Chemistry of southeast asian plants 2

types of carbon skeleton, have been isolated or detected in the Hepaticae so far.112-11380% of these sesquiterpenoids are enantiomeric to those found in higher plants.114The most widespr

Chapter 3 The Hepaticae

3.1 Introduction

Bryophytes are taxonomically placed between pteridophytes and algae They are the simplest terrestrial plants and the most primitive higher plants They are divided morphologically into three classes: Hepaticae (liverworts, 6000 species), Musci (mosses, 14000 species) and Anthocerotae (hornworts, 3000 species) Hepaticae is further divided into two subclasses: the Jungermanniidae and the Marchantiidae These two subclasses together contain 61 families with about 6000 species found worldwide.111

Chemical investigations of liverworts (and the bryophytes in general) have been neglected for a long time, for two main reasons First, they are tiny plants and tend to grow intermingled with other liverwort or moss species; therefore, it is difficult to collect them in adequate amounts as pure samples for chemical investigations Second, their identification is difficult and they are considered to have no nutritional value for human Started from 1980s, the development of superconducting magnets for NMR spectroscopy and the advances in computing technology have helped scientist to overcome the difficulties, by reducing the minimum amount of pure sample needed for analysis

Among the bryophytes, the phytochemistry of Hepaticae has been studied in greater detail because they possess oil bodies, while the other two classes lack such features

Up to 2001, more than 700 terpenoids and 220 aromatic compounds (excluding flavonoids) have been isolated from or detected in the Hepaticae, while the number of terpenoids found in Musci is signficantly lower.112

Monoterpenoids are responsible for the distinctive smell that many liverworts produce when crushed Among approximately 60 monoterpenoids found in liverworts, α-pinene

(164), β-pinene (165), and limonene (160) are the most abundant compounds.113

Structures of representative compounds are shown in Fig 3-1

types of carbon skeleton, have been isolated or detected in the Hepaticae so far.112-11380% of these sesquiterpenoids are enantiomeric to those found in higher plants.114

The most widespread sesquiterpene in liverworts is ent-bicyclogermacrene (170),

although the number of bicyclogermacranes isolated from liverworts is rather small It

is considered to be the biosynthetic precursor of many type of sesquiterpenoids, including aromadendranes, maalianes, aristolanes, and vitranes

The most common type of sesquiterpenoids in liverworts is the eudesmanes, of which

ent- α-selinene (171) and ent-β-selinene (172) are the two most widespread

compounds The family of Frullaniaceae, which comprises more than 500 species, is a rich source of eudesmanolides, which contribute to about three-quarters of eudesmanes

found in liverworts Among them, (-)-frullanolide (173) and dihydrofrullanolide (174)

are the most frequently encountered compounds.113

The Panamanian liverwort Plagiochila moritziana produces four rare C-35

sesquiterpene lactones (sesquiterpene + diterpene), plagiospirolides A-D (175-178),

and a rare C-30 sesquiterpene lactone (sesquiterpene + sesquiterpene), plagiospirolide

E (179) The C-35 sesquiterpene lactones could be the products of a Diels-Alder

cycloaddition reaction between the dienophiles diplophyllolide (180) and diplophylline (181) (both were also found in P moritziana) and either the diterpene

ent-diene (182) (in case of plagiospirolides A & B) or (183) (in case of plagiospirolides C

& D) For the C-30 lactone dimer, plagiospirolide E, the two suggested sesquiterpene

moieties are (181) and an aromadendrene derivative (184).115-116

O H

O

O H

H H

Aromadendranes comprise another widespread class of liverwort sesquiterpenoids

Ent-aromadendranes are present in many species of the Jungermanniales, while secoaromadendranes are found largely in Plagiochila species Among the

aromadendranes, anastreptene (185) and ent-spathulenol (186) are the most regularly

isolated compounds, especially the latter which has been found in more than 80

species Mylia taylorii was found to be a rich source of aromadendranes and

secoaromadendranes.114 This species also produces dimeric sesquiterpenoids,

bitaylorione (187), myltayloriones A (188) and B (189) The latter two are probably products of Diels-Alder reaction of mylione (190) and (191), an isomer of taylorione (192).117

