Axially Chiral Dimeric Naphthalene and Naphthoquinone Metabolites, from Root Cultures of the West African Liana Triphyophyllum peltatum

Article abstract of DOI:10.1021/acs.jnatprod.6b00439

Root cultures of the West African liana Triphyophyllum peltatum were initiated from stem explants of in vitro cultivated shoots. From these organ cultures, three new binaphthalenes, one binaphthoquinone, and two (bi)naphthalene glucosides were isolated, with substitution patterns related to those of the naphthylisoquinoline alkaloids, which are the normal main metabolites of T. peltatum. The structures of the diglucoside dioncoquinoside A (1) and of the axially chiral biaryls triphyoquinols A₁ (3), A₂ (4), and B (5), triphyoquinoside A (6), and triphyoquinone A (7) were elucidated by spectroscopic analysis (HRESIMS, 1D and 2D NMR) and by application of electronic circular dichroism (ECD) spectroscopy in combination with the exciton chirality method and quantum-chemical ECD calculations. The root cultures likewise produced the known alkaloids dioncophylline A (8), 5′-O-demethyldioncophylline A (9), dioncopeltine A (10), habropetaline A (11), and 5′-O-methyldioncophylline D (12a/b), the naphthalene glucoside plumbaside A (2), and the naphthoquinones plumbagin (13), droserone (14), and 8-hydroxydroserone (15).

Full text of DOI:10.1021/acs.jnatprod.6b00439

Article
pubs.acs.org/jnp
Axially Chiral Dimeric Naphthalene and Naphthoquinone
Metabolites, from Root Cultures of the West African Liana
Triphyophyllum peltatum
Gerhard Bringmann,*,† Andreas Irmer,† Tobias Buttner,† Anu Schaumloffel,† Guoliang Zhang,†
̈
̈
Raina Seupel,† Doris Feineis,† and Karin Fester‡
†Institute of Organic Chemistry, University of Wurzburg, Am Hubland, D-97074 Wurzburg, Germany
̈
̈
‡Institute of Pharmacy, Research Group Pharmaceutical Biology, University of Leipzig, Johannisallee 21-23, D-04103 Leipzig,
Germany
S
* Supporting Information
ABSTRACT: Root cultures of the West African liana
Triphyophyllum peltatum were initiated from stem explants of
in vitro cultivated shoots. From these organ cultures, three new
binaphthalenes, one binaphthoquinone, and two (bi)-
naphthalene glucosides were isolated, with substitution patterns
related to those of the naphthylisoquinoline alkaloids, which are
the “normal” main metabolites of T. peltatum. The structures of
the diglucoside dioncoquinoside A (1) and of the axially chiral
biaryls triphyoquinols A1 (3), A2 (4), and B (5), triphyoquino-
side A (6), and triphyoquinone A (7) were elucidated by
spectroscopic analysis (HRESIMS, 1D and 2D NMR) and by
application of electronic circular dichroism (ECD) spectroscopy
in combination with the exciton chirality method and quantum-
chemical ECD calculations. The root cultures likewise produced the known alkaloids dioncophylline A (8), 5′-O-
demethyldioncophylline A (9), dioncopeltine A (10), habropetaline A (11), and 5′-O-methyldioncophylline D (12a/b), the
naphthalene glucoside plumbaside A (2), and the naphthoquinones plumbagin (13), droserone (14), and 8-hydroxydroserone
(15).
In this paper, we describe the isolation and structural
uring the past years, axially chiral binaphthalenes have
Dattracted great attention as potent ligands in stereo- elucidation of seven natural products with naphthalene basic
selective total synthesis (e.g., BINAP, BINOL, or BINAM),1 in
structures from the root cultures of the West African liana
Triphyophyllum peltatum, among them new axially chiral
binaphthalenes and binaphthoquinones as well as related
(bi)naphthalene glucosides. Due to their occurrence in T.
peltatum (Dioncophyllaceae), they were named dioncoquino-
side A (1), triphyoquinols A1 (3), A2 (4), and B (5),
triphyoquinone A (7), and triphyoquinoside A (6). Only
plumbaside A (2) had already been known from previous
isolation work on various plant genera (Figure 1).19−24 These
mono- and dimeric naphthalenes were not identified in the
plant itself, which is, rather, a rich source of naphthylisoquino-
line alkaloids.25 Some of the isolated alkaloids, e.g., dioncophyl-
line A (8),25,26 dioncopeltine A (10),25,27 and 5′-O-methyl-
dioncophylline D (12a/b)28 (Figure 2), had previously also
been detected in the root cultures of T. peltatum.
Naphthylisoquinolines are exclusively produced by Dionco-
phyllaceae and Ancistrocladaceae plants, which are indigenous
to tropical rain forests in West, Central, and East Africa and
particular for transition-metal-catalyzed cross-coupling reac-
tions,2 or as building blocks for the construction of chiral
supramolecular and polymeric materials.3 In nature, however,
binaphthalenes or related glucosides have only rarely been
discovered, e.g., in plants belonging to the Ebenaceae
(Diospyros species4−10) or the Liliaceae (Hemerocallis,4,11
Dianella,4,12,13 or Stypandra4,13,14 taxa) and in the mold fungus
Chrysosporium meridarium.4,15 Dimeric naphthoquinones and
“mixed dimers” consisting of a naphthoquinone and a
naphthalene portion were isolated from several Diospyros
species.4,5,7,16,17 Some of the Ebenaceae and Liliaceae plants
are used in folk medicine for the treatment of parasitic diseases
(e.g., schistosomiasis), but their application is accompanied by
severe toxic side effects.4−6,10,11,13,15,16 Surprisingly, in contrast
to the many investigations on binaphthalenes of synthetic
origin, in the reports on naturally occurring binaphthalenes,5−14
the phenomenon of axial chirality was taken into consideration
only for some of these biaryls;18 for others of those compounds,
no ECD investigations were performed, and frequently not
even optical rotation values were measured.4
Received: May 13, 2016
© XXXX American Chemical Society and
American Society of Pharmacognosy
A
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
Figure 1. Mono- and dimeric naphthalene derivatives (1−7) from root cultures of T. peltatum.30
RESULTS AND DISCUSSION
■
Root cultures of T. peltatum were initiated from stem explants
of in vitro cultivated shoots31 on a modified Gamborg B5
(G5S2) medium (Supporting Information).32 Adventitious
roots were cut and transferred to medium of the same
composition, but without gelling agent. The roots were kept in
a submerged state with aeration brought about by continuous
shaking. The shake cultures showed a marked acceleration in
growth as compared to the corresponding stationary cultures,
the maximum dry weight of the root material being attained
after about 4 weeks. The roots were then removed from the
medium, lyophilized, ground, and exhaustively extracted with
CH2Cl2/MeOH (1:1). The resulting extracts were filtered and
subjected to cation-exchange chromatography using the resin
Amberlyst 15. The fractions thus obtained were finally purified
by chromatography on a semipreparative RP-18 HPLC column.
LC hyphenation with UV and ESIMS/MS analysis permitted
rapid dereplication of five known naphthylisoquinoline
alkaloids, viz., dioncophylline A (8),26 5′-O-demethyldionco-
phylline A (9),33 dioncopeltine A (10),27 habropetaline A
(11),34 and 5′-O-methyldioncophylline D (12a/b).28 The latter
had recently been identified as the first 7,8′-coupled (D-type)
naphthylisoquinoline alkaloid from a Dioncophyllaceae plant.28
Moreover, similar to the callus cultures,35,36 the root cultures
were found to also produce the naphthoquinones plumbagin
(13), droserone (14), and 8-hydroxydroserone (15). The
identity of metabolites 8−15 was confirmed by coelution with
authentic reference compounds;26−28,33−36 in the case of 5′-O-
methyldioncophylline D (12), an ultimate structural proof was
achieved by preparative isolation of this alkaloid, which is a 1:1
mixture of its two slowly interconverting atropo-diastereomers,
12a and 12b.28
Figure 2. Naphthylisoquinoline alkaloids (8−12) and naphthoqui-
nones (13−15) identified in the root cultures of T. peltatum.
Southeast Asia.25 In the plants themselves, metabolites
structurally related to the naphthalene moieties of the
naphthylisoquinolines, e.g., ancistronaphthoic acid B,29 had so
far been identified quite rarely; only three representatives had
as yet been discovered in two Congolese Ancistrocladus lianas.29
Far more frequently, naphthalene-derived tetralones such as cis-
isoshinanolone or naphthoquinones such as plumbagin (13)
were found to be present in these tropical lianas, and even
naphthoquinone dimers such as 3,3′-biplumbagin were
isolated.25 Here, we describe the elucidation of the metabolic
pattern of root cultures of T. peltatum, with particular emphasis
on the chiroptical analysis of the axially chiral constituents
triphyoquinol A1 (3), triphyoquinone A (7), and triphyoquino-
side A (6).
