Phytochemistry Reviews

, Volume 10, Issue 4, pp 459–469 | Cite as

Biosynthesis of saponins in the genus Medicago

Article

Abstracts

Saponins from Medicago species are glycosidic compounds with an aglycone moiety formed through the enzymatic cyclization of 2,3-oxidosqualene by the β-amyrin cyclase. All the saponins from Medicago genus possess the triterpenic pentacyclic nucleus belonging to the class of β-amyrin. The so formed β-amyrin skeleton can be further modified by oxidative reactions, mediated by cytochromes belonging to the class of cytochrome P450, to give different saponin compounds, characterized by the presence of hydroxyl or carboxyl groups located in specific positions of the triterpenic skeleton. Based on the position and the oxidation degree of the substituents, it is possible to distinguish two groups of saponins (sapogenins) in Medicago spp: (1) sapogenins possessing an OH group on C-24 (soyasapogenols A, B and E) without any substituent at the C-28 atom, and (2) sapogenins possessing the COOH group at C-28 that are associated with different oxidation degrees (zero, OH, CHO, COOH) at C-23. These results seem to indicate that the oxidation at C-24 and the presence of the COOH group at C-28 are mutually exclusive. The subdivision in the aglycone moiety is reflected also in the sugar moiety, operated by glycosyltranferases, as the saponins of the two groups differ for the position and the nature of the sugar chains. Based on these findings, new considerations on the biosynthesis of saponins in the genus Medicago can be drawn and a biosynthetic scheme is proposed.

Keywords

Triterpenes Sapogenins Biosynthesis Cytochrome P450 Medicagenic acid 

Introduction

In the recent years the number of studies (Hostettmann and Marston 1995; Dewick 2001; Haralampidis et al. 2002) on the biosynthesis of saponins has increased, due to the particular chemical, physical and physiological characteristics of these compounds making them important starting material for industrial applications. The biological properties of these plant secondary metabolites have been extensively investigated (Milgate and Roberts 1995; Lacaille-Dubois and Wagner 2000; Francis et al. 2002; Sprag et al. 2004). Within the genus Medicago (Leguminosae or Fabaceae) saponins isolated from the different species have been reported to possess antimicrobial, cytotoxic and insecticidal activities (Tava and Avato 2006). More recently, their nematicidal potential (Argentieri et al. 2008; D’Addabbo et al. 2009) has awakened interest on these substances as important bio-active compounds to be employed also in the agro-industry.

Saponins from the genus Medicago are triterpenic pentacyclic glycosides (Tava and Avato 2006) and their aglycone moieties belong to the class of oleanane, characterized by the presence of the Δ12(13) double bond and the E ring joined downwards to ring D (in α position at C-17 and C-18). All the sapogenins possess an oxygen atom at the 3β position, that originates from the precursor oxidosqualene (Fig. 1).
Fig. 1

General structure of Medicago sapogenins. R=H:β-amyrin

Studies on the biosynthesis of those saponins in Medicago spp. based on labelled precursors, such as 14C-acetate (Nowacki et al. 1976) and 2-14C-mevalonate (Peri et al. 1979), showed that the highest rate of radioactivity incorporated in M. sativa seedlings was associated with medicagenic acid.

A genomic approach in the model species M. truncatula allowed the identification of three enzymes involved in the formation of the triterpene aglycone: squalene synthase, squalene epoxidase and β-amyrin synthase (Suzuki et al. 2002; Iturbe-Ormaetxe et al. 2003) confirming that, as in other plant species, squalene is the starting substrate for the biosynthesis of the pentacyclic triterpenes also in the genus Medicago.

The synthesis of saponins in Medicago was furthermore investigated by means of chemical methods to confirm the chemical structures of newly isolated saponins, or to obtain new compounds with different biological activities. Thus, synthetic saponins were obtained by reaction of activated sapogenins from M. sativa saponins and sugars (Morris and Tankersley 1963; Levy et al. 1989; Zehavi et al. 1993).

In the present paper chemical and biological information on saponins obtained from different Medicago spp. are examined in order to (i) define the pattern of the modifications (nature and position of the substituents) in the aglycone moiety and propose, on this basis, a biosynthetic pathway for sapogenols in the genus Medicago and (ii) review the position and the nature of the sugar moiety and their relationships with the aglycone nature.

Aglycones

As reported by several authors (Xu et al. 2004; Phillips et al. 2006; Vincken et al. 2007), the triterpenic pentacyclic nucleus originates from the cyclization of squalene or oxidosqualene involving several steps, catalysed by the enzyme β-amyrin synthase.

