HAZARD ASSESSMENT FOR CHLORINATED PARAFFINS:
EFFECTS ON FISH AND WILDLIFE
Health and Environmental Review Division
Environmental Effects Branch
Toxicology Branch
January, 1985
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PRELIMINARY HAZARD ASSESSMENT ON. CHLORINATED PARAFFINS
Executive Summary
The following hazard assessment on chlorinated paraffins is based on the
testing data submitted by the Chlorinated Paraffins Consortium in response to
the negotiated testing as published in the Federal Register on January 8, 1982.
o Results from the Phase I 60-day toxicity tests on rainbow trout and
mussels indicate that the 58% chlorinated short chain length (Cio_i2)
y*
n-paraffins is the most toxic of the four formulations tested.
o The Phase I testing matrix failed to adequately identify toxicological
relationships between the wide array of chlorinated paraffin mixtures
which vary both in the degree of chlorination and in the length of the
carbon chain.
o Results of the Phase II life-cycle studies on a variety of test species
^indicate statistically significant (P £ 0.05) toxic effects at measured
concentrations of less than 10 ug/1 for sheepshead minnow, daphnids,
and mussels, and at less than 20 ug/1 for rainbow trout, mysid shrimp,
and a marine alga.
- o Chronic effects of the 58% chlorinated short chain length n-paraffins
^
include abnormal behavior, growth effects, reduced reproduction, and
lethality.
o The extent of the effects and the maximum acceptable toxicant concen-
trations (MATC) for most studies are obscured by shortcomings and
erratic test results resulting from either poor testing procedures or
husbandry problems. Adverse effects appear at test concentrations as
low as 2.4 ug/1 and the results of adequate studies might indicate
significant adverse chronic effects below 1 ug/1.
o Studies on rainbow trout and mussels indicate extremely high levels of
bioconcentration in both species for the 58% chlorinated short chain
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length paraffins. The potential for bioconcentration of 'other chlorin-
ated paraffins is indicated by the results of both the Phase I tests
and published data.
o Additional testing is necessary on other chlorinated paraffin fornula-
tions to determine the extent of their toxicity and bioconcentration
potential.
o Test results of the Avian Reproduction Study indicate no significant
effects on mallard ducks fed 28 and 166 ppm 58 percent chlorinated
short chain length (Cio-12) n-paraffins. Significant effects reported
at 1000 ppm include: 1) statistically significant decrease in eggshell
thickness, and 2) a slight reduction in the percent of viable embryos
per egg set, which was statistically significant for Weeks 3 and 6
only.
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PRELIMINARY HAZARD ASSESSMENT CM CHLORINATED PARAFFINS
The following hazard assessment on chlorinated paraffins is based on the
testing data submitted by the Chlorinated Paraffins Consortium in response to
the negotiated testing as published in the Federal Register on January 8, 1982.
The testing agreement involved a two tiered testing scheme with Phase I tests
conducted on four representative compounds selected from a matrix (Table 1).
Phase II testing on additional aquatic species would be conducted on the most
toxic of the four compounds. As seen from the Phase I test results presented
in Table 2 for rainbow trout (Salmo gairdneri) and mussels (Mytilus edulis),
the 58% chlorinated short carbon chain (Cio-i2) length paraffin was identified
as the test material for the Phase II aquatic tests. This hazard assessment
includes the results from the nineteen ecotoxicity studies (8 Phase I and 11
Phase II) submitted by the consortium and an avian reproduction study submitted
by American members of the Consortium. Table 1 indicates what tests have been
received and validated by OTS. The Avian Study and the Reproductive Study on
ducks refer to the same test.
The format followed in presenting the hazard assessment is as follows:
each section begins with a discussion of the test results as submitted by the
consortium, followed by a comparison of the results to information found in
the published literature. For consistency, all test concentrations and their
effect levels cited in this assessment are measured concentrations, since in
many tests the measured concentration is considerably less than the nominal
level. The summary at the end of each section is an assessment ofthe import-
ance of the data and its potential impact on fish and wildlife. Due to the
large number of complex studies, the strengths and weaknesses have been
summarized, but not discussed in detail in this document under the Section
titled - Adequacy of Test Data. A detailed description of the weaknesses and
the validation for each study are available upon request.
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Acute Toxicity
The only chlorinated paraffin for which acute studies were submitted is
the 58% chlorinated short chain (€10-12) length n-paraffin. The acute aquatic
LC50/EC50 values for this chlorinated paraffin range from less than 14.1 ug/1
to greater than 162 ug/1 (Appendix). High acute toxicity was reported in
organisms from three of the six taxonomic groups. The three acutely sensitive
species included mysid shrimp (Mysidopsis bahia), 96-hour LC50 < 14.1 ug/1;
marine alga (Skeletonema costatum), 48-hour EC50 31.6 ug/1; and the waterflea
(Daphnia magna), 48-hour LC50 about 46 ug/1. The 48-hour EC50 value of 530
ug/1 reported for daphnid is considered excessively high based on the mortality
data reported during the chronic study. From mortality data on Day 2 reported
in the chronic study, the LC50 could be estimated as about 46 ug/1.
