Phytochrmistry, 1975, Vol. 14, pp. 2073-2076. Pergamon Press. Printed in England.

ALKALOIDS, COUMARINS, TRITERPENES AND A

FLAVANONE FROM THE ROOT OF
ZANTHOXYLUM DIPETALUM*
FRANCIS FISH, ALEXANDER I. GRAY and PETER G. WATERMAN
Division of Pharmacognosy and Forensic Science, Department of Pharmaceutical Chemistry, University of Strathclyde,
Glasgow, Gl lXW, Scotland

(Revised Received 21 February 1975)

Key Word Index-Zanthoxylum dipetalum; Rutaceae; alkaloids; canthin-6-one; chelerythrine; nitidine; tembe-
tarine; coumarins; avicennol; xanthoxyletin; triterpenes; lupeol; sitosterol; flavanones; hesperidin; chemotax-
onomy.

Abstract-The root bark of Zanthoxylum dipetalum contained the alkaloids canthin-6-one, cheleryth-
rine, nitidine and tembetarine, the pyranocoumarins avicennol and xanthoxyletin, the triterpene lupeol
and the flavanoid hesperidin. The MS fragmentation pattern for avicennol is discussed and a tentative
structure is proposed for a third coumarin, designated ZD/l. The root wood of the type species
and the stem bark of the y variety contained most of the above compounds plus sitosterol and, in the
root wood only, magnoflorine. The chemotaxonomic implications of these findings are briefly dis-
cussed.

INTRODUCTlON chemotaxonomic significance [4] we have exam-

The large pan-tropical genus Zunthoxylum L (in- ined samples of the root of the type species and
cluding Fugara L) (fam. Rutaceae) is represented of the stem of Z. dipetulum var y Hillebr. (F. dipe-
in the Hawaiian Islands by six endemic species tulu v. mannii Sherf).
most of which have been sub-divided into several
varieties [ 11. Zanthoxylum dipetalum H. Mann RESULTS AND DISCUSSION
(syn. Fuguru &pet& Engl.), is a small to medium
The major component of the petrol extract
sized tree, distinguished from other Hawaiian spe-
from the root bark of Z. dipetulum was the pyr-
cies by a reduction in the number of petals to
ano[2,3-flcoumarin, avicennol (1), which has
two due to the coalescence of two contiguous.
been isolated previously only from the Asian spe-
petals; by the occurrence of a pair of reduced
cies Z. uvicennue (Lam.) DC [5,6]. Again it
basal leaflets adherent to the lowest pair of nor-
occurred in high concentration (in excess of 1%).
mal leaflets and by the mode of inflorescence [Z].
Avicennol was identified by direct comparison of
Several varieties have been recorded differing
physical and spectral data with that previously
from the type species in that they retain the ori-
reported and, in addition, a detailed study was
ginal tetramerous state [l-3].
made of the MS, for which accurate mass
As part of a continuing study on the secondary
measurements of significant ions were obtained.
metabolites of Zunthoxylum and their possible
The major fragmentation pattern can be rational-
*Part 5 in the series Chemosystematics in the Rutaceae. ized as dehydration of the 3-hydroxy-3-methyl-
For Part 4 see Waterman. P. G. (1975) A Review of the but-1-enyl side-chain and loss of a Me radical
Chemosystematics of the genus Zanthoxylum (Rutaceae) in from the pyran ring to give a fragment at m/e
South East Asia in, Roles and Goals of Tropical Botanic
Gardens (Stone, B. C. ed.), University of Malaya, Kuala Lum-
309. This is in agreement with previously pub-
pur. lished MS for pyranocoumarins [7]. A minor
2073
PHYTO141% L
3074 F. FIW, A. 1. GRAY and P. G. WATFR~MAN

&
I O
21 ’
0 0 0
/
m/e 310
OH
(lb) (1) OH

-Me*

m/e 295

Scheme I.

