estudo quÍmico das espÉcies psychotria nuda cham
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ESTUDO QUÍMICO DAS ESPÉCIES Psychotria nuda Cham. & Schltdl. E Psychotria suterella Müll. Arg. (RUBIACEAE) E AVALIAÇÃO DE
ATIVIDADES BIOLÓGICAS
UNIVERSIDADE ESTADUAL DO NORTE FLUMINENSE DARCY RIBEIRO
CAMPOS DOS GOYTACAZES-RJ
FEVEREIRO-2018
ALMIR RIBEIRO DE CARVALHO JUNIOR
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ESTUDO QUÍMICO DAS EPÉCIES Psychotria nuda Cham. & Schltdl. E Psychotria suterella Müll. Arg. (RUBIACEAE) E AVALIAÇÃO DE
ATIVIDADES BIOLÓGICAS
“Tese apresentada ao Centro de Ciência e
Tecnologia da Universidade Estadual do
Norte Fluminense Darcy Ribeiro, como parte
das exigências para obtenção do título de
Doutor em Ciências Naturais”
Orientador: Prof. Ivo Jose Curcino Vieira
Co-Orientador: Prof. Raimundo Braz-Filho
Co-Orientador: Prof. Mário Geraldo de Carvalho
Campos dos Goytacazes-RJ
Fevereiro-2018
ALMIR RIBEIRO DE CARVALHO JUNIOR
iii
Carvalho Junior, Almir Ribeiro de
Estudo químico das espécies Psychotria nuda Cham. & Schltdl. e Psychotria suerlla Müll.
Arg. (Rubiaceae) e avaliação de atividades biológicas / Almir Ribeiro de Carvalho Júnior. –
Campos dos Goytacazes, 2018.
x, 206 f. : il.
Tese (Doutorado em Ciências Naturais) -- Universidade Estadual do Norte
Fluminense Darcy Ribeiro. Centro de Ciência e Tecnologia. Laboratório de
Ciências Químicas. Campos dos Goytacazes, 2018.
Orientador: Ivo José Curcino Vieira.
Coorientação: Raimundo Braz-Filho.
Área de concentração: Bio-orgânica.
Bibliografia: f. 201-205.
1. Psychotria 2. IRIDÓIDES 3. ALCALÓIDES 4. MTT I. Universidade Estadual do
Norte Fluminense Darcy Ribeiro. Centro de Ciência e Tecnologia. Laboratório de
Ciências Químicas lI. Título
CDD
583.93
iv
ESTUDO QUÍMICO DAS EPÉCIES Psychotria nuda Cham. & Schltdl. E Psychotria suterella Müll. Arg. (RUBIACEAE) E AVALIAÇÃO DE
ATIVIDADES BIOLÓGICAS
“Tese apresentada ao Centro de Ciência e
Tecnologia da Universidade Estadual do
Norte Fluminense Darcy Ribeiro, como parte
das exigências para obtenção do título de
Doutor em Ciências Naturais”
Aprovada em: ___/___/___
Comissão Examinadora:
__________________________________________________
Prof. Daniela Barros de Oliveira (D.Sc., Química de Produtos Naturais)-UENF
__________________________________________________
Prof. Edmilson José Maria (D.Sc., Química)-UENF
__________________________________________________
Prof. Antônio Sérgio Nascimento Moreira (D.Sc., Ciências Naturais)-IFF
__________________________________________________
Prof. Ivo Jose Curcino Vieira (D.Sc., Química)-UENF
(Orientador)
ALMIR RIBEIRO DE CARVALHO JUNIOR
v
Aos meus pais, Irani e Almir, irmãos, Cristiano, Adriano e
Fabiano e Noiva, Rafaela, pelo amor e incentivos.
vi
AGRADECIMENTOS
Aos Professores Drs. Ivo Jose Curcino Vieira, Raimundo Braz-Filho e Mário Geraldo
de Carvalho pela orientação, ensinamentos e contribuição na minha formação
profissional e moral.
Aos professores membros da banca por dedicarem seus tempos à melhoria do
trabalho.
Aos amigos do laboratório, deixo meus agradecimentos pelas contribuições e por
tornarem a rotina laboratorial bem mais agradável.
Pelos experimentos de RMN, infravermelho e espectrometria de massas, agradeço a
Márcio e Marcelo.
Registro minha gratidão aos professores Drs. Francisco José Alves Lemos, Milton
Masahiko Kanashiro e Gil Rodrigues dos Santos pelos ensaios biológicos.
vii
SUMÁRIO
RESUMO .............................................................................................................................. ix
ABSTRACT ........................................................................................................................... x
1. INTRODUÇÃO ................................................................................................................... 1
2. REVISÃO DA LITERATURA .............................................................................................. 2
2.1 Informações sobre a família Rubiaceae e o gênero Psychotria .................................... 2
2.2 Composição química do gênero Psychotria .................................................................. 3
2.2.1 Alcaloides pirrolidinoindólicos ................................................................................ 3
2.2.2 Alcaloides indólicos monoterpênicos (AIM) ............................................................ 5
2.3 As espécies Psychotria nuda e Psychotria suterella ..................................................... 7
3. TRABALHOS ..................................................................................................................... 9
3.1 trabalho 1: psychotria genus: chemical constituents, Biological Activities And Synthetic Studies ............................................................................................................................... 9
3.1.1 Introduction ............................................................................................................ 9
3.1.2 Chemical Constituents ......................................................................................... 10
3.1.2.1 Dimeric and Polyindoline Alkaloids Isolated from Psychotria Species. .............. 11
3.1.2.2 Monoterpene Indole Alkaloids Isolated from Psychotria Species. ...................... 14
3.1.2.3 Other classes of Alkaloids Isolated from Psychotria Species ............................. 20
3.1.2.4 Triterpenoids from Psychotria Species .............................................................. 21
3.1.2.5 Other classes of Metabolites from Psychotria Species ...................................... 23
3.1.2 Biological Activities .............................................................................................. 27
3.1.3 Synthesis Of Some Compounds From Psychotria Species .................................. 33
3.1.4 Concluding Remarks ............................................................................................ 36
3.1.5 Abbreviations ....................................................................................................... 36
3.1.6 References .......................................................................................................... 37
3.2 Trabalho 2: 13C-NMR Spectral Data of Alkaloids Isolated from Psychotria Species (Rubiaceae)...................................................................................................................... 40
3.2.1 Introduction ................................................................................................................. 40
3.2.2 Discussion .................................................................................................................. 41
3.2.2.1 13C Chemical Shifts of Monoterpene Indole Alkaloids Isolated from Psychotria Species. ........................................................................................................................ 41
3.2.2.2 13C Chemical Shifts of Pyrrolidinoindoline Alkaloids Isolated from Psychotria Species. ........................................................................................................................ 56
3.2.2.3 13C Chemical Shifts of Benzoquinolizidine Alkaloids Isolated from Psychotria Species. ........................................................................................................................ 67
3.2.3 Conclusions ......................................................................................................... 70
3.2.4 Acknowledgments ................................................................................................ 70
3.2.5 Author Contributions ............................................................................................ 70
viii
3.2.6 Conflicts of Interest .............................................................................................. 71
3.2.7 References .......................................................................................................... 71
3.3 Trabalho 3: Metabolites from Psychotria suterella Müll. Arg. and Psychotria nuda Cham. & Schltdl (Rubiaceae) and Evaluation of Cytotoxic Activity ................................... 77
3.3.1 Introduction .......................................................................................................... 78
3.3.2 Results and discussion ........................................................................................ 79
3.3.2.1 Metabolites from P. suterella ............................................................................. 79
3.3.2.2 Metabolites from P. nuda .................................................................................. 79
3.3.2.3 Assessment of cell viability by MTT assay ........................................................ 81
3.3.3 Experimental ........................................................................................................ 82
3.3.3.1 Apparatus and instruments ............................................................................... 82
3.3.3.2 Plant material .................................................................................................... 82
3.3.3.3 Extraction and isolation ..................................................................................... 82
3.3.3.4 P. suterella ........................................................................................................ 82
3.3.3.5 9-epi-geniposidic acid ....................................................................................... 83
3.3.3.6 P. nuda ............................................................................................................. 83
3.3.3.7 Culture of cells .................................................................................................. 84
3.3.3.8 MTT assay ........................................................................................................ 84
3.3.4 Conclusion ........................................................................................................... 85
3.3.5 Disclosure statement ........................................................................................... 85
3.3.5 Acknowledgements .............................................................................................. 85
3.3.6 Funding ................................................................................................................ 85
3.3.6 References .......................................................................................................... 85
3.3.7 Supplemental online material ............................................................................... 88
4. CONCLUSÕES ....................................................................................................... 201
5. REFERÊNCIAS BIBLIOGRÁFICAS ........................................................................ 201
6. ANEXOS ................................................................................................................. 205
6.1 Metodologia do Ensaio Inseticida .......................................................................... 205
6.2 Metodologia do Ensaio Antifúngico ....................................................................... 206
ix
RESUMO
CARVALHO JUNIOR, Almir Ribeiro; D.Sc. Universidade Estadual do Norte
Fluminense Darcy Ribeiro. Fevereiro de 2018. Estudo Químico das Epécies
Psychotria nuda Cham. & Schltdl. e Psychotria suterella Müll. Arg. (Rubiaceae) e
Avaliação de Atividades Biológicas.
Espécies do gênero Psychotria (Rubiaceae) são reconhecidas por seus usos na
medicina popular e pela produção de metabólitos com potencial biológico. Em face
disto, objetivou-se com esta pesquisa realizar o estudo químico das espécies P.
nuda e p. suterella e avaliar atividades biológicas de seus extratos, frações e
compostos isolados. Das folhas e galhos de P. nuda foram isolados e identificados
dezessete compostos, que são: sitosterol, estigmasterol, campesterol, fitol, -
sitosterol e -estmasterol glucosilados, cinchonaina Ia, cinchonaina Ib, N,N,N-
trimetiltriptamônio, lialosídeo, lawsofrutose, roseosídeo, estrictosamida,
escopoletina, ácido rotungênico, estrictosidina e 5-carboxiestrictosidina. Um novo
iridoide inédito, ácido 9-epi-geniposídico, foi identificado das folhas de P. suterella,
juntamente com ácido geniposídico, sacarose, ácido 3-O-acetiloleanólico, ácido
pomólico, ácido espinósico, ácido maslínico, ácido tormêntico, metil oleanolato,
ácido lialosídico e ácido estrictosidínico. Neste trabalho foram avaliadas as
atividades inseticida, frente às larvas do mosquito Aedes aegypti, antifúngica, frente
aos fungos Fusarium oxysporum, Curvularia lunata, Colletotrichum musae,
Rhizoctonia solani, and Sclerotium rolfsii, e citotóxica frente às células cancerígenas
das linhagens THP-1 e U937. Os extratos e frações testados nos dois primeiros
ensaios não apresentaram resultados promissores. O alcaloide estrictosamida,
dentre os compostos testados, foi o que apresentou os melhores resultados quanto
a citotoxidade, com valores de EC50 de 120,0 ± 1 e 21,9 ± 1 g/mL frente a células
THP-1 e U937, respectivamente.
x
ABSTRACT
CARVALHO JUNIOR, Almir Ribeiro; D.Sc. Universidade Estadual do Norte
Fluminense Darcy Ribeiro. Fevereiro de 2018. Estudo Químico das Epécies
Psychotria nuda Cham. & Schltdl. e Psychotria suterella Müll. Arg. (Rubiaceae) e
Avaliação de Atividades Biológicas.
Psychotria species are recognized by their use in folk medicine and by the production
of biologically actives metabolites. Owing to it, the aim of this research was to
perform the chemical study of P. nuda and P. suterella as well as to assess biological
activities of its extracts, fractions and isolated compounds. Seventeen compounds
were isolated and identified from leaves and twigs of P. nuda, named sitosterol,
stigmasterol, campesterol, phytol, -sitosterol-3-O--D-glucoside, -stmasterol-3-O-
-D-glucoside, cinchonain Ia, cinchonain Ib, N,N,N-trimethyltryptamonium, lyaloside,
lawsofrutose, roseoside, strictosamide, scopoletin, rotungenic acid, strictosidine, and
5-carboxystrictosidine. The new iridoid named 9-epi-geniposidic acid, along with the
known compounds geniposidic acid, sucrose, 3-O-acethyloleanolic acid, pomolic
acid, spinosic acid, maslinic acid, tormentic acid, methyl oleanolate, lyalosidic acid,
and strictosidinic acid (11) were isolated and identified from leaves of P.suterella. In
this work, insecticidal, against Aedes aegypti larva, antifungal, against Fusarium
oxysporum, Curvularia lunata, Colletotrichum musae, Rhizoctonia solani, and
Sclerotium rolfsii, and citotoxic, THP-1 and U937 cancer cell lines, activities were
assessed. Extracts and fractions tested in the first two assays did not show promising
results. The alkaloid strictosamide, among the tested compounds, showed relevant
citotoxicity with IC50 values 120 ± 1 and 21.9 ± 1 g/mL, against THP-1 and U937 cell
lines, repectively.
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1. INTRODUÇÃO
A utilização de produtos naturais, principalmente da flora, com fins medicinais,
nasceu com a humanidade. Registros do uso de plantas medicinais e tóxicas datam
das civilizações mais antigas, sendo considerada uma das práticas mais remotas
empregadas para cura, prevenção e tratamento de doenças, atuando como
importante fonte de substâncias biologicamente ativas (Firmo et al. 2011).
Nos últimos anos, registrou-se um aumento expressivo no interesse em
substâncias derivadas de espécies vegetais, micro-organismos, insetos e
organismos marinhos. Newman & Cragg (2016) destacam que a prática da utilização
de produtos naturais e seus derivados estruturais na descoberta e desenvolvimento
de novos fármacos ainda está viva e progredindo. Por exemplo, na área de câncer,
considerando-se o período de 1940 até o final de 2014, das 175 pequenas
moléculas aprovadas, 131 ou 75% eram não sintéticas, com 85 ou 49% sendo
produtos naturais ou derivados diretamente deles.
O crescente interesse em novas substâncias biologicamente ativas está
diretamente relacionado à riqueza da biodiversidade. O Brasil ocupa posição
privilegiada em termos de biodiversidade, em diferentes aspectos. Considerando-se
apenas o restrito universo de espécies catalogadas no mundo, o país detém a maior
quantidade total (13%) e a segunda maior quantidade de espécies endêmicas em
valores absolutos. O território brasileiro é composto por sete biomas principais:
Amazônia, Cerrado, Caatinga, Mata Atlântica, Pampa, Pantanal e Zona Costeira e
Marinha. Desses, Mata Atlântica e Cerrado são exclusivos do território brasileiro
(Pimentel et al. 2015). Entretanto, poucas espécies da flora nativa foram
investigadas do ponto de vista químico e farmacológico.
Espécies do gênero Psychotria, pertencente à família Rubiaceae, se
destacam por sua importância na medicina tradicional, onde são utilizadas para uma
grande variedade de indicações terapêuticas como afecções do aparelho reprodutor
feminino, distúrbios gastrointestinais, úlceras, “tumores”, distúrbios oculares, no
tratamento de febres, dores de cabeça e ouvidos, sendo ainda empregadas em
rituais religiosos devido a suas propriedades alucinógenas (Faria 2009; Lima 2011).
O uso medicinal destas espécies estimulou a avaliação do potencial farmacológico
de extratos, frações semi-purificadas e substâncias isoladas, destacando-se
propriedades antivirais, anti-inflamatórias, antibióticas, antifúngicas, antitumorais,
dentre outras (Carvalho Junior et al. 2016).
2
O gênero Psychotria é considerado de taxonomia complexa, em função das
poucas características morfológicas diferenciadoras. Por esse motivo, alcaloides
indólicos são considerados marcadores químicos, importantes para os estudos
quimiotaxônomicos de espécies deste gênero (Porto et al. 2009; Carvalho Junior et
al. 2017).
A potencialidade químico-farmacológica descrita para o gênero Psychotria
justifica os estudos químicos e biológicos de espécies deste gênero. Neste contexto,
objetivou-se com esta pesquisa realizar o estudo químico das espécies Psychotria
nuda e Psychotria suterella e avaliar as atividades inseticida, antifúngica e citotóxica
de extratos, frações e de algumas das substâncias isoladas.
2. REVISÃO DA LITERATURA
2.1 Informações sobre a família Rubiaceae e o gêner o Psychotria
A família Rubiaceae é composta por mais de 600 gêneros, totalizando
aproximadamente 13000 espécies, distribuídas pelo mundo (Rydin et al. 2009;
Barbhuiya et al. 2014). Suas espécies são classificadas em quatro subfamílias:
Cinchonoideae, Ixorideae, Antirheoideae e Rubiodeae, na qual o gênero Psychotria
é incluído (Tomaz et al. 2008). Este gênero possuí mais de 2000 exemplares
encontrados principalmente em regiões tropicais e subtropicais do globo, sendo
considerado o maior da família Rubiaceae (Oliveira et al. 2013). Com base em
características morfológicas e distribuição geográfica, o gênero Psychotria é dividido
em três subgêneros: Psychotria (pantropical), Heteropsychotria (espécies
neotropicais) e, Tetramerae (algumas encontradas na Africa e Madagascar) (Moraes
et al. 2011).
Alguns relatos têm apontado que espécies deste gênero têm sido utilizadas na
medicina popular como alternativa de tratamento de diversas doenças. Flores de P.
colorata, por exemplo, são usadas por caboclos da Amazônia para tratamento de
dor de ouvido, enquanto que seus frutos são empregados em casos de dores
abdominais (Verotta et al. 1998). Na Malásia, folhas de P. rostrata são empregadas
para o tratamento de constipação (Takayama et al. 2004). Infecção intestinal, tosse,
distúrbios respiratórios e estomacais são outros exemplos de doenças combatidas
pelo uso de outras espécies do gênero (Benevides et al. 2004). O uso
etnofarmacológico de suas espécies, provavelmente, estimulou o desenvolvimento
3
de diversas pesquisas voltadas à investigação química e avaliação do potencial
biológico de seus metabólitos.
2.2 Composição química do gênero Psychotria
Em relação à composição química do gênero, vários trabalhos vêm sendo
realizados, destacando-o como uma potencial fonte de alcaloides. Aproximadamente
53 % dos metabólitos isolados de suas espécies são alcaloides, dois quais 87 % são
do tipo indólico. Esta classe de metabólitos secundários apresenta papel
fundamental no ponto de vista quimiotaxômico. Alcaloides pirrolidinoindólicos são
característicos de espécies do subgênero Psychotria (Lopes et al. 2004), enquanto
que alcaloides indólicos monoterpênicos são marcadores quimiotaxômicos do
subgênero Heteropsychotria (Kerber et al. 2008). Triterpenos (12 %) e flavonoides (6
%) são outras classes de metabólitos frequentemente isolados do gênero.
2.2.1 Alcaloides pirrolidinoindólicos
Os alcaloides pirrolidinoindólicos são caracterizados pela presença de várias
unidades de N-metiltriptamina em suas estruturas (Lopes et al. 2004), cujas
diferentes unidades apresentam, comumente, ligações do tipo C-3a/C-3a’ e C-3a/C-
7’ (Takayama et al. 2004). Os alcaloides deste tipo isolados do gênero têm
apresentado de dois a sete meros. Psicotridina, quadrigemina C e hodgkinsina são
os alcaloides mais comumente isolados de diversas espécies (Verotta et al. 1999)
(Hart et al. 1974) (Libot et al. 1987) (Roth et al. 1986) (Verotta et al. 1998) (Adjibade
et al. 1992) (Zhou et al. 2010).
Além da importância no ponto de vista quimiotaxônomico, estes alcaloides
despertaram interesse de pesquisadores no que tange às suas propriedades
biológicas. Diversos estudos têm apontado uma série de atividades biológicas, como
é o caso de quadrigemina B. Este alcaloide isolado da espécie P. rostrata
apresentou atividade citotóxica frente a células HEp-2 e atividade antibacteriana
frente a Escherichia coli e Staphulococcus aureus (Mahmud et al. 1993).
Psicotridina, isolado de P. forsteriana, também apresentou citotoxidade contra
células leucêmicas (Adjibade et al. 1989). Atividades analgésica (Amador et al.
2000) e antiparasitária são outros exemplos de atividades biológicas apresentadas
por este tipo de metabólitos (Muhammad et al. 2003). A Figura 1 apresenta alguns
exemplos de alcaloides pirrolidinoindólicos isolados de espécies Psychotria.
4
N
N NH
NH
H Me
Me
NN
HMe H
N N
H MeH
N
NHMe
NN
HMe
H
N N
H Me
N
N
H
Me
N
N
H
Me
NN
HMeN
NH
Me
N N
H MeH
N
N
H
Me
H
N
N
H
Me
H
NN
HMe H
N N
H MeH
N
N
N
Me
N
Me
NN
HMe H
NN
HMe H
N N
H MeH
N
NH
Me
H Me
N
NN
NN
N
MeH
N
NMeH
N
N
NNH
Me
H
MeH
Figura 1 : Alcaloides pirrolidinoindólicos isolados do gênero Psychotria.
Psicotridina
(Verotta et al. 1999)
Quadrigemina C
(Verotta et al. 1998)
Psicoleina
(Rasolonjanahary et al. 1995)
Psicotridiasina
(Zhou et al. 2010)
(+) Quimonantina
(Verotta et al. 1999)
Psicohenina
(Liu et al. 2014)
Psicotrimina
(Takayama et al. 2004)
Hodgkinsina
(Hart et al. 1974)
Psicotripina
(Li et al. 2011)
5
2.2.2 Alcaloides indólicos monoterpênicos (AIM)
Este tipo de alcaloide é característico de espécies encontradas no território
brasileiro. Sua biossíntese envolve reação entre a triptamina (oriunda do triptofano)
e o iridoide secologanina, levando a formação da estrictosidina. A diversidade
estrutural destes metabólitos está relacionada, pricipalmente, a modificações
envolvendo N1 e C-22, N4 e C-22. Os alcaloides correantinas A-C (Achenbach et al.
1995) exemplificam o primeiro caso enquanto que estrictosamida representa um
caso de ciclização entre N4 e C-22 (Faria et al. 2010). Oxidação de C-10 não é tão
comum, porém, 10-hidroxi-iso-depeaninol e 10-hidroxi-antirhina, isolados de P.
prunifolia (Kato et al. 2012) e 10-hidroxi-correantosideo (P. Correa) são exemplos de
alcaloides com esta característica.
N,-D-glucopiranosilvincosamida, isolado das folhas de P. leiocarpa,
apresentou característica peculiar por ser considerado, segundo os autores, o
primeiro relato de AIM N-glicosilado (Henriques et al. 2004). Bahienosídeos A e B,
isolados de P. bahiensis e P. acuminata, são exemplos raros de alcaloides que
apresentam uma porção terpênica adicional ligada ao N4 (Berger et al. 2012) (Paul et
al. 2003). A Figura 2 apresenta exemplos de AIMs isolados do gênero.
N
N
H
HO
H H OH
H
N
N
OHHH
H
HO
HO
N
NMeH
O
O
OGlcH
HH
HO
N
N
O
O
HOGlu
H
Glu
10-hidroxi-correantosídeo
(Achenbach et al. 1995)
10-hidroxi-iso-depeaninol
(Kato et al. 2012)
10-hidroxi-antirhina
(Kato et al. 2012)
N,-D-glucopiranosilvincosamida
(Henriques et al. 2004)
6
N
NMeH
O
HOCO2Me
H
HMe
H
N
NMeH
O
OH
MeH
H CHO
N
NH C
OH
OGlc
H
H
O2Me
N
NMeH
O
HCHO
H
HOH
Figura 2 : Acaloides indólicos monoterpênicos isolados do gênero Psychotria.
Correantina A
(Achenbach et al. 1995)
Correantina B
(Achenbach et al. 1995)
Correantina C
(Achenbach et al. 1995)
Estrictosidina
(Berger et al. 2012)
7
Glc
N
N
H
O
O
O
Glc
Glc
CO2Me
O
O
H
H
N
N
H
H
O2Me
O
C
H
H OGlc
Glc
CO2Me
O
O
H
H
N
N
H
H
O2Me
O
C
H
H O
N
N
O
O
H
H
OGlc
H
Figura 2 : Continuação.
2.3 As espécies Psychotria nuda e Psychotria suterella
A espécie P. nuda (Figura 3 ), conhecida popularmente como casca d’ anta, é
encontrada, pricipalmente, na forma de arbustos, medindo de 1 a 5 metros de altura
(Miguel et al. 2009) principalmente nos estados do Rio de Janeiro, Minas Gerais, até
o estado de Santa Catarina (Ferreira et al. 2014).
A espécie P. suterella (Figura 4 ) vulgarmente conhecida como grandiuva-de-
anta cafezinho-roxo-da-mata, apresenta porte arbustivo-arbóreo, podendo alcançar
até 6 m de altura (Lopes & Buzato 2005). O período de floração desta espécie
ocorre de janeiro a março e frutificação de setembro a maio (Bertani 2006), períodos
que facilitam a sua identificação, viabilizando sua coleta.
Bahienosídeo B
(Berger et al. 2012)
Bahienosídeo A
(Paul et al. 2003)
Estrictosamida
(Van De Santos et al. 2001)
Estaquiosídeo
(Pimenta et al. 2010)
8
Figura 3 : Fotografia da espécie P. nuda.
Fonte: https://www.flickr.com/photos/gustavolf/7291965490
(Acessado dia 27/01/2018)
Figura 4 : Fotografia da espécie P. suterella.
Fonte: http://www.ufrgs.br/fitoecologia/florars/open_sp.php?img=11745
(Acessado dia 27/01/2018).
Em relação à química destas espécies, há relatos escassos a esse respeito.
Referente à espécie P. nuda há o relato apenas de isolamento de um alcaloide
indólico monoterpênico: estrictosamida (Farias et al. 2008). Já das folhas de
P.suterella há o relato de identificação do alcaloide mencionado anteriormente,
outros dois: lialosídeo, e naucletina (Van De Santos et al. 2001).
9
3. TRABALHOS
3.1 Trabalho 1:
Psychotria Genus: Chemical Constituents, Biological Activitie s And
Synthetic Studies**
Almir Ribeiro de Carvalho Junior1, Mario Geraldo de Carvalho2, Raimundo Braz
Filho1,2, Ivo Jose Curcino Vieira1*
1. Laboratório de Ciências Químicas, Centro de Ciências e Tecnologia, Universidade
Estadual do Norte Fluminense Darcy Ribeiro, Campos dos Goytacazes, RJ-28013-
602, Brazil
2. Departamento de Química, ICE, Universidade Federal Rural do Rio de Janeiro,
Seropédica, RJ-23890-000, Brazil
Abstract: Natural products have been used by humankind for thousands of years in applications such as pigments, flavourings, and drugs. Since antiquity, the use of natural products has been the best or the only alternative adopted by many people worldwide, in the treatment of several diseases. In fact, plants are a potential source of bioactive compounds, but most of the world’s biodiversity has not been evaluated for any biological activity. In this context, several studies have been performed regarding the chemical composition and biological properties of various species from different genera such as Psychotria L. (Rubiaceae). This genus is the largest of the Rubiaceae family, comprising about 2000 species, mainly found in tropical and subtropical regions of the globe. Several works have been reported concerning the chemical composition and biological activities of species of this genus. The aim of this overview is to summarise the advances in knowledge on Psychotria species, compiling reports related to chemical composition and biological activities of the genus.
** Trabalho publicado como capítulo do livro Studies i n Natural Products
Chemistry, volume 48, páginas 231-261, do ano 2016, doi: 10.1016/B978-0-
444-63602-7.00007-2.
Introduction
Natural products have been employed in the treatment of several diseases for
thousands of years [1]. Even recently, despite availability of synthetical drugs, plants
remain widely used for medicinal purposes [2]. The diversity of biologically active
compounds in plants has motivated chemical studies of various species. Recently,
many plant-derived drugs, including semi-synthetics compounds, have either been
10
introduced to the market or are involved in clinical trials [3,4], highlighting the
importance of medicinal plants in drug discovery.
The family Rubiaceae Juss. comprises 620 genera totaling about 13526 species,
distributed worldwide [5]. Psychotria L. is the largest of the Rubiaceae, possessing
more than 2000 species, mainly found in tropical and subtropical regions [6]. Based
on its morphological features and geographical distribution, the genus was divided
into three subgenera: Psychotria (pantropical), Heteropsychotria (neotropical
species) and, Tetramerae (some African and Madagascan species) [7]. However,
Nepokroeff et al. (1999) proposed the reorganization of the genus based on a
molecular phylogenetic study [8].
Several Psychotria species are widely used in folk medicine around the world for
the treatment of various illnesses. Flowers and fruits of P. colorata, for example, are
used by “caboclos” from the Amazon to treat earache and abdominal pain,
respectively [9]. In Malaysia, leaves of P. rostrata are employed for the treatment of
constipation [10]. P. viridis is used as an ingredient in the hallucinogenic beverage
called ayahuasca [11], owing to the presence of N,N-dimethyltriptamine, an indole
alkaloid structurally related to neurotransmitter serotonin [12]. Intestinal infections,
coughs, respiratory and stomach disorders are other examples of illnesses which
have been treated using other Psychotria species [13].
