Synthesis of novel chelating agents and their effect on cadmium decorporation

Article abstract of DOI:10.1021/tx970134z

A series of novel dithiocarbamates, disodium salts of N-glucamyl-N- dithiocarboxyl-amino acids, were synthesized, and their usefulness as an antagonist of cadmium intoxication was investigated. These chelating agents were found to be effective in both acute and repeated exposure cadmium poisoning. The results showed that the cadmium mobilizing properties of disodium N-(2,3,4,5,6-pentahydroxylhexyl)-N-dithiocarbamate-L-threoninate and disodium N-(2,3,4,5,6-pentahydroxylhexyl)-N-dithiocarbamate-L-cysteinate are clearly superior to those of sodium N-(4-methoxybenzyl)-D-glucamine-N- carbodithioate (MeOBGDTC) revealed in the experiments described here. The toxicity of these novel compounds is modest, and their effect on the concentrations of essential metal ions in the renal cortex is quite small in comparison with that of a group treated with cadmium only. The new dithiocarbomates were identified by MS, rather than by elemental analysis, as they were extremely hygroscopic.

Full text of DOI:10.1021/tx970134z

Chem. Res. Toxicol. 1999, 12, 331-334
331
Syn th esis of Novel Ch ela tin g Agen ts a n d Th eir Effect on
Ca d m iu m Decor p or a tion
Chao Wang, Yiou Fang, and Shiqi Peng*
School of Pharmaceutical Sciences, Beijing Medical University, Beijing 100083, PRC
Dongxin Ma and J inyuan Zhao
Research Center of Occupational Medicine, Beijing Medical University, Beijing 100083, PRC
Received J uly 29, 1997
A series of novel dithiocarbamates, disodium salts of N-glucamyl-N-dithiocarboxyl-amino
acids, were synthesized, and their usefullness as an antagonist of cadmium intoxication was
investigated. These chelating agents were found to be effective in both acute and repeated
exposure cadmium poisoning. The results showed that the cadmium mobilizing properties of
disodium N-(2,3,4,5,6-pentahydroxylhexyl)-N-dithiocarbamate-L-threoninate and disodium
N-(2,3,4,5,6-pentahydroxylhexyl)-N-dithiocarbamate-L-cysteinate are clearly superior to those
of sodium N-(4-methoxybenzyl)-D-glucamine-N-carbodithioate (MeOBGDTC) revealed in the
experiments described here. The toxicity of these novel compounds is modest, and their effect
on the concentrations of essential metal ions in the renal cortex is quite small in comparison
with that of a group treated with cadmium only. The new dithiocarbomates were identified by
MS, rather than by elemental analysis, as they were extremely hygroscopic.
But it enhanced the cadmium concentration in the brain,
because Cd-DEDTC complexes are so lipid soluble that
they can penetrate the blood-brain barrier (8). It was
also shown that this could be prevented by appropriate
substitution of polar groups in the dithiocarbamate (9).
The studies showed that sodium N-methyl-D-glucamine-
N-carbodithioate (MGDTC) possessed low toxicity and
was able to remove deposited cadmium from kidney and
liver in mice (10).
Sodium N-benzyl-D-glucamine-N-carbodithioate (BG-
DTC) and a series of the corresponding substituted
aromatic compounds were prepared, and the results show
that they were superior to MGDTC for cadmium decor-
poration (11). Unfortunately, the toxicity of these aro-
matic compounds was higher than that of MGDTC (10,
11). It is obvious that the toxicity stems from the moiety
of the aromatic structure.
In view of these reasons, we decided to prepare com-
pounds of an analogous series which used amino acids
as the source of the amine groups. The structures of the
compounds prepared are shown in Figure 1. Compared
with BGDTC, the glucamine moiety was retained and the
benzylamine moiety was replaced by an amino acid-
derived group. When cysteine is the amino acid, the
chelating agent produced also contains a mercapto group,
which may also act to coordinate the metal ion.
The purpose of this study was to develop novel dithio-
carbamate chelating agents for in vivo cadmium mobi-
lization. Specifically, we hoped to obtain compounds
which were more effective and less toxic than those pre-
viously reported. The reactions used in the preparation
of these novel chelating agents are shown in Figure 1.
In tr od u ction
Of the various metals which are environmentally signi-
ficant as toxic species, cadmium plays an unusual role
because of the obvious accumulation in the human body.
