Synthesis and antifeedant properties of N-benzoylphenylisoserinates of Lactarius sesquiterpenoid alcohols

Article abstract of DOI:10.1016/S0031-9422(01)00294-1

The esterification of various sesquiterpenoid alcohols of Lactarius origin with N-benzoyl-[2R,3S]-phenylisoserine (side chain of Taxol) produced compounds whose antifeedant properties against storage pests Tribolium confusum, Trogoderma granarium and Sitophylus granarius were measured. The introduction of the taxol side chain in these molecules, in comparison to original compounds, moderately enhanced their antifeedant activities, as well as changed their selectivity of activity towards the test insects.

Full text of DOI:10.1016/S0031-9422(01)00294-1

Phytochemistry 58 (2001) 775–787
www.elsevier.com/locate/phytochem
Synthesis and antifeedant properties of N-benzoylphenylisoserinates
of Lactarius sesquiterpenoid alcohols
Piotr Kopczackia, Maria Gumu•kaa, Marek Masnyka, Halina Grabarczykb,
Gerard Nowakb, W•odzimierz M. Daniewskia,*
aInstitute of Organic Chemistry, Polish Academy of Sciences, Kasprzaka 44/52, 01-224 Warsaw, Poland
bDepartment of Medicinal Plants, Academy of Medicine, Mazowiecka 33, 60-623 Poznan˜, Poland
Received 5 February 2001; received in revised form 18 June 2001
Abstract
The esterification of various sesquiterpenoid alcohols of Lactarius origin with N-benzoyl-[2R,3S]-phenylisoserine (side chain of
Taxol1) produced compounds whose antifeedant properties against storage pests Tribolium confusum, Trogoderma granarium and
Sitophylus granarius were measured. The introduction of the taxol side chain in these molecules, in comparison to original com-
pounds, moderately enhanced their antifeedant activities, as well as changed their selectivity of activity towards the test insects.
# 2001 Elsevier Science Ltd. All rights reserved.
Keywords: N-Benzoylphenylisoserinates; Lactarius sesquiterpenes; Paclitaxel; Antifeedant activity; Storage pests; Tribolium confusum; Trogoderma
granarium; Sitophilus granarius; Rhizopertha dominica
1. Introduction
tylbaccatin III, we decided to attach the paclitaxel side
chain to a series of sesquiterpenoid alcohols to produce
N-benzoylphenylisoserine esters and to investigate their
biological properties.
In our recent paper (Daniewski et al., 1998) we have
identified a group of compounds responsible for the
resistance of Taxus baccata to the attack of insects. It
appeared that the taxanes 10-deacetylbaccatin III (1)
and its 7a isomer (2), contained in the needles of T. bac-
cata and possessing a very strong antifeedant activity,
protect them from the attack of insects (Fig. 1).
This fact prompted us to investigate the antifeedant
activity against the storage pests, of paclitaxel (3), the very
important anticancer compound isolated from Taxus
brevifolia. The antifeedant activity tests of paclitaxel (3)
revealed its very potent activity, and perhaps, its role in
the bark of T. brevifolia in defence against insects. In our
previous papers (Daniewski et al., 1993, 1995) we studied
the antifeedant properties of sesquiterpenoid alcohols
from Lactarius species. The sesquiterpenes, as in the case
of 10-deacetylbaccatin III (1) or paclitaxel (3), are
responsible for chemical defence of the mushrooms
against several predators. Since paclitaxel (3) is the 10-
acetyl-13-N-benzoylphenylisoserine ester of 10-deace-
2. Results and discussion
There are many methods of attachment of N-ben-
zoylphenylisoserine to the baccatin core to produce
Taxol1 (Farina, 1995); among them, a very simple one,
involves the synthesis of the oxazoline derivative (5) of
RS (natural) phenylisoserine (4) (Scheme 1):
Originally the synthesis of oxazoline (5) required as
starting material the phenylisoserine with unnatural
configuration at asymmetric center 2. In order to sim-
plify the procedure and to use the phenylisoserine with
natural configuration, which is commercially available,
we decided to use trimethyl ortho-benzoate for the cycli-
sation, as recently described (Kamata et al., 1998). The
hydrolysis of the ester 5 to the free acid 6 as well as its
esterification with properly protected baccatin III was
described by Kingston et al. (1994) in the semisynthesis of
paclitaxel (3). The same esterification procedure was
applied for the acid 6 and sesquiterpenoid alcohols and it
is exemplified by the following scheme where lactarorufin
A (31) was used as sesquiterpenoid alcohol (Scheme 2).
* Corresponding author. Tel.: +48-22-6323221 x. 2330; fax: +48-
22-6326681.
E-mail address: daniewsk@icho.edu.pl (W.M. Daniewski).
0031-9422/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0031-9422(01)00294-1

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P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
Fig. 1. 1, R=b-OH (10-deacetylbaccatin III); 2, R=a-OH (7-epi-10-deacetylbaccatin III).
The esterification reaction went smoothly to give the
Hz. The geminal methyl groups at C-11 appear as singlets
at ca. ꢀ 0.90–1.1, while the C-12 methyl resonates at lower
field (ca. ꢀ 1.15–1.5). Similarly isolactarane esters (11, 12,
14, 15) exhibited 1H NMR spectra showing three methyl
group singlets, one of them (C-12) resonating at lower
field. The cyclopropane protons at C-5, as in the case of
marasmane sesquiterpenes produced a pair of doublets
(gem. coupling J=5.5 Hz). An important and char-
acteristic 1H NMR spectral feature of isolactarane
esters is involving the ABq system with J=10 Hz, typi-
cal of saturated g-lactone ring, In case of 1H NMR
spectra of lactarane esters, signals of methyl groups
appear as in the previously discussed groups. Depending
on heterocyclic rings (furans or lactones), characteristic
signals are exhibited. The C-13 or C-5 methylene pro-
tons of lactones form an ABq system with a large cou-
pling constant (15–20 Hz). The value of H-8, H-9
cyclic ester 32. The hydrolysis of the cyclic ester to form
the N-benzoylphenylisoserinate (33) required different
conditions from those used for the synthesis of pacli-
taxel. To avoid elimination of the side chain low tem-
perature and methanol as solvent had to be used.
Using the above general method described in the
experimental part a series of N-benzoylphenylisoser-
inates of various Lactarius sesquiterpenoid alcohols was
prepared. (Scheme 2). The series included esters of
marasmane, isolactarane and lactarane sesquiterpenes.
The structures of all the compounds were substantiated
by their 1H NMR and 13C NMR spectra which are
included in the experimental part. The marasmane
esters (8, 9) possess very characteristic signals which
allow their identification. The geminal cyclopropane
protons at C-4 give rise to a pair of doublets with J=4–5
Scheme 1.
Scheme 2.

P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
777
coupling constants are indicative of the cis or trans ste-
reochemistry. As general feature the signals of protons
on carbons bearing the ester groups, in the H NMR
spectra, are down field shifted in comparison with those
of free alcohols (Fig. 2).
The 13C NMR spectra of all the phenylisoserinates
confirmed their structures and are included in the
experimental part.
The phenylisoserinates were prepared from sesqui-
terpenoid alcohols isolated from mushrooms (Lactarius
rufus, Lactarius vellereus, voucher numbers 33550, and
32260 respectively, specimen deposited at the Depart-
ment of Systematics and Geography of Plants of the
University of Warsaw) or synthesized by transformation
of natural products. The references on the procedures of
isolation or preparation of all the sesquiterpenes (7, 10,
13, 16, 19, 22, 25, 28, 31, 34, 37, 40) can be found in the
recent review (Daniewski and Vidari, 1999).
1
3. Antifeedant properties
As it was already mentioned in the introduction, we
wanted to check the antifeedant properties of the syn-
thesised compounds in order to find the influence of
introduction of paclitaxel side chain into their mole-
cules. First we checked paclitaxel (3) (Item 1, R=c,
R0=Ac) that has a very potent antifeedant activity to
the storage pests mentioned in Table 1. Therefore one
can assume that its role in Taxus brevifolia is to protect
the plant against insects. 10-Deacetyl baccatin III (1)
(Item 1, R=H, R0=H), which is a good antifeedant,
demonstrated lower activity than paclitaxel, especially
against Trogoderma granarium and Sitophylus granarius.
The bonding of paclitaxel side chain to the molecule of
isovellerol (Item 2) decreased its activity. Introduction
of paclitaxel side chain into the molecule of iso-
lactarorufin (10) (Item 3, R=H) (which is an attractant)
enhanced its antifeedant properties. Change of stereo-
chemistry at C-8 (Item 4) did not influence activity.
Similarly esterification of furanol (Item 5) did not have
important effect on antifeedant activity. In case of fur-
andiol (19) (Item 6), the esterification gave a compound
21 with twice as high activity. Introduction of the
paclitaxel side chain into ‘‘artifact’’ (Item 7) as well as
into lactarorufin A (Item 8) influenced only the selec-
tivities of antifeedant activity. Esterification of 8-epi-
lactarorufin A (34) (Item 9), which is inactive, caused
some activity. Introduction of the side chain into 5-
deoxy-lactarolid B (Item 10) did not increase its activity
but changed its selectivity towards storage pests. Our
procedure was verified with azadirachtin (Item 11) test.
4.2. Preparation of oxazoline methyl ester (5)
Phenylisoserine (R,S) (2.87 g, 0.013 mol) was sus-
pensed in dry toluene (50 ml) in round bottom flask (200
ml ) equipped with magnetic stirrer and a still head. The
reaction was heated to boiling and 15 ml of toluene was
distilled off. Subsequently trimethylorthobenzoate (5 ml,
0.029 mol) was added and the volatile components were
removed slowly by distillation during 8 h. Residual solids
were filtered off and the filtrate evaporated in rotavap.
The resulting oil was purified by chromatography over
silica-gel in hexane–ethyl acetate gradient solvent system.
Proper fractions were collected and gave pure ester 5 (1.5
g, 41% theoretical yield). [a]D=+14.2ꢀ (EtOH, c 1.0); 1H
NMR (200 MHz): ꢀH 3.90 (3H s), 4.96 (1H, d, J=6.5 Hz,),
5.50 (1H, d, J=6.5 Hz,), 7.30–7.62 (8H, m), 8.10–8.20
(2H,m); 13C NMR (CDCl3, 50 MHz): ꢀC 52.68, 74.59,
83.07, 126.39, 126.72, 127.97, 128.15, 128.22, 128.39,
128.64, 128.79, 131.86, 141.04, 163.89, 170.55.
The unidentified residual solids, by treatment with
another portion of trimethyl ortho benzoate in presence
of p-toluenesulphonic acid in toluene, can be converted
into the desired ester 5. In this way overall yield can be
improved up to 66%.
4.3. (4S,5R) 2,4-Diphenyl-4,5-dihydro-oxazol-5-
carboxylic acid (6)
Compound 5 (1.52 g, 0.0054 mol) dissolved in
methanol (8 ml) was treated with sodium hydroxide
(0.33 g, 0.00835 mol) dissolved in methanol (20 ml) and
water (0.5 ml) at room temperature. The reaction was
followed by TLC and when no starting material was
detected (5 h), hydrochloric acid (6 ml, 5%) was added
to the reaction mixture to obtain the pH value equal 6.
Precipitated solid was filtered, washed with water and
dried to give 1.23 g of the desired product with 86%
4. Experimental
4.1. General
All melting points were measured on a Koffler hot
plate and are uncorrected. H and 13C NMR spectra
yield. mp.=207–211 C, [a]D=À9.97ꢀ (0.05 N NaOH
1
ꢀ
were run at 500 and 125 MHz respectively (Bruker 500
MHz spectrometer) in CDCl3 as otherwise noted, using
TMS as internal standard. Column chromatography
(CC) was carried out using silica gel, and the TLC were
conducted on silica gel or RP-18.
aq., c 1.01),1H NMR (200 MHz, DMSO-d6,): ꢀH 5.00
(1H, d, J=6.4 Hz,), 5.41 (1H d, J=6.4 Hz,), 7.20–7.70
(8H,m), 7.90–8.10 (2H,m); 13C NMR (DMSO-d6, 50
MHz) ꢀC 73.78, 82.42, 126 53, 126.64, 127.79, 128.18,
128.71, 128.80, 132.09, 141.54, 162.86, 171.45.

