Ginseng Evaluation Program
Ginseng Evaluation Program
Ginseng Evaluation Program
Council
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In order to ensure fairness and objectivity, ABC has developed strict protocols for obtaining and
storing products, shipping samples to the testing laboratories, as well as reviewing results. All
products analyzed were purchased from retail establishments (including health food stores,
grocery stores, drugstores, and ethnic grocery stores), mail order, multilevel marketing
distributors, and/or health care professionals. ABC has not accepted in this program any ginseng
products that were sent directly by the manufacturer, in order to preclude the possibility that
uncharacteristic products could be evaluated.
GEP has three phases: the Regular Testing Phase, New Lot Testing Phase, and the Standardized
Testing Phase. The Regular Testing Phase is the original idea behind the GEP. Products were
purchased throughout the U.S. and Canada, analyzed up to four times, and given a status of Pass
or Fail.
Upon completion of the analyses, ABC notified manufacturers with products that had failed of
the results of the analyses for their products. Many of those manufacturers expressed concern
that the products analyzed in GEP were older products that had since been improved or had been
discontinued. In some cases manufacturers explained that they had discovered problems and
corrected them. Many manufacturers urged ABC to test the new version of their product
currently on the market. After assessing all the input and based on the fact that many of the
original products were manufactured prior to the passing of Dietary Supplement Health and
Education Act of 1994 (DSHEA), ABC decided to initiate the New Lot Testing Phase. For each
product that had failed in the Regular Lot Testing Phase, a new lot was purchased and analyzed
one time at one lab. Each of these products received a Pass or Fail status. The results for both the
Original lot and the New lot will be published in the final GEP report.
In the process of acquiring products for the Regular Testing Phase, it became evident that many
products claimed to be standardized and some of those claimed to be standardized to a specific
level of ginsenosides. Not knowing what each manufacturer means by standardized, ABC
decided to expand the GEP to review a small group of standardized Panax ginseng products.
Fourteen different products were selected and 5 different lots of each product were purchased.
These products do not receive a Pass or Fail status. Rather, the GEP report will include a
comparison of the different standardized products plus a comparison of the different lots of the
standardized products to determine any variation in the total quantity of key constituents and/or
the relative ratios or profiles of these compounds. This second comparison will show if a
standardized product is consistent on a lot-by-lot basis.
Laboratories & Methodologies
ABC has been working with the University of Illinois at Chicago (UIC) and University of
Ottawa (UO). Both labs worked together to develop the methodologies for analysis of
ginsenosides and eleutherosides. Both laboratories were involved in the analysis of the products.
In addition, UIC isolated many of the necessary chemical reference standards used in the
analyses.
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Results
The dissemination of information in the GEP Report will have a significant impact on the health
and well being of the general public as it impacts the future policy of research, manufacturing,
and claims about natural products. The level of responsibility on the part of industry will
continue to increase. In addition, due to the rapidly expanding interest in the use of herbal
products, the industry and the public will see the great value in utilizing this pioneering study as
a model to conduct further studies analyzing herbal dietary supplements.
The GEP reports will be featured in multiple issues of HerbalGram. The first report appears in
the Summer 2001 issue of HerbalGram and discusses the consistency of standardized Asian
ginseng products. The remaining reports will cover eleuthero (aka Siberian ginseng) and
American and Asian ginseng products and whether or not they meet their label claims. In
addition to the GEP reports, these special ginseng issues will include additional articles on
ginseng covering such topics as characteristics, research, taxonomy, conservation, history, etc.
If you would like to order HerbalGram 52 containing the first GEP report and do not want to
become a member at this time, call the ABC toll free order line at 800-373-7105.
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Ginsenoside Methodology
HPLC Methods for Separation and Quantitative Determination of GinsenosidesUsed in the
American Botanical Council's Ginseng Evaluation Program
P.N. Yat and J.T. Arnason
Ottawa-Carleton Institute of Biology, Department of Biology,
University of Ottawa, Ottawa, Ontario K1N 6N5, Canada.
