Diterpenoids from Euphorbia pithyusa subsp. cupanii

Article abstract of DOI:10.1021/np990209u

The aerial parts of Euphorbia pithyusa subsp, cupanii collected in Sardinia afforded eleven novel diterpenoids belonging to the lathyrane (1a), premyrsinane (4a-g), and tigliane (5a-c) types. Compounds 4a-g and 5a are esters of two new parent alcohols, na

Full text of DOI:10.1021/np990209u

J . Nat. Prod. 1999, 62, 1399-1404
1399
Diter p en oid s fr om Eu ph or bia pith yu sa su bsp . cu pa n ii
Giovanni Appendino,,*,† Emanuela Belloro,† Gian Cesare Tron,† J asmin J akupovic,*,‡ and Mauro Ballero§
Dipartimento di Scienza e Tecnologia del Farmaco, Universita` di Torino, Via Giuria 9, 10125 Torino, Italy,
Dipartimento di Scienze Botaniche, Universita` di Cagliari, Viale San Ignazio 13, 09123 Cagliari, Italy, and
Analyticon AG, Hermannswerder Haus 17,14473 Potsdam, Germany
Received May 3, 1999
The aerial parts of Euphorbia pithyusa subsp. cupanii collected in Sardinia afforded eleven novel
diterpenoids belonging to the lathyrane (1a ), premyrsinane (4a -g), and tigliane (5a -c) types. Compounds
4a -g and 5a are esters of two new parent alcohols, named premyrsinol and 4,12,20-trideoxyphorbol,
respectively. Structures were elucidated by spectroscopic and chemical methods. Puzzling differences
between the NMR data of lathyrol (1c) and its esters were rationalized in terms of flipping of the
exomethylene around the mean plane of the macrocycle.
A spurge named pityusa was held in great esteem in the
Greek, Latin, and medieval systems of medicine and was
mentioned in many ancient medical treatises.1 Tradition-
ally, this plant has been identified as Euphorbia pithyusa
L. (Euphorbiaceae), a species native to the Western Medi-
terranean region.2 The name pithyusa refers to its resem-
blance to a small pine,2 and the latex and seeds of this plant
were prescribed for a bewildering array of conditions, some
typical of spurges (constipation, removal of warts), but
others peculiar for this drug (breast ailments, general
tonic).1,2 The roots were also used as a cheap substitute
for turbith (Convolvolus turpethum L.), a practice which
caused dramatic and often lethal effects in patients.3
Euphorbia pithyusa bears no obvious morphological simi-
larity with the other spurges of the Mediterranean area
and is considered a paleoendemism.4 Despite this, and the
historical relevance of this plant in medicine and pharmacy,
no chemical study has been performed so far on its
constituents. We report here the isolation of eleven novel
diterpenoids from E. pithyusa L. subsp. cupanii (Guss.) A.
R. Sm. () E. cupanii Guss. ex Bertol.), a plant endemic to
the dry areas of Sardinia, Corsica, and Sicily.2,4
oxyrane carbons (δ 58.98, s and 55.43, t) were replaced by
two signals in the double bond region (δ 144.3, s, and 115.6,
t). To confirm that 1a was an ester of lathyrol, 1a and the
Resu lts a n d Discu ssion
The plant material (aerial parts) came from the area of
Gennargentu, in central Sardinia. An acetone extract was
separated by column chromatography to afford three major
(yield > 0.02%) crystalline constituents. The isolation of
the minor compounds required further purification by
HPLC, owing to their very similar chromatographic be-
havior.
Compound 1a (C32H40O7, HRMS) was the least polar of
the three major constituents. Its 1H NMR spectrum was
similar to that of the Euphorbia factor L1 (2),5 the first
Euphorbia diterpenoid obtained in pure form (Table 1).6
The major difference between the 1H NMR spectra of 1a
and 2 was the replacement of the AB system of the epoxide
protons of 2 by two olefin protons (δ 4.99 and 4.72, s). This
suggested that 1a was the deoxy derivative of 2, an
observation in accordance with the molecular formula and
the 13C NMR spectrum, where the resonances of the
known lathyrol ester Euphorbia factor L3 (1b)7 were
hydrolyzed, affording an identical triol (1c), having the
melting point of lathyrol.7 Deoxygenation of 2 with I2-and
polymer-supported triphenylphosphine8 afforded a com-
pound identical to 1a , confirming the results of the hy-
drolysis and establishing that the esterification patterns
of 1a and 2 are identical. Despite the important role of
lathyrol (1c) in the chemistry of the genus Euphorbia, no
data besides its melting point7 have been reported, prompt-
ing us to characterize lathyrol also from the spectroscopic
point of view. The 1H- and 13C NMR spectra of 1c, the
hydrolysis product of 1a and 1b, confirmed the atom
connectivity and the configuration expected for lathyrol
(Table 1), but several unexpected and puzzling differences
with 1a and 1b were noticed. Thus, in the 1H NMR
spectrum of 1c, the olefinic proton H-12 resonated ca. 0.50
ppm upfield than in 1a and 1b (δ 6.04 vs δ 6.50 and 6.49,
respectively), while the allylic methyl (H-20) was moved
downfield (ca. 0.25 ppm), and J 4,5 decreased, in absolute
value, from ca. 10 Hz to ca. 3 Hz. Shifts of this type are
not uncommon in macrocyclic compounds, and the change
in J 4,5 closely parallels that observed in ∆11, 6(17)-jatropha-
dienes upon interconversion of “endo” and “exo” conforma-
* To whom correspondence should be addressed. Tel.: (G.A.) +39 011
670 7684; (J .J .) +49 331 230 0213. Fax: (G.A.) + 39 011 670 7687; (J .J .) +
49 3331 230 0263. E-mail: (G.A.) appendin@pharm.unito.it, (J .J .) jjakupovic@
Analyticon-ag.com.
† Dipartimento di Scienza e Tecnologia del Farmaco.
‡ Analyticon AG.
§ Dipartimento di Scienze Botaniche.
10.1021/np990209u CCC: $18.00
© 1999 American Chemical Society and American Society of Pharmacognosy
Published on Web 09/03/1999

1400 J ournal of Natural Products, 1999, Vol. 62, No. 10
Appendino et al.
