全 文 ：Study on the Antifungal Activities of Mikania micrantha
Extracts
HAO Cai-qin1，2，FENG Jun-tao1* ，ZHANG Xing1
1． Research and Development Center of Biorational Pesticide，Northwest A ＆ F University /Shaanxi Research Center of Biopesticide Engineering and Technology，
Yangling 712100，China;2． Dearment of Biological and Pharmaceutical Technology，Ningxia Vocational and Technical College，Yinchuan 750002，China
Abstract The stems and leaves of Mikania micrantha were successively extracted with petroleum ether，ethyl acetate and ethanol． In vitro and in vivo test method
was adopted to determine the inhibition activity of three solvent extracts against three plant pathogens． Growth rate method showed that ethyl acetate extract could
significantly inhibit the mycelial growth of Botrytis cirerea，Glomerella cingulata and Fusarium bulbigenum under the given concentration of 0． 09 g /ml in dry sam-
ple，and the inhibition rates were all greater than 90% ． Organization test showed that the curative effect of petroleum ether extract against B． cirerea was 63． 55%
under the given concentration of 0． 18 g /ml in dry sample，and the curative effect of ethanol extract was 71． 47% ． In the potting test against Erysiphe graminis，the
curative effect of petroleum ether extract was 81． 26%，while the curative effect of ethyl acetate extract was 62． 07% ．
Key words Mikania micrantha;Botanical fungicides;Antifungal activity
Received:February 22，2011 Accepted:March 7，2011
Supported by Special Research for National Public Service Sectors (Agricul-
ture) (200903052)．
* Corresponding author． E-mail:hao0418@ 126． com
Mikania micrantha(Mikania，Asteraceae)originated in Cen-
tral and South America is the perennial heliophilic herbaceous
vine． Because of its rapid growth and strong destruction，it is
known as plant killer［1］． In China，M． micrantha occurred in
Hong Kong in 1970s，belonging to invasive species． At present，
M． micrantha has widely distributed in the regions of Shenzhen，
Zhuhai，Guangzhou，Dongguan，Panyu and Neilingding Island，
and causes serious impact on balance of native species［2 －5］． From
the perspective of species evolution， the rapid expansion of
M． micrantha is closely related to its strong competitiveness and
strong resistance，and this is the right theoretical basis for devel-
opment of botanical pesticide． Early preliminary test has shown
that the inhibitory effect in vitro of ethanol extract of M． micrantha
with the concentration of 0． 1 g /ml in dry sample against eight
plant pathogens such as Glomerella cingulata reached 100%，and
its protection effect against Erysiphe graminis was 83． 3% under
the concentration of 0． 05 g /ml in dry sample［6］． To further
confirm the polar parts of antifungal active components in M． mi-
crantha，the author tested in vitro and in vivo antifungal activities
of three organic solvent extracts of M． micrantha，in order to pro-
vide basis for further research，development and utilization of
M． micrantha．
1 Materials and Methods
1． 1 Test materials
1． 1． 1 Preparation of M． micrantha extracts． The stems and
leaves of M． micrantha collected from Shenzhen Fairy Lake Botan-
ical Garden were naturally dried，and crushed to 40 mesh． 200 g
powder was weighed and placed in 3 000 ml clean Erlenmeyer
flask，then extracted with industrial petroleum ether for three
times under room temperature (24 h each time)． The filtrate was
combined together，and carried out vacuum concentration． The
residues were successfully extracted with industrial ethyl acetate
and 95% industrial ethanol for three times，respectively． The re-
spective filtrates were combined together，and a total of three cop-