Chemical Constituents
Many studies have examined the chemical composition of the species of the genus
Psychotria (Rubiaceae). Since 1974 several works have shown that Psychotria is a
potential source of alkaloids. Approximately 52 % of the metabolites reported were
characterised as alkaloids (about 87 % belong to the subgroup of indole alkaloids),
followed by triterpenes (12 %), flavonoids (6 %), along with constituents of other
classes. Since Psychotria is taxonomically complex, alkaloids can be an important
tool to distinguish its species from others which belong to genera with similar features
such as Cephaelis Sw. and Palicourea Aubl. [14]. Moreover, these metabolites have
shown a range of biological activities, increasing interest in the study of this genus,
with the aim of discovering new natural medicines.
11
Dimeric and Polyindoline Alkaloids Isolated from Psychotria Species.
The main alkaloids found in pantropical Psychotria (subgenus Psychotria) are
polyindole alkaloids, which are characterized by the presence of several N-methyl
triptamine moieties in their structures [15], such as psychotridine (1). This alkaloid,
derived from five N-methyltriptamine units, was isolated from P. beccarioides, P.
forsteriana, P. oleoides, and P. colorota [16-19]. Quadrigemine C (2) is another
example of a polyindole alkaloid identified in this genus having been isolated from P.
colorata and P. oleoides [9,16,18,20–22].
Four polyindoline alkaloids, named quadrigemines A (4) and B (5), psychotridine
(1), and isopsychotridine C (6), were isolated from leaves of P. forsteriana [19, 23].
Besides these compounds, meso-chimonathine (7), a dimeric indole alkaloid, was
also isolated from the same species. It was the first isolation of a dimeric isomer of
calycanthine, which are commonly present in the genus Calycanthus, from
Psychotria [24].
The chemical study of P. rostrata leaves led to the isolation of two new alkaloids:
psychopentamine (8) and psychotrimine (9). Compound 8 was the first example of a
polymeric pyrrolidinoindoline alkaloid which contains a C-3a/C-5’ bond. This group of
compounds generally display two types of common linkages: C-3a/C-3a’ bond and C-
3a/C-7’ bond [10].
Other examples of polyindoline alkaloids isolated from Psychotria, along with the
compounds mentioned above, are summarised in Table 1 and their structures are
shown in Fig. (1).
Table 1 . Dimeric and polyindoline alkaloids isolated from Psychotria species Compound Species Part Reference
Psichotridine (1) P. forsteriana
P. oleoides
P. colorata
P. beccarioides
Leaves
Leaves
Leaves
Leaves
[16–19]
Quadrigemine C (2) P. colorata P. oleoides
Flowers and leaves
Leaves
[9, 15, 18, 20–22]
Psycholeine (3) P. oleoides Leaves [21]
12
Quadrigemine A (4) P. forsteriana Leaves [19]
Quadrigemine B (5) P. forsteriana P. colorata P. rostrata
Leaves
Leaves
Leaves and twigs
[16, 19,25]
Isopsychotridine C (6) P. forsteriana Leaves [19]
Meso-chimonanthine (7) P. forsteriana P. muscosa
Not specified
Leaves
[16, 24]
Psichopentamine (8) P. rostrata Leaves [10]
Psychotrimine (9) P. rostrata Leaves [10]
Psichotriasine (10) P. calocarpa Leaves [26]
Hodgkinsine (11) P. colorata P. oleoides P. lyciiflora P. muscosa; P. beccarioides P. rostrata
Flowers and leaves
Leaves
Leaves
Leaves
Branches and twigs
[9, 16, 17, 22, 25]
(+)-Chimonanthine (12) P. colorata P. muscosa P. rostrata
Flowers
Leaves
Branches and twigs
[9, 16, 25]
(13) P. henryi Leaves and twigs [27]
(14) P. henryi Leaves and twigs [27]
Nb-demetyl-meso-Chimonantina (15)
P. lyciiflora Leaves [22]
Psychohenin (16) P. henryi Leaves and twigs [28]
Quadrigemine I (17) P. oleoides Leaves [22]
Isopsychotridine B (18) P. oleoides Leaves [18, 22]
Oleoidine (19) P. oleoides Leaves [22]
Caledonine (20) P. oleoides Leaves [22]
Psychotripine (21) P. pilifera Leaves [29]
13
N N
H MeH
N
N
H
Me
H
N
N
H
Me
H
NN
HMe H
(2)
N N
H MeH
N
N
N
Me
N
Me
NN
HMe H
(3)N N
H Me
N
N
H
Me
N
N
H
Me
NN
HMeN
NH
Me
(1)
NN
H
Me
NN
H
Me
NN H
Me
N
NH
Me
N
N
H
Me
N
N
Me
H
(6)N N
H Me
N
N
H
Me
N
N
H
Me
NN
HMe
(4) N
N
H
Me
N
N
H
Me
N
N
H
Me
NN
HMe
NNHMe
(5)
N N
NN
MeH H
H MeH
(7)
N
NMeH
N
N
NNH
Me
H
MeH
(9)
Me
N
NMe
HN N
HMe
N
N
H
N
N
H
Me
N
N
H
Me
(8)
(12)
NN
HMe H
N N
H MeH
N
NHMe
NN
HMe
H
(10)
NN
HMe H
N N
H MeH
N
NH
Me
H
(11)
Fig. (1). Dimeric and polyindoline alkaloids isolated from Psychotria species
14
N N
Me
N
H
(13)
N N
MeH
N
N Me
H
(14)
N N
H Me
NN
HMe NN H
Me
N
N
H
Me
(17) (21)
Me
N
NN
NN
N
MeH
N N
H Me
NNMe N
N H
Me
n = 3 (18)n = 4 (19)n = 5 (20)
N N
H H
NNHMe
(15)
N
N NH
NH
MeH
Me(16)
H
n
Fig. (1) continued
Monoterpene Indole Alkaloids Isolated from Psychotria Species.
Monoterpene indole alkaloids (MIAs) are biosynthesised by the coupling of
tryptophan and the iridoid terpene secologanin. These kinds of metabolites have
exhibited several biological activities, being used, for example as anti-cancer, anti-
malaria and anti-arrhythmia agents [30]. MIAs are the main type of alkaloids found in
the subgenus Heteropsychotria (neotropical species) [31], being considered
chemotaxonomic markers for this subgenus [32].
The chemical investigation of P. correae led to the isolation of seven MIAs, along
with other kinds of metabolites. Among them, six alkaloids were reported for the first
time: correantoside (22), 10-hydroxycorreantoside (23), correantine A (24),
correantine B (25), 20-epi-correantine B (26), and correantine C (27) [33].
Henriques et al. (2004) reported the isolation of N--D-glucopyranosyl
vincosamide (28) from leaves of P. leiocarpa. It was the first report of a N-
glycosylated monoterpenoid indole alkaloid. The authors also concluded that the
accumulation of this alkaloid depends on the age of the plant and light exposure,
being restricted to the aerial parts of P. leiocarpa [34].
The chemical study of P. brachyceras led to the isolation of a new MIA named
brachycerine (29). According to Nascimento et al. (2013), this alkaloid belongs to a
new subclass of MIAs since its terpenoid moiety is probably derived from epiloganin
rather than secologanin, as is the usual case. Subsequent studies also showed that
15
the concentration of brachycerine (29) increases on UV-B radiation exposure and
osmotic/oxidative stress, suggesting that this compound may play a role in plant
defence mechanisms [35,36].
From leaves of P. umbellata, an unusual alkaloid named psychollatine (30) was
isolated. This compound is mainly accumulated in aerial parts of the plant but low
amounts were also found in its roots [37,38]. Other studies regarding P.
umbellatahave beenmade leading to the isolation of three new MIAs: 3,4-Dehydro-
18,19--epoxy-psychollatine (31), N4-[1-((R)-2-hydroxypropyl)]-psychollatine (32),
and N4-[1-((S)-2-hydroxypropyl)]-psychollatine (33) [39].
Table 2 provides information concerning these compounds, as well as other MIAs
isolated from Psychotria species. Their structures are displayed in Fig. (2).
Table 2 . Monoterpene indole alkaloids isolated from Psychotria species
Compound Species Part Reference
Correantosideo (22) P. correae Leaves [33]
10-hidroxicorreantosideo (23) P. correae Leaves [33]
Correantine A (24) P. correae Leaves [33]
Correantine B (25) P. correae Leaves [33]
20-epi-correantine B (26) P. correae Leaves [33]
Correantine C (27) P. correae Root [33]
N--D-glucopiranosil vincosamide (28) P. leiocarpa Aerial parts [34]
Brachycerine (29) P. brachyceras Leaves [40]
Psichollatine (30) P. umbellata Leaves [37, 39]
3,4-Dehydro-18,19--epoxy-Psychollatine (31)
P. umbellata Leaves [39]
N4-[1-((R)-2-hydroxypropyl)]-psychollatine (32)
P. umbellata Leaves [39]
N4-[1-((S)-2-hydroxypropyl)]-Psychollatine (33)
P. umbellata Leaves [39]
5-carboxystrictosidine (34) P. acuminata P. bahiensis
Leaves
Aerial parts
[41, 42]
Bahienoside B (35) P. acuminata P. bahiensis
Leaves
Aerial parts
[41, 42]
Desoxicordifoline (36) P. acuminata Leaves [41]
Lagamboside (37) P. acuminata Leaves [41]
(E/Z)-vallesiachotamine (38 + 39) P. acuminata P. bahiensis
Leaves [41–44]
16
P. laciniata P. suterella
Aerial parts
Leaves
Leaves
Strictosidinic acid (40) P. acuminata P. barbiflora P. myriantha P. myriantha
Leaves
Leaves
Leaves
Aerial pars
[6, 41, 45,46]
Strictosidine (41) P. acuminata Leaves [41]
Palicoside (42) P. acuminata; Leaves [41]
Bahienoside A (43) P. bahiensis Aerial parts [42]
Angustine (44) P. bahiensis P. laciniata
Aerial parts
Leaves
[42, 43]
Strictosamide (45) P. bahiensis; P. nuda; P. prunifolia; P. suterella P. laciniata
Aerial parts
Leaves
Leaves
Leaves
Leaves
[14, 42, 44, 47, 48]
Isodolichantoside (46) P. correae Leaves [33]
10-hydroxy-iso-deppeaninol (47) P. prunifolia Branches [49]
10-hydroxy-antirhine (48) P. prunifolia Branches [49]
N-oxide-10-hydroxyantirhine (49) P. prunifolia Branches [49]
14-oxoprunifoleine (50) P. prunifolia Branches [49]
17-Vinil-19-oxa-2-azonia-12-azapentaci-clo[14.3.1.02,14.05,13.06,11]icosa-2(14),3,5(13),6(11),7,9-hex-aeno (51)
P. prunifolia Leaves [47]
17-vinil-19-oxa-2-azonia-12-azapentaci-clo[14.3.1.02,14.05,13.06,11]icosa-2(14),3,5(13),6(11),7,9-hex-aeno (52)
P. prunifolia Leaves [47]
N-demethyl-correantoside (53) P. stachyoides Leaves [50]
Naucletine (54) P. suterella Leaves [48]
Croceaine A (55) P. umbellata Leaves [37]
Umbellatine (56) P. umbellata Leaves [51]
Correantosine E (57) P. stachyoides Leaves [32]
Correantosine F (58) P. stachyoides Stem bark [32]
Stachyoside (59) P. stachyoides Aerial parts [52]
Nor-methyl-23-oxo-correantoside (60) P. stachyoides Aerial parts [52]
17
Lyaloside (61) P. laciniata P. suterella
Leaves [44]
Myrianthosine (62) P. myriantha Aerial parts [46]
R= H (22)R= OH (23)
N
NMeH
O
O
OGlcH
HH
R
N
NMeH
O
HOCO2Me
H
HMe
H
(24)
N
NMeH
O
OR1
MeH
R2H
R1 = H; R2 = CHO (25)R1 = CHO; R2 = H (26)
GlcN
N
O
O
HOGlu
H
(28)
N
N
OH
H
H CO2Me
O
HO
Glc
H
H
(29)
N
NMeH
O
HCHO
H
HOH(27)
Fig.(2) . Monoterpene indole alkaloids isolated from Psychotria species
18
(30)
N
N
GlcOH
H
CO2CH3
H
H
H
N
N
O
O
H
H
CO2Me
GlcOH
(31)
N
N
O
H
H
CO2Me
H
HO
GlcOH
N
N
O
H
H
CO2Me
H
HO
GlcOH
(32)
(33)
N
NH C
OH
OGlc
H
CO2H
H
O2Me
(34)
N
N C
OH
OGlc
H
H
O
OGlcMeO2C
H
O2Me
(35)
N
N C
CO2H
O
OGlc
H
H
O2Me
(36) (37) (38 + 39)
N
N
H
CO2Me
CHOH
N
NH C
OH
OGlc
H
H
O2H
(40)
GlcN
N
H
CO2Me
CH2OHH
N
NH C
OH
OGlc
H
H
O2Me
N
N C
OH
OGlc
H
H
MeO2H
(41) (42)
H
Fig.(2) continued.
19
(43)
Glc
Glc
O
O
H
H
MeO2C
N
N
H
H
O2Me
O
C
H
H O
(45)
Glc
N
N
H
O
O
OH
N
NMeH
OMeO2C OGlc
HH
(46) (48)
N
N
H
HO
H H OH
H
N
N
OHHH
H
HO
HO
(47)
N
N
N
H
O
(44)
ON
N
H O(49)
-
+N
N
H
HO
H H
O
OH
H
ON
N
H
(51)
(52)
N
N
N
H
O
O(54)
+
(50)
N
N
H
O
HO
H
H
Glc
N
N H
O
H
O O
H
H
H
(53)
Fig.(2) continued.
20
N
N H
GlcO H
H
CO2CH3
H
H
(55) (57)
(58)
H
N
N
OH
H
HOGlc
CO2CH3H
(56)
N
N
O
O
H
H
OGlcH(59)
N
N
O
O
H
H
OGlcH
H
O
(60)
N
N
OH
H OGlu
H3CO2C
H
(61)
N
NH
HO
O
OOGluH
(62)
Glc
N
N
O
O
H
O
H
Fig.(2) continued.
Other classes of alkaloids Isolated from Psychotria Species
Besides the above-mentioned compounds, other kinds of alkaloids have also been
reported for the genus Psychotria. From aerial parts of P. glumerulla, for example,
three new quinoline alkaloids, named glomerulatine A to C (63–65, Table 3 , Fig. 3 )
were isolated [53].
The chemical study of P. klugii led to the isolation of two new benzoquinolizidine
alkaloids: klugine (66) and 7’-O-demethylisocepaheline (67). In addition, cephaeline
(68), isocephaeline (69), and 7-O-methyllipecoside (70) were also isolated from stem
bark of P. klugii [54].
Table 3. Other classes of alkaloids isolated from Psychotria species. Compound Species Part Reference
Glomerulatine A (63) P. glumerulata Aerial parts [53]
Glomerulatine B (64) P. glumerulata Aerial parts [53]
Glomerulatine C (65) P. glumerulata Aerial parts [53]
Klugine (66) P. klugii Stem bark [54]
21
7'-O-demethylisocefaeline (67)
P. klugii Stem bark [54]
Cephaelina (68) P. klugii Stem bark [54]
Isocephaelina (69) P. klugii Stem bark [54]
7-O-methylipecoside (70)
P. klugii Stem bark [54]
Harmane (71) P. barbiflora P. suerrensis
Leaves [6, 55]
N
N
MeH
N
NN
N
M
Me
e
N
NN
N
H
MeN
NN
N
M
Me
He
(63) (66)(64)
N
NHO
MeO
H
H
OHHH
OMe
OH(66)
N
NMeO
MeO
H
H
HHH
OMe
OH
N
NMeO
MeO
H
H
HHH
OR1
R2OR1= R2= H (68)R1= Me, R2= H (69)
N
O
MeO
O H
COCH3
OH
HO
MeO
(67)
Glc
(70) (71)
Fig. (3) . Other classes of alkaloids isolated from Psychotria species.
Triterpenoids from Psychotria Species
Some studies have also reported the isolation of tritepenoids from Psychotria
species, such as psychotrianosides A to G (72–78) and other common compounds
as lupeol (83), betulin (84), friedelin (88), and so on. In Table 4 we can find
information regarding these and other triterpenoids and Fig. 4 shows their structures.
22
Table 4 . Triterpenoids isolated from Psychotria species.
Compound Species Part Reference
Psychotrianoside A (72) P. sp Whole plant [56]
Psycotrianoside B (73) P. sp Whole plant [56]
Psycotrianoside C (74) P. sp Whole plant [56]
Psycotrianoside D (75) P. sp Whole plant [56]
Psycotrianoside E (76) P. sp Whole plant [56]
Psycotrianoside F (77) P. sp Whole plant [56]
Psycotrianoside G (78) P. sp Whole plant [56]
Ardisianoside D (79) P. sp Whole plant [56]
Barbinervic acid (80) P. stachyoides Leaves [50]
-amirin (81) P. stachyoides P. adenophylla
Leaves
Leavese
[50, 57]
Ursolic acid (82) P. adenophylla
P. mariniana
Leaves [57, 58]
Lupeol (83) P. mariniana;
P. vellosiana
Aerial parts
[58, 59]
Betulin (84) P. adenophylla
P. mariniana
Leaves [57, 58]
Betulinic acid (85) P. adenophylla Leaves [57]
Bauerenol (86) P. adenophylla Leaves [57]
Bauerenol acetate (87) P. adenophylla Leaves [57]
Friedelin (88) P. adenophylla Leaves [57]
23
HO
OH
COOH
OH
H
H
(78)
(86) R= H (87) R= Ac
O
(88)
R4O
R2R1
O
R3H
H
R1 R2 R3
(81) H CH3 CH3 (82) CH3 H CO2H
HO
R3
R1
R2
HO
R
(83) R= H(84) R = OH (85) R = CO2H
RO
H
H
R1 R2 R3 R4 (72) CH2OH CH3 H Ara (73) CH3 CHO -OH -D-Xyl(1 2)-b-D-Glc(1 4)--L-Ara (74) CH3 CH3 =O -D-Xyl(1 2)-b-D-Glc(1 4)--L-Ara (75) CH3 CH3 -OH -L-Rha(1 4)--D-Glc --D-Xyl(1 2)--D-Glc--L-(1 4)-Ara (76) CH3 CH3 -OH -D-Xyl(1 4)--D-Glc(1 2)--D-Glc(1 2)--L-Ara (77) CH2OH CH3 -OH -D-Xyl(1 4)--L-Rha(1 2)--6-Acethyl-Glc(1 4)--L-Ara (78) CHO CH3 -OH -L-Ara (79) CH3 CH3 -OH -D-Xyl(1 2)--D-Glc(1 4)--L-Ara
Fig. (4) . Triterpenoids isolated from Psychotria species.
Other Classes of Metabolites from Psychotria Species
Other types of metabolites isolated from this genus are summarised in Table 5 and
their structures are displayed in Fig. (5).
Table 5 . Other classes of metabolites isolated from Psychotria species. Compound Species Part Reference
Blumenol A (89) P. yunnanensis Aerial parts [60]
Drummondol (90) P. yunnanensis Aerial parts [60]
3-hydroxy-5�, 6�-epoxi-7-megastimen-9-one (91)
P. yunnanensis Aerial parts [60]
Salicylic acid (92) P. yunnanensis Aerial parts [60]
Resorcinol (93) P. yunnanensis Aerial parts [60]
(-)-Loliolide (94) P. yunnanensis Aerial parts [60]
(6S)-Menthiafolic acid (95) P. yunnanensis Aerial parts [60]
4-hydroxybenzoic acid (96) P. yunnanensis Aerial parts [60]
Vanillic acid (97) P. yunnanensis Aerial parts [60]
Siringic acid (98) P. yunnanensis Aerial parts [60]
Ethyl protocatechuate (99) P. yunnanensis Aerial parts [60]
hydroxy-1-(3,5-dimethoxy-4hydroxyphenyl)propan-1-one (100)
P. yunnanensis Aerial parts [60]
24
-hydroxypropiovanillone (101) P. yunnanensis Aerial parts [60]
(-)-Butin (102) P. yunnanensis Aerial parts [60]
2-(4-hydroxy-3-metoxyphenil)-3-(2-hydroxy-5-metoxyphenyl)-3-oxo-1-propanol (103)
P. yunnanensis Aerial parts [60]
(+)-siringaresinol (104) P. yunnanensis Aerial parts [60]
Feoforbídeo A (105) P. acuminata Leaves [61]
Pirofeoforbídeo A (106) P. acuminata Leaves [61]
-sitosterol (107) P. adenophylla P. hainanensis P. mariniana; P. vellosiana
Leaves Leaves Aerial parts
[57–60, 62]
-sitosterol glycosylated (108) P. stachyoides Leaves [50]
Stigmasterol glycosylated (109) P. stachyoides Leaves [50]
Stigmasterol (110) P. vellosiana Aerial parts [59]
Psicotramida A (111) P. sp. Stem [63]
Psicotramida B (112) P. sp. Stem [63]
Psicotramida C (113) P. sp. Stem [63]
Psicotramida D (114) P. sp. Stem [63]
Psicorubrina (115) P. rubra Stem [64]
Stearic acid (116) P. hainanensis Leaves [62]
6-hydroxy-luteolin-7-O-rutinoside (117) P. rubra Aerial parts [65]
Luteolin-7-O-rutinoside (118) P. rubra Aerial parts [65]
Quercetin (119) P. hainanensis; P. spectabilis
Leaves
Leaves
[13, 62]
Kaempferol-7-O-glucopyranoside (120) P. hainanensis Leaves [62]
Kaempferol-3-O-glucopyranoside (121) P. hainanensis Leaves [62]
Rutin (122) P. hainanensis Leaves [62]
Psychorubrin (123) P. rubra Aerial parts [65]
6-hydroxygeniposide (124) P. rubra Aerial parts [65]
Daucosterol (125) P. hainanensis Leaves [62]
Psycacoraone (126) P. yunnanensis Aerial parts [66]
Scopoletin (127) P. vellosiana Aerial parts [59]
Squalene (128) P. vellosiana Aerial parts [59]
Cyclopsychotride A (129) P. longipes Whole plant [67]
Deoxysolidagenone (130) P. spectabilis Leaves [13]
Solidagenone (131) P. spectabilis Leaves [13]
25
Coumarin (132) P. spectabilis Leaves [13]
Umbelliferone (133) P. spectabilis Leaves [13]
Psoralene (134) P. spectabilis Leaves [13]
Benz[g]isoquinoline-5,10-dione (135) P. camponutans Wood [68]
1-hydroxybenzoisochromanquinone I (136)
P. camponutans Wood [68]
OH
O
OH
(89)
OH
O
H3CO
HO
OCH3(98)
OH
O
OCH3
HO
(97)
OHHO
(93)
OH
CO2H(92)
O
O
O
(91)
OH
O
OH
(90)
HO
O
H3COOH
OCH3
HO
O
H3COOH
HO
O
O
HO
(99)
CO2H
OH
(95)
HO
OH
O
(96)
OHO
O
(94)
Fig. (5) . Other classes of metabolites isolated from Psychotria species
26
O
OH
OH
HO
O(102)
OH OH
H3CO
OH
OCH3
O(103)
O
O
OCH3
OH
H3CO
OH
OCH3H3CO
(104)
NNH
HNN
RHO
O
O
RO
(107) R = H(108) R = Glu
(109) R = Glu,
(110) R = H R(105) CO2Me(106) H
(111) n= 16 (113) n= 15(112) n= 14 (114) n= 13
( )14
( )n
HO
NH
OH
OH
O
OH
O
O
O
OH
(115)
H3CCH2( )
O
OH16
(116)
O
O
R3
OH
R2
OH
R1O
R
R R1 R2 R3
(117) OH Rut; OH; H (118) H Rut; OH; H (119) H H OH; OH (120) H Glc H OH (121) H H H O-Rut(122) H H OH O-Rut
Fig. (5) continued
27
O
O
O
OOH
OHO
OH
OH
OH(123)
HO O
OOH
O
OHO
OH
OH
OH(124)
OO OH
O
(127) (128)
RO
(125)
OH
OHO
(126)
OR
O R(130) H(131) OH
O OR
R(132) H(133) OH
O OO
(134)
N
O
O(135)
O
OHO
O(136)
Fig. (5) continued
Biological Activities
Many studies have evaluated the biological properties of extracts, fractions, and
isolated compounds from Psychotria species. These plants have been shown to have
mostly cytotoxic, analgesic, antioxidant, and antimicrobial activities, as described in
the next sections.
Cytotoxic Activity
Roth et al. (1986) evaluated cytotoxic activities of four polyindoline alkaloids, isolated
from P. forsterianaon rat hepatoma cells (HTC line). Quadrigemines A (4) and B (5),
psychotridine (1), and isopsychotridine C (6) exhibited higher cytotoxity than
vincristine , used in antitumor chemotherapy.
CYS GLY
GLU SER
CYS
PRO
PHE
VAL
ILE
THR
VAL
ALA LEU LEU GLY
CYS
CYS
THR
CYS
LYS
SER
LYS
VAL
CYS
TYR
SER
SER
LYS
ASN
PRO ILE
(129)
28
N
H
N
MeO2C
OH
NMeO
N
OAcHHO CO2Me
OH
Vincristine
In this essay, the authors concluded that the concentrations necessary to promote
100 % cellular mortality (after 24 hours of incubation) were 2.5, 5, 5, and 10 �M for
compounds 1, 4, 6, and 5, respectively [14]. In a subsequent study, quadrigemine B
(5) also showed time- and dose-dependent cytotoxic activity against HEp-2 cells [69].
Hayashi, Smith and Lee (1987) reported that psychorubrin (123), a new
naphthoquinone isolated from P. rubra, showed cytotoxic activity in the KB cell assay
(ED50 = 3.0 g/mL). In addition, another four naphthoquinone derivatives (137–140)
were prepared as a way to establish its structure-activity relationships. All derivatives
exhibited higher cytotoxity than psychorubrin (ED50 ranging from 0.3 to 0.6 g/mL).
The authors concluded that extension of conjugation (observed for compounds 137
and 140) is not sufficient to increase cytotoxic activity, since compound 141 (another
naphthoquinone tested) was not active. Thus, other factors must be considered [64].
O
O
O
O
O
O
O
O
O
(137) (138) (139)
O
O
OO
O
O
(140) (141)
Fig. (6) . Structures of compounds 137–141
The in vitro cytotoxic activity of klugine (66), cephaelin (68), and isocephaelin (69),
isolated from P. klugii, was evaluated against four human cancer cells lines, SK-MEL,
KB, BT-549, and SK-OV-3. In this assay, doxorubicin and 5-fluorouracil were used
as positive controls, whereas DMSO was used as a negative control.
29
O
O
OOMe
OH
OH
OHO
OH
H O
Me
NH2
HO
Doxorubicin
N
N
OO
F H
H
5-fluorouracil
Compound 66 was more potent against these human cancer cell lines (IC50 values
of 0.25, 0.3, 0.86, and 0.18 g/mL, respectively) than doxorubicin (IC50 values of
1.57, 1.7, 1.0, and 1.3 g/mL, respectively). On the other hand, compounds 66 and
69 did not show cytotoxic activity against these cell lines [54].
Analgesic Activity
Aiming at discovering new painkillers, some researchers have investigated the
analgesic properties of extracts and isolated compounds (mostly alkaloids) from
Psychotria species, such as P. colorata, used by Amazonian Cablocos to treat
earache and abdominal pain. The analgesic activity of an alkaloid extract from P.
colorata was assessed by the formalin, writhing, and tail-flick methods, confirming the
opioid-like analgesic activity of this species [70]. In a subsequent study, it was
reported that the alkaloids from this plant exhibited inhibitory activity on [3H]naloxone
binding in rat striata membranes [71].
Other work related to P. colorata was carried out by Both et al. (2000) in order to
evaluate the analgesic activity of hodgkinsine (11), a major alkaloid isolated from this
plant. The authors concluded that this compound presented dose-dependent
analgesic activity in mice, probablymediated by opioid and glutamate receptors,
suggesting that it participates in the analgesia previously reported for P. colorata [72].
Umbellatine (56), an alkaloid from P. umbellata, is other example of a compound
which showed analgesic properties [51].
Both et al. (2002) evaluated the analgesic activity of alkaloid extracts from three
Psychotria species classified as P. myriantha, P. nuda, and P. pubigera. In this work,
it was reported that only P. myriantha showed this property (hot plate method) [73].
Antioxidant Activity
Fragosos et al. (2008) evaluated antioxidant and antimutagenic potentials of
psychollatine (30) and the crude foliar extract of P. umbellata. Antioxidant properties
30
were assessed in strains of Saccharomyces cerevisiae deficient in superoxide
dismutase and/or catalase (exposed to H2O2 and paraquat) and by the
hypoxanthine/xanthine oxidase assay. Psychollatine (30) was more efficient in
protection of strains treated with paraquat, whereas the crude foliar extract showed
better results for strains treated with H2O2. In the hypoxanthine/xanthine oxidase
assay both psychollatine (30) and the crude extract showed a marked dose-
dependent antioxidant activity, but the crude extract was more active (possibly owing
to the presence of flavonoids) than the isolated compound. Both the crude extract
and psychollatine (30) showed antimutagenic effects on strains of S. cerevisiae
(mutagenesis was induced by H2O2) [74]. Brachycerine (29), isolated from P.
brachyceras, is another example of an MIA which possessed antioxidant and
antimutagenic activities [75].
