In vitro evaluation of antifungal properties of phenylpropanoids and related compounds acting against dermatophytes

Article abstract of DOI:10.1021/np9805443

Thirty-four arylpropanoids and related compounds were evaluated in vitro for antifungal properties. Among them, 22 phenyl-, 4 naphthyl-, and 4 phenanthrylpropanoids; naphthalene; phenanthrene; and 2-chloro-1-hexyl-1- propanone were tested against dermatop

Full text of DOI:10.1021/np9805443

©
Copyright 1999 by the American Chemical Society and the American Society of Pharmacognosy
Volume 62, Number 10
October 1999
Full Papers
In Vitr o Eva lu a tion of An tifu n ga l P r op er ties of P h en ylp r op a n oid s a n d Rela ted
Com p ou n d s Actin g Aga in st Der m a top h ytes
,
†
†
†
†
†
Susana A. Zacchino,* Silvia N. L o´ pez, Germ a´ n D. Pezzenati, Ricardo L. Furl a´ n, Carina B. Santecchia,
†
‡
‡
‡
Lorena Mu n˜ oz, Fernando A. Giannini, Ana M. Rodr ´ı guez, and Ricardo D. Enriz
Area Farmacognosia, Facultad de Ciencias Bioqu ı´ micas y Farmac e´ uticas, Universidad Nacional de Rosario, Suipacha 531,
2
000 Rosario, Argentina, and Area de Qu ´ı mica General, Facultad de Qu ´ı mica, Bioqu ı´ mica y Farmacia,
Universidad Nacional de San Luis, Chacabuco y Pedernera, 5700 San Luis, Argentina
Received November 30, 1998
Thirty-four arylpropanoids and related compounds were evaluated in vitro for antifungal properties.
Among them, 22 phenyl-, 4 naphthyl-, and 4 phenanthrylpropanoids; naphthalene; phenanthrene; and
2
-chloro-1-hexyl-1-propanone were tested against dermatophytes by the agar dilution method. R-Halo-
propiophenones exhibited a broad spectrum of activities against Microsporum canis, Microsporum gypseum,
Trichophyton mentagrophytes, Trichophyton rubrum, and Epidermophyton floccosum, with MIC values
between 0.5 and >50 µg/mL. Keto, alcohol, and R-haloketo propyl derivatives of naphthalene and
phenanthrene also showed very good activity against all dermatophytes tested, clearly showing that in
these series, a halogen atom is not necessary for activity. Phenanthryl derivatives were more active (MICs,
3
-20 µg/mL) than naphthyl ones (MICs, 3-50 µg/mL). A structure-activity relationship study was carried
out and aided in establishing the structural requirements of arylpropanoids for antifungal activities.
Because dermatophytes are a group of fungi that characteristically infect the keratinized areas of the
body, these new series of antifungal compounds open the possibility of discovering new topical antifungal
drugs for the treatment of dermatomycoses, which are frequently very difficult to eradicate.
During the past two decades, the incidence of fungal
infections, especially involving immunocompromised pa-
tients, has increased dramatically.1 In particular, some
forms of dermatomycoses are the cause of a great morbidity
in patients receiving antineoplastic chemotherapy, under-
going organ transplants, or suffering from AIDS. These
infections are produced by dermatophytes, a group of fungi
that characteristically infect the keratinized areas of the
body. Although imidazole compounds such as clotrimazole,
miconazole, and econazole have proven to be effective for
the treatment of dermatomycoses, these infections are
In the course of our screening program for antifungal
activity, we reported that 8.O.4′-neolignans (a small group
among the great structural variety of neolignans) possess
moderate but significant antifungal activity against der-
matophytes.3
In addition, other types of neolignans and structurally
4-6
related lignans showed antifungal activities. Considering
that lignans and neolignans are plant compounds formed
by two C-6-C-3 units7 and that dimerization of phenyl-
propanoids through dehydrogenation produces the skeleton
of all natural and synthetic lignans known to date, we
decided to carry out a systematic study of the antifungal
properties of their phenylpropanoid moieties.
2
frequently very difficult to eradicate, and more effective
new topical antifungal agents are still needed.
Studies have reported contradictory evidence of antimi-
crobial activities of some phenylpropanoids. Zemek et al.,
for example, reported that isoeugenol exhibits an inhibitory
effect on the growth of bacteria, yeasts, yeast-like organ-
*
To whom correspondence should be addressed. Tel.: +54 (341) 437-
8
5
315. Fax: +54 (341) 437-5315. E-mail: szaabgil@citynet.net.ar.
†
Area Farmacognosia.
Area de Qu ´ı mica General.
‡
1
0.1021/np9805443 CCC: $18.00
© 1999 American Chemical Society and American Society of Pharmacognosy
Published on Web 09/18/1999

1
354 J ournal of Natural Products, 1999, Vol. 62, No. 10
Zacchino et al.
