全 文 ：天然产物研究与开发 Nat Prod Res Dev 2011，23:1064-1068
文章编号:1001-6880(2011)06-1064-05
Received January 6，2010;Accepted June 12，2010
* Corresponding author Tel:86-22-87401895;E-mail:pharmgao@ tju．
edu． cn
三七叶、人参叶和西洋参叶皂苷类成分的研究
张兰兰1，2，高文远1* ，周水平2，朱永宏2，宋兆辉2，黄芝娟2
1天津大学药学院，天津 300072;2 天津天士力集团研究院中药所，天津 300410
摘 要:三七叶、人参叶和西洋参叶其皂苷类成分相近，但专属性成分各异，皂苷类成分的分布比例也各不相
同。本文建立了 HPLC-UV法测定上述皂苷成分的方法，经过方法学考察，各种皂苷成分精密度好、加样回收率
高，方法可靠。11 种皂苷成分总含量顺序为:西洋参叶 ＞人参叶 ＞三七叶;二醇组皂苷成分含量:西洋参叶 ＞三
七叶 ＞人参叶;三醇组皂苷成分含量:人参叶 ＞西洋参叶 ＞三七叶。西洋参叶中二醇组皂苷和人参叶中三醇组
皂苷含量明显高于其他。西洋参叶中人参皂苷 Rb3 和 Rd的含量之和占 11 种皂苷成分的 60%以上。鉴于其中
人参皂苷的高含量，三七叶、人参叶和西洋参叶应该作为皂苷来源得到充分利用;不同的皂苷成分有不同的药
理活性，应基于它们的皂苷组成和比例选择性进行研究和开发。
关键词:三七;人参;西洋参;叶;人参皂苷
中图分类号:R284. 1 文献标识码:A
Ginsenosides in Leaves of Panax notoginseng，P． ginseng and P． quinquefolius
ZHANG Lan-lan1，2，GAO Wen-yuan1* ，ZHOU Shui-ping2，ZHU Yong-hong2，SONG Zhao-hui2，HUANG Zhi-juan2
1College of Pharmaceutical ＆ Biotechnology，Tianjin University，Tianjin 300072，China;
2Modern TCM R＆D Institute，Tianjin Tasly Group Co． Ltd．，Tianjin 300410，China
Abstract:Leaves of Panax notoginseng，P． ginseng and P． quinquefolius have similar saponin components but with differ-
ent exclusive components． In this work，the contents of saponins in the leaves of Panax notoginseng，P． ginseng and P．
quinquefolius were determined with reversed-phase high-performance liquid chromatography． The chromatographic sepa-
ration was achieved by using a Kinetex-C18 column (100 × 4． 6 mm，2． 6 μm，phenomenex，at 30 ℃)． Eleven compounds
were separated with a gradient elution and detected at 203 nm． The results showed that the order of total content of sapo-
nins，protopanaxadiol saponins and protopanaxatriol saponins in three kinds of leaves was P． quinquefolius ＞ P． ginseng
＞ P． notoginseng，P． quinquefolius ＞ P． notoginseng ＞ P． ginseng，and P． ginseng ＞ P． quinquefolius ＞ P． notogin-
seng，respectively． The content of protopanaxadiol saponins in the leaves of P． quinquefolius and the content of proto-
panaxatriol saponins in the leaves of P． ginseng obviously were higher than those in others． And the total content of Gin-
senosides Rb3 and Rd was accounted for over 60% of the total 11 ginsenosides in P． quinquefolius leaves． Conclusively，
the leaves of Panax notoginseng，P． ginseng and P． quinquefolius should be well used as a wonderful source for saponins
due to their high contents． In addition，all the three kinds of saponins should be exploited on the basis of their contents
and compositions．
Key words:Panax notoginseng;P． ginseng;P． quinquefolius;leaves;ginsenoside
Instruction
Panax ginseng，P． notoginseng and P． quinquefolius are
valuable agriculture commodity used in many tradition-
al medicinal therapies． In the past years，much attention
has been paid to analyze the active saponins in the root
of P． ginseng，P． notoginseng and P． quinquefolius． For
example，P． notoginseng has been used as creditable
drug for the treatment of haemoptysis，haemostatic and
haematoma in China for more than 400 years． Modern
pharmacological studies have shown that P． notoginseng
has anticarcinogenic and hepatoprotective activities，as
well as protective effects on cardiovascular and cerebro-
vascular systems［1］． P． ginseng has been a tonic，seda-
tive，anti-fatigue drug，and also has anti-diabetic and
anti-tumor activities［2］． P． quinquefolius，which is a
popular herb for patients suffering from nervous system
and cardiovascular，is also a dietary supplement． Its ex-
tract has been included as a component in many sup-
plement formulations ［3］． Drammarane triterpene sapo-
nins are considered as the major bioactive constitutes in
Panax genus． According to the different aglycone，these
neutral saponins were classified into two groups:proto-
panaxatriol saponins (PTS，such as ginsenoside Re，
Rg1，Rf，Rh1，and notoginsenosideR1)and protopanax-