The in vitro antioxidant activity of fruits, stems, and leaf extracts of P. nilgiriensis
was investigated by DPPH, ABTS.+, and FRAP assays. An acetone extract of fruits,
had the highest total phenolics (505.74 �g GAE/g extract), tannin (460.78 g GAE/g
extract), and flavonoids (67.78 g RE/g extract). In addition, this extract presented
higher values for DPPH (IC50 = 20.0 g/mL), ABTS (41,343.51 mol TE/g extract),
and FRAP (4,713.33 mol Fe (II)/mg extract) assays [76].
Antimicrobial Activity
The antimicrobial activity of methanolic extracts of leaves, roots, and stem barks of P.
microlabastra against bacteria, protozoa, and fungi, was evaluated by the disk
diffusion method. In addition, fractions obtained by partition of these extracts with
petrol, dichloromethane, and ethyl acetate were also tested. In this work, the authors
reported that all extracts displayed activity against all bacteria and protozoa tested,
especially ethyl acetate fractions [77].
Three Psychotria species, along with other Rubiaceae and Meliaceae species,
were studied in order to investigate their antimicrobial properties by the disc diffusion
method. Extracts of leaves and bark of P. gardineri, P. nigra, and P stenophylla were
prepared using n-hexane, dichloromethane, and methanol as solvents and tested
against Saccharomyces cerevisiae, Ustilago maydis, Escherichia coli, Micrococcus
luteus, Bacillus subtilis, Bacillus cereus, and Aspergillus niger. P. gardineri and P.
stenophyllashowed a broad antimicrobial activity against six of the seven
31
microorganisms tested while P. nigra was active against four species [78]. The
biological activities reported for Psychotria species are summarized in Table 6 .
Table 6 . Biological activities reported for Psychotria species
Species Plant part Extract or compound Activity Ref erence
P. rubra Stem Psychorubrin Cytotoxic activity [64]
P. rostrata Bark and twigs Quadrigemine B Cytotoxic activity [69]
P. forsteriana Leaves Psychotridine,
auadrigemines A and B,
isopsychotridine, and
chimonanthine
Cytotoxic activity
[19,79]
P.
camponutans
Wood Benz[g]isoquinoline-
5,10-dione
1-
hydroxybenzoisochroma
nquinone
Cytotoxic activity
[68]
P. spectabilis Leaves Solidagenone and
psoralene
Cytotoxic activity [13]
P. colorata Leavesand
flowers
Aqueous and alkaloid
extracts
Analgesic activity [70, 71]
P. colorata Flowers Hodgkinsine Analgesic activity [72]
P.
brachypoda
Leaves ethanol extract Analgesic activity [80]
P. umbellata Leaves Umbellatine Analgesic activity [51]
P. myriantha Leaves Alkaloid extract Analgesic activity [73]
P. nilgiriensis Stem and fruit Acetone extract Analgesic and
antioxidant activities
[76]
P.
sarmentosa
Leaves and stems Aqueous extract Analgesic activity [81]
P. umbellata Leaves Methanol extract and
umbellatine
Antioxidant and
antimutagenic
activities
[74]
P. Leaves Methanol extract and Antioxidant and [75]
32
brachyceras brachycerine antimutagenic
activities
P. leiocarpa Leaves N,-D-glucopyranosyl
vincosamide
Antioxidant activity [82]
P.
microlabastra
Leaves, stem,
and roots bark
Methanol extract, petrol
and ethyl acetate
fractions
Antimicrobial activity [77]
P. gardineri Branches and
leaves
Dichloromethane and
methanol extracts
Antimicrobial activity [78]
P. nigra Branches and
leaves
Dichloromethane,
hexane and methanol
extracts
Antimicrobial activity [78]
P. reevesii Aerial parts Methanol extract Antimicrobial activity [83]
P. spectabilis Leaves Coumarin,
deoxysolidagenone,
psoralene, and
solidagenone
Antifungal activity [13]
P. prunifolia Branches Ethanol extract,
strictosamide, and 14-
oxoprunifoleine
Antiprotozoal activity [49]
P. serpens Not specified Ethanol extract Inhibition of herpes
simplex virus (HSV-
1) replication
[84]
P. klugii Stem bark Klugine, 7’-O-
demethylisocephaeline,
cephaeline,
isocephaeline, and 7-O-
methylipecoside
Antiparasitic activity [54]
P. laciniata Leaves Alkaloid fraction,
lyaloside, and
strictosamide
Monoamine oxidase
inhibition
[44]
P. suterella Leaves Alkaloid extract and
(E/Z)-vallesiachotamine
Monoamine oxidase
inhibition
[44]
P. myriantha Leaves Strictosidinic acid Monoamine oxidase
inhibition
[45]
33
P. myriantha Aerial parts Alkaloid extract,
myrianthosine, and
strictosidinic acid
Antichemotactic
activity
[46]
P. leiocarpa Leaves Aqueous extract Allelopathic activity [85]
P. capitata Leaves Ethanol extract
P. leiocarpa Leaves Ethanol extract Antimycobacterial
activity
[86]
P. glaziovii Leaves Ethanol extract Antimycobacterial
activitiy
[86]
P. nuda Leaves Ethanol extract Antimycobacterial activitiy
[86]
P. pubigera Leaves Ethanol extract Antimycobacterial activitiy
[86]
P. racemosa Leaves Ethanol extract Antimycobacterial activitiy
[86]
P. ruelliifolia Leaves Ethanol extract Antimycobacterial activitiy
[86]
P. suterella Leaves Ethanol extract Antimycobacterial activitiy
[86]
P. vellosiana Leaves Ethanol extract Antimycobacterial activitiy
[86]
Synthesis Of Some Compounds From Psychotria Species
Quadrigemine C (2) and Psycholeine (3)
The total synthesis of quadrigemine C (2) and psycholeine (3), isolated from P.
oleoides [15], was performed by Lebsack et al. (2002) [87]. The route reported starts
with meso-chimonanthine (7) which was obtained by reaction of oxindole with isatin
in 13 steps, as had been reported in previous work [88]. After some steps,
quadrigemine C (2) was obtained and acid-catalysed isomeration led to the formation
of psycholeine (3), as described in Fig. (7).
34
N N
N N
H
H
MeH
MeH
NO
H
NO
O
H
+
13 Steps
N N
N N
H
HI
I MeH
MeH
Bn
MeH
N N
N N
H
H
ON
TfO
TsMeN
O
NMeTs
N
TfO
MeH
Bn
Bn
Bn
MeH
N N
N N
H
H
N O
TsMeN
NO
TsMeN
MeH
Bn
Bn
MeH
N N
N N
H
H
NO
TsMeN
N O
TsMeN
MeH
H
Me
H
Me
MeR
N N
N N
H
HN
N
N
N
MeR
N
N
N
Me
N
Me
N N
H MeH
NN
HMe H
H
H
1. BuLi, TMEDA,Et2O 78 °C
2. ICH2CH2I, Et2O-78 °C to 0 °C
XPd2(dba)3.CH3, P(2-furyl)3, CuI, NMP, rt
X =
N
O
SnBu3
NMeTs
Bn
Pd(OAc)2, R-ToI-BINAP,
PMP, MeCN, 80 °C
Pd(OH)2, EtOH, MeOHH2 (100 psi), 80 °C
Na, NH3, THF, -78 °C
NH4Cl
0.1 N AcOH
100 °C
oxindole
isatin
meso-chimonanthine
(2) (3)
Fig.(7) . Synthesis of quadrigemine C (2) and psycholeine (3) proposed by Lebsack et
al. (2002)
Psychotrimine (9)
The first total synthesis of psychotrimine (9) was proposed by Matsuda, Kitagima and
Takayama (2007), involving 16 steps [89]. On the other hand, Newhouse and Baran
(2008) carried out the total synthesis of (±)-psycotrimine in four steps, using 7-
bromotryptamine as starting material. According to them, there had been no
methodology, up to that point, to construct that kind of C-N bond [90], as can be seen
in Fig. (8).
35
1. NIS, Et3N
NH2
I
H
N
NH
Br
HN
I
H
CO2Me
N
NH
Br
NNH
H
CO2MeMeO2C
N
NH
NNH
N
NHCO2Me
H
CO2MeMeO2C
N
NH
NNH
N
NHMe
MeMe
Br
N
NHCO2Me
H
Br NNHCO2Me
HN
I
2. Pd(OAc) Na2CO3
LiCl, X
R = CH2CH2NHCO2Me
N
R
H
NHCO2Me
TMS
= X
4. Red-Al
NHMe
NHMe
3. CuI
K2CO3
Fig .(8). Synthesis of psychotrimine (9) proposed by Newhouse and Baran (2008)
1-Hydroxybenzoisochromanquinone
This benzoquinone was isolated from the wood of P. camponutans. Its synthesis,
performed by Jacobs, Claessens and Kimpe (2008), was achieved with a phthalide
annulation reaction using 3-cyano-1(3H)-isobenzofuranone (142) and 5,6-
dihydropyran-2-one (143), followed by reduction of the lactone moiety. This synthetic
route is described in Fig. (9) [91].
O
O
CN
+O
O OH
OH
O
O OMe
OMe
O
O
OMe
OMe
O
OH O
O
O
OH
(136)
1.1 equiv. t-BuOLi
THF-60 °C, 3h; rt, 14h
5 equiv. K2CO3
2.2 equiv. Me2SO4
Acetone,, 2h
OMe
OMe
O
O
1.2 equiv. DIBAL-H
Toluene,-60 °C, 2.5 h
3 equiv. CAN
CH3CN/H2O 1/2rt, 30 min
(142) (143)
Fig.(9) . Synthesis of 1-hydroxybenzoisochromanquinone proposed by Jacobs,
Claessens and Kimpe (2008)
36
Concluding Remarks
The genus Psychotria presents a wide chemical diversity, comprising mainly
alkaloids. The more abundant alkaloids of the subgenus Psychotria are polyindole
alkaloids whereas MIAs are predominant in the subgenus Heteropsychotria.
Terpenoids, flavonoids, and other compounds are well known for their biological
properties and although a suite of compounds belonging to these phytochemical
classes has been isolated from the Psychotria genus, few have been subjected to
pharmacological assays. From two thousand species, only forty-seven have been
examined so far. There is a perception that extensive research work has been done
with some species of this genus; however, a large number of species are still
chemically and/or pharmacologically unknown. While this review has attempted to
unite the relevant information about Psychotria species, the bioactivity profiles from
the genus, and its alkaloids as the main bioactive compounds, clearly suggest future
research priorities. The presence of alkaloids makes the species of Psychotria
extremely promising, considering that this class of metabolites has shown a range of
biological activities. Moreover, these compounds can be used as models to obtain
more potent and effective synthetic derivatives.
Abbreviations
Ac acetyl
CAN Cerium (IV) ammonium nitrate
Et Ethyl
MIAs Monoterpene indole alkaloids
NIS N-iodosuccinimide
Pd2(dba)3 tris(dibenzylidineacetone)dipalladium (0)
PMP 1,2,2,6,6-pentamethylpiperidine
Rt room temperature
37
THF Tetrahydrofuran
TMEDA Tetramethylethylenedramine
TMSOTf trimethylsilyl trifluoromethanesulfonate
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40
3.2 Trabalho 2:
13C-NMR Spectral Data of Alkaloids Isolated from
Psychotria Species (Rubiaceae)**
Almir Ribeiro de Carvalho Junior 1, Ivo Jose Curcino Vieira 1* , Mario Geraldo de Carvalho 2,
Raimundo Braz-Filho 1,2, Mary Anne S. Lima 3, Rafaela Oliveira Ferreira 4, Edmilson José Maria 1,
Daniela Barros de Oliveira 5 1Laboratório de Ciências Químicas, CCT, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Campos dos
Goytacazes, RJ - 28013-602, Brazil;
2Departamento de Química, ICE, Universidade Federal Rural do Rio de Janeiro, Seropédica-RJ - 23890-000,
Brazil;
3Departamento de Química Orgânica e Inorgânica, Centro de Ciências, Universidade Federal do Ceará,
,Fortaleza–CE - 60021-940, Brazil;
4Colegiado de Ciências Agrárias e Biotecnológicas, Universidade Federal do Tocantins, Gurupi-TO – 77402-970,
Brazil; 5Laboratório de Tecnologia de Alimentos, CCTA, Universidade Estadual do Norte Fluminense Darcy Ribeiro,
Campos dos Goytacazes, RJ - 28013-602, Brazil.
*Correspondence: [email protected]; Tel.: +55-22-27486504
Academic Editor: Prof. Dr. Thomas J. Schmidt Received: date; Accepted: date; Published: date
Abstract: The genus Psychotria (Rubiaceae) comprises more than 2,000 species, mainly found in
tropical and subtropical forests. Several studies have been conducted concerning their chemical
compositions, showing that this genus is a potential source of alkaloids. At least 70 indole alkaloids
have been identified from this genus so far. This review aimed to compile 13C-NMR data of alkaloids
isolated from the genus Psychotria as well as describe the main spectral features of different skeletons.
Keywords: Rubiaceae; Psychotria; 13C-NMR Spectral Data.
** Trabalho publicado no periodico Molecules, volume 22, páginas 1-22, no ano 2017,
doi: 10.3390/molecules22010103.
1. Introduction
In phytochemistry and related areas, structural elucidation techniques play a key
role because precise knowledge of the chemistry of plants requires unequivocal
structural characterization of its metabolites to obtain information related to the
taxonomy of plant groups. Moreover, correct identification of biologically active
41
compounds is important, both to understand their possible mechanisms of action and
propose chemical modifications aimed at enhancing their activity.
The characterization of natural products requires, apart from patience and
dedication, knowledge about spectroscopic techniques (interpretation of these data)
and the biosynthesis of different types of metabolites. Comparison with literature data
is another important auxiliary tool that aids the structural characterization of a given
compound. In this context, finding a material that provides as much information as
possible about the spectral data of metabolites isolated from a genus (such as
Psychotria) may enable saving time.
The genus Psychotria (Rubiaceae) comprises more than 2,000 species, which
occur mostly in tropical and subtropical regions[1], with many of these species being
employed in folk medicine to treat several diseases [2,3]. The biological potential of the
chemical constituents of the species of this genus has possibly motivated several
studies regarding the chemical composition of such species. Most of these have
focused on investigating alkaloid fractions obtained by acid–base extraction, probably
owing to the biological importance of this type of metabolite. Such efforts have led to
the isolation and/or identification of various alkaloids, primarily indole-type. Some of
them exhibit some biological properties such as analgesic [4,5], antioxidant [6],
antiparasitic [7], and cytotoxic[8,9] activities.
This review aimed to compile 13C NMR spectral data of alkaloids isolated from
Psychotria species as well as to discuss the main spectral features observed for the
different types of skeletons.
2. Discussion
2.1. 13C Chemical Shifts of Monoterpene Indole Alkaloids Isolated from Psychotria
Species.
Monoterpene indole alkaloids (MIAs) comprise a wide group of secondary
metabolites, found mainly in the Apocynaceae, Loganiaceae, and Rubiaceae families
[10]. Their biosynthesis involves a reaction between tryptamine (derived from
42
tryptophan) and the iridoid secologanin, catalyzed by strictosidine synthase [11]. This
initial step leads to the formation of strictosidine (1) (Table 1 , Figure 1 ), the key
precursor of other MIAs.
Strictosidine (1) was isolated from P. elata [12] and P. nuda (data not reported),
and presents an ortho-substituted ring system (as do most of the MIAs isolated from
this genus), characterized by the presence of four methine carbon signals at δC 118.8
(CH-9), 120.1 (CH-10), 122.7 (CH-11), and 112.0 (CH-12), and two quaternary carbon
signals at δC 127.9 (C-8) and 137.9 (C-13). The signals of two quaternary carbons at δC
133.2 (C-2) and 107.7 (C-7), along with a methine carbon at δC 52.4 (CH-3) and two
methylene carbons at δC 42.9 (CH2-5) and 21.0 (CH2-6) complete the
tetrahydro-β-carboline system. The secologanin moiety is confirmed by the presence
of signals resonating at δC170.6 (C-22), δC 109.9 (C-16), and 156.1 (C-17), relative to
an α,β-unsaturated carboxyl group, a terminal vinyl at δC135.7 (CH-19) and 119.5
(CH2-18), besides signals at δC97.5 (CH-21), 45.6 (CH-20), 35.9 (CH2-14), and 32.5
(CH-15). The carbon signals of the glucose unit are observed at δC100.3 (CH-1’), with
four mono-oxygenated methines in the interval from δC78.6 to 71.7 and one
oxygenated methylene at δC 62.9 [13].
Strictosidine (1) may function as a precursor of other biosynthetic pathways,
leading to different skeletons and consequently changes in spectral properties.
Carbonylation at C-5 (δC = 176.5 ppm), as observed for 5-carboxystrictosidine (4), for
example, promotes a chemical shift displacement of CH2-6 (Δδ = 4.2 ppm, β effect)
when compared with 1, as can be seen in Table 2. A similar pattern was observed for
methylation of N-4 on correantoside (7) isolated from P. correa [14], where
Δδvariations of 5.4 and 3.5 ppm are observed for CH-3 and CH2-5, respectively (β
effect). For 10-hydroxycorreantoside (8), it is possible to observe the electronic
influence of a hydroxyl by the inductive effect at the ipso carbon (C-10) and an increase
in the electron densities at the ortho (CH-9 and CH-11) and para (C-13) positions by
the mesomeric effect. On the basis of this mesomeric effect, the signals corresponding
to carbon atoms at the ortho, CH-9 [δC118.8 (1) and 104.4 (8), ΔδC= −14.8 ppm] and
43
CH-11 [δC 122.7 (1) and 114.2 (8), ΔδC= −8.5 ppm], and para positions, C-13 [δC 137.9
(1) and 131.4 (8), ΔδC= −6.5 ppm], are displaced upfield.
Other metabolic pathways of this class of alkaloid revealed cyclization reactions
involving N-1(5 and 6) or N-4 (7–14) with C-22, or N-1 with C-18 and N-4 with C-22, as
particularly observed for stachyoside (30) isolated from P. stachyoides [15] (Figure 2 ).
Strictosamide (5), isolated from four different species,[16-19] is an example of lactam
formation between N-4 and C-22. By examining Table 2 ,it is possible to notice, apart
from the absence of a methoxyl group (carbomethoxy function) signal at δC 52.4, a
slight difference in the chemical shift of C-22 (δC 167.1 ppm), when compared with
compound 1 (δC 170.6 ppm), as well as a ΔδC variation of 6.9 ppm for C-17. In contrast,
correantoside (7) exemplified the first possibility involving cyclization between N-1 and
C-22. It is possible, in this case, to observe the variation in the chemical shifts of the
ortho CH-12 (Δδ = 4.0 ppm) and para CH-10 (Δδ = 4.1 ppm) atoms, promoted by the
inductive and mesomeric effects of the carboxyl group at C-22. These effects were
also observed for compounds 8 [ΔδC = 4.8 (CH-12) ppm], 13 [ΔδC = 4.4 (CH-12) and
4.1 (CH-10) ppm], 14 [ΔδC = 4.4 (CH-12) and 4.5 (CH-10) ppm], 18 [ΔδC = 7.8 (CH-12)
and 6.2 (CH-10) ppm], and 19 [ΔδC = 7.3 (CH-12) and 5.4 (CH-10) ppm], showing that
the downfield displacements of the CH-12 and CH-10 signals may be used to suggest
that N-1 is attached to C-22.
There are some examples of alkaloids isolated from this genus, whose
biosynthesis involves hydrolysis of a glycoside moiety such as
(E/Z)-vallesiachotamines, 23 and 24, isolated from P. bahiensis [17],
and10-hydroxy-iso-deppeaninol (27) and N-oxide-10-hydroxyantirhine (29) isolated
from P. prunifolia [20]. These types of skeletons may be suggested by analysis of the
region of the 13C spectrum that is typical of sugar, revealing the absence of the typical
signal of the anomeric carbon around δC 100.0, apart from additional signals of the
oxy-carbons characteristic of this unit.
Kerber et al. (2001) reported the isolation of a new MIA from P. brachyceras leaves
[21], named brachycerine (33), which showed a new alkaloid skeleton. Its biosynthesis
44
involved the coupling of tryptamine to a 1-epi-loganin derivative. Psychollatine (34), a
new MIA from P. umbellate [22], presented a terpenoid derivative from geniposide.
Both alkaloids as well as compounds 21, 22, and 35 revealed an important
characteristic in their 13C spectra: the absence of typical signals of a terminal vinyl
group (~δC 119 ppm). In contrast, bahienosides A (38) and B (37), isolated from P.
bahiensis[17], showed duplicate signals relative to two secologanin moieties. Figure 2
shows typical carbon assignments, which may indicate some different structural
possibilities in comparison with those values observed for strictosidine (1).
Table 1 . Monoterpene indole alkaloids from Psychotria species.
Compounds Species Reference
s
13C NMR Data
Strictosidine (1) P.elata [12] [13]
Strictosidinic acid (2) P. acuminata
P. barbiflora
P. myriantha
[1, 23-25] [25]
Palicoside (3) P. racemosa [12] [26]
5α-carboxystrictosidine (4) P. acuminata
P. bahiensis
[17, 23] [27]
Strictosamide (5) P. bahiensis
P. nuda
P. prunifolia
P. suterella
[16-19] [18]
N,β-D-glucopyranosilvincosamide (6) P. leiocarpa [28] [28]
Correantoside (7) P. correae [14] [14]
10-hydroxycorreantoside (8) P. correae [14] [14]
Correantine B (9) P. correae [14] [14]
20-epi-correantine B (10) P. correae [14] [14]
Correantine A (11) P. correae [14] [14]
Correantine C (12) P. correae [14] [14]
N-desmethyl-correantoside (13) P. stachyoides [29] [29]
Nor-methyl-23-oxo-correantoside (14) P stachyoides [15] [15]
14-oxoprunifoleine (15) P. prunifolia [18, 20] [18]
17-vinyl-19-oxa-2-azonia-12-azapentacy-clo
[14.3.1.02,14.05,13.06,11]icosa-2(14),3,5(13),6(
11),7,9-hex-aene (16)
P. prunifolia [18] [18]
Naucletine (17) P. suterella [19] [30]
Correantosine E (18) P stachyoides [31] [31]
Correantosine F (19) P stachyoides [31] [31]
45
Lagamboside (20) P. acuminata [23] [23]
N4-[1-((R)-2-hydroxypropyl)]-psychollatine
(21)
P. umbellata [32] [32]
N4-[1-((S)-2-hydroxypropyl)]-psychollatine
(22)
P. umbellata [32] [32]
(E/Z)-vallesiachotamine (23 + 24) P. bahiensis
P laciniata
[17, 33] [34]
Isodolichantoside (25) P. correae [14] [14]
Angustine (26) P. bahiensis
P laciniata
[17, 33] [35]
10-hydroxy-iso-deppeaninol (27) P. prunifolia [20] [20]
10-hydroxy-antirhine (28) P. prunifolia [20] [20]
N-oxide-10-hydroxyantirhine (29) P. prunifolia [20] [20]
Stachyoside (30) P stachyoides [15] [15]
Lyaloside (31) P. laciniata
P. suterella
[19, 36] [37]
Myrianthosine (32) P. myriantha [25] [25]
Brachycerine (33) P. brachyceras [21] [21]
Psychollatine (34) P. umbellata
P umbellata
[5, 22, 38] [22]
3,4-Dehydro-18,19-β-epoxy-psychollatine
(35)
P. umbellata [32] [32]
Desoxycordifoline (36) P. acuminata [23] [39]
Bahienoside B (37) P. acuminata
P. bahiensis
[17, 23] [17]
Bahienoside A (38) P. bahiensis [17] [17]
46
R1 R2 R3(1) H H Me(2) H H H(3) H Me H(4) CO2H H Me
C
OH
OGlc
N
NR2
H
H
R1
O2R3
2216
1520
19
17
1432
7
6 5
89
10
11
1213
18
21
22
16
15
20
19
17
14
3
2
7
6 5
89
10
11
1213
18
21
1''
1'
22
16
15
20
19
17
14
3
2
7
65
89
10
11
1213
18
21
Glc
N
N
R2
O
O
OR1
R1 R2(5) H 3a-H (6) Glc 3b-H
N
NMeH
O
O
OGlcH
H
R
H
R (7) H (8) OH
2216
1520
1917
1432
7
6 5
89
10
11
1213
18
212216
1520
1917
1432
7
6 5
89
10
11
1213
18
N
NMeH
O
OR2
MeH
R1H
R1 R2 (9) CHO H (10) H CHO
2120
1516
1719
1432
7
6 5
89
10
11
1213
18
22N
NMeH
O
CO2MeH
H
HHO
(11)
N
NMeH
CHOH
HO
HO
H
(12)
ON
N
R
H
N
N
N
H
O
O
(17)
N
NR
O
O
H
H
OGlc
H R(13) H(14) CHO
+16
15
20
19
17
1432
7
6 5
89
10
11
1213
18
21
2216
1520
19
17
1432
7
6 5
89
10
11
1213 18
21
22 16
15
2019
17
14
32
7
6 5
89
10
11
1213
18
21
R (15) C=O (16) CH2
(18)
3,4 (19)
Glc
N
N
O
O
H
O
H N
N
R
O
OGlc
CO2Me
H
H
H
H
22
16
1520
19
17
14
3
2
7
6 5
89
10
11
1213
18
21
21
2015
16
17
19
1432
7
6 5
8
9
10
11
1213 18
22 2120
15
16
17
19
14
32
7
6 5
89
10
11
1213
18
22
24
25 26
R(21) 25a-OH(22) 25b-OH (20)
GlcN
N
H
CO2Me
CH2OHH
Figure 1. Structures of monoterpene indole alkaloids from Psychotria species
47
(23 + 24)
N
N
H
CO2Me
CHOHH
(28)
N
N
OHHH
H
HO
HO
(27)
2216
15
20
19
17
14
3
2
7
6 5
89
10
11
1213
1821
22
16
15
20
19
17
14
3
2
7
6 5
89
10
11
1213
18
1''
2216
15
20
19
17
1432
7
6 5
89
10
11
1213
18
21
1''
16
15
20 19
17
14
3
2
7
6 5
89
10
11
1213
1821
16
15
20
19
17
1432
7
6 5
89
10
11
1213
18
21
N
N
H
HO
H
H
OHH
N
NMeH
OMeO2C OGlc
HH
(25)
N
N
N
H
O
(26)
(29)
16
15
20 19
17
14
3
2
7
6 5
8
9
10
11
1213
1821
-
+N
N
H
HO
H
O
H
OH
H
N
N
OH
H
H CO2Me
O
HO
H
H
GlcH
(33) (34)
N
N
O
GlcOH
H
CO2CH3
H
H
H
N
N
O
O
H
H
CO2Me
GlcOH
(35)
N
N C
CO2H
O
OGlc
H
H
O2Me
(36)
22
16
15
20
19
17
14
32
7
6 5
89
10
11
1213
18
21
22
16
1520
19
17
14
32
7
6 5
89
10
11
1213
18
21
22
16
1520
19
17
14
32
7
6 5
89
10
11
1213 18
21
1616
21
20
1516
17
19
14
32
7
6 5
89
10
11
1213
18
22
(31)
N
N
O
H
H OGlu
H3CO2CH
22
16
15
20
19
17
14
32
7
6 5
89
10
11
1213
18
21
22
16
15
20
19
17
14
3
2
7
6 5
89
10
11
1213
18
21
b
N
N
O
O
H
H
OGlc
H(30)
20
1516
17
19
1432
7
6 5
89
10
11
1213 18
21
22
16
15
20
19
17
14
3
2
7
6 5
89
10
11
1213
18
21
3b
1415
16 17
b
19
18
2021
bb
b
b
b b
1' '
Glc
Glc
O
O
H
H
MeO2C
N
N
H
R
O2Me
O
C
H
H O
R (37) 3-OH(38) 3-OH
N
NH
HO
O
OOGlu
H
(32)
Figure 1 . Continued
48
C
OH
OGlc
N
NH
H
H
O2Me22
16
15
20
19
17
14
3
2
7
6 5
89
10
11
1213
18
21
(1)
4
1
CH
14N
NH
22
32
789
10
11
1213
O(30)
CH CH2
GlcH
N
NHH
H
O
HO
H
H
(33)
21
2015
19
14
32
7
6 5
18
C
CH
(27)
16
15
20
19
17
1432
7
6 5 18
21
N
N
OHHH
HHO
C
N4-O+
(28)(29)
16
15
20 19
17
14
3
2
7
6 5
1821
N
N
HH
H
OHH
4
(5)(6)
56
7
2
3
14
17
15
16
22O
N
N
H
H
(34)
H
H
15
20
19
14
32
7
6 5
N
N
GlcOH
H
H
21
GlcOHN
NO
H
H
(35)
1520
19
14
32
7
6 5
18
21
18
OCH3
Figure 2 . Structural signaling based on specific signals compared with values
for strictosidine (1): absence of a given signal in red and presence in green.
Table 2 . 13C NMR data of MIAs from Psychotria species.