Cadmium possesses an ability to settle intracellularly by
binding with a low-molecular weight protein, metallo-
thionein (MT) (1). The accumulation of cadmium in the
human body, mainly in liver and kidney, will lead to
serious cadmium intoxication, especially the toxic renal
damage. Therefore, it is important to develop effective,
safe chelating agents for the therapy of cadmium intoxi-
cation.
Ethylenediaminetetraacetic acid (EDTA), 2,3-dimer-
captopropanol (BAL), and penicillamine (PA) are widely
used for the treatment of metal intoxication (2). Mercapto
compounds, BAL or PA, can chelate cadmium strongly
by the low-molecular weight ligands, but this kind of
cadmium complex will be reabsorbed by renal tubules.
It may increase the level of kidney deposition and also
enhance the kidney toxicity of cadmium (3, 4). On the
other hand, the commonly used hydrophilic aminopoly-
carboxylic acids, such as EDTA and DTPA (diethylene-
triaminepentaacetic acid), can form more stable com-
plexes with cadmium, but they only have a very slight
effect on the cadmium deposited in kidney, because these
chelating agents are unable to enter the intracellular
sites (5), and have a low selectivity for cadmium as metal
ions (6).
The ability of sodium diethyldithiocarbamate (DEDTC)
to act as an antagonist for acute cadmium chloride
intoxication was first reported by Gale (7). Subsequent
studies have confirmed and extended these results and
showed that dithiocarbamates are capable of reacting
with and mobilizing intracellular deposition of cadmium.
Exp er im en ta l Section
Ch em ica l Syn th esis. L-Amino acids were purchased from
* To whom correspondence should be addressed.
Sigma Chemical Co. Spectroscopic data and physical and
10.1021/tx970134z CCC: $18.00 © 1999 American Chemical Society
Published on Web 03/20/1999

332 Chem. Res. Toxicol., Vol. 12, No. 4, 1999
Wang et al.
Ta ble 1. Liver Ca d m iu m Levels a fter Tr ea tm en t w ith
Ch ela tin g Agen ts d u r in g Acu te In toxica tion a
4 h
24 h
48 h
Cd only
Cd and
MeOBGDTC
Cd and 3a
Cd and 3b
Cd and 3c
Cd and 3d
Cd and 3e
24.4 ( 0.69
24.4 ( 3.82
20.4 ( 0.48
20.1 ( 0.59
15.4 ( 1.79b,d 13.5 ( 0.86b,f
24.1 ( 1.36
21.9 ( 1.23
18.7 ( 4.56
28.5 ( 0.92
14.8 ( 0.52b,f 15.3 ( 0.55b,f
17.9 ( 0.90b,d 14.6 ( 0.99b,e
16.6 ( 1.03b,e 14.3 ( 1.02b,e
22.1 ( 0.58
9.9 ( 0.38b,c,e,f
13.0 ( 1.42b-d,f 15.8 ( 0.56b,f 11.0 ( 0.38b-d,f
a
Micrograms per gram of tissue weight (X ( SE) where n ) 5.
b Significantly less than that with Cd only. c Significantly less than
that with MeOBGDTC. P < 0.05. e P < 0.01. f P < 0.001.
d
Ta ble 2. Ren a l Cor tica l Ca d m iu m Levels a fter Tr ea tm en t
w ith Ch ela tin g Agen ts d u r in g Acu te In toxica tion a
4 h
24 h
48 h
Cd only
Cd and
MeOBGDTC
Cd and 3a
Cd and 3b
Cd and 3c
Cd and 3d
Cd and 3e
25.5 ( 0.66
20.6 ( 0.66
18.4 ( 1.32
14.6 ( 0.34b,f
15.0 ( 1.23b,e 13.1 ( 0.71b,e
17.6 ( 0.67b,d 16.6 ( 0.64
17.4 ( 0.62b,f
20.2 ( 1.01b,e
15.8 ( 0.61b,f
21.2 ( 0.85
20.9 ( 0.62
12.9 ( 0.47b,f
10.8 ( 0.42b-d,f
12.2 ( 0.47b,e
F igu r e 1. Synthetic scheme used for the preparation of the
novel chelating agents.