Ta ble 1. 1H NMR Data for Compounds 1a , 1c, and 3b (δ,
CDCl3 for 1a and 1b, CD3OD for 3b)a-c
position
1a
1c
3bd
1R
1â
2
3.36 dd (14, 8)
1.42 dd (14, 11) 1.81 dd (14, 10) 1.63 dd (13, 11)
2.20 m
2.80 dd (14, 10) 2.94 dd (13, 8.5))
2.17 m
1.93 m
3
5.58 t (3.5)
4.37 br d (3)
4.27 t (4)
4
5
2.76 dd (10, 3.5) 2.26 br t (3)
2.22 dd (8.5, 4)
4.86 d (8.5)
4.20 dd (8.5, 4))
-
1.97 ddd (15, 4, 4)
1.84 ddd (15, 12, 8.5)
1.32 ddd (12, 8.5, 4)
6.11 d (10)
2.20 m
4.44 br s
2.53 m
1.73 m
1.84 m
1.18 m
1.12 m
7R
7â
8R
8â
9
2.06 m
2.02 m
1.74 m
1.14 m
11
12
16
17a
17b
18
19
20
1.37 dd (11, 8)
6.49 d (11)
0.71 d (6.5)
4.99 br s
4.72 br s
1.18 s
1.39 dd (10, 8) 1.47 dd (11.5, 8.5)
6.04 br d (10)
1.20 d (6.5)
5.11 br s
4.96 br s
1.16 s
7.26 d (11.5)
1.08 d (7)
5.11 br s
4.90 br s
1.18 s
1.16 s
1.68 br s
1.19 s
1.14 s
1.99 br s
1.67 br s
a
b
500 MHz. J values are given in Hz in parentheses. Other
signals (δ), for 1a : 7.33-7.21 (m, OPhAc), 3.63 (d, J ) 15 Hz,
OPhAc), 3.61 (d, J ) 15 Hz, OPhAc). For 1c: 4.53 (s, OH-1), 3.47
(s, OH-3), 3.38 (s, OH-5). c Selected NOEs, for 1c: H-20, H-11;
H-20, H-4; H-20-1R; H-18, H-11; H-19, H-12; H-5, H-7â; H-5, H-12;
H-4, H-17a; H-4, H-17b. For 3b: H-7, H-4; H-7, H-9; H-7, H-1;
F igu r e 1. Calculated conformation of lathyrol (1c, A) and 7-hydroxyl-
athyrol (3b, B).
H-18, H-9; H-18, H-11; H-19, H-8â; H-19, H-12; H-20, H-11. 1H
d
NMR data in CDCl3: δ 3.10 (1H, dd, J ) 14, 10 Hz, H-1R), 4.38
(1H, t, J ) 3 Hz, H-3), 2.28 (1H, dd, J ) 8.5, 3 Hz, H-4), 4.83 (1H,
d, J ) 8.5 Hz, H-5), 4.24 (1H, dd, J ) 8.5, 4); 6.98 (1H, d, J ) 11.4
Hz, H-12), 5.12 (1H, br s, H-17a), 4.93 (1H, br s, H-17b), 1.71 (3H,
br s, H-20).
almost orthogonal relationship, and makes possible the
formation of a web of intramolecular hydrogen bondings
which offsets the energy loss due to the decrease planarity
within the enone system (Figure 1A). The observation that
all the three hydroxyls of 1c are involved in intramolecular
hydrogen bonding is consistent with the observation that
their chemical shift was not changed by dilution, while a
decreased conjugation with the endocyclic double bond
underlies the dramatic downfield shift of the ketone
carbonyl in 1c compared to 1a and 1b (∆δ ca. 10 ppm).
Indeed, lathyrol (1c) behaves spectroscopically as an un-
conjugated ketone, as judged from the chemical shift of the
enone â-proton (δ 6.04) and the enone carbonyl (δ 206.7).12
In sharp contrast to lathyrol, 7-hydroxylathyrol (2b) retains
the conformation of its ester, since in this conformation the
5-hydroxyl is ideally located to interact with the 7â-
hydroxyl, and a 3,5,7-hydrogen bonding web can be formed
(Figure 1B). This web is attained without decrease of
conjugation within the enone system, and its formation is
therefore favored over the 3, 5, 15 web.
The detection of a small J 4,5 value in lathyrol (1c) and
of a larger value in its esters might have contributed to
the confusing situation regarding the configuration at C-5
of lathyrol and 5-hydroxyisocharaciol13 derivatives. In the
publications describing the X-ray analysis of Euphorbia
factor L1 (1a )11 and 7-hydroxylathryol (3b),10 no specific
configurational data were in fact reported, and, even now,
the configuration of the 5-oxygen function of 1a is some-
times still incorrectly reported as â.14 Overall, the confor-
mational changes of ∆6(17),12-lathyradienes bear striking
similarities with those detected in ∆6(17),11-jatrophadienes.
Both are triggered by flipping of the exomethylene around
tions.9 Yet, their detection in 1c stands in sharp contrast
to what was observed during the hydrolysis of the 7-hy-
droxylathyrol ester Euphorbia factor L2 (3a ).10 In this case,
no significant change was observed for J 4,5, while the
chemical shift of the allylic methyl was moved upfield (∆δ
-0.16, CDCl3) and H-12 underwent a downfield shift (+
0.43 ppm, CDCl3) (Table 1). As a result of opposite shifts
compared to their esters, overall differences of almost 1
ppm (δ 6.04 vs 6.98, CDCl3) for the signal of H-12, and of
almost 0.30 ppm (δ 1.99 vs 1.71, CDCl3) for the signal of
the allylic methyl were observed between lathyrol (1c) and
its 7-hydroxy derivative (3b) (Table 1).
To rationalize these puzzling observations, a detailed
investigation on the conformation of lathyrol (1c) was
undertaken. The results of NOE difference experiments
(Table 1) revealed that lathyrol adopts a conformation with
the C-7 exomethylene on the mean plane of the macrocycle,
and the allylic methyl perpendicular to it (Figure 1, A). The
esters of lathyrol and 7-hydroxylathyrol, as well as 7-hy-
droxylathyrol itself (3b), adopt instead a conformation
having these groups approximately perpendicular to the
mean plane of the macrocycle, and syn-oriented on its
R-face (Figure 1B). A similar geometry was found in the
X-ray analysis of the Euphorbia factor L1 (1b).11 The
changes in the NMR spectra of lathyrol and its esters are
presumably related to a shielding effect of the C-15 ester
carbonyl on H-12, and to a different extent of planarity
within the enone moiety. Thus, removal of the C-15 ester
carbonyl is expected to move the signal of H-12 downfield,
as observed in the hydrolysis of 7-hydroxylathyrol esters,
while in the hydrolysis of lathyrol esters this effect is offset
by the reduced conjugation within the enone system,
eventually resulting in an upfield shift for H-12. Pivotal
to this effect is the flipping of the C-7 exomethylene from
a perpendicular to a parallel orientation toward the mean
plane of the macrocycle. This brings H-4 and H-5 into an
the macrocycle, and exhibit as hallmark changes in the
9
values of J 4,5
.
