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DDE formation: Dehydrochlorination or dehypochlorination
Chemosphere No. 2, pp 67 - 70, 1972.
PerL-amonPress.
Printed in Great Britain.
DDE FORMATION: DEHYDROCHLORIr~ATIONOR DEHYPOCHLORINATION James D. McKinney and Lawrence Fishbein National Institute of Environmental Health Sciences, P. O. Box 12233 Research Triangle Park, North Carolina 27709 (~eceived in USA 9 January 1972~ received in UK for publication 21 February 1972)
I t has been known for some time that p,p-DDT [l,l,l-trichloro-2,2-bis(E-chlorophenyl)ethane] is detoxified to p,p'-DDE [l,l-dichloro-2,2-bis(E-chlorophenyl)ethylene] in humans and is stored primarily in this form in the adipose tissue, l
Recent work2 has confirmed that DDE
can be an intermediate in the rat metabolism of DDT to DDA [bis(E-chlorophenyl)acetic acid], the primary form that is ultimately excreted.
Furthermore, i t appears that DDE is a product or
at least a short-lived intermediate in the slow degradation of DDT by the action of l i g h t and air. 3 Although the dehydrochlorination (-HCI) of DDT to DDE in l i v i n g organisms is generally considered to be a one-step process catalyzed by the "DDT dehydrochlorinase" enzyme,4'5 there is very l i t t l e information to substantiate this, especially in mammalian systems.6 Recent chemical work7 in our laboratory has revealed that the tertiary-carbon atom of alkane type analogues of DDT bearing the ~-chlorophenyl groups is uniquely sensitive to oxidation.
This observation is supported by the fact that such analogues are metabolized to the
corresponding series of hydroxy derivatives by Drosophila melanogaster8 whose metabolism of DDT seems to resemble in some ways the mammalian metabolism. In addition, the photooxygenation3 of DDT affords major products which are reasonably explained by the i n i t i a l substitution of oxygen for hydrogen at the tertiary-carbon.
Since
several classes of oxygenases produce oxygenation with a chemistry resembling that of photooxygenation (singlet oxygen),9 i t seemed essential to investigate the possibility that hydroxy derivatives such as kelthane could produce unsaturated compounds such as DDE via dehypochlorination (-HOCI). DDE and kelthane occur together in some metabolism studies lO of DDT although their relative concentrations may vary a great deal, e.g., from susceptible to resistant strains, etc.
67
CC13 DDT o
-
00'c
Kelthane I t was immediately evident from studies of the chromatographic and chemical properties of these hydroxy analogues that certain substituents favored decomposition to DBP (4,4'-dichlorobenzophenone) which is knownI I to be a decomposition product of kelthane under various conditions.
The decomposition is greatest when the t e r t i a r y hydroxy-bearing carbon is adjacent to
an electron deficient carbon such as contained in the trichloromethyl group of kelthane and even more so in an aldehyde or carboxyl group. Fewer chlorine atoms at the adjacent position would slow the decomposition to DBP. I f the 2-hydroxy aldehyde or 2-hydroxy carboxylic acid occurred in metabolism, they could function as one-carbon dono~at the oxidation levels of formaldehyde and formic acid.
Consequently, one might expect increased a c t i v i t y of certain f o l i c acid
dependent enzyme systems. On the other hand, i t appears that the 2-hydroxy ethanol derivative is not a one-carbon donor at the oxidation level of methanol but surprisingly functions now as an alkylating agent since dimers and polymers can be isolated from i t s solutions. 7 Alkylation reactions during the metabolism of DDT could explain the presence of yet uncharacterized metabolite conjugates12 of unusual s t a b i l i t y . I f kelthane is a precursor of DDE, i t must gain further activation toward elimination, instead of decomposition,possibly through conjugation of the hydroxy group. Conjugation is a common biological process for detoxicating and hence f a c i l i t a t i n g elimination of hydroxylic substances. Acetylation of kelthane provided a model system13 for examining the possibility that a conjugate would have an increased propensity toward elimination or an increased oxidizing ability.