As a general mechanism, oxidosqualene is activated by protonation. A proton-initiated cyclization of the ‘chair–chair–chair’ conformation results in the formation of the tetracyclic dammarenyl carbocation. This cyclization is generally drawn as concerted, although a series of step-by-step, ring-by-ring formations is also possible. As shown in Fig. 2, the dammarenyl cation, after D ring expansion via C-16 migration, forms the baccharenyl cation that, after 18β E ring expansion, gives the lupenyl cation, that in turn, after E ring expansion via C-21 migration, forms the germanicenyl cation, a triterpenic structure with a six membered ring alone.
Fig. 2

Cyclization of oxidosqualene to β-amyrin

By a series of 1,2-H shifts of the germanicenyl cation, the oleanyl cation is formed. This cation then loses a proton to give the corresponding Δ12(13) unsaturated compound β-amyrin (Xu et al. 2004; Vincken et al. 2007). As reported above, all the saponins (sapogenins) from Medicago until now isolated possess the OH group at the 3β position, presumably originated from the oxidosqualene. To our knowledge no other saponin skeleton, such as for example cholesterol leading to steroid type saponins, has been isolated from this genus (Tava and Avato 2006).

A further evidence that β-amyrin is the first compound formed in the biosynthesis of Medicago saponins comes from transgenic approaches. The functional characterization of squalene synthase, squalene epoxidase and β-amyrin synthase genes was accomplished in M. truncatula and the genes were expressed in yeast to obtain β-amyrin (Suzuki et al. 2002). A cDNA encoding a β-amyrin synthase from Aster sedifolius was expressed in yeast to produce β-amyrin (Cammareri et al. 2008). When this gene was used to transform M. truncatula, the obtained transgenic plants showed a significant increase of total saponins, expressed as sapogenins, and in particular of bayogenin, medicagenic and zanhic acid (Confalonieri et al. 2009). This study confirmed that the increased production of β-amyrin can promote the biosynthesis of the other sapogenins, derived from oxidative reactions on its triterpenic pentacyclic nucleus.

It is known that oxidative reactions modifying the β-amyrin skeleton are mediated by cytochromes belonging to the class of cytochrome P450 (Hayashi et al. 1993). Cytochrome P450 consists of a group of monooxygenases belonging to a gene superfamily and plays important roles in the metabolism of a series of physiologically important compounds. Most of the known P450 families in Dicot species exist in the model legume plant M. truncatula (Li et al. 2007); however, no gene involved in the oxidative reactions of the β-amyrin skeleton has yet been isolated from this species.

Oxidative modification of β-amyrin gives origin to the different sapogenins, characterized by the presence of hydroxyl or carboxyl groups in particular and specific positions of the triterpenic pentacyclic structure. In Medicago the regio- and stereospecific oxidation of the oleanane triterpenic skeleton originates several compounds that are reported in Fig. 3.
Fig. 3

Sapogenins detected in Medicago species

A summary of the various substituents and their position in the saponins characterized in the Medicago species until now studied is reported in Table 1.
Table 1

Substituents attached to carbon atoms of the oleanane skeleton of Medicago aglycones

 

23

(4α)

24

(4β)

28

(17β)

16α

21β

22β

30

(20β)

Soyasapogenol B

OH

OH

Soyasapogenol E

OH

=O

Soyasapogenol A

OH

OH

OH

Oleanolic acid

COOH

2β-Hydroxy oleanolic acid

COOH

OH

Queretaroic acid

COOH

OH

2β-Hydroxy queretaroic acid

COOH

OH

OH

Hederagenin

OH

COOH

Caulophyllogenin

OH

COOH

OH

Bayogenin

OH

COOH

OH

2β,3β-Dihydroxy-23-oxo olean-12-en-28-oic acid

CHO

COOH

OH

Medicagenic acid

COOH

COOH

OH

Zanhic acid

COOH

COOH

OH

OH

For number position see Fig. 2

Based on the position and the oxidation degree of these functional groups, a series of considerations can be drawn. A first classification of sapogenins can be assumed considering C-23, C-24 and C-28 positions (substitutions on 4α, 4β and 17β): sapogenins with an OH group on C-24 (–CH2OH substituted in 4α position) such as soyasapogenols A, B and E, do not show any kind of oxidation at C-28. The C-28 (17β substitution) shows the presence of a COOH group that can be associated to a different oxidation degree (zero, OH, CHO, COOH) of C-23 (4α substitution) only.

These evidences suggest that the oxidation at C-24 is not compatible with the presence of any kind of substituent at C-28, while the COOH group at C-28 (17β substitution) is compatible with the presence of different substituents (from no oxidation, CH3, to the maximum oxidation degree, COOH) only at C-23.