F
Many researchers have reported low acute toxicity fpr fish for chlorinated
paraffins. Johnson and Finley (1980) reported 96-hour LC50 values in excess of
300 nvg/1 for a series of static tests for various chlorinated paraffins to
rainbow trout (Salmo gairdneri) and bluegill sunfish (Depends macrochirus)
(Table 3). Acute data on channel catfish (Ictalurus punctatus) and fathead
minnow (Pimephales promelas) also indicated low toxicity (greater than 300 and
100 mg/1, respectively). Variations in test temperature between 5° and 25°C
did not reduce the static toxicity of Chlorowax 500C below a 96-hour of 300
mg/1. However, in flow-through tests with rainbow trout, sublethal effects
were noted at concentrations as low as 40 ug/1. The effects primarily involved
a progressive loss of motor function to the point of immobilization after 15
and 20 days of exposure. Death, when it occurred, resulted from debilitation
and other secondary effects. These sublethal effects were not present or
slightly expressed in bluegills and channel catfish. Madeley and Birtley
(1980) exposed rainbow trout to Cereclor 42 (C20-30 42 * cl w/w) for 96 hours
at 15°C. No mortality or abnormal behavior was noted at a mean measured level
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of 770 mg/1 (range 520-1630 mg/1). Linden et al. (1979) also tested several
chlorinated paraffins on bleaks (Albumus alburnus), a European cyprinid minnow
found in brackish water. Again, the 96-hour LC50 toxicity levels were low,
greater than 5000 or 10,000 mg/1 at 10°C (Table 4).
Recent tests with the harpacticoid copepod Nitocra spinipes have resulted
in 96-hour LC50 values below 1 mg/1 (Tarkpea et al., 1981) for several chlorin-
ated paraffin formulations having short carbon chains (<_ Ci7). These results
summarized in Bengtsson and Ofstad (1982), were unavailable. As a result, the
toxicity levels and identity of the tested chloroparaffins are unknown.
The acute LD50 data on laboratory mammals as summarized by Howard et al.
(1975) are greater than 10 gAg (Table 5). Acute LD50 and subacute LC50 data
on mallard ducks (Anas platyrynchos) and the ring-necked pheasant (Phasianus
colchicus) indicate that the chlorinated paraffin, Cereclor S52 (€14-17 an<^
52% chlorination), is not acutely or subacutely toxic to birds (Table 6).
While low acute toxicity of chlorinated paraffins has been indicated for
^
fish, tests with 58% chlorinated short chain length (Cio-12^ n-paraffins have
shown high acute toxicity to roost tested aquatic invertebrates. High acute
toxicity of other formulations is indicated in aquatic tests in which there are
acute LCso values of less than 1 mg/1 for copepods, but the information on the
identity of the tested formulations is not yet available. No toxicity informa-
tion is available on sensitive test species identified in Consortium studies
for formulations other than the 58% chlorinated short carbon chain (Cio_i2)
length n-paraffins. Furthermore, the importance of a conclusion about low
acute fish toxicity of chlorinated paraffins must be tempered by the extraordi-
narily chronic nature of these chemicals. Mortality that begins in an acute
test continues during longer exposures. For example, the 48-hour daphnid LCSO
value was reported as 530 ug/1, while chronic test data indicate an incipient
6-day LCSO value of greater than 8.9 ug/1 and less than 16.3 ug/1. The chronic
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data also indicate a 48-hour LC50 value of about 46 ug/1. When the chronic
daphnid LC50 values are plotted versus time, a near perfect chronicity curve
is produced (Figure 1). Perfect chronicity is indicated when "effect concen-
tration x time" yields a constant value. Chronic data on mysid shrimp also
indicate some chronicity. Available chronic toxicity tests indicate that
initial mortality is slow developing in some species and may even occur after
the organism has been removed from the exposure. Thus, acute toxicity results
may greatly underestimate the hazard of chlorinated paraffin formulations.
r The low solubility of chlorinated paraffins in water raiseda few questions
A
concerning the validity of some of the high LC50 values reported in the litera-
ture. None of the published articles indicated whether the test material was
fully dissolved or even which, if any, solvent was used. High levels of test
material added to water may underestimate the toxicity of a chemical, unless an
organism is exposed to a toxicant which is either dissolved or dispersed into
small enough particles as to be available.
