J Nutr Health.  2014 Aug;47(4):221-228. 10.4163/jnh.2014.47.4.221.

Fatty acid analysis and regulatory effects of citron (Citrus junos Sieb. ex TANAKA) seed oil on nitric oxide production, lipid accumulation, and leptin secretion

• Kim TW
• Kim KK
• Kang YH
• Kim DJ
• Choe M

• KMID: 2327155
• DOI: http://doi.org/10.4163/jnh.2014.47.4.221

Abstract

PURPOSE
Citron seed oil (CSO) has been reported to have high antioxidant activity. However, the composition and other biologically activities of CSO have not been reported. In this study, we confirmed the fatty acid composition of CSO, which may be beneficial to vascular disease and obesity.
METHODS
We investigated the oil composition of CSO using gas chromatography coupled with mass spectrometry (GC-MS) analysis, and cytotoxicity was confirmed by Cell Counting Kit-8 (CCK-8) assay. Nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs) was measured using Griess reagent, and lipid accumulation and leptin secretion in 3T3-L1 cells were measured by Oil-Red O staining and commercial ELISA kit, respectively.
RESULTS
GC-MS analysis indicated that CSO contains several components, including linoleic acid, oleic acid, palmitic acid, stearic acid, linolenic acid, palmitoleic acid, and arachidic acid. In physiological activity analysis, CSO did not induce cytotoxic effects in HUVECs and 3T3-L1 cells. Further, CSO significantly induced nitric oxide and leptin secretion as well as inhibited lipid accumulation.
CONCLUSION
CSO increased NO release, inhibited lipid accumulation, and induced leptin secretion, suggesting it may be useful for the management of vessels and weight gain. Although further studies are required to investigate the safety and mechanism of action of CSO, our results show that the composition and physiological activity of CSO are sufficient for its use as functional edible oil.

Keyword

citron seed oil; fatty acid composition; nitric oxide; leptin; lipid accumulation

MeSH Terms

3T3-L1 Cells
alpha-Linolenic Acid
Cell Count
Chromatography, Gas
Enzyme-Linked Immunosorbent Assay
Human Umbilical Vein Endothelial Cells
Leptin*
Linoleic Acid
Mass Spectrometry
Nitric Oxide*
Obesity
Leptin
Linoleic Acid
Nitric Oxide
Oleic Acid
Palmitic Acid
alpha-Linolenic Acid

Figure

• 

  Fig. 1 Concentration-dependent effects of citron seed oil (CSO) on human umbilical vein endothelial cells (HUVECs) and 3T3-L1 cell growth. A: Cytotoxicity of CSO on HUVECs. B: Cytotoxicity of CSO on 3T3-L1. Cell viability was analyzed using the CCK-8 assay kit. Each bar is the Mean ± S.D. from three independent experiment.

• 

  Fig. 2 GC chromatograms and MS fragments of individual fatty acids extracted from citron seed oil (CSO). 1: undecanoic acid, 2: palmitic acid, 3: palmitoleic acid, 4: stearic acid, 5: oleic acid, 6: Unknown, 7: linoleic acid, 8: arachidic acid, 9: linolenic acid.

• 

  Fig. 3 Effect of citron seed oil (CSO) on nitric oxide (NO) secretion in HUVACs. Reductions of NO by TNF-α were recovered by CSO (0.1 mg/ml) treatment. Results are presented as Mean ± S.D. of three independent experiments. #: p < 0.05, *: p < 0.01 as compared to the TNF-α (100 ng/ml), different letters on the bars (a and b) indicate significant differences (p < 0.05) by Duncan's multiple range test.

• 

  Fig. 4 Citron seed oil (CSO) inhibits lipid accumulation in 3T3-L1 cells. Adipocytes were incubated with two concentrations (0.05 and 0.1 mg/ml) of CSO and lipid accumulation was determined by Oil Red O staining. Results are presented as Mean ± S.D. of three independent experiments. **: p < 0.01 as compared to the control by unpaired two-tailed t-tests.

• 

  Fig. 5 Citron seed oil (CSO) induces leptin secretion in 3T3-L1 cells. Adipocytes were incubated with two concentrations (0.05 and 0.1 mg/ml) of CSO. Leptin secretion was determined by enzyme-linked immunosorbent assay (ELISA). Results are presented as Mean ± S.D. of three independent experiments. ***: p < 0.001 as compared to the control by unpaired two-tailed t-tests.

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