The most polar substance among the metabolites of the root
cultures of T. peltatum exhibiting a UV spectrum typical of
naphthalenes was found to possess a molecular formula of
C23H30O13 (m/z 537.1579, [M + Na]+) as evidenced by
HRESIMS. 1H and 13C NMR investigations indicated the
presence of a 1,4,5-trioxy-substituted 2-methylnaphthalene
derivative: Its 1H NMR spectrum (Experimental Section)
revealed resonances corresponding to a methyl group (δ 2.42)
and four aromatic protons (δ 6.88, 7.12, 7.41, and 7.82), among
them the signals (two doublets, one multiplet) of three
B
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
contiguous hydrogens. The 13C NMR spectrum (Experimental
Section) showed three low-field-shifted quaternary aromatic
carbons resonating at 147.2, 152.6, and 154.8 ppm, suggesting
the presence of three oxygen functions, two of them being
located at C-1 and C-4 in the “eastern” ring system. The third
one, a hydroxy function, was deduced to be located at C-5,
which was in agreement with HMBC couplings to C-5 (154.8
ppm) observed from the proton signals of both H-6 and H-7.
That the oxygen functions at C-1 and C-4 were each linked to a
glucopyranosyl residue became evident from the characteristic
Similar to dioncoquinoside A (1), plumbaside A (2) was also
found to undergo a spontaneous oxidation reaction, giving rise
to plumbagin (13) after cleavage of the glucopyranosyl residue.
In the case of 2, this reaction was observed to proceed readily,
even without enzymatic assistance.
The two naphthalene glucosides, dioncoquinoside A (1) and
plumbaside (2), presumably serve as storage products in the
roots (like pro-drugs), releasing the toxic plumbagin (13) as a
phytoalexin for protection of the plants, e.g., when attacked by
herbivores.
Further isolation work provided three new naphthalene
dimers with substitution patterns similar to those of the
isocyclic parts of napththylisoquinoline alkaloids. They were
named triphyoquinols A1 (3), A2 (4), and B (5).
1
1
signals in the H and 13C NMR spectra, from H,1H−COSY
and HMBC interactions, and in particular from ROESY
correlations between 2-Me and H-1′ and between H-3 and
H-1″ (Figure 3). The β-configurations at the anomeric C-1′
Triphyoquinol A1 (3) (Figure 4) had a molecular formula of
C24H23O4 (m/z 375.1591, [M + H]+), as deduced from
1
Figure 3. Key HMBC (single arrows) and H,1H-COSY and ROESY
(double arrows) interactions of dioncoquinoside A (1).
and C-1″ centers (and hence also the attachment to the
aglycone) were assigned by the large coupling constants (3J =
7.9 Hz) from H-1′ to H-2′ and from H-1″ to H-2″. Thus, the
structure of the new compound was assigned as 1,4-di-β-D-
glucopyranosyloxy-5-hydroxy-2-methylnaphthalene (1) and
named dioncoquinoside A (Figure 3).
Treatment of 1 with a commercial β-glucosidase at room
temperature or with methanolic HCl at 60 °C led to its
complete deglucosidation, providing D-glucose (identical to an
authentic sample by cochromatography on TLC, 1H NMR, and
[α]D), along with the respective naphthalenetriol, which
underwent immediate spontaneous oxidation to give the
naphthoquinone plumbagin (13), identical in all respects with
a reference sample of 13.
Figure 4. Triphyoquinol A1 (3): (A) Selected 1H and 13C NMR shifts
(methanol-d4, δ values in ppm), (B) key HMBC (single arrows) and
1H,1H-COSY and NOESY (double arrows) interactions, and (C)
assignment of the absolute configuration at the biaryl axis by ECD
spectroscopy in combination with the exciton chirality method38
(double arrows mark relevant electronic transition dipole moments;
the red arrow shows the clockwise array responsible for the positive
exciton couplet).
A second, likewise polar and water-soluble naphthalene
glucoside isolated from the root cultures showed a behavior
similar to that of dioncoquinoside A (1), chromatographically
1
and also spectroscopically, according to its H and 13C NMR
HRESIMS and from the number of signals in the 13C NMR
spectrum (Table 1). Similar to the results of the monomeric
dioncoquinoside A (1), the binaphthalene 3 displayed 1H NMR
signals (Table 1) for eight aromatic protons, viz., two sets of
three contiguous hydrogens and two one-proton singlets,
together with signals for two methoxy and two methyl groups.
Furthermore, the appearance of two signals at 153.6 and 154.5
ppm in the 13C NMR spectrum hinted at the additional
presence of two phenolic functions (Figure 4A). Specific
NOESY correlations from H-8 to Me-2′ and from H-8′ to Me-2
data. With a molecular formula of C17H20O8 (m/z 375.1050 [M
+ Na]+), as determined by HRESIMS, its structure differed
from that of 1 by the absence of one of the glucopyranosyl
residues. This compound, called plumbaside A (2) (Figure
1),19,20,24 is a well-known metabolite, previously isolated from
various plant genera such as Plumbago,19,20 Nepenthes,21
Drosophyllum,22 Drosera,23 and Dionaea,24 among them some
carnivorous species, which are the closest phylogenetic
neighbors of the likewise carnivorous Dioncophyllaceae.31,37
C
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
1
Table 1. H (600 MHz) and 13C (150 MHz) NMR Data of 3−5 and 7 in Methanol-d4 (δ in ppm, J in Hz)
triphyoquinol A1 (3)
δH (J in Hz) δC, type
triphyoquinol A2 (4)
δH (J in Hz) δC, type
triphyoquinol B (5)
δH (J in Hz) δC, type
7.26, s
triphyoquinone A (7)
no.
δH (J in Hz)
δC, type
1
128.9, C
129.9, C
136.1, C
107.9, CH
156.9, C
156.0, C
110.4, CH
128.6, CH
117.7, CH
137.4, C
115.0, C
120.3, CH
139.2, C
121.5, C
152.0, C
157.5, C
104.5, CH
126.7, CH
122.6, CH
137.5, C
115.0, C
123.7, C
138.3, C
113.4, CH
156.1, C
158.1, C
105.2, CH
128.3, CH
119.6, CH
137.1, C
114.9, C
127.9, C
135.8, C
123.2, CH
153.4, C
183.3, C
182.1, C
128.4, CH
144.0, CH
134.8, C
115.1, C
20.5, CH3
21.1, CH3
2
134.8, C
3
6.86, s
106.4, CH
155.4, C
6.96, s
6.84, s
4
5
153.6, C
6
6.58, dd (7.6, 1.1)
6.93, dd (7.7, 8.9)
6.31, dd (8.4, 1.1)
108.9, CH
127.1, CH
116.3, CH
136.2, C
6.69, dd (7.6, 1.0)
7.03, dd (7.6, 8.4)
6.41, dd (8.5, 1.1)
6.73, d (7.6)
6.93, d (7.5)
7
7.29, dd (7.6, 8.4)
7.39, d (8.4)
7.26, dd (7.5, 8.3)
6.73, d (8.4)
8
9
10
1′
2′
3′
4′
5′
6′
7′
8′
9′
113.5, C
126.0, C
136.3, C
6.73, s
112.0, CH
154.5, C
7.12, s
156.5, C
6.74, d (7.8)
6.97, s
103.1, CH
125.7, CH
118.9, CH
135.7, C
6.16, d (10.3)
6.80, d (10.4)
6.45, dd (8.5, 0.9)
10′
2-CH3
113.3, C
1.85, s
1.79, s
4.05, s
18.9, CH3
18.7, CH3
55.2, CH3
1.95, s
4.15, s
20.3, CH3
56.7, CH3
2.16, s
4.00, s
21.7, CH3
57.3, CH3
2.06, s
1.86, s
2′-CH3
4-OCH3
5-OCH3
5′-OCH3
4.10, s
56.9, CH3
3.99, s
55.3, CH3
indicated that the biaryl axis was located at C-1 in both
naphthalene moieties. The position of the methoxy group at C-
4 was deduced from its NOESY interaction with H-3, while the
other methoxy function was established to be at C-5′ by a
NOESY correlation with H-6′ (Figure 4B). According to these
findings, triphyoquinol A1 was assigned to possess the
constitution of the unsymmetric binaphthalene 3, as presented
in Figure 4A,B.