12.1 ( 0.68b,c,e,f 15.5 ( 0.61b,f
9.8 ( 0.90b,c,e,f 11.2 ( 0.94b-d,f 11.7 ( 0.06b,f
a
Micrograms per gram of tissue weight (X ( SE) where n ) 5.
analytical data were obtained on the instruments listed: IR,
Perkin-Elmer 983; NMR, Bruker AM-500; MS, VG-ZAB-MS;
elemental analysis, PE-2400; molecular rotation, Polartronic-
D; and HPLC, Waters 510.
Sod iu m N-(2,3,4,5,6-P en t a h yd r oxylh exylid en e)glyci-
n a te (1a ). D-(+)-Glucose (5.94 g, 30 mmol), glycine (2.25 g, 30
mmol), and NaOH (1.2 g, 30 mmol) were allowed to react in
water (3 mL) at 60 °C for 5 h to give a gel: FAB/MS (m/e) 260
[M + H]+.
b Significantly less than that with Cd only. c Significantly less than
that with MeOBGDTC. P < 0.05. e P < 0.01. f P < 0.001.
d
Sod iu m N-(2,3,4,5,6-p en ta h yd r oxylh exylid en e)-L-th r eo-
n in a te (1d ): FAB/MS (m/e) 304 [M + H]+.
Sod iu m N-(2,3,4,5,6-p en t a h yd r oxylh exylid en e)-L-cys-
tein a te (1e): FAB/MS (m/e) 306 [M + H]+.
N-(2,3,4,5,6-P en ta h yd r oxylh exyl)-L-a la n in e (2b): yield
57% (two steps); mp 201-203 °C; FAB/MS (m/e) 254 [M + H]+;
1H NMR (D2O) δ 3.51 (m, 2H, 6-H), 3.62 (m, 1H, 5-H), 3.68 (dd,
J ) 10.5, 3.0 Hz, 1H, 4-H), 3.69 (t, J ) 2.6 Hz, 1H, 3-H), 3.96
(m, 1H, 2-H), 3.03 (dd, J ) 12.8, 9.7 Hz, 1H, 1-H), 3.14 (dd, J )
12.8, 3.2 Hz, 1H, 1-H), 3.61 (q, J ) 3.1 Hz, 1H, Ala-CH), 1.38
(d, 3H, CH3); IR (KBr) 3410, 3270, 2972, 2940, 1903, 1621, 1585,
N-(2,3,4,5,6-P en ta h yd r oxylh exyl)glycin e (2a ). The crude
imine (1a ) was reduced to amine by NaBH4 (3.0 g) at room
temperature for 120 h. The reaction mixture was acidified by
HCl (6 N) to pH 2 at 0 °C. This solution was concentrated to a
syrup. Then, 30 mL of methanol was added, and the inorganic
salts were separated by filtration. The filtrate can be recrystal-
lized several times in H2O/EtOH to give a white crystalline solid
(yield of 60%, two steps): mp 182-183 °C; FAB/MS (m/e) 240
[M + H]+; 1H NMR (D2O) δ 3.52 (m, 2H, 6-H), 3.62 (m, 1H, 5-H),
3.68 (dd, J ) 11.0, 3.5 Hz, 1H, 4-H), 3.70 (t, J ) 2.0 Hz, 1H,
3-H), 3.99 (m, 1H, 2-H), 3.07 (dd, J ) 13.5, 9.5 Hz, 1H, 1-H),
3.16 (dd, J ) 13.0, 3.0 Hz, 1H, 1-H), 3.54 (d, J ) 2.0 Hz, 2H,
Gly-CH2); IR (KBr) 3222, 3020, 2926, 1617, 1563, 1467, 1377,
1245, 1122, 1092, 1059, 1037 cm-1. Anal. Calcd for C8H17N1O7:
C, 40.17; H, 7.16; N, 5.86. Found: C, 40.37; H, 6.90; N, 5.64.
Disod iu m N-(2,3,4,5,6-P en t a h yd r oxylh exyl)-N-d it h io-
ca r ba m a te-glycin a te (3a ). 2a (3.0 g, 12.6 mmol), NaOH (0.5
g, 12.6 mmol), and 10 mL of water were stirred under argon at
0 °C. CS2 (1.9 g, 25.0 mmol) in dioxane (10 mL) and NaOH (0.5
g, 12.6 mmol) in water (5 mL) were added dropwise to the clear
reaction mixture, while it was being stirred at 0 °C over the
course of 1 h, and then further stirred overnight at room
temprature. The mixture was put under reduced pressure to
remove excess CS2, and then the resulting solution was frozen
and lyophilized to obtain the crude product. It was recrystallized
from H2O/EtOH to give a yellow solid (yield of 89%): mp 100-
102 °C dec; FAB/MS (m/e) 358 [M - H]-; IR (KBr) 3408, 2960,
1479, 1421, 1396, 1367, 1287, 1267, 1146, 1131, 1014, 1003 cm-1
.