The crystalline alcohol 4a had a molecular weight 636
daltons (HRMS), corresponding to the molecular formula
C35H48O12. The 1H NMR spectrum (Table 2) showed signals
diagnostic of three acetates (δ 2.08, 2.07, 2.05, s), one
isobutyrate (δ 2.37, m; 1.09, d, J ) 7 Hz; 1.06, d, J ) 7
Hz), and one propionate residue (δ 2.33, q, J ) 7 Hz; 1.08,
t, J ) 7 Hz), accounting for all the extra carbons of the

Diterpenoids from E. pithyusa supsp. cupanii
J ournal of Natural Products, 1999, Vol. 62, No. 10 1401
Ta ble 2. 1H NMR Data (δ, CDCl3) for Compounds 4a -ga-c
position
4a
4b
4c
4d
4e
4f
4g
1R
1â
2
3.13 dd (14,8)
1.60 dd (14,13)
1.80 m
3.17 dd (14,8)
1.63 dd (14,13)
1.85 m
3.17 dd (14,8)
1.62 dd (14,13)
1.80 m
3.14 dd (14,8)
1.60 dd (14,13) 1.60 dd (14,13) 1.64 dd (14,13)
1.78 m
3.15 dd (14,8)
3.16 dd (14,8)
3.16 dd (14,8)
1.60 dd (14,13)
1.80 m
1.78 m
1.80 m
3
4
5.25 t (3.5)
5.23 t (3.5)
5.26 t (3.5)
5.25 t (3.5)
5.26 t (3.5)
2.40 m
5.38 t (3.5)
2.39 dd (11, 3.5) 2.33 m
5.25 t (3.5)
2.32 dd (11, 3.5) 2.38 dd (11, 3.5) 2.44 dd (11, 3.5) 2.41 m
5
7
8R
8â
9
6.17 d (11)
4.50 d (7)
2.07 m
1.78 br d (17)
0.71 m
6.23 d (11)
4.71 d (7)
2.09 m
1.90 br d (17)
0.77 m
6.11 d (11)
4.42 d (7)
2.06 m
1.80 br d (17)
0.71 m
6.18 d (11)
4.41 d (7)
2.07 m
1.78 br d (17)
0.69 m
6.18 d (11)
4.48 d (7)
2.05 m
1.80 br d (17)
0.71 m
6.38 d (11)
4.79 d (7)
2.05 m
1.85 br d (17)
0.72 m
6.21 d (11)
4.53 d (7)
2.00 m
1.81 br d (17)
0.70 m
11
12
16
17a
17b
18
19
20
15-OH
0.71 m
3.36 m
0.86 d (6.5)
4.41 d (12)
4.34 d (12)
1.03 s
0.89 s
1.67 s
4.46 s
0.83 m
3.48 m
0.92 d (6.5)
4.88 d (12)
4.51 d (12)
1.07 s
0.95 s
1.74 s
4.41 s
0.75 m
3.38 m
0.89 d (6.5)
4.35 d (12)
4.31 d (12)
1.05 s
0.89 s
1.72 s
4.48 s
0.74 m
3.35 m
0.86 d (6.5)
4.34 d (12)
4.31 d (12)
1.03 s
0.89 s
1.69 s
4.42 s
0.74 m
3.38 m
0.88 d (6.5)
4.41 d (12)
4.35 d (12)
1.04 s
0.91 s
1.69 s
4.45 s
0.76 m
3.56 m
0.86 d (6.5)
4.69 d (12)
4.31 d (12)
1.05 s
0.94 s
1.71 s
4.41 s
0.73 m
3.38 m
0.91 d (6.5)
4.43 d (12)
4.35 d (12)
1.05 s
0.92 s
1.69 s
4.52 s
a
b
500 MHz. J are given in Hz in parentheses. Other signals (δ), for 4a : O-prop: 2.33 (q, J ) 7 Hz), 1.08 (t, J ) 7 Hz); 5-O-iBu, 2.37
(m), 1.09 (d, J ) 7 Hz), 1.06 (d, J ) 7 Hz); OAc-7, 2.07 (s); OAc-13, 2.08 (s); OAc-17, 2.05 (s). For 4b: O-prop, 2.33 (q, J ) 7 Hz), 1.09 (t,
J ) 7 Hz); O-iBu, 2.38 (m), 0.88 (d, J ) 7 Hz); OAc-7, 2.11 (s); OAc-13, 2.11 (s); O-Nic, 9.16 (d, J ) 1.5 Hz), 8.82 (dd, J ) 6, 1.1 Hz), 8.19
(dt, J ) 8, 1.5 Hz), 7.44 (dd, J ) 8.5, 1.5 Hz). For 4c: O-prop, 2.35 (q, J ) 7 Hz), 1.10 (t, J ) 7 Hz); O-MeBu, 2.52 (m), 1.35 (m), 1.10 (t,
J ) 6.5 Hz), 1.06 (d, J ) 6.5 Hz); OAc-7, 2.09 (s); OAc-13, 2.11 (s)); O-iBu, 2.35 (m), 1.19 (d, J ) 7 Hz), 1.17 (d, J ) 7 Hz). For 4d : O-prop,
2.32 (m), 1.08 (t, J ) 7 Hz); 5-O-iBu, 2.37 (m); 1.09 (d, J ) 7 Hz); 1.07 (d, J ) 7 Hz); OAc-7, 2.07 (s); OAc-13, 2.08 (s): 17-O-iBu, 2.50 (m),
1.17 (d, J ) 7 Hz), 1.14 (d, J ) 7 Hz); 2.05 (s). F or 4e: O-prop, 2.36 (q, J ) 7 Hz), 2.34 (q, J ) 7 Hz), 1.16 (t, J ) 7 Hz), 1.10 (t, J ) 7
Hz); O-iBu, 2.37 (m); 1.09 (d, J ) 7 Hz); 1.07 (d, J ) 7 Hz); OAc, 2.10 (s), 2.08 (s). For 4f: O-Prop, 2.30 (q, J ) 7 Hz), 0.96 (t, J ) 7 Hz);
OBz, 7.87 (AA′), 7.50 (C), 7.37 (BB′); OAc, 2.15 (s), 2.12 (s), 2.12 (s). For 4g: O-iBu, 2.41 (m), 1.12 (2 x Me, d, J ) 6.5 Hz); OAc, 2.11 (s),
2.11 (s), 2.09 (s), 2.03 (s). c Selected NOEs for 4a : H-12, OAc-13; H-3, H-4; H-17a, H-20; H-17a,b-H-7; H-5, H-12.