(Supporting Information). As in the case of 3, the ECD
spectrum of 4 displayed a strong positive couplet, with
sequential positive and negative Cotton effects at 234 (Δε,
+56.6) and 218 nm (Δε, −44.6); thus, according to the exciton
chirality method38 its biaryl axis was P-configured, like that of 3.
1H and 13C NMR investigations (Table 1) in combination
with HRESIMS measurements suggested the third binaph-
thalene, triphyoquinol B (5) (Figures 1 and 5), again to be C2-
With its high steric hindrance at the central biaryl axis, 3
should show the phenomenon of axial chirality. HPLC analysis
of 3 on a chiral phase (Phenomenex Lux Cellulose-1) gave only
one peak, suggesting a high enantiomeric purity of the
compound. This 1,1′-linked30 binaphthalene 3 should be a
classic case of a molecule where the exciton chirality method38
is applicable due to the fact that it contains two electronically
very similar, nearly identical chromophores. The ECD
spectrum of 3 exhibited sequential positive and negative
Cotton effects at 235 and 219 nm, i.e., a positive couplet
(Figure 4C, left). Application of the exciton chirality method
led to the conclusion that the electronic transition dipole
moments defined a “positive chirality”, and, thus, the absolute
axial configuration of triphyoquinol A1 (3) was attributed to be
P (Figure 4C, right).
Figure 5. Selected NMR data of triphyoquinol B (5): (A) 1H and 13
C
NMR shifts (methanol-d4, δ values in ppm) and (B) HMBC (single
1
arrows) and H,1H-COSY and NOESY (double arrows) interactions.
The second dimer, triphyoquinol A2 (4) (Figure 1), was
symmetric, like 4. Its constitution was evidenced from a series
of NOESY correlations {Me-2 ↔ H-1 ↔ H-8 ↔ H-7 ↔ H-6
↔ 5-OMe ↔ 4-OH} and from an NOE interaction between 4-
OH and 2′-Me and the low-field shift of C-3 (120 ppm). The
latter suggested the biaryl linkage30 to be located between C-3
and C-3′ of the constitutionally identical naphthalene halves.
The ECD spectrum of this binaphthalene dimer 5 displayed a
negative first Cotton effect at 242 nm (Δε, −25.1) and a
second positive one at 224 nm (Δε, +39.0), i.e., a negative
couplet; thus, in contrast to the P-configured dimers 3 and 4,
1
found to be C2-symmetric. From H and 13C NMR measure-
ments (Table 1) it became evident that this binaphthalene was
a regioisomer of triphyoquinol A1 (3), consisting of two
structurally identical naphthalene portions, which are con-
nected via a 1,1′-linkage30 as deduced from the low-field-shifted
quaternary aromatic carbons (C-1/C-1′) resonating at 129.9
ppm. The positions of the methoxy groups were established
from its HMBC correlations with C-4 (and thus also with C-4′)
and from the specific NOESY interactions with H-3 (and H-3′)
D
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
the absolute configuration at the biaryl axis of triphyoquinol B
(5) was assigned to be M.
structurally closely related 1,1′-coupled triphyoquinols A1 (3)
and A2 (4).
A further new metabolite, triphyoquinone A (7) (Figure 6),
exhibited a UV profile similar to those of the three
Although triphyoquinone A (7) thus consisted of two
structurally and electronically different molecular portions, the
exciton chirality method was still applicable to the assignment
of the absolute configuration at the biaryl axis provided that the
dihedral angle at the configurationally stable axis was known
either from an X-ray structure analysis or from quantum-
chemical calculations. A conformational analysis of 7 (B3LYP/
TZVP), arbitrarily for the P-atropisomer, indicated a dihedral
angle of 92° between the planes of the two chromophores. In
addition, the electronic transition dipole moments of the
naphthalene and the ortho-quinone portions were calculated,
showing that the relevant transitions for the exciton coupling
were polarized along the long axes of the chromophores.
According to the exciton chirality method, the coupling of the
electronic transition dipole moments led to a positive first and
negative second Cotton effect and, thus, to a positive ECD
couplet (Figure 7A). Since the experimental ECD spectrum of
triphyoquinone A (7) showed a positive couplet, 7 was
assigned to have the absolute P-configuration. This result was
confirmed by the unambiguous match of the experimental ECD
curve of triphyoquinone A with the spectrum calculated for the
P-atropisomer of 7. As expected, the curve predicted for the M-
configured atropisomer of 7 was opposite compared to the
experimental ECD spectrum of 7, thus giving rise to mirror-
image-like curves (see Figure 7B).
Figure 6. (A) Selected 1H and 13C NMR shifts (methanol-d4, δ values
in ppm) and (B) HMBC (single arrows) and 1H,1H-COSY and
NOESY (double arrows) interactions of the “southern”, quinoid
molecular moiety of triphyoquinone A (7).
binaphthalenes described above, but with an additional
maximum at 440 nm, hinting at the presence of a quinoid
compound. This assumption was confirmed by 13C NMR
measurements exhibiting two downfield-shifted signals at 182.1
and 183.3 ppm typical of carbonyl functions (Figure 6A). The
two quinoid carbonyl groups were located ortho to each other,
at C-5′ and C-6′, as established by HMBC interactions between
H-7′ and H-8′ and by NOE correlations between H-8′ and Me-
2 (Figure 6B), likewise evidencing this binaphthoquinone 7 to
possess a 1,1′-biaryl linkage. All other NMR data of this new
dimer were similar to those reported above for the two
Finally, triphyoquinoside A (6), a structurally more complex
binaphthalene, possessing a molecular formula of C40H48O19
(m/z 855.2699 [M + Na]+), was isolated from the root cultures
of T. peltatum. The 1D and 2D NMR spectra exhibited signals
Figure 7. Assignment of the absolute axial configuration of triphyoquinone A (7) (A) by applying the exciton chirality method (double arrows mark
the relevant electronic transition dipole moments; the red arrow shows the clockwise array responsible for the positive exciton couplet) and (B) by
comparison of the experimental ECD curve of isolated 7 with the ECD spectra of P-7 and M-7 predicted by ECD calculations with TD-B3LYP/6-
31G* based on the B3LYP/TZVP optimizations.
E
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
Figure 8. HMBC (single green arrows), 1H,1H−COSY/ROESY (double red arrows), and TOCSY (double blue arrows) interactions relevant for the
constitution of triphyoquinoside A (6): (A) the binaphthalene core and (B) the “southern” and (C) “northern” glucopyranosyl residues; (D)
assignment of the absolute axial configuration by comparison of the experimental ECD spectrum of 6 with that calculated for P using ZINDO/S-Cl
(PM3 geometries).
typical of a 1,1′-coupled naphthalene dimer suggesting the
presence of three glucopyranosyl residues (Figures 1 and 8).
From HMBC and ROESY correlations (Figure 8A−C) it
became evident that the two oxygen groups at C-4 and C-5′ of
the binaphthalene core were attached to sugar moieties, each
with a β-configuration at the anomeric sugar carbon atoms, C-
1″ and C-1‴, as deduced from the large coupling constants (3J
= 7.8 and 7.9 Hz), from H-1″ to H-2″ and from H-1‴ to H-2‴,
respectively. Furthermore, the 1H NMR spectrum showed
signals suggesting the presence of a third anomeric proton,
which displayed no interactions with the binaphthalene core
according to HMBC and ROESY investigations, but showed
strong correlations with a CH2 group of one of the
glucopyranosyl residues. Complete attribution of the sugar
signals was finally achieved by TOCSY measurements, which
permitted clear assignment of geminal, vicinal, and long-range
couplings within one spin system, here, for example, within a
glucopyranosyl subunit (Figure 8B,C). Treatment of 6 with
methanolic HCl at 60 °C afforded D-glucose (identical to an
authentic sample by cochromatography on TLC, 1H NMR, and
[α]D), as the only carbohydrate, while the air-sensitive aglycone
got decomposed immediately.
