老鹳草乙醇提取物中黄酮类化合物和挥发性组分及其抗菌活性

李宇; 刘海华; 刘翠君; 由金文; 赵仁君; 石爱华; 何美军

doi:10.19303/j.issn.1008-0384.2020.12.014

老鹳草乙醇提取物中黄酮类化合物和挥发性组分及其抗菌活性

doi: 10.19303/j.issn.1008-0384.2020.12.014

1.
湖北省农业科学院中药材研究所，湖北　恩施　445000
2.
恩施职业技术学院，湖北　恩施　445000

基金项目: 湖北省农业科学院中药材研究所2019年青年科学基金项目（2019ZYCJJ05）

详细信息

作者简介:
李宇（1990−），男，研究方向：中药材天然产物研究（E-mail：977956996@qq.com）

通讯作者:
何美军（1974-），男，副研究员，研究方向：中药材天然产物研究（E-mail：840940513@qq.com）

中图分类号: Q 939.13; R 914.4
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- 被引次数: 0
出版历程
- 收稿日期: 2020-03-06
- 修回日期: 2020-08-28
- 刊出日期: 2020-12-31

Volatiles, Flavonoids, and Antibacterial Activity of Ethanol-extract from Geranium wilfordii

1.
institute of Chinese Herbal Medicines, Hubei Academy of Agricultural Sciences, Enshi, Hubei　445000, China
2.
Enshi Polytechnic, Enshi, Hubei　445000, China

摘要

摘要: 目的研究老鹳草乙醇提取物对12株临床致病菌的抑菌活性，为开发新型植物来源的抗生素提供重要的参考。方法以老鹳草全草乙醇提取物为研究对象，运用气相色谱-质谱联用仪、高效液相色谱仪研究其化学成分；运用滤纸片法测试12株临床致病菌的抑菌活性。结果从老鹳草乙醇提取物中鉴定出：18个挥发性组分包括脂肪烃、脂肪酸和生物碱，3个黄酮化合物杨梅苷、槲皮素-3-O-β-D-葡萄糖、花旗松素。老鹳草乙醇提取物对粪链球菌（MIC 1.0 mg·mL⁻¹）、耐甲氧西林金黄色葡萄球菌（MIC 1.0 mg·mL⁻¹）、大肠杆菌（MIC 1.0 mg·mL⁻¹）及鲍曼不动杆菌（MIC 2.0 mg·mL⁻¹）有高敏抑制活性。结论恩施地区老鹳草乙醇提取物主要分挥发性组分为脂肪烃、脂肪酸、生物碱；含有杨梅苷、槲皮素-3-O-β-D-葡萄糖、花旗松素3个黄酮类化合物；对常见临床致病菌具有优良抗菌活性和较广的抗菌谱覆盖面。
- 老鹳草 /
- 乙醇提取物 /
- 挥发组分 /
- 临床致病菌 /
- 黄酮
Abstract: Objective Antibacterial activity and mechanism of the ethanol-extract of Geranium wilfordii Maxim on clinical pathogens were studied for development of a natural antibiotic. Method The chemical compositions of the extract were analyzed by GC-MS and HPC. The filter paper method was used to test the antimicrobial activity of the extract against 12 clinical samples of the resistant bacteria. Result GC-MS detected 18 volatile substances, including aliphatic hydrocarbons, fatty acids, and alkaloids (1.15%). The 3 flavonoid compounds identified were myricitrin, quercetin-3-O-β-D-glucose, and taxifolin. High antibacterial activities against Enterococcus faecalis ATCC 29212 (MIC 1.0 mg·mL⁻¹), methicillin-resistant Staphylococcus aureus (MIC 1.0 mg·mL⁻¹), Escherichia coli ATCC 25922 (MIC 1.0 mg·mL⁻¹), and Acinetobacter baumannii ATCC 29212 (MIC 2.0 mg·mL⁻¹) of the extract were observed. Conclusion In the G. wilfordii ethanol-extract, aliphatic hydrocarbons, fatty acids, and alkaloids were the major volatiles, and myricitrin, quercetin-3-O-β-D-glucose, and taxifolin the 3 substantial flavonoid compounds found. The extract demonstrated a significant antibacterial activity with wide spectrum against the commonly known clinical pathogens.
- Geranium wilfordii Maxim /
- ethanol-extract /
- volatile substances /
- clinical resistant bacteria /
- flavonoids

