Synthesis and Antifungal Activity of Novel Bis(dithiocarbamate) Derivatives of Glycerol

Article abstract of DOI:10.1021/jf950751y

Three bis(dithiocarbamate) derivatives of glycerol were obtained in 30-95% yields by reaction of a dithiocarbamic acid salt with 1,3-dichloro-1,3-dideoxyglycerol. The in vitro antifungal activity of the new compounds was evaluated against Alternaria brassicae, Pseudocercosporella herpotrichoides, Septoria nodorum, and Phytophtora cinnamomi. Some of the compounds displayed greater activity than commercial fungicides. Bis(N,N-diethyldithiocarbamoyl)-1,3-dideoxyglycerol (4) was found to maintain its pesticidal activity up to 24 days in open field tests against A. brassicae.

Full text of DOI:10.1021/jf950751y

2856
J. Agric. Food Chem. 1996, 44, 2856−2858
Syn th esis a n d An tifu n ga l Activity of Novel Bis(d ith ioca r ba m a te)
Der iva tives of Glycer ol
Christophe Len, Anne-S. Boulogne-Merlot, Denis Postel, Gino Ronco, and Pierre Villa*
Laboratoire de Chimie Organique et Cine´tique, Universite´ de Picardie-J ules Verne, 80000 Amiens, France
Christel Goubert, Eric J eufrault, Bernard Mathon, and Herve´ Simon
Service Re´gional de la Protection des Ve´ge´taux, 80000 Amiens, France
Three bis(dithiocarbamate) derivatives of glycerol were obtained in 30-95% yields by reaction of a
dithiocarbamic acid salt with 1,3-dichloro-1,3-dideoxyglycerol. The in vitro antifungal activity of
the new compounds was evaluated against Alternaria brassicae, Pseudocercosporella herpotrichoides,
Septoria nodorum, and Phytophtora cinnamomi. Some of the compounds displayed greater activity
than commercial fungicides. Bis(N,N-diethyldithiocarbamoyl)-1,3-dideoxyglycerol (4) was found to
maintain its pesticidal activity up to 24 days in open field tests against A. brassicae.
Keyw or d s: Fungicides; bis(1,3-dithiocarbamoyl)-1,3-dideoxyglycerol; in vitro antifungal activity;
in vivo antifungal activity; open field test
Sch em e 1
Dithiocarbamates are important as all-purpose fun-
gicides used for the treatment of soil and seed and as
foliar sprays for the control of a large variety of diseases
affecting several types of crops. They are commonly
separated into two classes: (i) dialkyldithiocarbamates
including thiurame sulfide such as Ziram (zinc dimeth-
yldithiocarbamate); (ii) alkylene bis(dithiocarbamates)
and their oxidation products, such as Nabam (disodium
ethylenebis(dithiocarbamate)) (Thorn et al., 1962). A
problem associated with such salts is the possibility of
their containing or releasing trace toxic elements or
compounds (Woodrow et al., 1995; Buzasi-Gyorfy, 1992).
Also, it is possible that such compounds may not be used
to their full advantage due to the problems of transport
in the plant. Therefore, we have investigated an
alternative approach involving bis(dithiocarbamic es-
ters) of glycerol which are not associated with metal ions
and can act as propesticides.
throughout unless otherwise stated. 1H and 13C NMR chemical
shifts were reported in δ values based on the internal reference
tetramethylsilane. Column chromatography was carried out
with a Matrex silica gel 60 Å (70-230 mesh, Merck).
Syn th esis of N,N-Dieth yld ith ioca r ba m ic Acid Lith iu m
Sa lt (1). The synthesis of 1 was accomplished by adding
dropwise CS2 (10.6 g, 0.14 mol) to a solution of N,N-diethyl-
amine (10.0 g, 0.14 mol) in acetone (112 mL) at 8 °C. The
mixture was stirred for 10 min, and LiOH‚H2O (5.9 g, 0.14
mol) was added. Stirring was continued to ensure complete
salt formation (15 min). The solution was filtered to remove
solids. The organic filtrate was concentrated under reduced
pressure. The crude product was recrystallized from dioxane
to yield 18.2 g (86% yield) of 1. The structure was confirmed
by spectral analyses.
The derivatization of known pesticides to produce
novel propesticides having, for example, greater residual
effectiveness, continues to be a fruitful area of develop-
ment (Fahmy et al., 1981; Mallipudi et al., 1994).