Ta ble 1. In Vitro Antifungal Activities of Phenylpropanoids and a Nonaromatic Analogs
MIC (µg/mL)
T. m.c
compd
type
-R1
-R2
-R3
-R4
-R5
M. c.a
M. g.b
T. r.d
E. f.e
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
A
A
A
A
A
A
A
A
A
A
A
A
A
B
B
B
B
B
C
D
D
D
D
E
OCH3
-O-CH2-O-
OCH3
H
OCH3
-O-CH2-O-
OCH3
OCH3
-O-CH2-O-
OCH3
H
OCH3
H
OCH3
OCH3
-O-CH2-O-
OCH3
OCH3
H
H
OCH3
H
H
H
OCH3
H
H
OCH3
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
CH3
H
H
H
H
H
H
>50
>50
>50
>50
12.5
3.12
15
3.12
12.5
6.25
6.25
>50
>50
>50
>50
>50
50
>50
>50
>50
>50
40
25
50
12.5
12.5
12.5
20
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
6.25
6.25
>50
>50
>50
>50
25
>50
>50
>50
>50
>50
12,5
>50
10
10
10
12.5
50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
>50
25
50
40
>50
>50
3
6.25
8
0.15
6.25
0.5
15
50
50
50
50
>50
50
>50
>50
>50
>50
>50
>50
>50
0.3
25
OCH3
OCH3
OCH3
Br
Br
Br
Cl
Cl
Cl
Cl
CH3
CH3
Br
H
20
OCH3
OCH3
12.5
12.5
25
15
10
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
OCH3
H
OCH3
H
OCH3
OCH3
>50
>50
>50
>50
>50
>50
>50
50
>50
>50
>50
>50
>50
6.25
12.5
f
H
H
CH3
Br
OCH3
OCH3
H
OCH3
H
H
H
OCH3
H
OCH3
-O-CH2-O-
OCH3
>50
50
OH
>50
>50
>50
>50
>50
>50
15
H
OCH3
OCH3
OCH3
H
-O-CH2-O-
OCH3
OH
g
amp
ket.
h
15
a
M. canis C 112. M. gypseum C 115. c T. mentagrophytes ATCC 9972. d T. rubrum C 113. e E. floccosum C 114. f threo isomer. g Amp.
b
h
)
amphotericin B. Ket. ) ketoconazole.
isms, and molds twice as effective as its dimer, which
consists of two isoeugenol units. Contrary to this, Hattori
et al. reported that dimeric phenylpropanoids show stron-
phillus12 (Myrtaceae); methyleugenol 21, 5′-methoxyeu-
genol 22, elemicin 23, and isoeugenol 24 were isolated from
9
13
M. fragrans; and compounds 22 and 23 were isolated from
1
4
ger antimicrobial action than the corresponding monomer
unit. On the other hand, Himejima et al.10 reported that
safrole, methyleugenol, eugenol, and anethole were inactive
against Saccharomyces cerevisiae, Candida utilis, Pity-
rosporum ovale, and Penicillum chrysogenum at concentra-
tions up to 100 µg/mL.
A more systematic investigation of the antifungal activi-
ties of phenylpropanoids utilizing a large number of
compounds seems in order and will provide information on
the role that phenylpropanoids play in the antifungal
activity of their dimers against human pathogenic and
opportunistic fungi and will establish whether the anti-
fungal activity reported for 8.O.4′-neolignans can be as-
cribed to their phenylpropanoid moieties. In addition, this
systematic study could aid in the development of new and
more potent topical antifungal agents.
Virola surinamensis (Myristicaceae) and Diphlolophium
15
buchanani (Umbelliferae), respectively. Compounds 1-3,
5-11, and 14-19 are synthetic analogues. Compounds 4
and 13 are commercial samples (Aldrich Chemical Co). To
the best of our knowledge, compounds 9, 10, 15, 18, and
29-32 have not been reported previously in the literature.
16
In this study, agar dilution assays were used to
determine the minimal inhibitory concentrations (MICs)
of compounds by using a panel of human pathogenic fungi
consisting of yeasts as well as dermatophytes.
Resu lts a n d Discu ssion
To carry out the antifungal evaluation, concentrations
of arylpropanoids up to 50 µg/mL were incorporated into
3
growth media according to reported procedures. Com-
pounds producing no inhibition of fungal growth at that
level were considered inactive.
We describe here the antifungal properties of phenyl-
propanoids 1-18, 20-24, and hexylpropanoid 19 against
dermatophytes. In addition, results obtained with naph-
thalene, phenanthrene, naphthyl- and phenanthryl-pro-
panoids 25-32, and a structure-activity relationship
The agar dilution method showed that none of the
compounds tested possessed any activity against the yeasts
Candida albicans, S. cerevisiae, or Cryptococcus neofor-
mans or the filamentous fungi Aspergillus niger, Aspergil-
lus fumigatus, or Aspergillus flavus (results not shown).
In contrast, different results were obtained for compounds
of the series against dermatophytes, with some compounds
displaying strong activities. These results are shown in
Tables 1 and 2. All of the dermatophytes tested were
(SAR) study considering all the evaluated compounds are
also reported.
Among phenylpropanoids tested, aromatic ether 12 was
isolated from Wedelia forsteriana11 (Compositae); eugenol
2
0 was isolated from Myristica fragrans (Myristicaceae),
Cinnamomum zeylanicum (Lauraceae), and Eugenia caryo-

Antifungal Phenylpropanoids against Dermatophytes
J ournal of Natural Products, 1999, Vol. 62, No. 10 1355
Ta ble 2. In Vitro Antifungal Activities of Naphthyl- and
Phenanthrylpropanoids; Naphthalene and Phenanthrene
response through electronic or steric factors, we compared
the antifungal activities of compounds 8 and 12 (the last
one obtained through a bioisosteric substitution of chlorine
by a methyl group in compound 8). Compound 12 was
inactive, suggesting that the biological response is governed
by electronic rather than steric factors.