diterpenoid skeleton. One exchangeable proton at δ 4.46
showed the presence of a free hydroxyl, confirmed by the
presence of a IR absorption band at 3475 cm-1. Apart from
the signals of the ester groups and of three methyl singlets,
the 1H NMR spectrum could be interpreted in terms of
three independent aliphatic spin systems. One started from
a pair of diastereotopic methylene protons (δ 3.13, dd, J )
14, 8 Hz, H-1R, δ 1.60, dd, J ) 14, 13 Hz, H-1â), and
continued with one methine adjacent to a methyl (δ 1.80,
m, H-2), and with three additional methines (δ 5.25, t, J
) 3.5 Hz, H-3; δ 2.32, m, H-4; δ 6.17, d, J ) 11 Hz, H-5),
two of which were oxygenated on account of their downfield
resonance. The second spin system was an isolated AB
system of an oxygenated methylene (δ, 4.41 and 4.34, d, J
) 12 Hz, H-17a,b), while the last spin system started from
an oxymethine (δ 4.50, d, J ) 7 Hz, H-7), and proceeded
through a methylene (δ 2.07, m, H-8R; δ 1.78, br d, J )
17, H-8â) to three aliphatic methines (δ 0.71, m, H-9; δ
0.71, m, H-11; δ 3.36, m, H-12). The high-field chemical
shift of two of them suggested the presence of a cyclopro-
pane moiety. Three methyl singlets (δ 1.03, C-18; δ 0.89,
C-19 and δ 1.67, C-20) and one methyl doublet (δ 0.86, d,
J ) 6.5 Hz) were also present. The remaining nonproto-
nated carbons were one carbonyl (δ 204.3, C-14), two
aliphatic oxygenated quaternary carbons (δ 85.9 and 83.9,
s, C-13 and C-15), and one aliphatic quaternary carbon (δ
47.3, s, C-6). HMBC correlations allowed the assembly of
these features into a premyrsinane topology. In this
context, the most relevant feature was the detection of
HMBC correlations between the methine H-12 and both
C-6 and C-7, an observation requiring the connection of
C-12 to the quaternary carbon C-6. Location of the ester
groups was assessed by analysis of the HMBC correlations
between the ester carbonyls and the oxymethine (oxy-
methylene) protons, while the remaining ester group (an
acetate) was located at C-13 on account of a NOE effect
between H-12 and the acetyl signal at δ 2.07. NOE effects,
summarized in Table 2, were also pivotal to assess the
configuration as depicted in 4a . The polyol corresponding
to this structure is new, and we have named it premyrsinol.
The third crystalline product obtained by column chroma-
tography of the extract (4b) was closely related to 4a , the
only differences being related to the replacement of one
acetyl with one nicotinate group. Differences in the chemi-
cal shift of the diastereotopic methylene at C-17, backed
up by HMBC measurements, showed that the nicotinate
group was located at this position.
1H NMR spectroscopic analysis of the noncrystalline
fractions of E. pithyusa subsp. cupanii revealed the pres-
ence of mixtures of premyrsinanes containing small amounts
of phorbol-related compounds (characteristic broad singlet
for H-1 around 7.50 ppm). HPLC separation of these
mixtures afforded five additional premyrsinanes (4c-g)
and three tiglianes (5a -c). Compounds 4c-g showed the
same spin-systems of the diterpenoid core of 4a ,b, and
differed only for the esterification pattern, which contained,
besides acetyl residues, various combinations of propionic-,
R-methylbutyric, isobutyric, and benzoic acids. In all cases,
location of the ester groups could be assessed in a straight-
forward way by the inspection of diagnostic HMBC cor-

1402 J ournal of Natural Products, 1999, Vol. 62, No. 10
Appendino et al.
relations between the carbonyl carbons and their corre-
sponding oxymethines (oxymethylene), and by NOE-
difference experiments for the tertiary ester groups. The
results showed that compounds 4a -g shared the same
esterification pattern at C-7 and C-13, while various
combinations of aliphatic and aromatic acids were present
at the oxygenated carbons C-3, C-5, and C-17.
amplified by esterification with a diverse array of acids,
as exemplified very well by the structure of compounds
4a -g.
Exp er im en ta l Section
Gen er a l Exp er im en ta l P r oced u r es. Melting points were
determined on a Bu¨chi SMP-20 apparatus and are uncorrected.
IR spectra were recorded on
a Perkin-Elmer model 237
Compound 5a (C26H38O4, HRMS) displayed NMR signals
indicative of a diterpenoid esterified with 2,3-dimethylbu-
tyric acid. The olefinic signals at δ 7.56 (s, H-1) and 5.22
(br s, H-7) were typical of a tigliane derivative. No other
1
spectrophotometer. H and 13C NMR spectra were taken on a
1
Bruker DRX instrument (500 and 125 MHz, respectively). H
and 13C NMR chemical shifts refer to CHCl3 at 7.26 ppm, and
to CDCl3 at 77.0 ppm, respectively. HRMS were obtained on
a MAT 95ST Finnigan-MAT apparatus (70 eV, EI mode). CIMS
were carried out on a Finnigan-MAT apparatus TSQ 70 using
isobutane as a reactant gas. Si gel 60 (70-230 mesh) was used
of open column chromatography. A Hibar Lichrosorb column
(2.5 × 25 cm, Merck) was used for HPLC separations. Figure
1 was generated with PCMODEL, Serena Software, Version
4.0 (Serena Software, Bloomington, IN). Compounds 1b, 2, and
3a were available from previous work on E. lathyris L.5b
P la n t Ma t er ia l. Aerial parts (leaves and stems) of E.
pithyusa subsp. cupanii were collected around Arzana (Nuoro,
Sardinia, Italy) in J une 1998. The plant material was identi-
fied by Mauro Ballero, and a voucher specimen (1212) is kept
at the Dipartimento di Scienze Botaniche, University of
Cagliari.
1
nonexchangeable downfield signals were present in the H
NMR spectrum, in accordance with the quaternary nature
of the two oxygenated sp3 carbons detected in the 13C NMR
spectrum (δ 75.2 and 62.8, s, C-9 and C-13, respectively).
Since a signal diagnostic for H-4 (δ 2.40, ddd, J ) 10, 9, 4
Hz) could be detected, and H-20, resonated as an allylic
methyl (δ 1.71, br s), 5a was a derivative of 4,12,20-
trisdeoxyphorbol, a new type of tigliane polyol representing
the most deoxygenated form of phorbol reported to date.
It is not known whether this compound and the other less
oxygenated analogues of phorbol are derived from the
parent polyol by removal of oxygen atoms, or, alternatively,
from the transannular cyclization of less oxygenated
lathyrane precursors. The â-configuration of H-4 was
evident from its splitting pattern and the chemical shift of
H-1,15 while the site of esterification was located at the 13-
hydroxyl by comparison of the 13C NMR resonances of the
ring C carbons and those of known 12-deoxyphorbol
esters.15 This was further supported by the detection of a
low-field 1H NMR signal for the 9-hydroxyl (δ 5.54, s), a
feature diagnostic of an intramolecular hydrogen bonding
between the carbonyl of the C-13 ester group and the
tertiary 9-hydroxyl.16 The two remaining tiglianes (5b and
5c) were isolated in trace amounts (less than 0.1% w/w of
the purified extract), and could not be obtained completely
free of cyclomyrsinane impurities (ca. 15% in both cases).
