This paper not to oe eited \vithout prior rderence to the authors.
C.M. I9811E : 56
Marine Environmental
Quality Committee
INTERNATIONAL COUNCIL FOR
TUE EXPLORAnON OF TUE SEA
BIODEGRADABILITY IN TUE MARINE ENVIRONMENT.
I. TUE CUEMOSTAT AS A MODEL SYSTEM FOR TUE AQUATIC ENVIRONMENT.
by
R. v.d. Berg.
Mrs. J .M.P. Buntsma-Hamers and
B.L. Verboom
Netherlands Institute for Fishery Investigations
Haringkadc I. Postoox 68.
1970 AB IJmuiden. The Netherlands
This paper notto be cited without prior reference to the authors.
International Council for the
Exploration of the Sea.
C.M. 1981/E : 56
Marine Environmental
Quality Committee.
BIODEGRADABILITY IN TEE MARINE ENVIRONMENT.
I. THE CHEMOSTAT AS A MODEL SYSTEM FOR THE
AQUATIC ENVIRONMENT.
•
by
Ir. R. v.d. Berg, Mrs. J.M.P.
Buntsma-Hamers and B.L. Verboom
Netherlands Institute for
Fishery Investigations
Haringkade 1, Postbox 68
IJmuiden, The Netherlands .
Summary
A test method is described for measuring biodegraßability in the aquatic
marine environment. On the basis of the characteristic properties of the
sea the chemostat has been chosen as the model system. In the chemostat test
the shortest possible residence time for a test compound is established.
On the basis of this residence time a turn-over time in the natural environment is estimated.
•
.
Introduction
As man continues to develop and utilize new organic chemieals, the potential
for environmental pollution increases accordingly. Once a chemical has entered the environment,its eventual transport to the aquatic environment becomes an obvious possibility.
At the moment morp than 50 000 chemieals are produeed eommercially and
each year about 100 new chemieals are added to this list •
Before a chemieal is manufaetured commercially, a hazard assessment has
to be made to establish the environmental risk of this chemieal.
Besides, in ease of an inadmissible risk, a risk benefit analysis could be
performed. Two key quest ions in the environmental hazard assessment are:
• what is the fate of the chemical in the environment~
• which effects may be expected.
A thorough knowledge of the fate of a chemieal is required and particularly
with respect to its ultimate removal from the environment by biological or
chemieal (12) degradation:
Miero-organisms are eommonly believed to play t~e most important role in
determining the fate of ehemieal substances in aquatic systems. In the
marine environment most of the organic matter produeed photosynthetically
in the surface waters is decomposed in the upper few hundred meters (9)
espeeially by heterotrophie baeteria (1). In eoastal and open waters 90 %
of.the measurable heterotrophie uptake occurred in the fraetion less than
1 ~m in size or was assoeiated with bacteria on particles (1,9) •
Literature studies reveal few research coneerning the rates and products
cf microbial degradation in the aquatie environment of most of the commonly used chemieals. The greater part of the research has been pointed at
,
'
- 2 -
the metabolie pathways by which eertain chcmically. pure substances were
transformed by pure or enriehed eultures in a weIl defined medium (3,
10, 13).
.
Althougha large number of bio-degradability testing methods are available (4), at the moment not yet one suitable method for the measuring of
the aquatic environmental degradation exists; perhaps because of the
great variety in aquatie compartments (freshversus marine, estuaries,
eoastal zones, oeeans,:deep sea).
Particularly considering the marine environment no specific methods have
been developped. One has to rely on existing methods for the fresh water
environment.
This study has been aimed at the development of a test method for the
prediction of the bio-degradability potential of organic chemieals in the
marine environment.
•
•
'.
Choice of the test system
To prediet the fate of xenobiotic substances in the marine environment
a model system has been developped resembling the sea. Important eharaeteristie properties of the marine environment are:
• a continuous input of organic substanees by means of the primary produetion of phytoplankton,
• a eontinuous decomposition of organie material by mixed.populations of
heterotrophie micro-organisms,
· a dynamic equilibrium during this continuous carbon cycle,
• a wide range of turn-over times for al~ kinds of eompounds ~n various
areas (6).