Previous workers14 have approached the study of the oxidizing a b i l i t y of organic
polyhalides by investigating the products under reductive dechlorinating conditions. Reductive dechlorination of acetylkelthane, kelthane, and DDT in hot dimethylformamide (125-140°C) in the presence of zinc powder for 30 minutes gave markedly different products. Acetylkelthane was converted in 74 percent yield to DDE along with small and approximately equal amounts of two other compounds, one of which was identified as DDMU[l-chloro-2,2-bis(E-
No. 2
69
chlorophenyl)ethylene]. conditions.
In contrast, kelthane was quantitatively converted to DBP under these
Under identical conditions, DDT gave a mixture of at least five products of which
DDE and DDD [l,l-dichloro-2,2-bis(~-chlorophenyl)ethane] were major. The reductive dechlorination of DDT to DDD in preference to DDE has been previously reported 15 in uivo in the rat.
One
infers from these results that (a) the electronic distribution of acetylkelthane and kelthane are appreciably different, favoring elimination in one case and decomposition in the other; (b) acetylkelthane is at least as good, i f not a better, precursor of DDE than is DDT under reducing conditions.
Consequently, conversion of DDT toDDE can be envisioned as a one-step
process involving dehydrochlorination,or alternatively as a two-step process involving oxidation and reduction. Therefore, qualitative and quantitative differences in the metabolism of DDT, e.g., the presence or absence of saturated metabolites,such as DDD, may be explained on the basis of different physiological responses brought about by varying degrees of enzyme induction which are dependent on the amount of toxicant used to challenge the organism. Organismswhich have been conditioned with the toxicant where enzyme induction is at i t s maximum (resistant) produce primarily the unsaturated metabolites,2 DDE, DDMU, etc.
On the other hand, when preconditioning
is not done (susceptible) saturated metabolites such as DDD are found, 6 and in some systems the hydroxy derivatives such as kelthane may occur along with or in place of their saturated precursors.
I t is possible that the preferential formation of unsaturated metabolites is a
measure of altered enzyme a c t i v i t y of the normal oxidizing and reducing type with saturated metabolites occurring only in steady-state concentrations.
The unsaturated metabolites in turn
can be converted to more polar and excretable metabolites such as DDA.2
In support of this
argument, there is at least one report 17 of the in vitro and in uivo conversion of kelthane to DDE and DDA in which DDD was shown to be an intermediate.
These workers proposed that reduced
porphyrin systems are involved in the formation of DDE from kelthane.
This is consistent with
the rapid chemical Oxidation 18 of iron ( I I ) porphyrins by alkyl halides. that conjugation may be prerequisite for DDE formation.
Our work suggests
Furthermore, i t is clear from previous
work that for maximum DDT metabolizing a c t i v i t y5'19 the presence of a cofactor with reducing properties is essential. The unique chemical properties found for some of the possible hydroxy metabolites should provide an impetus for further inquiry into t h e i r occurrence and interaction products in biological systems.
79
'~;n. 2
REFERENCES I.
W. J. Hayes, Jr., G. E. Quinby, K. C. Walker, J. W. E l l i o t t , and W. M. Upholt, Arch. Ind. Health, 18, 398 (1958).
2.
P. R. Datta, Pesticides Svmoosia qf ~he Sixth Ini~er-American Conference on Toxicoloq.y and Occupational Medicine. 41 (i970).
3.
J. R. Plimmer, U. I. Klingebiel, and B. E. Hummer, Science, 167, 67 (1970).
4.
H. Lipke and C. W. Kearns, J. Biol. Chem., 234(8), 2123 (1959).
5.
H. Lipke and C. W. Kearns, J. Biol. Chem., 234(8), 2129 (1959).
6.