778
P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
Fig. 2. N-benzoylphenyllsoserinates of Lactarius sesquiterpenoid alcohols and their oxazdine synthatic intermediates.

P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
779
Table 1
Antifeedant activity of 10-deacetyl baccatin III (1) and Taxol1 (3) and N-benzoyl phenylisoserinates of several sesquiterpenoid alcohols of Lactarius
origin
Compound
Item 1
Trogoderma
granarium,
larvae
Sitophilus
granarius,
adults
Tribolium
confusum,
larvae
Rhizopertha
dominica,
adults
Average
activity
classa
R=H
R1=H
(1)
115.5
178.9
185.0
190.8
149.4
122.6
–
150.0 III
156.0 IV
R=b
R1=Ac
(3)
131.5
Item 2
R=H
(7)
200
176
161.9
95.1
–
179.3 IV
123.1 III
R=b
(9)
99.2
165.5
132.7
Item 3
R=H
(10)
À50.4
À7.0
15.3
–
-42.1 atr.
106.9 III
R=b
(12)
70.1
122.4
164.9
70.0
Item 4
R=H
(13)
42.6
92.9
35.3
À22.8
–
18.4 I
R=b
(15)
111.1
67.9
À23.0
62.2 II
Item 5
R=H
(16)
21.0
43.5
137.0
102.3
152.7
103.6
–
103.6 III
97.7 II
R=b
(18)
141.2
Item 6
R=H
(19)
9.2
60.0
95.7
84.1
–
55.0 II
R=b
(21)
78.1
118.5
125.1
101.5 III
Item 7
R=H
(25)
41.6
173.7
115.9
163.8
91.0
–
126.4 III
124.7 III
R=b
(27)
119.6
172.3
(continued on next page)

780
P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
Table 1 (continued)
Compound
Trogoderma
granarium,
larvae
Sitophilus
granarius,
adults
Tribolium
confusum,
larvae
Rhizopertha
dominica,
adults
Average
activity
classa
Item 8
R=H
(31)
148.6
118.9
88.6
144.9
11.7
–
127.4 III
86.2 II
R=b
(33)
162.9
51.2
Item 9
R=H
(34)
À17.4
À11.9
59.3
42.7
–
10.0 I
42.7 I
R=b
(36)
79.9
94.6
-46.6
Item 10
R=H
(40)
93.3
73.4
150.0
66.4
–
105.6 III
113.1 III
R=b
(42)
123.0
129.3
133.8
Item 11 Azadirachtin
190.0
170.0
185.0
–
181.7 IV
a
Below ‘‘0’’, attractant; class I (0–50), poor antifeedant; class II (51–100), medium antifeedant; class III (101–150), good antifeedant; class IV
(151–200), very good antifeedant.
4.4. General method for the synthesis of esters of
(4S,5R)-2,4-diphenyl-4,5-dihydro-oxazol-5-carboxylic
acid and sesquiterpenoid alcohols of Lactarius origin
31.6 (q, C-15), 31.7 (q, C-14), 35.5 (s, C-11), 36.8 (s, C-
3), 37.4 (s, C-6), 38.2 (d, C-2), 42.5 (d, C-9), 44.8 (t, C-
10), 47.5 (t, C-1), 67.8 (t, C-13), 74.5 (d, C-30), 83.0 (d,
C-20), 126.4 (d, C-Ph), 126.8 (s, C-Ph), 127.9 (d, C-Ph),
128.4 (d, C-Ph), 128.7 (d, C-Ph), 128.8 (d, C-Ph), 130.2
(s, C-7), 131.2 (d, C-8), 131.8 (d, C-Ph), 141.1 (s, C-Ph),
164.0 (s, NCO), 169.6 (s, C-10), 200.1 (s, C-5); EIMS 70
eV, m/z (rel. int.): 483 [M]+ (19.3), 269 (17.7), 268
(100.0), 22 (6), 193 (11), 173 (2.4), 165 (2.5), 146 (2), 120
(6), 119 (8), 105 (22), 91 (17), 90 (7), 89 (4), 77 (5), 69
(2), 55 (2), 43 (1) 41 (4); HR EIMS m/z: [M]+ calc. for
C31H33O4N: 483.24096, found: 483.23978.
A sesquiterpenoid alcohol (1 mmol), the acid (6) (1.1
mmol), DMAP (18.3 mg, 0.15 mmol) and DCC (247.2
mg, 1.2 mmol) were dissolved in methylene chloride (25
ml) in a round bottom flask. The reaction was carried out
at room temperature and was monitored by TLC. When
the reaction was completed (3–12 h) the content of the
flask was fitered, the solution evaporated in vacuo and
the residue purified by chromatography to give the
desired esters.
4.4.2. Compound 11
TLC Rf=0.24, (hexane–EtOAc 77:23); mp 170–
4.4.1. Compound 8
20
ꢀ
TLC Rf=0.72 (CH2Cl2–Me2CO 96:4); Oil;
[a]2D0=+2.62ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 204
(" 9664); IR ꢁmax (CHCl3) cmÀ1: 2955, 1753, 1702,1654;
1H NMR: ꢀ 0.97 (3H, s, H-15), 1.00 (3H, s, H-14), 1.23
(3H, s, H-12), 1.25 (1H, d, J10b,10a=12.2 Hz, H-10b), 1.27
(1H, d, J4b,4a=4.4 Hz, H-4b), 1.35 (1H, dd, J1b,1a=13.3
Hz and J1b,2a=7.9 Hz, H-1b), 1.60 (1H, dd, J10a,10b=12.2
Hz and J=7.4 Hz, H-10a), 1.68 (1H, d, J4a,4b=4.4 Hz, H-
4a), 1.77 (1H, dd, J1a,1b=13.3 Hz and J1a,2a=1.1 Hz, H-
1a), 2.46–2.52 (2H, m, H-2a and H-9a), 4.86–4.89 (1H, m,
172 C; [a]D =À43.7ꢀ (CHCl3 c 1.1); UV lmax (EtOH)
nm: 243 (" 20541); IR ꢁmax (KBr) cmÀ1: 3475, 2957,
1
1957, 1895, 1769, 1740, 1656; H NMR: ꢀ 0.95 (3H, s,
H-15), 1.10 (3H, s, H-14), 1.11 (1H, d, J5b,5a=6.2 Hz,
H-5b), 1.36 (1H, t, J10b,10a=11.3 Hz, H-10b), 1.50 (1H,
dd, J1b,1a=12.4 Hz and J1b,2a=7.8 Hz, H-1b), 1.64 (1H,
d, J5a,5b=6.2 Hz, H-5a), 1.68–1.74 (2H, m, H-1a and H-
10a), 2.08–2.19 (2H, m, H-2a and H-9a), 4.16 (1H, d,
J
13b,13a=9.4 Hz, H-13b), 4.41 (1H, d, J13a,13b=9.4 Hz,
H-13a), 4.90 (1H,0 d, J2 ,3 =6.6 Hz, H-20), 5.37 (1H, d,
0
0
H-13b and H-20), 5.28 (1H, d, J13a,13b=12.2 Hz, H-13a),
5.39 (1H, br s, H-8), 5.40 (1H, d, J2 ,3 =6.0 Hz, H-3 ),
7.28–7.55 (8H, m, H-Ph) 8.08–8.11 (2H, m, H-Ph), 9.45
(1H, s, H-5); 13C NMR: ꢀ 19.9 (q, C-12), 30.7 (t, C-4),
J3 ,2 =6.6 Hz, H-3 ), 5.66 (1H, d, J8b,9a=8.0 Hz, H-8b),
7.30–7.65 (8H, m, H-Ph), 8.07–8.12 (2H, m H-Ph); 13C
NMR: ꢀ 18.3 (t, C-5), 24.6 (q, C-12), 26.8 (q, C-15), 28.7
(q, C-14), 31.6 (s, C-11), 38.5 (s, C-6), 38.9 (s, C-7), 39.9
0
0
0
0
0