ies of extracts were obtained． The solvent was fully evaporated in
water bath，and the residue was under cryopreservation for later
use［6］．
1． 1． 2 Test pathogens． Glomerella cingulata，Fusarium bulbige-
num，Erysiphe graminis and Botrytis cirerea used in the test were
provided by Research and Development Center of Biorational Pes-
ticide in Northwest A ＆ F University．
1． 1． 3 Control reagents． The control reagents used in the test in-
cluded 15% Triadimefon WP (produced by Hubei Tianmen pesti-
cide factory) ，40% Pyrimethanil SC(produced by Germany Aige-
fu Company)and emulsifier OP-10 (produced by Changde Xun-
lian Fine Chemicals Co．，Ltd． in Hunan Province)．
1． 2 Experimental methods
1． 2． 1 Growth rate method． In vitro antifungal activity of M． mi-
crantha extracts was determined according to the growth rate meth-
od by WU Wen-jun et al．［7］，crossing method was adopted to
measure colony diameter，and the inhibition of mycelial growth
rate was calculated．
1． 2． 2 Organization determination method． The inhibition effect
of samples against B． cinerea was determined according to tomato
fruit acupuncture method［8］．
(1)Determination method of curative effect． The immature
tomato fruit was rinsed with running water． After the surface water
was dried，the surface of fruit was wiped with alcohol，and the in-
oculation was started as the alcohol evaporated． The fungal cakes
should be inoculated in the symmetric parts in shoulder of tomato．
The inoculation area would be first selected (diameter was the
same as fungal cake，about 5 mm) ，then the sterile inoculation
needle was used to prick small holes． The side of fungal cake with
mycelium should be stick to the small hole regions． Each fruit was
Plant Diseases and Pests 2011，2(2) :61 －63
inoculated with two symmetrical fungal cakes． The reagents were
sprayed an hour later，and the fruits were cultured in training
room under moisture． When the incidence of control was relatively
serious，the results were checked．
(2)Determination method of protective effect． The reagents
were sprayed first，and inoculation was carried out an hour later．
The other operations were the same as the determination method of
curative effect．
1． 2． 3 Potting test． Potting method［9］ was used to determine in
vivo control effect of samples against E． graminis． Samples were
dissolved in acetone，then dripped with 1% OP-10 emulsifier．
The treatments sprayed with 15% Triadimefon WP 1 000 times liq-
uid was set as standard reagent control． The treatment of acetone
added with emulsifier was set as solvent control． Each treatment
contained three pots of wheat seedling，and each pot contained 6
－8 wheat seedlings． Curative effect was sprayed with reagents at
24 h after inoculation，and protective effect was inoculated with
pathogen at 24 h after spraying． All the seedlings were cultured
under the conditions of (18 ±1)℃，light∶ dark =8 h∶ 16 h． Clas-
sification investigation was carried out according to the occurrence
condition of disease after 8 d，the disease index and control effect
of various treatments were calculated．
2 Results and Analysis
2． 1 Antifungal effects in vitro of three solvent extracts of
M． micrantha The inhibition results of M． micrantha extracts
against mycelial growth of three pathogens(G． cingulata，F． bul-
bigenum and B． cirerea)were shown in Table 1．
Table 1 Inhibition rates of three solvent extracts of Mikania micrantha
against mycelial growth of three pathogens %
Samples Botrytiscirerea
Glomerella
cingulata
Fusarium
bulbigenum
Petroleum ether extract 30． 1 b 37． 6 c 26． 0 b
Ethyl acetate extract 94． 4 a 98． 6 a 99． 0 a
Ethanol extract 24． 9 b 54． 8 b 24． 8 b
