Leaf flavonoids
The leaf flavonoids of Geranium are typical of the Geraniaceae and related dicotyledon-ous families. They are predominantly flavonols and the commonly occurring quercetin (6) is universally present. In a survey of acid-hydrolysed leaf tissue of 78 species, Bate-Smith (1972) reported that quercetin is generally accompanied by the lower homologue kaempferol (7) (in 93 per cent of the sample) and by the higher homologue myricetin (8) (Figure 4.2) (in 13 per cent of the sample). Variation in this basic flavonol pattern is to some extent correlated with the geography of the genus. A primitive pattern, including myricetin, predominates in plants from the central Eurasian area, while an advanced pattern, represented by high concentrations of kaempferol, is present in Mediterranean and American species.
Although it is apparent that the above three flavonols occur in Geranium in glyco-sidic combination, relatively little is known of the glycoglyco-sidic pattern of most species.
However, there is one report of an high performance liquid chromatography (HPLC) survey of Geranium leaves by Okuda et al. (1980). These authors found that quercetin occurs regularly in the genus as the 3-galactoside, called hyperin (9) Figure 4.2. This was detected in direct alcoholic leaf extracts of 12 out of 15 species surveyed (Table 4.1). The content of hyperin varied from 0.03 to 1.6 per cent dry weight, with an average value of 0.43 per cent.
One species apparently lacking in quercetin 3-galactoside is the Japanese G. thun-bergii (Table 4.1). Instead, leaves of this plant contain either kaempferol 3-rhamnoside
( ) Germacrone3 ( ) -Elemene4 α
( ) -Curcumene5 α
( ) Geraniol1 ( ) -Citronellol2 α
Figure 4.1 Essential oil components of Geranium.
22 Jeffrey Harborne and Christine Williams
or a mixture of kaempferol 3-arabinoside-7-rhamnoside and kaempferol 3,7-dirhamno-side (Kawamura et al., 1995). There are thus two chemical races in flavonoid content, but the occurrences of the two races do not correspond with any other variable features of this plant species.
( ) Quercetin6 ( ) Kaempferol7
( ) Myricetin8
( ) Hyperin9
( ) Kaempferol 3-rutinoside 4
10
′-glucoside
(11) Quercetin 3-glucuronide (12) Vitexin
Figure 4.2 Leaf flavonoids of Geranium.
Phytochemistry of the genus Geranium 23
Herb Robert, G. robertianum, a plant of medicinal interest, has been examined in some detail for its flavonol glycosides. Aerial parts yield a mixture of six mono-glucosides: kaempferol and quercetin 3-galactoside, quercetin 3-glucoside, quercetin 4-glucoside, quercetin 7-glucoside and quercetin 7-rhamnoside. Accompanying these monoglycosides are seven 3-diglycosides. Only four of the seven were fully characterised as the 3-rutinosides and 3-rhamnosylgalactosides of kaempferol and quercetin (Kartnig and Bucar-Stachel, 1991). Whether Herb Robert varies in its flavonol glycoside content is not yet clear, but it may be noted that Okuda et al.
(1980) failed to find the quercetin 3-galactoside reported by Kartnig and Bucar-Stachel, 1991) in their particular sample (Table 4.1).
A further four flavonol glycosides, not described so far, have been characterised vari-ously in five Geranium species native to Egypt (Table 4.2). The most distinctive is kaempferol 3-rutinoside-4-glucoside (10), recorded in G. yemense and G. rotundifolium.
The presence of quercetin 3-glucuronide (11) in G. dissectum is noteworthy (Saleh et al., 1987) Figure 4.2. The apparent absence of quercetin 3-galactoside from these five Egyptian species should also be noted.
Table 4.1 Geraniin and hyperiin content of the dry leaves of Geranium species
Species Geranin (%) Hyperin (%) Month of collection
G. eriostemon Fisch. var reinii Maxim 7.5 0.15 July
G. erianthum DC. 7.6 0.13 August
G. soboliferum Komar. 6.8 0.16 October
G. krameri Franch. et Savat. 6.8 0.19 October
G. yoshinoi Makino 9.8 0.55 September
G. yesoense Franch. et Savat. 12 0.18 August
G. yesoense Franch. et Savat. var. nipponicum Nakai 12 0.09 October
G. shikokianum Matsum. 6.0 0.59 August
G. sibiricum L. var. glabrius Ohwi 8.1 – October
G. thunbergii Sieb. et Zucc. 12 – August
G. wilfordii Maxim. 9.5 0.21 September
G. wilfordii Maxim. var. hastatum Hara* 0.50 0.03 September
G. tripartitum R. Knuth 12 1.3 September
G. robertianum L. 9.8 – September
G. carolianum L. 11 1.6 May
Note
* Fresh aerial tissue.
