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Biology and Chemistry of Ginkgo biloba

Article in Fitoterapia · September 2008

DOI: 10.1016/j.fitote.2008.05.007 · Source: PubMed

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 Fitoterapia 79 (2008) 401 – 418
www.elsevier.com/locate/fitote

Review
Biology and chemistry of Ginkgo biloba
Bikram Singh ⁎, Pushpinder Kaur, Gopichand, R.D. Singh, P.S. Ahuja
Institute of Himalayan Bioresource Technology, Palampur, HP — 176 061, India
Received 7 January 2008; accepted 12 May 2008
Available online 27 June 2008

Abstract

Ginkgo biloba has been existing on earth since 200 million years and is considered as a “living fossil”. It is among the most sold
medicinal plants in the world. A number of secondary metabolites representing terpenoids, polyphenols, allyl phenols, organic
acids, carbohydrates, fatty acids and lipids, inorganic salts and amino acids have been isolated from the plant. However, the main
bioactive constituents are terpene trilactones and flavonoid glycosides which are considered responsible for the pharmacological
activities of its standardized leaf extract. Scattered information is available on the extraction and analysis of these pharma-
cologically important constituents which have been compiled in the present review.
© 2008 Elsevier B.V. All rights reserved.

Keywords: Ginkgo biloba; Biology; Chemistry

Contents

1. History . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 402
2. Nomenclature and systematics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 402
3. Distribution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 403
4. Ecology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 403
5. Sexual reproduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 403
6. Seed dispersal and establishment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 404
7. Tree architecture . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 404
8. Seed production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 405
9. Leaf production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 405
10. Cultivation and propagation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 405
11. Pharmacological importance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 409
12. Economical importance of the plant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 409
13. Secondary metabolites . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 412
13.1. Flavonoid glycosides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 416
Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 416
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 416

⁎ Corresponding author. Fax: +91 1894 230433.

E-mail address: bikram_npp@rediffmail.com (B. Singh).

0367-326X/$ - see front matter © 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.fitote.2008.05.007
402 B. Singh et al. / Fitoterapia 79 (2008) 401–418

1. History

Ginkgo biloba L.(Syn.: Salisburia adiantifolia, Salisburia biloba Hoffmag-Common names: maidenhair-tree,
ginkgo), has identity as a valuable plant for mankind since more than 2000 years and is considered a “living fossil”
(Fig. 1). Though its natural habitat is in China, Japan and Korea, its place of origin is believed to be remote
mountainous valleys of Zhejiang province of eastern China [1,2]. Up to 350 years ago, knowledge about this plant
resource was limited to China. More than 100 plants of G. biloba ageing over 1000 years still exist around temples in
China [2] as its nuts are used in worship. Ginkgo probably originated about 200 million years ago, though its
swimming sperm (spermatozoids) were discovered hundred years ago [2].
Currently, G. biloba does not have any close relative in the plant kingdom. Thus, it is classified in a separate
division, the Ginkgophyta. This taxon is distinguished from the Coniferophyta (Conifers) on the basis of its
reproductive structures, its multiflagellated sperm cells, and from the Cycadophyta (Cycads) owing to its vegetative
anatomy [3]. Molecular characterization of the genome has indicated that Ginkgo is distant from conifers and rather
much closer to the Cycads [2]. The uncertainty has recently been resolved in Henan Province, China, by the discovery
of fossils belonging to the Middle Jurassic period (180 million years ago) with Ginkgo like ovule bearing organs [4].
Ginkgo yimaensis is a new species that differs from G. biloba in having more highly dissected leaves and much
smaller ovules clustered attached on branched peduncles. Other extinct species G. adenoids had an extensive distribution
in the northern hemisphere from the lower Cretaceous through the Pliocene and many researchers consider it to be the
ancestor of G. biloba because it had a similar type of leaf and ovule structure. About 98–65 mya, during the upper
Cretaceous period the fossil record for Ginkgo shows a decline in diversity and distribution, particularly due to lowering
the temperature. The genus disappeared from polar areas through the end of the Miocene. By the end of the Pliocene, as
temperature declined and the rainfall slightly shifted from summer-wet to summer-dry, the genus Ginkgo was not
traceable in Europe. From south western Japan, the G. biloba was reported in Pleistocene record. Based on leaf
characteristic, these fossils were classified and admitted as an extinct species G. biloba [2]. In terms of the reproductive
organs, the trend within the genus Ginkgo appears to be on the decline in the number of ovules with a contemporaneous
increase in the size of the ovules, loss of the pedicels resulting in sessile ovules connected directly to the peduncle [4–6].

2. Nomenclature and systematics

Linnaeus (1771) was the first pioneer who described G. biloba. Some other names like S. adiantifolia Smith and
Pterophyllus salisburiensis Nelson are not considered its valid synonyms as per the rules of priority. The German
name is not in practical use. The English term “Maidenhair tree” is based on the resemblance to the foliage of the
“Maidenhair fern” (Adiantum). In Japan, it is known by “Ginkyo”. In France, it is known as “L'arbre aux Quarante
ecus” and “Noyer Du Japon”. Earlier the Ginkgo spp. was included in Taxaceae. In fact this family included
Podocarpaceae and Cepholotaxaceae and has always been considered very artificial. A discovery in 1895 revealed that

Fig. 1. Ginkgo biloba L.

 B. Singh et al. / Fitoterapia 79 (2008) 401–418 403

Ginkgo possesses multicoated spermatozoid, and it formed the basis of Engler's classification of a particular family and
class of Ginkgoopsida [7] which dates back as old as the lower Jurassic [8]. This group reaching at its highest position
in upper Jurassic and bottom in Cretaceous period had also disappeared rapidly before the end of the Cretaceous. In the
Oligocene time, only two genera out of the 19 genera comprising about 60 species of the family became extinct.
Currently, these spp. are non-traceable from all but one continent, in which only one single species exists owing to
natural or cultural reasons [9]. Thus, G. biloba is the prime botanical example of a “living fossil”. This term was earlier
used by Darwin for the King Crab (Limulus sp.). The systematic classification of G. biloba is as follows [10].
Division: Spermatophyta

1. Sub-division: Coniferophytina
i) Class: Ginkgoopsida
ii) Class: Pinopsida (with three sub-classes).
2. Sub-division: Cycadophytina
i) Class: Lyginopteridopsida (Seed fern)
ii) Class: Cycadopsida (Cycadaceae a. o.)
iii) Class: Pinopsida (Only known in fossil form)
iv) Class: Gnetopsida (Ephedra, Welwitschia, Gnetum)
3. Sub-division: Magnoliophytina (Angiospermae).