The experimental ECD spectrum of 6 showed a positive first
and a negative second Cotton effect. As expected for these
types of structures, the ECD behavior of 6 was dominated by
the exciton coupling of the long-axis-polarized transitions of the
naphthalene chromophores.38 Semiempirical calculations on
the P-atropo-diastereomer of 6 predicted a dihedral angle of
92° at the biaryl axis. Within this geometry, the naphthalene
moieties constituted a clockwise array. According to the exciton
chirality method, the experimentally observed positive ECD
couplet around 235 nm thus indicated a “positive chirality” in
triphyoquinoside A (6), here corresponding to the P-
configuration. ZINDO/S-Cl calculations of the ECD spectrum
of the PM3 minimum of 6 corroborated the assignment of the
absolute configuration at the biaryl axis of 6 to be P (Figure
8D).
thalenes, the triphyoquinols A1 (3), A2 (4), and B (5), along
with the binaphthoquinone triphyoquinone A (7), and the
glucosides dioncoquinoside A (1) and triphyoquinoside A (6)
(Figure 1). The substitution patterns of these compounds
resemble those of the isocyclic parts of the biosynthetically
related naphthylisoquinoline alkaloids. The mono- and dimeric
naphthalenes were not identified in the plant itself andexcept
for triphyoquinol B (5)not even in callus cultures of T.
peltatum. On the other hand, some highly oxygenated
naphthoquinones, the dioncoquinones A−E,35 were detected
exclusively in solidified callus cultures, but not in the root
cultures, thus showing that the availability of different systems
(plants, callus cultures, root cultures) of the same species, here
T. peltatum, provides a valuable extension of the chemodiversity
of this productive plant species.
The discovery of new (bi)naphthalenes and related gluco-
sides demonstrates the remarkable ability of T. peltatum to
produce a broad variety of structurally interesting secondary
metabolites. Similar to the naphthylisoquinoline alka-
loids25−28,33,34 and dioncoquinones,35 the naphthalenes and
binaphthalenes identified in the root cultures are likely to be
formed from acetate/malonate units. The basic structures of
the metabolites previously isolated from Ebenaceae and
Liliaceae plants are assumed to arise from heptaketide
precursors, too.4,15,39 The unique biosynthesis of naphthyliso-
quinoline alkaloids,25,28,36 which are the first (and as yet only!)
acetogenic di- and tetrahydroisoquinoline alkaloids found in
nature, is known to be extremely susceptible to all sorts of
chemical, physical, or biotic stress.25,35,40 As a reaction to stress,
the introduction of nitrogen by the postulated transamination
step40 is easily blocked. As a consequence, the formation of the
tetrahydroisoquinoline portion of the alkaloids is inhibited, thus
leading to a predominant production of naphthoquinones and
(bi)naphthalenes in the root cultures of T. peltatum, instead.
EXPERIMENTAL SECTION
■
General Experimental Procedures. Melting points were
obtained on a Reichert Kofler hot-stage microscope and are
uncorrected. IR spectra were recorded on a Jasco FT/IR-410
spectrometer, UV spectra on a Varian Cary 50 Conc spectropho-
tometer, ECD spectra on a Jasco J-715 spectropolarimeter, and optical
Summarizing, like the intact West African liana T. peltatum
itself, its root cultures have become a promising new source of
naphthylisoquinoline alkaloids, among them dioncophylline A
(8),25,26 habropetaline A (11),34 and 5′-O-methyldioncophyl-
line D (12a/b)28 (Figure 2). Furthermore, these cultures,
representing differentiated tissues of a complete organ,
furnished a series of structurally new axially chiral binaph-
rotations on a Jasco P-1020 polarimeter. 1H NMR (600 MHz) and 13
C
NMR (150 MHz) were recorded on a Bruker DMX 600 instrument,
using D2O (δ 4.79) and methanol-d4 (δ 3.31 and 49.15 ppm) as the
solvents and the internal 1H and 13C standards. For the ROESY
F
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
experiments the mixing time was set to 1 s. Proton-detected,
heteronuclear correlations were analyzed using HMQC (optimized
purification of compounds 13−15, to give the naphthylisoquinoline
alkaloids habropetaline A (11) (retention time 18.0 min) and
dioncopeltine A (10) (retention time 20.5 min), along with a mixture
of dioncophylline A (8), 5′-O-demethyldioncophylline A (9), and 5′-
O-methyldioncophylline D (12a/b) (retention time 22.1 min).
Further resolution and purification of the coeluting compounds 8, 9,
and 12a/b was done as reported previously,28 by repeatedly performed
crystallization and preparative HPLC.
Structural elucidation of the known compounds 2 and 8−15 was
achieved by coelution with authentic reference material (for 8−15)
and by comparison of their physical and spectroscopic data with
reported data.19,20,26−28,33−35
Dioncoquinoside A (1): pale brown solid (MeOH); mp 164 °C;
[α]2D0 −230.1 (c 0.20, MeOH); UV (MeOH) λmax (log ε) 227 (1.01),
303 (0.18), 319 (0.14), 331 (0.11) nm; IR (ATR) νmax 3332 (br, m),
2968 (w), 2921 (m), 1617 (m), 1508 (w), 1428 (w), 1365 (m), 1236
(m), 1070 (s), 1039 (s) cm−1; 1H NMR (D2O, 600 MHz) δ 7.82 (1H,
d, 3J = 8.40 Hz, 8-H), 7.41 (1H, dd, 3J = 7.8 Hz, 3J = 8.3 Hz, 7-H), 7.12
(1H, s, 3-H), 6.88 (1H, d, 3J = 7.6 Hz, 6-H), 5.24 (1H, d, 3J = 7.9 Hz,
1″-H), 4.88 (1H, d, 3J = 7.9 Hz, 1′-H), 3.92 (1H, dd, 2J = 12.6 Hz, 3J =
2.3 Hz, 6″β-H), 3.74 (1H, dd, 2J = 12.4 Hz, 3J = 2.4 Hz, 6″α-H), 3.79−
3.73 (1H, m, 2′-H), 3.71−3.69 (1H, m, 2″-H), 3.66−3.63 (2H, m, 6′-
CH2), 3.63−3.61 (1H, m, 5″-H), 3.60−3.58 (1H, m, 3″-H), 3.55−3.48
(1H, m, 3′-H), 3.55−3.48 (1H, m, 4″-H), 3.55−3.48 (1H, m, 4′-H),
3.18−3.16 (1H, m, 5′-H), 2.42 (3H, s, 2-CH3); 13C NMR (D2O, 150
MHz) δ 154.8 (C, C-5), 152.6 (C, C-4), 147.2 (C, C-1), 133.4 (C, C-
9), 131.3 (CH, C-3), 131.3 (C, C-2), 130.4 (CH, C-7), 117.1 (C-10),
116.9 (CH, C-8), 113.2 (CH, C-6), 106.5 (CH, C-1′), 104.9 (CH, C-
1″), 79.1 (CH, C-5″), 78.5 (CH, C-5′), 78.3 (CH, C-3′), 78.3 (CH, C-
3″), 76.4 (CH, C-2′), 75.5 (CH, C-2″), 71.9 (CH, C-4″), 71.8 (CH,
C-4′), 63.1 (CH2OH, C-6″), 62.8 (CH2OH, C-6′), 19.2 (CH3, 2-
CH3); EIMS m/z 514 [M]+ (100), 352 [M − Glc]+ (78), 190 [M −
2Glc]+ (29); HRESIMS m/z 537.1579 [M + Na]+ (calcd for
C23H30NaO13, 537.1579).
1
n
for JHC = 145 Hz) and HMBC (optimized for JHC = 7 Hz)
experiments. HRESIMS analyses were performed on a Bruker
micrOTOF-focus mass instrument. Solvents were either purchased
from commercial suppliers or purified by standard techniques. Thin-
layer chromatography plates were visualized by exposure to ultraviolet
light at 254 and 365 nm. Column chromatography was carried out
utilizing silica gel (0.063 mm, Merck). Preparative HPLC was
performed on a Jasco System (PU-1580 Plus) in combination with
UV/vis detection at 195−650 nm (Jasco MD-2010 Plus diode array
detector). For isolation and purification of the root culture extracts a
Chromolith Semi-Prep RP-18e HPLC column (100 × 10 mm) from
Merck and a Symmetry RP18 column (19 × 300 mm, 7 μm) from
Waters were used; flow rate 10 mL/min; mobile phase (A) H2O
(0.05% TFA, (B) CH3CN (0.05% TFA); room temperature.
Sterile Root Cultures. Aseptic root cultures of T. peltatum were
initiated by transferring stem explants of in vitro cultivated shoot
cultures of T. peltatum31 to a modified Gamborg B5 (G5S2) medium32
with full strength of micro- and macroelements, but supplemented
with 2% sucrose, 0.1% casein hydrolysate, and 0.3% Gelrite.
Adventitious roots were transferred to Erlenmeyer flasks containing
liquid medium of the same composition, but without gelling agent. To
further increase adventitious root formation, 0.2 mg/L of the
phytohormone 1-naphthalene acetic acid was added to the medium.
The resulting root cultures were kept in the dark at 24 2 °C, in 300
mL Erlenmeyer flasks (containing 80−100 mL of liquid medium) in a
submerged state with aeration brought about by continuous shaking of
the cultures (130 rpm). For maintenance and propagation, the root
cultures were transferred to fresh medium every 4 weeks over a couple
of years.
