荷叶提取物在抑制口腔细菌和改善牙龈炎症的有效性分析

doi:10.3969/j.issn.2097-2806.2024.10.011

摘要/Abstract

摘要：

分析了荷叶提取物（LLE）在抑制口腔细菌和改善牙龈炎症的有效性。使用不同质量浓度的LLE处理10种常见口腔细菌24 h，分别检测最小抑菌浓度（MIC）和抑菌圈直径。使用CCK-8法检测LLE对人牙龈上皮细胞活力的影响。将人牙龈上皮细胞分为对照组（Control）、脂多糖（LPS）组和LPS+1~500 μg/mL LLE组，分别用1 μg/mL LPS和不同质量浓度的LLE处理人牙龈上皮细胞24 h。RT-qPCR检测白介素（IL）-1β、IL-6、IL-8、肿瘤坏死因子-α （TNF-α）、细胞间黏附分子-1 （ICAM-1） mRNA水平。Western blotting检测核因子-κB（NF-κB） p65和p38丝裂原活化蛋白激酶（MAPK）的磷酸化水平。结果显示，LLE对10种常见口腔细菌的MIC在300~500 μg/mL之间。与Control组相比，LPS组人牙龈上皮细胞中IL-1β、IL-6、IL-8、TNF-α和ICAM-1 mRNA相对水平均升高，NF-κB p65和p38 MAPK磷酸化水平均升高（P<0.05）。与LPS组相比，LPS+50~500 μg/mL LLE组人牙龈上皮细胞中IL-1β、IL-6、IL-8、TNF-α和ICAM-1 mRNA相对水平均呈LLE剂量依赖性降低，NF-κB p65和p38 MAPK磷酸化水平均呈LLE剂量依赖性降低（P<0.05）。本研究表明，LLE对10种常见口腔细菌和LPS诱导的人牙龈上皮细胞过度炎症具有抑制活性，其可能是口腔护理产品的有效添加成分。

关键词: 荷叶提取物, 口腔细菌, 牙龈炎症, 人牙龈上皮细胞, 核因子-κB

Abstract:

This study aimed to analyze the effectiveness of Lotus leaf extract （LLE） in inhibiting oral bacteria and improving gingival inflammation. LLE of different mass concentrations was used to treat 10 common oral bacteria for 24 hours, and the minimum inhibitory concentration （MIC） and inhibition zone diameter were measured respectively. The CCK-8 method was used to detect the effect of LLE on the viability of human gingival epithelial cells. Human gingival epithelial cells were divided into control group （Control）, lipopolysaccharides （LPS） group and LPS+1-500 μg/mL LLE group. Human gingival epithelial cells were treated with 1 μg/mL LPS and different mass concentrations of LLE respectively for 24 h. RT-qPCR were used to detect the mRNA levels of interleukin （IL）-1β, IL-6, IL-8, tumor necrosis factor-α （TNF-α）, and intercellular adhesion molecule-1 （ICAM-1）. Western blotting were used to detect the phosphorylation levels of nuclear factor-κB （NF-κB） p65 and p38 mitogen-activated protein kinase （MAPK）. The results show that the MICs of LLE against 10 common oral bacteria are 300-500 μg/mL, respectively. Compared with the Control group, the relative mRNA levels of IL-1β, IL-6, IL-8, TNF-α and ICAM-1 in human gingival epithelial cells in the LPS group are all increased, and the phosphorylation levels of NF-κB p65 and p38 MAPK are all increased （P<0.05）. Compared with the LPS group, the relative mRNA levels of IL-1β, IL-6, IL-8, TNF-α and ICAM-1 in human gingival epithelial cells in the LPS+50-500 μg/mL LLE group are all decreased in a dose-dependent manner, and the phosphorylation levels of NF-κB p65 and p38 MAPK are decreased in a dose-dependent manner （P<0.05）. This study shows that LLE has inhibitory activity against 10 common oral bacteria and LPS-induced excessive inflammation of human gingival epithelial cells, and it may be an effective ingredient in oral care products.

Key words: Lotus leaf extract, oral bacteria, gingival inflammation, human gingival epithelial cells, nuclear factor-κB

中图分类号:

TQ658

刘娟丽, 徐慧萍, 刘艳芳, 董振花, 王美艳. 荷叶提取物在抑制口腔细菌和改善牙龈炎症的有效性分析[J]. 日用化学工业（中英文）, 2024, 54(10): 1235-1242.

Juanli Liu, Huiping Xu, Yanfang Liu, Zhenhua Dong, Meiyan Wang. Analysis of the effectiveness of Lotus leaf extract in inhibiting oral bacteria and improving gingival inflammation[J]. China Surfactant Detergent & Cosmetics, 2024, 54(10): 1235-1242.

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Gene	Primer sequence
IL-1β	F: 5′-TACAAGGAGAAGAAAGTAATGACAA-3′
IL-1β	R: 5′-AGCTTGTTATTGATTTCTATCTTGT-3′
IL-6	F: 5′-AGCCACTCACCTCTTCAGAACG-3′
IL-6	R: 5′-GGTTCAGGTTGTTTTCTGCCAG-3′
IL-8	F: 5′-CTTGGCAGCCTTCCTGATTTC-3′
IL-8	R: 5′-TTCTGTGTTGGCGCAGTGTG-3′
TNF-α	F: 5′-CTCCTCACCCACACCATCAGCCGCA-3′
TNF-α	R: 5′-ATAGATGGGCTCATACCAGGGCTTG-3′
ICAM-1	F: 5′-AGAACCTTACCCTACGCTGC-3′
ICAM-1	R: 5′-CAGTATTACTGCACACGTCAGC-3′
GAPDH	F: 5′-GGAGCGAGATCCCTCCAAAAT-3′
GAPDH	R: 5′-GGCTGTTGTCATACTTCTCATGG-3′

ρ（LLE）/（μg/mL）	抑菌圈直径/mm
ρ（LLE）/（μg/mL）	F. nucleatum	E. coli	P. aeruginosa	E. faecalis	S. mutans	E. faecium	S. oralis	P. gingivalis	A. oris	C. albicans
0	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00
1	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00
10	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00	0.00±0.00
50	6.14±0.25	5.91±0.29	0.00±0.00	0.00±0.00	5.94±0.24	0.00±0.00	6.88±0.38	0.00±0.00	5.92±0.44	0.00±0.00
100	8.27±0.35	8.34±0.34	6.98±0.26	5.71±0.28	7.74±0.32	6.41±0.23	9.00±0.23	5.01±0.26	8.17±0.38	5.12±0.32
200	10.96±0.47	10.82±0.52	8.95±0.44	8.50±0.31	9.83±0.39	8.63±0.48	11.66±0.49	7.11±0.31	10.34±0.42	6.99±0.31
300	14.88±0.83	14.46±0.49	10.86±0.56	11.72±0.61	13.17±1.06	11.83±0.36	15.94±0.48	9.00±0.44	13.08±0.89	10.00±0.36
400	17.82±0.46	16.48±1.10	15.99±1.16	15.20±0.75	17.35±0.80	16.41±0.89	18.96±0.83	11.83±0.38	17.03±0.48	13.61±0.70
500	19.75±1.05	20.06±1.34	19.12±1.09	18.15±0.80	22.36±1.72	19.36±0.71	21.52±1.03	15.80±0.85	21.00±0.82	17.40±0.52
F	1 351.092	840.555	960.773	1 612.197	692.052	1 833.000	1 553.356	1 561.894	1 462.331	2 170.473
P	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001	<0.001