3. Distribution

G. biloba was introduced in Europe in the 18th century, sometime around 1730. The tree surviving in the botanical
garden of Utrecht is among the first ones that arrived in Europe. On September 27, 1815 Goethe wrote a poem in
Heidelberg, which is attributed to the glory of this tree. On the basis of material contained in certain publications [11],
proper prescription of G. biloba leaves for internal medical use in China seems to have been first mentioned in the book
of Liu Wen-Tai in 1505 A.D [12]. The fruit (baigo) is in use since ages and remark on its leaf extract is included in the
medical Dictionary of the Republic of China.
G. biloba is a member of the mixed mesophytic forest community that covered the hill country bordering the
Yangtze River Valley in China [13,14]. One of the last wild plants of G. biloba is reported to be in Zhejiang Province,
China on the west peak of Tianmu Mountain (1506 m amsl). Ginkgo was introduced in Europe around 1730 and is
widely cultivated as an ornamental tree in streets and parks and as a medicinal plant particularly in China, Korea,
France, Germany and the United States. The medicinal properties of Ginkgo seeds are reported in ‘Pen Ts'o Kang Mu’
[15]. A recent survey by the authors has revealed that there are about 30 plants of G. biloba planted all over India,
mostly in hilly tracts from north-west to north-east.

4. Ecology

The Tianmu Mountain in China the Ginkgo population is considered about 244 individuals with a mean diameter of
45 m at breast height (DBH) and a mean height of 18.4 m. Most of the population of Ginkgo was growing on degraded
sites, along stream beds, on rocky slopes and on the edges of soil beds. Ginkgo seedlings are quite rare on Tianmu
Mountains and are found in areas of the forests that have been highly disturbed. In some areas, the exceptional species
richness of the surrounding forest and the large size of many of its trees suggest that they may be well developed [16,17].
Recent studies on its population biology indicate a relatively low degree of genetic diversity among the plants. About
40% of the Tianmu Ginkgos have more than one trunk with DBH more than 10 cm. Most of these secondary trunks have
originated from lignotubers located at or just below ground level. Secondary trunk formation is most apparent in
specimens that were damaged due to severe stress. The sprouting ability of Ginkgo is an important property that has
supported the species to perpetuate on the eroded mountains and in survival and morphological stability of the species.

5. Sexual reproduction

Ginkgo is a dioecious species, with male and female plants occurring in about 1:1 ratio, with sporadic report of
monoecious individual [18]. Its leaves are deciduous, alternate or in clusters of 3–5 on short twigs, petiolate, fan or
404 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Adiantum shaped, bilobed, thickened at the margin, 5–10 cm, across, broader than long with venation dichotomously
branching seemingly parallel.
The name of Ginkgo has been derived from a wrong transcription of the Japanese name Yin-Kwo, meaning Silver
fruit. The name of the species biloba, recalls the bilobed shape of the leaves. The resemblance of the shape and vein of
its leaf with those of the leaf of maidenhair fern has established its common English name. In China and Japan, it is
considered sacred and is cultivated around temples. The male trees are more desirable for planting, because female
plants produce foul-smelling seeds. G. biloba is a tree of great beauty with a long life span, highly resistant to insects,
bacterial and viral infections and air pollution. Ginkgo shows a long juvenile period, typically not reaching sexual
maturity until 20 to 30 years. Male and female sex organs are produced on short shoots, in the axils of bud scales and
leaves. The male catkins emerge before the leaves and fall off immediately after shedding their pollen. Wind pollination
occurs during early April in the regions with mild winters, and during late May in areas with severe winters. As the
pollination ceases, it is difficult to say what is the maximum limit of dispersal of pollens, however in the Boston area no
setback to seed setting was recorded even when the male and female trees were 400 m apart [19]. Its ovules are 2 to
3 mm long and produced in pairs at the end of stalks 1.5 to 2.0 cm long. In receptive stage, it secretes a somewhat
mucilaginous fluid from its micropyle, which facilitates trapping airborne pollen chamber. Once inside the ovule, the
male gametophyte commences a 4 month long development period that culminates into production of a pair of
multiflagellated spermatozoids, one of which fertilizes a waiting egg cell [20] while the ovules are still on the tree [21].
Depending upon the date of pollination, the union can occur anytime from late August to late September. Ginkgo seed
at maturation is large and consists of an embryo encircled, in the tissue of the female gametophyte covered by a thick
seed-coat layer. The seed-coat consists of a soft, fleshy outer layer (sarcotesta), a hard, stony middle layer and a thin
membranous inner layer. The seed contains fleshy sarcotesta, which is generally known as the Ginkgo nut. The
developing ovules of Ginkgo are green in color up to maturation, when owing to cold temperature they attain yellow
color as that of the leaves. The seeds drop from the tree about a month after fertilization. The foul odor from Ginkgo
seeds comes out only after they are fully mature. The sarcotesta contains two volatile compounds viz., butanoic and
hexanoic acids. The seeds also contain phenolic compounds causing contact dermatitis in humans [2].