H HO

O O

O

O

O O

O O

(188), (189)

O

The biosynthetically interesting pinguisane sesquiterpenoids are found in liverworts only, and they are also restricted to a number of families, namely Lejeuneaceae, Trichocoleaceae, Ptilidiaceae, Porellaceae and Riccardiaceae.112 The carbon skeleton

of pinguisanes does not conform to the biogenetic isoprene rule Acutifolones A (193) and B (194), together with three dimeric pinguisane sesquiterpenoids, bisacutifolones

A (195), B (196) and C (197) have been isolated from Porella acutifolia subsp

Tosana.118-119 Further modifications of the pinguisane skeleton lead to the

norpinguisanes (198, 199)247 and pinguisanolides (200, 201),248 isolated from the

liverworts Porella recurva (Taylor) Kuhnemann and Trocholejeunea sandvicensis,

respectively

COOMe

O O

MeOOC

R H

O

(196) R = α-OH (197) R = H

CHO CHO

OH

(199) R = O

The biosynthesis pathway for pinguisone (203) from trans,cis-farnesol (202) has been

proposed, based on labeling experiment in the cultured liverwort Aneura pinguis,

120

(203)

Fig 3-2 Proposed biosynthetic pathway of pinguisone

The main constituent of Saccogyna viticulosa is the alcohol saccogynol (204) with the

zierane carbon skeleton.121 This type of sesquiterpene is very rare in nature, and

zierone (205) is the only other known member of this group

Liverworts are also rich sources of diterpenoids More than 200 diterpenoids, representing about 20 carbon skeletons, have been found in the Hepaticae Among them, clerodanes, labdanes, and kauranes are the most widepread types

Clerodanes are distributed in both higher plants and liverworts They have a vast stereochemical and functional variability More than half of the clerodanes found in

liverworts were isolated from Jungermannia and Schistochila species The liverwort

Anastrepta orcadensis produced two bitter clerodanes anastreptin (206) and orcadensin

(207).122 The latter possess a structure similar to that of gymnocolin (208) from

Gymnocolea inflata 1H-NMR data suggested that gymnocolin was a trans-clerodane,

but X-ray crystallographic study revealed that (208) was actually a cis- clerodane.123

These cis-clerodane lactones with a β-substituted furan ring exhibit intense

bitterness.114

O

O H

O

O O

O O

O O

O

O O

O O

H

OAc

Kaurane diterpenoids encountered in liverworts belong to the ent-series, and all of

them were isolated from the Jungermanniales ent-Kaurene (209) is the most common

compound of this series ent-(16S)-11α-Hydroxykauran-15-one (210) was isolated from Jungermannia infusca, together with a number of known ent-kauranes, including

(213).124 These known ent-kauranes were also found in Nardia species,125 another rich source of this type of diterpenoids

(209) R = H2 (210) R = OH, R' = α-Me, β-H (211) R = α-H, β-OH (213) R = H, R' = β-Me, α-H (212) R = O

Jungermannia infusca also produced a number of infuscasides, including infuscasides

A (214) and B (215), the first terpene glucosides found in bryophytes.126-127

OH O

OAc

OH OH OH

HO

OH

O O

HO

OH H

The majority of labdane diterpenoids were isolated from the Jungermanniales

Scapania undulata and Ptychanthus striatus are rich sources of these compounds: 22

labdane diterpenoids have been found in these species prior to 2001, including

labda-12E,14-dien-8α,11ζ-diol (216), scapanins A (217) and B (218), and ptychantins E-G