Because R-haloketones 5-11, which have substantial
intrinsic chemical and biological reactivity,17 were the only
phenylpropanoids active against dermatophytes, we syn-
thesized and tested structures 25-31 (Table 2). These
compounds do not possess R-halo groups, and benzene rings
were replaced by two different extended π-systems (naph-
thalene and phenanthrene).
The replacement of the benzene ring with or without
substituents (compounds 1-4, 12, and 15-18) by naph-
thalene and phenanthrene rings (compounds 25-28 and
2
9-31) showed that an increase in the number of rings
respective to benzene results in better activity. The com-
parison of MICs of keto compounds 12, 26, and 30 il-
lustrates this point well. Among naphthyl and phenanthryl
derivatives, results showed that compounds 25-31 inhib-
ited all dermatophytes tested with MICs between 3 and
50 µg/mL, phenanthrene derivatives 29-31 being the most
active ones (MICs 3-20 µg/mL).
It is interesting to note, that whereas the presence of
both halogen and keto groups were essential for antifungal
activities in the phenylpropanoid series, the same seems
not to be true for naphthyl and phenanthryl compounds.
Keto compounds 25, 26, 29, and 30 and alcohol derivatives
MIC(µg/mL)
compd type
R
M. c.a
M. g.b
T. m.c
T. r.d
E. f.e
2
2
2
2
2
3
3
3
5
6
7
8
9
0
1
2
G
G
H
H
J
J
K
J
H
CH3
H
CH3
H
CH3
CH3
Cl
50
40
50
25
3
20
6.25
3
>50
>50
>50
15
40
25
40
25
25
25
40
>50
40
12.5
6.25
20
10
1.5
>50
>50
25
40
40
40
25
20
20
10
15
>50
>50
0.3
25
25
12.5
3
6.25
6.25
3
6.25
20
6.25
10
f
napht.
phen.
amp.
ket.
>50
>50
6.25
6.25
>50
>50
6.25
12.5
g
h
2
7, 28, and 31 all possess antifungal activities. In addition,
i
15
it is noteworthy that even though R-halo ketone 32 displays
strong antifungal activity, ketone 29, which does not
possess a halogen atom in its structure, shows similar
antifungal properties. In addition, although the bioisosteric
substitution of the chlorine atom by a methyl group
a
M. canis C112. M. gypseum C115. c T. mentagrophytes
b
d
e
f
ATCC 9972. T. rubrum C113. E. floccosum C114. Napht. )
g
h
naphthalene. Phen. ) phenanthrene. Amp. ) amphotericin B.
i
Ket. ) ketoconazole.
(
compounds 32 vs 30) results in a decrease in antifungal
inhibited at 50 µg/mL and most often at lower concentra-
tions.
activity (compare MICs of both compounds), compound 30
does show some antifungal properties, in some cases
similar to or better than the antifungal agents amphoteri-
cin or ketoconazole.
To test the actual role of naphthyl or phenanthryl
structures in the antifungal activities of their propyl
derivatives, naphthalene and phenanthrene were evaluated
with the agar dilution method. Results indicated that these
compounds were completely devoid of activity, clearly
showing that the presence of naphthyl or phenanthryl
framework alone is not enough to produce activity.
Regarding fungi tested with phenanthrene derivatives,
Trichophyton mentagrophytes was the most susceptible
species (4 MICs e 6.25 µg/mL) followed by Microsporum
canis (3 MICs e 20 µg/mL).
According to the results obtained here, we could not
attribute to phenylpropanoid moieties the activities re-
ported previously for 8.O.4′-neolignans. In addition we
found that among phenylpropanoids tested, only R-haloke-
toderivatives possess strong antifungal activities against
dermatophytes. Because they are known reactive com-
Among the phenylpropanoids tested, compounds 1-3,
1
5-17, and 20-22, constitutive moieties of 8.O.4′-neolig-
3
nans reported previously, and compounds 4 and 12-14,
were all inactive. However, it is interesting to note that
phenylpropanoids 5-11 showed strong antifungal activities
comparable to those of amphotericin B and ketoconazole
(Table 1). Compounds 6 and 8-11 inhibited all tested
dermatophytes with MICs ranging between 0.5 and 25 µg/
mL, and compounds 5 and 7 were active against all
dermatophytes except Tricophyton rubrum, with MICs
ranging between 3 and 50 µg/mL. The analysis of active
structures reveals that all of them possess a halogen in
C-2 and a keto group in C-1, and bromine derivatives 5-7
were less active (MICs between 3 and >50 µg/mL) than
chlorine derivatives 8-10 (MICs between 0.5 and 25 µg/
mL). By comparing these results with those obtained with
compounds 1-4, 12, and 13 (MICs ) 40 µg/mL against all
dermatophytes tested), which have only a keto group joined
to a benzene ring in their structures, it would appear that
the presence of a halogen atom at C-2 seems to be crucial
to produce the biological response. Nevertheless, the lack
of activity found for threo-bromohydrin 14 indicates that
the presence of a halogen group is necessary but not
sufficient for antifungal activity of phenylpropanoids.