6. Seed dispersal and establishment

It is a mystery about the animal which might have dispersed Ginkgo seeds during the course of its evolution. Several
workers perceive that dinosaurs might have dispersed Ginkgo seeds, but none of them have provided any evidence to
support the claim [2]. It is believed that the early mammals in the extinct family Mutituberculata could have been
effective dispersal agents [19]. These marsupials like creatures, often referred to as the ‘rodent of the Mesozoic’, were
found in the temperate parts of the northern hemisphere from the late Jurassic through the Oligocene. At present, so
many mammals are observed feeding on, and presumably dispersing, the odoriferous, nutrient-rich seeds of G. biloba.
In Rodentia order, these include the red-bellied squirrel (Callosciurus flavimanus var. ning poemsis, family Sciuridae)
in the region of Tianmu Mountain [16], and gray squirrel (Sciurus carolinensis, family Sciuridae) in eastern North
America [19]. In Carnivora order, potential dispersal agents assumed the masked palm civet (Paguma larvata, family
Viveridae) in the Tianmu Mountain area. The leopard cat (Felis bengalensis, family Felidae) in Hubei Province, China
[19] and the raccoon dog (Nyctereutes procyonoides, family Conidae) in Japan [22].
The existence of three independent reports of carnivores consuming whole Ginkgo seeds and defecating intact nuts
raises the possibility that the foul foul-smelling sarcotesta may be attracting these animals [16]. Ginkgo seeds are
dormant when they fall from the tree because the embryo is not fully developed, and are only 4 to 5 mm in length. In
case seeds are collected shortly after dispersal, cleaned, and placed in a warm greenhouse, the embryo will grow to its
full size i.e. 10 to 12 mm in length and germinate within 8 to 10 weeks [21] In the warm-temperate climate, Ginkgo
seeds are shed in late summer, whereas under cold temperature seeds are shed late in the season. The colder temperature
delays full embryo development until the ensuing spring.

7. Tree architecture

G. biloba is admitted as a long lived deciduous tree. Its stem diameter has been recorded to range from 1 to 4 m [23],
depending upon the agro-climatic conditions. A mature tree may attain up to 20 to 40 m height. However, one
exceptional specimen in Korea is reported to be 64 m tall with 14 m diameter [24]. The appearance of a fully developed
 B. Singh et al. / Fitoterapia 79 (2008) 401–418 405

young Ginkgo tree is distinctly pyramidal with a dominant central trunk and widely spaced diagonal orientated
branches in whorls. The sexual maturity takes about 25 years. Vertical growth generally slows down and the tree fills in
its sparsely branched Juvenile structure, with a broad, spreading canopy [25,26].
After about 10 years of growth, Ginkgo produces two types of shoots; long shoots with widely spaced leaves that
subtend axillary buds, and short shoots with clustered leaves that lack both internodes and axillary buds. The long
shoots are basically responsible for over all growth of the tree and generating new growing points. The short shoots
produce clusters of leaves with reproductive structures. In due course of time, the effect of environmental and
physiological factors on the growth pattern of these shoots can be changed i.e. short shoot can produce a long shoot,
and the terminal growth of a long shoot may be restricted to produce a short shoot [25]. This alternation provides a
simple mechanism for modulating carbohydrate distribution between sexual reproduction and vegetative growth [26].
Under adverse growing conditions, both in stress and in higher accumulation of food material, Ginkgo is capable of
producing secondary trunks, may be 2–3 in number, just below ground level. Generally, these secondary stems
originate from root like positively geotropic shoots known as lignotubers or ‘basal chichi’ or by simple buds just below
the soil surface. Anatomically lignotubers are developed in all Ginkgo seedlings as part of their normal ontogeny from
buds located in the axils of the two cotyledons, and also with vegetative propagated plants by using stem cuttings.

8. Seed production

The seed of G. biloba contains a thin-layer nut, which is consumed as food and a medicine throughout the world.
Fresh nuts contain about 41% moisture. Dry nuts contain 6% sucrose, 68% starch, 13% protein and 3% fat [27]. The
commercial production of Ginkgo nuts has been reported in China for more than 600 years. About 44 cultivars with
best performance in growth have been selected, based on quality, size and shape of their nuts and productivity [2,28].
These cultivars are propagated by grafting technique using seedling rootstocks. These grafted plants start bearing nuts
after 5 years of age. In China, a widely grown cultivar, Dafushon, in Jiangsu Province, produces 5 to 10 kg of nuts by a
15 year old tree and between 50 to 100 kg by a 50 year old tree. One recent production from 700,000 to 800,000 trees
produced an average of 6000–7000 tons of dry nuts per year [2].

9. Leaf production

Recently (1982), G. biloba has been planted on a large scale in France (480 ha) and the United States (460 ha). For
this purpose, the saplings are planted at a closer spacing of 40 cm in rows at 1 m distance accommodating 25,000 plants
per hectare. Height of the plantation is maintained at about 3 m by deploying special pruning technique to facilitate
mechanical harvesting of leaves. Green leaves were harvested in summer season. In a rotation of 4 to 5 years of crop
season, the plants are pruned to near ground level, after harvesting. After such pruning or cutback operation, new
shoots sprout from buds located at the base of the stem or on basal lignotubers. With proper irrigation and fertilizers
application in growing season, the annual vertical growth from the point of cutting is generally 1 m or even more [25].
In Eastern China, after 1992, Ginkgo seedlings have been established about 2000 ha area specifically for commercial
leaf production. These plantations began producing marketable quantities of leaves in 1996, and the estimated yield
will be increased dramatically over the next 5 to 10 years [2,29].