Combining these eharaeteristics a continuous flow culture aeeording to the
ehemostat prineiple (16) has been chosen as model system for the marine
environment.
The primary produetion is simulated by the input of a large number of
fairIy bio-degradable organie ehemieals, in relatively high eoneentrations
regarding to the natural mar~ne environment, but in about the same relative
amount. In this way a relatively highcr biomass is obtained with relatively higher bio-degradation rates than in natural seawater. The 6rganie
substrates together eonstitute as carbon and energy source t~e growth
limiting faetor for the heterotrophie micro-organisms.
.
- The natural environment eontains a lot of heterogeneous assemblies of
microbes and the promotion of mixed eultures may give better results,
espeeially for xenobiotic ehemicals (14). The eontinuous input of a large
number of organie substances promotes the maintenance qf a weIl mixed
eommunity of micro-organisms in the ehemostat and may stimulate eometaboIic transformations (8). Co~etabolism is the metabolism by a miero-organism
of a chemieal which this speeies 'cannot use as a nutrient or energy source
(7, 11). Consequenees of eometabolism are a slow mierobial transformation
rate and the accumulation of strueturally similar products. IIowever, the
combination of a cometabolic step and mixed cultures may still lead to
complete degradation, in which reaction sequence
the cometabolic step(s)
is(are) likely to be rate limiting.
In a ehemostat just as in the sea adynamie equiiibrilim arises which cannot
be disturbed easily beeause this system is self-stabilizing.
The turn-over time of a ehemieal is the time needed for the complete degradation of this ehemieal. The turn-over time in the sea can be compared .
with the residence time in the ehemostat. The residenee time is the mean
time during which the substrates and the miero-organisms stay in the eulture vessel and during which degradation by the community of microbes may
- 3 oeeur. In the ehemostat a wid~ range of residenee times is possible.
A short applieable residenee time for a ehemical dces also m~an a short
turn-over time.
The study for the mierobiologieal fate ofaxenobiotie eompound is performed by adding the xenobiotic to the nutrient solution in minor, nontoxie, but analytieally suitable eonc~ntrations. The degradation is established as function of the residenee time. Figure 1 gives adegradation
eurve. As eharaeteristie data from this eurve the range of residenee times
(Tl to T2) for the transition from 0 to 100 %degradation is defined.
•
Interpretation of test results.
The degradation of organie eompounds in the natural aquatic compartments
ean be estimated by assuming that the deeomposition of the,organie substanees equals the produetion of these compounds by the primary produetion,
beeause thc standing erop oforganic earbon in the oeeans is in a steady
state. If the biomass present is known the degradation rate, the amount
of dissolved organic, carbon degraded per unit of time and biomass, ean be
ealculated. The turn-over of the organie substances of the nutrient solution in the chemostat is calculated for the highest possible degradation
rate (point A in figure 1), situated in the eharaeteristic residenee time
range Tl - T2. From the performed experiments (2) this range may be estimated at 0.75 to 1.0 h. Table I shows the degradation rates for organic
carbon in the various natural aquatic compartments and the chemostat.
From the data in table I a theoretical ratio between the degradation
rates of organic compounds in the chemostat and in the other aquatic
compartments ean be calculated (table 11).
For glucose some literature data are available of turn-over times in various
aquatic compartments (6, 15) and for the chemostat the minimum necessary
residence time for eomplete degradation may be estimated at 0.75 h(2).
A lärge range is seen in the turn-over data for glucose, but in comparison
with these experimental data the ratio between chemostat and "naturally"
measured data is about the same as theoretieally calculated (table 11).
Conclusions
It seems possible to predict residence times or turn-over times in aquatic
marine compartments on the basis of the data obtained in chemostat experiments .
•
- 4-
Referenees
1. Azam, F. and R.E. Hodson, 1977
Size distribution and aetivity of
marine mieroheterotrophs.
Limnol. Oeeanogr. 22: 492-501.
2.
Biodegradability in the marine
environment. 11. Development of the
ehemostat testmethodology.
(in preparation).
Berg~
R. van den
3. Furukawa, K., N. Tomizuka and
A. Kamibayashi, 1979
Effeet of ehlorine substition on the
baeterial metabolism of various
polyehlorinated biphenyls.