(219-221).112,114,122,128,129

OAc OR

AcO

AcO

AcO

H OAc

O

H OAc

OH

HO AcO

(221) R = Ac

A few rare classes of diterpenoids have been found in liverworts, e.g pleuroziol (222)

and levierol (223), which belong to the chettaphanins (rearranged labdane),129-130 and

the cembrane diterpenoid, setiformenol (224).131-132

The bis(bibenzyls) are a new class of natural products that is restricted to the

liverworts Since the first bis(bibenzyl), marchantin A (225) was isolated from

Marchantia polymorpha,133 more than 60 compounds of this class have been reported

Structurally, bis(bibenzyls) can be divided into four groups, depend on the type of the linkages between the two bibenzyl units

The first group, the marchantins, includes compounds with two biphenyl ether bonds between rings A/C and B/D The marchantins are widespread in the Marchantiales species, more than 20 compounds of this group have been characterized Beside the

major component marchantin A, M polymorpha also produced marchantins B-G

(225) R = H, R' = OH, R" = H2 (228) R = R' = H

(226) R = R' = OH, R" = H2 (229) R = H, R' = Me

(227) R = R' = H, R" = H2 (230) R = OH, R' = H

(231) R = H, R' = OH, R" = O

It is noteworthy that marchantin C (227) is not only isolated from the Marchantiales,

but also from the Jungermanniales, including Plagiochila sciophila and Schistochila

glaucescens. 137-138

Riccardin A (232) is representative of the second group, the riccardins, which possess

one biphenyl ether bond and one biphenyl bond between the two bibenzyl units

Riccardin C (233) has been isolated from various species, such as Reboulia

hemisphaerica,139 M polymorpha,136,140 and Monoclea forsteri.141 Lunularic acid (234)

or lunularin (235), could be the biosynthetic precursors of riccardins, since these two

bibenzyls are widespread in leafy and thalloid liverworts

OH

O

OH RO

Plagiochins form the third group of cyclic bis(bibenzyls), consisting of those with two

biphenyl bonds between rings A/C and B/D Its representative, isoplagiochin C (236),

was isolated from Plagiochila fruticosa, together with isoplagiochin D (237).142-143

The fourth group, perrottetin, contains a single o,p-ether linkage between two bibenzyl

units Its first member, perrottetin E (238) was isolated from Plagiochila sciophila,137

Radula perrottettii,144 R kojana,145 and M polymorpha.136 R perrottettii also produces

cultures of M polymorpha has been reported.146-147 It involved two specific

cytochromes P-450: one catalyzes the coupling of two lunalaric acid (234) molecules

to form marchantin C, while the other is responsible for the hydroxylation of marchantin C to marchantin A The detailed pathway is shown in Fig 3-4

COOH

HO

HO HO

O

S CoA COOH

NADP NADPH

NADP NADPH

O OH O

3.2 Chemistry of Pallavicinia

The Pallaviciniaceae family is one of the most diversified families of simple thalloid liverworts, comprising approximately 150 species in 10 (currently accepted) genera:148

Greeneothallus, Hattorianthus, Jensenia, Moerckia, Pallavicinia, Podomitrium,

Seppeltia, Symphyogyna, Symphyogynopsis, Xenothallus Of these, the most

chemically studied genus is Pallavicinia, and only 3 out of 23 species of this genus

have been studied so far

Sacculatal (241), a pungent diterpene dialdehyde, has been isolated from Pallavicinia

levieri, together with pallavicinol (223), a rare chettaphanin diterpenoid.149 It is

interesting to note that sacculatal is a significant chemical marker of Pellia

endiviifolia,114 hence these two species are chemically very close Labdanes and sacculatanes are considered chemical markers of the family Pallavicinaceae.150

Pallavicinia lyellii is considered chemically close to P subciliata ( = P longispina),

since bicyclogermacranes and aromadendranes were detected as the major components

of both species.114

From the diethyl ether extract of the Japanese P subciliata, Toyota et al 151 isolated

six novel secolabdanoids (242-247) This was the first isolation of such complex

diterpenoids from nature The structures of these highly oxygenated diterpenoids have

been established by 2D-NMR and/or X-ray crystallographic analysis Wu et al 152 also

isolated pallavicinin (248), a 7,8-secolabdanoid, from P subciliata collected in

Taiwan These results proved that liverworts biosynthesize many complex and interesting new diterpenoid structures