1
7,18
pounds, in some cases vesicant or lachrymatory,
they
are not suitable for developing new antifungal topical
compounds. Nevertheless, when we investigated the effect
of replacing the phenyl group by naphthyl or phenanthryl
structures in the monomers, we found a novel series of
arylpropanoids not possessing halogen in their structures
(compounds 25-31), with a strong antifungal effect against
dermatophytes. Among them, structures 29-31 displayed
antifungal behavior similar to or better than those of
amphotericin B or ketoconazole, and therefore they could
3
To evaluate if a halogen in a [-COCH(CH )X] system is
actually responsible for the observed activity, we tested
compound 19. The fact that it showed no activity against
all tested dermatophytes also suggests that this system
alone is not enough to produce the antifungal response.
To determine if the halogen atom effects the antifungal

1
356 J ournal of Natural Products, 1999, Vol. 62, No. 10
Zacchino et al.
be leads for the development of new topical antifungal
agents. In addition, results reported here open the pos-
sibility of synthesizing new dimers that may have better
antifungal properties than the previously reported anti-
fungal 8.O.4′-neolignans.
19.9 (q, C-3), 52.5 (d, C-2), 101.9 (t, -OCH
2
O-), 107.9 (d, C-2′),
108.6 (d, C-5′), 125.6 (d, C-6′), 128.5 (s, C-1′), 148.2 (s, C-3′),
+
1
52.2 (s, C-4′), 191.8 (s, C-1); EIMS m/z [M ] 212 (17), 149
100), 121 (50), 91 (20), 65 (35); HREIMS m/z 212.0243 (calcd
for C10 ClO , 212.0240); HPLC (MeOH-H O) t ) 8.68 min
99.8%); HPLC (MeCN-H O) t ) 7.82 min (99.7%).
r-Ch lor o-1-(3′,4′,5′-tr im eth oxyph en yl)-1-pr opan on e (10).
(
H
9
3
2
R
(
2
R
Exp er im en ta l Section
Compound 10 was obtained following the same method used
for compound 9 with ketone 3 (255.4 mg, 1.14 mmol), copper-
(II) chloride dihydrate (340 mg, 2 mmol), lithium chloride (50.4
mg, 1.2 mmol, and DMF (6 mL). R-Chloro-1-(3′,4′,5′-trimethox-
yphenyl)-1-propanone (10) (196 mg, 0.76 mmol, 67% yield) was
obtained as white crystals (hexane): mp 71.5-72.5 °C; IR
Gen er a l Exp er im en ta l P r oced u r es. Melting points were
obtained on an electrothermal apparatus and are uncorrected.
1
13
H and C NMR spectra were recorded on a Bruker model
AC 200 spectrometer in CDCl solutions. Carbon chemical
shifts are expressed in the δ scale in parts per million, using
CDCl as a reference signal at 76.9. J values are given in
3
-
1 1
(
KBr) νmax 2941, 1685, 1128 cm ; H NMR (CDCl
δ 1.70 (3H, d, J ) 6.6 Hz, H-9), 3.88-3.90 (3 × OCH
1H, q, J ) 6.6 Hz, H-8), 7.25 (2H, s, ArH); 13C NMR (CDCl
0 MHz) δ 19.9 (q, C-3), 52.4 (d, C-2), 56.2 (q, -OCH on C-3′
and C-5′), 60.8 (q, -OCH on C-4′), 106.6 (d, C-2′, C-6′), 129.0
s, C-1′), 143.2 (s, C-4′), 153.0 (s, C-3′, C-5′), 192.3 (s, C-1);
3
, 200 MHz)
3
3
), 5.19
Hertz. Elemental analyses were carried out at Atlantic Mi-
crolab, Inc. (Norcross, GA), and all compounds submitted for
testing had analytical results within (0.4% of the theoretical
values. Reversed-phase HPLC was performed on a Beckman
chromatograph (model 332) equipped with an UV detector of
(
5
3
,
3
3
(
+
EIMS m/z [M ] 258 (9), 195 (100), 167 (18), 152 (21), 109 (19),
2
54 nm, on a C18 ODS2 analytical column (0.46 × 25 cm) using
MeOH-H O 70:30 and MeCN-H O 60:40 as the eluents, with
a flow rate of 1 mL/min. Preparative TLC was done on Si gel
7
7 (26), 66 (26); HREIMS m/z 258.0654 (calcd for C12
58.0659); anal. C 55.78%, H 5.86%, Cl 13.52%, calcd for
Cl, C 55.80%, H 5.86%, Cl 13.55%.
-(3′,4′-Dim eth oxyp h en yl)-1-p r op a n ol (15). An ethereal
solution of ketone 1 (428 mg, 2.2 mmol) was gradually added
to a stirred suspension of LiAlH (456 mg, 12 mmol) in dry
15 4
H O Cl,
2
2
2
12 15 4
C H O
1
6
2
0 F254 1 mm. IR spectra were measured with a Bruker IFS
5 FT IR spectrophotometer. MS were measured on a MS
Shimadzu QP-5000 and a ZAB-SEQ4F70 spectrometer.
