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长叶榧叶挥发油成分分析及其抗菌作用研究
http://www.100md.com 《第二军医大学学报》 1998年第2期
     作者:陈振德 郑汉臣 田美兰△ 张 虹▲ 朱志明□

    单位:

    关键词:长叶榧;叶;挥发油;抗细菌作用;抗真菌作用

    第二军医大学学报980215 摘要 目的:分析长叶榧叶挥发油成分,并观察其抗菌作用。方法:用GC-MS法进行挥发油成分分析,用其挥发油对4种细菌和3种真菌进行体外抗菌作用研究。结果:长叶榧叶挥发油中分离到64个GC峰,已鉴定了54个成分。该挥发油对金黄色葡萄球菌、大肠杆菌、绿脓杆菌、淋病奈瑟菌、白色念珠菌、热带假丝酵母和酿酒酵母均有明显的抑制作用,其最低抑菌质量浓度分别为1.25,1.25,2.50,0.63,5.00,2.50和5.00 g/L。结论:长叶榧叶挥发油具有抑制细菌和真菌的作用。

    中国图书资料分类法分类号 R 282.710.5

, http://www.100md.com     Constituents analysis and antibacterial and antifungous actions of the essential oil in leaves of Torreya jackii Chun Constituents analysis and antibacterial and antifungous actions of the essential oil in leaves of Torreya jackii Chun

    Chen Zhende,Zheng Hanchen,Tian Meilan, Zhang Hong,Zhu Zhiming (Department of Pharmacognosy, College of Pharmacy, Second Military Medical University,Shanghai,200433)

    Abstract Objective: To analyse the constituents of the essential oil in leaves of Torreya jackii Chun(EOLT) and to observe its antibacterial and antifungous actions. Methods: The constituents were analysed by GC-MS.The inhibitory effects of EOLT on four species of bacteria and three species of fungi were tested in vitro. Results: Sixty-four constituents were firstly isolated by GC and 54 constituents were identified in EOLT.EOLT could effectively inhibit Staphylococci aureus,Escherichia coli, Pseudomonas aeruginosa, Neissia gonorrhoeae, Candida albicans, Candida tropicalis and Saccharomyces cerevisiae, and its MIC were 1.25,1.25,2.50,0.63,5.00,2.50,5.00 g/L, respectively. Conclusion: EOLT has the antibacterial and antifungous actions.
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    Key words Torreya jackii Chun;leaves; essential oil;antibacterial actions; antifungous action

    红豆杉科植物长叶榧(Torreya jackii Chun)是中国特有的裸子植物,产于浙江和福建[1],动物实验表明其茎的粗提物具有降压、舒张血管、抗肿瘤、抗菌等作用[2~6]。自从红豆杉科植物中发现抗癌成分紫杉醇后,国内外学者对其研究越来越关注。为了对榧属植物进行系统研究,我们对长叶榧叶挥发油成分进行GC-MS分析,并对该挥发油进行了抗细菌和抗真菌的体外实验。

    1 材料和方法

    1.1 挥发油的GC-MS分析

    1.1.1 材料 1996年5月采自浙江省仙居县,经鉴定为红豆杉科植物长叶榧的叶。
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    1.1.2 提取挥发油 按《中国药典》(1995年版)附录挥发油测定项进行,水蒸气蒸馏而得,挥发油得率为3.00 ml/kg。

    1.1.3 测定仪器及条件 仪器:Finnigan MAT ITD 800 GC/MS/DS。GC条件:HP-5MS石英毛细管柱(30 m×0.25 mm),进样口温度260℃,柱温150℃(3 min)~250℃(10 min),升温速度4℃/min,载气压力0.25 MPa,进样量0.1 μl。MS条件:电离方式EI,电离电压70 eV,灯丝电流80 μA,倍增器电压2 200 V,离子焰温度220℃,传输线温度230℃。

    1.1.4 数据处理及联机检索 用气质联用仪计算机的NBS谱库自动检索各组分的质谱数据,参考有关标准图谱对机检结果进行核对和补充检索,经色谱峰面积归一化法计算,得出各组分的质量分数。