O O

O

HO

O O

R

(248) R = H

Although many studies have been performed on the phytochemistry of the liverworts

in general, it is estimated that only about 10% of all liverwort species have been

investigated chemically As to the genus Pallavicinia, so far only the species collected

in Japan and Taiwan have been studied in detail The present study will focus on the

constituents of Pallavicinia species collected in Vietnam and Singapore

3.3 Constituents of Singaporean Pallavicinia cf lyellii

P lyellii and P levieri are quite similar, and it is difficult to distinguish them without

reference to the male plant The sample collected for this study lacked the male

reproductive organ and hence could only be identified as Pallavicinia cf lyellii

Phytochemical studies of the EtOAc extract of Pallavicinia cf lyellii collected in

Singapore led to the isolation of ten compounds, seven of which are novel:

pallavicinins B-F (249-253), 8-hydroxymarchantin C (267), and 7-oxo-riccardin D (268)

O

O

O

O H

H H

1

18 19

7

8 9 10

20

15

16 17

11 12 13 14

17

18 19

3

5 1

7

9 10

20 11

16 13 14 15

3

5 1

7

9 10

20 11

16 13 14 15

18

H H

19

H 20

3'

12' 14

1'

7' 5

5'

12

14' 10

10'

H H 1

18 19

7

8 9 10

20

15

16 17

starting points to establish the gross structure The six-membered ring was built first starting from two geminal tertiary methyl groups Me-18 [δC 33.5] and Me-19 [δC 24.3] Since both H-18 and H-19 showed HMBC correlations to each other and to C-3, C4 and C-5 (a methylene, a quaternary carbon and a methine, respectively), these two methyl groups must be attached to the quaternary carbon C-4 The cross-peaks ot the equatorial proton H-2α [δH 1.77 (dq, J = 12.2, 3.8 Hz)] with C-1, C-3, C-4 and C-10,

indicated that C-2 is linked to C-3, and helped to identify the last two members of the ring The fact that H-1 [δH 3.78 (dd, 12.2, 3.7 Hz)] correlated to C-3 and C-5 showed that this oxygenated methine is linked directly to C-2 The tertiary methyl Me-20 is attached to C-10, since its protons [δH 1.09 (s)] showed correlations to C-1 and C-5 The methine proton at C-5 correlated to both C-6 and C-7, therefore this double bond

is attached to C-5 The observed correlations of C-9 [δC 61.5] with C-1, C-5 and C-20 suggested that the methine C-9 is attached to C-10 The tertiary Me-17 showed correlations with the ketal carbon C-8 [δC 118.3] and C-9, revealed that C-8 is between C-17 and C-9

1 3

18 19

7 10

20

O

Fig 3-5: HMBC correlations of ring A

The HMBC correlations, as well as the coupling constants, of H-14 [δH 7.00 (qd, J =

7.2, 1.0 Hz)] and H-15 [δ

group is attached to C-14 Thus, C-12 and C-16 are linked directly to C-13, since H-14 showed HMBC correlations with both the oxygenated methine C-12 and the carboxyl C-16 Finally, H-11 correlated to C-8 and C-12; therefore, it is linked to C-9 and C-12 Since the degree of unsaturation indicated a tetracyclic system, there must be two ether links and one ester link in the molecule, as shown in Fig 3-6

O

O

O

OH

H1

18 19

H

Fig 3-6: Selected HMBC correlations of (249)