Anal. Calcd for C9H19N1O7: C, 42.68; H, 7.56; N, 5.53. Found:
C, 42.39; H, 7.33; N, 5.55.
N-(2,3,4,5,6-P en ta h yd r oxylh exyl)-L-p h en yla la n in e (2c):
yield 38% (two steps); mp 213-215 °C; FAB/MS (m/e) 330 [M +
H]+; 1H NMR (D2O) δ 3.48 (m, 2H, 6-H), 3.58 (m, 1H, 5-H), 3.65
(dd, J ) 11.2, 3.0 Hz, 1H, 4-H), 3.66 (t, J ) 2.2 Hz, 1H, 3-H),
3.91 (m, 1H, 2-H), 2.97 (dd, J ) 12.5, 9.5 Hz, 1H, 1-H), 3.05
(dd, J ) 12.5, 3.2 Hz, 1H, 1-H), 3.82 (t, J ) 6.0 Hz, 1H, Phe-
CH), 3.10 (d, J ) 5.5 Hz, 2H, CH2), 7.15-7.30 (5H, Ar-H); IR
(KBr) 3361, 3105, 3025, 2922, 2655, 1617, 1491, 1430, 1371,
1297, 1249, 1218, 1122, 1081, 1043 cm-1. Anal. Calcd for
C
15H23N1O7‚1/2HCl: C, 51.83; H, 6.82; N, 4.03. Found: C, 52.20;
H, 6.97; N, 3.99.
N-(2,3,4,5,6-P en ta h yd r oxylh exyl)-L-th r eon in e (2d ): yield
51% (two steps); mp 219-221 °C; FAB/MS (m/e) 284 [M + H]+;
1H NMR (D2O) δ 3.44 (m, 2H, 6-H), 3.55 (m, 1H, 5-H), 3.61 (dd,
J ) 11.7, 2.8 Hz, 1H, 4-H), 3.63 (t, J ) 2.1 Hz, 1H, 3-H), 3.94
(m, 1H, 2-H), 2.99 (dd, J ) 13.7, 10.0 Hz, 1H, 1-H), 3.09 (dd, J
) 12.9, 3.2 Hz, 1H, 1-H), 3.30 (d, J ) 7.7 Hz, 1H, Thr-CH), 3.88
(m, 1H, Thr-CH), 1.14 (d, J ) 6.5 Hz, 3H, CH3); IR (KBr) 3383,
3250, 2982, 2944, 1570, 1446, 1391, 1325, 1303, 1208, 1133,
1109, 1057, 1035 cm-1. Anal. Calcd for C10H21N1O8: C, 42.38;
H, 7.47; N, 4.95. Found: C, 42.35; H, 7.05; N, 4.74.
N-(2,3,4,5,6-P en ta h yd r oxylh exyl)-L-cystein e (2e): yield
35% (two steps); mp 197-199 °C; FAB/MS (m/e) 286 [M + H]+;
1H NMR (D2O) δ 3.41 (m, 2H, 6-H), 3.49 (m, 1H, 5-H), 3.55 (dd,
J ) 10.5, 3.6 Hz, 1H, 4-H), 3.58 (t, J ) 2.5 Hz, 1H, 3-H), 3.90
(m, 1H, 2-H), 2.95 (dd, J ) 12.9, 8.4 Hz, 1H, 1-H), 3.09 (dd, J )
12.8, 3.6 Hz, 1H, 1-H), 3.72 (t, J ) 5.1 Hz, 1H, Cys-CH), 2.86
(dd, J ) 9.0, 4.8 Hz, 2H, Cys-CH2); IR (KBr) 3271, 2932, 1603,
1588, 1460, 1377, 1209, 1169, 1055 cm-1
.
Compounds 1b-e, 2b-e, and 3b-e were prepared in a
manner similar to that described above, and the corresponding
yield, physical data, and analytical results for each are shown
as follows.