adiol saponins (PDS，such as ginsenoside Rc，Rd，Rb1，
Rb2，Rb3)． However，the biological activities of ginsen-
osides are related with their structures［4］． Therefore，it
is valuable to separate different types of saponins for
further pharmacological research． Pharmacological stud-
ies suggested that ginsenoside Rb3 may contribute to
improve learning and memory ［5］，and ginsenoside Rg3
and Rh2 showed anticancer effects
［6］． However，gin-
senoside Rg1，Rb1，Re showed strong protective effects
on cardiovascular and cerebrovascular systems ［7］．
Nowadays，several studies have demonstrated that the
leaves of P． ginseng，P． notoginseng and P． quinquefoli-
us also contain ginsenosides． Liu et al． reported the sep-
aration of ginsenoside Rb3 and Rc from extracts of leav-
es of P． notoginseng［8］． Shi et al． compared the ginsen-
oside content of the leaves with the root of P． ginseng
and investigated their changes with age［9］． Qu et al．
studied on ginsenosides in different parts and ages of
P． quinquefolius． In recent years，their leaves extract
have been frequently used as food additives and raw
materials of healthy food ［10］． Therefore，more systemat-
ic studies and comparisons on these Panax leaves be-
come necessary，which will provide more data and in-
formation in the field of food and medicine．
In the recent years，the macroporous resins have been
extensively applied to separate and purify the bioactive
components of herbs［11，12］． In this paper，we used the
macroporous resins D101 to purify and prepare the gin-
senoside extract． As for the analytical method，HPLC-
UV，which uses acetonitrile and water as elution sol-
vents and measured the absorbance at the wavelength of
203 nm，is a general method to determine the content of
ginsensosides，because of its good linear and precision．
Experiment
Materials and instruments
The standards of ginsenoside Rg1，Re，Rb1，Rb2，Rb3，
Rc，Rd，Rh2，Rg3，Rf and notoginsenoside R1 were pur-
chased from Chinese Medical and Biological Products
Institute (Beijing China)． Methanol and acetonitrile
were products of Merck． The deionized water was puri-
fied by Milli-Q purification system (Millipore Bedford，
USA)． Macroreticular resins D101 were products of
Tianjin Nankai Haiguang Chemicals Co．，Ltd． All the
other reagents were of analytical grade． Agillent 1100
liquid chromatography (Agilent Technologies，USA)e-
quipped with quaternary gradient pump was used to
separate the ginsenosides，UV detector coupled with a
Chromatograph Workstation was used for detecting the
ginsenosides． The fresh leaves of P． ginseng，P． notog-
inseng，P． quinquefolius materials were harvested in
fall，and P． notoginseng leaves obtained from Wenshan
in Yunnan Province，P． ginseng leaves and P． quinque-
folius leaves were obtained from Jingyu in Jilin Prov-
ince． The leaves were dried at 38 ℃ in ventilated dri-
er． The dried leaves were crushed into small pieces of
sizes of 1-2 cm．
Sample preparation
Heat reflux extraction
Samples of the leaves (1 kg)was mixed with 12 kg
water in a 20 L extractor，heated in 100 ℃ for 2 h． The
extraction process repeated for 3 times，with the extract
filtrated each time，then all the extracts were combined
and cooled down to room temperature．
Macroreticular resins separation and purification
The 800 g macroreticular resins，which were pretrea-
ted，were placed into a 10 cm id glass column，and then
washed several times with deionized water． The extract
solution was flowed through the column at the rate of
0. 5 BV /h． After adsorption equilibrium，the adsorbents
were flushed with water for 6 BV at 1BV /h，70% aque-
ous ethanol for 7 BV at 1 BV /h． All the 70% aqueous
ethanol were collected and condensed to 1000 mL，then
3000 mL 95% aqueous ethanol was added，placed o-
vernight，filtered and evaporated by a rotary evaporator
at 50 ℃ ． The dried residue was sealed to preserve until