Extr a ction a n d Isola tion . Dried and powdered plant
material (400 g) was extracted with Me2CO at room temper-
ature (1 × 1.5 L, 2 × 1 L). The pooled extracts were evaporated
in vacuo, and the residue was suspended in EtOH (400 mL)
and treated with an equal volume of 3% Pb(OAc)2. After about
3 h, the suspension was filtered on a bed of Celite, and the
clear filtrate was concentrated in vacuo to remove most of the
EtOH, and then extracted with EtOAc. After washing with
brine, drying (Na2SO4), and evaporation, a brown residue (4.2
g) was obtained. The latter was purified by open column
chromatography on Si gel (ca. 30 g), using mixtures of hexane
and EtOAc (from 9:1 to 3:7). According to differences in
composition indicated by TLC, 10 crude fractions were ob-
tained. Fractions A-C contained triterpenoids and fats and
were not further investigated. Fractions D, G, and I crystal-
lized from diethyl ether, affording 275 mg 1a (0.069%), 171
mg 4a (0.043%), and 99 mg 4b (0.025%). The mother liquors
from the crystallization of 1a were further separated by HPLC
(hexanes-EtOAc 7:3) to give 5 mg 5a . After HPLC (hexanes-
EtOAc 7:3), fraction E afforded 3 mg 4c and 36 mg 4d , and
fraction F 3 mg 5c and 23 mg 4e. Fraction H was further
purified by HPLC (hexanes-EtOAc 6:4) to afford 4f (12 mg)
and 4g (6 mg). Fraction L (120 mg) was first chromatographed
on Sephadex LH-20 (5 g). Elution with hexanes-EtOAc 4:6
afforded 19 mg of a colorless gum, which was further purified
by HPLC (hexanes-EtOAc 4:6) to afford 2 mg 5b.
1
Their structure elucidation was based on MS and H NMR
data alone, but was helped greatly by the similarity of the
spectra with those of 5a . Indeed, the differences could be
rationalized in terms of oxygenation of C-20 to an hy-
droxymethyl-(5b) and to an acetoxymethyl group (5c).The
detection of a downfield singlet for the 9-hydroxyl (δ 5.70
in 5b and δ 5.68 in 5d ) is in accordance with the location
of the ester group at C-13, as is the virtual identity of all
the proton resonances around ring C. The presence of a
hydroxyl at C-20, and of the 13-acyloxy-9-hydroxy group
on ring C make 5b a potential protein kinase C (PKC)
ligand, while 5c should be considered its corresponding
“cryptic” form.17 These compounds are presumably respon-
sible for the cathartic activity of the plant mentioned in
the ancient literature.1,3
La th yr ol-3-p h en yla ceta te-5,15-d ia ceta te () d eoxy Eu -
25
ph or bia fa ctor L1) (1a ): white powder, mp 125-127 °C; [R]D
+195° (c 0.9, CHCl3); IR (KBr) νmax 1740, 1728, 1647, 1622,
1263, 1238, 1128, 1010, 1005 cm-1; H NMR data, see Table
1
1; 13C NMR (CDCl3) δ 48.3 (t, C-1), 37.4 (d, C-2), 80.5 (d, C-3),
52.2 (d, C-4), 65.8 (d, C-5), 144.3 (s, C-6), 35.0 (t, C-7), 21.3 (t,
C-8), 35.3 (d, C-9), 25.2 (d, C-10), 28.4 (d, C-11), 146.7 (d, C-12),
134.1 (s, C-13), 196.8 (s, C-14), 92.3 (s, C-15), 13.7 (q, C-16),
115.6 (t, C-17), 29.0 (q, C-18), 16.8 (q, C-19), 12.4 (q, C-20);
PhAc: 169.8 (s), 135.4 (s), 129.5 (d), 128.5 (d), 127.1 (d), 41.5
(t); OAc: 171.3 (s), 170,7 (s), 22.0 (q), 22.0 (q); EIMS m/z
536.2780 [M]+ (5) (calcd for C32H40O7, 536.2774).
Compounds 4a -g belong to the premyrsinane group of
diterpenoids. Nine examples were previously known,18 but
the presence of an acyloxy group at C-17 in place of an
intramolecular ether function18a-c or a methyl group,18d sets
4a -g apart from all the other compounds of this class.
The detection of structurally unique diterpenoids in E.
pithyusa subsp. cupanii is not surprising on account of the
unique taxonomic position of this species and the geo-
graphical isolation of Sardinia. The detection of new
diterpene polyols shows that spurges are a source of
structurally diverse isoprenoids much richer than assumed
from bioactivity-directed fractionation with the mouse ear
erythema assay, which selectively targets compounds with
phorbol ester-like activity. This skeletal diversity is further
P r em yr sin ol-3-p r op a n oa te-5-isobu tyr a te-7,13,17-tr ia c-
25
eta te (4a ): white powder, mp 176-178 °C; [R]D -15° (c 1.1,
MeOH); IR (KBr) νmax 3475, 1736, 1728, 1653, 1367, 1289,
1
1245, 1159, 1038 cm-1; H NMR data, see Table 2; 13C NMR
data, see Table 3; EIMS m/z 636.3137 [M]+ (1) (calcd for
C
35H48O12, 636.3145), 71 (100).
P r em yr sin ol-3-pr opan oate-5-isobu tyr ate-7,13-diacetate-
17-n icotin a te (4b): white powder, mp 179-181 °C; [R]D
-18° (c 1.2, MeOH); IR (KBr) νmax 3470, 1732, 1714, 1650,
25

Diterpenoids from E. pithyusa supsp. cupanii
J ournal of Natural Products, 1999, Vol. 62, No. 10 1403
Ta ble 3. 13C NMR (δ, CDCl3) Data for Compounds 4a -ga,b
4,12,20-Tr ideoxyph or bol-13-(2,3-dim eth yl)bu tyr ate (5a):
gum, [R]D25 +35° (c 0.5, MeOH); IR (liquid film) νmax 3510, 1715,
position
4a
4b
4c
4d
4e
4f
4g
1
1660, 1390, 1119, 1100, 1016, 970 cm-1; H NMR (CDCl3) δ
C-1
C-2
C-3
C-4
C-5
C-6
C-7
C-8
42.8 t 42.8 t 42.8 t 42.8 t 42.8 t 42.8 t 42.7 t
37.3 d 37.4 d 37.4 d 37.3 d 37.3 d 37.2 d 37.3 d
78.2 d 78.3 d 78.2 d 78.2 d 78.2 d 78.1 d 78.2 d
50.2 d 50.5 d 50.5 d 50.4 d 50.4 d 50.3 d 50.1 d
68.7 d 69.0 d 68.6 d 68.6 d 68.7 d 69.8 d 68.7 d
47.3 s 47.6 s 47.2 s 47.2 s 47.4 s 47.7 s 47.4 s
70.5 d 70.7 d 71.2 d 71.1 d 70.8 d 70.6 d 70.5 d
22.0 t 22.3 t 21.9 t 21.9 t 22.0 t 22.0 t 22.1 t
18.7 d 18.9 d 18.7 d 18.8 d 18.8 d 18.9 d 18.8 d
18.0 s 18.3 s 18.1 s 18.0 s 18.1 s 18.2 s 18.1 s
23.8 d 23.8 d 24.0 d 23.7 d 24.3 d 23.8 d 23.8 d
34.7 d 35.0 d 34.7 d 34.7 d 34.7 d 35.0 d 34.7 d
85.9 s 85.7 s 86.0 s 86.0 s 86.0 s 85.7 s 85.9 s
7.56 (1H, br s, H-1), 5.54 (1H, s, OH-9), 5.22 (1H, br s, H-7),
3.29 (1H, br s, H-10), 2.81 (1H, dd, J ) 18, 9 Hz, H-5b), 2.40
(1H, ddd, J ) 10, 9, 4 Hz, H-4), 2.19 (1H, m, H-2′), 2.10 (1H,
dd, J ) 15, 6 Hz, H-12a), 2.07 (1H, br dd, J ) 6.5, 4 Hz, H-8),
1.98 (1H, dd, J ) 18, 10 Hz, H-5a), 1.93 (1H, m, H-3′), 1.71
(3H, br s, H-20), 1.53 (1H, dd, J ) 15, 4, H-12b), 1.19 (3H, s,
H-17), 1.09 (3H, d, J ) 6.5 Hz, Me-2′), 1.02 (3H, s, H-16), 0.93
(3H, d, J ) 6.5 Hz, Me-3′), 0.91 (3H, d, J ) 6.5 Hz, H-18), 0.90
(3H, d, J ) 6.5 Hz, Me-3′), 0.75 (1H, d, J ) 5 Hz, H-14); 13C
NMR (CDCl3) δ 161.0 (d, C-1), 138.3 (s, C-2), 203.0 (s, C-3),
44.4 (d, C-4), 34.0 (t, C-5), 136.2 (s, C-6), 126.8 (d, C-7), 41.9
(d, C-8), 75.2 (s, C-9), 53.9 (d, C-10), 46.2 (d, C-11), 31.8 (t,
C-12), 62.8 (s, C-13), 32.0 (d, C-14), 22.5 (d, C-15), 15.2 (q,
C-16), 22.9 (q, C-17), 19.0 (q, C-18), 10.0 (q, C-19), 25.2 (q,
C-20), 178.0 (s, C-1′), 35.3 (d, C-2′), 30.5 (d, C-3′), 20.7 (d, C-4′),
19.1 (q, C-5′), 13.3 (q, C-6′); EIMS m/z 414.2981 [M]+ (1) (calcd
for C26H38O4, 414.2982), 281 (100).