Note:① The concentration of test plant samples was 0． 09 g /ml in dry sam-
ple;② Each treatment contained three repeats，data in the table were
the average value of three repeats;③ The same lowercase letter in the
same column represented no significant difference at 0． 05 level at vari-
ance analysis test (Duncan new multiple range method)．
As shown in Table 1，three extracts of M． micrantha all
showed certain inhibition effect against three test pathogens under
the given concentrations． The inhibitory effect of ethyl acetate ex-
tract was the best，and its inhibitory effects against G． cingulata，
F． bulbigenum and B． cirerea had reached more than 90%，which
significantly higher than the inhibitory effect of petroleum ether
and ethanol extracts．
2． 2 Inhibition effect of M． micrantha extracts against B． ci-
rerea As shown in Table 2，the curative effect of petroleum ether
extracts against B． cirerea was the best under the given concentra-
tions，reaching 63． 55% ． However，the curative effect of ethyl ac-
etate and ethanol extracts was not obvious． The protective effect of
ethanol extract against B． cirerea was the best of 71． 47%，which
was close to control reagents． The protective effect of ethyl acetate
and petroleum ether extracts was not very good．
Table 2 Inhibition effects of three solvent extracts of Mikania micrantha
against Botrytis cirerea
Samples
Curative effect
Diameter
of lesion
mm
Inhibition
rate
%
Protective effect
Diameter
of lesion
mm
Inhibition
rate
%
Petroleum ether extract 1． 61 63． 55 a 3． 09 30． 85 b
Ethyl acetate extract 3． 33 11． 75 b 2． 91 35． 48 b
Ethanol extract 3． 01 21． 39 b 1． 52 71． 47 a
Pyrimethanil 0． 79 88． 25 a 0． 88 77． 38 a
Control 3． 72 － 4． 29 －
Note:The concentration of test plant samples was 0． 18 g /ml in dry sample，
the concentration of control reagent pyrimethanil was 0． 05 mg /L;②
Each treatment contained three repeats，data in the table were the aver-
age value of three repeats;③ The same lowercase letter in the same col-
umn represented no significant difference at 0． 05 level at variance anal-
ysis test (Duncan new multiple range method)．
2． 3 Potting experiment The potting experiment results of
using M． micrantha extract to control E． graminis was shown in
Table 3． The curative effect of ethyl acetate extracts was the best
under the given concentrations，being 62． 07% ． The protective
effect of petroleum ether extracts was the best，reaching 81． 26% ．
Both of them was close to or higher than control reagents．
Table 3 Control effects of three solvent extracts of Mikania micrantha
against Erysiphe graminis in pot
Samples
Curative effect
Disease
index
Control
effect∥%
Protective effect
Disease
index
Control
effect∥%
Petroleum ether extract 22． 73 47． 36 c 12． 56 81． 26 a
Ethyl acetate extract 16． 37 62． 07 a 32． 58 51． 40 b
Ethanol extract 18． 71 56． 67 b 35． 71 46． 73 c
Triadimefon 14． 95 65． 38 a 38． 11 43． 16 c
Control 43． 18 － 67． 06 －
Note:The concentration of test plant samples was 0． 18 g /ml in dry sample，
the concentration of control reagent triadimefon was 1． 0 mg /L;② Each
treatment contained three repeats，data in the table were the average
value of three repeats;③ The same lowercase letter in the same column
represented no significant difference at 0． 05 level at variance analysis
test (Duncan new multiple range method)．
3 Discussion
Studies have reported that the ethyl acetate extract of aerial
parts of M． micrantha has strong inhibition effect against the seed-
ling growth of test plant，which can hinder the germination process
of seeds，and the inhibition degree of seedling growth is more than