Another strategy has been to derivatize carbamates
with molecules such as carbohydrates, amino acids,
plant hormones, and organic acids. These molecules
have relatively high polarity (Ziegler et al., 1975) and
are phloem mobile. Such derivatives have been shown
to increase plant systemic activity (J ojima et al., 1983).
By a similar approach, we have synthesized a series of
novel dithiocarbamic esters possessing the hydrophilic
moiety glycerol and two substituted dithiocarbamoyl
groups following Scheme 1.
Syn th esis of Mor p h o-4-yl- (2) a n d 1-P ip er id yl- (3)
Dith ioca r ba m ic Acid Lith iu m Sa lts. The above method
applied to morpholine and piperidine yielded the corresponding
morpho-4-yl- (2) and 1-piperidyldithiocarbamic lithium salt (3)
in 97 and 96% yield, respectively.
MATERIALS AND METHODS
Syn th esis of Bis-1,3-S-(N,N-d ieth yld ith ioca r ba m oyl)-
1,3-d id eoxyglycer ol (4). The synthesis of 4 was accom-
plished as shown in Scheme 1 adding dropwise 1 (3.6 g, 23.2
mmol) to a stirred solution of 1,3-dichloro-1,3-dideoxyglycerol
(1.0 g, 7.8 mmol) in 43 mL of acetone. Upon completion of
the addition, the reaction mixture was stirred at 56 °C for 1
h. The mixture was filtered and the solvent removed by
distillation under reduced pressure to yield a viscous oil. This
oily residue was treated with ether-hexane (1:1) and washed
twice with water and dried over anhydrous sodium sulfate.
Gen er a l. Technical grade of Carbendazime (I), Maneb (II),
and 1,3-dichloro-1,3-dideoxyglycerol obtained from a com-
mercial source (Aldrich) were used as starting material after
purification by recrystallization from appropriate solvents or
by distillation. Melting points were measured with an elec-
trothermal melting point apparatus and were uncorrected. 1H
(at 300.13 MHz) and 13C (at 75.47 MHz) NMR spectra were
recorded on a Bruker AM 300 spectrometer. Deuteriochloro-
form (99.8% atom enriched, Aldrich) was used as NMR solvent
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Bis(dithiocarbamate) Derivatives of Glycerol
J. Agric. Food Chem., Vol. 44, No. 9, 1996 2857
Ta ble 1. P h ysicoch em ica l Da ta for th e Bis(d ith ioca r ba m a tes) 4-6
compd MW (g‚mol-1
)
mp ( °C)
NMR data
1H δ 3.58 (dd 2H, J 1,1′ J 3,3′ 14.2 Hz, H-1-3); 3.45 (dd 2H, J 1,2 J 2,3 4.5 Hz, H-1′-3′); 4.07 (m 1H,
4
354.62
syrupy
liquid
J 1′,2 J 2,3′ 6.9 Hz, H-2); 3.86 (q 4H, J CH
7.1 Hz, NCH2); 3.66 (q 4H, J CH
7.1 Hz,NCH2);
2,CH3
2,CH3
1.17 (t 6H, CH3); 1.12 (t 6H, CH3); 13C δ 42.4 (2C C1-3); 70.2 (1C C2); 195.5 (2CCS); 49.9 (2C
NCH2); 47.0 (2C NCH2); 12.5 (2C CH3); 11.5 (2C CH3)
5
6
382.59
378.64
136-138 1H δ 3.79 (dd 2H, J 1,1′ J 3,3′ 14.2 Hz, H-1-3); 3.57 (dd 2H, J 1,2 J 2,3 4.7 Hz, H-1′-3′); 4.21 (m 1H,
J 1′,2 J 2,3′ 7.2 Hz, H-2); 4.29 (4H, NCH2); 3.97 (4H, NCH2); 3.73 (4H, CH2 p); 13C δ 42.2(2C C1-3);
70.0 (1C C2); 197.5 (2C CS); 51.0 (4C NCH2); 66.2 (4C OCH2)
122-124 1H δ 3.58 (dd 2H, J 1,1′ J 3,3′ 14.1 Hz, H-1-3); 3.43 (dd 2H, J 1,2 J 2,3 4.5 Hz, H-1′-3′); 4.06 (m 1H,
J 1′,2 J 2,3′ 6.9 Hz, H-2); 4.12 (4H, NCH2); 3.78 (4H, NCH2); 1.55 (12H, CH2); 13C δ 42.4 (2C C1-3);
70.0 (1C C2); 195.3 (2C CS); 53.3 (2C NCH2); 51.5 (2C NCH2); 25.6 (2C CH2 m); 24.1 (4C CH2 p
)
Ta ble 2. P er cen t Gr ow th In h ibition of A. br a ssica e
Ca u sed by th e Bis(d ith ioca r ba m a tes) 4-6
study were compared with those obtained from a parallel trial
in which III (100 g‚ha-1) and Kidan (Iprodione) (IV) (500
g‚ha-1) were used in commercialy available formulation. III
and IV were imposed as national homologation references for
fungicides against A. brassicae of colza (Regnault, 1983;
Bourdin and al., 1995). Different parcels of colza (45 m2) in
an open field were treated with four repetitions of 4, III, and
IV, respectively. After 3 days, all the treated parcels and the
four control parcels were artificially contaminated with A.