4
1
9,20
20-22
Test Com p ou n d s. Phenylpropanoids 1, 5, and 8,
2,
Et O (36 mL). After addition was complete, the mixture was
2
1
4-23
22,24
18
25-28
29
3
, 7, and 17,
7,
11, 12,
threo-bromohydrin 14,
refluxed for 8 hs. Excess LiAlH4 was carefully destroyed by
3
30
14,25
31,32
alcohol 16 , hexylpropanoid 19, and compounds 22,
27,
addition of EtOAc. The mixture was extracted with Et O (2 ×
2
3
3
33
2
6, and 28 were synthesized following general methods
50 mL). The combined Et O extracts were washed with 10%
2
1
8-30
1
13
described previously.
Their H NMR, C NMR, IR, mp,
HCl, saturated aqueous NaCl and H O, dried (Na SO ),
2
2
4
and MS spectra were all in agreement with previously reported
decanted, and evaporated under vacuum, yielding 356 mg (1.82
mmol, 83%) of 1-(3′,4′-dimethoxyphenyl)-1-propanol (15) as a
1
8-33
data.
Compounds 4, 13, 20, 21, 23, and 24 were com-
-
1 1
mercial samples.
colorless oil; IR (film) νmax 3482, 2968, 2874, 1594 cm
H
;
2
-Met h yl-1-(3′,4′-d im et h oxyp h en yl)-1-p r op a n ol (18).
NMR (CDCl , 200 MHz) δ 0.89 (3H, t, J ) 7.3 Hz, H-3), 1.78
3
1
4
Compound 18 was prepared through a Grignard reaction
(2H, m, H-2), 1.96 (1H, brs, C-1, -OH), 3.86-3.87 (2 × OCH ),
3
1
3
with 3,4-dimethoxybenzaldehyde (Aldrich) (539.5 mg, 3.25
mmol) in anhydrous Et O (5 mL) and a 2.0 M solution of
4.51 (1H, t, J ) 6 Hz, H-1), 6.83-6.87 (m, ArH); C NMR
2
(CDCl , 50 MHz) d 10.1 (q, C-3), 31.7 (t, C-2), 55.6 and 55.7
3
isopropylmagesium chloride (Aldrich) (6.7 mL, 13.34 mmol).
The mixture was stirred for 8 h, then cooled in an ice bath
and poured into a cold saturated ammonium chloride solution.
The ethereal layer was washed with aqueous 1% NaOH and
(q, -OCH on C-3′ and C-4′), 75.7 (d, C-1), 108.8 (d, C-2′), 110.7
3
(d, C-5′), 118.1 (d, C-6′), 137.1 (s, C-1′), 148.2 (s, C-4′), 148.8
+
(s, C-3′); EIMS m/z [M ] 196 (19), 167 (80), 139 (100), 124 (52),
95(30), 77 (65), 53 (69), 39 (80); HREIMS m/z 196.1093 (calcd
for C H O , 196.1099); HPLC (MeOH-H O) t ) 4.57 min
brine and thoroughly washed with 0.1 N HCl and H
2
O, dried
1
1
16
3
2
R
(
Na SO ), and concentrated under reduced pressure. The
2
4
(100%); HPLC (MeCN-H O) t ) 3.92 min (98.4%).
2
R
residue was purified by column chromatography on Si gel 60H
using hexane-EtOAc (9:1) as the eluent; 491.7 mg (2.34
mmol, 72% yield) of 2-methyl-1-(3′,4′-dimethoxyphenyl)-1-
propanol (18) was obtained as white crystals: mp 67-68 °C;
1-(9′-P h en a n th r yl)-2-m eth yl-1-p r op a n ol (31). Compound
31 was obtained from 9-phenanthraldehyde (Sigma) (383.2 mg,
1.86 mmol) and a 2.0 M solution of isopropyl Mg chloride
(Aldrich) (5 mL, 10 mmol), following the procedure described
for compound 18, as a white solid (386.8 mg, 1.54 mmol, 83%
-
1
1
IR (KBr) νmax 2970, 2920, 2890, 1605 cm ; H NMR (CDCl
00 MHz) δ 0.75 (3H, d, J ) 6.8 Hz, H-3), 0.98 (3H, d, J ) 6.8
Hz, H-3), 1.90 (1H, m, H-2), 2.04 (1H, brs, C-1, -OH), 3.83-
.84 (2 × OCH ), 4.23 (1H, d, J ) 7.1 Hz, H-1), 6.78-6.84 (3H,
ArH); C NMR (CDCl , 50 MHz) δ 18.4 and 18.9 (q, C-3 and
on C-3′ and C-4′), 79.8
3
,
2
yield): mp 106-107 °C; IR (KBr) ν
max
3589, 3577, 3064, 2956,
1473, 1010 cm ; H NMR (CDCl , 200 MHz) δ 1.00 (6H, d, J
-
1 1
3
3
3
) 6 Hz, H-3), 2.05 (1H, d, H-1, J ) 4 Hz), 2.35 (1H, m, H-2),
1
3
13
3
5.22 (1H, brs, 1H, -OH), 7.26-8.76 (m, 9 H, ArH); C NMR
2
-methyl), 35.1 (d, C-2), 55.7 (q, -OCH
3
(CDCl , 50 MHz) δ 16.9 and 20.2 (q, C-3 and 2-methyl), 33.7
3
(
(
(
d, C-1), 109.2 (d, C-2′), 110.4 (d, C-5′), 118.7 (d, C-6′), 136.2
s, C-1′), 148.0 (s, C-4′), 148.6 (s, C-3′); EIMS m/z [M ] 210
63), 168 (100), 139 (92), 108 (72), 77 (22) 71 (45), 65 (18);
(d, C-2), 76.5 (d, C-1), 122.3, 123.2, 124.0, 124.2, 126.0, 126.3,
126.4, 126.5, 128.6 (d, C 1′-8′, C-10′), 129.7, 129.8, 130.6, 131.2,
+
+
137.6 (s, C-4′a, 4′b, 8′a, 9′, 10′a); EIMS m/z [M ] 250 (22), 207
HREIMS m/z 210.1260 (calcd for C12 , 210.1256); anal.