C. F. Rothe, A. M. Mattson, R. M. Nueslein, and W. J. Hayes, Jr., Arch. Ind. Health, L6, 82 (1957).
7.
J. D. McKinney, R. Hawk, E. Boozer, and J. E. Suggs, Canadian J. Chem., 49, 3877 (1971).
8.
M. Tsukamoto, Bochu-Kaqaku, 26, 74 (1961).
9.
O. Hayaishi, p. 151.
Oxyqen in the Animal Orqanism, The Macmillan Co., New York, N. Y., 1964,
lO.
M. Agosin, D. Michaeli, R. Miskus, S. Nagasawa, and W. M. Hoskins, J. Econ. Entomology, 54, 340 (1961).
II.
F. A. Gunther, J. H. Barkley, R. C. Blinn~ and D. E. Ott, Pesticide Research Bull., Stanford Research Inst., 2(2), 3 (1962).
12.
J. A. Jensen, C. Cueto, W. E. Dale, C. F. Rothe, G. W. Pearce, and A. M. Mattson, J. A~H c. Food Chem., 5, 919 (1957).
13.
This compound has been previously prepared [E. D. Bergmann and A. Kaluszyner, ~ . Chem., 23, 1306 (1958)] as an intemediate in the synthesis of kelthane from DDT.
14.
T. A. Cooper and T. Takeshita, J. Org. Chem., 36(23), 3517 (1971).
15.
A. K. Klein, E. P. Laud, P. R. Datta, J. O. Watts, and J. T. Chen, J. Assoc. Off. Anal. Chemists, 4__7_7(6),1129 (1964).
16.
J. E. Peterson and W. H. Robison, Toxicol. and Appl. Phamacol., 6_, 321 (1964).
17.
H. Hughes, I. Bazant, and J. R. Brown, Canadian J. of Public Health, 60(I), 41 (1963).
18.
C. A. Castro, J. Amer. Chem. Soc., 8_~6,2310 (1964).
19.
A. S. Perry, S. Miller, and A. J. Buckner, J. A~ric. Food Chem., Ll(6), 457 (1963).
PerL-amonPress.
Printed in Great Britain.
DDE FORMATION: DEHYDROCHLORIr~ATIONOR DEHYPOCHLORINATION James D. McKinney and Lawrence Fishbein National Institute of Environmental Health Sciences, P. O. Box 12233 Research Triangle Park, North Carolina 27709 (~eceived in USA 9 January 1972~ received in UK for publication 21 February 1972)
I t has been known for some time that p,p-DDT [l,l,l-trichloro-2,2-bis(E-chlorophenyl)ethane] is detoxified to p,p'-DDE [l,l-dichloro-2,2-bis(E-chlorophenyl)ethylene] in humans and is stored primarily in this form in the adipose tissue, l
Recent work2 has confirmed that DDE
can be an intermediate in the rat metabolism of DDT to DDA [bis(E-chlorophenyl)acetic acid], the primary form that is ultimately excreted.
Furthermore, i t appears that DDE is a product or
at least a short-lived intermediate in the slow degradation of DDT by the action of l i g h t and air. 3 Although the dehydrochlorination (-HCI) of DDT to DDE in l i v i n g organisms is generally considered to be a one-step process catalyzed by the "DDT dehydrochlorinase" enzyme,4'5 there is very l i t t l e information to substantiate this, especially in mammalian systems.6 Recent chemical work7 in our laboratory has revealed that the tertiary-carbon atom of alkane type analogues of DDT bearing the ~-chlorophenyl groups is uniquely sensitive to oxidation.
This observation is supported by the fact that such analogues are metabolized to the
corresponding series of hydroxy derivatives by Drosophila melanogaster8 whose metabolism of DDT seems to resemble in some ways the mammalian metabolism. In addition, the photooxygenation3 of DDT affords major products which are reasonably explained by the i n i t i a l substitution of oxygen for hydrogen at the tertiary-carbon.
