DDT Uptake and Metabolism by a Marine Diatom by JULIAN E. KEIL
Preventive Medicine Section and LAMAR E. PRIESTER
Chemistry Department Medical College of South Carolina Charleston, South Carolina
Introduction It had been o b s e r v e d
(3) that shrimp in
aquaria were sensitive to small amounts of certain insecticides,
but that similar levels of these
chemicals w h e n applied in a natural environment, a p p a r e n t l y had no effect on the shrimp.
This
i n d i c a t e d the p o s s i b i l i t y of the presence of natural d e t o x i f i e r s w h i c h m i g h t be r e s p o n s i b l e this m o d i f i c a t i o n of toxicity. detoxifiers
The most probable
are the m i c r o o r g a n i s m s
and e s p e c i a l l y the diatoms,
for
in the w a t e r
since they are known
to store food as oil and leucosin rather than starch and m i g h t be e x p e c t e d to serve as storage or "pickup" o r g a n i s m s
for oil soluble pesticides.
Methods Twelve one liter e r h l e n m e y e r flasks each c o n t a i n i n g 200 ml of culture m e d i a were i n o c u l a t e d w i t h the d i a t o m C ~ l i n d r o t h e c a closterium, and Lewin.
Reimann
Four flasks served as u n t r e a t e d
Bulletinof EnvironmentalContamination&~xicology, Yd. % No. 3, 1969, publi~ed by SpringerWerlag New York Inc.
controls,
four as DDT t r e a t m e n t s
.i p p m DDT),
(containing
and four as acetone controls,
since
acetone was the vehicle for the DDT. At the c o n c l u s i o n of a three week i n c u b a t i o n period, (0.46
the contents were m i l l i p o r e filtered pore size),
stored t e m p o r a r i l y at -70~
l y o p h i l i z e d and a n a l y z e d for DDT and its m e t a b o lites by gas c h r o m a t o g r a p h y using a M i c r o - T e k Model MT 220 e q u i p p e d w i t h a Nickel 63 e l e c t r o n capture detector.
Results and D i s c u s s i o n It had b e e n p o s t u l a t e d in the i n t r o d u c t i o n of this paper that there m i g h t be m a r i n e o r g a n i s m s w h i c h served as storage sites for oil soluble p e s t i c i d e s or w h i c h m i g h t be capable of d e t o x i f i cation of these chemicals.
Results shown in Table 1
tend to support this hypothesis.
Gas c h r o m a t o g -
raphy showed that C. c l o s t e r i u m adsorbed or a b s o r b e d DDT from culture m e d i a and on the average c o n c e n t r a t e d it about 265 times.
Data in Table 1
also showed that there was a pickup of DDT by diatoms not d e l i b e r a t e l y exposed to this insecticide.
It is t h o u g h t that these amounts came from
DDT that was n a t u r a l l y o c c u r r i n g in the w a t e r or from m i n u t e r e s i d u a l s
found on the glassware. 170
TABLE
i
DDT U p t a k e and C o n v e r s i o n to DDE by C y l i n d r o t h e c a c l o s t e r i u m after 21 Days C u l t u r e P e r i o d
DDT and DDE C o n t e n t p-p'DDT p-p'DDE (ppm) (ppm)
Treatment and R e p l i c a t e
Control 1 2 3 4
Acetone 1 2 3 4 x2
DDE
as P e r c e n t p-p'DDT
7.5064 9.6078 4.5183 7.6404 8.0682
1.0795 .6209 .3502 .3370 .5969
14.4 6.5 7.8 4.4 7.4
2.4228 2.1671 1.7319 3.7347 2.5141
.2296 trace trace .3554 .1462
9.5 --9.5 5.8
26.4066 41.6924 15.6668 26.5044 27.5675
1.9502 3.1407 .5286 4.1036 2.4307
7.4 7.5 3.4 15.5 8.8
Control
DDT E x p o s e d 1 1 2 3 4 x3
i.i p p m p - p ' D D T a d d e d to media; no DDT a d d e d to m e d i a of c o n t r o l s ; DDT p r e s e n t in c o n t r o l f r o m n a t u r a l o c c u r r e n c e in w a t e r or c o n t a m i n a t i o n . Confidence
Levels
~i - ~2
NSD
NSD
Xl - x3
.01>p>.001**
x2 - x3
pp>.01* .01>p>.001***
* = S i g n i f i c a n t (95% c o n f i d e n c e ) ** = H i g h l y S i g n i f i c a n t (99% c o n f i d e n c e ) *** = H i g h l y S i g n i f i c a n t (99.9% c o n f i d e n c e )
171
There was no s i g n i f i c a n t d i f f e r e n c e
in amounts of
DDT b e t w e e n the control and acetone control flasks of diatoms.
The amounts of DDT in each of these
treatments were fairly consistent.
