Medicinal Plant Research in Africa. Pharmacology and Chemistry

891 Pages·2013·16.503 MB·English

by  Victor Kuete (Eds.)

1
Monoterpenes and Related
Compounds from the Medicinal
Plants of Africa
Michel Kenne Tchimenea, Christopher O. Okunjia,c,
Maurice Mmaduakolam Iwua and Victor Kueteb
aInternationalCentreforEthnomedicineandDrugDevelopment,Nsukka,
Nigeria,bDepartmentofBiochemistry,UniversityofDschang,Dschang,
Cameroon,cUSPHeadquarter,Rockville,MD
1.1 Introduction
Monoterpenes are a class of terpenes that consist of two isoprene units and have
the molecular formula C H . They are predominantly products of the secondary
10 16
metabolism of plants, although specialized classes occur in some animals and
microorganisms, and are usually isolated from the oils obtained by steam distilla-
tion or solvent extraction of leaves, fruits, some heartwoods, and, rarely, roots, and
bark [1]. In favorable cases they occur to the extent of several percent of the wet
weight of the tissue. Conjugated nondistillable forms, e.g., terpene-β-D-glucoside,
are also frequently found, especially in the floral organs. They are the most repre-
sentative molecules, constituting 90% of the essential oils, and have a great variety
of structures. Monoterpenes may be linear (acyclic) or they may contain rings.
Biochemical modifications such as oxidation or rearrangement produce the related
monoterpenoids. They are known for their many biological activities such as anti-
microbial, hypotensive, antiinflammatory, antipruritic, antigerminative, antiplasmo-
dial, antiesophageal cancer, and anticandidal. The compounds are inexpensive and
have been widely used in flavoring and fragrances since the beginning of the nine-
teenth century. More recently, they have played a great role in the pharmaceutical
industry because of their potential. Monoterpenes are also included in the category
of nutraceuticals, which represent an industry in excess of US$75.5 billion with
prospectsofgrowingtoUS$167billionby2010[1].
1.2 Biosynthesis and Structural Diversity
Modernmethods ofseparation and structure determination, aswell as the advent of
radioisotopetechniques,haveledtoaveryrapidadvanceinknowledgeoftheroute
ofbiosynthesisofthisclassandtheothertypesofterpenoidsoverthelast30years.
MedicinalPlantResearchinAfrica.DOI:http://dx.doi.org/10.1016/B978-0-12-405927-6.00001-1
©2013ElsevierInc.Allrightsreserved.
2 MedicinalPlantResearchinAfrica
Several reviews, of differing completeness, have outlined the routes to terpenoids
and steroids [2(cid:1)8] in general and monoterpenes in particular [9,10]. One important
conclusion that emerges is the accuracy with which chemical theory can predict
the course of the biochemical processes. Enzymes exploit the innate reactivity of
their substances, and the biosynthetic routes can be dissected into unit steps such
as elimination, electrophilic addition, and Wagner(cid:1)Meerwein rearrangement that
are controlled by the stereoelectronic factor known to operate in nonbiological
systems. Even the reactivity of apparently nonactivated atoms can usually be
rationalized in terms of conformational and electronic changes imposed by
postulated substrate(cid:1)enzyme or substrate(cid:1)cofactor linkages. The well-established
patterns found can be used to asses feasible structures for novel terpenoids and to
designbiogenetic-typesynthesis.