P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
781
(t, C-1), 40.7 (d, C-2), 45.9 (d, C-9), 47.3 (t, C-10), 71.0
(s, C-3), 71.4 (t, C-13), 75.0 (d, C-8), 79.4 (d, C-30), 83,1
(d, C-20), 126.5 (d, C-Ph), 126.7 (s, C-Ph), 128.2 (d, C-
Ph), 128.5 (d, C-Ph), 128.7 (d, C-Ph), 128.9 (d, C-Ph),
132.1 (d, C-Ph), 140.9 (s, C-Ph), 164.2 (s, NCO), 170.1
(s, C-10), 175.1 (s, C-4); LSIMS m/z (rel. int.): 516
[M+H]+ (38), 498 (15), 460 (4), 307 (29), 268 (29), 231
(6), 222 (6), 154 (100); HR LSIMS m/z: [M+H]+ calc.
for C31H34O6N: 516.23859, found: 516.23813.
Ph), 135.7 (s, C-2), 135.7 (s, C-Ph), 137.8 (d, C-13),
141.1 (s, C-Ph), 141.8 (d, C-5), 164.1 (s, NCO), 170.1 (s,
C-10); EIMS 70 eV, m/z (rel. int.): 481 [M]+ (6), 268
(100), 223 (8), 222 (42), 215 (14), 214 (72), 199 (41), 193
(33), 158 (28), 145 (5), 143 (7), 119 (10), 105 (23), 91
(17), 77 (8); HR EIMS m/z: [M]+ calc. for C31H31O4N:
481.22531, found: 481.22887.
4.4.5. Compound 20
TLC Rf=0.63 (C6H6–Me2CO 8:2); mp 58–63 ꢀC;
[a]2D0=À31.38ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 242
(" 24950); IR ꢁmax (KBr) cmÀ1: 3439, 3063, 2952, 2869,
4.4.3. Compound 14
TLC Rf=0.25 (hexane–EtOAc 8:2); mp 86–91 ꢀC;
[a]2D0=+35.24ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm:
243 (" 16937); IR ꢁmax (CHCl3) cmÀ1: 3555, 3434, 2957,
1
1758, 1655; H NMR: d 1.09 (3H, s, H-15), 1.15 (3H, s,
H-14), 1.29 (3H, s, H-12), 1.58–1.59 (2H, m, H-10), 1.63
(2H, dd, J1a,1b=13.5 Hz and J=5.9 Hz, H-1), 2.35–2.41
(1H, m, H-2a), 2.49–2.55 (1H, m, 9a), 2.56 (1H, d,
1
1772, 1746,1655; H NMR: ꢀ 0.67 (3H, s, H-15), 0.85
(3H, s, H-14), 0.95 (1H, t, J10b,10a=11.8 Hz, H-10b),
1.18 (1H, d, J5b,5a=6.3 Hz, H-5b), 1.30–1.42 (3H, m, H-
1a, H-1b and H-10a), 1.60 (3H, s, H-12), 1.61 (1H, d,
J
4b,4a=15.0 Hz, H-4b), 2.82 (1H, d, J4a,4b=15.0 Hz, H-
0
0
0
4a), 5.00 (1H, d, J2 ,3 =6.2 Hz, H-2 ), 5.52 (1H, d,
0
0
0
J
5a,5b=6.3 Hz, H-5a), 1.99–2.05 (1H, m, H-2a), 2.25-
J3 ,2 =6.2 Hz, H-3 ), 6.06 (1H, d, J8b,9a=10.8 Hz, H-
8b), 7.13 (1H, br s, H-5), 7.24 (1H, br s, H-13), 7.28–7.57
(8H, m, H-Ph), 8.08–8.13 (2H, m, H-Ph); 13C NMR: ꢀ
30.6 (q, C-12), 31.8 (q, C-14), 32.1 (q, C-15), 32.5 (t, C-
4), 35.3 (s, C-11), 42.6 (t, C-1), 43.6 (t, C-10), 44.8 (d, C-
9), 53.9 (d, C-2), 70.7 (d, C-8), 71.3 (s, C-3), 74.7 (d, C-
30), 83,1 (d, C-20), 118.1 (s, C-6), 125.1 (s, C-7), 126.4 (d,
C-Ph), 126.6 (s, C-Ph), 128.1 (d, C-Ph), 128.5 (d, C-Ph),
128.7 (d, C-Ph), 128.9 (d, C-Ph), 132.0 (d, C-Ph), 138.9
(d, C-5), 140.9 (s, C-Ph), 141.9 (d, C-13), 164.1 (s,
NCO), 169.1 (s, C-10); EIMS 70 eV, m/z (rel. int.): 499
[M]+ (3), 481 (6), 464 (7), 446 (5), 268 (16), 232 (32), 223
(16), 222 (100), 217 (14), 199 (7), 193 (58), 189 (13), 165
(6), 145 (2), 137 (3), 119 (4), 105 (15), 91 (9), 77 (6), 43
(4); HR LSIMS m/z: [M+H]+ calc. for C31H34O5N:
500.24371, found: 500.24270.
2.32 (1H, m, H-9a), 4.08 (1H, d, J13b,13a=9.4 Hz, H-
13b), 4.21 (1H, d, J13a,13b=9.4 Hz, H-13a), 5.02 (1H, 0d,
0
0
0
0
0
J2 ,3 =5.9 Hz, H-2 ), 5.56 (1H, d, J3 ,2 =5.9 Hz, H-3 ),
5.83 (1H, d, J8a,9a=4.5 Hz, H-8a), 7.30–7.56 (8H, m, H-
Ph), 8.11–8.13 (2H, m, H-Ph); 13C NMR: ꢀ 19.5 (t, C-5),
24.7 (q, C-12), 26.5 (q, C-14), 28.1 (q, C-15), 32.0 (s, C-11),
35.2 (s, C-6), 37.6 (d, C-9), 37.6 (s, C-7), 39.6 (t, C-1), 42.1
(t, C-10), 44.3 (d, C-2), 68.0 (t, C-13), 69.3 (s, C-3), 71.2 (d,
C-8), 74.7 (d, C-30), 82.9 (d, C-20), 125.9 (s, C-Ph), 126.4
(d, C-Ph), 128.2 (d, C-Ph), 128.5 (d, C-Ph), 128.9 (d, C-
Ph), 129.0 (d, C-Ph), 132.2 (d, C-Ph), 140.6 (s, C-Ph),
163.4 (s, NCO), 170.1 (s, C-10), 173.6 (s, C-4); LSIMS
m/z (rel. int.): 516 [M+H]+ (100), 498 (26), 474 (2), 403
(2), 307 (12), 289 (7), 268 (100), 225 (52), 193 (8), 154
(92); HR LSIMS m/z: [M+H]+ calc. for C31H34O6N:
516.23861, found: 516.23963.
4.4.6. Compound 23
4.4.4. Compound 17
TLC Rf=0.62 (hexane–EtOAc 8:2); mp 127–131 C;
TLC Rf=0.27, (hexane–EtOAc 7:3); mp 64–68 ꢀC;
[a]2D0=À21.2ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 242 ("
16768); IR ꢁmax (CHCl3) cmÀ1: 3066, 2955, 2867, 1960,
1748, 1654; 1H NMR: ꢀ 0.95 (3H, s, H-15), 1.00 (3H, s, H-
14), 1.05 (3H, s, H-12), 1.41 (1H, dd, J10a,10b=12.6 Hz and
ꢀ
[a]2D0=À57.83ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 241
(" 15984); IR ꢁmax (CHCl3) cmÀ1: 3066, 2955, 2867, 1960,
1748, 1654; 1H NMR: ꢀ 0.90 (3H, s, H-15), 1.12 (3H, s, H-
14), 1.46 (1H, dd, J10b,10a=12.6 Hz, J10b,9a=8.3 Hz, H-
10b), 1.75 (3H, s, H-12), 1.91 (1H, ddd, J10a,10b=12.6 Hz,
J
10a,9b=10.6 Hz, H-10a), 1.50 (1H, dd, J1b,1a=12.9 Hz
and J1b,2a=10.8 Hz, H-1b), 1.70 (1H, dd, J10b,10a=12.6
Hz and 10b,9b=7.0 Hz, H-10b), 1.80 (1H, dd,
1a,1b=12.9 Hz and J1a,2a=8.1 Hz, H-1a), 2.02–2.10
(1H, m, H-9b), 2.13–2.19 (1H, m, H-2a), 2.59 (1H, d,
J
J
10a,9a=7.9 Hz and J=2.0 Hz, H-10a), 2.07 (1H, d,
1b,1a=15.0 Hz, H-1b), 2.20 (1H, d, J1a,1b=15.0 Hz, H-
J
J
1a), 2.91 (1H, d, J4b,4a=16.1 Hz, H-4b), 3.25–3.31 (1H,
m, H-9a), 3.40 (1H, d, J4a,4b=16.1 Hz, H-4a), 4.94 (1H,
J
4b,4a=14.1 Hz, H-4b), 2.69 (1H, d, J4a,4b=14.1 Hz, H-
0
0
0
0
0
0
0
0
0
d, J2 ,3 =6.3 Hz, H-2 ), 5.47 (1H, d, J3 ,2 =6.3 Hz, H-3 ),
5.82 (1H, d, J8b,9a=11.4 Hz, H-8b), 7.13 (1H, t, J=1.5
Hz, H-5a), 7.28–7.55 (9H, m, H-13 and H-Ph), 8.07–
8.12 (2H, m, H-Ph); 13C NMR: ꢀ 21.5 (q, C-12), 27.5 (q,
C-14), 28.7 (q, C-15), 29.4 (t, C-4), 37.6 (s, C-11), 44.6
(d, C-9), 45.8 (t, C-10), 46.5 (t, C-1), 73.6 (d, C-8), 74.7
(d, C-30), 83,2 (d, C-20), 121.3 (s, C-6), 124.2 (s, C-7),
126.4 (d, C-Ph), 128.0 (d, C-Ph), 128.4 (d, C-Ph), 128.7
(d, C-Ph), 128.8 (d, C-Ph), 130.0 (s, C-3), 131.9 (d, C-
4a), 4.98 (1H, d, J2 ,3 =6.4 Hz, H-2 ), 5.49 (1H, d,
0
0
0
J3 ,2 =6.4 Hz, H-3 ), 5.86 (1H, d, J8a,9b=10.3 Hz, H-
8a), 7.16 (1H, s, H-5), 7.20 (1H, s, H-13), 7.30–7.57 (8H,
m, H-Ph), 8.09–8.14 (2H, m, H-Ph); 13C NMR: ꢀ 20.7
(q, C-12), 31.0 (q, C-15), 31.2 (q, C-14), 34.7 (s, C-11),
40.1 (t, C-4), 43.2 (t, C-1), 46.1 (t, C-10), 46.5 (d, C-9),
55.1 (d, C-2), 73.3 (s, C-3), 74.8 (d, C-30), 75.7 (d, C-8),
83.3 (d, C-20), 119.1 (s, C-6), 124.9 (s, C-7), 126.5 (d, C-
Ph), 126.7 (s, C-Ph), 128.2 (d, C-Ph), 128.5 (d, C-Ph),