Table 4.2 Flavonol glycosides of five Egyptian species of Geranium
Species Major flavonols present Source: Data from Saleh et al. (1983).
24 Jeffrey Harborne and Christine Williams
Other classes of flavonoid such as glycosylflavones are also present in Geranium accord-ing to Bate-Smith (1977) but they have not in general been investigated further. They have been found to dominate in the case of Geranium phaeum. Here, the flavonol glycosides based on quercetin are minor components, compared to the five glycosyl flavones: vitexin (12) (Figure 4.2), isovitexin, orientin, iso-orientin and vicenin (Boutard and Lebreton, 1975).
Floral flavonoids
Most Geranium species have attractive flowers, with colours ranging from blue, purple and red to pink and white. Anthocyanins, together with co-occurring flavonol glyco-sides are presumably responsible for those flower colours, but surprising little work has been carried out on these pigments in the genus. A major study has, however, been devoted to the bluish-purple or bluish-magenta flowers in G. pratense and G. sanguinea and in the cultivar ‘Johnsons Blue’, a hybrid derived from G. himalayenseG. pratense.
The same major anthocyanin is present in the flower of all three plants. It is malvidin 3,5-diglucoside (13), with a labile acetyl substituent at the 6-position of the glucose residue attached to the 5-hydroxyl. Thus, it is malvidin 3-glucoside-5-(6-acetylgluco-side) (Markham et al., 1997).
In the petals of two of these plant species, the anthocyanin co-occurs with four flavonol glycosides, namely the 3-glucosides and 3-sophorosides of kaempferol and myricetin.
Colour tests in vitro indicate that kaempferol 3-sophoroside (14) is the most important co-pigment, shifting the mauve colour of the malvidin glycoside (13) Figure 4.3 towards the blue region. An additional and unusual feature of flower colour production in these
(13) Malvidin 3,5-diglucoside
(14) Kaempferol 3-sophoroside (15) Quercetin 3,7,3 ,4 -tetramethyl ether′ ′ Figure 4.3 Floral and exudate flavonoids of Geranium.
Phytochemistry of the genus Geranium 25
Geranium petals is the presence of a cell sap pH of between 6.6 and 6.8, instead of the more usual pH at 5.6. This appears to be a very rare feature in nature but is important for the full colour intensity observed in these flowers (Markham et al., 1997).
Exudate flavonoids
Geranium species regularly have glandular hairs or trichomes on the upper leaf surface.
The chemical constituents of these trichomes can be examined separately from the internal leaf components by brief rinsing of leaf surfaces in a solvent such as acetone.
Besides the terpenoids and hydrocarbons that are commonly present at the surface, leaf washes occasionally yield mixtures of lipophilic flavonoids, usually flavonol methyl ethers. Such compounds have been obtained from leaf surfaces of G. macrorrhizum and G. lucidum. The structures present are almost identical in both plants and consist of some 14 kaempferol, quercetin or myricetin methyl ethers (Table 4.3) (Ivancheva and Wollenweber, 1989).
The above report supercedes an earlier paper by Ognyanov and Ivantcheva (1972) in which a so-called novel flavonol, 3,5,7,2,4,6-hexahydroxyflavone and kaempferol 3-methyl ether were reported at the surface of G. macrorrhizum. Re-examination of the evidence for the ‘new’ flavonol suggests that this was a mistaken identification of a known flavonol. It may also be noted that quercetin 3,7,3,4-tetramethyl ether (15) was independently identified as a major lipophilic constituent of G. macrorrhizum by Nakashima et al. (1973). This compound (15) Figure 4.3 crystallised out in 0.4 per cent yield from the essential oil of this plant.