1.2.1 Biosynthetic Pathways
1.2.1.1 Isoprene Rule
Theearliestattempttorationalizethe patternofstructuresofthemonoterpeneswas
the rule proposed by Wallach in 1887, who envisaged such compounds as being
constructed from an isoprene unit (1) (Figure 1.1). Thirty years later, Robinson
extended this isoprene rule by pointing out that in monoterpenes, and such higher
terpenes as were then known, the units were almost invariably linked in a head-
to-tail fashion, as shown for limone (2) and camphor (3). However, many higher
terpenes and a few monoterpenes were later found not to obey this amended rule,
andRuzickaandhiscollaborators[11,12]proposed abiogeneticisoprenerule.This
generalization, which is now universally accepted, states that naturally occurring
terpenoids are derived either directly or by way of predictable stereospecific cycli-
zation, rearrangement, and dimerizations form acyclic C-10, C-15, C-20, and C-30
precursors geraniol, farnesol, geranylgeraniol, and squalene, respectively. This rule
implies a common pathway of biosynthesis for the whole family and any proposal
forirregularbiogeneticroutesmustbetreatedwithreservations.
Although isoprene has been formed on pyrolytic decomposition of some mono-
terpenes, it is not found in plants, and much speculation has occurred around the
nature of the active isoprene of the condensing unit, ranging from apiose to tiglic
acid. The C-5 unit was also postulated to arise from degradation of carbohydrates,
proteins,aminoacids,andmanyotherclassesofplantmetabolitesorbyelaboration
of acetic acid, ethyl acetoacetate, or acetone. These early views have been well
Figure1.1 Chemicalstructureofisopreneunit(1),
limone(2),andcamphor(3).
O
1 2 3
MonoterpenesandRelatedCompoundsfromtheMedicinalPlantsofAfrica 3
summarized [13,14]. Many C-10 compounds have been implicated as progenitors
of monoterpenes including citral [15], geraniol [16], nerol [17], limonene [18],
linalool [19], ocimene [20], and others [21(cid:1)24]. None of these speculations were
backedbyexperimentalevidenceofanykind.
1.2.1.2 Acyclic Compounds and Cyclohexane Derivatives
1.2.1.2.1 Hypotheses
TheproposalsofRuzickaandhiscoworkers[11]forthepatternofmonoterpenebio-
genesis are outlined in Figure 1.2. Several of the intermediates are formally repre-
sented as carbonium ions, but structurally equivalent species such as alcohols,
phosphateesters,terpeneglycosides,orsulfoniumsalts,eitherfreeorbondedtopro-
teins, may bethereactantsinvivo.Theschemeisextremelyattractive;theformation
OPP
OH
4 5 6
OPP
9
7 8
OH
Borane Pinane 11
OH 10 skeleton skeleton Carane Thujane
skeleton skeleton
Figure1.2 Formationofacyclicmonoterpene:myrcene(4),ctronellol(5),cis-ocimene(6),
α-terpineol(10),andterpinen-4-ol(11).
4 MedicinalPlantResearchinAfrica
of acyclics such as myrcene (4), citronellol (5), or cis-ocimene (6) from geranyl
pyrophosphate(GPP)hasmanyinvitroanalogies,andmonocyclizationoftheion(7)
formedfromnerylpyrophosphate(NPP)togiveα-terpineol(10),orterpinen-4-ol(11)
isalsochemicallyreasonable,althoughthebiochemicaldetailsareopentoconjecture.
Forthelatterprocess,eitherepoxides(whichhavebeenisolatedfromseveralessential
oils)[25]orsulfoniumcompoundsformedwithathiolgroupofanenzyme[26]may
beinvolvedasoutlinedinEq.(1.1)and(1.2).Bothofthesetypesofintermediatesare
knowntobeimplicatedintheformationofringsinhigherterpenoids,andinteresting
modelsystemsforthesynthesisofmonoterpenesinvitrousingsulfoniumylideshave
beendeveloped;the elucidationoftheimportance(ifany)ofsuchroutesintheplant
mustawaittheadventofsuitablecell-freesystems.