10. Cultivation and propagation

Perennial plantations are raised with a plant to plant spacing of 3 to 6 m. It prefers full sunlight and well drained soil
with mean soil temperature 15 to 27 °C. Ginkgo exhibits resistance to infection and air pollution. Fifty million G.
biloba trees are grown especially in China, France and South Carolina, USA, producing over 8000 tons of dried leaves
each year to meet the commercial demand.
A tree of Ginkgo is known to be over 5000 years old [30]. A study on effect of physical treatment on germination of
G. biloba seeds has confirmed that the seed viability decreased over time [31]. There are studies on effect of foliar
application of nitrogen on 3 year old G. biloba seedlings [32] and on effect of nitrogen, phosphorus and potassium on
leaf and vegetative growth of G. biloba [33,34]. Fang Ren et al. [35] have examined the microclimatic observations
and intercropping system with G. biloba. Youchao et al. [36] have studied the coppice management to improve the
yield and quality of G. biloba leaf producing plantation. Quan BingYan et al. [37], Wang et al. [38] and Kang Zhi et al.
406 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Table 1
Secondary metabolites reported from G. biloba root bark, wood, and leaves
Material Reference
Terpenes
Monoterpenes [64]
Cymene (1)
Isopropyl-phenol (2)
Thymol (3)
Linalool oxide (4)
Ionone (5)
Diterpenes [68,69]
Ginkgolide A (6)
Ginkgolide B (7)
Ginkgolide C (8)
Ginkgolide J (9)
Ginkgolide M (10)
Ginkgolide K (11)
Ginkgolide L (12)
Sesquiterpenes [64,69]
Bilobalide (13)
Bilobanone (14)
E- and Z- forms of 10-11-Dihydroatlantone (15)
E-10-11-Dihydro-6-oxoatlantone (16)
Elemol (17)
Eudesmol (18)
Steroids, phytosterols [64]
β-Sitosterol (19)
Stigmasterol (20)
Campesterol (21)
Dihydrobrassicasterol [64]
Carotenoids [64]
α-Carotene (22)
γ-Carotene (23)
Lutein (24)
Zeaxanthin (25)
Polyprenols [64,69]
Ginkgo polyprenols (26)
Ginkgo polyprenol acetates (27)
Flavonoids
Glycosides of [64,69]
Kaempferol (28)
Quercetin (29)
Myricetin (30)
Apigenin (31)
Isorhamnetin (32)
Luteolin (33)
Aglycones
Kaempferol(28)
Quercetin (29)
Myricetin (30)
Apigenin (31)
Isorhamnetin (32)
Luteolin (33)
Tamarixetin (34)
4′-OMe Apigenin (35)
3′-Methylmyricetin (36)
Catechin (37)
Epicatechin (38)
Epigallocatechin (39)
Gallocatechin (40)
 B. Singh et al. / Fitoterapia 79 (2008) 401–418 407

Table 1 (continued )
Material Reference
Dimers [64]
Catechin–catechin
Epicatechin–catechin
Epigallocatechin–catechin
Gallocatechin–catechin
Anthocyanidin [69]
Procyanidin (41)
Prodelphinidin (42)
Biflavones [64,69]
Amentoflavone (43)
7-Methoxyamentoflavone (44)
Bilobetin (45)
5′-Methoxybilobetin (46)
Sequojaflavone (47)
Ginkgetin (48)
Isoginkgetin (49)
Sciadopitysin (50)
Biflavone glucosides [70]
Ginkgetin
Isoginkgetin
Alkyl phenols and alkyl phenolic acids [64]
Cardanols
3-Tridecylphenol (51)
3-Tetradecylphenol (52)
3-Pentadecylphenol (53)
3-Heptadecylphenol (54)
Ginkgol (55)
Cardols [64]
5-Tridecylresorcinol (56)
5-Tetradecylresorcinol (57)
5-Pentadecylresorcinol (58)
5-Heptadecylresorcinol (59)
Bilobolol (60)
Anacardic acids [64]
6-Tridecylsalicyclic acid (61)
6-Tetradecylsalicylic acid (62)
6-Pentadecylsalicylic acid (63)
6-[8-Pentadecenyl] salicylic acid (64)
6-Hexadecylsalicylic acid (65)
6-[9, 12-Heptadecadienyl] salicylic acid (66)
6-[8-Heptadecenyl] salicylic acid (67)
Resorcylic acids [64,69]
6-[8-Pentadecenyl] resorcylic acid (68)
6-Tridecylresorcylic acid (69)
Alkyl coumarin (70) [64,69]
Organic acids
Ascorbic acids (71)
D-glucaric acid (72)
Quinic acid (73)
Shikimic acid (74)
6-Hydroxykynurenic acid (6-HKA) (75)
p-Hydroxybenzoic acid (76)
Protocatechuic acid (77)
Vanillic acid (78)
Acetic acid
Butyric acid
Formic acid
Hexanoic acid
(continued on next page)
408 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Table 1 (continued )
Material Reference
Organic acids
Valeric acid
Carbohydrates [64]
Sucrose
Pinitol (79)
Sequoyitol (80)
Polysaccharides
Long chain hydrocarbons and lipids [64]
1-Hexacosanol
2-Hexenal
n-Nonacosane (81)
n-Nonacosan-10-ol (ginnol) (82)
n-Nonacosan-10-one (ginnon) (83)
n-Octacosan-1-ol
Linolenic acid
Linoleic acids
14-Methylhexadecanoic acid
5, 9, 12-Octadecatrienoic acid
Oleic acid
Palmitic acid
Eicosanedicarboxylic acid
Phellonic acid
Inorganic salts/complexes [64]
Selenium
Copper
Zinc
Chromium
Iron
Manganese
Calcium oxalate
Phosphoric acid
Miscellaneous organic compounds [64]
N-containing compounds
Aminobutyric acid
Asparagine
Aspartic acid
Glutamine
Arginine
Proline
Glycine
4-O-Methylpyridoxine (84)
Proline
Pyroglutamic acid
Serine
Threonine
Valine
Isopentenyladenine
Ribosylzeatin
Zeatin and dihydrozeatin (85)
Zeatin glucoside
Hexosamines
6-Hydroxykynurenic acid
Lectins
Pentadien-1, 5-diyl-diphenol
Others [64]
Tannins
Condensed tannins
Glycerol derivatives
1-Hydroxypyrene conjugates
 B. Singh et al. / Fitoterapia 79 (2008) 401–418 409

[39] have studied the effect of density and stump height on leaf yield of G. biloba. Huang et al. [40] and Sung Chiung
Huei et al. [41] have evaluated it for absorption and tolerance to ozone and nitrogen dioxide. Li Jian et al. [42] and
Leng-PS et al. [43] have studied the effect of intensity and quality of light on photosynthesis of G. biloba.
Ginkgo tree is an extremely slow growing one and its regeneration through seeds is very poor. Therefore, vegetative
multiplication through cuttings is the only possible option for augmenting regeneration. The stimulation of adventitious
root formation in stem cuttings treated with auxins and commercial rooting mixture containing auxin is well known
[44]. Li et al. [45] have obtained best result when G. biloba cuttings were treated with carbendazim. However,
crystalline potassium permanganate and IAA (Indole acetic acid) containing powder have also been recommended for
stimulating rooting of G. biloba [46]. In China, dipping the base of cutting into ABT-1 (aminobenzotriazole) rooting
powder for 1 h gave the best results [47,48]. In addition to auxins, phenolic compounds either alone or in combination
of auxins [49,50] have also stimulated adventitious root formation in cuttings of several species. Shen Xiao Qing et al.
[51] have studied the propagation by shoot cutting of G. biloba in clay pots. Om Parkash et al. [52] have tried IBA
(Indole-3-butyric acid) along with certain phenolics for root induction in stem cutting of G. biloba. Schmid [53] has
studied cultivation of G. biloba. Yu Xin et al. [54] have studied storage technology of G. biloba seeds at normal
atmospheric temperature. Wu [55] have has studied the techniques for growing seedlings of G. biloba. Lee and Lee
[56] have reported the optimal planting spacing on the basis of growth condition of landscape trees.