These differences seem to be correlated with one of the most peculiar biological property of the corresponding saponins, their haemolytic activity. In fact soyasapogenol saponins do not show any kind of haemolysis, while all the other saponins have haemolytic properties, although at a different degree (Jurzysta and Waller 1996; Oleszek 1996).

The different sapogenins are characterised by the presence of other OH substituents in specific positions of the triterpenic nucleus. All the sapogenols (non haemolytic saponins) show the presence of an OH group at β-C-22 position, with soyasapogenol E derived from photooxidation of soyasapogenol B (Kitagawa et al. 1988). An additional OH group is present at β-C-21 position in soyasapogenol A. These sapogenins are common constituents of the Leguminosae family (Hostettmann and Marston 1995) and have been isolated from almost all the studied Medicago species (Tava and Avato 2006).

As for the C-28 carboxylic group containing sapogenins (haemolytic saponins), the C-23 position, as reported above, can be oxidated to a different degree giving the corresponding OH, CHO and COOH derivatives. Other OH groups can be present on the triterpenic skeleton, alone or in combination, in specific and selected positions. A most favourable position from a mechanistic point of view is the 2β, giving the 2β-OH derivatives, bayogenin, medicagenic and zanhic acids, representing the most abundant structural types of sapogenins isolated until now from the analyzed Medicago species (Tava and Avato 2006).

The 16α derivatives were detected as caulophyllogenin in M. polymorpha (Kinjo et al. 1994) and as zanhic acid in M. arborea (Tava et al. 2005), M. sativa (Oleszek et al. 1992; Bialy et al. 1999) and M. truncatula (Huhman et al. 2005; Kapusta et al. 2005). More recently, two 30-OH substituted sapogenins, named queretaroic acid and 2β-hydroxy queretaroic acid, were characterized in M. arabica (Tava et al. 2009).

Taking into account the most favourable positional substitutions on the triterpenic nucleus, it can be noticed that the two mutually exclusive substitutions at C-24 and C-28 (Table 1, Fig. 4) in the Medicago saponins are located on the same side of the molecule suggesting that different cytochromes P450 might be responsible for the specific oxidation in A ring (C-24) or E ring (C-28).
Fig. 4

Position of substituents on the triterpenic pentacyclic nucleus of β-amyrin

The 24β oxidation is only correlated with the presence of an alcoholic group (–CH2OH) and it is associated with 22β oxidation to give the soyasapogenol B, and eventually also in 21β to give soyasapogenol A, as shown by dotted arrows in Fig. 4. Carboxylation at C-28 can also be concerted with a substitution on both the left part of the molecule (ring A, positions 2 (and 4α) and the right part (ring D, position 16α and ring E, position 20β), as shown by non-dotted arrows in Fig. 4.

A very interesting structural feature of saponins from Medicago, is the presence of an aldehyde group at the C-23 position in 2β,3β-dihydroxy-23-oxo-olean-12-en-28-oic acid (Fig. 1), a new saponin aglycone isolated for the first time from M. arborea (Tava et al. 2005) and M. hybrida (Bialy et al. 2006). Chemical characterization of this metabolite gave new insights to understand saponin biosynthesis in Medicago. It might in fact represent an interesting biosynthetic intermediate in the oxidative steps that lead from a methyl group to the corresponding carboxylic acid. That is, if we consider the following sapogenins found in the genus Medicago—2β-hydroxyoleanolic acid, bayogenin, 2β,3β-dihydroxy-23-oxo-olean-12-en-28-oic acid and medicagenic acid–all the oxidative products at C-23 can be observed. The above sapogenins all possess the same stereochemistry (2β,3β) in the hydroxylated triterpene carbons with the different functional groups at the C-23 position. The presence of an aldehyde group in 2β,3β-dihydroxy-23-oxo-olean-12-en-28-oic acid, identified for the first time in Medicago, indicates a possible biosynthetic pathway for the sapogenins of this genus. Accordingly, medicagenic acid may originate from bayogenin by subsequent oxidative enzymatic steps involving the formation of 2β,3β-dihydroxy-23-oxo-olean-12-en-28-oic acid while bayogenin may originate by a selective oxidative demethylation at C-23 from 2β-hydroxyoleanolic acid. In a similar way, the two 16α-hydroxy triterpenes found in this genus, caulophyllogenin and zanhic acid probably originate by enzymatic oxidation of hederagenin and medicagenic acid, respectively. Based on these considerations, a possible biosynthetic pathway can be hypothesized, as shown in Figs. 5, 6.
Fig. 5

Proposed biosynthetic pathway for soyasapogenols in Medicago ssp. A series of step-by-step oxidations from β-amyrin are assumed. See the text for details

Fig. 6

Proposed biosynthetic pathway for sapogenols in Medicago ssp. A series of step-by-step oxidations from β-amyrin are assumed. See the text for details

The mechanism of formation of soyasapogenol B (Fig. 5), is supported by the identification in soyabean (Glycine max) of a cytochrome P450 gene CYP93E1 encoding for a 24-hydroxylase using as a substrate both β-amyrin and sophoradiol (Shibuya et al. 2006).