pathway, giving fragments at m/e 310 and 295 the major alkaloid of Z. semi-articulatum St. John
may be explained by elimination of methanol & Hosaka [9], the only other Hawaiian Zanthor-
with subsequent cyclization to the pyranocou- y/urn species investigated to date. The MeOH
marin (lb) followed by loss of a Me radical extract also yielded large quantities of the fla-
(Scheme 1). vanone glycoside hesperidin.
Four other compounds were isolated from the The root wood and the stem of the 1’variety
petrol extract by PLC; lupeol, canthin-6-one and contained a similar range of compounds but niti-
two coumarins one of which was identified as dine was absent from the root wood and hesperi-
xanthoxyletin. The second coumarin, designated din and tembetarine were not present in the y
ZD/l, gave UV and IR spectra typical of pyrano- variety. Sitosterol and magnoflorine were
coumarins [7], the IR spectrum showing a detected; t-he former in both samples, the latter
marked similarity to that of avicennol. Accurate only in the root wood.
mass measurement indicated the empirical for- In view of the disparity between the plant parts
mula C19H1804 with a fragmentation pattern examined little chemotaxonomic value can as yet
showing loss of only a Me radical. These data be placed on these findings from the intraspecific
suggest ZD/l may be a coumarin of novel type viewpoint although the absence of hesperidin
with a di-pyran ring system. Although no cou- from the stem of the 1’variety does seem unusual.
marins with such a ring system are known an At the generic level however. the occurrence
analogous dihydro-dipyrano acridone alkaloid of avicennol in Z. dipctalum (sect. Blackburnia)
has recently been reported from Atalantia mono- and Z. NZ;~C~WW (sect. Paniculatae-Gerontogaeae)
phylla Correa (Rutaceae) [8]. If these tentative and of canthin-6-one in widespread and seemingly
conclusions are correct the most likely structure distantly related species [IO] would appear to cast
for ZDjl would seem to be (lb). Unfortunately further doubt on Englerian classification of the
this compound has proved difficult to purify and, Zanthoxylum/Fayara complex [4].
as yet, very little material is available.
Investigation of the CHCl, and MeOH extracts EXPERIMETT.41,
yielded three quaternary alkaloids, the benzo-
phenanthridines chelerythrine and nitidine and UV spectra were recorded in EtOH and IR spectra in KC?.
NMR (60 M HZ) spectra were recorded with TMS as internal
the l-benzyltetrahydroisoquinoline tembetarine. standard. MS were determined at 70eV. Mps (uncorr) were
Chelerythrine has previously been reported to be determined on a Kofer hot stage.
 Alkaloids and coumarins of Zanthoxylum dipetalum 2075

