X.-L. Zhang et al. / Phytochemistry Letters 16 (2016) 82–86
83
+
Compound 2 exhibited a [M + Na] peak at m/z 691.4548 in the
HRESIMS (calcd 691.4544), consistent with a molecular formula of
1
13
C
41
H
64
O
7, requiring 10 degrees of unsaturation. The H and
NMR data (Tables 1 and 2) of 2 were similar to those of compound
. After hydrolysis with 0.1 M NaOMe, methyl hexadecanoate was
C
1
+
R
assigned by GC–MS (t 17.2 min, m/z 270 [M] ). In addition, NMR
signals for a hexadecanoyl group in 2 were observed to replace
those for an acetyl moiety in 1. Other characteristic resonances
included signals for a tiglyl group (Zhang et al., 2013; Chumkaew
3
et al., 2003). In the HMBC spectrum, H-12 proton showed a
J
correlation with the carbonyl carbon of the tiglyl group, confirming
the location of the tiglyl group at C-12. Thus, the structure of 2 was
determined to be 12-O-tiglylphorbol-4-deoxy-4b-phorbol-13-
hexadecanoate.
Fig. 1. Chemical structures of compounds 1–4.
Compound 3 was assigned the molecular formula C40H O on
62 7
+
the basis of the HRESIMS (m/z 677.4396 [M + Na] , calcd 677.4388)
0
0
0
1
carbons. The signals at
d
C
209.7 (C-3), 179.4 (C-1 ) and 167.6 (C-1 )
in the C NMR spectrum were attributed to carbonyl carbons. The
obvious HMBC correlations from a methyl singlet at 1.73 (3H, s,
CH -19) to carbons at 159.7 (C-1), 136.4 (C-2) and 209.7 (C-3)
suggested the presence of an -unsaturated carbonyl group. The
H– H COSY correlations of H-1/H-10/H-4/H-5, of H-7/H-8/H-
4 and H-12/H-11/H-18 showed the connection patterns of C-1–C-
0–C-4–C-5, C-7–C-8–C-14 and C-12–C-11–C-18 (Fig. 2). These
signals suggested that compound 1 possessed the 4-deoxy-4
phorbol or 4-deoxy-4 -phorbol backbone (Taylor et al., 1981;
Chumkaew et al., 2003). The chemical shifts of H-1 ( 7.55), H-4
2.48), H-5 ( 2.15 and 2.84) illustrated the 4-deoxy-4
phorbol skeleton of 1 rather than 4-deoxy-4 -phorbol skeleton, in
which the chemical shifts of these protons H-1, H-4 and H-5 were
at around H-1 ( 7.00), H-4 ( 2.70), H-5 ( 2.50 and 3.30)
Zhang et al., 2013). The chemical shift values of C-1, C-2 and C–
0 at 159.7, 136.4 and 54.1 also verified the configuration of H-
in compound 1 (Chumkaew et al., 2003). In addition, the
proposed stereochemistry of compound 1 was supported from
NOE correlations between H-4/H-8/H-11 on the face and H-1, H-
, H-14 and H-18 on the other side of the molecule (Fig. 2). The
coupling constant between H-11 and H-12 was 9.7 Hz, confirming
the configuration of H-12 (Chumkaew et al., 2003). Other
characteristic resonances included signals for a tiglyl group and an
and NMR data, indicating 10 degrees of unsaturation. H NMR and
1
3
13
C NMR data (Tables 1 and 2) for compound 3 indicated that
compounds 3 and 1 shared the same backbone. After hydrolysis
d
H
with 0.1 M NaOMe, methyl oleate was identified by GC–MS (t
R
3
d
C
+
a
,
b
21.9 min, m/z 296 [M] ). Other characteristic resonances included
1
1
signals for an acetyl group and an oleyl moiety (Zhang et al., 2013).
3
1
1
In the HMBC spectrum, H-20 showed a J correlation with the
carbonyl carbon of the oleyl group, confirming the location of the
oleyl group at C-20. Thus, the structure of 3 was determined to be
a
-
b
13-O-acetylphorbol-4-deoxy-4b-phorbol-20-oleate.
d
H
Compound 4 was assigned the molecular formula C40H O on
60 7
+
(d
H
d
H
b
-
the basis of the HRESIMS (m/z 675.4220 [M + Na] , calcd 675.4231)
a
and spectroscopic data, indicating 11 degrees of unsaturation. The
1
13
H and C NMR data (Tables 1 and 2) of 4 was quite similar to those
d
H
d
H
d
H
of 3, except that the oleoyl group in 3 was replaced by a linoleoyl
(
1
4
group in 4. Hydrolysis of 4 with 0.1 M NaOMe gave methyl
+
d
C
b
linoleate, which was verified by GC–MS (t
R
23.0 min, m/z 294 [M] )
analysis. Furthermore, In the HMBC spectrum, H-20 proton
3
showed a J correlation with the carbonyl carbon of the linoleyl
b
group, confirming the location of the linoleyl group at C-20. Hence,
the structure of 4 was determined to be 13-O-acetylphorbol-4-
7
deoxy-4
The skeletons of the isolated phorbol diesters can be divided
into three categories: 4 -hydroxy-phorbol (I), 4-deoxy-4 -phor-
bol (II) and 4-deoxy-4 -phorbol (III). Although their chemical
b-phorbol-20-linoleate.
a
b
a
3
acetyl moiety. In the HMBC spectrum, H-12 proton showed a
correlation with the carbonyl carbon of the tiglyl group ( 167.6),
confirming the location of the tiglyl group at C-12. Thus, the
structure of 1 was determined to be 12-O-tiglylphorbol-4-deoxy-
J
b
1
d
C
structures are very similar, the H NMR is still a powerful method
to distinguish them. For type I skeleton, the chemical shift of H-1is
2
usually in the range of 7.58–7.62 ppm, while the H -5 often appear
4
b-phorbol-13-acetate.
in the region of 2.5 and 2.6 ppm, respectively (Zhang et al., 2013).
For type II and III backbones, the chemical shifts of H-1, H-4 and
Fig. 2. Key 1H– H COSY (—), HMBC (!) and NOESY ($) correlations of 1.
1