Sugar chains

The saponins isolated from the different Medicago species contain several sugars or sugar chains that are generally linked at the 3-O position of the triterpene aglycone, giving the corresponding monodesmosides, and additionally at the 28-O position, giving the corresponding bidesmosides. Sugars at the 3-O position are linked by an ethereal linkage, while sugars at the 28-O position are linked by an ester linkage to the triterpene structure. A tridesmoside saponin (extra sugar at the 23-O position) has only been identified in M. sativa (Oleszek et al. 1992) and M. truncatula (Huhman et al. 2005).

A complete list of saponins until now detected in Medicago species has been reported in Tava and Avato (2006).

M. arabica leaves are characterized by the presence of short sugar chain saponins, including mono and bidesmosides of 2β-hydroxyoleanolic acid, hederagenin and bayogenin (Bialy et al. 2004; Tava et al. 2009). M. arborea leaves produce saponins containing up to seven sugars, identified as mono- and bidesmosides of medicagenic and zanhic acid (Tava et al. 2005). Saponins from M. hybrida roots are characterized by the presence of short sugar chain bidesmosides of hederagenin and medicagenic acid (Bialy et al. 2006). M. lupulina leaves contain mono and disaccharide saponins of hederagenin and medicagenic acid (Oleszek et al. 1988), while saponins from the leaves of M. polymorpha predominantly consist of short sugar chain bidesmosides of hederagenin and caulophyllogenin (Kinjo et al. 1994).

Saponins from the roots and the aerial parts of M. sativa are instead a complex mixture of both short and long sugar chains of mono- and bidesmosidic compounds with hederagenin, medicagenic acid, zanhic acid and soyasapogenols as the most representative aglycones (Massiot et al. 1991; Timbekova et al. 1996; Bialy et al. 1999). M. truncatula saponins from both roots and aerial parts consist of long sugar chain bidesmosides of medicagenic and zanhic acid (Huhman et al. 2005; Kapusta et al. 2005). Branched sugar chain saponins were also identified in this species, as in M. arborea and M. sativa.

The nature of the saccharide units, their position in the molecule and the similarity of the sugar chains in saponins from the different Medicago species, suggest enzymatic specificity for the sugar and regiospecificity for the sugar position. Sugar or sugar chains are inserted on the triterpenic nucleus through a series of reactions catalyzed by a group of glycosyltransferases (GTs). Plant GTs are members of a multigene superfamily that can typically transfer single or multiple activated sugars from nucleotide sugar donors to a wide range of metabolites (Kalinowska et al. 2005; Wang and Hou 2009).

High specificity for the sugar was demonstrated for two GTs involved in the biosynthesis of saponins in M. truncatula (Achnine et al. 2005) and one GT in soybean (Kurosawa et al. 2002).

Monodesmosides are glycosylated at the 3-O position: this glycosylation seems to be more related to the involved genin, as summarized in Table 2 in which only the most representative sapogenins of the Medicago species are reported.
Table 2

Sugars attached at the 3-O position of the triterpene skeleton of Medicago aglycones

Hederagenin

α-l-Ara

M. arabica, M. hybrida

ß-d-Glc(1 → 2)α-L-Ara

M. sativa

α-l-Rha(1 → 2)α-l-Ara

M. polymorpha

β-d-Glc or β-d-GluA

M. hybrida

Medicagenic acid

Zanhic acid

β-d-Glc or β-d-GluA

β-d-Glc(1 → 2)β-d-Glc

β-d-Glc(1 → 2)β-d-Glc(1 → 2)β-d-Glc

M. arborea, M. hybrida, M. lupulina, M. sativa

β-d-Glc(1 → 3)β-d-Glc

M. truncatula

Soyasapogenols

α-l-Rha(1 → 2)β-d-Gal(1 → 2)β-d-GluA

 

Hederagenin often contains an α-l-arabinopyranose unit as the first sugar in its 3-O position. Alternatively a β-d-glucopyranose, or the corresponding uronic derivative, are present as, for example, in root saponins isolated from M. hybrida (Bialy et al. 2006). The second monosaccharide unit linked at the C-2 position of α-l-arabinopyranose can be α-l-rhamnopyranose, as in M. polymorpha (Kinjo et al. 1994), or β-d-glucopyranose as in M. arabica (Tava et al. 2009) and M. sativa (Massiot et al. 1988; Timbekova et al. 1996).