    1.2 挥发油的体外抗细菌和抗真菌实验
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    1.2.1 受试样品 取长叶榧叶挥发油,加2倍量吐温-80,搅匀,加蒸馏水稀释成2%的溶液。

    1.2.2 受试菌株 金黄色葡萄球菌(Staphylococci aureus)、大肠杆菌(Escherichia coli)、绿脓杆菌(Pseudomonas aeruginosa)、淋病奈瑟菌(Neisseria gonorrhoeae)、白色念珠菌(Candida albicans)、热带假丝酵母(Candida tropicalis)和酿酒酵母(Saccharomyces cerevisiae)。

    1.2.3 受试条件[7] 细菌用水解酪蛋白液体培养基(淋病奈瑟菌用淋病液体培养基), 真菌用改良沙保培养基。细菌在35℃孵箱培养18~24 h,真菌在37℃孵箱培养72 h。

    1.2.4 接种物的准备 取已分纯的上述菌株数个菌落分别移种于上述培养基,孵育4~6 h,校正菌液的浓度,使其相当于0.5麦氏管的混浊度,再稀释1∶200后备用。
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    1.2.5 抗菌实验方法 排列试管10支,除第1管外,每管加入培养基1 ml。先吸取药物原液5.12 ml加入培养基4.88 ml,混匀,第1管和第2管中分别加入1 ml。第2管经混匀后吸出1 ml,加入第3管中,依次类推至第10管后,吸出1 ml弃去。另设对照管仅含培养基1 ml。将已校正浓度的待检菌悬液加入各含药管内,每管0.05 ml,混匀。各试管置孵箱过夜后观察结果(淋病奈瑟菌试管置于有10%CO2环境中培养)。抗菌实验重复3次。

    2 结 果

    2.1 挥发油成分的GC-MS分析 从长叶榧叶的挥发油中分离到64个GC峰,已鉴定出54个成分(见表1),其中含量较高的有γ-荜澄茄烯(γ-cadinene,16.233%)、石竹烯(caryophyllene,13.270%)、罗勒烯(ocimene,8.410%)、反式-β-法呢烯(trans-β-farnesene,6.052%)等。
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    2.2 挥发油的抗细菌和抗真菌作用 长叶榧叶挥

    表 1 长叶榧叶挥发油成分

    Tab 1 Constituents of the essential oil in the leaves of Torreya jackii No.

    Constituent

    Formula

    Mr

    Content(%)

    1

    3-carene

    C10H16
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    136

    0.056

    2

    ocimene

    C10H16

    136

    8.410

    3

    1,5-dimethyl-6-methylene-

    C10H16

    136
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    0.207

    spiro[2,4] heptane

    4

    unidentified

    0.068

    5

    4-methylene-1-(1-methyl-

    C10H16

    136

    0.340

    ethyl)-cyclohexene
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    6

    1-methylene-4-(1-methylethyl)-

    C10H16

    136

    2.481

    cyclohexene

    7

    methyl-(1-methylethyl)-benzene

    C10H14

    134

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    8

    1-methyl-4-(1-methylethenyl)-

    C10H16

    136

    0.479

    cyclohexene

    9

    1,1-dimethyl-2-(3-methyl-1,3-

    C10H16

    136
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    0.558

    butadienyl)-cyclopropane

    10

    3,7-dimethyl-1,3,7-octatriene

    C10H16

    136

    0.578

    11

    3-methyl-6-(1-methylidene)-

    C10H16
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    136

    0.222

    cyclohexene

    12

    2-methyl-6-methylene-1,7-

    C10H14O

    150

    0.156

    octadien-3-one

    13

    unidentified
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    0.071

    14

    6,6-dimethyl-2-methylene-

    C10H16O

    152

    0.090

    bicyclo[3,1,1]heptan-3-ol

    15

    unidentified

    0.198

    16
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    4-methyl-1-(1-methylethyl)-

    C10H18O

    154

    0.167

    3-cyclohexene-1-ol

    17

    1-(methylphenyl)-ethenone

    C 9H10O

    134

    0.099
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    18

    α,α,4-trimethyl-3-cyclo-

    C10H18O

    154

    0.289

    hexene-1-methanol

    19

    6,6-dimethyl-bicyclo[3,1,1]