11. Pharmacological importance

Although Ginkgo tree has been around for 200 million years, it's only during the last couple of decades that its true
value has been recognized. Its amazing vitality has attracted an increasing exploration into potential application in
health, foods and supplements. The medicinal parts of Ginkgo tree are fresh or dried leaves and seeds separated from
their fleshy outer layer. It contains large number of active compounds, the most important of which are flavanol
glycosides and terpene trilactones. The extract of Ginkgo leaves are standardized for these two components. Active
compounds in Ginkgo extract improve blood circulation, discourage clot formation, reinforce the walls of the
capillaries, and protect nerve cells from harm when deprived of oxygen. The leaves extracts are used to treat demential
disorder, such as concentration difficulties and memory impairment. The extract also possesses antioxidant [57],
antiasthmatic [58], scavenge radicals [59], wound healing [60] and neuroprotective properties as well as it improves
mental capacities in Alzheimer's patients [61,62].

12. Economical importance of the plant

Western medical interest in G. biloba has grown dramatically since the 1980s owing to its potent action on
cardiovascular system of human beings, particularly on the cerebral vascular activity. Over 7 billion dollars are spent
annually on botanical medicines and Ginkgo ranks first among herbal medications [57]. Fifty million G. biloba trees
are grown, especially in China, France, and South Carolina, USA, producing 8000 tons of dried leaves each year to

Fig. 2. Structures of secondary metabolites of G. biloba.

410 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Fig. 2 (continued ).

meet the commercial demand for G. biloba products [57]. Ginkgo is among the most sold medicinal plants and the
annual consumption in 2001 was between 4.5 million pounds and 5.1 million pounds of dried leaves [63]. The use of
G. biloba has been growing at a very rapid rate world wide at 25% per year in the open world commercial market.
Germany, Switzerland and France have respectively 31%, 8% and 5% of the world commercial market. Presently there
are around 142 G. biloba products on the global market and it is estimated that in the coming five years its utilization is
expected to grow threefold. G. biloba is sold in the form of leaf, powder extract, and as a tincture to the pharmaceutical
B. Singh et al. / Fitoterapia 79 (2008) 401–418 411

Fig. 2 (continued ).
412 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Fig. 2 (continued ).

and herbal companies [64]. Here in this report we have attempted to compile the information available on extraction
protocols and on the estimation of terpene trilactones and flavonoids in G. biloba.

13. Secondary metabolites

G. biloba has been well investigated chemically for various classes of constituents. It is reported to contain a number
of secondary metabolites including terpenoids, polyphenols, allyl phenols, organic acids, carbohydrates, fatty acids and
lipids, inorganic salts and amino acids. However terpene trilactones and flavonoids are considered the main bioactive
constituents. Composition and analytical procedures have also been well documented in different pharmacopoeias
[65–67]. These metabolites are listed in Table 1.
Ginkgolides, a terpene trilactones, are compounds with cage like structure. Furukawa (1932) isolated for the first
time ginkgolide A, B, C and M from G. biloba root bark. The ginkgolide A, B, C and J are also reported from the leaves
of G. biloba [64]. It is interesting to note that it was found only in leaves whereas ginkgolide M is found only in the root
 B. Singh et al. / Fitoterapia 79 (2008) 401–418 413

Fig. 2 (continued ).

bark of G. biloba [57]. In addition to the ginkgolides (diterpenes), a bilobalide (sesquiterpene) is also found in G.
biloba. Bilobalide is closely related to the ginkgolides [64]. Structures of ginkgolides K and L were established
recently. Ginkgolides and bilobalide are collectively known as terpene trilactones (TTLs). Terpene trilactones are
reported to be present only in G. biloba. To the best of the knowledge until now terpene trilactones are the only natural
products possessing t-butyl group in their structure [71]. Therefore, terpene trilactones has received more attention
among other constituents of G. biloba. Ginkgolides are the diterpenes having cage like structure consisting of six 5-
membered carbocyclic rings, three lactones and a tetrahydrofuran. Bilobalide is a sesquiterpene and differs by the
absence of tetrahydrofuran ring [64] (Fig. 2).
414 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Fig. 2 (continued ).

Terpene trilactones are the cage like compounds and are extraordinarily stable despite the presence of the multiple
oxygen functionals. They are unaffected by boiling in HNO3 [57]. The crude leaf extract contains significant amount of
more apolar compounds like ginkgolic acid, biflavones and chlorophyll along with active ingredients. Various methods
of extraction have been published for the enrichment of bioactive constituents and are listed in Table 2.
The quantities of terpene trilactones can vary greatly with small changes in parameters such as collection site,
harvest time and plant growth stage [64,76,82]. Analysis of terpene trilactones, including bilobalide and ginkgolides A,
 B. Singh et al. / Fitoterapia 79 (2008) 401–418 415

Table 2
Extraction and sample enrichment methods of terpene trilactones
Methods References
H2O2 followed by extraction with EtOAc. [72]
0.1% Na2HPO4, pH = 8 [73]
Pressurized water extraction [74]
Supercritical CO2 [75]
Polyamide, C18 SPE column [72,76]
Liquid–liquid Extraction with EtOAc–THF [77]
Amberlite XAD-7HP intermediate polar resin [78]
Supercritical fluid extraction [79]
Glass fiber membrane modified with [80]
Octadecyltrichlorosilane
Nanofiltration [81]
H2O2, hydrogen peroxide; EtOAc, Ethylacetate; Na2HPO4, di-sodium hydrogen phosphate; CO2, carbon dioxide; SPE, solid phase extraction; THF,
tetrahydrofuran.