Highly signifi-
cant d i f f e r e n c e s b e t w e e n either of the control groups and the DDT exposed diatoms, DDT content, was shown.
as regards p-p'
Even if the 8 ppm
average DDT content of the control diatoms was s u b t r a c t e d from the 27 ppm DDT content of the exposed diatoms,
there w o u l d still be a s i g n i f i c a n t
190-fold c o n c e n t r a t i o n of DDT by C. closterium. Data in Table 1 also i n d i c a t e d a fairly u n i f o r m c o n v e r s i o n of DDT to DDE dichloroethylene).
This is a d e t o x i f i c a t i o n
process by d e h y d r o h a l o g e n a t i o n . relatively
(dichlorodiphenyl-
While there were
small amounts of DDE detected,
the
amounts were u n i f o r m when e x p r e s s e d as a p e r c e n t age of p-p' DDT.
Since DDE is c o n s i d e r a b l y more
w a t e r soluble than DDT it w o u l d be less likely to a c c u m u l a t e in diatoms or other similar aquatic organisms. It is i n t e r e s t i n g that C. c l o s t e r i u m conv e r t e d DDT only to the less toxic m e t a b o l i t e DDE, and it w o u l d be r e a s s u r i n g to e c o l o g i s t s if this were typical of all diatoms. d e m o n s t r a t e d by Barker,
et al.
Indeed,
it has been
(i), S t e n e r s e n
(4),
Johnson,
et al.
(2) that certain b a c t e r i a may con-
vert DDT not only to DDE but to the equally or more toxic analogue DDD
(dichlorodiphenyldichloro-
ethane). It w o u l d be well if this work could be rep e a t e d on a wide spectrum of m a r i n e as well as fresh water diatoms and using labeled DDT,
since
the apparent u b i q u i t y of this p e s t i c i d e allows it to appear even in "controls".
Conclusion C y l i n d r o t h e c a closterium,
Reimann and Lewin, was
capable of absorbing and c o n c e n t r a t i n g DDT above the level in seawater.
DDT was m e t a b o l i z e d by
this o r g a n i s m only to DDE.
References i.
BARKER, Nature,
P.S., F.O. MORRISON, 205, 621 (1965).
and R.S WHITAKER,
.
JOHNSON, B.T., R.N. GOODMAN, Science, 157, 560 (1967).
.
KEIL, J.E., P r e s i d e n t i a l Address, Soc., (1967).
4.
STENERSEN,
J.H.V., Nature,
17~
and H.S. GOLDBERG,
S.C. Ent.
207, 660
(1965).
Preventive Medicine Section and LAMAR E. PRIESTER
Chemistry Department Medical College of South Carolina Charleston, South Carolina
Introduction It had been o b s e r v e d
(3) that shrimp in
aquaria were sensitive to small amounts of certain insecticides,
but that similar levels of these
chemicals w h e n applied in a natural environment, a p p a r e n t l y had no effect on the shrimp.
This
i n d i c a t e d the p o s s i b i l i t y of the presence of natural d e t o x i f i e r s w h i c h m i g h t be r e s p o n s i b l e this m o d i f i c a t i o n of toxicity. detoxifiers
The most probable
are the m i c r o o r g a n i s m s
and e s p e c i a l l y the diatoms,
for
in the w a t e r
since they are known
to store food as oil and leucosin rather than starch and m i g h t be e x p e c t e d to serve as storage or "pickup" o r g a n i s m s
for oil soluble pesticides.
Methods Twelve one liter e r h l e n m e y e r flasks each c o n t a i n i n g 200 ml of culture m e d i a were i n o c u l a t e d w i t h the d i a t o m C ~ l i n d r o t h e c a closterium, and Lewin.
Reimann
Four flasks served as u n t r e a t e d
Bulletinof EnvironmentalContamination&~xicology, Yd. % No. 3, 1969, publi~ed by SpringerWerlag New York Inc.
controls,
four as DDT t r e a t m e n t s
.i p p m DDT),
(containing
and four as acetone controls,
since
acetone was the vehicle for the DDT. At the c o n c l u s i o n of a three week i n c u b a t i o n period, (0.46
the contents were m i l l i p o r e filtered pore size),
stored t e m p o r a r i l y at -70~
l y o p h i l i z e d and a n a l y z e d for DDT and its m e t a b o lites by gas c h r o m a t o g r a p h y using a M i c r o - T e k Model MT 220 e q u i p p e d w i t h a Nickel 63 e l e c t r o n capture detector.
Results and D i s c u s s i o n It had b e e n p o s t u l a t e d in the i n t r o d u c t i o n of this paper that there m i g h t be m a r i n e o r g a n i s m s w h i c h served as storage sites for oil soluble p e s t i c i d e s or w h i c h m i g h t be capable of d e t o x i f i cation of these chemicals.
Results shown in Table 1
tend to support this hypothesis.