    C10H14O

    150

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    hept-2-ene-2-carboxaldehyde

    20

    2-dodecenal

    C12H22O

    182

    0.262

    21

    4-(2,6,6-trimethyl-2-cyclo-

    C13H20O

    192
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    1.206

    hexen-1-ylidene)-2-butanone

    22

    4,11,11-trimethyl-8-methylene-

    C15H24

    204

    0.171

    bicyclo[7,2,0]undec-4-ene

    23

    α-cubebene

    C15H24
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    204

    0.179

    No.

    Constituent

    Formula

    Mr

    Content(%)

    24

    copane

    C15H24

    204

    0.237
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    25

    β-cubebene

    C15H24

    204

    0.362

    26

    1,5,α-trimethyl-1,5,α-

    C15H24

    204

    0.882

    cyclododecatriene
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    27

    unidentified

    0.114

    28

    [S-(R*,S*)]-5-(1,5-

    C15H24

    204

    0.223

    dimethyl-4-hexenyl)-2-methyl-

    1,3-cyclohexadiene
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    29

    (1α,4aα,8aα)-1,2,3,4,4a,5,6,8a-

    C15H24

    204

    0.330

    octahydro-7-methyl-4-methylene-

    1-(1-methlethyl)-naphthalene

    30

    caryophyllene

    C15H24
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    204

    13.27

    31

    [1aR-(1aα,7α,7aα,7bα)]-1a,2,C15H24

    204

    1.038

    3,5,6,7,7a,7b-octahydro-1,1,7,7a-tetramethyl-1H-cyclo-

    propa[a]naphthalene

    32

    (1aα,3aα,7bα)-1a,2,3,3a,4,5,7a,C15H24
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    204

    0.915

    7b-octahydro-1,1,3a,7-tetra-

    methyl-1H-cyclopropa-

    [a]naphthalene

    33

    unidentified

    0.772

    34

    trans-β-farnesene

    C15H24
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    204

    6.052

    35

    (1α,4aα,8aβ)-1,2,3,4,4a,5,6,8a-

    C15H24

    204

    3.198

    octahytro-7-methyl-4-methylene-

    1-(1-methylethyl)-naphthalene

    36

    γ-cadinene
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    C15H24

    204

    16.233

    37

    (R)-2,4a,5,6,7,8,-hexahydro-

    C15H24

    204

    4.640

    3,5,5,9-tetramethyl-1H-

    benzocycloheptene

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    4-phenyl-bicyclo[2,2,2]octan-1-ol

    C14H18O

    202

    1.923

    39

    2,6-dimethyl-6-(4-methyl-3-pen-

    C15H24

    204

    1.046

    tenyl)-bicyclo[3,1,1]hept-2-ene
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    40

    (1α,4aβ,8aα)-1,2,3,4,4a,5,6,C15H24

    204

    1.944

    8a-octahydro-7-methyl-4-methy-

    lene-(1-methylethyl)-naphthalene

    41

    (1S-cis)-1,2,3,5,6,8a-hexahy-

    C15H24
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    204

    4.526

    (续表) No.

    Constituent

    Formula

    Mr

    Content(%)

    dro-4,7-dimethyl-1-(1-methyl-

    ethyl)-naphthalene

    42

    (E)-4-hexadecen-6-yne
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    C16H28

    220

    0.633

    43

    3,7,11-trimethyl-1,6,10-

    C15H26O

    222

    1.152

    dodecatrien-3-ol

    44

    (Z)-3-hexen-1-ol benzoate
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    C13H16O2

    204

    0.667

    45

    [4R-(4α,4aα,6β)]-4,4a,5,6,7,C15H22O

    218

    3.404

    8-hexahydro-4,4a-dimethyl-6-

    (1-methylethenyl)-2(3H)-

    naphthalenone
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    46

    [4R-(4α,4aβ,6β)]-4,4a,5,6,7,C15H22O

    218

    2.702

    8-hexahydro-4,4a-dimethyl-6-

    (1-methy-lethenyl)-2(3H)-

    naphthalenone

    47

    (E,E)-3,7,11-trimethyl-2,6,C15H26O
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    222

    0.834

    10-dodecatrien-1-ol

    48

    (Z)-4-hexadecen-6-yne

    C16H28

    220

    1.707

    49

    unidentified

    1.322

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    levomenol

    C15H26O

    222

    2.013

    51

    unidentified

    2.980

    52

    hexadecanal

    C16H32O

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    1.516

    No.