Carbons Compounds/ δC (ppm)
1I 2III 3III 4I 5I 6I 7I 8I 9II 10II
C
2 133.2 132.3 134.7 133.2 134.8 136.1 134.3 133.8 132.9 133.0
7 107.7 106.0 105.2 109.0 110.3 111.5 115.7 115.3 114.8 114.8
8 127.9 126.1 126.6 128.0 128.7 129.5 130.4 131.3 129.1 129.1
13 137.9 135.8 135.8 138.4 137.8 137.7 137.3 131.4 136.0 136.0
22 170.6 170.0 168.4 170.9 167.1 166.3 168.2 167.8 166.2 166.2
16 109.9 113.4 112.5 109.9 109.2 109.1 112.2 112.0 108.6 109.6
CH
3 52.4 49.6 56.1 53.2 55.1 54.5 57.8 58.2 56.4 56.7
5 - - - 60.1 - - - - - -
9 118.8 117.8 117.4 118.8 118.7 119.3 119.2 104.4 118.1 118.0
10 120.1 118.7 118.1 120.1 120.2 121.3 124.2 155.1 123.2 123.2
49
11 122.7 121.2 120.3 122.6 122.6 122.9 125.5 114.2 124.6 124.6
12 112.0 111.5 110.8 112.1 112.3 114.8 116.0 116.8 115.4 115.2
15 32.5 31.8 30.6 32.4 24.9 27.9 35.7 35.6 29.7 29.2
17 156.1 150.0 151.8 156.1 149.2 149.2 155.7 155.5 158.0 156.4
19 135.7 135.6 135.6 135.2 134.4 133.4 135.1 135.0 70.2 69.4
20 45.6 44.3 44.0 45.7 44.7 44.1 45.4 45.4 51.8 53.9
21 97.5 95.1 95.9 97.6 98.1 97.5 97.4 97.3 - -
CH2
5 42.9 40.0 45.2 60.1 44.8 41.6 46.4 46.7 45.5 45.5
6 21.0 19.2 15.9 25.2 22.1 22.3 18.8 18.8 17.6 17.7
14 35.9 33.7 35.3 35.6 27.3 35.6 34.4 34.1 39.1 35.3
18 119.5 117.8 117.8 119.6 120.6 120.7 119.2 119.3 - -
CH3
MeN- - - 39.8 - - - 41.4 41.2 - 41.5
Me - - - - - - - - 18.3 19.3
Gluc
ose
1’ 100.3 98.9 98.7 100.5 100.5 99.6 100.5 100.5 - -
2’ 78.6 69.8 73.0 74.7 74.3 74.9 74.7 74.7 - -
3’ 78.0 73.1 77.2 78.0 77.9 77.9 78.6 78.6 - -
4’ 74.6 77.2 70.0 71.9 71.3 71.6a 71.6 71.7 - -
5’ 71.7 76.5 76.6 78.6 78.2 78.3 78.0 78.0 - -
6’ 62.9 61.0 61.0 63.1 62.6 62.7 62.9 62.9 - -
1’’ - - - - - 87.6 - - - -
2’’ - - - - - 71.9 - - - -
3’’ - - - - - 75.1 - - - -
4’’ - - - - - 71.6a - - - -
5’’ - - - - - 81.2 - - - -
50
6’’ - - - - - 62.9 - - - -
CHO - - - - - - - - 199.5 199.2
CO2
Me
52.4 - - 52.6 - - - - - -
CO2H - - - 176.5 - - - - - -
I CD3OD, II CDCl3 e III DMSO-d6, letters (a–e) indicate signals that may be interchanged.
Table 2 . Continued.
Carbons Compounds/ δC (ppm)
11II 12I 13I 14I 15I 16I 17II 18I 19I 20I
C
2 136.2 134.6 136.0 132.4 134.4 132.2 127.4 145.7 134.4 136.0
3 - - - - 139.7 139.5 140.8 - 148.1 -
7 108.0 117.4 117.0 116.2 124.6 132.9 116.9 138.0 134.0 111.3
8 126.8 130.6 131.0 130.1 118.9 119.7 125.7 123.9 125.2 129.7
13 137.1 137.7 137.3 137.7 146.9 144.6 139.0 140.0 142.3 136.0
14 - - - - 191.6 - - - - -
15 - - - - - - 141.1 - - -
16 111.2 - 112.7 111.8 - - 117.1 113.5 114.5 95.3
19 - - - - - - 199.6 - - -
20 - - - - - - 138.8 - - -
21 - 194.3 - - - - - - - -
22 167.5 174.8 168.6 168.1 - - 161.6 167.9 168.8 171.8
CH
3 61.4 58.5 50.6 47.9 - - - 50.0 - 50.2
5 - - - - 134.1 132.5 - 137.0 142.6 -
6 - - - - 120.6 116.0 - 116.2 114.5 -
9 118.5 117.8 119.2 119.5 123.6 122.8 119.3 123.9 122.3 119.0
10 119.8 119.0 124.4 124.6 123.4 122.4 119.9 126.3 125.5 121.0
11 121.5 125.0 125.5 126.0 137.2 132.3 120.9 133.5 131.3 122.7
51
12 109.2 126.0 116.4 116.4 113.7 113.2 112.0 119.8 119.3 114.6
14 - - - - - - 95.6 - - -
15 30.8 34.5 35.7 35.6 42.8 25.6 - 30.5 21.2 33.7
16 - 52.0 - - - - - - - -
17 155.2 67.5 155.6 156.5 87.9 86.7 154.0 157.2 155.9 149.3
19 74.8 149.7 135.2 134.9 132.8 134.9 - 133.6 134.1 140.8
20 52.0 - 45.6 45.3 42.0 41.2 - 46.4 46.7 55.2
21 75.5 - 97.5 97.6 - - 155.4 97.9 97.9 -
CH2
5 52.0 48.0 40.0 41.6 - - 40.7 - - 53.0
6 20.9 19.6 23.2 23.2 - - 19.8 - - 23.4
14 36.7 35.9 36.7 34.8 - 24.8 - 36.7 39.8 35.1
16
- - - - 42.8 25.6 - - - -
18 - 33.8 119.3 119.5 118.9 117.9 - 121.8 121.3 116.9
21 - - - - 63.4 61.9 - - - 65.4
CH3
18 18.6 - - - - - 29.3 - - -
MeN- 43.0 41.9 - - - - - - -
-
Glucose
1’ - - 100.7 100.8 - - - 100.1 100.1 87.6
2’ - - 74.9 74.9 - - - 74.8 74.7 72.4
3’ - - 78.7 78.2 - - - 78.0 77.9 79.4
4’ - - 71.8 71.7 - - - 71.7 71.7 71.8
5’ - - 78.2 78.7 - - - 78.0 78.6 81.2
6’ - - 63.1 63.0 - - - 62.9 62.9 63.0
CHO - - - 163.9 - - - - - -
CO2Me 51.1 - - - - - - - - 51.2
I CD3OD, II CDCl3 e III DMSO-d6, letters (a–e) indicate signals that may be interchanged.
52
Table 2 . Continued.
Carbons Compounds/ δC (ppm)
21I 22I 23III 24III 25I 26III 27I 28I 29I
C
2 134.0 133.4 133.1 133.6 134.0 126.8 136.9 130.5 131.0
3 - - - - - 136.9 145.5 - -
7 108.4 108.4 106.6 107.4 106.5 114.8 130.6 106.0 105.7
8 138.6 138.1 126.2 127.0 128.1 125.5 123.1 128.6 128.3
10 - - - - - - 152.6 151.8 152.0
13 128.4 128.0 136.1 136.8 137.8 138.5 137.5 133.1 133.6
15 - - - - - 139.0 - - -
19 141.0 142.1 - - - - - - -
16 112.2 112.0 93.2 93.4 112.0 119.8 - - -
20 - - 146.1 143.9 - 127.8 - - -
22 169.7 169.0 166.9 167.6 169.8 161.1 - - -
CH
3 61.7 59.3 48.6 47.9 58.8 - - 57.0 71.6
5 - - - - - - 135.7 - -
6 - - - - - - 114.6 - -
9 118.6 118.0 117.4 118.4 118.7 119.9 106.6 103.2 103.3
10 119.5 120.0 118.3 119.2 119.9 119.9 - - -
11 121.9 122.0 120.7 121.6 122.3 124.6 120.4 112.9 113.2
12 112.0 112.0 110.8 111.8 111.8 112.0 113.7 112.8 113.0
14 - - - - - 93.8 - - -
15 33.0 35.3 27.4 30.5 30.5 - 36.4 31.1 30.6
17 153.3 153.0 147.2 148.5 154.0 149.7 - - -
18 132.6 131.0 - - - - - - -
19 - - 152.0 146.3 135.8 130.2 138.1 138.7 138.2
20 49.0 48.4 - - 45.5 - 51.0 50.8 52.3
21 95.6 97.0 - - 97.8 147.7 - - -
25 65.1 66.3 - - - - - - -
CH2
5 49.0 49.6 49.8 50.7 47.9 40.4 - 52.4 69.0
6 21.4 19.7 21.3 22.2 17.9 19.2 - 18.1 20.6
53
14 39.4 39.5 32.9 32.9 34.5 - 37.0 31.6 28.5
17 - - - - - - 61.4 48.0 59.1
18 - - - - 119.8 119.8 118.7 118.5 118.5
21 - - - - - - 64.4 64.0 63.8
24 62.4 61.6 - - - - - - -
CH3
18 - - 14.3 13.8 - - - - -
26 20.7 21.2 - - - - - - -
MeN- - - - - 40.6 - - - -
Gluc
ose
1’ 100.1 100.1 - - 100.5 - - - -
2’ 74.6 74.8 - - 74.7 - - - -
3’ 78.0 78.0 - - 78.6 - - - -
4’ 78.2 71.6 - - 71.6 - - - -
5’ 76.2 78.5 - - 78.0 - - - -
6’ 62.7 62.5 - - 62.9 - - - -
CHO - - 195.5 191.5 - - - -
CO2
Me
51.6 51.7 49.7 50.8 51.9 - - - -
I CD3OD, II CDCl3 e III DMSO-d6, letters (a–e) indicate signals that may be interchanged.
Table 2 . Continued.
Carbons Compounds/ δC (ppm)
30I 31 32III 33I 34I 35I 36I 37I 38I
C
2 137.1 140.3 134.8 130.7 131.1 128.8 135.6 135.0 138.0
3 - 143.8 - - - 158.9 142.9 - -
5 - - - - - - 135.6 - -
7 118.4 121.0 121.0 108.3 107.9 118.8 128.4 107.3 106.6
8 129.2 126.9 121.5 127.7 127.6 139.9 121.7 128.4 128.0
13 139.1 134.6 140.2 112.3 138.1 126.1 141.6 137.8 138.0
15
19 - - - - 140.0 67.3 - - -
54
16 114.8 109.9 112.0 111.8 112.2 110.2 108.7 112.1 111.5
21 169.0 - - - - - - - -
22 - 166.6 170.0 169.1 169.1 168.9 171.3 169.7 170.0
22
b
- - - - - - 169.5 169.4
C
H
3 51.7 - 48.5 54.7 53.7 - - 58.8 59.6
5 - 137.3 137.0 - - - - - -
6 - 112.6 118.0 - - - 114.2 - -
9 119.7 121.4 126.6 118.9 119.0 121.2 121.4 120.6 118.7
10 125.2 119.0 118.9 120.2 120.3 121.1 119.9 119.7 120.0
11 126.8 127.6 127.5 123.2 123.6 126.2 128.4 122.0 122.5
12 118.3 111.8 112.5 112.3 112.3 113.6 111.6 112.0 112.0
15 32.9 30.1 - 35.5 37.5 31.7 34.5 31.5 31.6
17 148.7 151.6 151.0 153.5 153.4 153.1 153.2 154.0 154.7
18 - - - 74.3 138.5 62.5 - - -
19 53.3 134.0 134.5 49.0 - - 133.8 136.2 136.1
20 95.5 42.9 45.5 41.9 49.0 43.8 44.4 45.5 45.4
21 - 95.9 95.4 99.0 99.4 95.2 96.1 98.2 97.9
15
b
- - - - - - 30.3 30.5
17
b
- - - - - - 153.2 153.5
19
b
- - - - - - 135.7 135.5
20
b
- - - - - - 44.8 44.8
21
b
- - - - - - 98.5 98.3
C
H2
5 48.0 - - 41.8 42.1 48.2 - 44.8 44.8
6 21.2 - - 24.4 20.5 20.1 - 17.6 17.4
14 43.7 32.1 45.6 43.5 40.5 34.7 34.0 36.9 36.7
55
15 - - 30.0 - - - - - -
17 - - - - - - - - -
18 72.4 118.6 118.9 - - - 117.6 119.8 119.8
3b - - - - - - 52.0 51.9
14
b
- - - - - - - 28.0 27.4
18
b
- - - - - - - 120.1 120.1
C
H3
M
e
- - 10.4 - - - - - -
Gluc
ose
1’ 100.3 98.79 98.6 100.6 101.5 99.4 99.0 100.4 100.4
2’ 74.9 73.1 73.0 74.0 71.0 74.7 73.2 74.6 74.8c
3’ 78.5 77.3 69.9 71.1 78.6 78.1 76.6 78.0 78.6a
4’ 71.9 71.10 77.3 78.3 74.6 72.1 70.4 71.6 71.7d
5’ 78.5 77.8 76.8 77.7 77.6 78.8 76.6 78.4 78.2b
6’ 63.0 61.2 61.0 62.1 61.8 63.3 61.8 62.9 62.9e
1’’ - - - - - - - 100.3 100.4
2’’ - - - - - - - 74.8 74.6c
3’’ - - - - - - - 78.1 78.4a
4’’ - - - - - - - 71.6 71.6d
5’’ - - - - - - - 78.3 78.0b
6’’ - - - - - - - 62.8 62.8e
C
O2
M
e
- 50.7 - 51.8 51.9 51.9 50.6 52.1 52.1
I CD3OD, II CDCl3 e III DMSO-d6, letters (a–e) indicate signals that may be interchanged.
2.2. 13C Chemical Shifts of Pyrrolidinoindoline Alkaloids Isolated from Psychotria
Species.
56
Some studies have also reported that the isolation of pyrrolidinoindoline alkaloids
seems to be specific to the Psychotria species (Table 3 ). As shown in Figure 3 , its
chemical structures present the condensation of some N-methyl-tryptamine units with
different connection patterns, mainly involving C-3a-C3’a, C-3a-C-7, and N-C-3 bonds
or containing N-methyl-tryptamine units linked to a bis-quinoline part. The compound
(+)-chimonanthine (40) was isolated from several Psychotria species[40-42] and is an
example of a dimer that presents a C-3a-C-3’a-type linkage between its two units. Its
13C spectrum exhibited 11 carbon-signal equivalents for both units. The signals at δC
52.4 (CH2-2) and 84.6 (C-8a) are typical of carbons bearing one and two nitrogen
atoms, respectively. The signals at δC 33.2 and 63.6 were attributed to C-3 and C-3a,
respectively, whereas the signal at δC 33.8 is consistent with a methyl carbon attached
to a nitrogen atom. The ortho-substituted aromatic rings are characterized by signals at
δC 124.9 (CH-4/CH-4’), 128.3 (C-4a/C-4a’), 122.3 (CH-5), 119.8 (CH-5’), 129.9
(CH-6/CH-6’), 110.5 (CH-7/CH-7’), and 150.5 (C-7a/C-7a’) [40] (Table 4 ).
Since some compounds with more than two units present a chimonanthine portion
in their structures, the monitoring of C-3a and C-7 (main binding sites) and their
neighborhood may be a good alternative, in order to determine the positions of the
other monomeric units. Hodgkinsine (52) occurs frequently in the genus [41-46] and
presents a third unit with a C-3’’a-C-7’ linkage. In this case, besides replacement of a
methine aromatic carbon by a quaternary carbon (C-7’), observing the upfield
displacements of C-6’ and C-4’ (Δδ around 3.0 ppm) is possible probably because of
the presence of a group that increases the electron densities of these positions
(comparison with compound 40). Takayama et al. (2004), however, reported the
isolation of psychopentamine (60) from P. rostrata2, which showed a new type of
linkage between C-3’’’a and C-5’’ [2].
The chemical study of P. calocarpa leaves [43] led to the isolation of a new alkaloid
named psychotriasine (45), which presents a tryptamine unit linked to a pyrroloindole
unit by an N-C3’a linkage. This type of junction was also observed for psychohenin (46)
and compound 48 isolated from P. henryi [47, 48] and may be indicated by the
57
presence of a quaternary carbon (C-3’a) that resonates at δC 79.4, 77.8, and 76.7 ppm,
in the three compounds, respectively. In contrast, psychotrimine (53), isolated from P.
rostrata [2] shows, besides the N-C-3’a bond, an N-C-7’ linkage indicated by the signal
of a quaternary aromatic carbon C-7’ at δC 121.5 ppm.
Alkaloids with more complex structures, containing from four to seven units, such
as quadrigemines A–C (55–57), psychotridine (61), oleoidine (64), and caledonine
(65), have also been isolated from this genus; however, the structural elucidation of
these compounds becomes more difficult as the number of units increases. Probably
owing to this, some studies did not provide detailed attributions of their carbon signals.
In such cases, mass spectrometry plays an important role in establishing the number
of units present in their structures as well as the pattern of the junctions.
Table 3 . Pyrrolidinoindoline alkaloids from Psychotria species.
Compounds Species References 13C NMR Data
Meso-chimonanthine (39) P. forsteriana
P. muscosa
[41, 49, 50] [50]
(+)-Chimonanthine (40) P. colorata
P. muscosa
P. rostrata
P. hoffmannseggiana
[40-42] [40]
Iso-calycanthine (41) P. forsteriana [50] [50]
Calycanthine (42) P. forsteriana [50] [50]
(8-8a),(8’-8’a)-tetradehydroisocalycanth
ine 3a(R), 3’a(R) (43)
P. colorata [42] [42]
Nb-desmethyl-meso-chimonanthine (44) P. lyciiflora [49] [49]
Psychotriasine (45) P. calocarpa [43] [43]
Psychohenin (46) P. henryi [47] [47]
Compound (47) P. henryi [48] [48]
Compound (48) P. henryi [48] [48]
Glomerulatine A (49) P. glumerulata [51] [51]
Glomerulatine B (50) P. glumerulata [51] [51]
58
Glomerulatine C (51) P. glumerulata [51] [51]
Hodgkinsine (52) P. colorata
P. oleoides
P. lyciiflora
P. muscosa
P. beccarioides
P. rostrata
[41-46] [42]
Psychotrimine (53) P. rostrata [2] [2]
Psychotripine (54) P. pilífera [52] [52]
Quadrigemine A (55) P. forsteriana [53] [53]
Quadrigemine B (56) P. forsteriana
P. colorata
P. rostrata
[41, 53] [53]
Quadrigemine C (57) P. colorata
P. oleoides
[41-43, 45, 46,
50, 54]
[45]
Quadrigemine I (58) P. oleoides [49] [49]
Psycholeine (59) P. oleoides [46, 54] [46]
Psychopentamine (60) P. rostrata [2] [2]
Psychotridine (61) P. forsteriana
P. oleoides
P. colorata
P. beccarioides
[41, 44, 45, 53] [45]
Isopsychotridine C (62) P. forsteriana [53, 55] [55]
Isopsychotridine B (63) P. oleoides [49, 50] [45]
Oleoidine (64) P. oleoides [49] [49]
Caledonine (65) P. oleoides [49] [49]
59
N N
NN
MeH H
H MeH
1'
2'3
3'a
8'a
4'a
4' 5'
6'
7'7'a
1
2
33a
8a
4a
45
6
7 7a
1'
2'
3'3'a
8'a
4'a
4'5'
6'
7'7'a
(39)
1
2
33a
8a
4a
45
6
77a N
NN
N
H
H
Me
Me1'
2'
3' 3'a
8'a
4'a
4'
5'
6'
7'7'a
1
2
33a
8a
4a
4
5
6
77a
(41)
N
NN
N
Me
Me1'
2'
3' 3'a
8'a
4'a
4'
5'
6'
7'7'a
1
233a
8a
4a
4
5
6
77a
(43)
1' 4
5
6
77a
6'4'5'
4'a7'
7'a
2
3
3'a
89
2'
3'
1
1'
(45)
N
NHMe
NN
HMe
H
(46)
1'2'
3'
3'a
8'a
4'a4'
5'
6'
7' 7'a
1
2
3
2'''
84a
4 5
77a
69
(40)
NN
HMe H
N N
H MeH
N
NN
N
H
HMe
Me
(42)
1'
2'3'
3'a
8'a
4'a
4'
5'
6'
7'7'a
1
23
3a
8a
4a
4
5
6
77a
N N
NN
MeH H
H HH
'
2'
3'3'a
8'a
4'a
4'5'
6'
7'7'a
(44)
b
2
33a
8a
4a
45
6
77a
6
45
7 7a6'
4' 5'
4'a
7'7'a
2
3
3'a
8
9
2'
3'
1
1'
N
NHMe
N NHMe
H
4a
N
NN
N
M
Me
e
N
NN
N
H
Me
N
NN
N
M
Me
He
(49)
(51)
(50)
1'
2' 3'
3'a
8'a
4'a
4'
5'
6'
7'7'a
1
23
3a
8a
4a4
5
6
77a
1'
2' 3'
3'a
8'a
4'a
4'
5'
6'
7'7'a
1
23
3a
8a
4a4
5
6
77a
1'
2' 3'
3'a
8'a
4'a
4'
5'
6'
7'7'a
1
23
3a
8a
4a4
5
6
77a
N N
Me
NN
H
Me
(47)
1'2'
3'
3'a
8'a
4'a
4'5'
6'
7' 7'a
1
2
3
2'''
84a
4 5
77a
69
N N
Me
NN
H
Me
H(48)
Figure 3 . Structures of pyrrolidinoindoline alkaloids from Psychotria species.
60
(53)
1
7'a
7
4
23a4a
6
5
8a7a
3
6'
7'
7''a
8''a
3'a
2'3'
1'8'a
6''
3''
5''4''a3''a
7''
4''
2''
2'''
5'
4'
4'a
1'''
4'''
3'''
7'''
3'''a
4'''a 6'''
5'''
8'''a 7'''a
1''
N N
H Me
N
N
H
Me
N
N
H
Me
NN
HMe
(55)
6''
4''5''
7'' 7''a6'
4' 5'
4'a
7'
7'a
2''
3''
3'a
8''
9''
2'
3'
1''
1'
N
NHMe
N NHMe
N
N
H
Me
H
1
2
3
7 6
5
44a
8
9
7a
N N
NN
MeH H
H MeH
N
N Me
HH
1'
2'
3'3'a
8'a
4'a
4'5'
6'
7'7'a
(52)
1
2
33a
8a
4a
45
6
77a
1'''2''
3''3''a
8''a
4''a
4''
5''
6''
7''a
(54)
Me
N
NN
NN
N
MeH
1
7'a
7
5'
4'
4
6'
7'
2
7''a
4'a
8''a 1''
3'a
2' 3'
3a
1'
4a
8'a
6 5
8a
7a
3
6''3''
5'' 4''a3''a
7''
4''
2''
8''
3'''a8'''a
(56)
7''a
8''a
3'a
2'3'
1'8'a
6''
3''
5''
4''a
3''a
7''
4''
2''
2'''
5'
4'
4'a
1'''
4'''
3'''
7'''
4'''a
6'''
5'''
7'''a
1''
N
N
H
MeN
N
H
Me
N
N
H
Me
N N
H Me1
7'a
7
4
23a4a
6
5
8a7a
3
6'
7'
1''
1
7'a
7
4
2
7''a
8''a
3'a
2'3'
3a
1'
4a
8'a
6
5
8a7a
3
6''
3''
5''4''a3''a
7''
4''
2''
2'''
5' 4'
6'
7'
4'a
1'''
4'''
3'''
7'''
3'''a4'''a 6'''
5'''
8'''a 7'''a
N N
H MeH
N
N
H
Me
H
N
N
H
Me
H
NN
HMe H
(57)
N N
H MeH
N
N
N
Me
N
Me
NN
HMe H
3
7a 8a
4a
3a
1
2
4
7
1''
7'''a8'''a
5'''
6'''4'''a
3'''a7'''
3'''
4'''
1'''
4'a4'
5'
2'''
2''
4''
7''
3''a4''a
5''
3''
6''8'a1'
3'2'3'a
8''a
7''a
7'6'5
6
7'a
(59)
6
5
7
4'
7'
6'
3
7a8a
4a
3a2
7'a
1
4
4'a
1'
3'a
5'
8'a
N N
H Me
NN
Me
NN
HMe
NN H
Me
H
1'''
4''a4''
7''
3''a
5''
6''
7''a
2'
3'
3''2''
8''a
7'''a
8'''a
5'''
6'''
4'''a
3'''a
7'''
3'''
4'''
2'''
1''
(58)
Figure 3 . Continued
61
1''''
7'''a
8'''a
5'''
6'''
4'''a
3'''a
7'''
3'''
4'''
1'''
4'a4'
2'''4''
7''
3''a 4''a 5''
3''
6''
1'
2''
2''''
8''a
7''a
7'
6'
3
7a8a
4a
3a 2
7'a
7''''a
4''''a 5''''1
3'a
2'3'
8'a
6''''
3''''
5'
8
7''''
4''''
9
Me
N
NMe
HN N
H Me
N
N
H
N
N
H
Me
N
N
H
Me
(60)
5
6
4
7
6
5
7
4'
7'
6'
3
7a8a
4a
3a2
7'a
1
4
1'''
4'a
4''a
1'3'a
5'
4''
7''
3''a5''
6''
7''a
2'3'3''2''
8''a
1''''
7'''a8'''a
5'''
6'''4'''a
3'''a7'''
3'''
4'''
2'''
8'a
2''''
7''''a
4''''a
5''''
6''''
3''''
7''''
4''''
3''''a
8''''a
1''
N N
H Me
N
N
H
Me
N
N
H
Me
NN
HMeN
NH
Me
(61)
6
5
7
4'
7'
6'
3
7a8a
4a
3a2
7'a
1
4
4'a
1'
3'a
5'
8'a
N N
H Me
NN
Me
NN
HMe
NN H
MeN
NH
Me
(62)
H
1'''
4''a4''
7''
3''a
5''
6''
7''a
2'
3'
3''2''
8''a
1''''
7'''a
8'''a
5'''
6'''
4'''a
3'''a
7'''
3'''
4'''
2'''
2''''
7''''a4''''a
5''''6''''
3''''
7''''4''''
3''''a
8''''a
1''
n
1
7'a
7
5'
4'4 6'
7'
2
7''a
4'a
8''a1''
3'a3'
3a4a
8'a
6
5
8a
7a3
3''
4''a
3''a
2''
2'
1'
5''
4'' 6''
7''
N N
H Me
NNMe N
N H
Me
n = 3 (63)n = 4 (64)n = 5 (65)
Figure 3 . Continued
Table 4 .13C NMR data of pyrrolidinoindoline alkaloids from Psychotria species.
Carbons Compounds/ δC (ppm)
39II 40II 41ns 42II 43II 44II 45I 46I 47II
C
3 - - - - - - 112.7 110.0 112.3
3a 64.7 63.6 37.8 36.8 48.9 62.8 - - -
4a 133.7 128.3 127.0 125.9 125.6 132.2 130.4 130.5 130.0
7a 152.5 150.5 145.3 146.2 145.8 151.7 137.7 138.0 135.0
8a - - - - 165.0 - - - -
3’a 64.7 63.6 37.8 36.8 48.9 63.9 79.4 77.8 75.3
4’a 133.7 128.3 127.0 125.9 125.6 130.0 131.3 131.3 128.9
62
7’a 152.5 150.5 145.3 146.2 145.8 150.3 152.5 152.7 152.4
8’a - - - - 165.0 - - - -
CH
2 - - - - - - 125.0 126.1 123.4
4 125.2 124.9 118.3 117.1 123.0 123.9 124.7 119.5 119.0
5 119.2 122.3 122.2b 122.1 118.5 119.9 119.3 118.8e 119.1f
6 128.9 129.9 127.7 127.3 128.2 128.2 130.7 123.0 121.4
7 109.5 110.5 112.9 112.8 123.9 109.1 120.1 117.4 112.8
8a 83.9 84.6 71.7 71.82 - 79.3 - - -
4’ 125.2 124.9 118.3 117.1 123.0 124.4 112.2 124.9 126.4
5’ 119.2 119.8 125.2 125.2 121.9 117.9 122.4 119.8 118.7
6’ 128.9 129.9 127.7 127.3 128.2 128.4 119.6 130.9 130.2
7’ 109.5 110.5 112.9 112.8 123.9 108.2 110.0 110.4 108.8
8’a 83.9 84.6 71.7 71.82 - 82.4 87.0 87.3 86.6
CH2
2 53.1 52.4 46.9 47.3 48.5 44.9 - - -
3 36.4 33.2 34.9 32.5 29.9 35.3 - - -
2’ 53.1 52.4 46.9 47.4 48.5 51.8 52.0 52.3 53.6
3’ 36.4 33.2 34.9 32.5 29.9 38.1 39.9 40.0 37.7
2’’ - - - - - - - - 69.1
CH3
Me-
N1-
nd 33.8 46.9 43.4 31.1 - 36.3 33.9 40.6
MeN
1’-
nd 33.8 46.9 43.4 31.1 35.12 35.7 36.4 37.1
I CD3OD, II CDCl3 e III benzene-d6, letters indicate signals that may be interchanged.