H
18
O
3
(100), 179 (55), 69 (20), 41 (30); HREIMS m/z 250.1429 (calcd
for C H18O, 250.1358); anal. C 86.28%, H 7.27%, calcd for
C 68.53%, H 8.68%, calcd for C12H O , C 68.53%, H 8.63%.
18 3
1
8
r-Ch lor op r op iop h en on es. A chlorination procedure suit-
able for preparing compounds 9, 10, and 32 was made
following the methodology described by Kosower et al.18 It is
illustrated for the case of R-chloro-1-(3′,4′-methylenedioxyphe-
nyl)-1-propanone (9).
r-Ch lor o-1-(3′,4′-m e t h yle n e d ioxyp h e n yl)-1-p r op a n -
on e (9). Piperonyl propiophenone 2 (320.5 mg, 1.8 mmol) was
added to a mixture of copper (II) chloride dihydrate (523 mg,
C
18
H
18O, C 86.35%, H 7.25%.
1-(9′-P h en a n th r yl)-1-p r op a n on e (29). Compound 29 was
obtained by oxidation of 1-(9′-phenanthryl)-1-propanol (330.5
mg, 1.4 mmol) (obtained in turn by a Grignard reaction of
9-phenanthraldehyde and ethyl Mg bromide) with J ones’
reagent as a white solid (320.7 mg, 1.37 mmol, 98% yield): mp
55.5-56.5 °C IR (KBr) νmax 2980, 2870, 1602 cm , H NMR
(CDCl , 200 MHz) δ 1.33 (3H, t, J ) 6 Hz, H-3), 3.13 (2H, q,
H-2, J ) 6 Hz), 7.63-8.65 (9 H, m, ArH); C NMR (CDCl
-
1
1
3
1
3
3
.10 mmol), lithium chloride (90 mg, 2.1 mmol), and DMF (7
3
, 50
mL) and heated for 6 h to 80 °C. R-Chloro-1-(3′,4′-methylene-
dioxyphenyl)-1-propanone (9) (252.3 mg, 1.19 mmol, 66% yield)
was obtained after column chromatography purification (Si gel
MHz) δ 6.6 (q, C-3), 35.4 (t, C-2), 122.6, 122.8, 126.5, 127.0,
127.3, 127.4, 128.5, 128.6, 129.5 (d, C 1′-8′, C-10′), 128.3, 130.0,
130.7, 131.6, 135.5 (s, C-4′a, 4′b, 8′a, 9′, 10′a), 205.4 (s, C-1);
+
6
0 H, eluent CHCl
3
) as a white solid: mp 32.5-33 °C; IR (KBr)
EIMS m/z [M ] 234 (32), 205 (100), 177 (56), 151 (10), 88 (32),
-
1 1
ν
max 3010, 2940, 1695 cm ; H NMR (CDCl
3H, d, J ) 6 Hz, H-3), 5.16 (1H, q, J ) 6 Hz, H-2), 6.05 (2H,
3
, 200 MHz) δ 1.71
69 (78); HREIMS m/z 234.1123 (calcd for C17
H
14O, 234.1045);
(
anal. C 87.05%, H 6.05%, calcd for C17H14O, C 87.14%, H
6.03%.
1
3
s, -OCH
2
O-), 6.87-7.63 (ArH); C NMR (CDCl
3
, 50 MHz) δ

Antifungal Phenylpropanoids against Dermatophytes
J ournal of Natural Products, 1999, Vol. 62, No. 10 1357
1
-(9′-P h en a n th r yl)-2-m eth yl-1-p r op a n on e (30). Com-
pound 30 was obtained by oxidizing the secondary alcohol 31
200 mg, 0.8 mmol) with J ones’ reagent (188.5 mg, 0.76 mmol,
was defined as the lowest compound concentration showing
no visible fungal growth after incubation time.