Since
several classes of oxygenases produce oxygenation with a chemistry resembling that of photooxygenation (singlet oxygen),9 i t seemed essential to investigate the possibility that hydroxy derivatives such as kelthane could produce unsaturated compounds such as DDE via dehypochlorination (-HOCI). DDE and kelthane occur together in some metabolism studies lO of DDT although their relative concentrations may vary a great deal, e.g., from susceptible to resistant strains, etc.
67
CC13 DDT o
-
00'c
Kelthane I t was immediately evident from studies of the chromatographic and chemical properties of these hydroxy analogues that certain substituents favored decomposition to DBP (4,4'-dichlorobenzophenone) which is knownI I to be a decomposition product of kelthane under various conditions.
The decomposition is greatest when the t e r t i a r y hydroxy-bearing carbon is adjacent to
an electron deficient carbon such as contained in the trichloromethyl group of kelthane and even more so in an aldehyde or carboxyl group. Fewer chlorine atoms at the adjacent position would slow the decomposition to DBP. I f the 2-hydroxy aldehyde or 2-hydroxy carboxylic acid occurred in metabolism, they could function as one-carbon dono~at the oxidation levels of formaldehyde and formic acid.
Consequently, one might expect increased a c t i v i t y of certain f o l i c acid
dependent enzyme systems. On the other hand, i t appears that the 2-hydroxy ethanol derivative is not a one-carbon donor at the oxidation level of methanol but surprisingly functions now as an alkylating agent since dimers and polymers can be isolated from i t s solutions. 7 Alkylation reactions during the metabolism of DDT could explain the presence of yet uncharacterized metabolite conjugates12 of unusual s t a b i l i t y . I f kelthane is a precursor of DDE, i t must gain further activation toward elimination, instead of decomposition,possibly through conjugation of the hydroxy group. Conjugation is a common biological process for detoxicating and hence f a c i l i t a t i n g elimination of hydroxylic substances. Acetylation of kelthane provided a model system13 for examining the possibility that a conjugate would have an increased propensity toward elimination or an increased oxidizing ability.
Previous workers14 have approached the study of the oxidizing a b i l i t y of organic
polyhalides by investigating the products under reductive dechlorinating conditions. Reductive dechlorination of acetylkelthane, kelthane, and DDT in hot dimethylformamide (125-140°C) in the presence of zinc powder for 30 minutes gave markedly different products. Acetylkelthane was converted in 74 percent yield to DDE along with small and approximately equal amounts of two other compounds, one of which was identified as DDMU[l-chloro-2,2-bis(E-
No. 2
69
chlorophenyl)ethylene]. conditions.
In contrast, kelthane was quantitatively converted to DBP under these
Under identical conditions, DDT gave a mixture of at least five products of which
DDE and DDD [l,l-dichloro-2,2-bis(~-chlorophenyl)ethane] were major. The reductive dechlorination of DDT to DDD in preference to DDE has been previously reported 15 in uivo in the rat.
One
infers from these results that (a) the electronic distribution of acetylkelthane and kelthane are appreciably different, favoring elimination in one case and decomposition in the other; (b) acetylkelthane is at least as good, i f not a better, precursor of DDE than is DDT under reducing conditions.
Consequently, conversion of DDT toDDE can be envisioned as a one-step
process involving dehydrochlorination,or alternatively as a two-step process involving oxidation and reduction. Therefore, qualitative and quantitative differences in the metabolism of DDT, e.g., the presence or absence of saturated metabolites,such as DDD, may be explained on the basis of different physiological responses brought about by varying degrees of enzyme induction which are dependent on the amount of toxicant used to challenge the organism. Organismswhich have been conditioned with the toxicant where enzyme induction is at i t s maximum (resistant) produce primarily the unsaturated metabolites,2 DDE, DDMU, etc.
On the other hand, when preconditioning
is not done (susceptible) saturated metabolites such as DDD are found, 6 and in some systems the hydroxy derivatives such as kelthane may occur along with or in place of their saturated precursors.