B, C and J has long been difficult because of poor UV absorption due to the lack of good chromophore and relatively
low concentrations in leaves. For the analysis of these compounds, various quantitative methods based on HPLC, GC,
LC–MS, capillary electrophoresis, TLC have been published and details are given here in Table 3.

Table 3
Methods of analysis of terpene trilactones
Methods Solvent system References
Thin-layer chromatography Toluene:acetone (7:3) [69]
C6H6:EtOAc (1:1) [69]
EtOAc:toluene:acetone:hexane (4:3:2:1) [69]
CHCl3:acetone:HCO2H (75:16.5:8.5) [69]
Toluene:EtOAc:acetone:MeOH (10:5:5:0.6) [69]
EtOAc:MeOAc (1:1) [65]
Liquid chromatography RP–HPLC–UV MeOH:iPrOH:H2O (15:10:75) (preparative) [83]
MeOH:ACN:0.01 M phosphate buffer (pH = 5.0) [84]
H2O:MeOH:iPrOH (72.5:17.5:10) [85]
H2O:CAN:THF (10:2:1) RP–HPLC–RI [78]
H2O:MeOH:THF (7:2:1) [72]
MeOH:H2O:H3PO4 (25:75:0.1) [86]
MeOH:H2O (33:67) RP–HPLC–ELSD [87]
NH4OAc:MeOH:iBuOH [88]
MeOH:H2O (gradient) [82]
HPLC–ELSD EtOAc:Pet. ether:MeOH (55:40:5) [89]
LC–ESI–MS THF:iPrOH:H2O (4:8:88) (+ve ion mode) [90]
LC–MS–MS H2O:MeOH (−ve ion mode) [91]
LC–APCI–MS H2O:MeOH (−ve ion mode) [92, 93]
LC–SSI–MS H2O:MeOH [94]
By derivatization benzylation–debenzylation [95]
Gas chromatography GC-FID Conditions: [15,96]
Column DB-5 Injector temp — 280 °C
Detector temp — 300 °C
Oven temp — 200 °C 0–1 min
280 °C 1–5 min
Super critical fluid chromatography CO2 modified with 10%MeOH [97]
H2O:THF:MeOH (68.5:10.5:21)
Oven temperature — 40 °C
Pressure — 280 atm
Capillary electrophoresis Capillary — 60 cm × 75μm, 30 °C [76] [76]
Buffer 25 mM phosphate and 90 mM SDS
NMR Acetone d6:benzene d6 (50:50) [98]
C6H6, cyclohexane; CHCl3, chloroform; HCO2H, formic acid; MeOH, methanol; MeOAc, methyl acetate; iPrOH, isopropyl alcohol; H2O, water;
ACN, acetonitrile; H3PO4, phosphoric acid; iBuOH, isobutyl alcohol; NH4OAc, ammonium acetate.
416 B. Singh et al. / Fitoterapia 79 (2008) 401–418

Table 4
List of extraction and analytical methods of flavonoids
Method References
Molecularly imprinted polymer (MIP) [99]
Leaching with aqueous two phase system [100]
Adsorption separation [101]
Ultrafiltration [102]
RP–HPLC–UV [69,82,66,67]
CE–ED [103]
HPTLC [104,105]
HPLC–PDA [106]
LC–MS–MS [107]
RP–HPLC–UV, Reverse phase–high performance liquid chromatography–ultra violet spectrometer; CE–ED, capillary electrophoresis–
electrochemical detection; HPTLC, high performance thin thin-layer chromatography, PDA, photo diode array; LC–MS, liquid chromatography–
mass spectrometry.

Table 5
List of methods of simultaneous analysis of terpene trilactones and flavonoids
Method References
RP–HPLC–ELSD [108]
RP–HPLC (inline UV–ELSD) [109]
GC–MS [110]
Reverse-flow micellar electrokinetic [111]
Chromatography (RF–MEKC)
Nano ESI–MS [112]
NMR [113]
ELSD, evaporative light scattering detector; GC, gas chromatography; ESI, electrospray ionizations; NMR, nuclear magnetic resonance.

13.1. Flavonoid glycosides

Flavonoids are the polyphenolic, low molecular weight compounds, probably found in all green plants. Currently,
more than 30 genuine flavonoids are known in G. biloba. The great number of different flavonoids is not a result of the
variability of the 2-phenylchromane framework but of the different glycosides found in Ginkgo. Nevertheless, only
glucose and rhamnose can be found as sugar molecules and the variety of mono-, di- and triglycosides in different
binding patterns [63]. The flavonoids comprise of large group of polyphenols and include flavones, flavonol glycosides,
acylated flavonol glycosides, biflavonoids, flavan-3-ols and proanthocyanidins. Of these flavonol glycosides present in
G. biloba leaves are more abundant than the other flavonoids and most of them being derivatives of quercetin,
kaempferol and isorhamnetin. The aglycones themselves occur only in relatively low concentration (Table 4).
A good number of publications are available on quantitative analysis of ginkgoterpenes and flavonoids individually
which is time consuming and uneconomical. Few reports which have appeared on simultaneous quantification are
listed below in Table 5.

Acknowledgements

Authors are grateful to the Director IHBT, for providing necessary facilities. Financial assistance to Pushpinder
Kaur from the National Medicinal Plant Board, DBT, Government of India, is gratefully acknowledged.

References

[1] Kubitzki K. In: Kramer K, Greech PS, editors. The families and genera of vascular plants— pteridophytes and gymnosperms. Berlin: Springer
Verlag; 1990. p. 284.
[2] Hori T. In: Ridge RW, Tulecke W, Del Tredici P, Tremouillaux Guiller, Tobe JH, editors. Ginkgo biloba— a global treasure. From biology to
medicine. Tokyo: Springer; 1997.
 B. Singh et al. / Fitoterapia 79 (2008) 401–418 417

[3] Wang FH, Chen ZK. Acta Bot Sinica 1983;25:199.