The relative stereochemistry was established by NOEDIFF experiments (Table 3-1 and Fig 3-7) Since H-1 [δH 3.78 (dd, J = 12.2, 3.7 Hz)] is axial and irradiation at this

proton caused NOE enhancements at H-2α, H-3α and H-5, H-1 and H-5 were deduced

to be syn Irradiation at H-20 caused NOE effects at H-2β, H-6 and H-9, therefore both H-9 and Me-20 were on the same face Irradiation at H-9 caused NOE enhancements at H-20, H-11 and H-12, suggesting that H-11 and H-12 were also on the same face as H-

9 and Me-20 Finally, an NOE enhancement was observed at H-9 when H-17 was irradiated, suggesting that the configuration of Me-17 was β The absolute

configuration of pallavicinin B was assumed to be the same as those of 242 and 243.151

O

O O

H H

H

CH3

CH3

CH31

H

H

H3C H

11 2

5

Fig 3-7: Observed NOE enhancements of (249)

Table 3-1 1H-NMR (500 M Hz), 13C-NMR (125 M Hz), HMBC and NOEDIFF

spectral data of compound (249) in CDCl3

A plausible biosynthesis of 249, which proceeds via cleavage of C7-C8 bond of labdane diterpenoids followed by cyclization, is shown in Fig 3-8

11 12 13 14

17

18 19

J = 10.0 Hz, H-9)], four tertiary methyl groups [δH 0.80, 0.86, 1.26 and 2.20 (each 3H,

s, H-18, 19, 20 and 17)] and one secondary methyl group [δH 2.07 (dd, J = 7.3, 2.7 Hz,

H-15)] The 13C-NMR spectrum contained twenty signals There was one ketone group [δC 210.8 (C-8)], a carboxyl group [δC 168.2 (C-16)], four olefinic carbons [δC 142.0 (C-14); 135.0 (C-6); 124.5 (C-13); and 120.2 (C-7)] and three oxygenated carbons [δC

78.4 (C-1); 75.6 (C-11); and 72.9 (C-12)] These data suggested that this compound is

a tricyclic diterpenoid

The tricyclic system was established using HMQC and HMBC experiments (Table

3-2) The first six-membered ring was built up in the similar manner as for 249 Since the

vinylic protons H2-7 showed HMBC correlations to C-6 and C-5, the vinyl group must

be attached to 5 The methine H-9 and the quaternary methyl H-20 are attached to

C-10, since both correlated to C-10 and C-5 The fact that H-12 showed 3J correlation

with C-9, while H-11 correlated to C-8 suggested that C-8 is linked directly to C-9, and then C-9 is connected to C-11 The cross-peaks of H-1 with C-9, C-20 and C-12, revealed that the second ring is formed by an ether link between C-12 and C-1 Since both H-12 and H-15 showed correlations to C-13 and C-14, the methyl group Me-15 must be connected to C-14, while C-12 is linked to C-13 The olefinic proton H-14 also correlated to C-16, therefore this carboxyl group must be attached to C-13 The correlations of H-11 to C-12, C13 and C-16 showed that there must be an ester link between C-11 and C-16, thus the third ring in this structure is a five-membered lactone ring Finally, the tertiary methyl H-17 is connected to C-8, since it correlated to this ketone group

O

O

O O

3 1

7

5

8

9 10

11 12

13 14

17

18 19

20

Fig 3-9: Selected HMBC correlations of (250)

The relative stereochemistry was established by NOEDIFF experiments (Table 3-2) The coupling constants of H-1 [δ

axial proton Irradiation at H-1 caused NOE enhancements at H-9, as well as H-3α,

therefore both H-1 and H-9 must be syn Since NOE enhancements at H-6 and Me-20

were observed when 19 was irradiated, 20 must be on the same face with

Me-19, while H-5 is on the opposite face Irradiation at H-11 caused NOE effects at Me-20 and H-12, suggested that both H-11 and H-12 were also on the same face as Me-20

Table 3-2 1H-NMR (500 M Hz), 13C-NMR (125 M Hz), HMBC and NOEDIFF

spectral data of compound (250) in CDCl3

The biosynthesis of compound 250 may proceed via the pathway shown below:

(250)