Sod iu m N-(2,3,4,5,6-p en ta h yd r oxylh exylid en e)-L-a la n i-
n a te (1b): FAB/MS (m/e) 274 [M + H]+.
Sod iu m N-(2,3,4,5,6-p en ta h yd r oxylh exylid en e)-L-p h en -
yla la n in a te (1c): FAB/MS (m/e) 350 [M + H]+.

Synthesis of Novel Chelating Agents
Chem. Res. Toxicol., Vol. 12, No. 4, 1999 333
FAB/MS (m/e) 402 [M - H]-, 380 [M - Na]- ; IR (KBr) 3334,
Ta ble 3. Blood Ca d m iu m Levels a fter Tr ea tm en t w ith
Ch ela tin g Agen ts d u r in g Acu te In toxica tion a
2923, 1588, 1388, 1227, 1079 cm-1
.
Disod iu m N-(2,3,4,5,6-p en t a h yd r oxylh exyl)-N-d it h io-
ca r ba m a te-L-cystein a te (3e): yield 62%; mp 104-106 °C dec;
FAB/MS (m/e) 404 [M - H]-; IR (KBr) 3368, 2923, 1600, 1386,
4 h
24 h
48 h
Cd only
Cd and
MeOBGDTC
Cd and 3a
Cd and 3b
Cd and 3c
Cd and 3d
Cd and 3e
0.06 ( 0.02
<0.01 ( 0.00 <0.01 ( 0.00
<0.01 ( 0.00 <0.01 ( 0.00
0.18 ( 0.03b,f
1224, 1178, 1083 cm-1
.
The preparations of MeOBGDTC were made following the
methods of J ones and co-workers (11). The product was char-
acterized by negative FAB/MS and elemental analysis.
An im a l Stu d ies. Acu te stu d ies. Adult male Wistar rats
weighing 250 ( 50 g were used for experiments. Each of them
was given a single 15 µmol/kg injection (ip) of cadmium chloride
mixed with mercaptoethanol (ME) at 300 µmol/kg in a final
volume of 2.5 mL of saline. Two hours later, they were further
treated (ip) with chelating agents, 0.25 mmol/kg in a final
volume of 2.5 mL of water. The cadmium-only group was given
a 0.9% saline injection (ip). Biosamples were collected 4, 24, and
48 h after cadmium injection. Rats were killed under pentobar-
bital anesthesia (ip); blood (by cardiac puncture), liver, and
kidney samples were then obtained, and urine samples were
periodically collected from individual metabolic cages.
Rep ea ted Exp osu r e Stu d ies. Male ICR mice weighing 25
( 2 g were loaded with cadmium via a series of injections (ip)
by administering four 3 mg injections of CdCl2‚2.5H2O/kg in 0.5
mL of 0.9% saline on four consecutive days. After a 3 day
interval, the animals were randomly divided into groups of seven
each. One group served as the cadmium-only group and was
given 0.9% saline injections (ip) instead of chelating agent. Each
member of the other groups was given an injection (ip) of one
of the chelating agents at a level of 1.0 mmol/kg in 0.5 mL of
water each day for five consecutive days. Two days after the
last injection of chelating agent, all the animals were sacrificed
by cervical dislocation and the livers and kidneys immediately
excised.
Biosamples were digested in H2SO4, HClO4, and HNO3 (1:1:
3) on a heating block, dried at 80 °C, redissolved in 1% nitric
acid, and analyzed for cadmium content using a HITACHI 180-
80 atomic absorption spectrometer in the flame mode. The
variety of trace metals were determined by synchrotron X-ray
fluorescence. The LD50 values were determined by single injec-
tion (ip) in mice.
All animals were kept in an AALAC approved animal care
facility during the course of the experiments and were provided
with free access to food and water. The statistical analysis of
the data was carried out by using standard analysis of variance
methods.