H
ð1:1Þ
O H OH
S Enz + HS Enz ð1:2Þ
H
H
Bicyclic skeletons of the pinane and borane series are (according to Ruzicka’s
scheme)derivedbyinternaladditionsofpositivecenterstodoublebondswithinmono-
cyclic frameworks in a direction governed either by electronic factors (Markovnikov
addition) or by steric factors. Hydride shift within the ion (8) followed by cyclization
of 9 gives rise to the thujane skeleton, and that of the caranes arises from an internal
electrophilic substitution at the allylic position of the former carbonium ion. This
latter reaction, as given, is biochemically improbable, and an internal displacement
(Figure1.3)inanintermediatesuchas12(X5ester)ortheintermediaryofanonclassi-
cal ion (13) has been suggested [27], but both proposals beg the question. A study of
themechanismofdecompositionofcertainunsaturatedepoxidessuggeststhatEq.(1.3)
X
12
13
Figure1.3 Suggestedinternaldisplacementinanintermediateinmonocyclicmonoterpenes.
MonoterpenesandRelatedCompoundsfromtheMedicinalPlantsofAfrica 5
is feasible and the mechanism could be modified to form other bicyclic monoterpenes
directlyfromacyclicprecursors;cf.Eq.(1.4).Thegenerationoftheintermediate
OH
OH
O ð1:3Þ
EnzSH NADPH
ð1:4Þ
O
SEnz SEnz
carbenes, or their formal equivalents, may be possible at the enzyme surface where
water and other potential scavengers may be locally excluded. No evidence is
availabletoassessthesehypotheses.
1.2.1.3 Cyclopentane Derivatives
1.2.1.3.1 General
Iridoids (Figure 1.4) are a family of compounds based on carbon skeleton (14) that
can be regarded as being formed by cyclization of 15. They were originally isolated
from the defensive secretions of Iridomyrmex, a genus of ant [28,29], but are now
known to be widely distributed in higher plants, usually, but not invariably, as the
OH
CHO
O O OR
c
CHO o
O
HO MeO C
2
OH
14 15 16
CHO 10 HO
OG OG
O
HO 9
8 1 O O
O 7 2
CO Me 5
2 3
6 HO
4
17
CO2Me 19
11
G=-β-D-glucose
18
Figure1.4 Structureofsomeiridoids.
6 MedicinalPlantResearchinAfrica
β-D-glucosides. Several hundred iridoids and related compounds have been isolated
fromleaf, seed, fruit,bark, and roottissueofdicotyledons. Thiswidespreaddistribu-
tion in plant tissues may be a consequence of the water solubility endowed by the
sugar residue, and contrasts with the storage and retention in specialized oil glands
of the largely water-insoluble monoterpenes of the types considered previously.
Few systematic studies of chemotaxonomy have been made [30], although a simple
fieldtestisavailabletodetectiridoids.
A decade ago it was suggested [31,32] that tetrahydropyranmethycyclopentane
monoterpenes of this then unusual type were possible biogenetic precursors of the
indolealkaloids; similar proposalswere madefor the formationofoleuropeine (16)
and elenolide (17). More recent work has amply confirmed these speculations, and
there is little doubt that loganin, or a close-related compound, does fulfill these
roles. Most of the biosynthetic studies on the iridoids have been concerned with
their function as intermediates en route to indole alkaloids, and it is only recently
thatthesemonoterpeneshavebeguntobestudiedintheirownright.
Loganin is also an intermediate in the biosynthesis of other iridoids and of
secoiridoids formed by rearrangement and functionalization of the skeleton (14)
[33,34]. Its agluconeis unstable and the sugar moietymay playa solubilizing, trans-
port-facilitating, and, very importantly, protective role; in particular, it may protect
the C -linked hydroxyl group (for numbering of the ring see 18; alternative systems
1
are sometimes used) from oxidation until the appropriate stage in the biosynthetic
scheme,whenthesugarresidueiscleavedoff.