brassicae. Measurements were taken 24 days after the treat-
ment, noting for each condition the number of spots per silique
per parcel over a total of 50 siliques.
days
concn
compd
(ppm)
4
7
14
21
4
50
20
2
50
20
2
50
20
2
58
58
35
49
53
35
49
51
25
38
58
45
6
29
55
16
42
29
13
23
54
32
4
28
37
15
34
23
6
37
8
0
5
16
0
8
0
0
0
5
6
I
RESULTS AND DISCUSSION
50
23
In unpublished work, we demonstrated that 1-S-(N,N-
diethyldithiocarbamoyl)-1-deoxy-D,L-glycerol and other
dithiocarbamic ester derivatives of itols and carbohy-
drates showed modest inhibition of growth of A. bras-
sicae at different concentrations. The synthesis of
glycerol derivatives permitted the introduction of more
than one dithiocarbamic ester into a molecule. It was
envisaged that a multiple graft of dithiocarbamoyl
groups on a low molecular weight polyol leads to an
enhanced activity. Thus dithiocarbamic salts were
converted into bis(dithiocarbamic esters) of glycerol
following Scheme 1, using very mild conditions.
The growth inhibitory effect of 4-6 derivatives was
tested in vitro on PDA plates using DMSO as solvent.
This solvent was used since the bis(dithiocarbamates)
4-6 are only weakly soluble in most other common
solvents and it is well established that DMSO can be
used in pesticide bioassays (Stratton, 1985). Com-
mercial dithiocarbamic compounds were used to inhibit
the growth of a number of fungal species. Four species
were chosen for this study on the basis of the potential
economical advantage of successful treatments of crops
such as colza, corn, and potato. Antifungal activity of
4-6 was compared to that of the commercial I and II
products. All inhibition data were normalized as per-
centage inhibition compared to the control plates using
only DMSO. The three bis(dithiocarbamates) 4-6 did
not cause statistically significant growth inhibition of
P. herpotrichoides and P. cinnamomi at the three
concentrations tested (nontoxic at 2 ppm; growth inhibi-
tion < 20% at 50 ppm). We also examined the effect of
4-6 compared to I for A. brassicae (Table 2) and II for
S. nodorum (Table 3). The results noted in Table 2
showed that 4-6, in concentrations of 50 and 20 ppm,
were more antifungal than the commercial dithiocar-
bamate I at 50 ppm. The results described in Table 3
show that 4-6 had good antifungal activity against S.
nodorum at 50, 20, and 2 ppm. At the same concentra-
tion range, 4-6 showed higher activity than commercial
dithiocarbamate II at 50 ppm during the 21 day
observation.
The crude product was purified by column chromatography
on silica gel eluted with hexane-acetone (85:15) to give 2.6 g
(95%) of 4 (physicochemical data are reported in Table 1).
Syn th esis of Bis-1,3-S-(m or p h o-4-yld ith ioca r ba m oyl)-
1,3-d id eoxyglycer ol (5) a n d Bis-1,3-S-(1-p ip er id yld ith io-
ca r ba m oyl)-1,3-d id eoxyglycer ol (6). The above method
applied to lithium salts 2 and 3 yielded the corresponding bis-
[1,3-S-(morpho-4-yldithiocarbamoyl)]- (5) and bis[1,3-S-(1-pi-
peridyl dithiocarbamoyl)]-1,3-dideoxyglycerol (6) in 42 and 30%
yield, respectively (corresponding physicochemical are reported
in Table 1).