Z.Z. Lu, J.F. Fitzloff and H.H.S. Fong
Department of Medicinal Chemistry and Pharmacognosy, and
Program for Collaborative Research in Pharmaceutical Sciences
University of Illinois at Chicago, Chicago, IL 60612-7231, U.S.A.
D.V.C. Awang
MediPlant Consulting Services
15876 Thrift Avenue, White Rock, British Columbia V4B 2M8, Canada.
INTRODUCTION
Ginseng is one of the most widely used herbal drugs and is reported tohave a wide range of
therapeutic and pharmacological activities (1-3). The twomajor species of commerce are Panax
ginseng C.A. Meyer (Asian ginseng),and Panax quinquefolius L. (North American ginseng).
Both speciescontain active ginsenoside saponins, but there are significant differences intheir
identity and distribution. It has been observed that over thirtyginsenosides have been identified
from Panax spp. (4), however six of these,Rg1, Re, Rb1, Rc, Rb2, and Rd constitute the major
ginsenosides accounting forover 90% of the saponin content of ginseng root (5).
Methods for the identification and quantitative analysis of ginsenosidesare available in the
literature (6). However, methods such as colorimetry mayoverestimate results and cannot give
information on individual ginsenoside levels,while gas chromatography has other reported
difficulties (7, 8). HPLC methodshave been the most successful and are now the most widely
accepted analyticalprocedure (7-9). A definitive HPLC/MS method is now available (10).
Based on a reviewof verified and widely used High Pressure Liquid Chromatography
(HPLC)methods reported in the scientific literature, such as the Soldati method (7,8),we updated,
adapted and verified an HPLC method for the quantification ofseven major ginsenosides
including Rf, whose presence or absence is the basisfor the HPLC differentiation between Asian
and North American ginseng, fromwhich Rf is absent. Rg2, also absent from North American
ginseng, for whichpurported ginseng products were initially screened, was not assayed for
because agood standard was not available in sufficient quantities.
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Page 6 of 16
At the UIC Collegeof Pharmacy laboratory: 2-6g of each sample was weighed accurately,directly
into a 10mL volumetric flask, diluted with methanol and made up to themark prior to HPLC
analysis. The sample solution was filtered directly into theHPLC sample insert just before HPLC
analysis.
Recovery of Ginsenosides.
At the University of Ottawa, replicate analyses were performedon recovery of standard
ginsenoside Rg1 in root sample, and of a mixtureof standard ginsenosides, Rg1, Re, Rb1, and Rd
in different ginsengformulations: powder, capsule, softgel, syrup and liquid. Two equal amounts
by weight ofa particular ginseng formulation were taken and standard ginsenoside(s)added to one
of these. Extraction procedures were identically performed on thetwo samples and the extract
analyzed. Analysis on the unaltered extract wasfollowed by analysis of the extract containing
standard(s). Recovery was determinedby taking the difference between the total ginsenoside
concentration(including added standard) and the detected ginsenoside in extracts without
standardadded. This spiking procedure also allowed verification of the peak identity inextracts,
since there was no development of split or shouldered peaks or change inonline spectra in the
rising or declining portion of the peak.
Recovery studies were performed similarly at the UIC Collegeof Pharmacy laboratory; the
number of recovery studies was limited by theamount of available pure ginsenoside standards;
these are reported in Table1.
HPLC Analysis.
The HPLC used at the University of Ottawa was a Beckman System Gold,consisting of a
Beckman Diode-array detector (module 168), a programmable binarysolvent delivery system
(module 126), an autosampler (module 502), capable ofinjecting up to 94 samples, a 486 Dell
(IBM-compatible) computer for dataprocessing, equipped with Beckman System Gold software.
The equipment was certifiedsemi-annually by Beckman technicians. Separations were achieved
with Beckman ultrasphereODS, 5um particle, 250 x 4.6mm column. For ginsenoside analysis,
the mobilephase was water (A) and acetonitrile (B) at a constant composition of 2l%B from0-20
minutes and flow rate of 1.3 mL/min; and a gradient elution rising to 42%Bfrom 20-60 minutes.