Table 4 .Continued.
Carbons Compounds/ δC (ppm)
48II 49III 50III 51III 52II 53II 54I + II 55II* 56II*
C
2 129.6 - - - - - - - -
3 109.4 - - - - 114.9 - - -
3a - 49.1 48.6 49.2 62.8 - 69.1 60.9c 60.1c
4a 128.0 126.4 126.1a 129.5 131.7 128.3 133.8 132.3d 133.2e
7a 137.4 177.3 147.1 148.6 150.8 136.1 152.2 150.9h 150.6h
63
8a - 165.1 164.7 166.5 - - 106.9 - -
3’a 76.7 49.1 48.6 45.3 63.0 76.7 37.0 63.2j 63.9i
4’a 130.5 126.4 125.4a 122.3 132.3 132.0 122.0 132.4d 132.9e
7’ - - - - - 121.5 130.9 108.9g -
8’a - 165.1 164.7 - - - - - -
3’’ - - - - - 112.5 - - -
3’’a - - - - 60.0 - 38.4 62.9j 63.3i
8’’ - - - - - 25.7b 68.0 - -
9’’ - - - - - 52.0 - - -
4’’a - - - - 131.7 129.8 122.3 132.6d -
7’’a - - - - 151.1 136.1 144.4 - -
3’’’a - - - - - - - 60.8c 60.9c
CH
2 - - - - - 126.0 - - -
4 117.9 123.7 120.9 123.0 126.4 119.4a 122.9 - 125.9d
5 119.2 122.3 122.2b 122.1 118.5 119.9 119.3 118.8e 119.1f
6 121.3 128.9 129.0c 128.8 127.9 122.4 128.2 127.9f 128.0g
7 112.1 125.0 125.2d 125.2 109.0 111.2 107.7 109.0g 108.9
8a - - - - 86.4 - - 86.9i 85.9i
4’ 124.5 123.7 124.0d 117.5 121.9 123.7 123.7 122.5 125.1d
5’ 119.0 122.3 122.6b 124.9 116.8 119.3a 122.0 116.3k 118.3f
6’ 129.6 128.9 128.8c 127.1 126.0 127.3 121.1 125.4 127.8g
7’ 108.9 125.0 124.4d 114.4 - - - - -
8’a 86.5 - - 76.5 81.7 86.1 69.7 86.1i 83.3j
2’’ - - - - - 124.3 - - -
4’’ - - - - 124.2 119.3a 125.4 - -
5’’ - - - - 117.5 119.3a 117.8 118.7e 117.2f
6’’ - - - - 127.4 121.7 127.6 - -
7’’ - - - - 108.1 112.2 112.5 - -
8’’a - - - - 82.3 - 69.4 - 82.3j
8’’’a - - - - - - - - 87.1i
5’’’ - - - - - - - 116.2k 116.8f
6’’’ - - - - - - - 126.4f -
64
CH2
2 - 48.2 48.1 48.5 51.7 - 54.9 52.6a 52.3a
3 - 30.3 30.3 31.7 37.6 - 36.3 38.8b 38.5b
2’ 51.2 48.2 48.1 50.5 51.9 51.7 42.3 52.5a 52.2a
3’ 40.6 30.3 30.3 34.0 36.7 39.1 33.1 38.7b 36.6b
2’’ - - - - 51.9 - 45.9 52.2a -
3’’ - - - - 38.0 - 33.7 38.5b -
3’’’ - - - - - - - 36.6b -
CH3
Me-N
1-
44.8 30.9 30.8 30.7 35.2 36.3 36.4 35.7l 35.8k
MeN1’
-
36.1 30.9 - 36.6 35.0 36.4 - 35.5l 35.7k
Me-N
1’’-
- - - - 35.1 36.4 41.8 35.0l 35.6k
Me-N
1’’’ -
- - - - - - - - 35.2k
I CD3OD, II CDCl3 e III benzene-d6, letters indicate signals that may be interchanged, * indicates cases for
which there was no complete detailed attribution of carbon signals.
Table 4 .Continued.
Carbons Compounds/ δC (ppm)
57II* 58II* 59II* 60II 61II* 62II* 63II* 64II** 65II*
C
3a 60.6 60.0 59.6b 61.1 60.1a 60.9c 63.0a 60.4c 60.0c
4a - 132.0 132.4c 132.9b - 132.7d - 132.8 132.1e
7a - - - 152.8 - 150.6f - 150.7e 150.5f
3’a 62.6 63.0 37.5f 63.1 62.9 63.7h 63.3a 63.3c 63.0
4’a - - - 132.8b - 132.0d - - 132.4e
7’ - 110.0c - 123.8 - - - - 108.8
7’a - - - 151.0 - 148.9f - 150.3e 148.9f
5’’ - - - 136.2 - 117.1 - - -
3’’a 62.6 - 38.0f 64.2 62.9 63.2h 59.8c 60.9c -
65
4’’a - - - 132.6 - - - - -
7’’a - - - 149.8 - - - - 148.6
3’’’a 60.6 - 60.6b 62.3 60.6a 60.1c 59.8c - 60.5c
4’’’a - - 133.8c 138.6 - - - - -
7’’’ - - - 120.4 - - - - -
7’’’a - - - 144.7 - - - - -
3’’’’ - - - 114.5 - - - - -
3’’’’a - - - 128.3 60.8a - 60.7c - -
4’’’’a - - - - - - - -
CH
4 - 126.0 - 126.9 - 123.6 - 126.1d 125.3d
4’ - 124.0 - 122.1 - 122.2 - 124.1 125.2d
5 - 117.0b - 118.8 - 119.1e - 116.3 118.9
6 - 129.5 - 128.0 - 128.2 - 128.7 128.1
7 - 109.0c - 110.5 - 109.0 - 109.3 107.7
8a 85.8a 88.0d 88.5d 87.3 86.0b 87.2g 81.8b 86.6 86.9
5’ - 118.5b - 116.2 - 118.4e - 119.4 117.3
6’ - 127.5 - 126.5 - 126.1 - 126.2 125.3
8’a 82.3 83.0d 74.0g 82.4 82.6c 85.8g 86.8b 82.8 86.0
4’’ - - - 121.4 - - - 125.7d 124.1
5’’ - 119.0b - - - 117.1 - - -
6’’ - - - 126.1 - 125.4 - 128.4 -
7’’ - - - 108.5 - - - - -
8’’a 82.3 - 72.0g 83.4 82.3c 83.1 86.8d 83.0 -
4’’’ - - - 123.3 - - - 122.7 123.6
5’’’ - 119.5b - 118.8 - - - - -
6’’’ - 128.5 - 125.1 - - - - -
7’’’ - - - 120.4 - - - - -
8’’’a 86.7a - 87.5d 88.4 86.9b 82.1 85.5d - -
2’’’’ - - - 126.1 - - - - -
4’’’’ - - - 119.3 - - - 125.7 123.2
5’’’’ - - - 111.2 - - - - -
6’’’’ - - - 122.3 - - - - -
66
7’’’’ - - - 119.7 - - - - -
8’’’’a - - - - 85.1b - 84.8d - -
CH2 - - - - - - - - -
2 - 53.0 47.17a 52.7a - 52.5a - 52a 52.1a
3 - 38.0a - 37.8 - 38.8b - 38.7b 38.3d
8 - - - - - - - -
9 - - - - - - - -
2’ - - 52.7a - 52.0a - 52.8a 52.4a
3’ 39.0a 32.8e 35.8 - 38.5b - 38.7b 38.6b
2’’ - - 52.6a - - - 52.9a -
3’’ - 32.4e 37.2 - - - - -
2’’’ - 48.0a 52.5a - - - - -
3’’’ - - 39.2 - - - - -
3’’’’ - - 114.5 - - - - -
CH3 - - - - - - - -
Me-N
1-
36.0 36.1h 34.8 - 35.6i - 35.7 35.4g
Me-N
1’-
- 42.6i 35.3 - 35.1i - - 35.6g
Me-N
1’’-
- 42.6i 35.8 - - - - -
Me-N
1’’’ -
- 36.1h 35.7 - - - - -
Me-N
1’’’’ -
- - 36.5 - - - - -
I CD3OD, II CDCl3 e III benzene-d6, letters indicate signals that may be interchanged, * indicates cases for
which there was no complete detailed attribution of carbon signals.
2.3. 13C Chemical Shifts of Benzoquinolizidine Alkaloids Isolated from Psychotria
Species.
Muhammad et al. (2003) reported the isolation of five benzoquinolizidine alkaloids
from Psychotria klugii [7] (Table 5 ). Among them, klugine (66) and
7’-O-demethylisocephaeline (67) were reported for the first time, whereas cephaeline
67
(68), isocephaeline (69), and 7-O-methylipecoside (70) were previously isolated from
Cephaelis species [56, 57].
Compound 68 (ipecac alkaloid) as along with compounds 66, 67, and 69
possesses an unusual skeleton with two tetrahydroisoquinoline ring systems [10]
characterized by the presence of four quaternary carbon signs at δC 147.2, 147.5 (C-9
and C-10, oxygenated ortho-substituted carbons), 126.8 (C-7a), and 130.1 (C-11a),
two methine carbons at δC 108.6 (CH-11) and 111.5 (C-8), and signals at δC 62.4
(CH-11b), 52.3 (CH2-6), and 29.2 (CH2-7). A similar system is observed for the lower
unit, with the exception of the absence of a methoxyl group attaching C-6’ (a hydroxyl
group in this position). The remarkable difference between compounds 68 and 69
(isomers) is associated with the chemical shift of carbon C-1' at δC 51.9 and 55.3
respectively, whereas compounds 66 and 67 differ from 68 and 69 in the number and
positions of the methoxyl groups. Interestingly, compound 70 exhibits carbon
assignments relative to a tetrahydroisoquinoline ring attached to a secologanin moiety
at C-1.
The chemical structures of compounds 68–70 are shown in Figure 4 , and their 13C
NMR data are listed in Table 6 .
Table 5 .Benzoquinolizidine Alkaloids from P. kluggi. Compound Species Reference 13C NMR Data
klugine (66) P. klugii [7] [7]
7’-O-demethylisocephaeline
(67)
P. klugii [7] [7]
cephaeline (68) P. klugii [7] [56]
isocephaeline (69) P. klugii [7] [56]
7-O-methylipecoside (70) P. klugii [7] [57]
68
N
R1O
R2O
N
R3
H
H OR4
OH
12
312
13
4
6
77a
89
1011
11a11b
14
1'
3'
4'4'a
5'6'
7'8'
8'a
(70)
NAc
HO
MeO
H
O
OGlc
MeO2C
6'5'
9'1'
10'
8'
3
44a
56
78
8a1
4'11'
3'
1''
R1 R2 R3 R4(66) H Me OH Me(67) Me Me H H (68) Me Me 1'-H Me(69) Me Me 1'-H Me
Figure 4 . Structures of Benzoquinolizidine Alkaloids from P. klugii
Table 6 .13C NMR Data ofBenzoquinolizidine Alkaloids from P. kluggi.
Carbons Compound/ C (ppm)
66ns 67ns 68II 69II 70I
C
6 - - - - 146.5a
7 - - - - 147.8a
9 146.5a 146.8 147.2a 147.2a -
10 147.8b 148.0 147.5a 147.4a -
4a - - - - 126.9
7a 127.8 126.9 126.8 126.5 -
8a - - - - 130.2
11a 129.7 127.9 130.1 129.9 -
1’ 79.5 - - - -
4’ - - - - 111.7
4’a 127.7 123.2 127.6 127.9 -
6’ 146.4a 145.6 143.9b 144.0 -
7’
11’ - - - - 169.2
8’a 129.7 126.0 131.1 131.0 -
CH
69
1 - - - - 50.6
2
3 42.5 41.3 41.7 61.5 -
5 - - - - 116.2
8 116.2 112.1 111.5 111.4 111.1
11 109.7 109.0 108.6 108.2 -
11b 63.8 62.7 62.4 62.8 -
1’ - 53.6 51.9 55.3 98.7
3’ - - - - 153.1
4’ 28.5 27.6 29.0 29.3 -
5’ 116.4 115.2 114.7 114.8 27.5
8’ 110.0 113.2 108.4 108.6 136.3
9’ - - - - 45.1
CH2
1 40.6 36.9 36.9 39.3 -
3 - - - - 36.1
4 62.2 61.6 61.3 52.6 29.1
6 53.3 51.9 52.3 52.6 -
7 29.3 25.3 29.2 29.1 -
12 24.4 23.3 23.6 24.0 -
14 37.0 38.0 40.9 40.4 -
3’ 41.0 39.5 40.1 41.4 -
4’ 28.5 27.6 29.0 29.3 -
6’ - - - - 41.1
10’ - - - - 120.1
CH3 11.5 10.1 11.2 11.3 -
13 11.5 10.1 11.2 11.3 -
Me7-O
-
- - - 56.5
70
Me9-O
-
- 55.4d 55.8e 55.8f -
Me10-
O-
56.8c 55.8d 56.0e 56.0f -
Me7’-
O-
56.6c - 56.3e 56.0f -
Gluco
se
1’’ - - - - 100.5
2’’ - - - - 74.8
3’’ - - - - 78.2b
4’’ - - - - 71.5
5’’ - - - - 78.3b
6’’ - - - - 62.7
CO2M
e
51.7
I CD3OD, II CDCl3 e nsnot specified, letters indicate signals that may be interchanged.
3. Conclusions
In this work, we attempted to compile 13C data of alkaloids isolated from the
Psychotria genus and provide information that may be useful in order to distinguish
different types of skeletons. For monoterpene indole alkaloids (MIAs), mainly found in
tropical species, a good strategy for their structural elucidation is to compare their
spectral data with those observed for strictosidine (1). The monitoring of differences in
specific parts of the spectrum, such as the signals of C-22, CH-17, CH-12, CH2-5, and
CH-1’, may suggest alternative structural possibilities. Note that all comparisons
performed in this work are restricted preferably to compounds whose 13C NMR
experiments were run in the same solvent.
The main pyrrolidinoindoline alkaloids found in this genus are chimonanthine
derivatives, with units linked mostly by C3a-C3’a or C-3a-C7a bonds. Some examples
have shown different patterns of linkages between N (from tryptamine terminal units)
71
and C-3a. For compounds with more than three units, such as quadrigemines A–C and
psychotridine and its isomer, obtaining detailed assignments of these carbons is not
possible owing to structural complexity.
The occurrence of benzoquinolizidine alkaloids in Psychotria species is less
common, comprising some compounds isolated from Psychotria klugii.
Acknowledgments: The authors are grateful to Fundação de Amparo à Pesquisa do
Estado do Rio de Janeiro (FAPERJ) for grants and a research fellowship, to Conselho
Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Coordenação de
Aperfeiçoamento de Pessoal de Ensino Superior (CAPES) for research fellowships.
Author Contributions: All authors contributed equally to the realization of the
manuscript.
Conflicts of Interest: The authors declare no conflict of interest.
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77
3.3 Trabalho 3:
Metabolites from Psychotria suterella Müll. Arg. and Psychotria nuda
Cham. & Schltdl. Wawra (Rubiaceae) and Evaluation of Cytotoxic
Activity
Almir Ribeiro de Carvalho Juniora*, Michel de Souza Passosa, Mario
Geraldo de Carvalhob, Raimundo Braz-Filhoa,b, Milton Masahiko
Kanashiroc, and Ivo Jose Curcino Vieiraa
The chemical study of P. suterella leaves led to the identification of a new iridoid named 9-epi-
geniposidic acid (1), along with the known compounds geniposidic acid (2), sucrose (3), 3-O-
acethyloleanolic acid (4), pomolic acid (5), spinosic acid (6), maslinic acid (7),tormentic acid
(8), methyl oleanolate (9), lyalosidic acid (10), and strictosidinic acid (11). From twigs and
leaves of P. nuda sitosterol (12), stigmasterol (13), campesterol (14), phytol (15), -sitosterol-
3-O--D-glucoside (16), -stmasterol-3-O--D-glucoside (17), cinchonain Ia (18), cinchonain
Ib (19), N,N,N-trimethyltryptamonium (20), lyaloside (21), lawsofrutose (22), roseoside (23),
strictosamide (24), scopoletin (25), rotungenic acid (26), strictosidine (27), and 5-
carboxystrictosidine (28) were identified. These structures were elucidated based on NMR,
HR-MS, IR spectrum as well as comparison with literature data. Furthermore, the cytotoxic
activity of compounds 20, 23, and 24 was evaluated against two cancer cell lines (THP-1 and
U937). Only compound 24 showed significant cytotoxic activity against the U937 cell line (IC50
= 29.1 ± 1 g/L).
Keywords: Psychotria; iridoid; 9-epi-geniposidic acid; alkaloids; MTT
** Este trabalho será submetido ao periódico Natural Product Research.
1. Introduction
The genus Psychotria comprises about 2000 species, occurring mostly in tropical and
subtropical regions of the world (Marques de Oliveira et al. 2013). Some of its species
are widely used in folk medicine for several purposes such as earache (Verotta et al.
1998), abdominal pain (Amador et al. 1996), constipation (Zhou et al. 2010), coughs
(Benevides et al. 2004), etc. Several biological activities have been reported for this
genus, such as cytotoxic (Zhang et al. 2013), analgesic (Both et al. 2002), and
antimicrobial (Jayasinghe et al. 2002) activities, highlighting the biological potential of
78
its species.
Chemical studies related to these species have been shown that this genus is
a potential source of monoterpene indole alkaloids whose the biosynthesis involves
the coupling between tryptamine and the iridoid secologanin (Runguphan et al. 2009).
The chemical diversity of this genus also includes flavonoids (Lu et al. 2014),
triterpenes (Zhang et al. 2013), coumarins (Benevides et al. 2004), iridoids (Lu et al.
2014), etc.
In order to contribute to the expansion of the knowledge about the chemistry of
this genus, in this paper we describe the isolation and identification of metabolites from
P. suterella and P. nuda (Rubiaceae). In addition, it also shows the cytotoxic activity of
some of these compounds against THP-1 and U937 cancer cell lines.
2. Results and discussion
2.1 Metabolites from P. suterella
Compound 1 (Figure 1) was obtained in mixture with compounds 2 and 3 as a brown
oil. The molecular formula C16H22NaO10 was determined based on HR-ESI-MS (m/z
397.1064, [M + Na]+, calculated for 397.1111). Its NMR data (Table S1, Supplemental
online material) showed a doublet at H 5.15 (H-1, J = 3.4 Hz) attached to a carbon
at C 92.5 (CH-1), typical of methinedioxy group. The doublet at H 4.51 (d, 7.7, H-1’)
was attributed to the anomeric proton of the glucose moiety. The signal at H 7.50 (H-
3), attached to the carbon at C151.5 (CH-3) suggested a ,-unsatured carboxyl
group (C170.4, C-11). Moreover, it was also possible to verify signals of another
double bound (H-7, H 5.82) and a terminal hydroxyl group (CH2-10, C 60.1). Its
relative stereochemistry was proposed based on the coupling constant value of H-1
(3.4 Hz), typical of an axial-equatorial coupling and the shielding effect on C-1’ (C
96.6, suggesting H-9 in equatorial position). All correlations observed by 2D NMR
experiments are summarized in Table S1.
The known compounds were identified as geniposidic acid (2, Güvenalp et al.
2006), sucrose (3), 3-O-acethyloleanolic acid (4, Itokawa et al. 1989), pomolic acid (5,
Chama et al. 2015), spinosic acid (6, Wang et al. 2011), maslinic acid (7, Pnou et al.
2011), tormentic acid (8, Taniguchi et al. 2002), methyl oleanolate (9, Mahato &
79
Kundu), lyalosidic acid (10, Lin et al. 2011), and strictosidinic acid (11, Berger et al.
2015).
2.2. Metabolites from P. nuda
The metabolites isolated from twigs and leaves of P. nuda were identified as pomolic
acid (5, Chama et al. 2015), spinosic acid (6, Wang et al. 2011), sitosterol (12),
stigmasterol (13), campesterol (14), phytol (15, Miranda et al. 2012), -sitosterol-3-O-
-D-glucoside (16), -stmasterol-3-O--D-glucoside (17, Kojima et al. 1990),
cinchonain Ia (18), cinchonain Ib (19, Nonaka & Nishioka 1982), N,N,N-
trimethyltryptamonium (20, Martins et al. 2009), lyaloside (21, Berger et al. 2015),
lawsofrutose (22, Uddin et al. 2013), roseoside (23, Otsuka et al. 1995), strictosamide
(24, Zhang et al. 2001), scopoletin (25, Darmawan et al. 2012), , rotungenic acid (26)
(Nakatani et al. 1989), strictosidine (27, Patthy-luka et al. 1997), and 5-
carboxystrictosidine (28, Ferrari et al. 1986).
R1 R2 R3 R4 R5 R6 R7
4 H Ac CH3 H H H H
5 H H CH3 CH3 OH H H
6 H H CH3 H OH CH3 H
7 OH H CH3 H H CH3 H
8 OH H CH3 CH3 OH H H
9 H H CH3 H H CH3 CH3
26 H H CH2OH CH3 OH H H
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
R1
R3
R2O
R6
CO2R7
R4
R5
H
O
O
HOHO
HO
OH
O
O OH
OH
OH
H
1
3
52
461'
2'
3'
4'
5'
6'
3
O
OH
HO
OH
OHO
H H
1
2
34
5
6
1'
2'
3'
4'
22
OH
15
1
23 4
5
67
8
9
10 11
1216
17
18
13
141520
19
1
2
3
4 6
7
8
9
10
11
12
13
14
19
18
21
17
16
15
20
23
24
25
27
5
22
R1 R2
12 H CH2CH3 22,23-dihydro
13 H CH2CH3 22,23
14 H CH3; 22,23 dihydro
16 Glc CH2CH3 22,23-dihydro
17 Glc CH2CH3 22,23
R2
R1
O
CO2H
OO
HO
OHOH
OH
HO H
H
1
2'
3
46
7
8
9
10
5
1'
3'
4'5'
6'
2
18
12
3
456
7
89
10
1'
2'
3'
4'
5'
6'
6'''
5'''
4'''
3'''
2'''
1'''9'' 7''
8''
O
OH
OH
OH
OH
O
O
OH
OH
O
CO2H
OO
HO
OHOH
OH
HO H
H
1
2'
3
46
7
8
9
10
5
1'
3'
4'5'
6'
1
Figure 1. Chemical structures of compounds 1-28.
80
NN
O
H
O
O
H
H
H
O
HO
OHOH
OH
8
9
10
111
7
14
15
16
17
19
18
1213
21
20
4
5
32
6
2'1'
3'
4'5'
6'
24
10 R = H
21 R = CH3
1
NN
CO2R
O
OO
HO
OHOH
OH
H
22
1615
2019
1714
32
7
6
58
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
N
H
N+
12
34
56
7
89
1'
2'
20
R1 R2
11 H H
27 H CH3
28 CO2H CH3
1
NN
CO2R2
O
OO
HO
OHOH
OH
H
R1
22
1615
2019
1714
32
7
6
5
8
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
19
12
3
456
7
89
10
1'
2'
3'
4'
5'
6'
6'''
5'''
4'''
3'''
2'''
1'''9'' 7''
8''
O
OH
OH
OH
OH
O
O
OH
OH
10
O
O
HOO
HO
OHOH
OH
2
651
3
4
11
7
8 9
13
4'
3'
12
1'2'
6'
5'
23
19
8
6
7OO
OCH3
OH
54
2
310
25
Figure 1. Continued.
2.3. Assessment of cell viability by MTT assay
The cytotoxic potential of compounds 20, 23, and 24 was assessed by 3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results of cell
viability of THP-1 and U937 cell lines, treated with these compounds, after 48 h of
incubation, are shown in Figures S89 and S90 show the. In both cases, compounds
20 and 23 did not promoted significant cell death and the IC50 values were higher than
200 g/mL. Compound 24 did not display promising result against THP-1 cell line (IC50
= 120 ± 1 g/mL). However, this compound showed significant cytotoxicity against
U937 cell line (IC50 = 21.9 ± 1). This result is in accordance with previous reports in
literature, since indole alkaloids have been recognized by their cytotoxicity against
different cancer cell lines (Chaturvedula et al. 2003; Figueiredo et al. 2010; Shao et al.
2015; Zhang et al. 2015; Wang et al. 2015).
81
3. Experimental
3.1. Apparatus and instruments
Fourier transform infrared (FTIR) spectra were recorded on an IRAffinity-1 Shimadzu
spectrometer using KBr disk. The NMR analysis were carried out on a Bruker Ascend
500 in pyridine-d5, CDCl3 or methanol-d4 at 500 MHz for 1H and 125 MHz for 13C, using
TMS as internal reference. Chemical shifts (δ) are expressed in ppm and coupling
constants (J) in Hz. HR-ESI-MS mass spectra were obtained on a micrOTOF-Q II
Bruker Daltonics mass spectrometer, using positive and negative ion mode of analysis.
Gas chromatography coupled to Mass Spectrometry (GC/MS) of low resolution
experiments were carried out on a GCMS-QP5050A Shimadzu, operating with an
ionization energy of 70 eV. Silica gel 60 and silica gel 60 silanized (0.063-0.200 mm,
MERCK) were used for column chromatography (CC) and silica gel 60 F254 for thin
layer chromatography (TLC, MERK).
3.2. Plant material
Leaves of P. suterella and leaves and twigs of P. nuda were collected at the Reserva
Biológica de Poço das Antas, Nova Iguaçu-RJ, Brazil, and identified by the botanist
Sebastião José da Silva Neto. Both voucher specimens (H9724 and H9726,
respectively) are deposited at the herbarium of UENF.
3.3. Extraction and isolation
3.3.1.1. P. suterella
The powdered air-dried leaves of P. suterella (345.4 g) were exhaustively extracted
with methanol, at room temperature, affording 50.1 g of crude extract. After suspended
in a MeOH-H2O (1:3) solution, part of this extract (40.0 g) was partitioned with
dichloromethane, ethyl acetate and n-butane, affording, respectively, 19.7 g, 5.1 g, and
8.4 g of each fraction. The dichloromethane fraction was suspended in a
hexane:MeOH (1:1) solution, obtaining a hexane fraction (11.0 g) and a methanol
fraction (7.8 g).
The methanolic fraction (7.0 g) was subjected to CC on silica gel and eluted with
CH2Cl2 and CH2Cl2:MeOH solutions, increasing polarity till 15% of MeOH, affording 10
82
subfractions. Subfraction 4 (700 mg) was similarly chromatographed, affording
compound 9 (25.3 mg). Subfraction 5 (840.0 mg) was analogously chromatographed
affording compound 4 (27.0 mg) and a mixture of 5 and 6 (246.0 mg). From fraction 8
(660 mg), by analogous chromatographic procedure, a mixture of compounds 7 and 8
(88.0 mg) was obtained. Fraction 9 (457.3 mg) was also similarly chromatographed,
yielding a mixture of compounds 1, 2, and 3 (67.2 mg). The n-butanol fraction was
subjected to CC on silica silanized, eluted with isocratic mixture of MeOH:H2O (1:1),
yielding six fractions. Fraction 3 (300 mg) was chromatographed on Sephadex LH-20
and eluted with MeOH, affording compounds 10 (27.0 mg) and 11 (34.2 mg).
3.3.1.2. 9-epi-geniposidic acid
Brown oil. IR (KBr) max cm-1: 3418, 2928, 2860, 1686, 1639, 1275. 1H NMR (500 MHz,
CD3OD): 5.15 (1H, d, 3.4, H-1), 7.50 (1H, d, 0.9, H-3), 3.10 (1H, m, H-5), 2.85 (1H, dd,
16.4, 6.2, 1H-6), 2.14-2.04 (1H, m, 1H-6), 5.82 (1H, br s, H-7), 2.76 (1H, t, 7.5, H-9),
4.33 (1H, br d, 13.3, 1H-10), 4.21 (1H, br d, 13.3, 1H-10), 4.51 (1H, d, 7.7, H-1’), 3.15
(1H, m, H-2’), 3.45 (1H, m, H-3’), 3.55 (1H, m, 1H-4’), 3.54 (1H, m, 1H-5’), 3.80 (1H,
m, 1H-6’), 3.70 (1H, m, 1H-6’). 13C NMR (125 MHz, CD3OD): 96.8 (CH-1), 151.5 (CH-
3), 112.1 (C-4), 35.4 (CH-5), 38.4 (CH2-6), 127.2 (CH-7), 143.3 (C-8), 45.6 (CH-9),
60.1 (CH2-10), 98.7 (CH-1’), 73.4 (CH-2’), 76.3 (CH-3’), 70.1 (CH-4’), 76.9 (CH-5’),
61.3 (CH2-6’).
3.3.2. P. nuda
The methanolic extract of twigs (1.45 Kg) and leaves (426.0 g) of P. nuda were
obtained as previously mentioned (section 3.3.1.1), affording, respectively, 36.7g and
46.5 g of each crude extract. Both extracts were also partitioned as previously.