By contrast, in all the studied species of Medicago, saponins of medicagenic and zanhic acids are always characterized by the presence of β-d-glucopyranose or β-d-glucuronopyranose units as the first sugar in the 3-O position. The second monosaccharide is β-d-glucopyranose, linked predominantly at the C-2 position, as in M. arborea (Tava et al. 2005), M. hybrida (Bialy et al. 2006) and in M. sativa (Massiot et al. 1988; Oleszek et al. 1992; Bialy et al. 1999). Only in M. truncatula the second β-d-glucopyranose moiety is linked at the C-3 position (Kapusta et al. 2005), and this can suggest the presence of a specific glycosyltransferase in this species.

Trisaccharides are numerically most abundant in M. sativa, and are predominantly constituted of 3-O-[β-d-glucopyranosyl(1 → 2)-β-d-glucopyranosyl(1 → 2)-β-d-glucopyranosyl] derivatives (Oleszek et al. 1992; Bialy et al. 1999).

Soyasapogenols are always characterized by the chain: 3-O-[α-l-rhamnopyranosyl-(1 → 2)-β-d-galactopyranosyl(1 → 2)-β-d-glucuronopyranosyl] (Kitagawa et al. 1988; Oleszek et al. 1988; Mahato, 1991; Massiot et al. 1992; Bialy et al. 1999; Huhman and Sumner 2002; Kapusta et al. 2005; Tava et al. 2005; Tava et al. 2009).

The nature of the sugar chain at the 3-O position, unique in soyasapogenols (Table 2), suggests a specificity not only for the A ring (Fig. 1) but also for the type of aglycone (soyasapogenol/haemolytic genins). Within the higher variation of the sugar chain of haemolytic saponins it is worth to note the unusual 1–3 linkage found only in M. truncatula (Table 2).

Glycosylation at 28-O position only concerns haemolytic saponins, characterized by the COOH substitution at C-28 and seems to be more species specific, as evidenced in Table 3. M. arabica (Tava et al. 2009), M. hybrida (Bialy et al. 2006) and M. lupulina (Oleszek et al. 1988) saponins are characterized by the presence of the β-d-glucopyranose unit esterified at the C-28 carboxylic group, while the disaccharide chain 28-O[-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside] was only found in M. polymorpha (Kinjo et al. 1994). Chains with more than two sugars, always characterized by α-l-arabinopyranose, directly linked at the C-28, and an α-l-rhamnopyranose in the central position, linked (1 → 4) with a β-d-xylopyranose and branching points formed by α-l-arabinopyranose or β-d-apiofuranose linked (1 → 3) at the β-d-xylopyranose unit, are typical of saponins extracted from M. arborea (Tava et al. 2005), M. sativa (Massiot et al. 1988; Oleszek et al. 1990; Massiot et al. 1991; Oleszek et al. 1992; Timbekova et al. 1996; Bialy et al. 1999) and M. truncatula (Huhman et al. 2005; Kapusta et al. 2005). In contrast to sugar linkage at 3-O, the C-28 glycosylation suggests a high selectivity for the nature of the saccharide unit independent of the involved sapogenin.
Table 3

Sugars attached to the 28-O position of the triterpene skeleton of Medicago aglycones

M. arabica, M. hybrida,

H

M. lupulina

βd-Glc

M. polymorpha

H

β-d-Glc

β-d-Glc(1 → 6)β-d-Glc

M. arborea, M. sativa,

M. truncatula

H

α-l-Rha(1 → 2)α-l-Ara

β-d-Xyl(1 → 4)α-l-Rha(1 → 2)α-l-Ara

β-d-Xyl(1 → 4)α-l-Rha(1 → 2)α-l-Ara

α-l-Ara(1 → 3)

β-d-Xyl(1 → 4)α-l-Rha(1 → 2)α-l-Ara

β-d-Api(1 → 3)

From all the structural data on Medicago saponins, it can be observed that the non-haemolytic saponins (soyasapogenol saponins) only occur as monodesmosides (sugar chain at C-3 position), whit a unique sugar chain (Table 2). Haemolytic saponins on the contrary can occur as mono-, bi- and, occasionally, tridesmosides (Oleszek et al. 1992)—with the additional sugar chains at C-28 and C-23 position, respectively—with variable sugar chains. Thus, haemolytic saponins seem to display a wider potential range of biological plasticity in consideration of the variation in the nature and position of the sugar moiety.