At 60-70 minutes, the column was flushed with 99%B thenreturned and equilibrated for 15
minutes with 21%B. Detection of ginsenosides wasachieved at 203nm.
At the UIC Collegeof Pharmacy laboratory, analysis was performed with a Waters HPLC
systemconsisting of model 510 EF pumps, model 717 autosampler, and model 486 UVVISdetector set at 202nm and equipped with a MillenniumR version 2.15 programmablesystem
controller and data processing software; the column used was also aBeckman Ultrasphere ODS
5um particle, 250 x 4.6mm.
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Page 8 of 16
Page 9 of 16
Ginseng
Syrup
formulations
96
107.4
104.1
98.3
96
(-)
(8.1)
124.1
(5.9)
96.0
(1.5)
101.2
(-)
Re
Rf
(5.5)
109.8
(3.7)
96.8
(1.7)
85.9
Rb1
(-)
108.3
(2.9)
102.3
(2.2)
95.5
Rc
(6.9)
104.0
(5.9)
107.3
(2.5)
98.7
Rb2
(1.1)
100.1
(6.3)
92.3
(1.6)
100.9
Rd
(1.7)
106.4
(3.17)
103.1
(0.9)
99.0
100
(3.9)
(7.4)
(3.1)
(-)
103
104
105
106
102
102
110
102
94
96
99
Chicago
Rg1
Rb1
Rc
Rd
95
(-)
Note: For Ottawa, n = 4-5 where s.e. is given, n = 1 where nos.e. is given. For Chicago, n = 2.
The response of the LC system to standard ginsenosides invarious ranges of concentration was
highly linear with an r2>0.99 in all cases (Table 2a). The procedure enabled detection of 12ng of
Rg2and 22ng Rg1 at above background noise levels.
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Table 2a.
Linearity of peak area to concentration (Ottawa)
Ginsenoside
Rg1
Re
Rf
Rg2
Rb1
Rc
Rb2
Rd
r2
0.9996
0.9980
0.9991
0.9997
0.9905
0.9971
0.9972
0.993
Similar linear response data was obtained in the Chicago laboratory; seeTable 2b.
Table 2b. Calibration curves and detection limit(Chicago)
In order to check the linearrelationships between UV and absorption intensity and amount
ofginsenosides, calibration curves are established for Rg1, Re, Rb1, Rc, Rb2, Rd.
Regression Analysis of Standard Ginsenosides:
Correlation Coefficient r2
Ginsenoside
Rg1
Re
Rb1
Rc
Rb2
Rd
r2
1.000
1.000
1.000
1.000
0.999
1.000
REFERENCES
1.H.Yamaguchi, H.Matsuura, R.Kasai, O.Tanaka, M.Satake, H.Kohda, H.Izumi, M.Nuno,
S.Katsuki, S. Isoda, J. Shoji an K. Goto, Chem. Pharm.Bull. 36, 4177(1988).