    Constituent

    Formula

    Mr

    Content(%)

    53

    (E,E)-3,7,11-trimethyl-

    C15H24O

    220

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    2,6,10-dodecatrienal

    54

    (Z,E)-3,7,11-trimethyl-2,6,C15H24O

    220

    0.348

    10-dodecatrienal

    55

    unidentified

    0.307

    56
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    benzoic acid phenyl methyl ester

    C14H12O2

    212

    0.448

    57

    3-(dodecyl)dihydro-2,5-

    C16H26O3

    266

    0.288

    furandione
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    58

    hexadecanoic acid

    C16H32O2

    256

    0.973

    59

    1-chloro-7-heptadecene

    C17H33C

    272.5

    0.219

    60
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    [4aS-(4aα,5α,4aβ)-decahydro-

    C20H32

    272

    0.529

    1,1,4a-trimethyl-6-methylene-

    5-(3-methylene-4-pentenyl)-

    naphthalene

    61

    unidentified

    0.237
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    62

    (E)-9-eicosene

    C20H40

    280

    0.210

    63

    unidentified

    0.238

    64

    [1S-(1α,4aα,5α,8αβ)]-decahydro-

    C20H34O2
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    220

    0.214

    5-(5-hydroxy-3-methyl-3-penten-

    yl)-1,4a-dimethyl-6-methylene-

    1-naphthalene methanol

    发油对金黄色葡萄球菌、大肠杆菌、 绿脓杆菌、淋病奈瑟菌、白色念珠菌、热带假丝酵母和酿酒酵母的最低抑菌质量浓度(MIC)分别为1.25,1.25,2.50,0.63,5.00,2.50,5.00 g/L。

    3 讨 论

    长叶榧为红豆杉科榧属植物,该属植物均含挥发油,具芳香气味,不为虫菌所害[8],其木材入土而不腐。我们对长叶榧叶的挥发油成分进行了GC-MS分析,从中分离到64个GC峰,鉴定出54种成分,为该药用植物的系统研究积累了资料。
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    对长叶榧叶挥发油进行的体外抗细菌和抗真菌实验结果表明,其挥发油对医院常见致病菌有较强的抑制作用。该挥发油对不同菌种的抑制强度依次为:淋病奈瑟菌>金黄色葡萄球菌、大肠杆菌>绿脓杆菌、热带假丝酵母>白色念珠菌、酿酒酵母。

    致谢 复旦大学中心实验室胡耀铭老师协助GC-MS测定,特致谢意!

    参 考 文 献

    1 中国科学院中国植物志编缉委员会.中国植物志(第7卷).北京: 科学出版社,1978.464~467

    2 倪 峰,王碧英,郑光中.长叶榧药理作用的研究.福建中医药,1989,20(3):18

    3 陈仁通,潘丽卿,林 温,等.长叶榧化学成分的研究( Ⅰ).中药通报,1982,7(6):31
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    4 陈仁通,潘丽卿,高 凌,等.长叶榧化学成分的研究(Ⅱ).中药通报,1985,10(4):32

    5 陈仁通,方圣鼎.长叶榧化学成分的研究(Ⅲ).长叶榧醇的结构.中草药,1986,17(12):38

    6 中国药材公司.中国中药资源志要.北京:科学出版社,1994.156

    7 徐淑云,卞如濂,陈 修,等.药理实验方法学.北京: 人民卫生出版社,1991. 1385~1389

    8 成俊卿,杨家驹,刘 鹏.中国木材志.北京: 中国林业出版社,1992.81~82

    (1997-08-16收稿,1997-12-20修回), 百拇医药