90%［10］． ZHU Mu-jin et al． preliminarily determined the antifun-
gal activity of ethanol extract of M． micrantha［6］，and the results
showed that its crude extracts demonstrated good antifungal activity
in vitro and in vivo． This study further confirmed the antifungal
26 Plant Diseases and Pests 2011
activity of crude extracts of M． micrantha in different polar parts，
and the results showed that different polar parts all had certain an-
tifungal effect，indicating that the stems and leaves of M． micran-
tha contained active substances with diverse structure types and
broad agricultural spectrum． The further exploration on agricultur-
al biological activity of M． micrantha not only is expected to de-
velop a new generation of botanical fungicides，but also can pro-
vide new ways for the comprehensive development and utilization
of M． micrantha．
References
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［6］ZHU MJ，FENG JT，FENG HL，et al． Fungistatic activities of the extrac-
tion from Mikania micrantha［J］． Acta Agriculturae Boreali-occidentalis
Sinica，2003，12(4):53 －55． (in Chinese)．
［7］WU WJ． Plant chemical protection experiment technology introduction［J］．
Xi’an:Shaanxi Science and Technology Publishing House，2003，12(4):
53 －55． (in Chinese)．
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and other plant samples［D］． Yangling:Northwest University of Farming
and Forestry Technology，2001． (in Chinese)．
［9］FANG ZD． FANG ZD． Research method of plant pathology［M］． Beijing:
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［10］SHAO H，PENG SL，ZHANG C，et al． Allelopathic potential of Mikania
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Chinese)．
小花假泽兰提取物抑菌活性研究
郝彩琴1，2，冯俊涛1* ，张 兴1 (1．西北农林科技大学无公害农药研究中心 /陕西无公害农药工程与技术研究中心，陕西杨陵 712100;2．
宁夏职业技术学院生物与制药技术系，宁夏银川 750002)
摘要 以石油醚、乙酸乙酯、乙醇对小花假泽兰(Mikania micrantha)茎叶进行依次提取。采用离体和活体试验法测定 3 种有机溶剂提取物对 3
种植物病原真菌的抑制活性。生长速率法试验结果表明:在干样0． 09 g /ml浓度下，乙酸乙酯提取物能显著抑制番茄灰霉病(Botrytis cirerea)、苹
果炭疽病(Glomerella cingulata)、南瓜枯萎病(Fusarium bulbigenum)3种病原真菌菌丝的生长，抑制率均在 90%以上。组织法试验结果表明:在
干样 0． 18 g /ml浓度下，石油醚提取物对番茄果实灰霉病(Botrytis cirerea)的治疗效果为 63． 55%，乙醇提取物保护效果为 71． 47%。对小麦白粉
病(Erysiphe graminis)的盆栽试验结果表明:石油醚提取物保护作用为 81． 26%，乙酸乙酯提取物治疗作用为 62． 07%。
关键词 小花假泽兰;植物源杀菌剂;抑菌活性
基金项目 国家公益性行业(农业)科研专项(200903052)。
作者简介 郝彩琴(1979 －)，女，陕西榆林人，讲师，硕士，从事天然药物提取分离及活性研究。* 通讯作者，教授，博士，硕士生导师，从事农药毒理学和
农药化学研究。
收稿日期 2011-02-22 修回日期
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2011-03-07
(Continued from page 60)
［8］MA GS． Physiological ecology of Phytophthora parasitica var． nicotianae and
resistance monitoring to metalaxy and Genetic research［D］． Hefei:Anhui
Agricultural University，2002:16． (in Chinese)．
［9］SUN YP，JOHNSON ER． Analysis of joint action of insecticides against house
flies［J］． Journal of Economic Entomology，1960，53(5):887 －892．
［10］CHENG YF，GAO ZM，SHI J，et al． Combined virulence of the mixed
preparations of metalaxyl and propamocab to Phytophthora boehmeriae［J］．
Plant Protection，2007，33(1):41 －43． (in Chinese)．
甲霜灵与其他杀菌剂复配对烟草黑胫病菌联合毒力测定
刘 铭1，2 (1．西昌学院农学系，四川西昌 615013;2．四川农业大学研究生院，四川雅安 625014)
摘要 ［目的］为克服烟草黒胫病对甲霜灵的抗药性，有效控制其危害，研究开发新型甲霜灵高效复配剂。［方法］采用生长速率法测定了 9 种
杀菌剂对烟草黑胫病菌的毒力，并在此基础上选取效果较好的杀菌剂与甲霜灵进行复配，确定最佳复配比。［结果］甲霜灵的毒力最强，EC50为
2． 130 5 μg /ml，其次为多菌灵、代森锰锌和烯酰吗啉，EC50分别为 2． 357 9、2． 639 8、2． 778 8 μg /ml。氰霜唑的效果最差，EC50为 6． 278 8 μg /ml。
效果较好的烯酰吗啉、多菌灵、代森锰锌 3种杀菌剂与甲霜灵的最佳复配比分别为 40∶60、30∶70 和 20∶80，其共毒系数分别为 138． 80、124． 25 和
115． 00。［结论］复配剂具有推广应用的价值，可进一步开展大田试验。
关键词 烟草黑胫病菌;甲霜灵;复配;毒性
基金项目 四川省教育厅重点项目“凉山州烟草黑胫病生理小种研究及四川烟草种质资源的抗性评价”(08ZA032)。
作者简介 刘铭(1978 － )，女，四川广安人，讲师，硕士，从事烟草教学科研工作。
收稿日期 2011-01-30 修回日期 2011-03-02
36HAO Cai-qin et al． Study on the Antifungal Activities of Mikania micrantha Extracts