Concluding remarks

The information here reported have been mainly derived from the chemical data published on saponins isolated over the years from different species within the genus Medicago. The comparison of the chemical structures of these saponins allowed to disclose the structural correlations among aglycones, by means of the study of all the positions and nature (state of oxidation) of the specific substituents. Thus saponins from Medicago can all be related to the basic structure of β-amyrin and fall into two groups: (1) with an OH on the aglycone at C-24 without any substituent at the C-28 atom and (2) with a COOH group at C-28, associated with substituents possessing different degrees of oxidation (zero, OH, CHO, COOH) at C-23. The synthesis of these different structures appears to be specifically regulated and connected with their biological activity (haemolytic/not haemolytic). This subdivision is also reflected in their sugar moieties and affects the position and the nature of the sugar chains in the two groups.

Little data are still available on the enzymatic machinery involved in the saponin biosynthetic pathway in the genus Medicago and in its regulation. The current molecular data and the chemical information here discussed suggest, however, the involvment of stereospecific P450 cytochromes in the biosynthesis of saponin aglycones in Medicago spp.

The saponin biosynthetic pathway proposed here should stimulate phytochemical and functional genomic joint approaches to improve our knowledge of the biosynthesis of these secondary metabolites.

References

  1. Achnine L, Huhman DV, Farag MA, Sumner LW, Blount JW, Dixon RA (2005) Genomics-based selection and functional characterization of triterpene glycosyltransferases from the model legume Medicago truncatula. The Plant J 41:875–887CrossRefGoogle Scholar
  2. Argentieri MP, D’Addabbo T, Tava A, Agostinelli A, Jurzysta M, Avato P (2008) Evaluation of nematicidal properties of saponins from Medicago spp. Eur J Plant Pathol 120:189–197CrossRefGoogle Scholar
  3. Bialy Z, Jurzysta M, Oleszek W, Piacente S, Pizza C (1999) Saponins in alfalfa (Medicago sativa L.) root and their structural elucidation. J Agric Food Chem 47:3185–3192PubMedCrossRefGoogle Scholar
  4. Bialy Z, Jurzysta M, Mella M, Tava A (2004) Triterpene saponins from aerial parts of Medicago arabica L. J Agric Food Chem 52:1095–1099PubMedCrossRefGoogle Scholar
  5. Bialy Z, Jurzysta M, Mella M, Tava A (2006) Triterpene saponins from the roots of Medicago hybrida. J Agric Food Chem 54:2520–2526PubMedCrossRefGoogle Scholar
  6. Cammareri M, Consiglio MF, Pecchia P, Corea G, Lanzotti V, Ibeas JI, Tava A, Conicella C (2008) Molecular characterization of a β-amyrin synthase gene from Aster sedifolius L. and analysis of triterpenoid saponins. Plant Sci 175:255–261CrossRefGoogle Scholar
  7. Confalonieri M, Cammareri M, Biazzi E, Pecchia P, Fevereiro MPS, Balestrazzi A, Tava A, Conicella C (2009) Enhanced triterpene sapogenin biosynthesis and root nodulation in transgenic barrel medic (Medicago truncatula Gaertn.) expressing a novel β-amyrin synthase (AsOXA1) gene. Plant Biotech J 7:172–182CrossRefGoogle Scholar
  8. D’Addabbo T, Avato P, Tava A (2009) Nematicidal potential of materials from Medicago spp. Eur J Plant Pathol 125:39–49Google Scholar
  9. Dewick PM (2001) Medicinal natural products—a biosynthetic approach. Wiley, New YorkGoogle Scholar
  10. Francis G, Keem Z, Makkar HPS, Becker K (2002) The biological action of saponins in animal systems. A review. Brit J Nutr 88:587–605PubMedCrossRefGoogle Scholar
  11. Haralampidis K, Trojanowska M, Osbourn AE (2002) Biosynthesis of triterpenoid saponins in plants. Adv Biochem Eng Biotech 75:31–49Google Scholar
  12. Hayashi H, Hanaoka S, Tanaka S, Fukui H, Tabata M (1993) Glycyrrhetinic acid 24-hydroxylase activity in microsomes of cultured licorice cells. Phytochemistry 34:1303–1307CrossRefGoogle Scholar