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图 1 老鹳草乙醇提取物与黄酮标准品保留时间对比

Figure 1. Retention times of G. wilfordii ethanol-extract and flavonoid standards

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图 2 老鹳草乙醇提取物与黄酮标准品光谱对比

Figure 2. Spectrograms on G. wilfordii ethanol-extract and flavonoid standards

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表 1 老鹳草乙醇提取物组分挥发性成分及其含量测定结果

Table 1. Volatiles in G. wilfordii ethanol-extract

化合物 Compounds	保留时间 Retention time/min	化学式 Chemical formula	分子质量 Molecular mass	相对含量 Relative amount/%	匹配度 Matching rate/%
二十二烷酸二十二烷基酯(1) docosyl docosanoate	5.157	C₄₂H₈₄O₂	621.13	0.42	93.5
6-甲基-正十八烷(2) 6-methyloctadecane	5.934	C₁₉H₄₀	268.53	0.56	94.5
2，6，10-三甲基-正四十烷(3) 2,6,10-trimethyltetradecane	6.757	C₁₇H₃₆	240.48	0. 62	95.6
11-(3-戊氧基-2-乙基)十一酸甲酯(4) methyl11-(3-pentyloxiran-2-yl)undecanoate	8.610	C₁₉H₃₆O₃	312.49	0.96	98.2
十八烷(5) Octadecane	9.004	C₁₈H₃₈	254.50	0.97	93.8
二十八烷(6) Octacosane	9.075	C₂₈H₅₈	394.45	1.85	97.2
聚十八烷基乙烯基醚(7) 1-(Vinyloxy)octadecane	9.310	C₂₀H₄₀O	296.54	1.50	93.7
2-溴十八醛(8) 2-bromooctadecanal	9.492	C₁₈H₃₅BrO	347.38	1.56	92.8
顺-7-甲基-十四碳8-烯-1-乙酸酯(9) (Z)-7-methyltetradec-8-en-1-yl acetate	9.616	C₁₇H₃₂O₂	268.44	0.92	91.8
10 -辛基-十九烷(10) 10-octylnonadecane	9.704	C₂₇H₅₆	380.75	0.32	99.0
棕榈酸甲酯(11) methyl palmitate	9.992	C₁₇H₃₄O₂	270.46	7.91	95.2
(5β)孕烷-3,20-β-二醇,14 α,18α-[4-甲基-3-氧基-(1-oxa-4-氮杂丁烷-1,4-二基)]- 二乙酸酯(12) (5.β.)Pregnane-3,20.β.-diol, 14α.,18.α-[4-methyl-3-oxo-(1-oxa-4-azabutane-1,4- diyl)]-, diacetate	10.210	C₂₈H₄₃NO₆	489.65	1.15	98.9
2-甲基-十六烷-1-醇(13) 2-methylhexadecan-1-ol	10.439	C₁₇H₃₆O	256.47	0.52	90.3
棕榈酸(14) palmitic acid	10.710	C₁₆H₃₂O₂	256.43	0.45	93.6
甲基(E)-十八烷基-9-烯酸酯(15) Methyl (E)-octadec-9-enoate	11.469	C₁₉H₃₆O₂	296.50	4.93	98.2
12-甲基十四烷酸甲酯(16) Methyl 12-methyltetradecanoate	11.663	C₁₆H₃₂O₂	256.43	0.53	99.5
油酸(17) oleic acid	13.333	C₁₈H₃₄O₂	282.47	0.97	90.1
邻苯二甲酸二辛酯(18) dioctyl phthalate	21.603	C₂₄H₃₈O₄	390.56	0.43	92.3