Part of the dichloromethane fraction of the twigs (1.2 g) was successively
chromatographed on silica gel and eluted with CH2Cl2 and CH2Cl2:MeOH solutions,
leading to the identification of a mixture of compounds 12-14 (153.0 mg), 15 (23.2 mg),
and a mixture of 16 and 17 (53.0 mg). The ethyl acetate fraction of leaves (790.0 mg)
was similarly chromatographed (except for the use of CH2Cl2:AcOET as eluent),
allowing the identification of compounds 18 and 19 (35.0 mg) whereas n-butanol
fraction yielded compound 20 (26.0 mg).
83
The n-butanol fraction of leaves (1.85 g) was successively chromatographed,
leading to the identification compounds 21 and 22 (in mixture, 64.2 mg). Besides,
another fraction (88.0 mg), obtained from this procedure, was purified on Sephadex
LH-20 and compounds 23 (13.0 mg) and 24 (11.0 mg) were identified. The ethyl
acetate fraction (920 mg) was similarly chromatographed, affording 18 fractions.
Fraction 5 (28.3 mg) was purified by preparative TLC and compound 25 (4.0 mg) was
obtained. Fraction 12 (114.5 mg) was rechromatographed leading to the identification
of a mixture of compounds 5 and 6 (21.0 mg). Fraction 16 (121.3 mg) was
rechromatographed leading to the isolation of compound 26 (26.0 mg).
The metanolic fraction o leaves (1.7 g) was fractionated by CC on silica gel and
eluted with CH2Cl2 and CH2Cl2:MeOH solutions (till 20 % of MeOH), affording 12
fractions. Fraction 5 (200 mg) was similarly chromatographed and, after purification by
CC on Sephadex LH-20 (eluted with MeOH), compound 27 (22.0 mg) was identified.
From fraction 10 (210 mg), compound 28 (32 mg) was, analogously, obtained.
3.3. Culture of cells
Human leukemia cell lines U937 (histiocytic lymphoma cell line) and THP-1 (acute
monocytic leukemia cell line were cultured in DMEM-F12 medium (Gibco, BRL),
supplemented with 20 mg/mL gentamycin (Gibco, BRL) and 10 % fetal bovine serum
(Gibco, BRL). The cultures were replicated every 2 days and incubated at 37 °C, with
5 % of CO2 and humidity control.
3.4. MTT assay
Cell lines were plated into a 100 L/well (1x106 cells/mL) in 96-well plates and treated
with compounds 20, 23, and 24 at concentrations of 0, 6.25, 12.5, 25.0, 50.0, 100.0
and 200 g/mL. The cells were kept at 37 °C, with 5 % of CO2 and humidity control.
Cell viability was measured by MTT assay after 48 h of incubation (Terra et al. 2013).
The assays were analyzed by ANOVA, followed by Tukey test using Graph Pad
Software 5.0 program.
84
4. Conclusion
This study led to the isolation and identification of 28 compounds from these two
species. To the best of our knowledge, besides the novel iridoid (compound 1),
compounds 2-10, 18-19, 20, and 23 are reported for the first time in this genus. This
work, then, may add relevant information related to the chemotaxonomy of this
complex genus. With respect to the MTT assay, only compound 24 showed significant
cytotoxicity.
Disclosure statement
No potential conflict of interest.
Acknowledgements
The authors are thankful to the botanist professor Dr. Sebastião José da Silva Neto
(UERJ) for the identification of plant material.
Funding
The authors are thankful to FAPERJ, CAPES, and CNPq for financial support.
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Supplemental online material
Compound 1
Table S1. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 1,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
HSQC HMBC
C H 2JCH 3JCH
C -
2 112.1 - H-3
4 143,3 - H-7; H-9; 2H-10 H-1
8 170.4 - H-3; H-5
11
CH H-3; H-1’
1 92.5 5.15 (d, 3.4) H-1; H-5
3 151.5 7.50 (d, 0.9) - -
4 - - H-1; H-3; H-7
5 35.4 3.22-3.15 (m) H-5; 2H-10
7 127.2 5.82 (brs) H-1 H-7
9 45.6 2.73 (t, 7.5) H-1
1’ 96.6 4.51 (d, 7.7)
2’ 73.4 3.22-3.15 (m)
3’ 76.3 3.45 (m)
4’ 70.1 3.55 (m)
5’ 76.9 3.54 (m)
CH2
6 38.4 2.85 (dd, 16.4, 6.2)
2.14 – 2.04 (m) H-5; H-7
10 60.1 4.33 (d, 13,3)
4.21 (d, 13.3) H-7
6’ 61.3 3.80 (m), 3.70 (m)
Compound 2
88
Table S2. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 2,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
HSQC HMBC
C H 2JCH 3JCH
C
2 112.1 - H-3
4 143,3 - H-7; H-9; 2H-10 H-1
8 170.4 - H-3; H-5
11
CH H-3; H-1’
1 96.8 5.16 (d, 7.5) H-1; H-5
3 151.5 7.50 (d, 0.9) - -
4 - - H-1; H-3; H-7
5 35.4 3.10 (m) H-5; 2H-10
7 127.2 5.82 (brs) H-1 H-7
9 45.6 2.73 (t, 7.5) H-1
1’ 98.7 4.74 (d, 7.9)
2’ 73.4 3.15 (m)
3’ 76.3 3.45 (m)
4’ 70.1 3.55 (m)
5’ 76.9 3.54 (m)
CH2
6 38.4 2.85 (dd, 16.4, 6.2)
2.14 – 2.04 (m) H-5; H-7
10 60.1 4.33 (d, 13,3)
4.21 (d, 13.3) H-7
6’ 61.3 3.80 (m), 3.70 (m)
Compound 3
89
Table S3. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 3,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
HSQC HMBC
C H 2JCH 3JCH
C
2 103.9 - 2H-1 H-1’
CH
3 77.9 4.13 (d, 8.3)
4 74.3 4.05 (t, 8.3) H-4; 1H-3
5 82.2 3.80 (m) H-4
1’ 92.5 5.42 (d, 3.8) H-3’
2’ 71.7 3.40 (m) H-1’
3’ 73.2 3.75 (m)
4’ 70.0 3.40 (m)
5’ 72.9 3.86 (m)
CH2
1 62.6 3.80 (m); 3.69 (m) H-3
6 62.0 3.80 (m); 3.70 (m)
6’ 60.9 3.82 (m); 3.75 (m)
Figure S1. Infrared spectrum of compounds 1-3.
90
CCAS2.003.001.1r.esp
8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
Methanol
0.1
115
0.8
752
0.8
8310
.89
560
.90
96
0.9
228
0.9
499
1.2
718
1.2
953
1.3
179
1.3
246
1.3
325
1.3
392
1.5
464
1.5
970
1.6
095
2.0
956
2.1
004
2.1
053
2.1
105
2.1
178
2.1
273
2.1
431
2.2
746
2.7
186
2.7
335
2.7
494
3.1
5153.1
671
3.1
698
3.1
854
3.2
659
3.2
686
3.2
845
3.3
974
3.4
569
3.4
840
3.5
384
3.6
097
3.6
467
3.6
677
3.7
510
3.7
891
3.8
657
3.8
892
3.9
283
4.0
354
4.0
518
4.1
229
4.1
394
4.2
224
4.3
188
4.3
466
4.5
098
4.5
253
4.7
380
4.7
539
5.1
450
5.1
517
5.1
667
5.4
135
5.4
211
5.8
168
7.4
997
Figure S2. 1H NMR spectrum (500 MHz, CD3OD) of compounds 1-3.
1
OO
H
O
O
OH
OHOH
HH
OH
H
HOOC
NOE H 5.15 (d, 3.4 Hz)
H 4.51 (d, 7.7 Hz)
1
4
5
7
2'
36
8
3'
4'5'
6'
9
10
1'
91
CCAS2.003.001.1r.esp
6.0 5.9 5.8 5.7 5.6 5.5 5.4 5.3 5.2 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3
Chemical Shift (ppm)
4.3
188
4.3
466
4.5
098
4.5
253
4.7
380
4.7
539
5.1
450
5.1
517
5.1
667
5.4
135
5.4
211
5.8
168
Figure S3. Expansion of 1H NMR spectrum (500 MHz, CD3OD) of compounds 1-3.
92
CCAS2.002.001.1r.esp
176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
35.3
69
4
38.3
99
2
45.6
50
4
60.1
05
86
0.7
84
76
1.2
18
16
2.0
45
06
2.5
86
7
69.8
92
17
0.0
94
37
1.7
41
07
2.9
50
77
3.1
96
37
3.4
56
37
4.2
94
17
4.8
53
8
76.8
68
87
7.8
83
58
2.2
49
4
92.2
37
99
2.5
26
89
6.7
30
19
6.8
81
89
8.9
04
1
103
.88
75
112
.11
73
127
.17
22
143
.31
40
151
.49
33
170
.38
32
Figure S4. DEPTQ spectrum (125 MHz, CD3OD) of compounds 1-3.
93
CCAS2.100.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
1
2
3
4
5
6
7
F1 C
hem
ica
l S
hift
(pp
m)
Figure S5. 1H-1H-COSY spectrum of compounds 1-3.
94
CCAS2.200.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
F1 C
he
mic
al S
hift (p
pm
)
Figure S6. HSQC spectrum of compounds 1-3.
95
CCAS2.300.001.2rr.esp
11 10 9 8 7 6 5 4 3 2 1 0 -1
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
200
F1 C
hem
ica
l S
hift
(pp
m)
Figure S7. HMBC spectrum of compounds 1-3.
96
CCAS2.500.001.2rr.esp
7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
-0.5
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
F1 C
hem
ica
l S
hift (p
pm
)
Figure S8. NOESY spectrum of compounds 1-3.
97
Figure S9. ESI-MS spectrum of compounds 1-3 (positive mode).
102.1257
203.0487
365.1018
487.2037
707.2135
1049.3313
+MS, 0.3-1.0min #(16-61)
0
1
2
3
4
5
4x10
Intens.
100 200 300 400 500 600 700 800 900 1000 m/z
98
Figure S10. ESI-MS2 (m/z 397.1064) spectrum of compound 1 (positive mode).
217.0408
235.0527
397.1064
+MS2(397.1064), 40eV, 0.1min #7
0
100
200
300
400
500
Intens.
100 150 200 250 300 350 400 m/z
99
Compound 4
Table S4. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 4,
including results of HSQC and HMBC experiments. Chemical shifts are given inppm
and coupling constants in Hz
4 Literature*
HSQC HMBC
C C H 2JCH 3JCH C**
4 37.8 3H-23; 3H-24 37.9
8 39.6 - H-9; 3H-26 2H-11; 3H-27 40.0
10 37.1 - H-7b; 3H-25 2H-11 37.3
13 142.6 - H-18 2H-11; H-19 144.9
14 41.2 - 3H-27 H-9; H-18; 3H-26 42.2
17 45.3 - H-18 H-19 46.1
20 34.6 35.7
28 184.8 - H-16a; H-18; H-22a 180.9
O-Ac 171.0 - H-3 170.6
CH
3 80.9 4.50 (dd, 10.2, 6.2) 3H-23; 3H-24 80.8
5 55.3 0.90 (m) H-7b; 3H-23; 3H-24;
3H-25
55.6
9 47.8 1.70 (m) 2H-11 3H-25; 3H-26 48.2
12 125.18 5.43 (brs) 2H-11 H-18 123.2
18 43.4 3.09 (brs) H-19 2H-16; H-22b 44.8
19 81.5 3.34 (d, 3.6) H-18 H-21; 3H-29; 3H-30 81.3
CH2
1 37.8 1.62 (m); 1.08 (m) 3H-25 38.1
2 23.7 1.70-1.60 (m) 24.1
6 18.3 1.58 (m); 1.42 (m) H-7b 18.6
7 32.3 1.70 (m); 1.30 (m) 3H-26 33.7
11 23.4 1.96 (dd, 8.5, 2.4) 23.9
15 28.0 1.60 (m); 1.05 (m) 2H-16 3H-27 28.4
16 27.3 2.28 (td, 13.3, 3.2),
1.68 (m)
H-18 29.2
21 28.0 1.80 (m) H-19; 3H-29; 3H-30 29.1
22 32.5 1.85 (m); 1.48 (m) H-16b 33.2
CH3
23 28.0 0.87 (s) 3H-24 28.2
24 16.6 0.87 (s) 3H-23 16.9
25 15.2 0.93 (s) H-9 15.3
26 17.3 0.71 (s) H-9 17.4
27 25.0 1.25 (s) 24.9
29 28.0 0.98 (s) H-19; 3H-30 28.9
30 24.3 0.97 (s) H-19; 3H-29 24.9
O-Ac 21.3 2.05 (s) 21.1
* Itokawa et al. 1989 **Pyridine-d5
100
AC4.001.001.1r.esp
9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
TMS
0.7
146
0.7
275
0.8
596
0.8
721
0.9
666
0.9
837
1.0
789
1.2
385
1.2
498
1.2
614
1.2
910
1.5
890
1.6
369
1.6
565
1.6
961
1.8
096
2.0
519
2.0
687
2.1
138
2.2
899
2.3
100
2.3
164
3.0
892
3.3
363
3.3
437
4.4
853
4.4
969
4.5
052
4.5
1775.3
076
5.4
317
7.2
837
Figure S11. 1H NMR spectrum (500 MHz, CDCl3) of compound 4.
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
O
CO2HH
O
4
101
AC4.002.001.1r.esp
220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 -20
Chemical Shift (ppm)
TMS
15.2
26
41
6.6
23
91
7.3
38
9
18.3
60
9
21.2
93
2
23.4
41
82
3.7
30
7
24.3
59
02
5.0
23
5
27.3
49
12
7.9
26
8
27.9
84
6
32.5
31
1
37.6
73
33
7.8
10
63
9.6
37
84
1.1
61
7
43.3
89
8
45.2
74
8
47.7
34
0
55.3
13
8
80.9
06
18
1.4
62
2
125
.17
88
142
.58
10
171
.04
77
184
.79
53
Figure S12. 13C NMR spectrum (125 MHz, CDCl3) of compound 4.
102
AC4.100.001.2rr.esp
7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
-0.5
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
F1 C
he
mic
al S
hift (p
pm
)
Figure S13. 1H-1H-COSY spectrum of compound 4.
103
AC4.200.001.2rr.esp
6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
F1 C
hem
ica
l S
hift
(pp
m)
Figure S14. HSQC spectrum of compound 4.
104
AC4.300.001.2rr.esp
6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift
(pp
m)
Figure S15. HMBC spectrum of compound 4.
105
Compounds 5 and 6
Table S5. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 5,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
5 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
4 40.1 - 40.4
8 39.7 - 39.0
10 37,3 - 37.4
13 138.7 - H-18 3H-27 137.9
14 41.9 - 3H-27 H-12 42.4
17 48.0 - 2H-16; H-18 47.8
19 72.5 - H-18; 3H-29 72.7
28 180.4 - H-18; 3H-27 180.7
CH
3 78.0 3.43 (dd, 10.7, 5.2) 78.2
5 55.6 0.89 (m) 3H-25 55.9
9 47.9 1.85 (m) H-12; 3H-25 48.3
12 127.6 5.60 (t, 3.4) H-18 128.1
18 54.3 3.06 (sl) H-12; 3H-29 54.7
20 41.9 1.50 (m) H-18; 3H-29 42.2
CH2
1 39.8 2.15 (m) e 2.06 (m) H-3 39.4
2 28.2 1.28 (m) 28.2
6 18.7 1.58 (m) e 1.41 (m) 19.0
7 33.4 1.64 (m); 1.40 (m) 33.6
11 23.8 2.03 (m) 24.1
15 28.9 1.28 (m) 2H-16 3H-27 29.4
16 26.6 3.12 (dt, 13.0, 4.5) 26.4
21 26.7 1.39 (m) H-18 27.0
22 38.6 1.58 (m) e 0.98 (m) 38.5
CH3
23 28.5 1.19 (s) H-3 28.8
24 16.1 0.99 (s) H-3 16.8
25 15.3 0.96 (s) 15.6
26 16.8 1.06 (s) 17.3
27 24.4 1.73 (s) 24.7
29 26.9 1.45 (s) 1H-18 27.2
30 16.8 1.06 (m) 16.5
* Chama et al. 2015.
106
Table S6. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 6,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
6 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
4 40.1 - 38.5
8 39.7 - 39.6
10 37,3 - 3H-25 37.2
13 144.6 - H-18 3H-27; H-19 143.4
14 41.9 - 3H-27 H-12 41.4
17 45.9 - 2H-16; 1H-18 45.5
28 180.7 - H-18; 3H-27 181.1
CH
3 77.9 3.43 (dd, 10.7, 5.2) 78.6
5 55.6 0.89 (m) 3H-25 55.7
9 47.9 1.85 (m) H-12; 3H-25 47.7
12 123.6 5.60 (t, 3.4) H-18 123.7
18 54.3 3.06 (sl) H-12; 3H-29 44.0
19 81.0 3.62 (m) H-18; 3H-29 81.3
20 41.9 1.50 (m) H-18; 3H-29 34.9
CH2
1 39.8 2.15 (m) e 2.06 (m) H-3 38.7
2 28.2 1.28 (m) 26.8
6 18.7 1.58 (m) e 1.41 (m) 18.5
7 33.4 1.64 (m) e 1.40 (m) 32.8
11 23.8 2.03 (m) 23.9
15 28.9 1.28 (m) 2H-16 3H-27 28.3
16 26.6 3.12 (td, 13.0, 4.5) 27.5
21 26.7 1.39 (m) H-18 28.4
22 38.6 1.58 (m) e 0.98 (m) 32.8
CH3
23 28.5 1.19 (s) H-3 27.6
24 15.1 0.88 (s) H-3 15.1
25 15.3 0.96 (s) 14.6
26 16.8 1.06 (s) 16.6
27 24.4 1.73 (s) 24.0
29 26.9 1.45 (s) H-18; 1H-19 27.5
30 24.2 1.14 (s) H-19 24.0
*Wang et al. 2011.
107
AC 1.003.001.1r.esp
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
0.7
3340.8
649
0.8
728
0.8
814
0.8
957
0.9
076
1.0
180
1.1
077
1.2
270
1.2
313
1.3
799
1.4
150
1.4
439
1.5
623
1.6
490
1.7
228
1.8
192
1.9
693
1.9
815
2.0
441
2.0
673
2.1
393
2.1
481
2.1
649
2.1
780
2.3
434
2.8
331
2.8
398
3.0
530
3.4
127
3.4
228
3.4
338
3.4
445
3.6
284
4.1
099
4.1
209
5.0
313
5.5
649
5.5
716
5.5
9785
.60
49
5.6
116
7.1
883
7.5
556
8.7
046
Figure S15. 1H NMR spectrum (500 MHz, Pyridine-d5) of compounds 5 and 6.
5
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
HO
CO2H
HO
H
6
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
HO
CO2H
HO
H
108
AC 1.002.001.1r.esp
184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
15.2
44
51
5.3
52
81
6.2
98
91
6.5
55
31
6.9
99
51
7.2
84
8
18.7
22
02
3.8
02
9
24.4
67
42
4.6
33
5
26.1
79
02
6.7
13
52
7.8
87
1
28.5
58
82
8.6
20
2
29.0
93
22
9.7
72
133.3
79
7
37.1
35
3
38.2
76
43
8.7
89
23
9.1
61
13
9.7
93
14
0.1
47
04
1.8
94
8
42.1
51
2
44.5
95
9
47.5
60
7
48.0
84
3
48.1
49
3
54.4
11
05
5.6
53
3
72.4
70
4
77.9
84
6
81.0
61
4
123
.58
631
27
.82
94
139
.73
90
144
.65
37
180
.46
19
180
.69
31
Figure S16. DEPTQ spectrum (125 MHz, Pyridine-d5) of compounds 5 and 6.
109
AC 1.100.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
1
2
3
4
5
6
7
8
F1 C
he
mic
al S
hift (p
pm
)
Figure S17. 1H-1H spectrum of compounds 5 and 6.
110
AC 1.200.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
F1 C
he
mic
al S
hift (p
pm
)
Figure S18. HHSQC spectrum of compounds 5 and 6.
111
AC 1.300.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift (p
pm
)
Figure S19. HMBC spectrum of compounds 5 and 6.
112
Compounds 7 and 8
Table S7. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 7,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
7 Literature*
HSQC HMBC
C C H 2JCH 3JCH C**
4 39.8 - H-3 40.7
8 40.2 - 40.9
10 39.6 - 3H-25 39.5
13 144.6 - H-18 3H-27; H-19 144.9
14 41.9 - 3H-27 H-12 42.8
17 45.8 - 2H-16; H-18 46.8
20 35.1 - H-18; 3H-29 36.2
28 180.7 - H-18; 3H27 182.5
CH
2 68.5 4.08 (m) H-3; 2H-1 69.6
3 83.7 3.37 (dd, 9.4, 2.4) 2H-1; H-5 84.7
5 55.8 1.05 (m) 3H-25 57.0
9 48.0 1.94 (m) H-12; 3H-25 49.4
12 123.1 5.54 (m) H-18 124.9
18 54.4 3.03 (brs) H-12; 3H-29 54.3
19 81.2 3.59 (brs) H-18; 3H-29 82.6
CH2
1 47.3 2.2 (m);1.48 (m) H-3 48.1
6 18.7 1.42 (m) 19.8
7 33.4 1.64 (m)/1.39 (m) 34.0
11 24.1 2.10 (m) 25.0
15 28.9 1.57 (m) 2H-16 3H-27 29.6
16 28.6 1.43 (m) 28.7
21 29.0 1.08 (m) H-18 29.6
22 33.4 1.58 (m) 34.2
CH3
23 28.6 1.18 (s) H-3 29.4
24 17.0 1.06 (s) H-3 17.5
25 17.3 1.00 (s) 17.1
26 17.4 0.90 (s) 17.9
27 24.5 1.62 (s) 25.2
29 28.6 1.18 (s) H-18; 1H-19 28.8
30 24.6 1.10 (s) H-19 25.3
*Ponou et al. 2011. ** CD3OD.
113
Table S8. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 8,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
8 Literature*
HSQC HMBC
C C H 2JCH 3JCH C**
4 39.8 - 1H-3 39.9
8 40.2 - 40.5
10 39.6 - 3H-25 38.5
13 139.7 - H-18 3H-27; H-19 140.0
14 41.9 - 3H-27 H-12 42.2
17 48.0 - 2H-16; H-18 48.3
19 72.5 - H-18; 3H-29 72.7
20 41.9 - H-18; 3H-29 42.4
28 180.4 - H-18; 3H27 180.6
CH
2 68.5 4.08 (m) H-3; 2H-1 68.6
3 83.7 3.37 (dd, 9.4, 2.4) 2H-1; H-5 83.9
5 55.8 1.05 (m) 3H-25 56.0
9 48.0 1.94 (m) H-12; 3H-25 47.9
12 127.7 5.54 (m) H-18 128.0
18 54.4 3.03 (brs) H-12; 3H-29 54.6
20 41.9 - H-18; 3H-29 42.4
CH2
1 47.3 2.2 (m)/1.48 (m) H-3 48.0
6 18.7 1.42 (m) 19.0
7 33.4 1.64 (m)/1.39 (m) 33.5
11 24.1 2.10 (m) 24.1
15 28.9 1.57 (m) 2H-16 3H-27 29.3
16 26.2 1.43 (m) 26.4
21 26.7 1.08 (m) H-18 27.1
22 38.5 1.58 (m) 38.5
CH3
23 28.6 1.18 (s) H-3 29.3
24 17.4 1.06 (s) H-3 17.7
25 17.3 1.00 (s) 16.9
26 17.4 0.90 (s) 17.2
27 24.5 1.62 (s) 24.7
29 26.7 1.18 (s) H-18; H-19 27.1
30 16.5 1.10 (d, 5.8) H-19 16.9
*Taniguchi et al. 2002. **Pyridine-d5.
114
AC2.003.001.1r.esp
10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
0.8
4000
.85
35
0.8
586
0.8
733
0.9
846
0.9
972
1.0
142
1.0
603
1.0
704
1.0
957
1.1
110
1.1
241
1.2
522
1.2
589
1.2
778
1.3
203
1.4
203
1.5
369
1.5
967
1.6
238
1.6
958
1.7
419
1.8
404
1.9
893
2.0
366
2.0
781
2.0
961
2.1
236
2.1
397
2.1
498
2.2
230
2.8
232
3.0
364
3.0
913
3.3
601
3.3
650
3.3
790
3.3
836
3.5
883
3.6
146
4.0
698
4.0
783
4.0
914
4.0
966
4.1
101
4.1
531
4.1
735
4.1
842
5.5
367
5.5
431
5.5
648
5.5
712
5.5
779
5.8
683
5.8
799
Figure S20. 1H NMR spectrum (500 MHz, Pyridine-d5) of compounds 7 and 8.
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
HO
HO
CO2HH
7
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
HO
HO
CO2HH
HO
8
115
AC2.002.001.1r.esp
184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
15.6
30
91
6.5
40
9
17.3
24
51
7.4
14
8
18.7
90
62
3.8
93
22
4.0
44
9
24.4
56
52
4.6
11
8
26.1
64
62
6.7
06
3
26.8
79
6
28.1
58
0
28.5
94
9
28.9
05
52
8.9
70
52
9.0
39
1
29.1
07
7
29.7
46
93
3.4
15
83
5.4
85
03
8.2
80
03
8.4
64
23
9.6
30
6
42.1
36
74
4.5
92
3
45.8
41
84
7.3
07
9
47.6
07
6
47.6
79
94
8.0
62
6
48.1
99
9
54.3
82
25
5.7
43
65
5.8
12
2
68.3
86
2
72.4
81
3
81.0
14
4
83.6
36
1
123
.56
10
127
.73
55
139
.73
90
144
.67
18
180
.41
86
180
.65
69
Figure S21. DEPTQ NMR spectrum (125 MHz, Pyridine-d5) of compounds 7 and 8.
116
AC2.100.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
1
2
3
4
5
6
7
8
F1 C
hem
ica
l S
hift
(pp
m)
Figure S22. 1H-1H-COSY spectrum of compounds 7 and 8.
117
AC2.200.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift (p
pm
)
Figure S23. HSQC spectrum of compounds 7 and 8.
118
AC2.300.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
180
F1 C
he
mic
al S
hift (p
pm
)
Figure S24. HMBC spectrum of compounds 7 and 8.
119
Compound 9
Table S9. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound 9,
including results of HSQC and HMBC experiments. Chemical shifts are given in ppm
and coupling constants in Hz
9 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
4 39.1 - 3H-23; 3H-
24
38.9
8 39.5 - 3H-26; H-9 3H-27 39.8
10 37.2 - 3H-25 2H-2 37.1
13 144.6 - H-18 3H-27; 2H-11 143.4
14 41.9 - 3H-27 3H-26 41.6
17 46.4 - H-18 46.6
20 30.7 - 3H-29; 3H-
30
30.6
28 180.1 - 181.0
CH
3 78.1 3.45 (m) 2H-2 3H-23;3H24 78.7
5 55.6 0.85 (m) 3H-23; 3H-25 55.2
9 48.1 1.65 (m) 3H-25; 3H-26 47.6
12 122.9 5.60 (m) 2H-11 H-18 122.1
18 41.9 3.30 (m) 41.3
CH2
1 38.7 1.61 (m); 1.02
(m)
3H-25 38.5
2 27.8 1.95 (m); 1.05
(m)
27.4
6 18.5 1.65 (m); 1.40
(m)
18.3
7 33.0 1.65 (m); 1.35
(m)
32.6
11 23.0 1.95 (m) 23.1
15 28.1 2.19 (m); 1.99
(m)
3H-27 27.7
16 23.4 2.20 (m); 2.00
(m)
23.4
19 46.2 1.85 (m), 1.30
(m)
3H-29; 3H-30 45.8
21 34.0 1.45 (m), 1.25
(m)
3H-29; 3H-30 33.8
22 33.0 2.00 (m), 1.78
(m)
32.3
CH3
23 28.8 1.12 (s) 3H-24; H-3 28.1
24 16.2 1.03 (s) 15.6
25 15.1 0.87 (s) 15.3
26 17.3 1.01 (s) H-9 16.8
120
27 25.9 1.18 (s) 26.0
29 33.0 .094 (s) 33.1
30 23.6 0.99 (s) 3H-29 23.6
Me-O-28 3.58 (s) -
*Mahato & Kundo, 1994.
121
AC 8.003.001.1r.esp
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
0.8
846
0.9
413
0.9
999
1.0
179
1.1
094
1.2
333
1.2
745
1.2
958
1.5
244
1.5
506
1.6
787
1.8
066
1.8
334
1.9
466
1.9
945
2.2
044
3.2
814
3.2
921
3.4
257
3.4
382
3.4
492
3.4
584
3.5
908
Figure S25. 1H NMR spectrum (500 MHz, Pyridine-d5) of compound 9.