782
P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
128.7 (d, C-Ph), 128.9 (d, C-Ph), 132.0 (d, C-Ph), 138.2
(d, C-5), 140.8 (s, C-Ph), 140.9 (d, C-13), 164.0 (s,
NCO), 169.3 (s, C-10); EIMS 70 eV, m/z (rel. int.): 499
[M]+ (1.5), 269 (17), 268 (98), 266 (20), 234 (51), 222
(40), 217 (10), 194 (12), 193 (71), 189 (28),165 (19), 145
(7), 143 (4), 137 (10), 119 (35), 105 (56), 91 (100), 77
(40), 43 (97); HR EIMS m/z: [M]+ calc. for C31H33O5N:
499.23587, found: 499.23743.
10), 69.6 (t, C-13), 73.3 (d, C-8), 74.8 (d, C-30), 82.8 (d,
C-20), 126.4 (s, C-2), 126.4 (s, C-3), 127.0 (s, C-6), 128.3
(d, C-Ph), 128.6 (d, C-Ph), 128.7 (d, C-Ph), 128.8 (d, C-
Ph), 129.0 (d, C-Ph), 132.2 (d, C-Ph), 135.5 (s, C-Ph),
140.6 (s, C-Ph), 155.9 (s, C-7), 163.9 (s, NCO), 169.8 (s,
C-5), 173.6 (s, C-10); LSIMS m/z (rel. int.): 498
[M+H]+ (60), 460 (2), 313 (2), 307 (23), 289 (11), 268
(67), 231 (6), 229 (6), 166 (6), 154 (100), 137 (76), 120
(18), 105 (26); HR LSIMS m/z: [M+H]+ calc. for
C31H32O5N: 498.22806, found: 498.23098.
4.4.7. Compound 26
TLC Rf=0.46 (hexane–EtOAc 8:2); mp 50–53 ꢀC;
[a]2D0=À46.43ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 242
(" 10297); IR ꢁmax (CHCl3) cmÀ1: 2955, 2872, 1755,
1654; 1H NMR: ꢀ 0.90 (3H, t, JCH CH =6.9 Hz,
4.4.9. Compound 32
TLC Rf=0.41 (C6H6-Me2CO 8:2); mp 83–86 ꢀC;
[a]2D0=À20.45ꢀ (CHCl3, c 1.0,); IR ꢁmax (CHCl3) cmÀ1
:
2
3
1
ÀOCH2CH3), 1.08 (3H, s, H-15), 1.16 (3H, s, H-14), 1.20
(3H, s, H-12), 1.57–1.61 (3H, m, H-1a, H-10a and H-10b),
1.69 (1H, t, J1b,1a=12.8 Hz, H-1b), 2.37-2.42 (1H, m, H-
2a), 2.58 (1H, dd, J4b,4a=15.2 Hz and J=1.2 Hz, H-4b),
2.59–2.64 (1H, m, H-9a), 2.75 (1H, d, J4a,4b=15.2 Hz, H-
4a), 3.28 (1H, qn, JCH CH =6.9 Hz, ÀOCH2CH3), 3.36
3592, 3507, 2958, 2871, 1756, 1655; H NMR: d 1.05
(3H, s, H-15), 1.14 (3H, s, H-14), 1.28 (3H, s, H-12),
1.39 (1H, dd, J10b,10a=13.3 Hz and J10b,9a=7.1 Hz, H-
10b), 1.55 (1H, t, J=12.9 Hz, H-1b), 1.66 (1H, dd,
J
10a,10b=13.3 Hz and J10a,9a=6.0 Hz, H-10a), 1.70 (1H,
dd, J1a,1b=12.9 Hz and J1a,2a=6.9 Hz, H-1a), 2.49-2.55
(1H, m, H-2a), 2.62 (2H, ABq, J=17.3 Hz, H-4a and H-
4b), 2.73-2.80 (1H, m, H-9a), 4.56 (1H, d, J13b,13a=17.3
2
3
(1H, qn, JCH CH =6.9 Hz, ÀOCH2CH3), 5.00 (1H, d,
2
3
0
0
0
0
0
0
J2 ,3 =6.4 Hz, H-2 ), 5.54 (1H, d, J3 ,2 =6.4 Hz, H-3 ),
6.06 (1H, d, J8b,9a=11.4 Hz, H-8b), 7.05 (1H, s, H-5), 7.13
(1H, t, J=1.3 Hz, H-13), 7.29–7.57 (8H, m, H-Ph), 8.09–
8.13 (2H, m, H-Ph); 13C NMR: ꢀ 15.7 (q, ÀCH2CH3), 25.5
(q, C-12), 28.9 (t, C-4), 32.2 (q, C-14), 32.7 (q, C-15),
34.9 (s, C-11), 41.7 (t, C-1), 43.5 (t, C-10), 44.5 (d, C-9),
52.5 (d, C-2), 55.6 (t, -CH2CH3), 75.4 (s, C-3), 74.8 (d,
C-30), 77.2 (d, C-8), 83.2 (d, C-20), 119.3 (s, C-6), 125.6
(s, C-7), 126.4 (d, C-Ph), 126.7 (s, C-Ph), 128.0 (d, C-
Ph), 128.4 (d, C-Ph), 128.7 (d, C-Ph), 128.9 (d, C-Ph),
131.9 (d, C-Ph), 137.7 (d, C-5), 140.3 (d, C-13), 141.1 (s,
C-Ph), 164.1 (s, NCO), 169.3 (s, C-10); EIMS 70 eV, m/z
(rel. int.): 527 [M]+ (4), 512 [M-CH3]+ (2), 498 [M-
C2H5]+ (2), 269 (18), 268 (100), 260 (12), 223 (11), 222
(40), 215 (20), 193 (32), 165 (6), 137 (6), 105 (20), 91 (26)
77 (11), 43 (16); HR EIMS m/z: [MÀC2H5]+ calc. for
C31H32O5N: 498.22805, found: 498.22811.
Hz, H-13b), 4.70 (1H, d, J13a,13b=17.3 Hz, H-13a), 4.96
0
0
0
0
0
(1H, d, J2 ,3 =6.2 Hz, H-2 ), 5.47 (1H, d, J3 ,2 =6.2 Hz,
H-30), 6.00 (1H, d, J8b,9a=10.3 Hz, H-8b), 7.30–7.57
(8H, m, H-Ph), 8.06–8.10 (2H, m, H-Ph); 13C NMR: ꢀ
28.6 (q, C-14), 29.5 (q, C-12), 30.7 (q, C-15), 35.8 (t, C-
4), 35.9 (s, C-11), 42.6 (d, C-9), 42.7 (t, C-10), 47.7 (t, C-
1), 52.1 (d, C-2), 69.5 (t, C-13), 71.5 (s, C-3), 72.5 (d, C-
8), 74.7 (d, C-30), 82.8 (d, C-20), 125.7 (s, C-6), 126.3 (s,
C-Ph), 126.4 (d, C-Ph), 128.3 (d, C-Ph), 128.6 (d, C-Ph),
128.7 (d, C-Ph), 129.0 (d, C-Ph), 132.2 (d, C-Ph), 140.5
(s, C-Ph), 157.1 (s, C-7), 163.9 (s, NCO), 169.5 (s, C-5),
173.9 (s, C-10); LSIMS m/z (rel. int.): 516 [M+H]+ (60),
498 (4), 307 (14), 289 (8), 268 (77), 225 (15), 154 (100),
136 (77), 105 (44); HR LSIMS m/z: [M+H]+ calc. for
C31H34O6N: 516.23859, found: 516.23648.
4.4.10. Compound 35
4.4.8. Compound 29
TLC Rf=0.28 (hexane–EtOAc 6:4); mp 83–87 ꢀC;
[a]2D0=À66.59ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 194
(" 52202); IR ꢁmax (CHCl3) cmÀ1: 3593, 3030, 2958,
TLC Rf=0.57 (hexane–EtOAc 7:3); mp 62–64 ꢀC;
[a]2D0=À47.90ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 196
(" 76709); 242 (" 20511); IR ꢁmax (KBr) cmÀ1: 3448,
1
2868, 1751, 1655; H NMR: ꢀ 0.91 (3H, s, H-15), 0.97
1
2952, 2866, 1758, 1655; H NMR: ꢀ 0.92 (3H, s, H-15),
1.13 (3H, s, H-14), 1.44 (1H, dd, J10b,10a=12.5 Hz and
(3H, s, H-14), 1.04 (1H, t, J1b,1a=12.7 Hz, H-1b), 1.12
(1H, t, J=12.2 Hz, H-10b), 1.28 (3H, s, H-12), 1.43 (1H,
dd, J10a,10b=12.2 Hz and J10a,9a=7.0 Hz, H-10a), 1.55
(1H, dd, J1a,1b=12.7 Hz and J1a,2a=7.9 Hz, H-1a), 2.47
(1H, d, J4b,4a=19.2 Hz, H-4b), 2.57–2.63 (1H, m, H-2a),
2.64 (1H, dq, J4a,4b=19.2 Hz and J=3.0 Hz, H-4a),
2.93–3.01 (1H, m, H-9a),0 4.65 (2H, br s, H-13), 4.93
J
10b,9a=9.0 Hz, H-10b), 1.86 (1H, dd, J10a,10b=12.5 Hz
and J10a,9a=4.7 Hz, H-10a), 1.77 (3H, s, H-12), 2.13
(1H, d, J1b,1a=15.8 Hz, H-1b), 2.28 (1H, d, J1a,1b=15.8
Hz, H-1a), 2.97 (1H, d, J4b,4a=20.3 Hz, H-4b), 3.18
(1H, d, J4a,4b=20.3 Hz, H-4a), 3.15-3.28 (1H, m, H-9a),
4.64 (2H, ABq, JAB=17.00Hz, H-13a and H-13b), 4.96
0
0
0
0
(1H, d, J2 ,3 =6.0 Hz, H-2 ), 5.39 (1H, d, J3 ,2 =6.0 Hz,
H-30) 6.96 (1H, br s, H-8a), 7.30–7.57 (8H, m, H-Ph),
8.06–8.09 (2H, m, H-Ph); 13C NMR: ꢀ 26.2 (q, C-15),
28.9 (q, C-14), 32.4 (q, C-12), 34.9 (t, C-4), 37.7 (s, C-
11), 43.9 (d, C-9), 42.2 (t, C-10), 44.6 (t, C-1), 49.1 (d, C-
2), 70.4 (t, C-13), 73.8 (s, C-3), 74.0 (d, C-8), 75.0 (d, C-
0
0
0
0
(1H, d, J2 ,3 =6.1 Hz, H-2 ), 5.47 (1H, d, J3 ,2 =6.1 Hz,
H-30), 5.69 (1H, d, J8b,9a=10.9 Hz, H-8b), 7.31–7.58
(8H, m, H-Ph), 8.08–8.11 (2H, m, H-Ph); 13C NMR: ꢀ
22.6 (q, C-12), 27.1 (q, C-15), 28.5 (q, C-14), 30.3 (t, C-
4), 37.2 (s, C-11), 45.0 (d, C-9), 45.4 (t, C-1), 46.5 (t, C-