TANNINS
Hydrolysable tannins
The major hydrolysable tannin of Geranium is the compound geraniin (16) Figure 4.4, first crystallised from leaf extracts of G. thunbergii. This plant has been used extensively in folk medicine in Japan. A boiling water extract of G. thunbergii has been taken by numerous people over many years as an antidiarrhetic and for controlling intestinal function (Okuda et al., 1992). Geraniin makes up more than
Table 4.3 Exudate flavonol methyl ethers of G. macrorrhizum and G. lucidum
Kaempferol Quercetin Myricetin
3-methyl ether – –
4-methyl ether* – –
3,7-dimethyl ether 3,7-dimethyl ether† –
3,4-dimethyl ether 3,3-dimethyl ether –
7,4-dimethyl ether 7,3-dimethyl ether –
3,7,4-trimethyl ether 3,7,3-trimethyl ether 7,3,4-trimethyl ether 7,3,4-trimethyl ether
3,7,3,4-tetramethyl ether 3,7,3,4-tetramethyl ether Notes
*Lacking in G. macrorrhizum;
†lacking in G. lucidum; otherwise all compounds present in both species.
26 Jeffrey Harborne and Christine Williams
10 per cent of the weight of the dried leaf. It forms yellow crystals and, remarkably for an ellagitannin, completely lacks the astringency normally associated with plant tannins.
In its chemical structure, geraniin (16) is based on a molecule of glucose, which is disubstituted in the 3,6- and 2,4-positions by two hexahydroxygallic acid residues.
Additionally, there is a galloyl ester group linked at C-1 of the sugar. Biosynthetically, geraniin is derived from gallic acid (17) via pentagalloylglucose as an intermediate (Haslam, 1989).
Geraniin would appear to be the characteristic hydrolysable tannin of the genus Geranium, since it has been detected by HPLC in every one of the 15 species surveyed (Okuda et al., 1980) (Table 4.2). The richest source is G. thunbergii, with over 12 per cent of its dry weight made up by geraniin. Other species range from 0.5 per cent up to
Figure 4.4 Tannins and their precursors in Geranium.
(16) Geraniin (17) Gallic acid
(18) Ellagic acid (19) Procyanidin
(20) (+)-Catechin (21) (–)-Epicatechin
Phytochemistry of the genus Geranium 27 12 per cent with an average of about 10 per cent dry weight. By comparison with the leaves, the stems of these plants have only 1–2 per cent dry weight.
In an earlier survey based on a colour test for ellagitannin developed with nitrous acid in the absence of oxygen, Bate-Smith (1972) found that some 30 species, representing 16 sections of the genus, contain ellagitannin (presumably geraniin) in amounts ranging from 1.3 to 20 per cent dry weight. It seems likely that most, if not all, known Geranium species contain geraniin or an ellagitannin of similar structure. The same compound, incidentally extends its distribution to other members of the Geraniaceae and to other families in the order Geraniales. It has been detected, for example, in 28 plant species of the Euphorbiaceae. It also occurs in the cocaine-containing Erythroxylon coca (Erythroxylaceae). Geraniin is however apparently absent from the closely related genus, Pelargonium, in spite of the fact that ellagitannins are also abundantly present in Pelargonium species (Okuda et al., 1980) (see Chapter 11).
Co-occurring with geraniin in the leaves of Geranium species is the related structure, ellagic acid (18). Bate-Smith (1962) records ellagic acid in the leaves of 4 out of 6 surveyed, in G. meeboldii, G. phaeum, G. robertianum and G. sylvaticum. A richer source of ellagic acid, is however the plant root and rhizome. Here, it has been recorded in some 61 species (Hegnauer, 1966). Moreover, so much is present in the roots that ellag-ic acid can be isolated in crystalline form. Gallellag-ic acid (3,4,5-trihydroxybenzoellag-ic), whellag-ich is a presumed precursor of ellagic, has also been recorded regularly in the leaf (Bate-Smith, 1962) and the root (Hegnauer, 1966). Gallic acid has been isolated, for example, from roots of G. maculatum, G. nepalensis and G. pratense.