C-9
C-10
C-11
C-12
C-13
C-14 204.3 s 204.3 s 204.5 s 204.4 s 204.4 s 204.3 s 204.3 s
C-15
C-16
C-17
C-18
C-19
C-20
83.9 s 84.1 s 84.1 s 84.0 s 84.0 s 84.1 s 84.0 s
14.0 q 14.1 q 14.1 q 14.0 q 14.0 q 13.8 q 14.0 q
63.4 t 64.3 t 63.7 t 63.5 t 63.3 t 62.8 t 63.4 t
29.4 q 29.4 q 29.5 q 29.4 q 29.4 q 29.4 q 29.5 q
14.7 q 14.8 q 15.0 q 14.8 q 14.8 q 14.8 q 14.8 q
24.5 q 24.5 q 23.9 q 24.5 q 23.8 q 24.8 q 24.5 q
4,12-Did eoxyp h or bol-13-(2,3-d im eth yl)bu tyr a te (5b):
gum, IR (liquid film) νmax 3600, 1713, 1660, 1375, 1120, 1060,
a
125 MHz; assignments aided by HMBC and HMQC experi-
ments. Selected HMBC for 4a as representative: H-12, C-5; H-12,
C-7; H-12, C-17; H-3, C-15; H-5, C-15; H-3, CdO (Prop, δ 174.1),
H-5, CdO (iBu, δ 175.0); H-7, CdO (Ac, δ 169.8)); H-17a,b, CdO
1
1011, 980 cm-1; H NMR (CDCl3) δ 7.56 (1H, br s, H-1), 5.70
(1H, s, OH-9), 5.24 (1H, br s, H-7), 4.06 (1H, br d, J ) 11 Hz,
H-20a), 4.03 (1H, br d, J ) 11 Hz, H-20b) 3.28 (1H, br s, H-10),
3.16 (1H, dd, J ) 18, 9 Hz, H-5â), 2.42 (1H, ddd, J ) 10, 9, 4
Hz, H-4), 2.19 (1H, m, H-2′), 1.94 (1H, m, H-3′), 1.71 (1H, br
s, H-19), 1.61 (1H, dd, J ) 15, 4 Hz, H-12b), 1.20 (3H, s, H-17),
1.09 (3H, d, J ) 6.5 Hz, Me-2′), 1.06 (3H, s, H-16), 0.97 (3H, d,
J ) 6.5 Hz, Me-3′), 0.91 (3H, d, J ) 6.5 Hz, Me-3′), 0.77 (1H,
d, J ) 5 Hz, H-14); CIMS m/z 431 [M]+ [C26H38O5 + H]+ (31).
4,12-Did eoxyp h or bol-13-(2,3-d im eth yl)bu tyr a te-20-a c-
eta te (5c): gum, IR (liquid film) νmax 3600, 1718, 1660, 1380,
1150, 1061, 1090, 990 cm-1; 1H NMR (CDCl3) δ 7.55 (1H, br s,
H-1), 5.68 (1H, s, OH-9), 5.56 (1H, br s, H-7), 4.42 (1H, br d,
J ) 12 Hz, H-20a), 4.35 (1H, br d, J ) 12 Hz, H-20b) 3.26
(1H, br s, H-10), 3.15 (1H, dd, J ) 18, 9 Hz, H-5â), 2.40 (1H,
m, H-4), 2.19 (1H, m, H-2′), 2.04 (3H, s, OAc), 1.93 (1H, m,
H-3′), 1.69 (3H, br s, H-19), 1.56 (1H, dd, J ) 15, 4 Hz, H-12b),
1.19 (3H, s, H-17), 1.09 (3H, d, J ) 6.5 Hz, Me-2′), 1.03 (3H, s,
H-16), 0.93 (3H, d, J ) 6.5 Hz, Me-3′), 0.90 (3H, d, J ) 6.5 Hz,
Me-3′), 0.75 (1H, d, J ) 5 Hz, H-14); CIMS m/z 475 [M]+
[C28H40O6 + H]+, (10).
b
(Ac, δ 170.2). Other signals (δ): for 4a : O-Prop, 174.1 (s), 27.7
(t), 8.8 (q); O-iBu, 175.0 (s), 34.0 (d), 18.7 (q), 18.5 (q); OAc-7, 169.8
(s), 21.2 (q); OAc-13, 170.5 (s), 21.2 (q); OAc-17, 170.2 (s), 21.1 (q).