[4] Zhou Z, Zhang B. Palaeontographica 1989;B 211:113.
[5] Zhou Z. Rev Palaeobot Palynol 1991;68:203.
[6] Zhou Z. Acta Palaeontologica Sinica 1994;33:131.
[7] Hirase S. J Coll Sci Univ Tokyo 1895;8:307.
[8] Tralau H. Bot Not 1967;120:409.
[9] Del Tredici P. Am J Bot 1992;79:522.
[10] Strasburger E, Noll F, Schenck H, Schimper AFW, Sitte P, Ziegler H, Ehrendorfer F, Bresinsky A. Lehrbuch der Botanik 34, vol. 697. Stuttgart:
Fischer Verlag; 1998. p. 658.
[11] Michel PM, Hosford D. Ginkgo biloba: from “living fossil” to modern therapeutic agent, vol. 1. Ginkgolides: Chemistry, Biology Pharmacology
and Clinical Perspectives; 1988. p. 1.
[12] Del Tredici P. In: Edelin C, editor. The architecture of Ginkgo biloba L. L'Arbre. Biologie et developpement. Naturalia Monspeliensia; 1991.
p. 155.
[13] Wang CW. The forests of China. Maria Moors Cabot Found, Harvard University. Cambridge: Mass; 1961.
[14] Zheng CZ. Preliminary analysis of flora in Tianmu Mountain Reserve. In: Yang F, editor. Comprehensive investigation report on natural
resource of Tianmu Mountain Nature Reserve. Hangzhou: Science and Technology Press; 1992. p. 89.
[15] Beek van TA, Bombardelli E, Morazzoni P, Peterlongo F. Fitoterapia 1998;69:195.
[16] Del Tredici P, Ling H, Yang G. Conserv Biol 1992;6:202.
[17] Zheng CZ. A catalogue of seed–plants in Tianmu Nature Reserve. In: Yang F, editor. Comprehensive investigation report on natural resource
of Tianmu Mountain Nature Reserve. Hangzhou: Science and Technology Press; 1992. p. 94.
[18] Santamour FS, He SA, Ewert TE. J Arboricult 1983;9:170.
[19] Del Tredici P. Biosystems 1989;22:327.
[20] Friedman WE. Am J Bot 1987;74:1797.
[21] Holt BF, Rotwell GW. Am J Bot 1997;84:870.
[22] Hori T. Microscopia 1996;13:184.
[23] Lin JX. Old Ginkgo tree in China. International Dendrological Society Yearbook; 1995. p. 32.
[24] Del Tredici P. Arnoldia 1991;51:2.
[25] Gunkle JE, Thimann KV, Wetmore RH. Am J Bot 1949;36:309.
[26] Del Tredici P. In: Edelin C, editor. The architecture of G. biloba L. L'Arbre, Biologie et Development; 1991. p. 155. Naturalia Monspeliansia
n0 h.s.
[27] Duke JA. Handbook of nuts. Boca Raton: Florida Press; 1989.
[28] Santamour FS, He SA, McArdle AJ. J Arboricult 1983;9:88.
[29] Agronomic aspects of G. biloba leave production. In: Balz JP, editor. Proceedings of The 97 International Seminar on Ginkgo, November 10–12
Beijing China; 1997. p. 101.
[30] Suma TS. Dinosaur of the plant kingdom heritage Amruth December; 2006. p. 35.
[31] Koczka N, Orloci L, Ferenczy AZ, Stefanovits BE, Dimeny J. Int J Horti Sci 2000;16:31.
[32] Son Y. J Plant Nutr 2002;30:93.
[33] Wei G, Wang J, Zhou JC, Zhao HL, Chen XG, Ma LB. J Beijing For Univ 1999;10:96.
[34] Qiao YS, Zhang Z, Fang JG, Wang WB, Xu XL. J Jiangsu For Sci Technol 2001;15:8.
[35] Peng FR, Li J, Huang BL, Zhang JL. J Plant Resour Environ 2001;8:16.
[36] Xie YC, Cao FL, Yao ZG, Wang GB, Zhang WX. J Nanjing For Univ 2000;24:11.
[37] Qian BY, Zhao HL, Chen XG. J Jiangsu For Sci Technol 1997;50:25.
[38] Wang CZ, Shen ZB, Tan WH, Yu Q. J Tropical Subtropical Bot 2000;5:329.
[39] Kang ZX, Lu AH, Jin MZ, Xu YQ, Zhang RZ, Wang XJ, et al. J Fujian coll For 2000;10:321.
[40] Huang YX, Lin SH, Han RZ, Yao YQ. Acta Bot Sinica 1990;32:380.
[41] Sung CH, Wang YN, Sun EJ, Ho LH. Quarterly J Exp For Nat Taiwan Univ 1998;23:269.
[42] Li J, Dong XC, Meng QW, Zhao SJ. J Shandong Agric Univ 1999;7:256.
[43] Leng PS, Su SC, Wang TH, Jiang XN, Wang SS. J Plant Resour Environ 2002;12:1.
[44] Blazich FA. Chemicals and formulations used to promote adventitious rooting. In: Avis TD, Haissag BE, Sankhla N, editors. Adventitious root
formation in cuttings. Portland: Dioscorides Press; 1998. p. 132.
[45] Li GP, Yang LS, Wu JC. South China Fruits 1998;27:44.
[46] Balabuska VK. Bull Glav Bot Sada 1990;156:65.
[47] Cai JB, Shen XK, Zhang RH, Lin DD, Lu YY. J Zhejiang Coll 1998;15:340.
[48] Diao AZ, Xu JS, Ling SG. J Jiangsu For Sci Tech 2000;27:33.
[49] Kling GJ, Meyer MN. Hort Sci 1983;8:352.
[50] Nandi SK, Rikhari HC, Nadeem M, Palni LMS. Physiol Mol Biol Plant 1997;3:15.
[51] Shen XQ, Shen LM, Wang XQ, Ling SG. J Jiangsu For Sci Tech 1999;2:32.
[52] Parkash O, Nagar PK, Brij Lal, Ahuja PS. Timber For Prod 2002;9:47.
[53] Schmid W. L 2001;3:165.
[54] Yu X, Li J. Trans Chin Soc Agricul Engin 2002;18:137.
[55] Wu QK. J Zhejiang For Sci Tech 1999;4:20.
[56] Lee OH, Lee KJ. Kor J Environ Eco 1999;6:34.
418 B. Singh et al. / Fitoterapia 79 (2008) 401–418

[57] Koji N. Bioorg Med Chem 2005;13:4987.