Extraction and Isolation. Lyophilized root material (9.2 g) was
ground and repeatedly extracted (3×) with MeOH/CH2Cl2 (1:1).
The combined extracts were filtered and concentrated in vacuo to give
2.65 g of a crude residue. The extract was dissolved in MeOH and
directly submitted to cation-exchange chromatography using the resin
Amberlyst 15 (Fluka) (column size: 8 × 25 cm). The resin was
successively eluted with MeOH (3 × 10 mL) to provide fraction A.
Subsequent elution with an aqueous saturated NaCl solution
(containing 0.05% TFA) (3 × 5 mL) and finally again with MeOH
(3 × 10 mL) afforded fraction B.
Fraction A was concentrated under reduced pressure, macerated
with water, and then repeatedly extracted with n-hexane to separate
polar compounds (water subfraction, A-1) from moderately polar and
unpolar metabolites (n-hexane subfraction, A-2). Resolution of
subfraction A-1 by preparative HPLC on a Chromolith Semi-Prep
RP-18e HPLC column, using a linear gradient (0 min 5% B, 10 min
40% B, 16 min 100% B, 18 min 100% B, 19 min 5% B, 25 min 5% B),
yielded 13.3 mg of dioncoquinoside A (1) (retention time 8.9 min),
5.6 mg of plumbaside (2) (retention time 12.5 min), and 10.5 mg of
triphyoquinoside A (6) (retention time 20.2 min). From subfraction
A-2, 1.2 mg of triphyoquinol A1 (3) (retention time 14.6 min), 3.0 mg
of triphyoquinol A2 (4) (retention time 13.5 min), 1.77 mg of
triphyoquinol B (5) (retention time 12.4 min), and 1.40 mg of
triphyoquinone A (retention time 11.3 min) were isolated by
preparative HPLC on a Chromolith Semi-Prep RP-18e HPLC
column, using a linear gradient (0 min 30% B, 10 min 70% B, 16
min 100% B, 18 min 100% B, 19 min 30% B, 25 min 30% B).
Furthermore, subfraction A-2 was found to contain the known
metabolites 8-hydroxydroserone (15)35 (retention time 11.8 min),
droserone (14)35,40 (retention time 27.5 min), and plumbagin
(13)35,40 (retention time 32.3 min), which were purified by
preparative HPLC on a Symmetry RP18 column, using a linear
gradient (0 min 5% B, 30 min 70% B, 35 min 100% B, 40 min 100% B,
41 min 5% B, 46 min 5% B), as described earlier.35
Triphyoquinol A1 (3): brown, amorphous solid (MeOH); mp 155
°C; [α]2D0 +27.1 (c 0.05, MeOH); UV (MeOH) λmax (log ε) 227
(1.39), 307 (0.35), 323 (0.24), 335 (0.22) nm; ECD (c 0.1, MeOH)
Δε219 −44.6, Δε235 +56.6, Δε294 −3.6, Δε320 +0.2, Δε339 −5.4 cm2
mol−1; IR (ATR) νmax 3399 (br, w), 2973 (w), 2927 (w), 2512 (br,
m), 2159 (s), 2026 (s), 1974 (s), 1683 (w), 1610 (w), 1511 (w), 1428
(w), 1361 (w), 1180 (w), 1124 (w), 1085 (w), 833 (w), 802 (w), 742
1
(m), 617 (w) cm−1; H NMR and 13C NMR see Table 1; EIMS m/z
374 [M]+ (100), 359 [M − CH3]+ (12), 187 [M/2]+ (23), 172 [M/2
− CH3]+ (9); HRESIMS m/z 375.1596 [M + H]+ (calcd for
C24H23O4, 375.1591).
Triphyoquinol A2 (4): white, crystalline solid (MeOH); mp 152 °C;
[α]2D0 +64.4 (c 0.05, MeOH); UV (MeOH) λmax (log ε) 227 (1.29),
307 (0.28), 323 (0.26), 255 (0.24) nm; ECD (c 0.1, MeOH) Δε218
−44.6, Δε234 +56.6, Δε294 −3.6 cm2 mol−1; IR (ATR) νmax 3659 (br,
w), 3409 (br, w), 2979 (s), 2361 (m), 2161 (br, w), 2029 (br, w),
1716 (w), 1656 (w), 1607 (s), 1457 (m), 1425 (m), 1379 (s), 1361
(s), 1245 (s), 1228 (s), 1118 (s), 1083 (s), 953 (m), 827 (m), 812
(m), 755 (s), 700 (w), 667 (w), 621 (w), 607 (m) cm−1; 1H NMR and
13C NMR see Table 1; EIMS m/z 374 [M]+ (100), 359 [M − CH3]+
(9), 187 [M/2]+ (21), 172 [M/2 − CH3]+ (5); HRESIMS m/z
373.1440 [M − H]+ (calcd for C24H21O4, 373.1440).
Triphyoquinol B (5): brown, amorphous solid (MeOH); mp 106
°C; [α]2D0 −30.4 (c 0.05, MeOH); UV (MeOH) λmax (log ε) 227
(0.89), 307 (0.15), 319 (0.13), 331 (0.10), 443 (0.08) nm; ECD (c
0.1, MeOH) Δε244 +13.22, Δε217 −10.04 cm2 mol−1; IR (ATR) νmax
3401 (br, w), 2975 (w), 2925 (w), 2854 (w), 2524 (br, m), 2159 (s),
2026 (s), 1974 (s), 1718 (w), 1671 (w), 1631 (w), 1430 (w), 1386
(w), 1353 (w), 1267 (w), 1178 (w), 1081 (w), 871 (w), 806 (w), 742
1
(m), 613 (w) cm−1; H NMR and 13C NMR see Table 1; EIMS m/z
374 [M]+ (100), 187 [M/2]+ (14), 172 [M/2 − CH3]+ (6);
HRESIMS m/z 397.1410 [M + Na]+ (calcd for C24H22NaO4,
397.1410).
After removal of the organic solvent from fraction B, the aqueous
solution was repeatedly extracted with CH2Cl2. The organic phases
were evaporated to dryness, then dissolved in MeOH, and directly
submitted to preparative HPLC on a Symmetry RP18 column, applying
the same chromatographic conditions described above for the
Triphyoquinone A (7): pale brown, amorphous solid (MeOH); mp
163 °C; [α]2D0 −324.5 (c 0.02, MeOH); UV (MeOH) λmax (log ε) 227
(0.89), 307 (0.15), 319 (0.13), 331 (0.10), 443 (0.08) nm; ECD (c
G
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
0.1, MeOH) Δε244 +13.22, Δε217 −10.04 mdeg; IR (ATR) νmax 3401
(br, w), 2975 (w), 2925 (w), 2854 (w), 2524 (br, m), 2159 (s), 2026
(s), 1974 (s), 1718 (w), 1671 (w), 1631 (w), 1430 (w), 1386 (w),
1353 (w), 1267 (w), 1178 (w), 1081 (w), 871 (w), 806 (w), 742 (m),
account, while 20 excitations were calculated for each of the
monomeric halves. The excitations of the monomers were then
compared in order to identify the relevant excited states along with
their electronic transition dipole moments. Comparison of the
calculated UV and ECD spectra with the experimental ones was
performed with SpecDis.47 A UV correction of 12 nm in combination
with a spectral bandwidth σ of 0.16 eV was applied to the calculated
curves.
1
613 (w) cm−1; H NMR and 13C NMR see Table 1; EIMS m/z 374
[M]+ (100), 359 [M − CH3]+ (53), 190 [M − naphthalene]+ (5), 188
[M − naphthoquinone]+ (5); HRESIMS m/z 397.1047 [M + Na]+
(calcd for C23H18NaO5, 397.1052).
The P-atropo-diastereomer of triphyoquinoside A (6) was
investigated with an automated conformational search using the
modified-Dreiding48 force field implemented in the Vconf49 program,
which was followed by the optimization of the thus identified
conformations, with PM3.50 The PM3 minimum geometry was taken
as a starting point for the calculation of the UV and ECD spectra of P-
6 using ZINDO/S-Cl51 (40 excited states). The calculated ECD
spectra were compared with the experimental ones by using SpecDis.47
For the calculation of the curves, a UV correction52 of −18 nm in
combination with a spectral bandwidth σ of 0.20 eV was used.