  13. Hostettmann K, Marston A (1995) Chemistry and pharmacology of natural products—Saponins. In: Phillipson JD, Baxter H (eds). Cambridge University Press, CambridgeGoogle Scholar
  14. Huhman DV, Sumner LW (2002) Metabolic profiling of saponins in Medicago sativa and Medicago truncatula using HPLC coupled to an electrospray ion-trap mass spectrometer. Phytochemistry 59:347–360PubMedCrossRefGoogle Scholar
  15. Huhman DV, Berhow MA, Sumner LW (2005) Quantification of saponins in aerial and subterranean tissues of Medicago truncatula. J Agric Food Chem 53:1914–1920PubMedCrossRefGoogle Scholar
  16. Iturbe-Ormaetxe I, Haralampidis K, Papadopoulou K, Osburn AE (2003) Molecular cloning and characterization of terpene synthases from Medicago truncatula and Lotus japonicus. Plant Mol Biol 51:731–743PubMedCrossRefGoogle Scholar
  17. Jurzysta M, Waller GR (1996) Antifungal and hemolytic activity of aerial parts of alfalfa (Medicago) species in relation to saponin composition. In: Waller GR, Yamasaky K (eds) Advances in experimental medicine and biology, vol 404, Saponins used in traditional and modern medicine. Plenum Press, New York, pp 565–574Google Scholar
  18. Kalinowska M, Zimowskim J, Paczkowski C, Zdzislaw A (2005) The formation of sugar chains in terpenoid saponins and glycoalkaloids. Phytochem Rev 4:237–257CrossRefGoogle Scholar
  19. Kapusta I, Stochmal A, Perrone A, Piacente S, Pizza C, Oleszek W (2005) Triterpene saponins from barrel medic (Medicago truncatula) aerial parts. J Agric Food Chem 53:2164–2170PubMedCrossRefGoogle Scholar
  20. Kinjo J, Uemura H, Nakamura M, Nohara T (1994) Two new triterpenoidal glycosides from Medicago polymorpha L. Chem Pharm Bull 42:1339–1341PubMedGoogle Scholar
  21. Kitagawa I, Taniyama T, Murakami T, Yoshihara M, Yoshikawa M (1988) Saponin and sapogenol. XLVI. On the constituents of aerial part of american alfalfa, Medicago sativa L. The structure of dehydrosoyasaponin I. Yakugaku Zasshi 108:547–551Google Scholar
  22. Kurosawa Y, Takahara H, Shiraiwa M (2002) UDP-glucuronic acid:soyasapogenol glucuronosyltransferase involved in saponin biosynthesis in germinating soybean seeds. Planta 215:620–629PubMedCrossRefGoogle Scholar
  23. Lacaille-Dubois MA, Wagner H (2000) Bioactive saponins from plants: an update. In: Rahman A (ed) Studies in natural product chemistry, vol 21. Elsevier, London, pp 633–687Google Scholar
  24. Levy M, Zehavi U, Naim M, Polacheck I, Evron R (1989) Structure-biological activity relationshipin alfalfa antimycotic saponins: the relative activity of medicagenic acid and synthetic derivatives thereof against plant pathogenic fungi. J Phytopathol 125:209–216CrossRefGoogle Scholar
  25. Li L, Cheng H, Gai J, Yu D (2007) Genome-wide identification and characterization of putative cytochrome P450 genes in the model legume Medicago truncatula. Planta 226:109–123PubMedCrossRefGoogle Scholar
  26. Mahato SB (1991) Triterpenoid saponins from Medicago hyspida. Phytochemistry 30:3389–3393PubMedCrossRefGoogle Scholar
  27. Massiot G, Lavaud C, Le Men-Olivier L, van Binst G, Miller SPF, Fales HM (1988) Structural elucidation of alfalfa root saponins by mass spectrometry and nuclear magnetic resonance analysis. J Chem Soc Perkin Trans I:3071–3079Google Scholar
  28. Massiot G, Lavaud C, Besson V, Le Men-Olivier L, van Binst G (1991) Saponins from aerial parts of alfalfa (Medicago sativa). J Agric Food Chem 39:78–82CrossRefGoogle Scholar
  29. Massiot G, Lavaud C, Benkhaled M, Le Men-Olivier L (1992) Soyasaponin VI, a new maltol conjugate from alfalfa and soyabean. J Nat Prod 55:1339–1341CrossRefGoogle Scholar
  30. Milgate J, Roberts DCK (1995) The nutritional and biological significance of saponins. Nutr Res 8:1223–1249CrossRefGoogle Scholar
  31. Morris RJ, Tankersley DL (1963) The synthesis of the β-d-glucoside of medicagenic acid, an alfalfa root saponin. J Org Chem 28:240–242CrossRefGoogle Scholar