P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
783
30), 83.1 (d, C-20), 122.6 (s, C-6), 126.4 (d, C-Ph), 126.4
(s, C-Ph), 128.3 (d, C-Ph), 128.6 (d, C-Ph), 128.6 (d, C-
Ph), 129.0 (d, C-Ph), 132.2 (d, C-Ph), 140.4 (s, C-Ph),
157.9 (s, C-7), 163.9 (s, NCO), 174.3 (s, C-5), 174.3 (s,
C-10); LSIMS m/z (rel. int.): 516 [M+H]+ (24), 369 (3),
307 (16), 268 (25), 222 (4), 154 (100), 137 (76); HR
LSIMS m/z: [M+H]+ calc. for C31H34O6N: 516.23861,
found: 516.23900.
45.2 (t, C-1), 50.6 (d, C-2), 73.1 (s, C-3), 69.6 (d, C-8),
71.9 (t, C-5), 74.3 (d, C-30), 83.1 (d, C-20), 122.9 (s, C-7),
126.6 (d, C-Ph), 126.6 (s, C-Ph), 128.0 (d, C-Ph), 128.3
(d, C-Ph), 128.7 (d, C-Ph), 128.8 (d, C-Ph), 131.9 (d, C-
Ph), 140.7 (s, C-Ph), 151.9 (s, C-6), 163.9 (s, NCO),
168.4 (s, C-13), 172.5 (s, C-10); LSIMS m/z (rel. int.):
516 [M+H]+ (58), 403 (1), 379 (1), 359 (1), 337 (1), 307
(3), 268 (85), 249 (11), 222 (12), 154 (42), 136 (38), 105
(73), 81 (100); HR LSIMS m/z: [M+H]+ calc. for
C31H34O6N: 516.23861, found: 516.23737.
4.4.11. Compound 38
TLC Rf=0.26 (hexane–Me2CO 7:3); mp 90–92 ꢀC;
[a]2D0=-41.2ꢀ (CHCl3, c 1.0); UV no peaks found; IR
4.5. General method for the hydrolysis of esters of
(4S,5R)-2,4-diphenyl-4,5-dihydro-oxazol-5-carboxylic
acid and sesquiterpenoid alcohols of Lactarius origin into
(2S,3R)-phenyl-isoserinates
ꢁ
max (KBr) cmÀ1: 3448, 3064, 2953, 2866, 1758, 1655; 1H
NMR: ꢀ 0.96 (3H, s, H-15), 1.06 (3H, s, H-14), 1.09 (3H,
s, H-12), 1.38 (1H, dd, 10a,10b=12.6 Hz and
10a,9b=10.7 Hz, H-10a), 1.53 (1H, dd, J1b,1a=13.3 Hz
and J1b,2a=10.5 Hz, H-1b), 1.66 (1H, dd, J10b,10a=12.6
Hz and 10b,9b=6.9 Hz, H-10b), 1.86 (1H, dd,
1a,1b=13.3 Hz and J1a,2a=8.3 Hz, H-1a), 2.13–2.21
(1H, m, H-9), 2.24–2.31 (1H, m, H-2a), 2.36 (1H, dq,
J
J
A sesquiterpenoid alcohol (4S,5R)-2,4-diphenyl-oxa-
zol-5-carboxylate (0.2 mmol) dissolved in methanol (25
J
ꢀ
J
ml) was chilled to 0 C and treated with 0.5 N aq. HCl
solution (2.5 ml). The reaction must be carried out at
ꢀ
J
J
4b,4a=15.1 Hz and J=3.0 Hz, H-4b), 2.81 (1H, d,
4a,4b=15.1 Hz, H-4a), 4.63 (1H, ddd, J13b,13a=17.6 Hz,
0 C. After completion of the reaction the solution was
neutralised with saturated aq. sodium hydrogen car-
bonate solution (pH=7) and left for 15 min. Subse-
quently the solution was diluted with water (75 ml) and
extracted with chloroform (3Â40 ml). The chloroform
extract was dried over magnesium sulphate, filtered and
evaporated to dryness leaving a residue, which was
purified by chromatography in proper solvent system
and gave the desired N-benzoylphenylisoserinate.
J=3.1 Hz and J=1.3 Hz, H-13b), 4.75 (1H, dq,
13a,13b=17.6 Hz and J=1.6 Hz, H-13a), 4.98 (1H, d,
J
0
0
0
0
0
0
J2 ,3 =6.2 Hz, H-2 ), 5.44 (1H, d, J3 ,2 =6.2 Hz, H-3 ),
5.89 (1H, dd, J8a,9b=10.7 Hz and J=1.9 Hz, H-8a),
7.31–7.59 (8H, m, H-Ph), 8.08–8.10 (2H, m, H-Ph); 13C
NMR: ꢀ 20.7 (q, C-12), 30.8 (q, C-15), 31.2 (q, C-14),
34.3 (s, C-11), 39.5 (t, C-4), 43.4 (t, C-10), 44.0 (d, C-9),
46.2 (t, C-1), 54.4 (d, C-2), 68.9 (t, C-13), 72.1 (s, C-3),
75.0 (d, C-30), 76.7 (d, C-8), 83.0 (d, C-20), 124.5 (s, C-6),
126.3 (s, C-Ph), 126.4 (d, C-Ph), 128.4 (d, C-Ph), 128.6
(d, C-Ph), 128.6 (d, C-Ph), 129.0 (d, C-Ph), 132.2 (d, C-
Ph), 140.3 (s, C-Ph), 159.1 (s, C-7), 163.8 (s, NCO),
169.4 (s, C-10), 173.6 (s, C-5); LSIMS m/z (rel. int.):
1031 [2M+H]+ (3), 516 [M+H]+ (95), 499 (18), 307
(11), 268 (100), 222 (18), 193 (5), 154 (77), 137 (53), 105
(45); HR LSIMS m/z: [M+H]+ calc. for C31H34O6N:
516.23861, found: 516.23779.
4.5.1. Compound 9
TLC Rf=0.21 (hexane–EtOAc 7:3); mp 60–63 ꢀC;
[a]2D0=+15.83ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm:
226 (" 17509); IR ꢁmax (CHCl3) cmÀ1: 3683, 3519, 3440,
2955, 2866, 1731, 1702, 1664, 1511; 1H NMR: ꢀ 0.95 (3H,
s, H-15), 0.98 (3H, s, H-14), 1.23 (1H, t, J10b,10a=13.0 Hz,
H-10b), 1.26 (3H, s, H-12), 1.29 (1H, d, J4b,4a=4.5 Hz, H-
4b), 1.30 (1H, dd, J1a,1b=13.3 Hz and J1a,2a=1.8 Hz, H-
1a), 1.59 (1H, dd, J10a,10b=13.0 Hz and J10a,9a=7.6 Hz,
H-10a), 1.72 (1H, dd, J1b,1a=13.3 Hz and J1b,2a=8.0 Hz,
H-1b), 1.80 (1H, d, J4a,4b=4.5 Hz, H-4a), 2.42–2.50 (2H,
4.4.12. Compound 41
TLC Rf=0.26 (hexane–EtOAc 4:6); mp 76–80 ꢀC;
[a]2D0=À36.29ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 194
(" 57717); IR ꢁmax (CHCl3) cmÀ1: 3566, 2958, 2869,
m, H-2a and H-9a), 4.58 (1H, d, J2 ,3 =2.2 Hz, H-20),
4.84 (1H, d, J13b,13a=12.1 Hz, H-13b), 5.29 (1H, d,
0
0
J
(1H, dd, J3 ,NH=8.9 Hz and J3 ,2 =2.2 Hz, H-3 ), 7.05
13a,13b=12.1 Hz, H-13a), 5.33 (1H, br s, H-8), 5.63
1
0
0
0
0
1764, 1654; H NMR: ꢀ 0.99 (3H, s, H-15), 1.06 (3H, s,
0
H-14), 1.16 (1H, t, J10b,10a=11.9 Hz, H-10b), 1.23 (3H,
s, H-12), 1.25 (1H, t, J1b,1a=7.2 Hz, H-1b), 1.62 (1H, t,
(1H, d, JNH,3 =8.9 Hz, NH), 7.26–7.52 (8H, m, H-Ph),
7.74–7.77 (2H, m, H-Ph), 9.40 (1H, s, H-5); 13C NMR: ꢀ
20.0 (q, C-12), 31.0 (t, C-4), 31.6 (q, C-14), 31.7 (q, C-
15), 35.5 (s, C-11), 36.9 (s, C-3), 37.3 (s, C-6), 38.3 (d, C-
2), 42.5 (d, C-9), 44.8 (t, C-10), 47.5 (t, C-1), 54.5 (d, C-
30), 68.5 (t, C-13), 73.2 (d, C-20), 126.9 (d, C-Ph), 127.0
(s, C-Ph), 127.9 (d, C-Ph), 128.5 (d, C-Ph), 128.6 (d, C-
Ph), 129.9 (s, C-7), 131.7 (d, C-Ph), 131.8 (s, C-8), 134.1
(s, C-Ph), 138.8 (s, C-Ph), 166.7 (s, NCO), 172.6 (s, C-
10), 200.6 (s, C-5); LSIMS m/z (rel. int.): 524 [M+Na]+
(9), 484 (4), 446 (12), 330 (5), 308 (28), 286 (14), 231
J
10a,10b=11.9 Hz, H-10a), 1.67–1.74 (1H, m, H-1a),
2.56 (1H, d, J4b,4a=18.8 Hz, H-4b), 2.62–2.69 (1H, m,
H-2a), 2.80 (1H, d, J4a,4b=18.8 Hz, H-4a), 2.85–2.91
(1H, m, H-9a), 4.61 (2H, ABq, JAB=17.0 Hz, H-5a and
0
0
0
H-5b), 4.97 (1H, d, J2 ,3 =6.5 Hz, H-2 ), 5.50 (1H, d,
0
0
0
J3 ,2 =6.5 Hz, H-3 ), 5.96 (1H, d, J8b,9a=5.2 Hz, H-8b),
7.26–7.55 (8H, m, H-Ph), 8.08–8.11 (2H, m, H-Ph); 13C
NMR: ꢀ 26.9 (q, C-15), 29.4 (q, C-14), 29.6 (q, C-12),
36.5 (s, C-11), 38.6 (t, C-4), 42.5 (d, C-9), 45.1 (t, C-10),