[58] Mahmoud F, Abul H, Onadeko B, Khadadah M, Hainea D, Morgan G. Jpn J Pharmacol 2000;83:241.
[59] Louajri A, Harraga S, Godot V, Toubin G, Kantelip JP. Biol Pharm Bull 2001;24:710.
[60] Bairy KL. J Nat Rem 2002;84:11.
[61] Kennedy DO, Scholey AB, Wesnes KA. Psychopharmacology 2000;151:416.
[62] Polich J, Gloria R. Hum Psychopharm Cli 2001;16:409.
[63] Van Beek TA. Ginkgo biloba– medicinal and aromatic plants – industrial profiles, vol. 12. Harwood academic publishers; 2000.
[64] Medicinal herb production guide, Ginkgo biloba L. http://www.naturalmedicinesofnc.org/Growers%20Guides/ginkgo-gg.pdf.
[65] The United States Pharmacopeia 31, vol. 1; 2008. p. 947.
[66] European Pharmacopoeia 5.0, vol. 2; 2005. p. 1657.
[67] British Pharmacopoeia, vol. 1; 2008. p. 994.
[68] Van Beek TA, Lelyveld GP. Planta Med 1992;58:413.
[69] Van Beek TA. J Chromatogr A 2002;967:21.
[70] Bedir E, Tatli I, Khan RA, Zhao J, Takamatsu S, Walker LA, et al. J Agric Food Chem 2002;50:3150.
[71] Van Beek TA. Bioorg Med Chem 2005;13:5001.
[72] Dirk L, John MB, Koji N. J Nat Prod 2002;65:1501.
[73] Lang Q, Wai CM. Anal Chem 1999;71:2929.
[74] Lang Q, Wai CM. Green Chem 2003;5:415.
[75] Yang C, Xu YR, Yao WX. J Agric Food Chem 2002;50:846.
[76] Hasler A, Meier B. Planta Med 1993;59:632.
[77] Lang Q, Yak HK, Wai CM. Talanta 2001;54:673.
[78] Lai SM, Chen RL, Suen SY. J Liq Chromatogr Rel Tech 2003;26:2941.
[79] Kou Lung C, Chuan CY, Hao CJ, Chiehming JC, Wen YP. J Supercrit Fluids 2002;24:77.
[80] Fang SI, Jen CL, Yi SS. J Sep Sci 2005;28:1211.
[81] Li X, Shiyong W. Desalination 2005;184:305.
[82] Flesh V, Jacques M, Cosson L, Tang B, Petiard V, Balz J. Phytochemistry 1992;31:1941.
[83] Yuping T, Fengchang L. J Liq Chromatogr Rel Tech 2000;23:2897.
[84] Lin CH, Ya LH, Thomas C, Cherng Y. Anal Bioanal Chem 2004;397:445.
[85] Mesbah MK, Khalifa SI, Gindy A, Tawfik KA. Farmaco 2005;60:583.
[86] Wang HF, Ju XR. Se Pu 2000;18:394.
[87] Cui T, Xiuli W, Chunhua Y. J Pharm Biomed Anal 2003;33:811.
[88] Ganzera M, Zhao J, Khan IA. Pharm Bull 2001;49:1170.
[89] Van Beek TA, Gerrit PL. J Nat Prod 1997;60:735.
[90] Pierluigi M, Barbara M, Piergiorgio P. J Mass Spectrom 1999;34:1361.
[91] Sun Y, Li W, Fitzloff FJ, Breemen BR. J Mass Spectrom 2005;40:373.
[92] Mauri P, Simonetti P, Gardana C, Minoggio M, Morazzoni P, Bombardelli E, et al. Rapid Commun Mass Spectrom 2001;15:929.
[93] Jensen AG, Ndjoko K, Wolfender JL, Hostettmann K, Camponovo F, Soldati F. Phytochem Anal 2002;13:31.
[94] Chen E, Ding C, Lindsay CR. Anal Chem 2005;77:2966.
[95] Stanislav J, Shahid M, Koji N. Phytochemistry 2004;65:2897.
[96] Qingyong L, Kwang YH, Chien MW. Talanta 2001;54:673.
[97] Strode III JBT, Taylor LT, Van Beek TA. J Chromatogr A 1996;738:115.
[98] Hae CY, Kyoon CH, Arno H, Bermejo B, Yvonne S, Cornelis E, et al. Chem Pharm Bull 2003;51:158.
[99] Xie J, Zhu L, Luo H, Zhou L, Li C, Xu X. J Chromatogr A 2001;934:1.
[100] Zhang CX, Hu XL. Bull Environ Contam Toxicol 2003;71:662.
[101] Yu FC, Lai SM, Suen SY. Sep Sci Technol 2003;38:1033.
[102] Xu Z, Li L, Wu F, Tan S, Zhang Z. J Membr Sci 2005;255:125.
[103] Cao Y, Chu Q, Fang Y, Ye J. Anal Bioanal Chem 2002;374:294.
[104] Li J, Yu L, Zeng Y. Zhongguo Zhongyao Zazhi 1996;21:106.
[105] Jamshidi A, Adjvadi M, Husain SW. J Planar Chromatogr Mod TLC 2000;13:57.
[106] Dubber MJ, Kanfer I. J Pharm Pharmaceut Sci 2004;7:303.
[107] Dubber MJ, Sewram V, Mshicileli N, Shephard GS, Kanfer I. J Pharm Biomed Anal 2005;37:723.
[108] Li W, Fitzloff JF. J Pharm Biomed Anal 2002;30:67.
[109] Gray DE, Messer D, Porter A, Ferguson S, Harris RK, Clark AP, et al. J AOAC Int 2005;88:1613.
[110] Fengxia D, William ZS. J Chromatogr A 2003;986:121.
[111] Dubber MJ, Kanfer I. J Chromatogr A 2006;1122:266.
[112] Liu C, Mandal R, Li XF. Analyst 2005;130:325.
[113] Li CY, Lin CH, Wu CC, Lee KH, Wu TS. J Agric Food Chem 2004;52:3721.

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