Condensed tannins
Plants of Geranium contain both hydrolysable and condensed tannin, but the distribu-tion in the different organs varies considerably. The main occurrence of condensed tan-nin (or proanthocyanidin) is in the root stock, according to Bate-Smith (1973). There is apparently a suppression of this chemical character in the leaves, where ellagitannins dominate (see above). Only a handful of 60 species of Geranium surveyed have signific-ant amounts of prosignific-anthocyanidins in the leaves. These are: G. polysignific-anthes (Eurasia), G. platypetalum (Armenia), G. renardii (Caucasus), G. sinense (China), G. incanum (South Africa) and G. lindenianum (Venezuela).
The proanthocyanidins in Geranium are based on either procyanidin or prodel-phinidin or a mixture of the two. No detailed chemistry has yet been carried out on the condensed tannins of these plants. However, it is likely that the procyanidins are of a common type (e.g. (19)), since the two procyanidin precursors, ()-catechin (20) and ()-epicatechin (21) Figure 4.4 have been detected in roots of G. pratense and G. palustre (Hegnauer, 1966).
The content of procyanidin in fresh rhizomes, as compared to the amount of ellagitannin, has been shown to be about the same in G. sylvaticum. By contrast, there is only one-seventh the amount of procyanidin, compared to six-sevenths ellagitannin, in G. pratense (Hegnauer, 1966). The high content of tannins in the roots has meant that Geranium species have been employed in the past as good sources of tanning material for the leather industry. At least two species, G. nepalense and G. wallichianum have been used in this way.
28 Jeffrey Harborne and Christine Williams MISCELLANEOUS CONSTITUENTS
Aerial parts of G. richardsonii and G. viscosissimum characteristically accumulate the organic acid, tartaric acid. This acid occurs regularly in members of the Geraniaceae (Stafford, 1961). However, it is not always present in every Geranium species. Thus, G. robertianum and G. sanguineum, when analysed, showed the presence of malic and citric acids, but tartaric acid was missing (Kinzel, 1964).
CONCLUSION
The chemistry of Geranium, as is clear from the above summary, is dominated by phen-olic constituents. Not only are these two classes of plant tannin – proanthocyanidin and ellagitannin – widely distributed in both aerial and underground tissues. But also there are a wealth of monomeric flavonoids, variously present in the leaf and petal. We are only just beginning to appreciate the chemical complexity bound up in the phenolic fraction of Geranium plants and much further work is required to establish the precise range of structures that are present in any given species.
Since both the monomeric flavonoids (Rice-Evans, 2000) and the various tannins (Haslam, 1989) have long been considered to be active components of many medicinal plants, it is likely that the useful properties in terms of human medicine associated with Geranium plants may be due to the type and quantity of particular phenolics present.
However, there is no doubt that much further biological experimentation is required before we can adequately explain the curative properties of these plants.
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Bate-Smith, E.C. (1977) Chemotaxonomy of Geranium. Bot. J. Linn. Soc., 67, 347–359.
Boutard, B. and Lebreton, P. (1975) The presence of C-glycoflavones in Geranium phaeum. Plantes Med. Phytotherapi., 9, 289–296.
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Kartnig, T. and Bucar-Stachel, J. (1991) Flavonoide aus den oberirdischen Teilen von Geranium robertianum. Planta Med., 57, 292–293.
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Markham, K.R., Mitchell, K.A. and Boase, M.R. (1997) Malvidin 3-glucoside-5-(6-acetylgluco-side) and its colour in Geranium ‘Johnson’s Blue’ and other ‘Blue’ Geraniums. Phytochemistry, 45, 417–423.
Nakashima, R., Yoshikawa, M. and Matsuura, T. (1973) Quercetin 3,7,3,4-tetramethyl ether from Geranium macrorrhizum. Phytochemistry, 12, 1502.
Ognyanov, I.V. and Ivantcheva, S. (1972) A new hexahydroxyflavone and isokaempferide in Geranium macorrhizum. Dokl. Bulg. Akad. Nauk., 25, 1057–1059.
Ognyanov, I., Ivanov, D., Herout, V., Hovak, M., Pliva, J. and Sorm, F. (1958) Structure of ger-macrone. Chem. Listy, 52, 1163–1173.
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Okuda, T., Yoshida, T. and Hatano, T. (1992) Pharmacologically active tannins from medicinal plants. In: Plant Polyphenolsi (Hemingway, R.W. and Laks, P.E., eds), pp. 539–569, Plenum Press, New York.
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Phytochemistry of the genus Geranium 29