Triphyoquinoside A (6): pale brown, amorphous solid (MeOH);
mp 215 °C; [α]2D0 −206.7 (c 0.2, MeOH); UV (MeOH) λmax (log ε)
227 (0.85), 307 (0.16), 335 (0.11) nm; ECD (c 0.1, MeOH) Δε236
+60.81, Δε221 −41.2 cm2 mol−1; IR (ATR) νmax 3370 (br, m), 2973
(w), 2518 (s, br), 2159 (s), 2026 (s), 1974 (s), 1673 (w), 1427 (w),
1357 (w), 1178 (m), 1139 (w), 1056 (m), 829 (w), 759 (w), 607 (w)
cm−1; 1H NMR (D2O, 600 MHz) δ 7.40 (1H, s, 3-H), 7.25 (1H, d, 3J
3
3
= 7.8 Hz, 6′-H), 7.05 (1H, dd, J = 8.5 Hz, J = 7.8 Hz, 7′-H), 7.03
(1H, dd, 3J = 8.3 Hz, 3J = 7.7 Hz, 7-H), 6.88 (1H, s, 3′-H), 6.72 (1H,
d, 3J = 7.7 Hz, 6-H), 6.62 (1H, d, 3J = 8.8 Hz, 8′-H), 6.45 (1H, d, 3J =
8.5 Hz, 8-H), 5.24 (1H, d, 3J = 7.8 Hz, 1‴-H), 5.13 (1H, d, 3J = 7.9 Hz,
1″-H), 4.39 (1H, d, 3J = 7.7 Hz, 1⁗-H), 4.23 (1H, dd, 2J = 11.7 Hz, 3J
= 2.0 Hz, 6‴-H), 3.94 (1H, m, 6″-H), 3.91 (1H, m, 6‴-H), 3.86 (1H,
ASSOCIATED CONTENT
* Supporting Information
■
S
2
3
m, 5‴-H), 3.85 (1H, m, 6⁗-H), 3.75 (1H, dd, J = 12.2 Hz, J = 6.4
Hz, 6″-H), 3.63 (1H, m, 2‴-H), 3.63 (1H, m, 6⁗-H), 3.61 (1H, m, 2″-
H), 3.56 (1H, m, 3‴-H), 3.52 (1H, m, 5⁗-H), 3.52 (1H, m, 5″-H),
3.51 (1H, m, 3″-H), 3.50 (1H, m, 4‴-H), 3.46 (1H, m, 4″-H), 3.33
(1H, m, 3⁗-H), 3.29 (1H, m, 4⁗-H), 3.25 (1H, m, 2⁗-H), 1.94 (3H,
s, 2-CH3), 1.91 (3H, s, 2′-CH3); 13C NMR (D2O, 150 MHz) δ 156.1
(C, C-5′), 155.3 (C, C-5), 155.0 (C, C-4), 154.4 (C, C-4′), 137.6 (C,
C-2′), 137.5 (C, C-9), 137.1 (C, C-9′), 136.6 (C, C-2), 132.0 (C, C-
1), 128.5 (CH, C-7), 127.7 (C, C-1′), 127.2 (CH, C-7′), 122.2 (CH,
C-8′), 118.0 (CH, C-8), 115.7 (C, C-10′), 115.6 (C, C-10), 114.3
(CH, C-3), 114.1 (CH, C-3′), 111.0 (CH, C-6′), 111.0 (CH, C-6),
105.2 (CH, C-1⁗), 104.5 (CH, C-1″), 104.3 (CH, C-1‴), 78.8 (CH,
C-5″), 78.2 (CH, C-3″), 78.1 (CH, C-5⁗), 78.1 (CH, C-3⁗), 78.1
(CH, C-3‴), 78.0 (CH, C-4⁗), 77.7 (CH, C-5‴), 75.2 (CH, C-2⁗),
75.1 (CH, C-2‴), 75.1 (C, C-2″), 71.5 (CH, C-4‴), 71.3 (C, C-4″),
70.5 (CH2, C-6‴), 62.8 (CH2OH, C-6⁗), 62.5 (CH2OH, C-6″), 20.5
(CH3, 2-CH3), 20.2 (CH3, 2′-CH3); EIMS m/z 832 [M]+ (100), 670
[M − Glc]+ (56), 508 [M − 2Glc]+ (37), 346 [M − 3Glc]+ (25);
HRESIMS m/z 855.2699 [M + Na]+ (calcd for C40H48NaO19,
855.2682).
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.jnat-
prod.6b00439.
1
NMR (1H, 13C, H,1H-COSY, HSQC, HMBC, NOESY,
and ROESY), HRESIMS, and IR spectra of compounds
1 and 3−7 (PDF)
AUTHOR INFORMATION
Corresponding Author
*Tel: +49-931-318-5323. Fax: +49-931-318-4755. E-mail:
bringman@chemie.uni-wuerzburg.de.
■
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This work was supported by the Deutsche Forschungsgemein-
schaft (Br 699/14-2). G.Z. was supported by a grant of the
German Excellence Initiative to the Graduate School of Life
Hydrolysis of the Glucosides 1, 2, and 6. Treatment of the
glucoside dioncoquinoside A (1), plumbaside (2), or triphyoquinoside
A (6) (ca. 1.5 mg) with methanolic HCl or with a commercial β-
glucosidase from almonds (Sigma) led to a complete deglucosidation
of the compounds. For this purpose, the compounds were dissolved in
HCl/MeOH (3 mL, 1:1) at 60 °C for 2 h and cooled to room
temperature, or stirred in 2 mL of DMSO/H2O (1:1) and incubated
with the enzyme at room temperature for 90 min. Ethyl acetate (2 × 5
mL) was added, and the phases were separated. In each case,
evaporation of the aqueous phase gave D-glucose, which was identical
with an authentic sample (Aldrich) by TLC comparison (silica gel,
CHCl3/MeOH/H2O, 9:6:1, detection by spraying with 10% vanillin in
concentrated H2SO4, followed by heating) and by 1H NMR and
optical rotation, [α]D20 +49.0 (c 0.05, H2O), lit.41 [α]2D0 +52.2 (c 0.05,
H2O). For dioncoquinoside A (1) and plumbaside (2), evaporation of
the organic phases afforded plumbagin (13) as the aglycone. Its
chromatographic (HPLC) and spectroscopic (1H NMR) properties
were in accordance with those of a sample isolated from callus cultures
of T. peltatum.35,40 The aglycone of triphyoquinoside A (6) was not
detectable; it was found to decompose during deglucosidation and
workup.
Sciences, University of Wurzburg. We thank Dr. M. Grune and
̈
̈
Mrs. E. Ruckdeschel for the NMR spectroscopy experiments
and Dr. M. Buchner and Mr. F. Dadrich for the mass spectra.
̈
Further thanks are due to Mrs. D. Thieme and Mrs. M. Michel
for technical assistance in establishing and maintaining the root
cultures and to Mr. K. Lutke for helpful advice.
DEDICATION
■
We dedicate this paper to Professor Ulrike Holzgrabe, Chair of
Pharmaceutical and Medicinal Chemistry of the University of
Wurzburg, on the occasion of her 60th birthday.
̈
REFERENCES
■
(1) (a) Arshad, N.; Kappe, C. O. Adv. Heterocycl. Chem. 2010, 99,
33−59. (b) Li, G. Q.; Gao, H.; Keene, C.; Devonas, M.; Ess, D. H.;
Kurti, L. J. Am. Chem. Soc. 2013, 135, 7414−7417. (c) Ma, B.; Miao,
̈
T.; Sun, Y.; He, Y.; Liu, J.; Feng, Y.; Cheng, H.; Fan, Q. H. Chem. -
Eur. J. 2014, 20, 9969−9978. (d) Bringmann, G.; Price Mortimer, A. J.;
Keller, P. A.; Gresser, M. J.; Breuning, M. Angew. Chem., Int. Ed. 2005,
44, 5384−5427.
(2) (a) Busacca, C. A.; Fandrick, D. R.; Song, J. J.; Senanayake, C. H.
Adv. Synth. Catal. 2011, 353, 1825−1864. (b) Magano, J.; Dunetz, J. R.
Chem. Rev. 2011, 111, 2177−2250.
(3) (a) Tu, T.; Maris, T.; Wuest, J. D. J. Org. Chem. 2008, 73, 5255−
5263. (b) He, Y.; Bian, Z.; Kang, C.; Gao, L. Chem. Commun. 2010, 46,
Computational Details. The conformational analysis of the P-
configured dimer triphyoquinone A (7) and the optimization of the
naphthalene and ortho-quinone building blocks, in which the free
binding site of the former biaryl axis was substituted by a hydrogen
atom, were performed with B3LYP/TZVP42,43 using ORCA.44 The
excited-state properties of the three compounds were simulated with
TD-B3LYP/6-31G*42,45 using the Gaussian 0346 software package.