784
P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
(28), 215 (16), 154 (20), 111 (21), 105 (100); HR LSIMS
m/z: [M+Na]+ calcd for C31H33O5NNa: 524.24129,
found: 524.24127.
(100); HR LSIMS m/z: [M+Na]+ calc. for C31H35
O7NNa: 556.23114, found: 556.23175.
4.5.4. Compound 18
TLC Rf=0.23 (hexane–EtOAc 8:2); mp 68–70 ꢀC;
[a]2D0=À4.55ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: no
4.5.2. Compound 12
TLC Rf=0.21 (hexane–EtOAc 1:1); mp 104–108 C;
ꢀ
[a]2D0=À9.82ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 223
peaks found above threshold; IR nmax (CHCl3) cmÀ1
:
1
(" 16399); IR nmax (KBr) cmÀ1: 3431, 2953, 2865, 1768,
3423, 2952, 2864, 1736, 1645; H NMR: ꢀ 0.78 (6H, s,
H-14 and H-15), 1.16 (1H, dd, J10b,10a=12.9 Hz and
1
1745,1651; H NMR: ꢀ 0.68 (3H, s, H-15), 0.80 (3H, s,
H-14), 1.07–1.12 (2H, m, H-5b and H-10b), 1.36–1.45
(2H, m, H-1b and H-10a), 1.56 (1H, dd, J=11.9 Hz and
J=2.5 Hz, H-1a), 1.58 (3H, s, H-12), 1.69 (1H, d,
J10b,9a=8.1 Hz, H-10b), 1.73 (3H, s, H-12), 1.83 (1H,
dd, J10a,10b=12.9 Hz and J10a,9a=7.9 Hz, H-10a), 1.94
(1H, d, J1b,1a=15.0 Hz, H-1b), 2.11 (1H, d, J1a,1b=15.0
Hz, H-1a), 2.89 (1H, d, J4b,4a=15.9 Hz, H-4b), 3.25–
J
5a,5b=6.2 Hz, H-5a), 2.03–2.12 (2H, m, H-2a and H-
9a), 4.10 (1H, d, J13b,13a=9.3 Hz, H-13b), 4.23 (1H, d,
3.31 (1H, m, H-9a), 3.42 (1H, d, J4a,4b=15.9 Hz, H-4a),
0
0
0
0
0
0
J
13a,13b=9.3 Hz, H-13a), 4.65 (1H, d, J2 ,3 =2.0 Hz, H-
4.60 (1H, d, J2 ,3 =2.0 Hz, H-2 ), 5.73 (1H, dd, J3 ,NH
0
=
20), 5.67 (1H, d, J8b,9a=8.0 Hz, H-8b), 5.77 (1H, dd,
8.8 Hz and J3 ,2 =2.0 Hz, H-30), 5.81 (1H, d,
0
0
0
0
0
J3 ,NH=9.1 Hz, J3 ,2 =2.0 Hz, H-3 ), 7.15 (1H, d,
0
J
8b,9a=11.1 Hz, H-8b), 7.12 (1H, d, JNH,3 =8.8 Hz,
0
JNH,3 =9.1, NH), 7.25–7.53 (8H, m, H-Ph), 7.76–7.80
NH), 7.13 (1H, br s, H-5), 7.23 (1H, s, H-13), 7.26–7.53
(8H, m, H-Ph), 7.79–7.81 (2H, m, H-Ph); 13C NMR: ꢀ
21.5 (q, C-12), 27.2 (q, C-14), 28.2 (q, C-15), 29.3 (t, C-
4), 37.5 (s, C-11), 44.4 (d, C-9), 45.8 (t, C-10), 45.8 (t, C-1),
54.7 (d, C-30), 73.8 (d, C-8), 74.9 (d, C-20), 121.6 (s, C-6),
124.8 (s, C-7), 126.8 (d, C-Ph), 127.1 (d, C-Ph), 127.8 (d, C-
Ph), 128.5 (d, C-Ph), 128.6 (d, C-Ph), 130.1 (s, C-3), 131.7
(d, C-Ph), 134.0 (s, C-2), 136.0 (s, C-Ph), 137.8 (d, C-5),
138.9 (s, C-13), 141.8 (d, C-13), 166.4 (s, NCO), 172.8 (s,
C-10); LSIMS m/z (rel. int.): 522 [M+Na]+ (14), 338 (5),
329 (26), 308 (43), 286 (48), 258 (4), 240 (4), 215 (44), 199
(6), 187 (4), 176 (100), 154 (78), 136 (56), 120 (12), 105 (37);
HR LSIMS m/z: [M+Na]+ calc. for C31H33O5NNa:
522.22565, found: 522.22465.
(2H, m, H-Ph); 13C NMR: ꢀ 18.4 (t, C-5), 24.4 (q, C-12),
26.2 (q, C-15), 28.4 (q, C-14), 31.3 (s, C-11), 38.1 (s, C-
6), 38.7 (s, C-7), 39.9 (t, C-1), 40.0 (d, C-2), 46.1 (d, C-9),
46.6 (t, C-10), 54.6 (d, C-30), 71.0 (s, C-3), 71.2 (t, C-13),
73.5 (d, C-8), 80,2 (d, C-20), 126.9 (d, C-Ph), 127.1 (d, C-
Ph), 127.8 (d, C-Ph), 128.6 (d, C-Ph), 128.6 (d, C-Ph),
131.9 (d, C-Ph), 133.7 (s, C-Ph), 138.6 (s, C-Ph), 166.4
(s, NCO), 172.6 (s, C-10), 175.1 (s, C-4); LSIMS m/z (rel.
int.): 534 [M+H]+ (4), 516 (3), 307 (8), 286 (100), 249
(3), 231 (12), 210 (20), 176 (2), 154 (56), 137 (46), 123
(24), 105 (60); HR LSIMS m/z: [M+Na]+ calc. for
C31H35O7NNa: 556.23114, found: 556.23308.
4.5.3. Compound 15
TLC Rf=0.25 (hexane–EtOAc 55:45); mp 176–183 ꢀC;
[a]2D0=+49.6ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 228
(" 14420); IR ꢁmax (CHCl3) cmÀ1: 3555, 3436, 2935,
4.5.5. Compound 21
TLC Rf=0.31 (C6H6–Me2CO 8:2); mp 72–75 ꢀC;
[a]2D0=À5.67ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 193
(" 64656); IR ꢁmax (KBr) cmÀ1: 3401, 3065, 2929, 2856,
1
2858, 1769, 1730,1657; H NMR: ꢀ 0.73 (3H, s, H-15),
1
0.80 (3H, s, H-14), 0.91 (1H, t, J10b,10a=11.7 Hz, H-
10b), 1.05 (1H, d, J5b,5a=6.2 Hz, H-5b), 1.20–1.30 (2H,
m, H-1a and H-10a), 1.31 (3H, s, H-12), 1.41 (1H, t,
1757, 1646; H NMR (CD3OD): ꢀ 0.80 (3H, s, H-15),
0.87 (3H, s, H-14), 1.21 (3H, s, H-12), 1.28–1.38 (2H, m,
H-10), 1.51 (1H, dd, J1b,1a=13.0 Hz and J1b,2a=7.4 Hz,
H-1b), 1.58 (1H, dd, J1a,1b=13.0 Hz and J1a,2a=1.3 Hz,
H-1a), 2.28–2.33 (1H, m, H-2a), 2.46–2.51 (1H, m, H-
9a), 2.57 (1H, d, J4b,4a=15.0 Hz, H-4b), 2.73 (1H, d,
J
1b,2a=12.1 Hz, H-1b), 1.48 (1H, d, J5a,5b=6.2 Hz, H-
5a), 1.86–1.92 (1H, m, H-2a), 2.14–2.21 (1H, m, H-9a),
3.87 (1H, d, J13b,13a=9.3 Hz, H-13b), 4.02 (1H, d,
0
0
0
0
J
13a,13b=9.3 Hz, H-13a), 4.86 (1H, d, J2 ,3 =2.1 Hz, H-
J
4a,4b=15.0 Hz, H-4a), 4.73 (1H, d, J2 ,3 =3.4 Hz, H-
20), 5.65 (1H, d, J8a,9a=4.5 Hz, H-8a), 5.83 (1H, dd,
3 ,NH=9.3 Hz, J3 ,2 =2.1 Hz, H-3 ), 7.23–7.51 (9H, m,
20), 5.65 (1H, d, J3 ,2 =3.4 Hz, H-3 ), 5.96 (1H, dd,
J8b,9a=11.0 Hz and J=1.2 Hz, H-8b), 7.19 (1H, br s, H-
0
0
0
0
0
0
0
J
NH and H-Ph), 7.80–7.84 (2H, m, H-Ph); 13C NMR: ꢀ
19.4 (t, C-5), 24.9 (q, C-12), 26.3 (q, C-14), 28.6 (q, C-15),
32.0 (s, C-7), 32.1 (s, C-6), 34.6 (s, C-11), 37.5 (d, C-9),
39.5 (t, C-1), 41.9 (t, C-10), 44.2 (d, C-2), 54.8 (d, C-30),
68.2 (t, C-13), 69.7 (s, C-3), 70.4 (d, C-8), 73,8 (d, C-20),
127.0 (d, C-Ph), 127.3 (d, C-Ph), 127.8 (d, C-Ph), 128.5
(d, C-Ph), 128.6 (d, C-Ph), 131.6 (d, C-Ph), 134.2 (s, C-
Ph), 138.7 (s, C-Ph), 166.9 (s, NCO), 172.0 (s, C-10),
174.2 (s, C-4); LSIMS m/z (rel. int.): 556 [M+Na]+
(22), 534 [M+H]+ (4), 516 (3), 338 ( 8), 312 (16), 286
(20), 240 (3), 225 (18), 210 (5), 154 (16), 136 (17), 105
5), 7.22 (1H, br s, H-13), 7.25–7.57 (8H, m, H-Ph), 7.85–
7.92 (2H, m, H-Ph); 13C NMR: ꢀ 31.1 (q, C-12), 32.0 (q,
C-15), 32.4 (q, C-14), 33.6 (t, C-4), 35.8 (s, C-11), 43.3 (t,
C-1), 44.4 (t, C-10), 45.9 (d, C-9), 54.7 (d, C-2), 54.7 (d,
C-30), 72.1 (d, C-8), 72.5 (s, C-3), 74.9 (d, C-20), 120.5 (s,
C-6), 126.6 (s, C-7), 128.2 (d, C-Ph), 128.5 (d, C-Ph),
128.6 (d, C-Ph), 129.5 (d, C-Ph), 129.6 (d, C-Ph), 133.0
(d, C-Ph), 135.2 (s, C-Ph), 139.9 (d, C-13), 140.6 (s, C-
Ph), 142.3 (d, C-5), 169.4 (s, NCO), 172.9 (s, C-10);
LSIMS m/z (rel. int.): 1057 [2M+Na]+ (1), 540
[M+Na]+ (24), 413 (5), 391 (3), 330 (22), 308 (100), 286