Pallavicinin D

Pallavicinin D (251), was obtained as a colorless powder, mp 195-197 oC, [α]D +32.9

(c 0.12, CHCl3) It was determined to have the molecular formula C22H32O6 by

HR-EIMS (m/z M+ 392.2206)

O

O

O O

3

5 1

7

9 10

20 11

16 13 14 15

12

21 H

(251)

The 1H-NMR spectrum contained signals due to an olefinic proton [δH 6.48 (qd, J =

7.5, 2.9 Hz, H-14)], four oxygenated methines [δH 3.16 (dd, J = 3.9, 11.5 Hz, H-1); 4.97 (dd, J = 7.8, 10.8 Hz, H-11); 5.09 (dt, J = 7.8, 2.7 Hz, H-12); and 5.51 (dt, J =

2.4, 3.1 Hz., H-6)], five tertiary methyl groups [δH 0.95, 0.97, 1.26, 1.50 and 2.07 (each 3H, s, H-18, 19, 20, 17 and 22)] and one secondary methyl group [δH 2.26 (dd, J = 2.9,

7.5 Hz, H-15)] The 13C-NMR spectrum contained twenty two signals There were two carboxyl groups [δC 166.2 (C-16) and 169.8 (C-21)], one trisubstituted double bond [δC

123.4 (C-13) and 141.4 (C-14)] and five oxygenated carbons [δC 69.4 6); 70.2 8); 73.4 (C-12); 73.9 (C-11); and 80.0 (C-1)] These data suggested that this compound has a tetracyclic structure

(C-HMQC and HMBC experiments (Table 3-3) were used to determine the structure of the compound In the HMBC experiment, ring A is built first, again starting from two tertiary methyl groups Me-18 and Me-19 (Fig 3-10)

C-10, therefore, it must be C-6, attached to C-5 The fact that H-7 showed correlations

to C-5, C-6, C-8, C-9, C-17, while the tertiary methyl H-17 correlated to C-7, C-8 and C-9, led to the structure of ring B (Fig 3-11), with C-17 attached to the oxygenated C-

8

O

OH10

Since H-11 correlated to C-8, it must be connected to C-9 The cross-peak of H-12 with C-1 revealed that there was an ether link between C-1 and C-12 The coupling

constant between H-11 and H-12 in this compound (7.8Hz) is the same as that of 250,

therefore both compounds must have the same stereochemistry at C-11 and C-12.The secondary methyl H-15 showed correlations to both olefinic carbons C-13 and C-14, therefore C-15 is attached to C-14 H-14 correlated to C-12 and C-16, showed that these two are attached to C-13 Finally, structure of the last ring is completed by the observed cross-peaks of H-11 with C-12, C-13 and C-16 Again, it is a five-membered lactone ring

O

OO

10 11

16 13

14 15

12

OH

17 8

Fig 3-12: Observed HMBC correlations of ring C and D

The relative configurations of 8 chiral centers were determined by NOEDIFF experiments (Table 3-3) Irradiation at H-6 caused NOE enhancements at H-5, H-7α

and H-18, therefore Me-18, H-5 and H-6 were syn Irradiation at H-5 caused NOE

effect at 1, suggested that 1 was also on the same face with 5 Irradiation at

H-20 caused NOE effects at H-11, H-12, H-17 and H-19, therefore H-11, H-12, Me-17,

Me-20, and Me-19 were deduced to be syn Finally, the fact that NOE enhancement

was not observed for H-9 when H-20 was irradiated, together with a diaxial coupling

(10.8 Hz) in H-11 proton signal, suggested that H-9 and Me-20 were anti

H H

O

Me

Me AcO

20 17

1 9 5

Biosynthesis of (251) should be similar to that of (250), without the cleavage of C7-C8

bond

Table 3-3 1H-NMR (500 M Hz), 13C-NMR (125 M Hz), HMBC and NOEDIFF

spectral data of compound (251) in CDCl3

1 3.16 (dd, J = 11.5, 3.9 Hz) 5 2, 9, 20 80.02α 1.73 (m) a

Pallavicinin E

Pallavicinin E (252), was obtained as a white powder, mp 204-205 oC, [α]D +95.5 (c