Gas c h r o m a t o g -
raphy showed that C. c l o s t e r i u m adsorbed or a b s o r b e d DDT from culture m e d i a and on the average c o n c e n t r a t e d it about 265 times.
Data in Table 1
also showed that there was a pickup of DDT by diatoms not d e l i b e r a t e l y exposed to this insecticide.
It is t h o u g h t that these amounts came from
DDT that was n a t u r a l l y o c c u r r i n g in the w a t e r or from m i n u t e r e s i d u a l s
found on the glassware. 170
TABLE
i
DDT U p t a k e and C o n v e r s i o n to DDE by C y l i n d r o t h e c a c l o s t e r i u m after 21 Days C u l t u r e P e r i o d
DDT and DDE C o n t e n t p-p'DDT p-p'DDE (ppm) (ppm)
Treatment and R e p l i c a t e
Control 1 2 3 4
Acetone 1 2 3 4 x2
DDE
as P e r c e n t p-p'DDT
7.5064 9.6078 4.5183 7.6404 8.0682
1.0795 .6209 .3502 .3370 .5969
14.4 6.5 7.8 4.4 7.4
2.4228 2.1671 1.7319 3.7347 2.5141
.2296 trace trace .3554 .1462
9.5 --9.5 5.8
26.4066 41.6924 15.6668 26.5044 27.5675
1.9502 3.1407 .5286 4.1036 2.4307
7.4 7.5 3.4 15.5 8.8
Control
DDT E x p o s e d 1 1 2 3 4 x3
i.i p p m p - p ' D D T a d d e d to media; no DDT a d d e d to m e d i a of c o n t r o l s ; DDT p r e s e n t in c o n t r o l f r o m n a t u r a l o c c u r r e n c e in w a t e r or c o n t a m i n a t i o n . Confidence
Levels
~i - ~2
NSD
NSD
Xl - x3
.01>p>.001**
x2 - x3
pp>.01* .01>p>.001***
* = S i g n i f i c a n t (95% c o n f i d e n c e ) ** = H i g h l y S i g n i f i c a n t (99% c o n f i d e n c e ) *** = H i g h l y S i g n i f i c a n t (99.9% c o n f i d e n c e )
171
There was no s i g n i f i c a n t d i f f e r e n c e
in amounts of
DDT b e t w e e n the control and acetone control flasks of diatoms.
The amounts of DDT in each of these
treatments were fairly consistent.
Highly signifi-
cant d i f f e r e n c e s b e t w e e n either of the control groups and the DDT exposed diatoms, DDT content, was shown.
as regards p-p'
Even if the 8 ppm
average DDT content of the control diatoms was s u b t r a c t e d from the 27 ppm DDT content of the exposed diatoms,
there w o u l d still be a s i g n i f i c a n t
190-fold c o n c e n t r a t i o n of DDT by C. closterium. Data in Table 1 also i n d i c a t e d a fairly u n i f o r m c o n v e r s i o n of DDT to DDE dichloroethylene).
This is a d e t o x i f i c a t i o n
process by d e h y d r o h a l o g e n a t i o n . relatively
(dichlorodiphenyl-
While there were
small amounts of DDE detected,
the
amounts were u n i f o r m when e x p r e s s e d as a p e r c e n t age of p-p' DDT.
Since DDE is c o n s i d e r a b l y more
w a t e r soluble than DDT it w o u l d be less likely to a c c u m u l a t e in diatoms or other similar aquatic organisms. It is i n t e r e s t i n g that C. c l o s t e r i u m conv e r t e d DDT only to the less toxic m e t a b o l i t e DDE, and it w o u l d be r e a s s u r i n g to e c o l o g i s t s if this were typical of all diatoms. d e m o n s t r a t e d by Barker,
et al.
Indeed,
it has been
(i), S t e n e r s e n
(4),
Johnson,
et al.
(2) that certain b a c t e r i a may con-
vert DDT not only to DDE but to the equally or more toxic analogue DDD
(dichlorodiphenyldichloro-
ethane). It w o u l d be well if this work could be rep e a t e d on a wide spectrum of m a r i n e as well as fresh water diatoms and using labeled DDT,
since
the apparent u b i q u i t y of this p e s t i c i d e allows it to appear even in "controls".
Conclusion C y l i n d r o t h e c a closterium,
Reimann and Lewin, was
capable of absorbing and c o n c e n t r a t i n g DDT above the level in seawater.
DDT was m e t a b o l i z e d by
this o r g a n i s m only to DDE.
References i.
BARKER, Nature,
P.S., F.O. MORRISON, 205, 621 (1965).
and R.S WHITAKER,
.
JOHNSON, B.T., R.N. GOODMAN, Science, 157, 560 (1967).
.
KEIL, J.E., P r e s i d e n t i a l Address, Soc., (1967).
4.
STENERSEN,
J.H.V., Nature,
17~
and H.S. GOLDBERG,
S.C. Ent.
207, 660
(1965).
