Australian Researchers Publish Simple Method to Detect the Adulteration of Ginkgo Extracts
Reviewed: Wohlmuth H,
Savage K, Dowell A, Mouatt P. Adulteration of Ginkgo biloba products and a simple method to improve its
detection. Phytomedicine. 2014;21(6):912-918.
Abstract available at http://dx.doi.org/10.1016/j.phymed.2014.01.010.
There is growing concern in the global
botanical industry regarding the adulteration of ginkgo (Ginkgo biloba, Ginkgoaceae) leaf concentrated
extracts with exogenously derived (from other plants) free flavonols (also
known as free flavonol aglycones) or flavonol glycosides.1,2
Accordingly, several methods have been developed to deal with the adulteration
of ginkgo extracts with admixtures of flavonols or flavonol glycosides. Since
the official monographs (e.g., the methods of the United States
Pharmacopeia-National Formulary [USP-NF] and the European Pharmacopoeia
[EP]) for quality control of ginkgo extracts specify a flavonoid content
of 22-27% measured by high-performance liquid chromatography (HPLC) after
hydrolysis** of the flavonol glycosides, it is relatively easy to bring a
low-quality extract into compliance with the specified levels of flavonoid
content by adding pure rutin, pure quercetin, or flavonol-rich extracts from
other plant material. An overview of the widespread and unethical practice was
published in 2008.3
Despite the increased awareness of the problem in the dietary supplement
industry, both in the United States and elsewhere, the adulteration of ginkgo
extracts has persisted, as evidenced by more recent publications.4,5
Possible solutions for detecting such adulteration are the following methods:
(1) calculation of quercetin (Q) to kaempferol (K) and isorhamnetin (I) ratios
(ginkgo material should have a Q:K ratio between 0.8 and 1.2 and a Q:I ratio ≥
0.1 according to the USP); (2) the use of an HPLC fingerprint of the
non-hydrolyzed extract; or (3) a combination of HPLC fingerprints and ultraviolet
(UV) spectral images analyzed by statistical means.3-6
The newly published method by Australian researchers Hans Wohlmuth, PhD, and
colleagues of Southern Cross University in Lismore describes a simple addition
to the current pharmacopeial methods. (Dr. Wohlmuth is the research and
development manager at Integria Healthcare in Australia and is a recently
appointed member of the ABC Advisory Board.) The authors analyzed ginkgo leaf
material and commercial extracts before and after hydrolysis using the HPLC
conditions detailed in the USP 36-NF 31
(the combined 36th edition of the United States Pharmacopeia and the
31st edition of the National Formulary).6
Pharmacopeial-grade ginkgo extract is made by concentrating ginkgo material by
approximately 50 times, thereby producing the compendially required levels of
ginkgolides and bilobalide (6-7%; these compounds are unique to ginkgo) and the
flavonol levels of 22-27% (these flavonols are relatively ubiquitous in
plants). Adulteration of substandard ginkgo material with exogenously obtained
flavonols creates the false impression that the adulterated extract has been
concentrated to the level of the pharmacopeial-grade extract.
Non-hydrolyzed ginkgo leaf raw material contains very little to no detectable
quercetin, kaempferol, or isorhamnetin (even a low-quality manufacturing
process is not expected to yield large amounts of free flavonols, according to
the authors); therefore, substantial amounts of these flavonols are an
indicator of adulteration. The total flavonoid content can be calculated using
the difference between pre- and post-hydrolysis aglycone content. An advantage
of using this approach is the ability to use the same HPLC conditions that are
specified in the pharmacopeial monographs, so a quality control laboratory does
not have to invest in a new analytical instrument or a new HPLC column. Samples
are prepared by sonicating (sonication is a process in which energy in the form
of ultrasonic waves is applied to a material in a solvent to accelerate
extraction or dissolution) 300-1000 mg of commercial product in 50 mL ethanol
for 15 minutes.
The authors discovered admixtures of free quercetin and kaempferol in three of
the eight commercial samples that were analyzed, even if the flavonoid contents
were within the specifications set by the USP
36-NF 31. In addition, the three adulterated samples contained genistein,
an isoflavone that has not been found in ginkgo leaves but has been reported in
one instance in a ginkgo leaf extract of Chinese provenance.7 The
authors hypothesize that the genistein could come from extracts of the fruit of
the Japanese pagoda tree (Styphnolobium japonicum, Fabaceae; syn. Sophora
japonica), a plant that previously has been identified as a likely
adulterant of ginkgo extracts.8
Comment: The proposed new method is a quick and easy way to detect the
admixture of free flavonols added to ginkgo extracts and is suggested as an
amendment to current pharmacopeial methods. Large amounts of quercetin,
kaempferol, or isorhamnetin in non-hydrolyzed materials indicate adulteration.
One drawback of the method is that adulteration with pure flavonol glycosides
(e.g., pure rutin, or flavonol glycoside-rich herbal extracts such as those of
buckwheat [Fagopyrum esculentum,
Polygonaceae] or Japanese pagoda) will remain undetected. In such cases,
the determination of the Q:K and Q:I ratios outlined in the USP 36-NF 31 provides a solution. In
addition, the published HPLC fingerprinting methods4,5 can be used
to obtain a definitive answer regarding the authenticity of the ginkgo extract.
** Hydrolysis
is a chemical reaction used in this case to remove the sugar moieties from the
flavonol aglycones quercetin, kaempferol, and isorhamnetin. The result is the
reduction of a highly complex chromatographic profile to a simple HPLC trace
consisting of only three major peaks. The total flavonoid contents are obtained
using a mathematical function that converts the amount of flavonol aglycones to
the corresponding glycosides.
References
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