Thefusedbicyclicsystemofloganinaccountsfor8ofthe10carbonatomsderived
fromtheacyclicmonoterpeneprecursor.Oneoftheremainingcarbonisabsentinsome
compounds that cooccur with, and are undoubtedly related to, the iridoids, although
there is no formal biosynthetic demonstration for these relationships saved in the case
of aucubin (19) [35]. Unedoside (20) (Figure 1.5) [36] is the only compound so far
characterized that has lost both peripheral carbons; none have been reported which
havelosttheC methylgroupbutnottheC carboxylgroup,whereasincontrastsev-
10 11
eralfamiliesofcompoundshavelostthelattergroupbutretainedtheformer,e.g.,aucu-
bin (19) and catalposide (21); R5p-hydroxybenzol) [37,38]. Secoiridoids such as
gentiopicroside(22)[39]maybederivedfromloganinoraclose-relatedcompoundby
cleavageoftheC7(cid:1)C8bondyieldinginitially,inthecaseofloganinitself[40],secolo-
ganin(23)[41].Theisolationofcompoundssuchasfoliamenthin(24)[42],sweroside
(25) [43], and ipecoside (26) [44], as well as biosynthetic studies, provide further evi-
dencethatthesegroupsofcompoundsarebiogeneticallyrelated.Otherrelativesarethe
alkaloids β-skytanthine (27) [45] and actinidine (28) [46]; most of these compounds
occur as their glucosides, but in addition to those described, genipin (29) and a few
others appear to be presenting plant tissues as their aglycones. A diglucoside and a
thioesterareamonginterestingiridoidsthathaverecentlybeencharacterized.
All the biosynthetic studies on this group of compounds have depended on
investigation of the fate in intact plant tissue of specifically labeled and carefully
chosen precursors, and these have often been supplemented by the isolation of
suspectedintermediatesfromthetissue.Onlyafewplantspecieshavebeeninvesti-
gated,especiallyyoungshootsofVincaroseaorCatharanthusroseus.
MonoterpenesandRelatedCompoundsfromtheMedicinalPlantsofAfrica 7
OG OHOG OG
O O
O O O
HO RO
20 21
O O
22
OG OG
O OH O
OHC
OG
CO2Me CO O O O
23 O O G=β-D-glucose
O 25
24
OHOH
HO
NAc N N
HO
OG CO2Me
29
O 27 28
MeOC
2
26
Figure1.5 Chemicalstructureofunedoside(20),captaposide(21),gentiopicroside(22),
secologanin(23),foliamenthin(24),sweroside(25),ipecoside(26),β-skytanthine(27),
actinidine(28),andgenipin(29).
Whereas the broad outlines of the biosynthetic pathways have undoubtedly been
unveiled,someoftheminordetailsmaybespeciesoreventissuespecific.Forexam-
ple, differences in labeling patterns between the same compound found in the leaves
andflowersmayoccur.Generallytheinfluenceofthis,andofotherphysiologicalpara-
meters, on biosynthetic routes has been ignored, but studies on the formation
ofverbenalin(30)(Figure1.6),β-skytanthine(27),andnepetalactone(31)havedemon-
stratedthecriticalimportanceofthesefactorsmayhaveonlabelingpatterns.Thesame
substratemayalsobeaneffectiveprecursorofaparticulariridoidinoneplantspecies
but not in another; for example, whole and sliced rhizomes of Menyanthes trifoliata
didnotincorporate[2-14C]MVAintologanin,whereasinV.roseatheadditivewasan
efficient and specific precursor [47]. Data based on several different experimental
approachesorproceduresarethusdesirableforinvestigationofanyonespecies.
Experiments using the 4R and 4S isomers of [2-14C, 4-3H ]MVA have confirmed
1
that the stereospecificity of formation of the two double bonds of geraniol used in
loganin formation is similar to that found in terpene synthesis in general, and that
directcondensationofisopentenylpyrophosphate(IPP)withdimethylallylpyrophos-
phate(DMAPP)togivenerolratherthangeranioldirectlyalsodoesnotoccurinthis
class of compounds. Geraniol, GPP, or some other derivative such as the enzyme-
bound intermediate previously discussed, appears to be an obligatory precursor. The
use of (1R)- and (1S)-[2-14C,1-3H ]GPP has demonstrated that conversion of the C
1 1
carbon into an aldehydic or equivalent oxidation level is also stereospecific, and the
hydrogens at rogens atC and C geraniolare retainedduring its transformation into
2 6
8 MedicinalPlantResearchinAfrica
OG O OH
O
O O
OG
O O
CO Me O
2
G= β-D-glucose 31
30 CHO Me
2
32
OH
N
O
MeO N OCOMe
HO CO Me
2
33
34
Figure1.6 Chemicalstructureofverbenalin(30),nepetalactone(31),plumieride(32),
iridodial(33),andloganin(34).