2. C-X.Liu, and P-G.Xiao, Journal of Ethnopharmacology, 36, 27(1992).
3. T.Kaku, Y.Kawashima, Chem.Abstr., 89, 70806d (1973).
4. M.Sakata et al, Proceedings of the 3rd International GinsengSymposium, 8 (1980).
Page 11 of 16
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Root
(Main)
0.91
Lateral
Root
Root
Fiber
Leaves
Leaf
stalks
Stem
5.19
0.77
0.76
3.72-9.03
Korean White
Korean Red
3.90-7.19
Ko, 1989 (4)
Liu,
1992
(5)
(rootstock, not normal root)
Ma, 1995 (6)
Soldati, 1980 (9)
Sollorz, 1985 (10)
3.3-7.2
4.19
1.20-3.95
1.35
3.53
6.15
6.750.69-1.80 1.54-2.93
8.56
The minimum level of ginsenosides found in the literature search was 0.69%. This value was
then adjusted for the coefficient of variation for the test method and for the representative
amount of ginsenosides assayed. The result is the Root Powder Minimum (RPM) of 0.59% for
Asian ginseng
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Root (whole)
Tea
3.09
8.65-24.00
1.17-8.60
6.2
1.70
0.23-9.85
The minimum level of ginsenosides found in the literature search for American ginseng was
1.17%. This value was then adjusted for the coefficient of variation for the test method and for
the representative amount of ginsenosides assayed. The result is the Root Powder Minimum
(RPM) of 1.00% for American ginseng
ELEUTHERO, AKA SIBERIAN GINSENG (ELEUTHEROCOCCUS SENTICOSUS)
Eleutheroside
B
Eleutheroside
E
Bladt,1990 (1)
Eleutherococci radix type A (China / Russia /
Korea)
Eleutherococci radix type B (China / Korea / Japan)
E. senticosus (Korea) root
E. senticosus (Korea) Root bark
E. senticosus (Korea) Wood
E. senticosus (Korea) Stem bark
0.030.15%
0.040.13%
(traces)
0.150.18%
0.090.11%
0.01%
0.020.05%
0.130.15%
0.090.10%
0.070.08%
0.05%
0.0550.178%
0.100.120%
1.23%
Since some types of eleuthero contain no eleutheroside B, the GEP team set no root powder
minimum for eleutheroside B content. The minimum level of eleutheroside E found in the
literature search for eleuthero was 0.02%. This value was then adjusted for the coefficient of
variation for the test method. The result is the Root Powder Minimum (RPM) of 0.019% for
eleuthero root powder.
For eleuthero root bark powder, the minimum level of eleutheroside E found in the literature
search was 0.09%. This value was then adjusted for the coefficient of variation for the test
method. The result is the Root Powder Minimum (RPM) of 0.088% for eleuthero root bark
powder.
Page 14 of 16
References
1.
2.
Chang YS. Chemical and pharmacological studies of Panax quinquefolius and Panax
ginseng [thesis]. Chicago (IL): Univ. of Illinois at Chicago; 1989.
3. Chuang WC, Wu HK, Sheu SJ, Chiou SH, Chang HC, Chen YP. A comparative study on
commercial samples of ginseng radix. Planta Med. 1995;61:459-465.
4. Ko SR, et.al., Saengyak Hakhoechi, 1989,20,170
5.
Journal of
6.
Page 15 of 16
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Page 16 of 16
Links
For more information on herbal product quality and testing of herbal products we invite you to
visit the following sites.
ConsumerLab.com (CL) provides independent test results and information to help consumers
and healthcare professionals evaluate health, wellness, and nutrition products. It publishes results
of its tests on-line at www.consumerlab.com. Products that pass CL's testing are eligible to bear
the CL certification seal. CL addresses a growing need of consumers and healthcare
professionals for better information to guide the selection of health, wellness, and nutritional
products.
Dietary Supplement Quality Initiative
The DSQI Web Site is the most publicly visible part of the Dietary Supplement Quality
Initiative. Beyond our consumer-oriented activities is a commitment to develop comprehensive
quality practices and standards for dietary supplements. We believe increased levels of research
in combination with widely accepted standards will benefit those who use, prescribe and make
dietary supplements. In promoting a science-based understanding of the health benefits
associated with supplementation we believe the quality, safety and availability of dietary
supplements can only increase.
The Institute for Nutraceutical Advancement
The INA is a non-corporate division of Denver-based Industrial Laboratories, an independent
laboratory that provides analytical and consulting services to the natural products industry. The
INA's mission is to support the production of high quality botanical products. To serve their
mission, in 1998, they launched the INA Methods Validation Program (MVP), an international
project designed to select, validate and publish scientific methods for use in analyzing raw
botanical materials. The INA MVP is funded through contributions from 30 companies,
including some of the industry's largest producers of dietary supplements and botanical raw
materials. These 30 companies along with seven industry organizations and the FDA comprise
the INA MVP Advisory Committee.