0.91 ( 0.10b,c,f
0.28 ( 0.04b,f
0.13 ( 0.03b,d
<0.01 ( 0.00 <0.01 ( 0.00
<0.01 ( 0.00 <0.01 ( 0.00
0.01 ( 0.00
0.05 ( 0.01
0.88 ( 0.04b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
0.49 ( 0.02b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
Micrograms per milliliter (X ( SE) where n ) 5. b Significantly
a
more than that with Cd only. c Significantly more than that with
MeOBGDTC. P < 0.05. e P < 0.01. f P < 0.001.
d
Ta ble 4. Ur in a r y Ca d m iu m Levels a fter Tr ea tm en t w ith
Ch ela tin g Agen ts d u r in g Acu te In toxica tion a
4 h
24 h
48 h
Cd only
Cd and
MeOBGDTC
Cd and 3a
Cd and 3b
Cd and 3c
Cd and 3d
Cd and 3e
0.03 ( 0.004
<0.01 ( 0.00 <0.01 ( 0.00
<0.01 ( 0.00 <0.01 ( 0.00
0.10 ( 0.001b,f
1.68 ( 0.37b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
0.59 ( 0.07b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
3.60 ( 0.41b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
1.82 ( 0.21b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
6.25 ( 0.26b,c,e,f <0.01 ( 0.00 <0.01 ( 0.00
Micrograms per milliliter (X ( SE) where n ) 5. b Significantly
a
more than that with Cd only. c Significantly more than that with
MeOBGDTC. P < 0.05. e P < 0.01. f P < 0.001.
d
Ta ble 5. Ca d m iu m Levels follow in g Ch ela te Tr ea tm en t of
Mice in Rep ea ted Exp osu r e Mod ea
liver
kidney
Cd only
Cd and MeOBGDTC
Cd and 3a
Cd and 3b
Cd and 3c
33.4 ( 0.30
20.1 ( 0.21
15.6 ( 0.22b,f
19.3 ( 0.25b,f
18.9 ( 1.07b,f
16.6 ( 0.48b,f
13.3 ( 0.24b-d,f
14.6 ( 0.38b-d,f
5.6 ( 0.35b,f
8.2 ( 0.43b,f
10.1 ( 0.27b,f
5.0 ( 0.89b,f
8.50 ( 0.069b,f
4.2 ( 0.23b,c,e,f
Cd and 3d
Cd and 3e
a
Micrograms per gram of tissue weight (X ( SE) where n ) 8.
b Significantly less than that with Cd only. c Significantly less than
d
that with MeOBGDTC. P < 0.05. e P < 0.01. f P < 0.001.
1565, 1414, 1388, 1350, 1290, 1131, 1082, 1036 cm-1. Anal.
Calcd for C9H19N1O7S1: C, 37.89; H, 6.71; N, 4.91. Found: C,
37.69; H, 6.43; N, 4.91.
Resu lts a n d Discu ssion
Disod iu m N-(2,3,4,5,6-p en t a h yd r oxylh exyl)-N-d it h io-
ca r ba m a te-L-a la n in a te (3b): yield 89%; mp 103-105 °C dec;
FAB/MS (m/e) 372 [M - H]-, 350 [M - Na]-; IR (KBr) 3378,
The results of the acute studies are summarized in
Table 1-4. The data in Tables 1 and 2 indicate that all
of the compounds were reasonably effective antagonists
under this experimental condition and significantly
reduced the level of cadmium in liver and kidney in
comparison with the control (Cd only) in most situations.
Compound 3d in 48 h and compound 3e in 4 or 48 h are
clearly superior to MeOBGDTC with respect to liver
2923, 1584, 1389, 1254, 1171, 1076 cm-1
.
Disod iu m N-(2,3,4,5,6-p en t a h yd r oxylh exyl)-N-d it h io-
ca r ba m a te-L-p h en yla la n in a te (3c): yield 80%; mp 145-147
°C dec; FAB/MS (m/e) 448 [M - H]-, 426 [M - Na]- ; IR (KBr)
3350, 2925, 1588, 1449, 1381, 1225, 1160 cm-1
.