Stock Cu ltu r es. The fungi Alternaria brassicae, Pseudocer-
cosporella herpotrichoides, Septoria nodorum, and Phytophtora
cinnamomi were used as test organisms. Cultures were
obtained from the Institut National de la Recherche Agron-
omique (INRA Paris, France) and the Service Re´gional de la
Protection des Ve´ge´taux de Picardie (SRPV Picardie, France)
and maintained on potato dextrose agar (PDA) at 18 °C in a
phytotron room. Fungal growth plugs were cut using a 4.5
cm cork borer and transferred from stock plates to fresh agar
biweekly to maintain actively growing fungi.
Ra d ia l F u n ga l Gr ow th Assa ys. In vitro assays were
conducted on PDA at 18 °C in continuous light into phytotron
room. Prior to the PDA being poured into plastic petri plates
(100 × 15 mm), both fungicide solutions and solvent were
added to the molten PDA (50 °C) and mixed on a rotary shaker
for 2 min. Agar was poured into five control and five test
plates for each of the conditions used. Bis(dithiocarbamates)
4-6 and commercial fungicides I and II were tested against
all four fungi at concentrations of 50, 20, and 2 ppm in
dimethyl sulfoxide (DMSO). I, usually associating with Fluz-
ilazole Such Punch (III) for treatment of Alternaria and
Cercosporella diseases (Index Phytosanitaire, 1993), was used
for in vitro carbamic reference against A. brassicae and P.
herpotrichoides. II, known for fungicide activity against
Septoria and Phytophtora diseases, was selected for reference
with S. nodorum and P. cinnamomi. Small samples of each
fungi (30 mm diameter) were taken from the outer margin of
fresh stock culture plates and transferred to the center of each
medium. Measurements were taken three times daily over a
period of 4-21 days, noting the distance from the edge of the
fungal plug to the edge of the actively growing fungus.
Measurements ceased when fungal growth reached the edge
of the DMSO control plates.
In Vivo Assa y in a n Op en F ield . Compound 4 was tested
at concentration of 8.46 mol‚ha-1, which is the concentration
chosen for prelimary investigations. The results from this
Compound 4 was selected for an in vivo assay in an
open field of colza contaminated by A. brassicae. This

2858 J. Agric. Food Chem., Vol. 44, No. 9, 1996
Len et al.
Ta ble 3. P er cen t Gr ow th In h ibition of S. n od or u m
Ca u sed by th e Bis(d ith ioca r ba m a tes) 4-6
in pesticides and treated vegetables. Microchem. J . 1992,
46, 30-35.
Fahmy, M. A. H.; Fukuto, T. R. N-Sulfinylated derivatives of
methylcarbamate esters. J . Agric. Food Chem. 1981, 29,
567-572.
days
concn
(ppm)
compd
4
7
14
21
Index Phytosanitaire 1993; Association de Coordination Tech-
nique Agricole: Paris, 1993.
4
50
20
2
50
20
2
50
20
2
55
55
50
50
50
50
27
29
29
36
57
58
58
59
56
63
42
35
32
29
67
66
58
62
64
64
42
43
50
23
62
62
53
61
60
58
29
25
45
5
J ojima, T.; Fahmy, M. A. H.; Fukuto, T. R. Sugar, glyceryl,
and (pyridylalkoxy)sulfinyl derivative of methylcarbamate
insecticides. J . Agric. Food Chem. 1983, 31, 613-620.
Mallipudi, N. M.; Lee, A. H.; Kapoor, I. P.; Hollingshaus, G.
L. Synthesis and insecticidal activity of novel N-oxalyl-N-
methylcarbamates. J . Agric. Food Chem. 1994, 42, 1019-
1025.
Regnault, Y. Me´thode d’essai d’efficacite´ pratique des produits
fongicides destine´s a` lutter contre la maladie des taches
noires du colza, Alternaria brassicae Berk. Commission des
Essais Biologiques; Association Nationale pour la Protection
des Plantes, No. 101; 1983.
5
6
II
50
bioassay was also conducted with the two commercial
fungicides III and IV. The trial showed that 4 main-
tained its antifungal activity at level of 30% contami-
nated surface reduction, using a nonoptimized fomula-
tion. This result is lower than those obtained for III
(62%) and IV (57%); however, these were measured
using an optimized formulation.
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ACKNOWLEDGMENT
We are grateful to G. Mackenzie for reading the
paper.
Ziegler, H.; Zimmerman, M. H.; Milburn, J . A. Transport in
plant I; Springer-Verlag: Berlin, 1975; Chapter 3.
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Received for review November 13, 1995. Revised manuscript
received May 30, 1996. Accepted J une 6, 1996.X We are
grateful to the Centre de Valorisation des Glucides and the
Biopoˆle de Picardie for financial support.
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