For 4b, O-Prop: 174.0 (s), 27.7 (t), 8.8 (q); O-iBu, 174.9 (s), 33.9
(d), 18.4 (q), 18.3 (q); O-Nic, 164.8 (s), 153.9 (d), 150.5 (d), 136.7
(d), 126.0 (s), 123.7 (d); OAc-7, 169.9 (s), 21.2 (q); OAc-13: 170.6
(s), 21.3 (q). For 4c: O-Prop, 174.1 (s), 27.7 (t), 8.8 (q); O-iBu, 174.7
(s), 34.1 (d), 18.8 (q), 18.7 (q); O-MeBu, 176.4 (s), 40.9 (d), 27.2 (t),
11.6 (q), 18.8 (q); OAc-7, 169.9 (s), 21.2 (q); OAc-13, 170.7 (s), 21.3
(q). For 4d : O-Prop, 174.1 (s), 27.7 (t), 8.8 (q); O-iBu, 175.1 (s),
34.1 (d), 18.7 (q), 18.5 (q); 7-OAc, 169.8 (s), 21.2 (q); OAc-13, 170.6
(s), 21.3 (q); 17-O-iBu, 176.4 (s), 34.1 (d), 19.3 (q), 18.6 (q). For
4e: O-Prop, 174.1 (s), 173.7 (s), 27.7 (t), 27.7 (t), 9.0 (q), 8.8 (q);
O-iBu, 175.1 (s), 34.1 (d), 18.7 (q), 18.5 (q); OAc-7, 169.7 (s), 21.3
(q); OAc-13, 170.6 (s), 21.3 (q). For 4f: O-Prop, 173.5 (s), 25.7 (t),
8.7 (q); O-Bz, 165.1 (s), 132.0 (d), 129.8 (s), 129.6 (d), 128.2 (d);
OAc, 170.7 (s), 170.6 (s), 170.1 (s), 21.3 (q), 21.3 (q), 20.5 (q). For
4g: O-iBu, 175.2 (s), 34.1 (d), 18.7 (q), 18.5 (q); OAc, 171.0 (s),
170.7 (s), 170.3 (s), 170.0 (s), 21.3 (q), 21.3 (q), 21.2 (q), 21.1 (q).
Deoxygen a tion of Eu ph or bia F a ctor L1 (2). To a solution
of iodine (270 mg, 1.07 mmol, 3.7 mol equiv) in CH2Cl2 (20
mL) was added polymer-supported triphenylphosphine (350
mg, 1.10 mmol, 3.8 mol equiv), resulting in the decoloration
of the solution and the formation of a black precipitate. After
stirring at room temperature for 15 min, a solution of Euphor-
bia factor L1 (2) (160 mg, 0.29 mmol) in CH2Cl2 (5 mL) was
added. After further stirring for 10 min, the reaction mixture
was worked up by filtration, and the filter cake was washed
with CH2Cl2 (10 mL). The pooled filtrates were washed
sequentially with 5% Na2S2O3 and brine. After drying (Na2-
SO4) and removal of the solvent, a yellowish solid was
obtained. Washing with diethyl ether gave 81 mg (82%) 1a as
a white powder, identical (1H NMR, TLC) to the natural
product.
1
1360, 1281, 1249, 1150, 1010 cm-1; H NMR data, see Table
2; 13C NMR data, see Table 3; EIMS m/z 699.3255 [M]+ (0.5)
(calcd for C37H49NO12, 669.3255), 124 (100).
P r em yr sin ol-3-p r op a n oa te-5(R-m eth yl)bu tyr a te-7,13-
25
d ia ceta te-17-isobu tyr a te (4c): gum, [R]D -16° (c 0.9,
MeOH); IR (liquid film) νmax 3450, 1740, 1729, 1650, 1376,
1295, 1245, 1150, 1030 cm-1; H NMR data, see Table 2; 13C
1
NMR data, see Table 3; EIMS m/z 678.3622 [M]+ (1) (calcd
for C36H54O12, 678.3615), 57 (100).
P r em yr sin ol-3-p r op a n oa te-5,17-d iisobu tyr a te-7,13-d i-
a ceta te (4d ): white powder, mp 72-75 °C; [R]D25 -11° (c 0.9,
MeOH); IR (KBr) νmax 3496, 1740, 1729, 1372, 1230, 1192,
1063, 670 cm-1 1H NMR data, see Table 2; 13C NMR data,
;
see Table 3; EIMS m/z 664.3457 [M]+ (2) (calcd for C35H52O12
,
Hyd r olysis of Deoxy Eu ph or bia F a ctor L1 (1a ). Com-
pound 1a (200 mg, 0.37 mmol) was suspended in 5% KOH in
MeOH (2 mL). After stirring at room temperature for 4 h, the
reaction was worked up by dilution with water (8 mL) and
extraction with EtOAc. The organic phase was washed with
brine and dried (Na2SO4). The residue was crystallized from
ether to give 68 mg (55%) lathyrol (1c) as a white powder,
identical to the compound obtained from the hydrolysis of
Euphorbia factor L3 (1b): mp 168-170 °C (lit.: 168-169 °C);7
664.3459), 71 (100).
P r em yr sin ol-3,17-d ip r op a n oa te-5-isobu tyr a te-7,13-d i-
25
a ceta te (4e): gum, [R]D -16° (c 0.9, MeOH); IR (liquid film)
νmax 3470, 1736, 1729, 1380, 1290, 1250, 1100, 1039 cm-1; H
1
NMR data, see Table 2; 13C NMR data, see Table 3; EIMS m/z
650.3301 [M]+ (1) (calcd for C34H50O12, 650.3302), 57 (100).
P r em yr sin ol-3-p r op a n oa te-5-ben zoa te-7,13,17-tr ia ce-
ta te (4f): gum, [R]D25 -15° (c 1.3, MeOH); IR (liquid film) νmax
3450, 1745, 1719, 1660, 1370, 1297, 1241, 1136, 1031 cm-1
;
25
[R]D +101° (c 1.3, MeOH): IR (KBr) νmax 3389, 3261, 1653,
1H NMR data, see Table 2; 13C NMR data, see Table 3; EIMS
m/z 670.2989 [M]+ (1) (calcd for C36H46O12, 670.2989), 105 (100).
P r em yr sin ol-3-p r op a n oa t e-5-isob u t yr a t e-7,13,17-t r i-
a a ceta te (4g): gum, [R]D25 -14° (c 0.9, MeOH); IR (liquid film)
1641, 1444, 1269, 1047, 910, 904 cm-1; 1H NMR data, see Table
1; 13C NMR (CDCl3) δ 46.8 (t, C-1), 38.1 (d, C-2), 77.2 (d, C-3),
53.2 d (C-4), 69.6 (d, C-5), 147.8 (s, C-6), 33.7 (t, C-7), 23.2 (t,
C-8), 34.8 (d, C-9), 24.0 (s, C-10), 26.0 (d, C-11), 139.9 (d, C-12),
137.0 (s, C-13), 206.7 (s, C-14), 87.9 (s, C-15), 13.9 (q, C-16),
110.9 (t, C-17), 28.6 (q, C-18), 15.5 (q, C-19), 13.6 (q, C-20);
EIMS m/z 334.2160 [M]+ (10) (calcd for C20H30O4, 334.2144).
νmax 3470, 1737, 1380, 1291, 1225, 1158, 1050 cm-1; H NMR
1
data, see Table 2; 13C NMR data, see Table 3; EIMS m/z
622.2981 [M]+ (2) (calcd for C32H46O12, 622.2989), 71 (100).