Disod iu m N-(2,3,4,5,6-p en t a h yd r oxylh exyl)-N-d it h io-
ca r ba m a te-L-th r eon in a te (3d ): yield 87%; mp 78-81 °C dec;
Ta ble 6. In flu en ce of Ch ela tin g Agen ts on th e Ren a l Cor tex Con cen tr a tion of Essen tia l Meta ls in Ra tsa
Zn
Cu
Fe
Mn
Ca
Rb
Cd only
4 h
24 h
4 h
24 h
4 h
24 h
4 h
24 h
4 h
21.8 ( 1.24
21.8 ( 3.27
22.0 ( 0.32
28.4 ( 1.03
21.0 ( 1.00
22.6 ( 0.68
22.0 ( 0.55
19.0 ( 0.84
19.8 ( 0.58
23.8 ( 0.49
5.4 ( 0.25
6.6 ( 0.51
5.5 ( 0.58
7.0 ( 0.97
6.0 ( 0.55
6.6 ( 0.25
10.4 ( 0.40
5.4 ( 0.25
5.4 ( 0.25
5.2 ( 0.37
50.2 ( 1.93
45.2 ( 3.60
45.4 ( 5.03
54.4 ( 5.84
47.2 ( 3.26
44.4 ( 2.21
37.8 ( 0.37
56.0 ( 1.45
51.4 ( 5.54
61.4 ( 3.06
1.2 ( 0.20
1.0 ( 0.00
1.0 ( 0.00
1.2 ( 0.20
1.0 ( 0.00
1.0 ( 0.00
1.2 ( 0.20
1.2 ( 0.20
0.8 ( 0.20
1.2 ( 0.20
62.4 ( 3.93
126.6 ( 47.3
51.8 ( 3.57
77.4 ( 18.9
53.8 ( 7.59
61.2 ( 4.22
66.2 ( 6.14
62.0 ( 2.72
65.0 ( 4.18
79.6 ( 7.36
9.6 ( 0.60
8.0 ( 0.71
10.0 ( 0.55
12.4 ( 0.68
14.4 ( 0.75
8.4 ( 0.40
16.6 ( 0.25
8.2 ( 0.37
9.0 ( 0.71
9.0 ( 0.00
Cd and MeOBGDTC
Cd and 3c
Cd and 3d
Cd and 3e
24 h
a
Micrograms per gram of tissue weight (X ( SE) where n ) 5.

334 Chem. Res. Toxicol., Vol. 12, No. 4, 1999
Wang et al.
cadmium decorporation. Compound 3c in 48 h, compound
3d in 4 h, and compound 3e in 4 or 24 h are clearly
superior to MeOBGDTC with respect to kidney cadmium
decorporation.
level of efficacy. In the structures of these new compounds
presented here, the additional groups come from the side
chain of the amino acids, and according to the theory of
coordination chemistry, some heteroatoms can participate
in coordination with heavy metals to form stable com-
plexes. That probably is the reason why the effects of 3d
and 3e are more beneficial than the effects of 3a and 3b
on cadmium excretion.
The further study on representative compounds showed
that the LD50 of 3d was 10.5 mmol/kg (ip) and that of 3e
8.6 mmol/kg (ip). The LD50 values for 3d and 3e are
greater than that of PA [2.53 mmol/kg (ip)] and DEDTC
[8.35 mmol/kg (ip)] (16).
The data in Tables 3 and 4 are the concentrations of
cadmium in blood and urine. As illustrated in Table 3,
the concentrations of cadmium in blood were increased
2 h after injection of the chelators, which presumably
reflects in part mobilization of cadmium from tissues. The
effects of all chelating agents on the urinary excretion of
cadmium in rats are shown in Table 4. It was a gross
increase in the level of urinary excretion of cadmium with
use of chelators in the 4 h group. The increased level of
urinary excretion of cadmium by all of the new com-
pounds is significantly more then that by MeOBGDTC.
These results suggest that such structures of disodium
salts with good water solubility can make cadmium
complexes more easily excreted in urine.
Thus, these novel dithiocarbamates deserve further
consideration as members of a small but growing group
of compounds which may eventually be proved to be
suitable antidotes for cadmium intoxication.
The all new dithiocarbamates also proved to be effica-
cious in repeated exposure cadmium intoxication as
shown in Table 5. All of the compounds were found to be
able to remove cadmium from the liver and kidney.
Compounds 3d and 3e are superior to MeOBGDTC in
removing cadmium from the liver, and 3e is clearly
superior in removing cadmium from the kidney. Within
24 h of the injection of cadmium into a rodent, the
majority of the element is present in intracellular sites
(12), and at the end of 3 days, most of this intracellular
cadmium is present as the complex with MT (13). So the
results in repeated exposure studies indicate that these
compounds are able to remove cadmium from its deposits
in both the liver and kidney.
Refer en ces
(1) Webb, M. (1986) Role of metallothionein in cadmium metabolism.
In Cadmium (Foulkes, E. C., Ed.) pp 281-337, Springer-Verlag,
Berlin.
(2) Andersen, O. (1989) Oral cadmium exposure in mice: Toxicoki-
netics and efficiency of chelating agents, Toxicology 20, 83-112.