12
3 4
2324
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
HO
CO2CH3H
9
122
AC 8.002.001.1r.esp
220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 -20
Chemical Shift (ppm)
15.3
09
51
6.3
13
41
7.2
16
2
18.5
66
72
3.4
70
7
23.5
32
1
23.5
93
5
25.9
33
5
27.8
29
32
8.0
89
3
28.5
48
0
30.7
29
13
2.9
71
6
33.0
33
0
33.9
86
43
7.1
49
73
8.7
17
03
9.1
50
33
9.5
29
5
41.7
90
1
41.9
49
04
6.2
60
74
6.4
66
5
47.8
96
5
55.5
91
9
65.5
62
3
77.8
87
1
122
.34
41
125
.43
16
144
.60
32
180
.01
42
Figure S26. 13C NMR spectrum (125 MHz, Pyridine-d5) of compound 9.
123
AC 8.100.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
1
2
3
4
5
6
7
8
F1 C
hem
ica
l S
hift
(pp
m)
Figure S27. 1H-1H-COSY spectrum of compound 9.
124
AC 8.200.001.2rr.esp
10 9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
200
F1 C
hem
ica
l S
hift (p
pm
)
Figure S28. 1H-1H-COSY spectrum of compound 9.
125
Compound 10
Table S10. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound
10, including results of HSQC and HMBC experiments. Chemical shifts are given in
ppm and coupling constants in Hz
10 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
2 136.7 - H-14b 138.2
3 145.2 H-14b H-5 144.8
7 129.7 - H-5; H-9 130.8
8 120.8 - H-6; H-12 122.1
13 141.7 - H-9; H-11 135.8
16 113.5 - H-17 114.8
22 171.8 - H-17 174.7
CH
5 134.57 8.22 (d, 5.6) H-6 136.7
6 113.52 8.10 (d, 5.6) H-5 114.9
9 121.54 8.11 (d, 8.1)
10 119.80 7.31 (t, 8.1) H-10 H-11 122.7
11 128.90 7.62 (t, 8.1) H-12 121.2
12 111.89 7.70(d, 8.1) H-9 130.3
15 34.02 3.57 (m) H-10 113.4
17 150.70 7.50 (s) H-17; H-21 36.0
19 133.75 5.87 (dd, 17.1, 9.5) H-21 151.2
20 44.67 2.61 (m) 2H-18; H-20 134.6
21 95.79 5.67 (d, 6.8) 2H-18 46.4
CH2 95.8 2.61 (m) H-20 H-17; H-1’A 97.6
14 34.5 3.70 (m)
3.23 (m)
36.6
18 117.6 4.95-4.80 (m) H-20 118.8
Glucose
1’ 98.9 4.76 (d, 8.4) H-2’A H-21 100.4
2’ 73.2 3.21 (dd) 74.6
3’ 76.5 3.45 (m) 78.4
4’ 70.2 3.30 (m) 71.7
5’ 77.1 3.35 (m) 78.1
6’ 61.5 3.94 (dd, 12.1, 1.9)
3.70 (m)
63.2
*Lin et al. 2011.
126
AC 1323.003.001.1r.esp
9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
1.3
076
2.6
091
2.6
229
3.1
904
3.2
068
3.2
791
3.2
981
3.3
295
3.3
676
3.3
899
3.4
082
3.6
834
3.6
8803
.71
94
3.8
4633.8
887
3.9
125
3.9
305
3.9
342
3.9
546
4.7
421
4.7
570
4.7
738
5.6
629
5.6
763
5.8
572
5.8
914
6.7
023
6.7
203
6.9
739
6.9
916
7.2
9577.3
110
7.3
256
7.5
020
7.6
185
7.6
914
7.7
076
8.0
902
8.1
015
8.2
1508
.22
66
Figure S29. 1H NMR spectrum (500 MHz, CD3OD) of compound 10.
10
1
NN
CO2H
O
OO
HO
OHOH
OH
H
22
1615
2019
1714
32
7
6
58
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
127
AC 1323.002.001.1r.esp
176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
34.5
42
5
44.6
64
5
61.1
60
361.4
85
3
70.0
43
77
0.2
35
17
3.2
46
87
6.5
43
87
6.6
55
77
7.0
71
0
95.7
87
6
98.8
93
2
102
.25
88
111
.88
61
113
.52
20
115
.19
76
117
.61
71
117
.98
18
119
.80
54
120
.82
74
121
.54
24
128
.90
19
129
.73
61
134
.56
78
134
.70
50
141
.71
43
150
.69
88
171
.83
49
Figure S30. 13C NMR spectrum (125 MHz, CD3OD) of compound 10.
128
AC 1323.100.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
1
2
3
4
5
6
7
8
F1 C
hem
ica
l S
hift
(pp
m)
Figure S31. 1H-1H-COSY spectrum of compound 10.
129
AC 1323.200.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
F1 C
he
mic
al S
hift (p
pm
)
Figure S32. HSQC spectrum of compound 10.
130
AC 1323.300.001.2rr.esp
9 8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
200
F1 C
he
mic
al S
hift (p
pm
)
Figure S33. HMBC spectrum of compound 10.
131
Compound 11
Table S11. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 11, including
results of HSQC and HMBC 2D experiments. Chemical shifts in ppm and coupling
constants in Hz
11 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
2 129.6 -- 130.4
7 105.9 -- 105.3
8 126.1 -- H-10 127.5
13 136.8 -- H-9; H-11 138.2
16 111.0 H-17 109.3
22 175.0 -- H-17 175.4
CH
3 50.9 4.47 (d, 11.4) 52.5
9 117.7 7.47 (d, 7.8) H-11 119.1
10 119.1 7.05 (t, 7.8) H-12 120.6
11 121.9 7.15 (dd, 7.8, 7.3) H-9 123.4
12 110.5 7.33 (d, 7.3) H-10 112.3
15 32.6 3.01 (m) 33.8
17 150.5 7.59 (s) 151.2
19 134.8 5.87 (m) 136.2
20 44.3 2.72 (m) 2H-18 45.6
21 95.2 5.84 (d, 9.3) H-17; H-1’ 96.7
CH2
5 41.6 3.35 (m) 43.0
6 18.2 3.10-2.90 (m) 19.6
14 33.7 2.39 (m)
2.14 (m)
35.1
18 118.0 5.83 (d, 17.4)
5.22 (d, 10.7)
119.1
Glucose
1’ 100.0 4.76 (d, 7.9) H-2’ H-21 100.3
2’ 73.3 3.21 (m) H-3’ 74.6
3’ 76.6 3.42 (m) H-4’ 77.9
4’ 70.4 3.29 (m) H-3’ 72.0
5’ 77.3 3.35 (m) H-4’ 78.1
6’ 61.7 3.94 (dd, 12.3, 2.0)
3.70 (m)
H-4’ 52.9
Berger et al. 2015.
132
AC 1321.003.001.1r.esp
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
Methanol
0.9
177
0.9
3720.9
519
0.9
595
0.9
668
0.9
741
1.2
734
1.3
125
1.4
211
1.4
354
1.5
605
1.5
709
2.1
100
2.3
913
2.6
012
2.6
159
3.0
021
3.0
271
3.1
882
3.2
703
3.2
794
3.2
880
3.2
978
3.3
673
3.4
085
3.4
256
3.7
081
3.7
194
3.7
655
3.7
707
3.8
582
3.9
327
4.0
260
4.0
776
4.4
824
4.7
567
4.7
726
5.2
077
5.2
293
5.3
144
5.3
492
5.4
121
5.6
565
5.6
702
5.8
340
5.8
526
5.8
691
5.8
895
5.9
072
7.0
556
7.0
709
7.1
475
7.3
034
7.3
204
7.3
372
7.4
803
7.4
922
7.5
947
7.6
872
7.7
030
8.0
719
8.0
829
8.2
034
8.2
156
Figure S34. 1H NMR spectrum (500 MHz, CD3OD) of compound 11.
1
NN
CO2H
O
OO
HO
OHOH
OH
H
22
1615
2019
1714
32
7
6
5
8
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
133
AC 1321.002.001.1r.esp
184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
18.2
02
0
32.6
17
7
33.7
11
9
41.6
31
2
44.3
07
04
4.7
15
1
50.9
29
9
61.4
88
9
70.2
35
1
73.2
50
4
76.5
47
47
7.0
63
87
7.3
31
07
7.8
94
4
92.2
23
49
5.1
99
09
5.7
73
2
98.8
93
2
105
.88
44
110
.80
64
111
.88
98
112
.00
53
113
.48
59
117
.54
84
117
.67
48
119
.13
37
119
.75
49
120
.83
46
121
.50
99
121
.94
68
126
.13
58
128
.82
97
129
.11
13
129
.63
86
133
.85
28134
.62
92
134
.71
22
134
.83
14
136
.79
59
141
.65
65
143
.70
04
150
.51
10
152
.03
13
175
.09
94
Figure S35. 13C NMR spectrum (125 MHz, CD3OD) of compound 11.
134
AC 1321.100.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
8.0
8.5
F1 C
hem
ica
l S
hift (p
pm
)
Figure S36. 1H-1H-COSY spectrum of compound 11.
135
AC 1321.200.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
F1 C
hem
ica
l S
hift
(pp
m)
Figure S37. HSQC spectrum of compound 11.
136
AC 1321.300.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift (p
pm
)
Figure S38. HMBC spectrum of compound 11.
137
Compounds 12-14
Figure S39. GC/MS chromatogram (a) and LRMS of compounds 12 (b), 13 (c), and 14
(d).
(b)
(c)
(d)
(a)
138
Compound 15
Table S12. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 15, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
15 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
3 140.3 - 3H-20 2H-1; 2H-4 140.3
CH
2 123.1 5.40 (td, 7.0, 1.3) 2H-1 3H-20 123.0
7 32.7 1.39 (m) 3H-19 32.7
11 32.8 32.7
15 28.0 1.54 (m) 3H-17 27.9
CH2
1 59.4 4.17 (d, 6.9) 59.4
4 39.9 2.01 (m) 3H-20 39.8
5 25.1 1.48 (m) 25.2
6 36.6 36.6
8 37.3 1.08 (m) 37.4
9 24.5 1.27 (m) 24.4
10 37.4 1.28 (m) 37.4
12 24.5 1.37 (m) 39.3
13 39.4 1.16 (m) 24.4
14 39.4 1.16 (m) 3H-17 39.3
CH3
16 22.7 0.86 (d, 6.5) 22.7
17 22.6 0.87 (d, 6.5) 22.6
18 19.7 0.88 (d, 6.5) 19.7
19 19.7 0.92 (s) 19.7
20 16.1 1.70 (s) 16.1
*Miranda et al. 2012.
139
AC12 2.001.001.1r.esp
9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
TMS
0.0
154
0.0
285
0.8
6020
.87
33
0.8
815
0.8
953
1.0
914
1.1
637
1.2
663
1.2
772
1.2
855
1.3
016
1.3
322
1.3
608
1.5
860
1.6
925
1.9
945
2.0
000
2.0
128
2.0
311
2.1
257
3.9
4294.1
702
4.1
839
5.4
201
5.4
342
5.4
455
5.4
479
7.2
834
8.2
191
Figure S40. 1H NMR spectrum (500 MHz, CDCl3) of compound 15.
OH
15
1
23 4
5
67
8
9
10 11
1216
17
18
13
141520
19
140
AC12 2.002.001.1r.esp
152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
16.1
83
41
9.7
22
31
9.7
54
8
22.6
29
32
2.7
23
2
24.4
74
62
4.7
99
62
5.1
39
0
27.9
84
6
29.7
03
5
32.6
97
23
2.7
98
3
36.6
65
83
7.2
94
23
7.3
62
83
7.4
35
03
9.3
74
23
9.8
76
2
59.4
48
6
63.2
07
8
88.8
43
4
112
.80
70
123
.06
99
140
.35
65
Figure S41. 13C NMR spectrum (125 MHz, CDCl3) of compound 15.
141
AC12 2.100.001.2rr.esp
7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
-0.5
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
F1 C
hem
ica
l S
hift
(pp
m)
Figure S42. 1H-1H-COSY spectrum of compound 15.
142
AC12 2.200.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
F1 C
hem
ica
l S
hift
(pp
m)
Figure S43. HSQC spectrum of compound 15.
143
AC12 2.300.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
F1 C
hem
ica
l S
hift
(pp
m)
Figure S44. HMBC spectrum of compound 15.
144
Compounds 16 and 17
Table S13. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 16 and 17,
including results of HSQC and HMBC experiments. Chemical shifts in ppm and
coupling constants in Hz
Literature Literature*
C C H C C H C
5 140,7 - 141,0 140,7 - 141,0
10 36,7 - 37,0 36,7 - 37,0
13 42,3 - 42,4 42,1 - 42,4
CH
3 78,4 3,98 (m) 78,2 78,4 3,98 (m) 78,2
6 121,7 5,34 (d, 2,9) 122,0 121,7 5,34 (d, 2,9) 122,0
8 31,8 32,1 31,8 32,1
9 50,1 50,4 50,1 50,4
14 56,7 56,9 56,7 57,0
17 57,6 56,3 56,7 56,2
20 36,7 36,5 36,6 36,5
22 - 138,6 5,21 (dd, 15,1, 8,9) 138,3
23 - 129,2 5,05 (m) 129,5
24 46,6 46,1 51,2 51,5
25 30,8 29,6 31,9 32,7
CH2
2 30,0 30,3 30,0 30,3
4 39,1 2,72 (dd, 13.3, 2.7);
2,47 (m)
39,4 39,1 2,72 (dd, 13.3, 2.7);
2,47 (m)
39,4
7 31,9 32,2 31,9 32,2
11 21,1 21,4 21,3 21,5
12 39,7 40,0 39,6 39,9
15 24,2 24,6 24,3 24,7
16 29,1 28,6 29,2 29,4
22 34,5 34,3 - -
23 26,2 26,5 - -
28 23,9 23,5 25,5 25,7
CH3 -
18 12,3 0,65 (s) 12,0 11,9 0,67 (s) 12,0
19 19,4 0,92 (s) 19,3 19,7 0.93 (s) 19,3
21 19,2 0,98 (d, 6.6) 19,1 21,3 1,07 (d, 6,6) 21,5
26 19,3 0,88 (d, 1.5) 19,5 21,1 0,88 (d, 1.5) 21,4
27 19,6 0,87 (d, 2.4) 20,0 19,8 0,87 (d, 2.4) 20,0
29 12,7 0,86 (m) 12,2 19,9 0,86 (m) 12,6
Glucose
1’ 102,4 5,05 (m) 102,6 102,4 5,05 (m) 102,6
2’ 78,3 4,27 (m) 75,4 78,3 4,27 (m) 75,4
3’ 75,8 4,05 (t, 8,2) 78,6 75,8 4,05 (t, 8,2) 78,6
4’ 72,2 4,27 (m) 71,7 72,2 4,27 (m) 71,7
5’ 79,1 4,27 (m) 78,5 79,1 4,27 (m) 78,5
6’ 63,3 4,41 (dd, 11.7, 2,4;
4,54 (dd, 11,7, 2,4)
62,9 63,3 4,41 (dd, 11.7, 2,4);
4,54 (dd, 11,7, 2,4)
62,9
Kojima et al. 1990.
145
EMMPN 23(8).003.001.1r.esp
10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
0.6
5000
.66
55
0.8
016
0.8
157
0.8
459
0.8
596
0.9
243
0.9
273
0.9
844
1.0
738
1.0
921
1.2
419
1.2
519
1.3
685
1.3
804
1.5
039
1.5
296
1.5
741
1.7
267
1.7
505
1.9
442
1.9
494
2.1
160
2.1
401
2.4
687
2.4
952
2.7
085
2.7
140
2.7
350
2.7
405
3.9
676
3.9
731
3.9
795
4.0
393
4.0
561
4.0
719
4.2
666
4.2
825
4.2
977
4.4
097
4.4
204
4.5
494
4.5
680
4.5
729
5.1
834
5.2
0115
.33
66
5.3
433
7.1
955
7.5
638
8.7
042
Figure S45. 1H NMR spectrum (500 MHz, Py-d5) of compounds 16 and 17.
1
2
3
4 6
7
8
9
10
11
12
13
14
19
18
21
17
16
15
20
23
24
25
27
5
22 R2
Glu16
1
2
3
4 6
7
8
9
10
11
12
13
14
19
18
21
17
16
15
20
23
24
25
27
5
22 R2
Glu17
146
EMMPN 23(8).002.001.1r.esp
152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
11.7
60
21
1.9
26
31
2.3
09
11
8.7
94
81
8.9
68
11
9.2
06
41
9.7
62
52
1.0
84
22
1.2
57
6
29.2
52
72
9.8
23
23
0.0
39
93
1.8
38
23
1.9
53
83
6.7
06
13
7.2
62
23
9.1
18
34
0.5
70
04
2.1
30
04
2.2
67
34
5.8
27
85
0.1
32
35
1.2
08
55
5.8
52
45
6.0
33
05
6.6
18
05
6.7
11
9
62.6
12
5
71.4
81
5
75.1
21
57
7.9
09
37
8.2
56
07
8.3
86
0
102
.35
68
121
.70
54
129
.24
55
138
.61
28
140
.70
01
Figure S46. 13C NMR spectrum (125 MHz, Py-d5) of compounds 16 and 17.
147
Compounds 18 and 19
Table S14. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 18, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
18 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
5 155.9 - H-6 2H-4 156.2
7 150.6 - H-6 H-7’’ 151.2
8 104.6 - H-7’’ H-6 105.2
9 152.1 - H-7’’ 152.5
10 103.8 - 2H-4 H-6 104.5
1’ 130.5 - H-2 H-5’ 131.2
3’ 144.6 - H-5’ 144.1a
4’ 144.4 - H-2’ 144.7b
1’’ 133.9 - H-7’’ H-5’’; 2H-8’’ 134.5
3’’ 144.9 - H-5 145.0b
4’’ 143.8 - H-6’’ 145.4b
9’’ 169.4 - 2H-8’’ H-7’’ 168.9
CH
2 78.3 4.90 H-2’; H-4; H-6’ 79.0
3 65.2 4.27 (m) 2H-4 65.8
6 94.9 6.23 (s) 95.8
2’ 113.7 6.99 (d, 1.8) H-2; H-6’ 114.4a
5’ 114.5 6.77 (d, 8.1) 144.1b
6’ 117.8 6.80 (dd, 1.8, 8.1) H-2; H-2’ 118.4
2’’ 113.6 6.55 (d, 2.1) H-6’’; H-7’’ 115.4a
5’’ 115.5 6.62 (d, 8.2) 115.8a
6’’ 117.8 6.46 (dd, 2.1, 8.2) H-2’’; H-7’’ 118.4
7’’ 33.9 4.56 (d, 5.6) 2H-8’’ H-2’’; H-6’’ 34.5
CH2
4 28.1 3.05-2.95 (m) 28.9
8’’ 37.2 3.07 (dd, 15.7, 7.0)
2.90 (m)
38.0
*Nonaka & Nishioka, 1982. **Acetone-d6 + D2O. Letters a and b indicate signals that
may be interchanged.
148
Table S15. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 19, including
results of HSQC and HMBC 2D experiments. Chemical shifts in ppm and coupling
constants in Hz
19 Literature*
HSQC HMBC
C C H 2JCH 3JCH C**
5 155.85 - H-6 2H-4 156.2
7 150.67 - H-6 H-7’’ 151.2
8 104.78 H-7’’ H-6 105.4
9 152.02 - H-7’’ 152.6
10 103.86 - 2H-4 H-6 104.7
1’ 130.30 - H-2 H-5’ 131.0
3’ 144.51 - H-5’ 144.3b
4’ 144.35 - H-2’ 144.8b
1’’ 133.85 - H-7’’ H-5’’; 2H-8’’ 134.4
3’’ 144.90 - H-5’’ 144.9b
4’’ 143.71 H-2’’; H-6’’ 145.5b
9’’ 169.37 - 2H-8’’ H-7’’ 168.9
CH
2 78.80 4.96 H-2’; H-4; H-6’ 79.4
3 65.61 4.22 (m) 2H-4 66.0
6 95.00 6.22 (s) 96.0
2’ 113.60 6.85 (d, 1.8) H-2; H-6’ 114.5a
5’ 114.59 6.70 (d, 8.1) 114.8a
6’ 117.91 6.63 (dd, 1.8, 8.1) H-2; H-2’ 118.7c
2’’ 113.94 6.65 (d, 2.1) H-6’’; H-7’’ 115.3a
5’’ 115.09 6.71 (d, 8.2) 115.9a
6’’ 117.9 6.56 (dd, 2.1, 8.2) H-2’’; H-7’’ 118.5c
7’’ 33.70 4.56 (d, 5.6) 2H-8’’ H-2’’; H-6’’ 34.2
CH2
4 27.89 2.90-2.85 (m) 28.8
8’’ 36.94 3.02 (dd, 15.7, 6.9)
2.92(m)
37.6
*Nonaka & Nishioka, 1982. **Acetone-d6 + D2O. Letters a-c indicate signals that may
be interchanged.
149
MPNA 15-29.003.001.1r.esp
8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
Methanol
0.1
223
0.8
481
0.9
179
1.0
348
1.1
794
1.2
493
1.2
539
1.2
667
1.2
941
1.3
057
1.3
640
1.5
297
1.6
056
1.6
188
1.9
300
1.9
327
2.0
758
2.2
454
2.2
808
2.2
958
2.3
104
2.8
602
2.8
632
2.8
681
2.8
910
2.8
947
2.9
035
2.9
069
2.9
398
3.0
664
3.7
910
3.8
182
3.8
264
3.8
499
3.8
606
3.8
637
3.9
210
3.9
250
4.2
212
4.2
615
4.2
676
4.4
677
4.5
687
6.2
187
6.2
211
6.2
257
6.2
285
6.4
701
6.4
744
6.5
461
6.5
500
6.6
181
6.6
419
6.6
946
6.7
041
6.7
819
6.7
904
6.8
509
6.9
891
7.0
193
Figure S47. 1H NMR spectrum (500 MHz, CD3OD) of compounds 18 and 19.
18
12
3
456
7
89
10
1'
2'
3'
4'
5'
6'
6'''
5'''
4'''
3'''
2'''
1'''9'' 7''
8''
O
OH
OH
OH
OH
O
O
OH
OH
19
12
3
456
7
89
10
1'
2'
3'
4'
5'
6'
6'''
5'''
4'''
3'''
2'''
1'''9'' 7''
8''
O
OH
OH
OH
OH
O
O
OH
OH
150
MPNA 15-29.002.001.1r.esp
168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
24.5
50
42
4.6
58
82
7.8
87
12
8.0
67
72
8.6
70
7
33.6
10
8
33.7
66
13
3.9
71
9
36.9
40
33
7.1
64
2
47.1
02
1
47.2
71
84
7.4
41
54
7.6
11
24
7.7
81
04
7.9
54
3
48.1
24
0
65.1
68
76
5.6
05
6
78.3
24
17
8.8
29
7
94.8
59
69
5.0
04
0
103
.80
44
103
.86
22
104
.64
22
104
.76
50
113
.60
51
113
.94
09
114
.51
87
114
.59
45
115
.09
29
115
.15
42
117
.79
04
117
.91
32
117
.99
62
130
.29
94
130
.53
78
133
.87
81
133
.99
72
143
.79
07
144
.35
40
144
.41
18
144
.56
71
144
.89
93
144
.95
35
150
.66
99
152
.02
05
152
.11
44
155
.85
55
155
.92
06
169
.36
85
Figure S48. 1H NMR spectrum (125 MHz, CD3OD) of compounds 18 and 19.
151
MPNA 15-29.200.001.2rr.esp
7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
F1 C
he
mic
al S
hift (p
pm
)
Figure S49. HSQC spectrum of compounds 18 and 19.
152
MPNA 15-29.300.001.2rr.esp
7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift
(pp
m)
Figure S50. HMBC spectrum of compounds 18 and 19.
153
Compounds 20
Table S16. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 20, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
20 Literature*
HSQC HMBC
C C H 2JCH 3JCH C**
3 107.97 - 2H-1’;
H-2
2H-2’; H-5 110.8
4 126.65 - 2H-1’; H-2; H-
6; H-8
127.0
9 136.78 - H-2; H-5; H-7 136.5
CH
2 122.95 7.25 (s) 2H-1’ 123.8
5 118.77 7.64 (d, 8,0 Hz) H-7 118.9
6 117.53 7.09 (t, 7,5 Hz) H-8 118.7
7 121.44 7.15 (t, 7,5 Hz) H-5 121.7
8 111.20 7.41 (d, 8,0 Hz) H-6 111.9
CH2
1’ 18.95 3.27 (m) 2H-2’ 19.0
2’ 66.47 3.62 (m) 2H-1’ NMe3 65.5
CH3 -
N-M3 52.27 3.23 (s) 52.5
*Martins et al. 2009. **DMSO-d6.
154
MPN 29.003.001.1r.esp
8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
3.2
592
3.2
735
3.2
921
3.6
018
3.6
192
3.6
332
7.0
717
7.0
854
7.1
501
7.2
474
7.3
972
7.4
131
7.6
269
7.6
425
Figure S51. 1H NMR spectrum (500 MHz, CD3OD) of compound 20.
N
H
N+
12
34
56
7
89
1'
2'
20
155
MPN 29A.002.001.1r.esp
160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
18.9
53
6
52.2
70
1
66.4
98
1
107
.96
86
111
.19
69
117
.53
09
118
.76
59
121
.43
82
122
.94
76
126
.64
55
136
.77
84
Figure S52. 13C NMR spectrum (125 MHz, CDCl3) of compound 20.
156
MPN 29A.100.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
8.0
F1 C
hem
ica
l S
hift
(pp
m)
Figure S53. 1H-1H-COSY spectrum of compound 20.
157
MPN 29.200.001.2rr.esp
11 10 9 8 7 6 5 4 3 2 1 0 -1 -2
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
200
F1 C
hem
ica
l S
hift
(pp
m)
Figure S54. HSQC spectrum of compound 20.
158
MPN 29 A.300.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
F1 C
hem
ica
l S
hift
(pp
m)
Figure S55. HMBC spectrum of compound 20.
159
Compounds 21 and 22
Table S17. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 21, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
21 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
2 135.1 -- H-6; 2H-14 135.5
3 143.5 -- 2H-14 H-5 142.9
7 128.9 -- H-5 128.4
8 121.1 -- H-6; H-12 121.5
13 141.1 -- H-9; H-11 141.2
16 109.5 - H-15; H-17 H-14b 110.8
22 167.9 -- H-17; CO2-
Me
167.3
CH
5 136.1 8.23 (d, 5.4) H-6 136.4
6 112.9 7.98 (d, 5.4) H-5 112.6
9 121.3 8.17 (d, 7.9) H-11 121.2
10 119.4 7.27 (dd, 7.9, 1.1) H-12 119.1
11 128.2 7.57 (m) H-10; H-12 H-9 127.8
12 111.5 7.60 (brd, 8.0) H-10 111.3
15 32.9 3.61 (m) 2H-14 H-17; H-21 34.9
17 152.8 7.55 (d, 0.9) H-21 152.2
19 134.0 5.58 (ddd, 16.5, 10.6, 8.9) H-18b; H-20 134.4
20 44.11 2.62 (m) H-15 2H-14; 2H-
18
44.2
21 96.04 5.75 (d, 6.7) H-20 H-15; H-17;
H-1’
96.4
CH2
14 33.80 3.60 (m); 3.35 (m) H-15 H-20 35.8
18 118.09 5.05 (dd, 10.6, 1.1)
4.94 (brs, 16.5)
H-20 117.9
CH3
CO2-Me 50.2 3.38 (s) 49.9
Glucose
1’ 96.78 4.77 (d, 7.9) H-2’ H-21 99.7
2’ 73.22 3.25(dd, 7.9, 9.2) H-3’ 73.0
3’ 76.56 3.41 (t, 9.2) 76.4
4’ 70.21 3.30 (m) 70.8
5’ 77.15 3.32 (m) 74.4
6’ 61.44 3.93, 3.87 (m) 62.9
*Berger et al. 2015.
160
Table S18. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 22, including
results of HSQC and HMBC 2D experiments. Chemical shifts in ppm and coupling
constants in Hz
22 Literature*
HSQC HMBC C C H 2JCH
3JCH C 2 103.78 - 2H-1 108.7
CH 3 77.00 4.16 (d, 8.1) H-4 2H-1’ 83.2 4 75.90 3.94 (dd, 8.1, 7.2) H-3 78.4 5 81.96 3.77 (dd, 7.2, 3.2) H-4;
2H-6 83.8
CH2 1 60.54 3.67 (d, 11.7)
3.54 (d, 11.7) 61.6
6 63.70 3.74 (d, 9.4) 3.60 (m)
H-4 62.6
1’ 60.78 3.71 (m), 3.51 (m) 2H-2’ 2H-3’ 61.9 2’ 32.05 1.55 (m) 2H-3’ 3H-4’ 33.4 3’ 18.96 1.40 (m) 2H-2’;
3H-4’ 2H-1’ 20.4
CH3 4 12.96 0.95 (t, 7.2) 2H-3 2H-2’ 14.2
*Uddin et al. 2013.
161
FB 70 71.003.001.1r.esp
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
Methanol
0.1
195
0.8
859
0.8
908
0.9
058
0.9
204
0.9
363
0.9
512
0.9
659
1.2
341
1.2
466
1.3
088
1.3
900
1.5
306
1.5
446
1.5
477
1.5
614
1.5
733
3.2
288
3.2
471
3.2
629
3.2
855
3.3
011
3.3
813
3.5
299
3.5
589
3.6
699
3.6
934
3.7
532
3.7
636
4.1
184
4.1
346
4.7
604
4.7
762
5.5
097
5.7
461
5.7
595
7.2
753
7.5
520
7.5
993
7.9
712
7.9
819
8.1
695
8.1
854
8.2
250
8.2
360
Figure S56. 1H NMR spectrum (500 MHz, CD3OD) of compounds 21 and 22.