  32. Nowacki E, Jurzysta M, Dietrych-Szostak D (1976) Zur biosynthese der medicagensaure in keimenden luzernesamen. Biochem Physiol Pflanz 156:183–186Google Scholar
  33. Oleszek W (1996) Alfalfa saponins: structure, biological activity and chemotaxonomy. In: Waller GR, Yamasaky K (eds) Advances in experimental medicine and biology, vol 405, saponins used in food and agriculture. Plenum Press, New York, pp 155–170Google Scholar
  34. Oleszek W, Price KR, Fenwick GR (1988) Triterpene saponins from the roots of Medicago lupulina L. (black medic trefoil). J Sci Food Agric 43:289–297CrossRefGoogle Scholar
  35. Oleszek W, Price KR, Colquhoun IJ, Jurzysta M, Ploszynski M, Fenwick GR (1990) Isolation and identification of alfalfa (Medicago sativa L.) root saponins: their activity in relation to a fungal bioassay. J Agric Food Chem 38:1810–1817CrossRefGoogle Scholar
  36. Oleszek W, Jurzysta M, Ploszynski M, Coloquhoun IJ, Price KR, Fenwick GR (1992) Zahnic acid tridesmoside and other dominant saponins from alfalfa (Medicago sativa L.) aerial parts. J Agric Food Chem 40:191–196CrossRefGoogle Scholar
  37. Peri I, Mor U, Heftmann E, Bondi A, Tencer Y (1979) Biosynthesis of triterpenoid sapogenols in soybean and alfalfa seedlings. Phytochemistry 18:1671–1674CrossRefGoogle Scholar
  38. Phillips DR, Rasbery JM, Bartel B, Matsuda SPT (2006) Biosynthetic diversity in plant triterpene cyclization. Curr Opin Plant Biochem 9:305–314CrossRefGoogle Scholar
  39. Shibuya M, Hoshino M, Katsube Y, Hayashi H, Kushiro T, Ebizuka Y (2006) Identification of b-amyrin and sophoradiol 24-hydroxylase by expressed sequence tag mining and functional expression assay. FEBS J 273:948–959PubMedCrossRefGoogle Scholar
  40. Sprag SG, Light ME, van Staden J (2004) Biological activities and distribution of plant saponins. J Ethnopharm 94:219–243CrossRefGoogle Scholar
  41. Suzuki H, Achnine L, Xu R, Matsuda SP, Dixon RA (2002) A genomic approach to the early stages of triterpene saponin biosynthesis in M. truncatula. The Plant J 32:1033–1048CrossRefGoogle Scholar
  42. Tava A, Avato P (2006) Chemical and biological activity of triterpene saponins from Medicago species. Nat Prod Comm 1:1159–1180Google Scholar
  43. Tava A, Mella M, Avato P, Argentieri MP, Bialy Z, Jurzysta M (2005) Triterpene saponins from leaves of Medicago arborea L. J Agric Food Chem 53:9954–9965PubMedCrossRefGoogle Scholar
  44. Tava A, Mella M, Avato P, Biazzi E, Pecetti L, Bialy Z, Jurzysta M (2009) New Triterpenic Saponins from the Aerial Parts of Medicago arabica (L.) Huds. J Agric Food Chem 57:2826–2835PubMedCrossRefGoogle Scholar
  45. Timbekova AE, Isaev MI, Abubakirov NK (1996) Chemistry and biological activity of triterpenoid glycosides from Medicago sativa. In: Waller GR, Yamasaki K (eds) Advances in experimental medicine and biology, Saponins used in food and agriculture, vol 405. Plenum Press, New York, pp 171–182Google Scholar
  46. Vincken JP, Heng L, de Groot A, Gruppen H (2007) Saponins, classification and occurrence in the plant kingdom. Phytochemistry 65:261–291Google Scholar
  47. Wang J, Hou B (2009) Glycosyltransferases: key players involved in the modification of plant secondary metabolites. Front Biol China 4:39–46CrossRefGoogle Scholar
  48. Xu R, Fazio GC, Matsuda SPT (2004) On the origin of triterpenoid skeletal diversity. Phytochemistry 65:261–291PubMedCrossRefGoogle Scholar
  49. Zehavi U, Ziv-Fecht O, Levy M, Naim M, Polacheck I, Evron R (1993) Synthesis and antifungal activity of medicagenic acid saponins on plant pathogens: modification of the saccharide moiety and the 23α-substitution. Carbohydr Res 244:161–169PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2010

Authors and Affiliations

  1. 1.CRA-FLC Centro di Ricerca per le Produzioni Foraggere e Lattiero CasearieLodiItaly
  2. 2.Dipartimento Farmaco-ChimicoUniversità di Bari Aldo MoroBariItaly

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