For the dimer, a total number of 40 vertical excitations were taken into
H
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
5695−5697. (c) Kumar, J.; Tsumatori, H.; Yuasa, J.; Kawai, T.;
Nakashima, T. Angew. Chem., Int. Ed. 2015, 54, 5943−5947.
O.; Tasler, S. In Progress in the Chemistry of Organic Natural Products;
Herz, W.; Falk, H.; Kirby, G. W.; Moore, R. E., Eds.; Springer: Wien,
2001; Vol. 82, pp 111−123. (d) Ibrahim, S. R. M.; Mohamed, G. A.
Fitoterapia 2015, 106, 194−225.
(4) Bringmann, G.; Gunther, C.; Ochse, M.; Schupp, O.; Tasler, S. In
̈
Progress in the Chemistry of Organic Natural Products; Herz, W.; Falk,
H.; Kirby, G. W.; Moore, R. E., Eds.; Springer: Wien, 2001; Vol. 82, pp
38−47.
(26) (a) Bringmann, G.; Rubenacker, M.; Jansen, J. R.; Scheutzow,
̈
D.; Ake
́
Assi, L. Tetrahedron Lett. 1990, 31, 639−642. (b) Bringmann,
(5) Mallavadhani, U. V.; Panda, A. K.; Rao, Y. R. Phytochemistry
1998, 49, 901−951.
G.; Jansen, J. R.; Reuscher, H.; Rubenacker, M. Tetrahedron Lett. 1990,
̈
31, 643−646.
(6) (a) Yoshihira, K.; Natori, S.; Kanchanapee, P. Tetrahedron Lett.
1967, 8, 4857−4860. (b) Sen, H. G.; Joshi, B. S.; Parthasarathy, P. C.;
Kamat, V. N. Drug Res. 1974, 24, 2000−2003.
́
(27) Bringmann, G.; Rubenacker, M.; Vogt, P.; Busse, H.; Ake Assi,
̈
L.; Peters, K.; von Schnering, H. G. Phytochemistry 1991, 30, 1691−
1696.
(7) Sankaram, A. V. B.; Narayana Reddy, V. V. Phytochemistry 1984,
23, 2039−2042.
(8) Okuyama, E.; Homma, M.; Satoh, Y.; Fujimoto, H.; Ishibashi, M.;
Yamazaki, M.; Satake, M.; Ghazali, A. B. A. Chem. Pharm. Bull. 1999,
47, 1473−1476.
(9) Tanaka, T.; Furusawa, M.; Ito, T.; Iliya, I.; Oyama, M.; Iinuma,
M.; Tanaka, N.; Murata, J. Nat. Prod. Commun. 2007, 2, 55−59.
(10) Paphassarang, S.; Becchi, M.; Raynaud, J. Tetrahedron Lett.
1984, 25, 523−526.
(11) Wang, J. H.; Humphreys, D. J.; Stodulski, G. B. J.; Middleton, D.
J.; Barlow, R. M.; Lee, J. B. Phytochemistry 1989, 28, 1825−1826.
(12) Colegate, S. M.; Dorling, P. R.; Huxtable, C. R. Phytochemistry
1986, 25, 1245−1247.
(13) Colegate, S. M.; Dorling, P. R.; Huxtable, C. R. Phytochemistry
1986, 26, 979−981.
(14) Colegate, S. M.; Dorling, P. R.; Huxtable, C. R.; Skelton, B. W.;
White, A. H. Aust. J. Chem. 1985, 38, 1233−1241.
(15) Tsipouras, A.; Goetz, M. A.; Hensens, O. D.; Liesch, J. M.;
Ostlind, D. A.; Williamson, J. M.; Dombrowski, A. W.; Ball, R. G.;
Singh, S. B. Bioorg. Med. Chem. Lett. 1997, 7, 1279−1282. In this
reference, the axial configuration was correctly determined as S (= P)
according to the exciton chirality method, but was erroneously drawn
as M.
(28) Bringmann, G.; Irmer, A.; Rudenauer, S.; Mutanyatta-Comar, J.;
̈
Seupel, R.; Feineis, D. Tetrahedron 2016, 72, 2906−2912.
(29) (a) Bringmann, G.; Hamm, A.; Gunther, C.; Michel, M.; Brun,
̈
R.; Mudogo, V. J. Nat. Prod. 2000, 63, 1465−1470. (b) Bringmann, G.;
Saeb, W.; Ruckert, M.; Mies, J.; Michel, M.; Mudogo, V.; Brun, R.
̈
Phytochemistry 2003, 62, 631−636.
(30) Note that, different from the IUPAC rules, a 2-methyl-4,5-dioxy
substitution pattern is applied in the numbering of the naphthalene
moiety, regardless of the coupling site, to ensure an easier comparison
of binaphthalenes of different coupling types.
(31) Rembold, K.; Irmer, A.; Poppinga, S.; Rischer, H.; Bringmann,
G. Carniv. Plant Newslett. 2010, 39, 71−77.
(32) Gamborg, O. L.; Miller, R. A.; Ojima, K. Exp. Cell Res. 1968, 50,
151−158.
(33) Bringmann, G.; Saeb, W.; God, R.; Schaffer, M.; Franco
̧
is, G.;
̈
Peters, K.; Peters, E. M.; Proksch, P.; Hostettmann, K.; Ake
Phytochemistry 1998, 49, 1667−1673.
́
Assi, L.
(34) Bringmann, G.; Messer, K.; Schwobel, B.; Brun, R.; Ake
́
Assi, L.
̈
Phytochemistry 2003, 62, 345−349.
(35) (a) Bringmann, G.; Rudenauer, S.; Irmer, A.; Bruhn, T.; Brun,
̈
R.; Heimberger, T.; Stuhmer, T.; Bargou, R.; Chatterjee, M.
̈
Phytochemistry 2008, 69, 2501−2509. (b) Bringmann, G.; Zhang, G.;
Hager, A.; Moos, M.; Irmer, A.; Bargou, R.; Chatterjee, M. Eur. J. Med.
Chem. 2011, 46, 5778−5789.
(16) (a) Zhong, S. M.; Waterman, P. G.; Jeffreys, J. A. D.
Phytochemistry 1984, 23, 1067−1072. (b) Maiti, B. C.; Musgrave, O.
(36) (a) Bringmann, G.; Wohlfarth, M.; Rischer, H.; Grune, M.;
C. J. Chem. Soc., Perkin Trans. 1 1986, 675−681. (c) Uc-Coachon
́
, A.
es, M.;
́
z, R. Nat. Prod. Res. 2013, 27, 1174−
̈
H.; Molina-Salinas, G. M.; Said-Fernan
Caceres-Farfan, M.; Borges-Argae
́
dez, S.; Men
́
dez-Gonzal
́
Schlauer, J. Angew. Chem., Int. Ed. 2000, 39, 1464−1466. (b) Bring-
́
́
mann, G.; Mutanyatta-Comar, J.; Greb, M.; Rudenauer, S.; Noll, T. F.;
̈
1178. (d) Li, X. C.; van der Bijl, P.; Wu, C. D. J. Nat. Prod. 1998, 61,
817−820. (e) Prajoubklang, A.; Sirithunyalug, B.; Charoenchai, P.;
Suvannakad, R.; Sriubolmas, N.; Piyamongko, S.; Kongsaeree, P.;
Kittakoop, P. Chem. Biodiversity 2005, 2, 1358−1367. (f) Wang, X.;
Irmer, A. Tetrahedron 2007, 63, 1755−1761.
(37) (a) Green, S.; Green, T. L.; Heslop-Harrison, Y. Bot. J. Linn. Soc.
1979, 78, 99−116. (b) Marburger, J. E. Am. J. Bot. 1979, 66, 404−411.
(c) Bringmann, G.; Schlauer, J.; Wolf, K.; Rischer, H.; Buschbom, U.;
́
Habib, E.; Leon, F.; Radwan, M. M.; Tabanca, N.; Gao, J.; Wedge, D.
́
Kreiner, A.; Thiele, F.; Duschek, M.; Ake Assi, L. Carniv. Plant
E.; Cutler, S. J. Chem. Biodiversity 2011, 8, 2331−2340.
(17) (a) Baker, R. W.; Liu, S.; Sargent, M. V.; Skelton, B. W.; White,
A. H. Chem. Commun. 1997, 451−452. (b) Baker, R. W.; Liu, S.;
Sargent, M. V. Aust. J. Chem. 1998, 51, 255−266.