Plant material. Root bark and root wood of Zanthoxylum nm: 271, 279(sh), 318, 339(sh) and 394. Identical in all respects
dipetalum H. Mann (Voucher: GS 8 at BISH, HLA, NBV) [1 I] (UV, IR, TLC, mmp, conversion to dihydrochelerythrine) with
were collected at the Pupukea-Paumalu Forest Reserve, Koo- an authentic sample of chelerythrine chloride.
lau Mountains, Oahu, Hawaii State. Stem bark of Z. dipetalum Nitidine chloride. Recrystallized from EtOH as yellow nee-
var. y (Voucher: Herbst 1030 at BISH, US, HLA, NBV) [11] dles mp 240” decomp. (lit. [I63 240” decomp.). UV km,, nm:
was collected at the junction of the Kokee and Halemanu 231, 272, 281, 301, 329 and 385. Identical in all respects (UV,
roads, Kokee State Park, Kauai, Hawaii State. IR, TLC, mmp, conversion to dihydronitidine) with an authen-
Extraction of root bark qf Z. dipetalum. Milled bark (55 g) tic sample of nitidine chloride.
was extracted in a Soxhlet separately and successively with Hesperidin. Recrystallized from MeOH as buff coloured nee-
petrol (bp 4&60”), CHCl, and MeOH. The extracts were cone dles mp 257’ decomp. (lit. [17] 255-260” decomp.). UV E.,,,
under red pres. On standing a ppt formed in the petrol con- nm: 283 and 329; I,,, AlCl, nm: 306 and 380 [IQ A posi-
centrate and it was collected by decanting the supernatant tive reaction was obtained with the Mg/HCl test for fla-
sol. The crystals were washed with n-hexane to yield avicennol vanones [18] and the material was identical in all respects
(560mg). The supernatant extract was furthe; cone and an (UV, IR, TLC, mmp) with an authentic sample of hesperidin.
aliquot (50”/,) subjected to PLC on alumina (Woelm. neutral Tembetarine iodide. Recrystallized from EtOH-EtOAc as
activity I) with n-hexane -EtOAc (4: 1) as eluting solvent. Xan- white plates mp 151-160”. Found M+ - 1 343.1774,
thoxyletin (R, @3; 12 mg), lupeol (RJ @4; 15 mg) and an un- C2,H2,N0, requires M -1 343.1783. [a]$’ + 91” (c
identified compound designated ZDjl (RJ 0.5, 10mg) were 0.32EtOH). UV I,,, nm: 286; npay nm: 296. IR ~~~~crn-‘:
isolated. A second aliquot was treated by PLC on Si gel G 3350 (OH), 1620, 1520, 1270, and 1240. MS m/e (rel. int.):
with CHCl, as eluting solvent to give ZD/l (R/ 0.5; 12mg) 343 (7), 192 (lOO), 177 (18), 137 (2). Temhetarine chloride. De-
and canthin-6-one (RI @ 1; 8 mg). The CHCl, concentrate, on posited from the purified EtOH extract before evaporation
shaking with 1% aq HCI, gave a yellow ppt in the aq phase and addition of KI and recrystallized from EtOH-EtOAc
which was filtered off to yield chelerythrine chloride (91 mg). (15:3) as white prisms mp 236” (lit. [193 237”). Identical spec-
The acid extract was basified with NH,OH and re-extracted tral data (UV, IR, MS) was obtained to that recorded for
with CHCI, to give canthin-6-one (15 mg). tembetarine iodide.
On standing the MeOH concentrate deposited an amor- Extraction of root wood of Z. dipetalum. Milled root wood
phous buff ppt of hesperidin (1.5g). The supernatant extract (80 g) was extracted as previously described for the root bark.
was purified by ion-exchange chromatography [12] and from Examination of the petrol extract on TLC (alumina, solvent
the resulting EtOH extract yellow crystals of nitidine chloride CHCI,) indicated the presence of traces of avicennol. xantho-
(13 mg) were obtained. The EtOH extract was then evan to xyletin, canthin-6-one. lupeol, sitosterol and compound ZCjl.
dryness and the alkaloid mixture dissolved in the minimum Examination of the CHCl, extract (alumina, CHCl,-MeOH,
H,O. A saturated soln of KI was added dropwise to ppt the 49: 1) showed further traces of canthin-6-one and of cheleryth-
alkaloids as the iodides from which, after repeated recrystalli- rine chloride whilst the MeOH extract (Cellulose, 0.1 N HCl)
zation from EtOH-EtOAc, tembetarine iodide (104mg) was contained tembetarine and a further quaternary alkaloid,
obtained. probably magnoflorine. Hesperidin (375 mg) was isolated from
Avicennol (1). Recrystallized from n-hexane-EtOAc (99: 1) the MeOH extract and identified as previously described.
as yellow plates mp 124.5-I 25.5” (lit. [S] 124.>125.5”). Found Extraction of the stem bark of Z. dipetalum var. Milled bark
M’ 342. 1477, C,,H,,05 requires A4 342.1467. UV A,,,,, nm (150 g) was extracted in the manner previously described. The
(log E): 250 (4.50) 257 (4.60) and 301 (4.26 IR v,,, cm-‘: petrol concentrate was subjected to column chromatography
3475 (OH), 1725 (C=O), 1585, 1140 and 825. NMR (CDCI,): on alumina (Woelm, activity IV). Elution with n-hexane gave
6 1.48 (12H, s, T-Me, and 3”-Me,), 2.15 (lH, s, D,O exchange, lupeol (20 mg), n-hexane-EtOAc (9: 1) gave xanthoxyletin
OH), 3.80 (3H, s, dMe), 5.69 aid 6.65 (2H, A& J 1OGz, (24 mg) and n-hexane-EtOAc (4: 1) gave sitosterol (30 mg). All
3’.H and 4’-H). 6.80 and 6.92 (2H. A&. J 16Hz. 2”-H and were identified by direct comparison with authentic material.
1”-H), 6.27 ani 8.06 (2 H, A&; J 10 Hz, 3-H and 4-H). MS Trace quantities of canthin-6-one and avicennol were detected
m/e (rel. int.): 342 (23), 327 (IOO), 324 (52), 310 (16), 309 (66), by TLC (systems as before). Similarly traces of chelerythrine
295 (6). On admixture with an authentic sample of avicennol and canthin-6-one were detected in the CHCl, extract.
from Z. auicennae there was no depression of mp.
Xanthoxyletin. Recrystallized from MeOH as colourless Acknowledgements-We thank Mr. R. T. Hirano and Mr. G.
prisms mp 132-133” (lit.1131 133”). UV /I,,, nm: 223, 268.5 Spence of the Lyons Herbarium, University of Hawaii for the
and 346. Identical in all respects (UV, IR, TLC, mmp) with supply and authentication of plant material, Dr. R. D. Waigh,
an authentic sample of xanthoxyletin. Department of Pharmaceutical Chemistry, University of
Comnound ZDil. Amoruhous yellow oowder from MeOH Strathclyde for NMR spectra, and the University of Strath-
with nb sharp mp. Found M’ ~10.1198. C,,H,,O, requires Clyde for a studentship awarded to one of us (AIG).
310.1205 UV i.,,, nm: 223, 243(sh), 250, 296, 306(sh) and 336.
IR v,,,, cm-‘: 1730 (C=O), 1610, 1360, 1135, 1025, 825, 740 REFERENCES
and 705. MS m/e (rel. int.): 310 (26). 295 (100).
Lupeol. Recrystallized from Me,CO as white needles mp 1. Sherf, E. E. (1958) Am. J. Botany 45, 461.
217-218” (lit. [I41 215-216”). Identical in all respects (IR, TLC, 2. Hillebrand, W. (1888) Flora of the Hawaiian Islands, p.
mmp) with an authentic sample of lupeol. 76. Williams and Noigate, London.
Canthin-6-one. Recrystallized from MeOH as pale cream 3. Rock. J. F. (1913) The Indiuenous Trees of the Hawaiian
needles mp 162-163” (lit. [Sl 162-163”). Found M+ 220.0613, Islands, p. 2b7. Published under patronage, Honolulu.
C,,H,N,b requires M 220.0637. UV i,,, nm: 227, 250, 259, 4. Fish, F. and Waterman, P. G. (1973) Taxon 22, 177.
268.299.346.362 and 379. Identical in all resnects. \
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as yellow needles mp 200-202” (lit. [IS] 202-203”). UV imax chemistry 14, 841.
2076 F. FISH. A. I. GRAY and P. G. WATERMAN

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