0.28, CHCl3) It was determined to have the molecular formula C22H32O6 by HR-EIMS

(m/z M+ 392.2208)

O

O

O O

3

5 1

7

9 10

20 11

16 13 14 15

The 1H-NMR spectrum contained signals due to an olefinic proton [δH 7.09 (dq, J =

1.7, 7.1 Hz, H-14)], four oxygenated methine [δH 3.40 (dd, J = 4.7, 11.5 Hz, H-1); 5.20 (m, H-11 and H-12); 5.57 (dt, J = 3.9, 2.8 Hz, H-6)], five tertiary methyl groups [δH

0.93, 1.01, 1.29, 1.37 and 2.08 (each 3H, s, H-18, H-19, H-20, H-17 and H-22)] and a secondary methyl group [δH 2.04 (dd, J = 7.1, 0.5 Hz, H-15)] The 13C-NMR spectrum contained twenty two signals There were two carboxyl groups [δC 168.4 (C-16) and 169.3 (C-21)], one trisubstituted double bond [δC 128.1 (C-13) and 144.9 (C-14)], and five oxygenated carbons [δC 70.4 (C-6); 72.9 (C-12); 78.0 (C-1); 80.0 (C-11); and 81.9 (C-8)] These data suggested a tetracyclic diterpenoid structure, similar to that of pallavicinin D

The gross structure of pallavicinin E was established using HMQC and HMBC experiments (Table 3-4), and since the process is the same as previous compound, pallavicinin D, it will not be discussed in detail here However, the major HMBC correlations are shown in Fig 3-13

O

O

OHO

17

7

11 12 13

14 15

16

18 19

20

H

HH

Fig 3-13: Observed HMBC correlations of (252)

The relative stereochemistry was established by NOEDIFF experiments (Table 3-4), and the only difference between pallavicinin D and E lies at the chiral C-12 Irradiation

at H-1 caused NOE effects at H-3α, H-5, H-9, and H-12, therefore H-1, H-5, H-9 and

H-12 were syn (H-1 and H-12 were anti in pallavicinin D) Irradiation at H-18 caused

NOE enhancements at 1 and 6 indicated that 6 was also on the same face as

H-1 Irradiation at Me-20 caused NOE enhancements at H-11, Me-17, and Me-19;

therefore Me-17, Me-20 and H-11 was syn

Biosynthesis of this compound should be similar to that of 251 The only difference is

in the stereochemistry of C-12 when the epoxide is formed

Table 3-4 1H-NMR (500 M Hz), 13C-NMR (125 M Hz), HMBC and NOEDIFF

spectral data of compound (252) in CDCl3

Pallavicinin F

The minor compound, pallavicinin F (253), was obtained as a colorless amorphous

powder, [α]D 4.2 (c 0.06, CHCl3) It was determined to have the molecular formula

C20H32O4 by HR-EIMS (m/z M+ 336.2294)

O

O OH

O

1

9 11 12 13 14 15 16

17

18

H H

19

H 20

H

(253)

The 1H-NMR spectrum contained signals due to three oxygenated methine protons [δH

3.42 (dd, J = 3.9, 11.5 Hz, H-1); 4.51 (dd, J = 3.7, 7.4 Hz, H-12); and 5.08 (dd, J =

7.3, 9.1 Hz, H-11)], four tertiary methyl groups [δH 0.83, 0.87, 0.95 and 1.22 (each 3H,

s, H-17, H-12, H-19, and H-16)] and a primary methyl group [δH 1.06 (t, J = 7.3 Hz,

H-15)] The 13C-NMR spectrum contained twenty signals There was a carboxyl group [δC 177.0 (C-16) and four oxygenated carbons [δC 77.5 (C-1); 79.4 (C-12); 80.9 (C-11); and 83.0 (C-8)] NMR and MS data suggested that this compound is a tetracyclic diterpenoid