21
1
NN
CO2CH3
O
OO
HO
OHOH
OH
H
22
1615
2019
1714
32
7
6
58
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
O
OH
HO
OH
OHO
H H
1
2
34
5
6
1'
2'
3'
4'
22
162
FB 70 71.002.001.1r.esp
176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
12.9
22
5
18.9
60
4
32.0
58
0
44.1
12
0
50.2
51
0
60.5
42
86
0.7
84
76
1.4
42
06
3.5
97
8
70.2
13
4
73.2
25
2
75.9
04
67
7.0
02
47
7.1
54
1
81.9
60
5
96.0
40
4
98.7
84
9
103
.78
63
109
.53
17
111
.47
45
112
.93
70
118
.08
65
119
.44
07
121
.07
29
121
.27
52
128
.24
47
133
.96
11
152
.75
72
167
.88
43
Figure S57. 13C NMR spectrum (125 MHz, CD3OD) of compounds 21 and 22.
163
FB 70 71.100.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
1
2
3
4
5
6
7
8
F1 C
hem
ica
l S
hift (p
pm
)
Figure S58. 1H-1H-COSY spectrum of compounds 21 and 22.
164
FB 70 71.200.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
F1 C
hem
ica
l S
hift
(pp
m)
.
Figure S59. HSQC spectrum of compounds 21 and 22.
165
FB 70 71.300.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
F1 C
hem
ica
l S
hift
(pp
m)
Figure S60. HMBC spectrum of compounds 21 and 22.
166
Compound 23
Table S19. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of 23, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
23 Literature*
HSQC HMBC C C H 2JCH
3JCH C 1 41.03 - 2H-2; 3H-12; 3H-13 42,5 3 199.82 - 2H-2 201,3 5 165.87 - 3H-11 H-8 167,4 6 78.61 - H-7 2H-2; H-8;
3H-11; 3H-12; 3H-13
80,0
CH 4 125.78 5.89 (s) 3H-11 127,2 7 130.14 5.87 (m) H-8 H-9 131,8 8 133.88 5.88 (m) H-7; H-9 3H-1-0 135,2 9 75.89 4.45 (m) 3H-10 H-1’; H-7 77,0
CH2 2 49.29 2.52 (d, 16.9
2.17 (d, 16.9) 3H-12; 3H-13 50,9
CH3 10 19.79 1.31 (d, 6.3) 21,2 11 18.16 1.94 (s) H-4 19,8 12 23.29 1.06 (s) 2H-2 24,7 13 22.03 1.05 (s) 2H-2 23,5
Glucose 1’ 101.33 4.36 (d, 7.7) H-2’ 102,7 2’ 73.84 3.19 (dd, 9.9, 7.7) H-1’; H-3’ 75,3 3’ 76.70 3.35 (m) H-2’; H-4’ H-5’ 75,1 4’ 70.24 4,27 (t, 9.6) H-3’; H-5’ H-6’a 71,6 5’ 76.63 3.25 (m) H-6’b 78,0 6’ 61.42 3.86 (dd,13.5, 2.0)
3.64 (dd, 13.5, 5.4) H-5’ 62,7
*Otsuka et al. 1995.
167
FB 77 90 - 15 27.003.001.1r.esp
10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
Methanol
0.1
190
0.9
190
0.9
361
0.9
507
1.0
511
1.0
569
1.1
140
1.3
025
1.3
153
1.3
355
1.3
410
1.5
530
1.9
386
1.9
411
2.1
522
2.1
858
2.2
489
2.5
220
2.5
558
3.1
737
3.1
895
3.1
920
3.2
078
3.2
588
3.2
747
3.3
573
3.3
750
3.6
518
3.6
627
3.7
799
3.8
559
3.8
592
3.8
791
4.0
621
4.3
541
4.3
696
4.4
398
4.4
520
4.4
630
4.6
970
4.7
129
5.2
865
5.3
069
5.3
176
5.3
493
5.5
696
5.5
733
5.8
756
5.8
841
5.8
912
Figure S61. 1H NMR spectrum (500 MHz, CD3OD) of compound 23.
10
O
O
HOO
HO
OHOH
OH
2
651
3
4
11
7
8 9
13
4'
3'
12
1'2'
6'
5'
23
168
FB 77 90 - 15 27.002.001.1r.esp
220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 -20
Chemical Shift (ppm)
19.7
90
92
2.0
33
42
3.2
90
1
24.5
21
5
27.0
38
5
41.0
31
7
42.4
25
6
49.2
94
0
61.4
20
3
70.2
42
37
3.8
42
77
5.8
90
27
6.6
34
1
96.5
89
39
8.3
08
2
101
.33
80
119
.42
99
125
.77
83
130
.13
69
133
.88
17
152
.55
50
165
.87
65
199
.82
13
Figure S62. 13C NMR spectrum (125 MHz, CD3OD) of compound 23.
169
FB 77 90 - 15 27.200.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
F1 C
hem
ica
l S
hift (p
pm
)
Figure S63. HSQC spectrum of compound 23.
170
FB 77 90 - 15 27.300.001.2rr.esp
7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
F2 Chemical Shif t (ppm)
0
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift
(pp
m)
Figure S64. HMBC spectrum of compound 23.
171
Compound 24
Table S20. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of 24, including
result 2D experiments (HSQC and HMBC). Chemical shifts in ppm and coupling
constants in Hz
24 Literature*
HSQC HMBC C C H 2JCH
3JCH C 2 133.37 - H-3 H-6b; 2H-14 134.9 7 108.96 - 2H-6 H-9 110.4 8 127.31 - H-10; H-12 128.8 13 136.37 - H-9; H-11 137.8 16 107.84 - H-17 2H-14; H-20 109.3 22 165.70 - H-3; 2H-5; H-17 167.2 CH 3 53.72 5.09 (d, 4.6) H-14b 2H-5 55.2 9 117.28 7.40 (d, 7.7) H-11 118.8 10 118.78 7.01 (ddd, 7.9, 7.7, 0.9) H-12 120.3 11 121.09 7.10 (ddd, 7.7, 8.1, 1.0) H-9 122.6 12 110.86 7.34 (d, 8.1) H-10 112.4 15 23.54 2.48 (ddd, 14.1, 4.4, 2.3) 2H-14; H-20 H-3; H-17; H-21 25.0 17 147.77 7.39 (d, 2.3) H-21 149.3 19 132.97 5.67 (dt, 17.1, 10.1) 2H-18; H-20 134.4 20 43.36 2.98 (dd, 9.1, 8.8) H-15; H-19 H-14b; 2H-18 44.8 21 96.67 5.43 (d, 1.8) H-1’; H-17 98.2
CH2 5 43.39 4.97 (dd, 12.8, 5.5)
3.12 td, 12.8, 4.6) 44.9
6 20.70 2.99 (t, 5.5) 2.95 (m)
22.2
14 25.93 2.81 (m) 2.06 (td, 13.9, 6.0)
27.4
18 119.13 5.38 (dd, 17.1, 1.6) 5.34 (dd, 10.1, 1.9)
H-20 120.8
Glucose 1’ 99.11 4.59 (d, 7.9) H-2’ H-21 100.6 2’ 72.92 2.71 (m) H-3’ 74.3 3’ 76.56 3.27 (t, 9.1) H-2’; H-4’ 78.2 4’ 69.95 3.20 (t, 9.1) H-5’ H-6’a 71.4 5’ 76.83 3.28 (m) H-6’b 78.0 6’ 61.20 3.87 (dd, 11.8, 2.1)
3.65 (dd, 11.8, 5.2) H-4’ 62.7
*Zhang et al. 2001
172
AR4.003.001.1r.esp
10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0
Chemical Shift (ppm)
Methanol
1.3
089
2.0
555
2.0
671
2.0
833
2.4
686
2.4
753
2.6
895
2.7
105
2.7
206
2.9
635
2.9
812
3.2
054
3.2
237
3.2
588
3.2
768
3.2
957
3.3
680
3.6
432
3.6
554
3.8
647
3.8
684
3.8
882
3.8
922
4.0
051
4.5
881
4.6
040
5.0
958
5.3
328
5.3
533
5.3
722
5.4
064
5.4
286
5.6
575
5.6
770
5.6
916
5.7
118
6.9
9997
.01
55
7.1
006
7.3
392
7.3
553
7.3
947
7.4
115
Figure S65. 1H NMR spectrum (500 MHz, CD3OD) of compound 24.
NN
O
H
O
O
H
H
H
O
HO
OHOH
OH
8
9
10
111
7
14
15
16
17
19
18
1213
21
20
4
5
32
6
2'1'
3'
4'5'
6'
24
173
AR4.002.001.1r.esp
176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
20.7
29
8
23.5
39
3
25.9
33
5
43.3
57
3
43.4
00
6
53.7
28
5
61.1
92
8
69.9
53
4
72.9
14
67
6.5
51
07
6.8
29
1
96.6
68
8
99.1
02
7
107
.83
81
108
.90
33
110
.85
70
117
.27
40
118
.74
73
119
.13
01
121
.08
74
127
.30
58
132
.96
08
133
.36
89
136
.36
98
147
.77
74
165
.71
04
Figure S66. 13C NMR spectrum (125 MHz, CD3OD) of compound 24.
174
AR4.100.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
8.0
F1 C
hem
ica
l S
hift
(ppm
)
Figure S67. 1H-1H-COSY spectrum of compound 24.
175
AR4.200.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
F1 C
hem
ica
l S
hift
(ppm
)
Figure S68. HSQC spectrum of compound 24.
176
AR4.300.001.2rr.esp
7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
170
180
F1 C
hem
ica
l S
hift (p
pm
)
Figure S69. HMBC spectrum of compound 24.
177
Compound 25
Table S21. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of 26, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
25 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
1 161.0 - 1H-3 160.8
4 111.6 - 1H-2 112.1
6 143.3 - 3H-10/1H-8 146.0
7 150.2 - 151.9
9 149.9 - 1H-8 1H-3 151.2
CH
2 113.6 6.24 (d, 9.4) 113.3
3 143.3 7.62 (dd, 9.5, 1.8) 144.7
5 107.7 6.84 (s) 1H-3 109.9
8 103.3 6.87 (s) 103.7
CH2
10 56.5 3.92 (s) 56.5
*Darmawan et al. 2012.
178
CCDS 1011.001.001.1r.esp
9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
-0.1
36
6-0
.03
01
-0.0
25
8-0
.02
34
-0.0
16
7-0
.01
24
0.0
007
0.0
056
0.0
123
0.0
590
0.0
999
0.1
041
0.8
281
0.8
385
0.8
501
0.8
641
0.8
779
0.9
069
1.2
397
1.2
699
1.3
163
1.5
979
2.0
037
2.0
351
2.0
412
2.2
895
2.3
057
2.3
130
2.5
751
3.3
864
3.3
894
3.9
224
3.9
294
5.2
941
6.2
290
6.2
476
6.8
462
6.8
687
7.2
815
7.2
858
7.6
089
7.6
123
7.6
278
7.6
315
Figure S70. 1H NMR spectrum (500 MHz, CDCl3) of compound 25.
19
8
6
7OO
OCH3
OH
54
2
310
25
179
CCDS 1011.002.001.1r.esp
168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
TMS
56
.411
6
10
3.2
15
8
10
7.4
80
5
11
1.6
76
71
13.4
57
0
14
3.2
67
1
14
9.9
40
51
50.5
00
2
Figure S71. 1H NMR spectrum (125 MHz, CDCl3) of compound 25.
180
CCDS 1011.200.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift (p
pm
)
Figure S72. HSQC spectrum of compound 25.
181
CCDS 1011.300.001.2rr.esp
8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5
F2 Chemical Shif t (ppm)
30
40
50
60
70
80
90
100
110
120
130
140
150
160
170
F1 C
hem
ica
l S
hift
(ppm
)
Figure S73. HMBC spectrum of compound 25.
182
Compound 26
Table S22. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data for 26, including
results of HSQC and HMBC experiments. Chemical shifts in ppm and coupling
constants in Hz
26 Literature*
HSQC HMBC C C H 2JCH
3JCH C 4 43.7 - 2H-24 43.2 8 40.8 - 3H-26 40.4 10 37.6 - 3H-25 37.2 13 139.7 - H-18 140.0 14 42.6 - 3H-27 42.1 17 48.8 - H-18 48.4 19 72.5 - H-18 2H-21; 3H-30 54.7 28 180.4 - H-18 180.0 CH 3 80.8 3.61 (m) 2H-24 80.3 5 56.1 0.96 (m) 56.5 9 48.4 1.80 (m) 47.9 12 126.7 5.59 (m) 1H-18 127.9 18 54.4 3.04 (brs) 54.7 20 42.9 1.49 (m) 3H-30 42.4
CH2 1 38.2 2.15 (m); 2.04 (m) 38.8 2 28.2 1.26 (m) 28.5 6 19.7 1.65 (m); 1.35 (m) 19.3 7 34.4 1.58 (m); 1.35 (m) 34.0 11 27.8 1.97 (m) 24.3 15 29.8 1.26 (m) 29.1 16 26.9 1.89 (m) 26.5 21 27.4 1.52 (m); 0.93 (m) 27.0 22 39.3 1.32 (m) 38.6 24 65.1 4.48 (d, 11.0); 3.65 (m) 64.6
CH3 23 23.3 1.53 (s) 2H-24 23.7 25 17.6 0.82 (s) 17.2 26 16.6 1.02 (s) 16.8 27 24.4 1,71 (s) 24.7 29 26.6 1.43 (s) 27.2 30 16.5 1.11 (d, 6.5) 16.1
*Nakatani et al. 1989.
.
183
PNFM5 2327.003.001.1r.esp
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
0.8
433
0.8
634
0.9
967
1.0
559
1.1
133
1.2
048
1.3
003
1.4
294
1.4
388
1.5
300
1.5
416
1.6
756
1.7
079
1.8
269
1.8
495
1.9
483
2.0
350
2.0
609
2.0
789
2.1
289
2.1
579
2.3
016
2.6
067
3.0
421
3.1
223
3.5
891
3.6
425
3.6
645
3.7
768
3.7
871
3.9
879
4.4
581
4.4
645
4.4
864
5.4
704
5.4
777
5.5
906
5.5
973
5.6
717
7.1
929
7.5
609
8.7
041
Figure S74. 1H NMR spectrum (500 MHz, Pyridine-d5) of compound 26.
26
12
3 4
23
56
7
89
10
11
1213
14
15
16
17
18
1920 21
22
HO
CO2HH
HO
OH
30
29
28
184
PNFM5 2327.002.001.1r.esp
184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
15.7
80
51
6.5
38
91
6.8
27
8
17.2
75
6
18.9
87
2
21.1
68
42
3.3
56
7
23.6
09
52
4.0
14
0
24.4
04
0
26.1
37
32
6.6
93
4
26.8
81
2
28.1
66
82
9.0
55
13
0.8
24
63
3.6
41
33
6.8
62
53
8.2
56
43
8.5
16
4
39.1
59
13
9.2
53
0
40.1
12
54
1.8
17
0
42.1
34
7
42.9
00
3
47.6
02
0
48.0
64
3
53.3
00
45
4.3
83
85
6.2
11
0
64.3
36
1
72.4
61
2
80.0
23
0
127
.69
02
139
.70
81
180
.46
36
Figure S75. 13C NMR spectrum (125 MHz, Pyridine-d5) of compound 26.
185
PNFM5 2327.100.001.2rr.esp
12 11 10 9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
1
2
3
4
5
6
7
8
9
10
11
12
F1 C
he
mic
al S
hift (p
pm
)
Figure S76. 1H-1H-COSY spectrum of compound 26.
186
PNFM5 2327.200.001.2rr.esp
9 8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
F1 C
he
mic
al S
hift (p
pm
)
Figure S77. HSQC spectrum of compound 26.
187
PNFM5 2327.300.001.2rr.esp
9 8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
170
F1 C
hem
ica
l S
hift (p
pm
)
Figure S78. HMBC spectrum of compound 26.
188
Compound 27
Table S23. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound
27, including results of HSQC and HMBC experiments. Chemical shifts are given in
ppm and coupling constants in Hz
27 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
2 128.8 - H-3 2H-14 133.2
7 105.7 - H-9 107.7
8 126.0 - 127.9
13 136.8 - H-12 H-9 137.9
16 107.6 - H-17 109.9
22 170.0 - H-17;Me-O-22 170.6
CH
3 51.6 3.81 (s) 52.4
9 117.7 7.48 (d, 7.9) 118.8
10 119.7 7.08 (t, 7.6) 120.1
11 122.1 7.16 (d, 7.7) 122.7
12 110.9 7.34 (d, 8.0) 112.0
15 31.0 3.10 (m) 2H-14 H-17;1H-19 32.5
17 155,5 7.83 (s) 165.1
19 134,0 5.86 (m) H-20 H-21 135.7
20 44.0 2.77 (m) 45.6
21 95.9 5.86 (m) H-17;1H-19 97.5
CH2
5 41.3 3.75 (m); 3.48 (m) 42.9
6 18.1 3.10 (m) 1H-5 21.0
14 33.4 2.37 (m) 35.9
18 118,4 2.25 (m) 119.5
CH3
Me-O-22 51.2 3.82 (s) 52.4
Glucose
1’ 99,0 4.82 (d, 8.2) H-2 H-3 100.3
2’ 77,4 3.39 (m) H-3 78.6
3’ 76,6 3.42 (m) H-4 78.0
4’ 73,2 3.23 (m) 74.6
5’ 70,3 3.25 (m) 2H-6 71.7
6’ 61,6 4.0 (dd, 11.8, 1.9);
3.66 (dd, 11.8, 7.1)
H-5 62.9
*Patthy-Lukáts et al. 1997.
189
AC14.003.001.1r.esp
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
1.0
221
1.1
057
1.3
055
1.3
397
2.0
689
2.2
501
2.2
733
2.3
645
2.3
883
2.7
678
2.7
782
3.1
266
3.1
336
3.2
349
3.2
441
3.2
523
3.2
627
3.3
286
3.4
354
3.4
537
3.6
737
3.8
198
3.9
839
4.0
053
4.6
796
4.7
021
4.8
181
4.8
336
5.2
885
5.3
099
5.3
651
5.3
999
5.8
423
5.8
575
5.8
633
5.8
707
5.8
884
5.8
978
5.9
134
7.0
477
7.0
630
7.1
548
7.1
695
7.3
400
7.3
562
7.4
767
7.4
923
7.8
230
Figure S79. 1H NMR spectrum (500 MHz, CD3OD) of compound 27.
1
NN
CO2CH3
O
OO
HO
OHOH
OH
H
22
1615
2019
1714
32
7
6
5
8
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
27
190
AC14.002.001.1r.esp
184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
18.1
51
5
31.0
07
2
33.4
15
8
41.3
09
8
44.0
10
9
51.2
26
05
1.7
24
3
61.5
82
8
70.3
39
8
73.2
72
17
6.5
83
57
7.3
38
2
95.9
61
0
99.0
01
6
105
.75
80
107
.57
08
110
.91
47
117
.71
82
118
.48
01
119
.21
68
122
.09
85
126
.03
10
128
.77
19
133
.98
64
136
.80
31
155
.46
19
169
.84
88
Figure S80. 13C NMR spectrum (125 MHz, CD3OD) of compound 27.
191
AC14.100.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
1
2
3
4
5
6
7
8
F1 C
he
mic
al S
hift (p
pm
)
Figure S81. 1H-1H-COSY spectrum of compound 27.
192
AC14.200.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
F1 C
hem
ica
l S
hift
(ppm
)
Figure S82. HSQC spectrum of compound 27.
193
AC14.300.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
180
F1 C
he
mic
al S
hift (p
pm
)
Figure S83. HMBC spectrum of compound 27.
194
Compound 28
Table S24. 1H NMR (500 MHz) and 13C NMR (125 MHz) spectral data of compound
28, including results of HSQC and HMBC experiments. Chemical shifts are given in
ppm and coupling constants in Hz
28 Literature*
HSQC HMBC
C C H 2JCH 3JCH C
2 129.0 - H-3 2H-6 133.2
7 106.9 - 2H-6 HN-1; H-9 109.0
8 126.1 - HN-1; H-10; H-12 128.0
13 137.1 - H-9; H-11 138.4
16 107.2 - H-17 2H-14 109.9
22 170.4 - H-17; MeO-22 170.9
23 172.2 - H-5 176.5
CH
3 51.7 4.62 (d, 11.7) 2H-14 H-5 53.2
5 58.3 3.94 (dd, 12.0, 5.0) 2H-6 60.1
9 117.7 7.48 (d, 7.0) H-11 118.8
10 119.2 7.05 (t, 7.0) H-12 120.1
11 122.0 7.14 (t, 7.0) H-9 122.6
12 110.9 7.33 (d, 7.0) H-11 H-10 112.1
15 31.6 3.11 (m) 2H-14; H-
20
H-17; H-19; H-21 32.4
17 155.8 7.84 (s) H-21 156.1
19 133.8 5.87 (ddd, 17.4, 10.7, 7.5) H-18; H-20 H-21 135.2
20 43.8 2.79 (m) H-19; H-21 H-18 45.7
21 96.0 5.94 (d, 9.2) H-20 H-15; H-17; H-1’ 97.6
CH2
6 22.7 3.45 (m), 3.05 (m) H-5 25.2
14 33.3 2.46 (m), 2.25 (m) H-3 35.6
18 118.4 5.39 (d, 17.7);
5.27 (d, 10.7)
H-20 119.6
CH3
Me-O-22 51.4 3.85 (s)
Glucose
1’ 99.0 4.85 (d, 8.0) H-2’ H-21 100.5
2’ 73.2 3.27 (dd, 9.1, 8.0) H-3’ 74.7
3’ 77.4 3.43 (t, 9.1) H-2’; H-4’ 78.0
4’ 70.4 3.28 (t, 9.1) H-3’; H-5’ 71.9
5’ 76.5 3.38 (m) H-6’b 78.6
6’ 61.7 4.04 (dd, 11.8, 1.8)
3.70 (dd, 11.8, 7.0)
H-5’ H-4’ 63.1
Ferrari et al. 1986.
195
AC 11.003.001.1r.esp
9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5
Chemical Shift (ppm)
Methanol
1.2
6701.3
043
2.2
501
2.4
578
2.7
821
2.7
910
3.0
445
3.0
820
3.2
560
3.2
743
3.2
804
3.2
993
3.3
878
3.4
357
3.4
537
3.4
723
3.7
057
3.8
167
3.8
790
4.0
275
4.0
312
4.0
510
4.0
547
5.2
675
5.2
888
5.3
776
5.4
124
5.8
465
5.8
529
5.8
673
5.9
307
5.9
490
7.0
3857
.05
32
7.1
429
7.1
578
7.3
256
7.3
421
7.4
736
7.4
892
7.8
379
Figure S84. 1H NMR spectrum (500 MHz, CD3OD) of compound 28.
1
NN
CO2CH3
O
OO
HO
OHOH
OH
H
CO2H
22
1615
2019
1714
32
7
6
5
8
9
10
11
1213
18
21
2'1'
3'
4'5'
6'
28
196
AC 11.002.001.1r.esp
176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0
Chemical Shift (ppm)
22.6
36
5
31.2
52
7
33.2
28
0
43.9
17
0
51.7
09
9
58.0
94
4
61.7
05
6
70.3
76
0
73.2
68
57
6.5
76
37
7.4
10
5
96.0
22
4
99.0
70
2
106
.93
89
107
.22
78
110
.88
95
117
.79
76
118
.45
48
119
.22
40
122
.01
91
126
.08
88
129
.04
64
133
.80
22
137
.11
37
155
.87
36
170
.42
66
172
.20
32
Figure S85. 13C NMR spectrum (125 MHz, CD3OD) of compound 28.
197
AC 11.100.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
8.0
F1 C
hem
ica
l S
hift (p
pm
)
Figure S86. 1H-1H-COSY spectrum of compound 28.
198
AC 11.200.001.2rr.esp
8 7 6 5 4 3 2 1 0
F2 Chemical Shif t (ppm)
0
10
20
30
40
50
60
70
80
90
100
110
120
130
140
150
160
F1 C
hem
ica
l S
hift (p
pm
)
Figure S87. HSQC spectrum of compound 28.
199
AC 11.300.001.2rr.esp
8 7 6 5 4 3 2 1
F2 Chemical Shif t (ppm)
20
40
60
80
100
120
140
160
180
F1 C
hem
ica
l S
hift
(pp
m)
Figure S88. HMBC spectrum of compound 28.
200
MTT assay
Figure S89. Evaluation of cell viability of U937 leukemic cell lines by MTT assay (n =
3), after 48 h of incubation with compounds 20, 23, and 24. The concentration 0 is the
negative control test (cells and culture medium). The concentration of DMSO was 0.2
%.
Figure S90. Evaluation of cell viability of THP-1 leukemic cell lines by MTT assay (n =
3), after 48 h of incubation with compounds 20, 23, and 24. The concentration 0 is the
negative control test (cells and culture medium). The concentration of DMSO was 0.2
%.
23
24
20
23
24
20
201
4. CONCLUSÕES
Com o desenvolvimento desta pesquisa foi possível, além de compreender a
química do gênero Psychotria, contribuir com mais informações que poderão ser úteis
no ponto de vista quimiotaxonômico. Neste trabalho, descrevemos os dados
espectrais de um novo iridoide (ácido 9-epi-geniposídico), além de outros dez
metabólitos isolados de P. suterella. Da espécie P. nuda outros dezessete metabólitos
foram isolados. Através de uma busca na literatura foi possível constatar que, além
do composto inédito, os ácidos geniposídico, 3-O-acetiloleanólico, pomólico,
espinólico, maslínico, tormêntico e lyalosídico, metil oleleanato, cinchonainas Ia e Ib,
roseosídeo e o alcaloide raro N,N,N-trimetiltriptamônio estão sendo, provavelmente,
relatados pela primeira vez no gênero.
Além do estudo químico das espécies mencionadas, este estudo também
consistiu na avaliação de atividades inseticida, antifúngica e citotóxica de extratos,
frações e compostos isolados. As amostras testadas não promoveram, de forma
considerável, a morte das larvas do mosquito A. aegypti, que é o vetor de doenças
como a dengue e febre amarela. Os extratos e frações testadas nos ensaios para
avaliação de atividade antifúngica não inibiram, consideravelmente, o crescimento
dos fungos fitopatogênicos Fusarium oxysporum, Curvularia lunata, Colletotrichum
musae, Rhizoctonia solani, and Sclerotium rolfsii, que causam prejuízos ao
agronegócio. Em relação ao ensaio de atividade citotóxica, o alcaloide estrictosamida
apresentou os melhores resultados frente às duas linhagens de células cancerígenas
THP-1 e U937, com valores de EC50 de 120 ± 1 e 21.9 ± 1 g/mL, respectivamente.
Este resultado aponta para a possibilidade de testar outros alcaloides e outras
linhagens de células cancerígenas em pesquisas futuras.
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6. ANEXOS
6.1 Metodologia do Ensaio para Avaliação de Atividade Inseticida
Os extratos e frações das espécies P. nuda e P. suterella, obtidos como
mencionados na seção 3.3.1.1 (páginas 82-83), foram solubilizados em DMSO / H2O
ou DMSO puro. Quinze larvas de terceiro instar (Aedes aegypti) foram adicionadas
aos potes contendo água destilada e adicionadas às soluções de teste à temperatura
ambiente, 27 ° C. Os testes foram realizados em triplicado e em duas repetições. O
controle negativo foi a água pura, DMSO puro e uma solução DMSO / H2O (2,5%).
Para o controle positivo foi utilizado o composto Imidacloprid, com concentrações
entre 0,01 μg / mL e 1,0 μg / mL. A avaliação da mortalidade foi feita 24 horas após a
exposição das larvas às soluções. Todas as amostras testadas não apresentaram
atividade larvicida numa escala considerável.
6.2 Metodologia do Ensaio para Avaliação de Atividade Antifúngica
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Os bioensaios foram conduzidos por adição ao meio de cultura PDA (Sigma-
Aldrich®), às soluções aquosas de extratos e frações das duas plantas testadas,
utilizando-se volumes apropriados para obter uma concentração de 3500 μg mL-1,
igualmente para cada microrganismo. Com o meio já vertido em placas de Petri, foram
inoculados discos de micélio-ágar de 5 mm de diâmetro feitos de culturas puras de
fungos na superfície de cada cultura com seus respectivos tratamentos.
Posteriormente, as placas foram seladas com película de plástico e incubadas em
uma câmara de crescimento (25 ± 1 ° C) e expostas a um fotoperíodo de 12 horas.
Para estimar a eficiência dos tratamentos, o diâmetro de cada uma das colônias foi
medido durante o crescimento micelial e comparado ao controle. O crescimento radial
foi medido com paquímetro, em dois eixos ortogonais um ao outro calculando uma
média para cada placa. As amostras testadas não apresentaram resultados
satisfatórios.