loganin.However,ifsaturationoftheC /C doublebondofgeraniolisaprerequisite
2 3
for the formation of loganin, then both reduction and subsequent removal of the
addedprotonoccurinastereospecificfashion[48,49].
The occurrence of foliamenthin (24) and related compounds also suggests that
oxidation of the isopropylidene group in geraniol is essential for its conversion into
loganin.However,evidencefromtheincorporationofdoublylabeledmevalonicacid
(MVA) into indole alkaloids suggests that incorporation of the intact propylidene
unitofgeranioltakesplace.Suchfindingsarenowreconciledbyourknowledgethat
oxidation occurs at both C and C of geraniol (Figure 1.7) and that equilibration
9 10
ofthesetwocarbonsofgeranioloccursduringthebiosynthesisofloganinandrelated
compounds from geraniol. Thus early studies [50] on the biosynthesis of plumieride
(32) [51]provedthat duringits formation fromgeraniolthe C and C atoms ofthe
9 10
latterbecamebiosyntheticallyequivalent,for25%ofthelabelpresentwaslocatedat
thestarredatomsin32when[2-14C]MVAwasusedasaprecursor.Asimilarpattern
in loganin (18) was obtained with the same precursor and with [3-14C]MVA, and
analogous results have been reported for all the iridoids, secoiridoids, and indole
alkaloids that have been studied. To account for the pattern in plumieride, iridodial
(or irodial)(33) was proposedasanintermediate, but this compoundis not a precur-
sor of loganin or vindoline (34) in V. rosea [52]. The equilibration of carbon atoms
equivalentto C and C ofgeraniolmay, however,not always be complete and can
9 10
vary with the physiological condition of the plant used. However, the point was
made that asymmetric labeling of the part of the molecule derived from IPP, com-
mon for the monoterpenes described in the previous section, is not as widespread a
phenomenon for these cyclopentane derivatives. 10-Hydroxygeraniol (35) and
10-hydroxynerol (36) (using the accepted numbering) have recently been shown to
be precursors of loganin and of the indole alkaloid, and a reasonable route for loga-
nin biosynthesis can be summarized in Figure 1.7. Complete randomization of 14C
label from C and C of 35 was observed. Several related monoterpenes—linalool,
9 10
MonoterpenesandRelatedCompoundsfromtheMedicinalPlantsofAfrica 9
HO CH3 4
HOH2C CO2H H5H3 2 C1H2OH
HA HD HB CH2OH
HB HC 9 810(Hc, HD)
OH
a OH a b
OH
OH
OH
OHC CHO OH CHO
35 37 36
a b
OG
HOHC7 HB OH HH 7 HOGO CHCOHO b OCHHO
HB18Co2R (Hc, HD) 41 38HCO2R Co2R
G=β-D-glucose
H H H and H refer to the 4S, 4R, 2R, and 2S hydrogens, respectively of MVA
A, B, C, D
Figure1.7 Formationofcyclopentanederivatives.