HMQC and HMBC experiments (Table 3-5) were used to determine the structure of this compound Structure of ring A was built similar to previous compound and this is where the similarity ended Structure of ring B was established starting from the

tertiary methyl Me-17 Since Me-17 showed correlations to C-7, C-8 and C-9, it must

be attached to C-8 C-9 is next to C-10, due to the fact that it showed 3J correlation to

Me-20 The methylene H-6 correlated to 8, while H-7 correlated to 5, therefore

C-6 must be in between C-7 and C-5 C-11 is attached to C-9, since its proton showed HMBC correlation to C-10 The primary methyl H-15 showed correlations to C-13 and C-14 (a methine and a methylene, respectively), showed that C-14 is in between C-13 and C-15 The observed cross-peaks of both H-14 and H-12 with C-16 indicated that both C-12 and C-16 must be attached to C-13 Since the oxygenated methine H-11 also correlated to C-16, ring D must be a five-membered lactone ring

O

OOH

O

13

17

1819

20

H

Fig 3-14: Observed HMBC correlations of (253)

The stereochemistry was established by NOEDIFF experiments (Table 3-5) Since irradiation at Me-20 caused NOE enhancements at H-11 and Me-17; therefore M-20,

Me-17 and H-11 were syn H-12 must be on the same face as H-11, since NOE

enhancements were observed at H-11 and Me-17 when H-12 was irradiated; this also

suggested that both H-13 and H-9 were anti to H-11 and H-12 The fact that proton

signals of H-1 has one diaxial coupling and it did not show NOE enhancement when

Me-20 was irradiated, suggested that H-1 and Me-20 were anti Similarly, Me-20 and

irradiated In addition, splitting pattern of one H-6 proton showed three large couplings (13.5 Hz), one of these coupling must be diaxial coupling with H-5, therefore H-5 is an axial proton The configurations of Me-20 and H-5 showed that the A/B ring junction

is trans

O O

O Me

12 11

20

17

Fig 3-14a: Key NOE correlations of (253)

Table 3-5 1H-NMR (500 M Hz), 13C-NMR (125 M Hz), HMBC and NOEDIFF spectral

Symphyogynolide (254) from the Venezuelan liverwort Symphyogyna brasiliensis 153

also has the same skeleton as 253 Structure of 253 bears a striking similarity to that of

the scapanins such as (217-218), which were isolated from the liverwort Scapania

undulata.122,128 Thus, pallavicinin F is probably derived from scapanin skeleton via lactonization

O

O

H H

H O

O

(254)

(-)-3,14-clerodadien-13-ol [ = (-)-kolavelool ]

Compound (255), (-)-3,14-clerodadien-13-ol, was obtained as a yellowish oil, [α]D -8.1

(c 0.21, CHCl3) (lit.140 –25.7) It was determined to have the molecular formula

groups [δH 1.57 (3H, d, J = 1.7 Hz), 1.27, 0.98, and 0.71 (each 3H, s)], and a secondary

methyl group [δH 0.77 (3H, d, J = 6.3 Hz, H-17)] The 13C-NMR spectrum contained twenty signals There were four olefinic carbons [δC 111.7, C-15; 120.4, C-14; 144.5, C-4 and 145.3, C-3) and one oxygenated carbon [δC 73.5, C-13) Based on MS and NMR data, it was clear that this compound is a bicyclic diterpenoid The structure of

(255) was determined to be (-)-kolavelool by comparison of its 1H and 13C-NMR data with those of the literature.140

Previously, (-)-kolavelool has been found in P subciliata,152,154 Macrolejeunia,120Nardia subclavata,125 Pleurozia,117,140 and Scapania species.135 However, it was not only confined in liverworts, in fact, it was originally isolated from higher plants.111,155

Many clerodanes with similar stereochemistry (256-265) has been isolated from