P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
785
(95), 240 (8), 215 (24), 176 (95), 136 (60), 158 (28), 105
(98); HR LSIMS m/z: [M+Na]+ calc. for
C31H35O6NNa: 540.23621, found: 540.23982.
CH2CH3), 72.6 (d, C-8), 73.6 (d, C-20), 75.4 (s, C-3),
119.4 (s, C-6), 125.2 (s, C-7), 126.9 (d, C-Ph), 127.1 (d,
C-Ph), 127.8 (d, C-Ph), 128.5 (d, C-Ph), 128.6 (d, C-Ph),
131.7 (d, C-Ph), 133.9 (s, C-Ph), 138.0 (d, C-5), 138.8 (s,
C-Ph), 140.4 (d, C-13), 166.3 (s, NCO), 172.1 (s, C-10);
LSIMS m/z (rel. int.): 568 [M+Na]+ (18), 413 (2), 338
(2), 330 (18), 308 (95), 286 (26), 261 (19), 240 (6), 215
(100), 176 (46), 154 (23), 136 (21), 105 (66), 95 (17); HR
LSIMS m/z: [M+Na]+ calc. for C33H39O6NNa:
568.26751, found: 568.26825.
4.5.6. Compound 24
TLC Rf=0.29 (hexane–EtOAc 1:1); mp 95–98 ꢀC;
[a]2D0=À17.80ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 195
(" 51825); IR ꢁmax (CHCl3) cmÀ1: 3597, 3516, 3438, 2955,
2867, 1733, 1664; 1H NMR: ꢀ 0.98 (3H, s, H-15), 1.01 (3H,
s, H-14), 1.02 (3H, s, H-12), 1.28 (1H, dd, J10a,10b=12.7
Hz and J10a,9b=9.9 Hz, H-10a), 1.50 (1H, dd, J1b,1a=13.0
Hz and
J
1b,2a=10.5 Hz, H-1b), 1.63 (1H, dd,
4.5.8. Compound 30
J10b,10a=12.7 Hz and J10b,9b=6.4 Hz, H-10b), 1.80 (1H,
TLC Rf=0.11 (hexane–EtOAc 7:3); mp 80–83 ꢀC;
[a]2D0=+5.23ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 204
(" 53790); 220 (" 41181); IR ꢁmax (CHCl3) cmÀ1: 3528,
3440, 2956, 2867, 1752, 1667; 1H NMR: ꢀ 0.75 (3H, s, H-
15), 0.77 (3H, s, H-14), 1.12 (1H, dd, J10b,10a=12.8 Hz and
dd, J1a,1b=13.0 Hz and J1a,2a=7.6 Hz, H-1a), 2.07–2.14
(2H, m, H-2a and H-9b), 2.53 (1H, d, J4b,4a=14.0 Hz, H-
4b), 2.67 (1H, d, J4a,4b=14.0 Hz, H-4a), 4.68 (1H, dd,
J2 ,3 =3.6 Hz and J2 ,OH=2.3 Hz, H-20), 5.81 (1H, d,
0
0
0
0
J
8a,9b=10.0 Hz, H-8a), 5.82 (1H, dd, J3 ,NH=9.1 Hz and
J
J
10b,9a=8.8 Hz, H-10b), 1.64 (1H, ddd, J10a,10b=12.8 Hz,
10a,9a=7.8 Hz and J=1.8 Hz, H-10a), 1.74 (3H, br s, H-
0
0
J3 ,2 =3.6 Hz, H-3 ), 7.12 (1H, t, J=1.5 Hz, H-5), 7.19
0
0
(1H, d, JNH,3 =9.1 Hz, NH), 7.22 (1H, t, J=1.0 Hz, H-
12), 2.01 (1H, d, J1b,1a=15.0 Hz, H-1b), 2.29 (1H, d,
1a,1b=15.0 Hz, H-1a), 2.89 (1H, d, J4b,4a=19.9 Hz, H-
4b), 3.13–3.24 (2H, m, H-4a and H-9a), 4.59 (1H, d,
13), 7.28–7.51 (8H, m, H-Ph), 7.74–7.79 (2H, m, H-Ph);
13C NMR: ꢀ 20.7 (q, C-12), 31.0 (q, C-15), 31.1 (q, C-14),
34.7 (s, C-11), 40.1 (t, C-4), 43.2 (t, C-1), 46.2 (t, C-10),
46.4 (d, C-9), 54.6 (d, C-30), 54.9 (d, C-2), 73.3 (s, C-3), 73.6
(d, C-20), 77.1 (d, C-8), 118.8 (s, C-6), 123.9 (s, C-7), 126.9
(d, C-Ph), 127.0 (d, C-Ph), 127.9 (d, C-Ph), 128.6 (d, C-
Ph), 128.7 (d, C-Ph), 131.8 (d, C-Ph), 133.9 (s, C-Ph),
138.9 (s, C-Ph), 139.2 (d, C-13), 140.7 (d, C-5), 166.9 (s,
NCO), 172.1 (s, C-10); LSIMS m/z (rel. int.): 540
[M+Na]+ (18), 481 (3), 425 (6), 329 (18), 308 (19), 286
(40), 225 (19), 176 (100), 154 (55), 136 (42), 105 (40), 91
(81); HR LSIMS m/z: [M+Na]+ calc. for
C31H35O6NNa: 540.23621, found: 540.23494.
J
0
0
J
13b,13a=17.3 Hz, H-13b), 4.69 (1H, d, J2 ,3 =3.8 Hz, H-
20), 4.70 (1H, dd, J13a,13b=17.3 Hz and J=3.6 Hz, H-13a),
0
0
0
0
5.60 (1H, dd, J3 ,NH=11.2 Hz and J3 ,2 =3.8 Hz, H-3 ),
5.61 (1H, d, J8b,9a=11.2 Hz, H-8b), 7.25–7.88 (9H, m, NH
and H-Ph), 8.10–8.15 (2H, m, H-Ph); 13C NMR: ꢀ 22.6 (q,
C-12), 27.2 (q, C-14), 28.6 (q, C-15), 30.9 (t, C-4), 36.2 (s,
C-11), 46.1 (d, C-9), 45.9 (t, C-10), 47.5 (t, C-1), 57.5 (d,
C-8), 57.5 (d, C-30), 71.4 (t, C-13), 74.8 (d, C-20), 127.2
(s, C-2), 128.3 (d, C-Ph), 128.6 (d, C-Ph), 128.7 (d, C-
Ph), 129.4 (s, C-6), 129.5 (d, C-Ph), 129.6 (d, C-Ph),
133.0 (d, C-Ph), 135.2 (s, C-Ph), 137.4 (s, C-Ph), 140.4
(s, C-3), 159.0 (s, C-7), 169.4 (s, NCO), 173.3 (s, C-10),
175.9 (s, C-5); LSIMS m/z (rel. int.): 538 [M+Na]+
(18), 516 [M+H]+ (8), 443 (2), 338 (5), 308 (29), 286
(100), 253 (5), 231 (15), 210 (8), 176 (70), 154 (59), 136
(42), 105 (83); HR LSIMS m/z: [M+Na]+ calc. for
C31H33O6NNa: 538.22056, found: 538.22162.
4.5.7. Compound 27
TLC Rf=0.31 (hexane–EtOAc 7:3); mp 69–74 ꢀC;
[a]2D0=À21.16ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 196
(" 60975); 218 (" 21513); IR ꢁmax (CHCl3) cmÀ1: 3517,
1
3440, 2956, 2871, 1731, 1667; H NMR: ꢀ 0.84 (3H, s,
H-15), 0.87 (3H, s, H-14), 0.90 (3H, t, J=6.9 Hz, -
OCH2CH3), 1.17 (3H, s, H-12), 1.32 (1H, dd, J10a,10b
14.0 Hz and J10a,9a=2.5 Hz, H-10a), 1.43 (1H, dd,
10b,10a=14.0 Hz and J10b,9a=6.8 Hz, H-10b), 1.49 (1H,
dd, J1a,1b=12.6 and J1a,2a=6.6 Hz, H-1a), 1.55 (1H, t,
J=12.6 Hz, H-1b), 2.34–2.40 (1H, m, H-2a), 2.56 (1H,
d, J4b,4a=15.4 Hz, H-4b), 2.58-2.63 (1H, m, H-9a), 2.74
(1H, d, J4a,4b=15.4 Hz, H-4a), 3.27 (1H, qn, J=6.9 Hz,
=
4.5.9. Compound 33
TLC Rf=0.33, (C6H6–Me2CO 8:2); mp 101–103 C;
ꢀ
J
[a]2D0=À18.40ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 218
(" 26218); IR ꢁmax (KBr) cmÀ1: 3428, 2955, 2867, 1746,
1649; 1H NMR: ꢀ 0.71 (3H, s, H-15), 0.90 (3H, s, H-14),
1.06 (1H, dd, J10b,10a=12.6 Hz and J10b,9a=10.0 Hz, H-
10b), 1.22 (3H, s, H-12), 1.20–1.27 (1H, m, H-1b), 1.48
(1H, dd, J10a,10b=12.6 Hz and J10a,9a=6.8 Hz, H-10a),
1.54 (1H, dd, J1a,1b=11.8 Hz and J1a,2a=7.2 Hz, H-1a),
2.44–2.50 (1H, m, H-2a), 2.53-2.62 (3H, m, H-4a, 4-b and
H-9a), 4.41 (1H, br d, J13b,13a=17.3 Hz, H-13b), 4.65 (1H,
-OCH2CH3), 3.36 (1H, qn, J=6.9 Hz, -OCH2CH3), 4.68
0
0
0
0
(1H, d, J2 ,3 =2.2 Hz, H-2 0), 5.79 (1H, dd, J3 ,NH=8.9
0
Hz and J3 ,2 =2.2 Hz, H-3 ), 6.01 (1H, d, J8b,9a=11.2
0
Hz, H-8b), 6.93 (1H, s, H-5), 7.13 (1H, t, J=2.0 Hz, H-
0
13), 7.23 (1H, d, JNH,3 =8.9 Hz, NH), 7.26-7.53 (8H, m,
dt, J13a,13b=17.3 Hz and J=2.5 Hz, H-13a), 4.71 (1H, d,
0
H-Ph), 7.78-7.83 (2H, m, H-Ph); 13C NMR: ꢀ 15.7 (q, -
CH2CH3), 25.5 (q, C-12), 29.1 (t, C-4), 31.6 (q, C-15),
32.1 (q, C-14), 34.7 (s, C-11), 41.8 (t, C-1), 43.8 (t, C-10),
44.2 (d, C-9), 51.9 (d, C-2), 54.7 (d, C-30), 55.6 (t, -
J2 ,3 =2.3 Hz, H-2 ), 5.64 (1H, d, J=5.9 Hz, H-8b), 5.78
0
0
0
0
0
0
(1H, dd, J3 ,NH=9.3 Hz and J3 ,2 =2.3 Hz, H-3 ), 7.25–
C
7.54 (9H, m, NH and H-Ph), 7.78-7.81 (2H, m, H-Ph); 13
NMR: ꢀ 22.6 (q, C-15), 27.1 (q, C-14), 29.4 (q, C-12),