5601Vol. 23 ZHANG Lan-lan，et al:Ginsenosides in Leaves of Panax notoginseng，P． ginseng and P． quinquefolius
used．
Analytical methods
The separation was carried out on a Kinetex-C18 column
(100 × 4． 6 mm，2． 6 μm，phenomenex，at 30 ℃)． For
HPLC analysis，a 20 μL sample was injected into the
column and eluted at 30 ℃ at a constant flow rate of 1．
0 mL /min． Acetonitrile (A)and 0． 05% Phosphoric
acid aqueous solution (B)with a gradient elution:0-15
min，19% A，81% B;15-30 min，19% -31% A，81% -
69% B;30-50 min，31% -53% A，69% -47% B;50-60
min，53% -70% A，47% -30% B． Then the elution sol-
vent was kept at 19% A and 81% B for 10 min at a
flow rate of 1． 0 mL /min before analyzing the next sam-
ple．
Notoginsenoside R1(0． 5075 mg /mL) ，ginsenoside Rg1
(0． 5156 mg /mL) ，Re (0． 5015 mg /mL) ，Rb1
(0. 5037 mg /mL) ，Rb2 (0． 5014 mg /mL) ，Rb3
(0. 5116 mg /mL) ，Rc (0． 4975 mg /mL) ，Rd
(0. 5125 mg /mL) ，Rh2 (0． 5081 mg /mL) ，Rg3
(0. 5103 mg /mL) ，and Rf(0． 5062 mg /mL)mixed
standard stork solution were prepared in methanol． A
series of standard operating solution of different con-
centrations were obtained by diluting the mixed stand-
ard stock solution． The concentrations of these com-
pounds in the samples were calculated according to the
regression parameters derived from the standard
curves．
Results
Evaluation of macroreticular resins for sample
preparation method
Macroreticular resins adsorption process for ginsen-
osides have been reported many times． In Fig． 1-3，elu-
tion curve was depicted to show the effect of 70% etha-
nol on the desorption behavior． The elution curves
Fig. 1 The elution curve of P． ginseng main ginsen-
osides from D101 desorpted by 70% ethanol．
Note:Y was the peak area and X was elution volume．6BVmeans that e-
lution volume was six times of the column volume．The same below．
showed that the column was successfully washed by
7BV 70% ethanol． The order of desorption efficiency
was Ginsenoside Re，-R1 ＞ GinsenosideRg1 ＞ Ginsen-
oside Rb1 ＞ Ginsenoside Rb3，-Rd．
Evaluation of the analytical methods
The calibration curves and precision were shown in Ta-
ble 1． All the correlation coefficients exceeded 0． 99，
suggesting good linearity． Precision was obtained by six
parallel experiments ． The accuracy of this method was
evaluated by the recovery experiment． The recoveries
rate were calculated by the following formula:recovery
(%) = (found amount-original amount)/ added a-
mount × 100% ． 50 mg of the extracts of P． quinquefo-
lius leaves，P． notoginseng leaves，P． ginseng leaves
were treated，respectively，as described in Sections
2. 3． to get the original amounts of these ginsenosides．
Then the fixed quantities of 11 ginsenosides were added
into 25 mg of the extract of leaves and analyzed in the
same way． All the data were the average values ob-
tained by six parallel experiments． The recovery rates of
the 11 ginsenosides were listed in Table1． The recover-
ies are between 96． 95% and 104． 17% ． The relative
standard deviations (RSD)were no more than 5% ．
These results demonstrated that the method proposed in
this paper was accurate for the quantitative determina-
tion of the 11 ginsenosides in samples． The standards
and typical sample chromatograms were shown in Fig．
6601 Nat Prod Res Dev Vol. 23
4．
Table 1 The calibration curves and recovery of ginsenosides determined by standard addition method．
Ginsenoside Calibration curves r Original
(mg)
Added
(mg)
found
(mg) Recovery RSD (%)
R1 Y = 257． 46X － 9． 8521 0． 9999 0． 11 0． 13 0． 24 0． 9998 2． 69
Rg1 Y = 273． 03X + 2． 1819 0． 9897 0． 39 0． 48 0． 89 1． 0416 3． 92
Re Y = 296． 76X － 10． 504 0． 9931 1． 53 1． 04 2． 59 1． 0192 2． 76
Rf Y = 301． 74X － 5． 888 0． 9896 0． 06 0． 05 0． 11 0． 9960 3． 32
Rb1 Y = 248． 13X － 0． 9699 0． 9989 0． 59 0． 43 1． 00 0． 9534 2． 99
Rc Y = 242． 73X + 7． 3315 0． 9985 0． 65 0． 50 1． 17 1． 0400 2． 75