(3) Dalhamn, T., and Friberg, F. (1955) Dimercaprol (2,3-dimercap-
topropanol) in chronic cadmium poisoning. Acta Pharmacol. 11,
68-78.
(4) Lyle, W. H., Green, J . N., Gore, V., and Vidler, J . (1968)
Enhancement of cadmium nephrotoxicity by penicillamine in the
rat. Postgrad. Med. J ., Suppl., 18-25.
(5) Waalkes, M. P., Watkins, J . B., and Klaassen, C. D. (1983)
Minimal role of metallothionein in decreased chelator efficacy for
cadmium. Toxicol. Appl. Pharmacol. 56, 392-398.
(6) Anderson, O. (1984) Chelation of cadmium. Environ. Health
Perspect. 54, 249-257.
The results in Table 6 showed that the concentration
of Zn, Cu, Fe, Mn, Ca, and Rb in the kidney of rats after
treatment with MeOBGDTC and 3c-e exhibited little
change as compared with that of the group treated with
cadmium only. The data in Table 6 were recorded for
cadmium-intoxicated animals which have been treated
with cadmium, described as acute studies.
A problem in this experiment is these new chelating
agents were extremely hygroscopic in air. Therefore, the
repeatability of elemental analysis was very low. Fortu-
nately, the structures of these compounds were identified
by negative FAB/MS, and the purity was ascertained by
high-performance liquid chromatography, with a purity
of >98%.
Compound 2e or 3e must be isolated from air to
prevent oxidation while it is being prepared because the
mercapto group can transform into disulfide easily under
alkaline conditions.
The experimental model with a single injection (ip) of
CdCl2 and ME in rats is simple, rapid, and useful for the
screening of the new chelators, and it can clearly reflect
cadmium-induced renal damage within 24 h of cadmium
exposure (14).
(7) Gale, G. R., Smith, A. B., and Walker, E. M. (1981) Diethyldithio-
carbamate in the treatment of acute cadmium poisoning. Ann.
Clin. Lab. Med. 11, 476-483.
(8) J ones, S. G., Basinger, M. A., J ones, M. M., and Gibbs, S. J . (1982)
Polarity and antidotal efficacy of dithiocarbamates in acute
cadmium intoxication. Res. Commun. Chem. Pathol. Pharmacol.
40, 155-164.
(9) J ones, S. G., and J ones, M. M. (1984) Structure-activity relation-
ships among dithiocarbamate antidotes for acute cadmium chlo-
ride intoxication. Environ. Health Perspect. 54, 285-290.
(10) Shinobu, L. A., J ones, S. G., and J ones, M. M. (1984) Sodium
N-methyl-D-glucamine dithiocarbamate and cadmium intoxica-
tion. Acta Pharmacol. Toxicol. 54, 189-194.
(11) J ones, S. G., Singh, P. K., and J ones, M. M. (1988) Use of the
Topliss scheme for the design of more effective chelating agents
for cadmium decorporation. Chem. Res. Toxicol. 1, 234-237.
(12) Shaikh, Z. A., and Lucis, O. J . (1972) Biological differences in
cadmium and zinc turnover. Arch. Environ. Health 24, 410-418.
(13) Cantilena, L. R., J r., and Klaassen, C. D. (1982) Decreased
effectiveness of chelation therapy with time after acute cadmium
poisoning. Toxicol. Appl. Pharmacol. 63, 173-180.
(14) Zhao, J . Y., Foulkes, E. C., and J ones, M. M. (1990) Delayed
nephrotoxic effects of cadmium and their reversibility by chela-
tion. Toxicology 64, 235-243.
(15) Singh, P. K., J ones, M. M., J ones, S. G., Gale, G. R., Atkins, L.
M., Smith, A. B., and Bulman, R. A. (1989) Effect of chelating
agent structure on the mobilization of cadmium from intracellular
deposits. J . Toxicol. Environ. Health 28, 501-518.
(16) Cantilena, L. R., J r., and Klaassen, C. D. (1981) Comparison of
the effectiveness of several chelators after single administration
on the toxicity, excretion and distribution of cadmium. Toxicol.
Appl. Pharmacol. 58, 452-460.
Singh et al. (15) indicated not only that the structure
of the chelator must have the proper type of chelating
group but also that the presence of appropriate additional
groups in the molecule is essential for an appreciable
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