(18) The 6,6′-coupled binaphthalene sporandol15 and the 8,6′-linked
binaphthoquinone isodiospyrin17 are two examples of naturally
occurring dimeric naphthalenes for which the issue of axial chirality
was investigated more closely.
Newslett. 1999, 28, 7−13. (d) Bringmann, G.; Wenzel, M.; Bringmann,
́
H. P.; Schlauer, J.; Ake Assi, L.; Haas, F. Carniv. Plant Newslett. 2001,
30, 15−21.
(38) (a) Berova, N.; Nakanishi, K. In Circular Dichroism: Principles
and Application, 2nd ed.; Nakanishi, K.; Berova, N.; Woody, R. W.,
Eds.; Wiley-VCH: New York, 2000; pp 337−490. (b) Harada, N.;
Nakanishi, K.; Berova, N. In Comprehensive Chiroptical Spectroscopy−
Applications in Stereochemical Analysis of Synthetic Compounds, Natural
Products, and Biomolecules; Berova, N.; Polavarapu, P. L.; Nakanishi, K.;
Woody, R. W., Eds.; John Wiley & Sons, Inc.: Hoboken, NJ, 2012;
Vol. 2, pp 115−166.
(19) Yue, J.; Lin, Z.; Wang, D.; Feng, Y.; Sun, H. Phytochemistry
1994, 35, 1023−1025.
(20) Serrilli, A. M.; Sanfilippo, V.; Ballero, M.; Sanna, C.; Poli, F.;
Scartezzini, P.; Serafini, M.; Bianco, A. Nat. Prod. Res. 2010, 24, 633−
639.
(39) One of the rare nonpolyketidic chiral binaphthalenes is the well-
known dimeric sesquiterpene gossypol from the cotton plant
Gossypium hirsutum: (a) Kenar, J. A. J. Am. Oil Chem. Soc. 2006, 83,
269−302. (b) Dodou, K. Expert Opin. Invest. Drugs 2005, 14, 1419−
1434.
(21) Rischer, H.; Hamm, A.; Bringmann, G. Phytochemistry 2002, 59,
603−609.
(22) Budzianowski, J.; Budzianowski, A.; Kromer, K. Phytochemistry
2002, 61, 421−425.
(40) (a) Bringmann, G.; Wohlfarth, M.; Rischer, H.; Ruckert, M.;
̈
(23) Budzianowski, J. Phytochemistry 1996, 42, 1145−1147.
(24) (a) Tokunaga, T.; Dohmura, A.; Takada, N.; Ueda, M. Bull.
Schlauer, J. Tetrahedron Lett. 1998, 39, 8445−8448. (b) Bringmann,
́
G.; Rischer, H.; Wohlfarth, M.; Schlauer, J.; Ake Assi, L. Phytochemistry
́
Chem. Soc. Jpn. 2004, 77, 537−541. (b) Kreher, B.; Neszmelyi, A.;
2000, 53, 339−343.
Wagner, H. Phytochemistry 1990, 29, 605−606.
(41) Myhre, D. V.; Smith, F. J. Agric. Food Chem. 1960, 8, 359−360.
(42) For B3LYP, see: (a) Becke, A. D. J. Chem. Phys. 1993, 98,
5648−5652. (b) Lee, C.; Yang, W.; Parr, R. G. Phys. Rev. B: Condens.
Matter Mater. Phys. 1988, 37, 785−789.
(25) (a) Bringmann, G.; Pokorny, F. In The Alkaloids; Cordell, G. A.,
Ed.; Academic Press: New York, 1995; Vol. 46, Chapter 4, pp 127−
́
271. (b) Bringmann, G.; Franco̧ is, G.; Ake Assi, L.; Schlauer, J. Chimia
1998, 52, 18−28. (c) Bringmann, G.; Gunther, C.; Ochse, M.; Schupp,
̈
I
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
(43) For TZVP, see: (a) Schafer, A.; Horn, H.; Ahlrichs, R. J. Chem.
̈
Phys. 1992, 97, 2571−2577. (b) Schafer, A.; Huber, C.; Ahlrichs, R. J.
̈
Chem. Phys. 1994, 100, 5829−5835.
(44) (a) Neese, F.; Becker, U.; Ganyushin, D.; Koßmann, S.; Hansen,
A.; Liakos, D.; Petrenko, T.; Riplinger, C.; Wennmohs, F. ORCA,
version 2.7.0; University of Bonn: Germany, 2010. (b) Neese, F.
WIREs: Comp. Mol. Sc. 2012, 2, 73−78.
(45) For 6-31G*, see: (a) Hehre, W. J.; Ditchfield, R.; Pople, J. A. J.
Chem. Phys. 1972, 56, 2257−2261. (b) Hariharan, P. C.; Pople, J. A.
Theor. Chim. Acta 1973, 28, 213−222.
(46) Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.;
Robb, M. A.; Cheeseman, J. R.; Montgomery, J. A., Jr.; Vreven, T.;
Kudin, K. N.; Burant, J. C.; Millam, J. M.; Iyengar, S. S.; Tomasi, J.;
Barone, V.; Mennucci, B.; Cossi, M.; Scalmani, G.; Rega, N.; Peterson,
G. A.; Nakatsuji, H.; Hada, M.; Ehara, M.; Toyota, K.; Fukuda, R.;
Hasegawa, J.; Ishida, M.; Nakajima, T.; Honda, Y.; Kitao, O.; Nakai,
H.; Klene, M.; Li, X.; Knox, J. E.; Hratchian, H. P.; Cross, J. B.;
Bakken, V.; Adamo, C.; Jaramillo, J.; Gomperts, R.; Stratmann, R. E.;
Yazyev, O.; Austin, A. J.; Cammi, R.; Pomelli, C.; Ochterski, J. W.;
Ayala, P. Y.; Morokuma, K.; Voth, G. A.; Salvador, P.; Dannenberg, J.
J.; Zakrzewski, V. G.; Dapprich, S.; Daniels, A. D.; Strain, M. C.;
Farkas, O.; Malick, D. K.; Rabuck, A. D.; Raghavachari, K.; Foresman,
J. B.; Ortiz, J. V.; Cui, Q.; Baboul, A. G.; Clifford, S.; Cioslowski, J.;
Stefanov, B. B.; Liu, G.; Liashenko, A.; Piskorz, P.; Komaromi, I.;
Martin, R. L.; Fox, D. J.; Keith, T.; Al-Laham, M. A.; Peng, C. Y.;
Nanayakkara, A.; Challacombe, M.; Gill, P. M. W.; Johnson, B.; Chen,
W.; Wong, M. W.; Gonzalez, C.; Pople, J. A. Gaussian 03, revision
E.01; Gaussian, Inc.: Wallingford, CT, 2004.
(47) (a) Bruhn, T.; Schaumloffel, A.; Hemberger, Y.; Bringmann, G.
̈
SpecDis, version 1.60; University of Wurzburg: Germany, 2012.
̈
(b) h t t p : / / w w w - o r g a n i k. ch em i e . u n i - w u e r z b u r g . d e /
lehrstuehlearbeitskreise/bringmann/specdis/. (c) Bruhn, T.;
Schaumloffel, A.; Hemberger, Y.; Bringmann, G. Chirality 2013, 25,
̈
243−249.
(48) For the original Dreiding force field, see: Mayo, S. L.; Olafson,
B. D.; Goddard, W. A. J. Phys. Chem. 1990, 94, 8897−8909.
(49) Vconf, version 1.0; VeraChem, LLC: Germantown, MD, USA,
2006.
(50) (a) Stewart, J. J. P. J. Comput. Chem. 1989, 10, 209−220.
(b) Stewart, J. J. P. J. Comput. Chem. 1989, 10, 221−264.
(51) Zerner, M. C. In Reviews in Computational Chemistry; Lipkowitz,
K. B.; Boyd, D. B., Eds.; Wiley-VCH Inc.: New York, 1991; Vol. 2,
Chapter 8, pp 313−365.
(52) (a) Bringmann, G.; Bruhn, T.; Maksimenka, K.; Hemberger, Y.
Eur. J. Org. Chem. 2009, 2009, 2717−2727. (b) Bringmann, G.; Gotz,
̈
D.; Bruhn, T. In Comprehensive Chiroptical Spectroscopy−Applications
in Stereochemical Analysis of Synthetic Compounds, Natural Products, and
Biomolecules; Berova, N.; Polavarapu, P. L.; Nakanishi, K.; Woody, R.
W., Eds.; John Wiley & Sons, Inc.: Hoboken, NJ, 2012; Vol. 2, pp
355−386.
J
DOI: 10.1021/acs.jnatprod.6b00439
J. Nat. Prod. XXXX, XXX, XXX−XXX