Rb2 Y = 264． 51X + 8． 5753 0． 9994 1． 22 1． 31 2． 57 1． 0417 0． 73
Rb3 Y = 269． 89X － 2． 6274 0． 9918 4． 28 5． 15 9． 49 1． 0305 0． 68
Rd Y = 297． 67X + 19． 386 0． 9997 2． 77 3． 25 6． 05 1． 0092 0． 70
Rg3 Y = 384． 43X － 8． 4466 0． 9995 0． 20 0． 29 0． 49 0． 9986 2． 36
Rh2 Y = 464． 04X + 11． 236 0． 9988 0． 09 0． 16 0． 25 1． 0005 4． 24
Note:Y was the peak area and X was the concentration of analysis．
Fig. 4 The standards (A)and typical sample (B)
chromatograms．
Ginsenoside contents in the leaves of different gin-
seng species
The ginsenoside contents in the leaves of different gin-
seng species were shown in Table 2，which revealed
that the total content of 11 ginsenosides followed this
order:P． quinquefolius leaves ＞ P． ginseng leaves ＞
P． notoginseng leaves． The contents of protopanaxadiol
saponins (Rb1，Rb2，Rb3，Rc，Rd，Rg3，Rh2)ranked
as:P． quinquefolius leaves ＞ P． notoginseng leaves ＞
P． ginseng leaves，The content of protopanaxatriol sapo-
nins (R1，Rg1，Re，Rf)ranked as:P． ginseng leaves ＞
P． quinquefolius leaves ＞ P． notoginseng leaves． Gin-
senosides Rb3 and Rd were the main constituents of P．
quinquefolius leaves． The total contents Ginsenoside
Rb3 and Rd accounted for above 60% of total the total
amount of the 11 ginsenoside in P． quinquefolius leav-
es． Ginsenoside Rg1 and Re were the main constituents
of P． ginseng leaves． The total contents of Ginsenoside
Rg1 and Re accounted for above 58% of the total a-
mount of 11 ginsenoside in P． gingseng leaves． As for
the individual ginsenoside，Ginsenoside Rb3 was the
dominant component in P． quinquefolius leaves and P．
notoginseng leaves． Ginsenoside Re was dominant com-
ponent in P． ginseng． The content of ginsenoside Rb3 in
P． quinquefolius leaves is more than 48 times in P． gin-
seng leaves，and more than 3 times in P． notoginseng
leaves． Ginsenoside Rd was of similar amount in P． no-
toginseng leaves and P． ginseng leaves，however，the a-
mount in P． quinquefolius leaves was 2 times higher．
Ginsenoside Rf，the chemical marker of P． ginseng was
also detected in P． ginseng leaves，but not found in the
two others． Only trace amount of ginsenoside Rh2 was
found in P． quinquefolius leaves，and notoginsenoside
R1 was undetectable in P． gingseng leaves and P． quin-
quefolius leaves．
Table 2 Contents of ginsenosides in the leaves of different
ginseng Species (mg·g-1)
Saponins P． quinquefoliusLeaves
P． ginseng
Leaves
P． notoginseng
Leaves
R1 － － 0． 40
7601Vol. 23 ZHANG Lan-lan，et al:Ginsenosides in Leaves of Panax notoginseng，P． ginseng and P． quinquefolius
Rg1 2． 20 8． 25 1． 56
Re 8． 59 16． 59 4． 89
Rf － 0． 26 －
Rb1 3． 26 5． 45 3． 20
Rc 3． 63 2． 66 0． 82
Rb2 6． 82 2． 28 1． 99
Rb3 23． 80 0． 49 7． 03
Rd 15． 48 6． 12 6． 88
Rg3 1． 11 0． 09 0． 23
Rh2 0． 50 － －
Protopanaxadiol 54． 60 17． 35 20． 15
Protopanaxatriol 10． 70 24． 84 6． 85
Total 65． 30 42． 19 27． 00
Conclusion and Discussion
The results revealed that not only the total saponins
content were different among different ginseng species
leaves，but also the proportions of individual ginsen-
oside were different． The result was similar with other
reports ［13］．
Our study indicated that P． quinquefolius leaves and P．
notoginseng leaves were good sources of protopanaxadi-
ol saponins，especially ginsenoside Rb3 and Rd，and P．
ginseng leaves was good sources of protopanaxatriol
saponins，especially ginsenoside Rg1 and Re．
Due to the the high content of ginsenosides in the leav-
es of different ginseng species，the leaves should be
throughtly used as a new ginsenoside supplier． Different
ginsenosides have different pharmacological effects，so
we can perform the extraction，separation and purifica-
tion based on the content differences in leaves of differ-
ent ginseng species．
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8601 Nat Prod Res Dev Vol. 23