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表 2 标准曲线、重复性及检测限（means±SD，n=6）

Table 2. Regression equation, correlation coefficient, and detection limit on measurements (means±SD, n=6)

标准品 Standard substance	标准曲线 Standard curves	R²	检出限 Detection limit	定量限 Limit of quantitation	回收率 Recovery/%	S/%
花旗松 Taxifolin	y=4.16×A−2.77	0.9996	0.136	0.725	98.56	1.50
槲皮素-3-O-β-D-葡萄糖 Quercetin-3-O-β-D-glucose	y=3.13×A−6.88	0.9994	0.128	0.915	101.05	1.23
杨梅苷 Myricetrin	y=3.44×A−4.08	0.9991	0.153	0.817	99.26	0.55
注：A为峰面积，单位mAU; y为浓度，单位μg·mL⁻¹；S为相对标准偏差。 Note: A was the peak area, unit was mAU; y was the concentration, unit was μg·mL^-1: S is the relative standard deviation.

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表 3 老鹳草乙醇提取物各黄酮化合物含量表（means±SD，n=6）

Table 3. Flavone contents in　G. wilfordii ethanol-extract

黄酮组分 Flavonoid components	n^b/（μg·mL⁻¹）	回收率 Recovery/%	S/%	C^a/（mg·g⁻¹）
花旗松素 Taxifolin	7.44±0.48	101.70	1.12	3.72
槲皮素-3-O-β-D-葡萄糖 Quercetin-3-O-β-D-glucose	11.12±0.73	101.98	0.85	5.56
杨梅苷 Myricetrin	5.12±0.33	99.24	1.05	2.56
注：a：以各黄酮组分质量与老鹳草样品的干重质量比，b：HPLC检测浓度 Note: a: individual flavone content in G. wilfordii (dry weight); b: concentration detected by HPLC.

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表 4 老鹳草乙醇提取物的抗菌活性（means±SD，n=3）

Table 4. Antibacterial activity of G. wilfordii ethanol-extract

细菌　　　　　　　　　 Bacteria	抑菌圈 Inhibition zone/mm	阳性对照 Positive control/mm	MIC/mg·mL⁻¹
克雷伯氏肺炎菌 K. pneumonia ATCC 13883	11.18±0.30	14.07±0.09	4.0
金黄色葡萄球菌 S. aureus ATCC 29213	14.95±0.26	16.26±0.28	4.0
粪肠球菌 E. faecalis ATCC 29212	19.35±1.24	6.50±0.26	1.0
枯草芽孢杆菌 B. subtilis	12.66±0.58	14.36±0.16	4.0
藤黄微球菌 M. luteus	8.77±0.16	17.35±0.87	16.0
苏云金芽孢杆菌 B. thuringiensis	11.06±0.28	21.41±0.27	4.0
耐甲氧西林金黄色葡萄球菌 methicillin-resistant S. aureus	16.77±0.95	—	1.0
铜绿假单胞菌 P. aeruginosa	11.23±0.97	14.23±0.44	4.0
抗甲氧西林表皮葡萄球菌 methicillin-resistant S. epidermidis	14.32±0.45	20.86±1.76	4.0
大肠埃希氏菌 E. coli ATCC 25922	20.30±0.30	27.74±0.32	1.0
鲍曼不动杆菌 A. baumannii ATCC 19606	18.24±0.50	20.44±0.32	2.0
溶藻弧菌 V. alginolyticus XSBZ14	14.91±0.34	12.08±0.19	8.0
注: “—”表示无抑菌圈；阳性对照为氨苄青霉素(100 mg·mL⁻¹) Note: “—” indicates no inhibition zone, ampicillin (100 mg·mL⁻¹) was the positive control

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