786
P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
34.7 (t, C-4), 36.0 (s, C-11), 42.4 (d, C-9), 44.6 (t, C-1), 45.0
(t, C-10), 50.2 (d, C-2), 54.7 (d, C-30), 70.6 (t, C-13), 71.7
(d, C-8), 72.3 (s, C-3), 73.2 (d, C-20), 126.9 (d, C-Ph), 127.1
(d, C-Ph), 127.8 (s, C-6), 127.9 (d, C-Ph), 128.6 (d, C-Ph),
128.7 (d, C-Ph), 132.1 (d, C-Ph), 133.3 (s, C-Ph), 138.0 (s,
C-Ph), 154.0 (s, C-7), 167.0 (s, NCO), 172.2 (s, C-10), 174.3
(s, C-5); LSIMS m/z (rel. int.): 534 [M+H]+ (10), 391 (3),
369 (5), 307 (19), 286 (24), 225 (10), 215 (8), 176 (5), 154
(100), 137 (71), 105 (39); HR LSIMS m/z: [M+H]+ calc.
for C31H36O7N: 534.24921, found: 534.25325.
m, H-Ph), 7.75–7.78 (2H, m, H-Ph); 13C NMR: ꢀ 20.6
(q, C-12), 30.8 (q, C-15), 31.1 (q, C-14), 34.2 (s, C-11),
39.3 (t, C-4), 43.4 (t, C-1), 44.0 (d, C-9), 46.2 (t, C-10),
54.4 (d, C-2), 54.9 (d, C-30), 69.3 (t, C-13), 72.1 (s, C-3),
73.5 (d, C-20), 77.5 (d, C-8), 124.1 (s, C-6), 127.0 (d, C-
Ph), 127.0 (d, C-Ph), 128.2 (d, C-Ph), 128.7 (d, C-Ph),
128.8 (d, C-Ph), 132.1 (d, C-Ph), 133.4 (s, C-Ph), 138.4
(s, C-Ph), 159.7 (s, C-7), 167.2 (s, NCO), 172.0 (s, C-10),
174.1 (s, C-5); ESI-MS m/z (rel. int.): 1623.7016
[3M+Na]+ (20), 1089.4559 [2M+Na]+ (100), 556.2276
[M+Na]+ (95); HR ESI-MS m/z: [M+Na]+ calc. for
C31H35O7NNa: 556.23112, found: 556.23060.
4.5.10. Compound 36
ꢀ
LC Rf=0.16 (hexane–Me2CO 7:3); mp 114–117 C;
[a]2D0=À11.63ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 194
(" 59567), 220 (" 25610); IR ꢁmax (CHCl3) cmÀ1: 3593,
3440, 2958, 2868, 1738, 1662; 1H NMR: d 0.88 (3H, s, H-
15), 0.99 (3H, s, H-14), 1.03 (1H, t, J=12.2 Hz, H-1b),
1.19 (1H, t, J=12.3 Hz, H-10b), 1.21 (3H, s, H-12), 1.46
(1H, dd, J10a,10b=12.3 Hz and J10a,9a=8.9 Hz, H-10a),
1.51 (1H, dd, J1a,1b=12.2 Hz and J1a,2a=7.8 Hz, H-1a),
2.43 (1H, d, J4b,4a=19.2 Hz, H-4b), 2.53–2.60 (1H, m, H-
4.5.12. Compound 42
TLC Rf=0.34 (hexane–Me2CO 6:4); mp 102–105 C;
ꢀ
[a]2D0=À3.80ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 194
(" 64173), 217 (" 23025); IR ꢁmax (CHCl3) cmÀ1: 3429,
2958, 2868, 1757, 1727, 1650; 1H NMR: ꢀ 0.52 (3H, s, H-
15), 0.78 (3H, s, H-14), 1.06 (1H, t, J=12.4 Hz, H-1b),
1.31 (3H, s, H-12), 1.51 (1H, ddd, J1a,1b=12.4 Hz,
J
J
1a,2a=7.8 Hz and J=2.1 Hz, H-1a), 1.54 (1H, ddd,
10a,10b=12.3 Hz, J10a,9a=6.8 Hz and J=2.2 Hz, H-10a),
2a), 2.60 (1H, dq, J4a,4b=19.2 Hz and J=2.9 Hz, H-4a),
0
0
0
2.83–2.91 (1H, m, H-9a), 4.61 (1H, d, J2 ,3 =2.2 Hz, H-2 ),
2.48-2.58 (2H, m, H-2a and H-9a), 2.56 (1H, d,
0
4.77 (2H, br s, H-13), 5.65 (1H, dd, J3 ,NH=9.0 Hz and
J4b,4a=19.4 Hz, H-4b), 2.76 (1H, d, J4a,4b=19.4 Hz, H-
0
0
0
0
0
J3 ,2 =2.2 Hz, H-3 ), 6.88 (1H, br s, H-8a), 7.15 (1H, d,
4a), 4.68 (2H, br s, H-5a and H-5b), 4.76 (1H, d, J2 ,3 =1.9
JNH,3 =9.0 Hz, NH), 7.28–7.48 (8H, m, H-Ph), 7.72–7.75
Hz, H-20), 5.83 (1H, d, J8b,9a=2.3 Hz, H-8b), 5.86 (1H, dd,
0
0
(2H, m, H-Ph); 13C NMR: ꢀ 26.2 (q, C-15), 28.9 (q, C-14),
29.4 (q, C-12), 34.8 (t, C-4), 37.7 (s, C-11), 42.3 (t, C-10),
43.8 (d, C-9), 44.6 (t, C-1), 48.9 (d, C-2), 55.0 (d, C-30), 70.8
(t, C-13), 73.3 (d, C-20), 73.6 (s, C-3), 74.8 (d, C-8), 122.2
(s, C-6), 127.0 (d, C-Ph), 127.0 (d, C-Ph), 128.0 (d, C-
Ph), 128.6 (d, C-Ph), 128.8 (d, C-Ph), 131.9 (d, C-Ph),
133.6 (s, C-Ph), 138.5 (s, C-Ph), 158.5 (s, C-7), 167.5 (s,
NCO), 174.8 (s, C-5), 174.8 (s, C-10); LSIMS m/z (rel.
int.): 534 [M+H]+ (2), 524 (1), 460 (1), 391 (2), 369 (1),
338 (3), 307 (14), 286 (12), 231 (2), 154 (100), 136 (77),
120 (14), 105 (42); HR LSIMS m/z: [M+Na]+ calc. for
C31H35O7NNa: 556.23112, found: 556.23110.
J3 ,NH=9.9 Hz, J3 ,2 =2.3 Hz, H-3 ), 7.21 (1H, d,
0
0
0
0
J3 ,NH=9.9 Hz, NH), 7.24–7.55 (8H, m, H-Ph), 7.81–7.83
(2H, m, H-Ph); 13C NMR: ꢀ 25.3 (q, C-15), 28.9 (q, C-14),
32.2 (q, C-12), 36.2 (s, C-11), 36.8 (t, C-4), 42.3 (d, C-9),
45.8 (t, C-10), 46.3 (t, C-1), 48.8 (d, C-2), 54.0 (d, C-30),
70.1 (d, C-8), 72.3 (t, C-5), 72.7 (d, C-20), 73.0 (s, C-3),
122.5 (s, C-7), 126.9 (d, C-Ph), 127.1 (d, C-Ph), 127.7 (d, C-
Ph), 128.6 (d, C-Ph), 128.8 (d, C-Ph), 132.3 (d, C-Ph),
133.3 (s, C-Ph), 137.9 (s, C-Ph), 164.2 (s, C-6), 167.3 (s,
NCO), 172.5 (s, C-10), 173.9 (s, C-13); LSIMS m/z (rel.
int.): 1067 [2M+H]+ (1), 534 [M+H]+ (26), 460 (1), 286
(100), 240 (8), 210 (20), 167 (7), 154 (76), 136 (56), 105 (76);
HR LSIMS m/z: [M+H]+ calc. for C31H36O7N:
534.24918, found: 534.24918.
4.5.11. Compound 39
TLC Rf=0.31 (hexane–Me2CO 6:4); mp 116–120 C;
ꢀ
[a]2D0=À47.20ꢀ (CHCl3, c 1.0); UV lmax (EtOH) nm: 221
(" 24542); IR nmax (KBr) cmÀ1: 3423, 2952, 2865, 1747,
1649; 1H NMR: ꢀ 0.97 (3H, s, H-15), 1.01 (3H, s, H-14),
1.07 (3H, s, H-12), 1.28 (1H, dd, J10a,10b=12.6 Hz and
4.6. Antifeedant activity test
The test is described in detail in B•oszyk et al. (1995).
Insects (adults and larvae) used for the test were reared
under laboratory conditions at temp. of 26 ꢀC and 75%
humidity. All compounds investigated were dissolved in
EtOH at concentration of 10 mg mlÀ1. Air dry wheat
wafer discs were used as test food. The discs (1 cm in
diameter) were saturated with EtOH solutions of pure
compounds to produce 0.5% (by weight) contamination
of the wafer in every test. Feeding of insects was recor-
ded under three conditions: (1) on pure food (control);
(2) on food with possibility of choice (choice test); on
food with the compounds tested (no choice test). The
wafer discs were weighed after saturation and drying in
J
10a,9b=10.3 Hz, H-10a), 1.51 (1H, dd, J1b,1a=13.2 Hz
and J1b,2a=10.1 Hz, H-1b), 1.60 (1H, dd, J10b,10a=12.6
Hz and 10b,9b=6.4 Hz, H-10b), 1.83 (1H, dd,
1a,1b=13.2 Hz and J1a,2a=7.8 Hz, H-1a), 2.14-2.25
J
J
(2H, m, H-2a and H-9b), 2.28 (1H, dd, J4b,4a=15.1 Hz
and J=2.3 Hz, H-4b), 2.75 (1H, d, J4a,4b=15.1 Hz, H-
4a), 4.56 (1H, dd, J13b,13a=18.1 Hz and J=2.9 Hz, H-
0
0
0
13b), 4.66 (1H, d, J2 ,3 =2.4 Hz, H-2 ), 4.78 (1H, br d,
0
J
and J3 ,2 =2.4 Hz, H-3 ), 5.79 (1H, br d, J8a,9a=8.7 Hz,
13a,13b=18.1 Hz, H-13a), 5.71 (1H, dd, J3 ,NH=9.0 Hz
0
0
0
0
H-8a), 7.21 (1H, d, JNH,3 =9.0 Hz, NH), 7.29–7.52 (8H,

P. Kopczacki et al. /Phytochemistry 58 (2001) 775–787
Daniewski, W.M., Gumu•ka, M., Ptaszyn
787
air for 30 min before the experiments and again after 7
days of feeding by beetles or larvae. On the basis of
eaten food, the index of activity of the compounds tes-
ted was calculated in the following way: three values of
the food eaten were obtained in the control KK, in the
no-choice test EE, and in the choice test K, E.
˜
ska, K., Skibicki, P., B•os-
.
.
zyk, E., Drozdz, B., Stromberg, S., Norin, T., Holub, M., 1993.
Antifeedant activity of some sesquiterpenoids of the genus Lactar-
ius. European Journal of Entomology 90, 65–70.
Daniewski, W.M., Gumu•ka, M., Przesmycka, D., Ptaszynska, K.,
´
.
.
B•oszyk, E., Drozdz, B., 1995. Sesquiterpenes of Lactarius origin,
antifeedant structure-activity relationship. Phytochemistry 38,
1161–1168.
Thus, the relative coefficient of antifeedancy:
Daniewski, W.M., Gumu•ka, M., Anczewski, W., Masnyk, M., B•os-
zyk, E., Gupta, K.K., 1998. Why the yew tree (Taxus baccata) is not
attacked by insects. Phytochemistry 49, 1279–1282.
K À E
R ¼
 100
K þ E
Daniewski, W.M., Vidari, G., 1999. Constituents of Lactarius (mush-
rooms). Progress in the Chemistry of Organic Natural Products 77,
69–171.
The total coefficient of antifeedancy is equal to
T=A+R, and the maximum value of the coefficient can
reach 200 for a perfect antifeedant.
Farina, V., 1995. The Chemistry and Pharmacology of Taxol and its
Derivatives. Elsevier Science B.V., Amsterdam. The Netherlands.
Kamata, K., Agata, I., Meyers, A.I., 1998. An efficient and versatile
method for the synthesis of optically active 2-oxazolines: an acid-
catalyzed condensation of ortho esters with amino alcohols. Journal
of Organic Chemistry 63, 3113–3116.
References
.
.
B•oszyk, E., Szafran
´
ski, F., Drozdz, B., Al-Shameri, A., 1995. African
Kingston, D.G.I., Chaudhary, A.G., Gunatilaka, A.A.L., Middleton,
M.L., 1994. Synthesis of taxol from baccatin III via oxazoline
intermediate. Tetrahedron Letters 35, 4483–4484.
plants as antifeedants against stored-product insect pests. Journal of
Herbs, Spices and Medicinal Plants 3, 25–36.