1404 J ournal of Natural Products, 1999, Vol. 62, No. 10
Appendino et al.
(8) The use of monomeric triphenylphosphine gave a product which could
not be satisfactorily separated from triphenylphosphine oxide by
crystallization or chromatography.
(9) Appendino, G.; J akupovic, S.; Tron, G. C.; J akupovic, J .; Milon, V.;
Ballero, M. J . Nat. Prod. 1998, 61, 749-756.
Hyd r olysis of Eu ph or bia fa ctor L2 (3a ). Compound 3a
(300 mg) was hydrolyzed as described for 1a , giving 89 mg
(54%) 7â-hydroxylathyrol (3b) as a white powder, mp 220-
25
222 °C; [R]D +52° (c 0.9, MeOH): IR (KBr) νmax 3435, 1676,
1615, 1153, 1076, 1059, 1007, 922 cm-1 1H NMR data, see
;
(10) Narayanan, P.; Ro¨hrl, M.; Zechmeister, K.; Engel, D. W.; Hoppe, W.;
Hecker, E.; Adolf, W. Tetrahedron Lett. 1971, 1325-1328.
(11) Zechmeister, K.; Ro¨rl, M.; Brandl, F.; Hechfischer, S.; Hoppe, W.;
Hecker, E.; Adolf, W.; Kubinyi, H. Tetrahedron Lett. 1970, 3071-
3073.
(12) The UV spectra of lathyrol (1c), hydroxylathyrol (3b), and their esters
are characterized by an anomalous nfπ* absorption [278 (ꢀ ) 14 400)
for 1c; 275 nm (ꢀ ) 12 600) for 2b]. These values are somewhat
reminiscent of those observed in taxinine and related taxoids (Ap-
pendino, G. In The Chemistry and Pharmacology of Taxol® and its
Derivatives; Farina, V., Ed.; Elsevier: Amsterdam, 1995; pp 56-58)
and rather different from the ones calculated from Woodward’s rules.
Ring strain and the presence of a vinyl group adjacent to the
endocyclic double bond probably underlay this remarkable batho-
chromic shift. For these reasons, the differences between the UV
spectra of 1c and 3b are difficult to rationalize.
Table 1; 13C NMR (CDCl3-DMSO-d6 1:1) δ 47.2 (t, C-1), 35.8
(d, C-2), 76.9 (d, C-3), 53.2 d (C-4), 63.4 (d, C-5), 147.8 (s, C-6),
76.2 (d, C-7), 22.8 (t, C-8), 30.3 (d, C-9), 26.6 (s, C-10), 27.4 (d,
C-11), 147.8 (d, C-12), 132.2 (s, C-13), 199.8 (s, C-14), 87.2 (s,
C-15), 12.8 (q, C-16), 111.3 (t, C-17), 30.0 (q, C-18), 14.8 (q,
C-19), 11.1 (q, C-20); EIMS m/z 350.2083 [M]+ (10) (calcd for
C20H30O4, 350.2093).
Ack n ow led gm en t. This work was supported by EU
(Contract ERB FAIR CT 1781) and MURST (Fondi 40%,
Progetto Chimica dei Composti Organici di Interesse Bio-
logico). We are grateful to Dr. Raffaele Sestu and the Pro Loco
Arzana (NU) for their hospitality and help during the collection
of the plant material.
(13) Seip, E. H.; Hecker, E. Phytochemistry 1984, 23, 1689-1694.
(14) Hecker, E. in Hagers Handbuch der Pharmazeutischen Praxis;
Blaschek, W., Ha¨nsel, R., Keller, K., Reichling, J ., Rimpler, H.,
Schneider, G., Eds.; Springer: Berlin, 1998; Folgeband 2, pp 644-
649.
Refer en ces a n d Notes
(15) Evans, F. J .; Taylor, S. E. In Progress in the Chemistry of Organic
Natural Products; Herz, W., Grisebach, H., Kirby, G. W., Eds.;
Springer: New York, 1983; Vol. 44, pp 1-99.
(16) This hydrogen bond is typical of all phorboid 13-esters, and was even
detected in the X-ray analysis of phorbol 13-acetate bound to a
fragment of PKC δ (Zhang, G.; Kazanietz, M. G.; Blumberg, P. M.;
Hurley, J . H. Cell 1995, 81, 917-924).
(17) Krauter, G.; Von Der Lieth, C. W.; Schmidt, R.; Hecker, E. Eur. J .
Biochem. 1996, 242, 417-127.
(18) (a) Euphoppins A-D: (Yang, L.; Shi, Y. P.; J ia, Z.; Saleh, S.; Lanham,
J . J . Nat. Prod. 1995, 58, 1883-1888. (b) Aleppicatines A and B:
O¨ ksu¨z, S.; Gu¨rek, F.; Lin, L.-Z.; Gil, R. R.; Pezzuto, J . M.; Cordell, G.
A. Phytochemistry 1996, 42, 473-478. (c) Karajinones A and B:
Ahmad, V. U.; J assbi, A. R. Phytochemistry 1998, 48, 1217-1220.
(d) 17-Deoxy-18-hydroxypremyrsinol-3,18-dinicotinate-5-benzoate-7-
acetate: O¨ ksu¨z, S.; Gu¨rek, F.; Qiu, S.-X.; Cordell, G. A. J . Nat. Prod.
1998, 61, 1198-1201.
(1) Pliny’s opening of the paragraph on pityusa in his Naturalis Historia
has often been quoted: Cum honore et pityusa simile de causa dicetur,
quamdam in tithymali genere numerant (Honorable mention will now
be made of pityusa, which some include in the same class as
tithymalus) (XXIV, XXXI).
(2) Turner, R. Euphorbias; Batsford: London, 1995; pp 148-149.
(3) Petrus Andreas Matthiolis. Pedacio Dioscorides, 1544; CLXVII.
(4) Valsecchi, F. Boll. Soc. Sarda Sci. Nat. 1980, 83, 336-342.
(5) (a) Adolf, W.; Hecker, E.; Balmain, A.; Lhomme, M. F.; Nakatani, Y.;
Ourisson, G.; Ponsinet, G.; Pryce, R. J .; Santhanakrishnan, T. S.;
Matyukhina, L. G.; Saltikova, I. A. Tetrahedron Lett. 1970, 2241-
2244. (b) Appendino, G.; Tron, G. C.; Cravotto, G.; Palmisano, G.;
J akupovic, J . J . Nat. Prod. 1999, 62, 76-79. (c) Itokawa, H.; Ichihara,
Y.; Yahagi, M.; Watanabe, K.; Takeya, K. Phytochemistry 1990, 29,
2025-2026.
(6) This compound was isolated as early as 1890 but was originally
considered a steroid and named “euphorbiasteroid” (Tahara, Y. Chem.
Ber. 1890, 23, 3347-3351).
(7) Adolf, W.; Hecker, E. Experientia 1971, 27, 1393-1394.
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