Appl. Environ. Mierobiol. 38: 301-310.
4. Gilbert, P.A. and G.K. Watson,
1977
Biodegradability testing and its
relevanee to environmental
aeeeptability.
Tenside Detergents 14: 171-177.
5. Hanson, R.B. and W.J. Wiebe, 1977
Heterotrophie aetivity assoeiated with
partieulate size fraetions in a
Spartina alterniflora salt marsh
estuary, Sapelo Island, Georgia, USA
and the eontinental shelf water,
Mar. Biol; 42: 321-333.
6. Hoppe, H.G., 1978
Relations between aetive baeteria
and heterotrophie potential in the
sea.
Neth. J. Sea Res. 12: 78-98.
7.
Horvath~
Mierobial eo-metabolism and the
degradation of organie compounds ln
nature.
Baeteriol. Rev. 36: 146-155.
R.S., 1972
8. Jaeobson, S.N., N.L. OlMara and
M. Alexander~ 1980
The Ultimate Sink. Proeeedings of the
workshop Mierobial degradation of
pollutants in Marine environments.
(A.W. Bourguinkeds and P.H. Pritehard,eds.)
pp 3-9 Report no. EPA-60o/9-79-012.
U.S. Environmental Proteetion AF,eney~
Gulf Breeze, Florida.
9. Jannaseh, H.W., 1979
10. Knaekmuss, H.J. andH.
1978
Evidenee for eometabolism in sewage.
Appl. Environ. Mierobiol. 40: 917-921.
,Hellwig~
11. Leadbetter, E.R. and J.W.
1959
Foster~
Utilization and eooxidation of
ehlorinated phenols by Pseudomonas
sp. B 13.
Areh. Mierobiol~ 117: 1~r.
Oxidation produets formed from
gaseous alkanes by the baeterium
Pseudomonas methanica.
Areh. Bioehem. Biophys. 82:491-492.
12. Mabey, W. and T. Mill, 1978
Critieal Review of hydrolysis ofOrganie
Compounds in Water Under Environmental
Conditions.
J. Phys. ehem. Ref. Data. 7: 383-415.
- 5-
13. Reineke,W. andH.J. Knackmuss,
1980
•
Hybrid pathway for chlorobenzoate
metabolism in Pseudomonas sp. B 13
derivatives.
J. Bacteriol. 42: 467-473.
14. Senior,E.,A.T. Bull and
J.H. SLater, 1976
Enzyme evolution in a microbial
community growing on the herbicide
Dalapon.
Nature 263: 476-479.
15. Sepers, A.B.J., 1977
The utilization of dissolved organic
compounds in aquatic environments.
Hydrobiologia 52: 39-54.
16. Tempest, D.W., 1970
The continuous cultivation of microorganisms. I; Theory of the chemostat.
In Methods in Microbiology, volume 2.
(J.R. Norris and D.W. Ribbons, eds.)
pp. 259-276. Academic Press, London •
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1. 0
•
residencetime [h] 1
•
Tl
FIGURE 1. Concentration and degradationrate of
a substrate in the chernostat as
a function 01 the residencetime.
T2
~---------------
T ABlE I. Degradationrate of organic carbon In
various aquatic compartments.
primary
depth
production
gC/m 2 a
50
o~eans
estuaria
coastal zones
100 - 500
a
b
biomass degrad.
degrad.
rate
m
gC/m 3 a
mg/m 3
rate
gC/g.a
200
0.25
5 - 50
5-50
20
5 - 25
6
10
10 -100
5
10
50-2500
4
10
.emostat
notes
a. depth in which the degradationprocesses take place
b. biomass' as dry weight
J ABlE U. Ratio of the degradationrates (A)
and experimental data for glucose(BL'
®
•
ratio degr. rate
chemostat compartment
ocean
estuaria
coastal zones
chemostat
200 - 2000
4 - 200
1
notes
a. turnovertime -literature data (6,15)
b. residencetime (2)
.
®
glucose degrad.rate
experimental data
[hJ
median
range
1'000 a
200-6000
20
a
O.7S
1 - 500
b
a
a