nematode. (A) SOD activity as a percentage of superoxide scavenged per control. (B) Average catalase activity of each group, calculated from the residual H2O2 concentration (determined spectrophotometrically). Differences compared with the control group were considered significant at *P<0.05 and ***P<0.001.
Fig. 3
Fig. 3
Effects of the ethyl acetate fraction of Cnidium officinale Makino on intracellular ROS accumulation of wild-type N2 nematodes. (A) Intracellular ROS accumulation spectrophotometrically quantified at an excitation wavelength of 485 nm and emission wavelength of 535 nm, recorded every 30 min for 120 min. (B) Average percentages of intracellular ROS levels. Differences compared with the control group were considered significant at **P<0.01 and ***P<0.001.
Fig. 4
Fig. 4
Effects of the ethyl acetate fraction of Cnidium officinale Makino on the stress tolerance of wild-type N2 nematodes. For oxidative stress assays, the worms were transferred to a 96-well plate containing 1 mM juglone liquid culture, and then their viability was scored. The statistical difference between the curves was analyzed using the log-rank test.
Fig. 5
Fig. 5
Effects of the ethyl acetate-soluble fraction of Cnidium officinale Makino on the thermal stress tolerance of wild-type N2 nematodes. To assess thermal tolerance, the worms were incubated at 36°C, and then their viability was scored. The statistical difference between the curves was analyzed using the log-rank test. All experiments were performed in triplicates. Differences compared with the control group were considered significant at *P<0.05 and ***P<0.001.
Fig. 6
Fig. 6
Effects of the ethyl acetate fraction of Cnidium officinale Makino on the expression levels of SOD-3 and HSP-16.2 as determined using transgenic nematodes. The mean GFP intensity of CF1553 (A) and CL2070 (B) mutants is presented as the mean±standard error of the mean using 100 organisms per experiment. The GFP intensity was quantified using ImageJ software by determining the average pixel intensity. Images of SOD-3::GFP (C) and HSP-16.2::GFP (D) expression taken from the corresponding mutants grown with or without Cnidium officinale Makino. Data are presented as the mean±standard deviation of three independent experiments (n=3). Differences compared with the control group were considered significant at **P<0.01 and ***P<0.001.
Fig. 7
Fig. 7
Effects of Cnidium officinale Makino on the lifespan of Caenorhabditis elegans. (A) Mortality of each group, which was determined by counting the number of dead organisms daily. (B) Mean lifespan of N2 nematodes, which was calculated from the survival curves. The statistical difference between the curves was analyzed using the log-rank test. The error bars represent the standard error of the mean. Differences compared with the control group were considered significant at *P<0.05, **P<0.01, and ***P<0.001.
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