citronellol,and citral—were notsignificantlyincorporated.These resultssuggest that
a further step after 35 and 36 in the biosynthesis of iridoids involves attack on C
9
of 35 or 36 (or of the corresponding aldehydes) to give a hypothetical species such
as 37 (route a, Figure 1.7). It is not known whether C or C is oxidized first, or
5 10
if indeed there is a specific order. 10-Hydroxynerol was a more efficient precursor
than its isomer, and this suggests that the immediate precursors of the iridoids and
indole alkaloids [53] have the cis double bond at C and C that is expected on
2 3
stereochemical grounds. The rate of isomerization of this double bond may play an
important roleindiverting GPP fromits alternative functionasa precursor ofhigher
terpenoids. It is also possible that cyclization may proceed prior to further oxidation
at C of 10-hydroxynerol (route b, Figure 1.7). The only other intermediates that
9
havebeendemonstratedbetweengeraniolandloganinorloganicacidaredeoxyloga-
nin and deoxyloganic acid, respectively ((38), R5Me, H), and both have been
shown to be specific precursors of loganin [54]. Deoxyloganin occurs together with
loganininV.roseaandStrychnosnux-vomica[55].Neithertheagluconeofdeoxylo-
ganin nor the isomers with the double bonds at the C /C or C /C positions were
6 7 7 8
incorporated into the final product. The final stage of loganin [56] biosynthesis is
therefore envisaged as hydroxylation of deoxyloganin at C , which data on loganic
7
acid biosynthesis suggest is stereospecific, like other biological hydroxylations. Both
10 MedicinalPlantResearchinAfrica
deoxyloganin and loganic acid occur in V. rosea, and a cell-free system from this
plant can convert the acid into loganin; thus a dual pathway is suggested in which
methylation can occur at different points (Figure 1.8). Similar and more complicated
metabolicgridshavebeenobservedinthebiosynthesisofotherterpenoids,especially
carotenoids,andotherswillbementionedshortly.
Recent work on loganic acid and gentiopicroside [57(cid:1)59] biosynthesized
from 14C and 3H doubly labeled isomers of MVA and geraniol has confirmed the
formation of geraniol and hence of the cyclopentane derivatives from MVA. The
stoichiometry of boththe decarboxylation of MVAPP to give IPP and of the addi-
tion of IPP to DMAPP to give GPP is similar to that reported previously for other
terpenoids and steroids. Deviations from the expected 14C/3H ratio of activities
of C of loganic acid were found that were similar to those reported in steroid
7
synthesis. Such results have been accounted for by the relatively slow rate of
removal of DMAPP by prenyl transferase as compared to the rate of establishing
the equilibrium between IPP and DMAPP by IPP isomerase. Conversion of
DMAPP into IPP in the latter equilibration would result in a partial loss of asym-
metry ofthe 3H/1HpairatC of IPP.
2
No preferential labeling of the two isoprene units of loganic acid was observed.
However, such patterns can occur at the monoterpene level; formation of menthia-
folin, a hydroxylated isomer of 24, from [2-14C]geraniol gave a product in which
the two C-10 moieties were labeled in the ratio of 3:1. This finding suggests that
either the monoterpene or its constituent units may be synthesized in different
pools, which may correspond to intra- and extrachloroplastic sites of synthesis
(both of which sites contain terpene synthesizing enzymes). The pools may be
connected at the monoterpene-glucoside level as these compounds are water
soluble. However, the stage at which glucose is coupled to a monoterpene remains
unknown;presentevidence suggeststhat itis notthe final step inloganin oriridoid
biosynthesis. The earlier findings indicate that iridoids may pass through several
intra- and extracellular compartments during biosynthesis, and the distribution of
iridoids in all types of plant tissues may provide further evidence for such tortuous
pathways. The changes in labeling pattern at C and C of certain iridoids and
3 11
related compounds dependent on the age of the plant material may also be related
to the need for the biosynthetic scheme to occur at several distinct sites. Indeed,
theobserved14C/3HisotoperatiosofactivitiesofC ofloganicacidbiosynthesized
7
from 4R and 4S isomers of [2-14C,-4-3H ]MVA that have been discussed earlier
1
may be the result of incomplete randomization at the two positions, since the
OG Figure1.8 Chemicalstructureofloganinderivative.
O
O
CO Me
2
G=β-D-glucose
41