(
9
2
5% yield) as a white solid: mp 47-48 °C; IR (KBr) νmax 2970,
Ack n ow led gm en t. This work was supported by respective
grants of the Universidad Nacional de Rosario (Project B050),
Universidad Nacional de San Luis (Project 81-01), Universidad
Nacional de Rosario (Res. 9405/95), and from CONICET (PEI
-1 1
920, 2890, 1605 cm ; H NMR (CDCl , 200 MHz) δ 1.26 (6H,
3
d, J ) 6 Hz, H-3), 3.60 (1H, m, H-2), 7.26-8.75 (m, 9 H, ArH);
1
3
C NMR (CDCl
d, C-2), 122.5, 122.7, 126.3, 126.9, 127.0, 127.1, 127.3, 128.2,
29.3 (d, C 1′-8′, C-10′), 128.7, 130.0, 130.6, 131.3, 136.1 (s,
3
, 50 MHz) δ 18.5 (q, C-3 and 2-methyl), 39.7
(
1
2
39/97). We thank Profs. Dr. Angel Guti e´ rrez Ravelo (Instituto
de Bioorg a´ nica “Antonio Gonzalez”, Universidad de La Laguna,
Tenerife, Espa n˜ a) and Dr. Manuel Gonzalez Sierra (IQUIOS,
+
C-4′a,4′b, 8′a, 9′, 10′a), 209.0 (s, C-1); EIMS m/z [M ] 248 (20),
2
2
8
05 (100), 177 (49), 151 (9), 88 (13), 69 (78); HRFABMS m/z
1
CONICET-Universidad Nacional de Rosario) for the H and
+
49.1274 (calcd for C18
6.82%, H 6.57%, calcd for C18
-(9′-P h en a n t h r yl)-2-ch lor o-1-p r op a n on e (32). Com-
H
17O [M+H] , 249.1279); anal. C
1
3
C NMR spectra; Dr. Clara L o´ pez (CEREMIC, Universidad
H
16O, C 87.05%, H 6.50%.
Nacional de Rosario), for providing cultures of dermatophytes
and for her valuable assistance in the mycological work.
Collaboration from international project X-2 of RIPRONAMED
1
pound 32 was prepared from ketone 29 (200 mg, 0.81 mmol),
Cu(II) chloride dihydrate (236.5 mg, 1.4 mmol), lithium
chloride (46.2 mg, 1.1 mmol), and DMF (3 mL), following the
procedure described previously for compound 9. Compound 32
(Iberoamerican Network on Medicinal Natural Products) from
CYTED (Iberoamerican Program of Science and Technology
for Development, Subprogram X-10) is gratefully acknowl-
edged.
(187.6 mg, 0.7 mmol, 87% yield) was obtained as a white
-
1 1
solid: IR (KBr) νmax 3010, 2940, 1695 cm
2
7
; 3
H NMR (CDCl ,
00 MHz) δ 1.64 (3H, d, 3H, J ) 7 Hz, H-3), 5.41 (1H, q, J )
Hz, H-2), 7.70-8.72 (9 H, m, H-1′-7′, 10′, ArH); 13C NMR
, 50 MHz) δ 20.3 (q, C-3), 56.4 (d, C-2), 122.7-122.8,
Refer en ces a n d Notes
(CDCl
3
(
1) Walsh, T. In Emerging Targets in Antibacterial and Antifungal
Chemotherapy; Sutcliffe, J ., Georgopapadakou, N., Eds.; Chapman
and Hall: New York, 1992; Chapter 13, pp 349-373.
1
1
1
26.3, 127.2, 127.3, 127.6, 128.6, 129.0, 129.7 (d, C 1′-8′, C-10′),
30.2, 131.1, 132.2, 133.8, 134.0 (s, C-4′a,4′b, 8′a, 9′, 10′a),
97.33 (s, C-1); EIMS m/z 268 (32), 205 (100), 177 (56), 90 (31);
(2) Li, E.; Clark, A.; Hufford, C. J . Nat. Prod. 1995, 58, 57-67.
(
3) Zacchino, S.; Rodr ´ı guez, G.; Pezzenati, G.; Orellana, G.; Enriz, R.;
Gonzalez Sierra, M. J . Nat. Prod. 1997, 60, 659-662.
4) Gottlieb, O. J . Ethnopharmacol. 1979, 1, 309-323.
HREIMS m/z 268.0729 (calcd for C17
C %, H %, calcd for C17 13OCl; C 76.10%, H 4.89%, Cl 13.04%;
HPLC analysis, the observed t values for compound 32 in two
different solvent systems (MeOH-H O, 70:30 and CH CN-
O 60:40) were 5.38 min (100%) and 4.56 min (98.7%),
respectively.
H13OCl 268.0655); anal.
H
(
R
(5) Stoessl, A. Tetrahedron Lett. 1966, 21, 2287-2292.
(6) MacRae, W.; Towers, N. Phytochemistry 1984, 23, 1207-1220.
2
3
(7) Yoshida, M. Mem. Inst. Oswaldo Cruz (Rio de J aneiro) 1991, 86, 39-
H
2
4
2.
(8) Zemek, J .; Kosikova, B.; Augustin, J .; Koniak, D. Folia Microbiol.
1979, 24, 483-486.
Micr oor ga n ism s a n d Med ia . The following microorgan-
isms used for the fungistatic evaluation were purchased from
American Type Culture Collection (Rockville, MD): C. albicans
ATCC 10231, S. cerevisiae ATCC 9763, C. neoformans ATCC
(9) Hattori, M.; Hada, S.; Watahaki, H.; Shu, Y.; Kakiuchi, N.; Mizuno,
T.; Namba, T. Chem. Pharm. Bull. 1986, 34, 3885-3893.