I t is possible that the preferential formation of unsaturated metabolites is a
measure of altered enzyme a c t i v i t y of the normal oxidizing and reducing type with saturated metabolites occurring only in steady-state concentrations.
The unsaturated metabolites in turn
can be converted to more polar and excretable metabolites such as DDA.2
In support of this
argument, there is at least one report 17 of the in vitro and in uivo conversion of kelthane to DDE and DDA in which DDD was shown to be an intermediate.
These workers proposed that reduced
porphyrin systems are involved in the formation of DDE from kelthane.
This is consistent with
the rapid chemical Oxidation 18 of iron ( I I ) porphyrins by alkyl halides. that conjugation may be prerequisite for DDE formation.
Our work suggests
Furthermore, i t is clear from previous
work that for maximum DDT metabolizing a c t i v i t y5'19 the presence of a cofactor with reducing properties is essential. The unique chemical properties found for some of the possible hydroxy metabolites should provide an impetus for further inquiry into t h e i r occurrence and interaction products in biological systems.
79
'~;n. 2
REFERENCES I.
W. J. Hayes, Jr., G. E. Quinby, K. C. Walker, J. W. E l l i o t t , and W. M. Upholt, Arch. Ind. Health, 18, 398 (1958).
2.
P. R. Datta, Pesticides Svmoosia qf ~he Sixth Ini~er-American Conference on Toxicoloq.y and Occupational Medicine. 41 (i970).
3.
J. R. Plimmer, U. I. Klingebiel, and B. E. Hummer, Science, 167, 67 (1970).
4.
H. Lipke and C. W. Kearns, J. Biol. Chem., 234(8), 2123 (1959).
5.
H. Lipke and C. W. Kearns, J. Biol. Chem., 234(8), 2129 (1959).
6.
C. F. Rothe, A. M. Mattson, R. M. Nueslein, and W. J. Hayes, Jr., Arch. Ind. Health, L6, 82 (1957).
7.
J. D. McKinney, R. Hawk, E. Boozer, and J. E. Suggs, Canadian J. Chem., 49, 3877 (1971).
8.
M. Tsukamoto, Bochu-Kaqaku, 26, 74 (1961).
9.
O. Hayaishi, p. 151.
Oxyqen in the Animal Orqanism, The Macmillan Co., New York, N. Y., 1964,
lO.
M. Agosin, D. Michaeli, R. Miskus, S. Nagasawa, and W. M. Hoskins, J. Econ. Entomology, 54, 340 (1961).
II.
F. A. Gunther, J. H. Barkley, R. C. Blinn~ and D. E. Ott, Pesticide Research Bull., Stanford Research Inst., 2(2), 3 (1962).
12.
J. A. Jensen, C. Cueto, W. E. Dale, C. F. Rothe, G. W. Pearce, and A. M. Mattson, J. A~H c. Food Chem., 5, 919 (1957).
13.
This compound has been previously prepared [E. D. Bergmann and A. Kaluszyner, ~ . Chem., 23, 1306 (1958)] as an intemediate in the synthesis of kelthane from DDT.
14.
T. A. Cooper and T. Takeshita, J. Org. Chem., 36(23), 3517 (1971).
15.
A. K. Klein, E. P. Laud, P. R. Datta, J. O. Watts, and J. T. Chen, J. Assoc. Off. Anal. Chemists, 4__7_7(6),1129 (1964).
16.
J. E. Peterson and W. H. Robison, Toxicol. and Appl. Phamacol., 6_, 321 (1964).
17.
H. Hughes, I. Bazant, and J. R. Brown, Canadian J. of Public Health, 60(I), 41 (1963).
18.
C. A. Castro, J. Amer. Chem. Soc., 8_~6,2310 (1964).
19.
A. S. Perry, S. Miller, and A. J. Buckner, J. A~ric. Food Chem., Ll(6), 457 (1963).