10) Himejima, M.; Kubo, I. J . Nat. Prod. 1992, 55, 620-625.
11) Bohlman, F.; Zdero, C. Chem. Ber. 1976, 109, 791-792.
(
(
3
2264, A. flavus ATCC 9170, A. fumigatus ATCC 26934, and
(12) Dewick, P. Medicinal Natural Products: A Biosynthetic Approach;
J ohn Wiley and Sons: London, 1997; Chapter 4, pp 120-128.
A. niger ATCC 9029. Strains were grown on Sabouraud
chloramphenicol agar slants for 48 h at 30 °C. Cell suspensions
in sterile distilled water were adjusted to give a final concen-
tration of 106 viable yeast cells/mL.34 Dermatophytes: M. canis
C 112, T. rubrum C 113, E. floccosum C 114, and M. gypseum
C 115 are clinical isolates and were kindly provided by
CEREMIC, Centro de Referencia Micol o´ gica, Facultad de
Ciencias Bioqu ´ı micas y Farmac e´ uticas (Suipacha 531, 2000
Rosario, Argentina). T. mentagrophytes was ATCC 9972.
Organisms were maintained on slopes of Sabouraud dextrose
agar (SDA, Oxoid) and subcultured every 15 days to prevent
pleomorphic transformations. Spore suspensions were obtained
(
13) Forrest, J .; Heacock, R. Lloydia 1972, 35, 440-449.
(
14) Barata, L.; Baker, P.; Gottlieb, O.; R u´ veda, E. Phytochemistry 1978,
1
7, 783-786.
(15) Marston, A.; Hostettmann, K. J . Nat. Prod. 1995, 58, 128-130.
(
16) Mitscher, L.; Leu, R.; Bathala, M.; Wu, W.; Beal, J . Lloydia 1972,
3
5, 157-166.
(
17) Rishton, G. DDT 1997, 2, 382-384.
(
18) Kosower, E.; Cole, W.; Wu, G.; Cardy, D.; Meisters, G. J . Org. Chem.
1963, 28, 630-633.
(
19) Perry, C.; Kalnins, M.; Deitcher, K. J . Org. Chem. 1972, 37, 4371-
4
376.
(
20) Fuson, R.; Gaertner, R.; Chadwick, D. J . Org. Chem.1948, 13, 489-
495.
(21) Alesso, E.; Tombari, D.; Moltrasio, G. Can. J . Chem. 1987, 65, 2568-
3
4
6
2574.
according to reported procedures and adjusted to 10 spores
with colony-forming ability per mL.
(
22) Biftu, T.; Hazra, B.; Stevenson, R.; Williams, J . J . Chem. Soc., Perkin
Trans. 1 1978, 10, 1147-1150.
An tifu n ga l Assa ys. The fungistatic activity of phenylpro-
panoids was evaluated with the agar dilution method by using
Sabouraud chloramphenicol agar for both yeast and dermato-
phyte species. The assay was carried out in 96-well microtiter
plates. Stock solutions of phenylpropanoids in DMSO were
diluted to give serial two-fold dilutions that were added to each
medium resulting in concentrations ranging from 0.10 to 50
µg/mL. The final concentration of DMSO in the assay did not
exceed 2%. Using a micropipet, an inoculum of 5 µL of the
yeast cell or spore suspensions was added to each Sabouraud
chloramphenicol agar well. The antifungal agents ketoconazole
(23) Hunter, M.; Cramer, A.; Hibbert, H. J . Am. Chem. Soc. 1939, 61, 516-
520.
(
24) Obase, H.; Nakamizo, N.; Takai, H.; Teranishi, M.; Kubo, K.; Shuto,
K.; Kasuya, Y.; Kato, H.; Shigenober, K.; Krerihara J . Chem. Pharm.
Bull. 1982, 30, 462-473.
(25) Bohlman, F.; Zdero, C. Chem. Ber. 1976, 109, 791-792.
(
(
(
(
26) Norcross, G.; Openshaw, H. J . Chem. Soc. 1949, 1174-1179.
27) Mattson, M.; Rapoport, H. J . Org. Chem.1996, 61, 6071-6074.
28) Izumi, J .; Mukaiyama, T. Chem. Lett. 1996, 739-740.
29) Zacchino, S. J . Nat. Prod. 1994, 57, 446-451.
(30) Matsuda, J .; Sato, S. J Organomet. Chem. 1986, 314, 47-52.
(
(
31) Dosa P.; Fou, G. J . Am. Chem. Soc.1998, 120, 445-446.
32) Vidal Ferran, A.; Bampos, N.; Moyano, A.; Pericas, M.; Riera, A.;
Sanders, J . J . Org. Chem. 1998, 63, 6309-6318.
(J anssen Pharmaceutica) and amphotericin B (Sigma Chemi-
(33) Pini, D.; Iuliano, A.; Salvadori, P. Tetrahedron Asymmetry 1992, 13,
693-694.
(
cal Co) were included in the assay as positive controls. Drug-
free solution was also used as a blank control. The plates were
incubated 24, 48, or 72 h at 30 °C (according to the control
fungus growth) up to 15 days for dermatophyte strains. MIC
34) Wright, L.; Scott, E.; Gorman, S. J . Antimicrob. Chemother. 1983,
1
2, 317-327.
NP9805443