Since the literature contains mostly low acute fish toxicity values and no
data on the more sensitive species identified in the Consortium studies, some
concern for the more sensitive species remains for all formulations. High
acute toxicity reported in copepods for several formulations indicate that not
all formulations are of low acute toxicity. Given a propensity for chronicity,
low water solubility, and the persistent nature of these chlorinated paraffins,
the significance of acute values are of limited importance and chronic effects
and incipient ££50 values should generally be considered more important to
naturally occurring populations than acute toxicity.
Chronic Aquatic Toxicity - Consortium Studies
Chronic effects were reported in all test species for most all chlorinated
paraffin formulations tested. The chronic effects include chronic mortality,
significantly (P = 0.05) increased and/or reduced growth, abnormal behavior,
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reduced filtration (feeding) activity, reduced offspring per female, offspring
mortality, reduced hatchability, reduced insect emergence, and reduced cell
growth in algae. The maximum acceptable toxicant concentration (MATC) levels
for these effects ranged as low as > 2.4 < 4.1 ug/1 and < 3.2 ug/1. The most
sensitive species were sheepshead minnow larvae, mussels, daphnids, rainbow
trout, mysid shrimp, and marine algae. All had no effect levels below 20 ug/1.
The results from the Phase I, 60-day toxicity tests on mussels and rainbow
trout exposed to four select chlorinated paraffin formulations, indicate that
the greatest chronic toxicity occurred from the 58% chlorinated short chain
(Cio-12) Ien9th n-paraffins (Table 2). The short chain paraffin with 58%
chlorination shows greater hazard potential based on all of the criteria
measured: abnormal behavior, mortality, and bioconcentration factor (BCF),
compared to the results of the other three formulations. With the exception of
the short chain length paraffins, mortality reported in the intermediate chain
length (C^-ig) paraffin (52% chlorination) and long chain length (€20-30)
paraffins of low (43%) and high (70%) chlorination was 7 percent or less for
all test levels and abnormal behavior in rainbow trout was either equivalent to
controls or transient on Days 5, 8, and 9. Abnormal behavior in mussels was
reported as reduced filtration (feeding) activity and generally occurred only
at highest test concentration. Patterns of toxicity related to carbon chain
length and/or percent chlorination could not be found, because the test concen-
trations were too low to yield effect levels for most studies.
./
Chronic effects of short chain length paraffins on mussels include chronic
mortality, reduction in shell and tissue growth, and reduction in filtration
(feeding) activity. Continual mortality throughout the test period produced a
60-day LC50 value of 74 (68 - 81) ug/1. Mortality at 13 ug/1 (0 percent) and
44 ug/1 (2 percent) are insignificant. Non-quantitative observations were made
on filtration and effects were reported on a number of occasions at 13 ug/1,
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the lowest test concentration. Significant (P'£ 0.05) reduction in the rate of
growth was reported in the growth effects study oh the 58% chlorinated short
chain length paraffins for both tissue weight and shell length." Both growth
rates were reduced 53 percent and gave a MATC of greater than 2.3 ug/1 and less
than 9.3 ug/1. Estimation of growth rates from Phase I 60-day, toxicity data
indicate values too small (0.2 - 2 percent) for comparison. Growth in Phase I
studies were lower than normal, because the mussels had been fed a diet suffi-
cient only for maintenance and not enough to support vigorous growth.
Chronic effects of short chain length paraffins on juvenile rainbow trout
include chronic mortality, abnormal behavior, and growth effects. Chronic
lethality occurred throughout the test period of 60 days with 33 percent death
at concentrations as low as 33 ug/1. Abnormal behavior in fish was reported as
persistent and slight to moderate at the lowest observed test level (33 ug/1).
The growth study reported significant (P £ 0.05) growth effects at > 3.4 ug/1
< 17.2 ug/1 (26 percent increase in weight). If the subsample weights are
assumed representative in each test group, increased growth was also evident
at 33 and 100 ug/1, while growth reductions occurred at test levels greater
than 350 ug/1 and less than 1,070 ug/1 (Table 7). The MATC reported for the
study is greater than 3.4 ug/1 and less than 17.2 ug/1 based on growth effects.
Study One on sheepshead minnow embyro-larvae indicates chronic effects
including significantly (P £ 0.05) increased growth levels at all test levels
(2.4 - 55 ug/1) for length and at all concentrations greater than 2.4 ug/1 and
s
less than 4.1 ug/1 through 55 ug/1 for weight. Growth levels for length and
weight in the second study were significantly increased at 36 and 71 ug/1 and
significantly reduced at 620 ug/1 (Table 7). The degree to which the toxicant
affected larval growth is partially obscured by the reduction in growth caused
by acetone, compared to controls. In the first study, length was significantly
(P £ 0.05) reduced 4.3 percent in acetone controls. Length reduction of 8.5
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percent in the second study, although larger was not statistically significant.
In the same studies weight in the acetone controls were reduced by 5.9 and 16.9
percent, but neither value was statistically significant. The effect of acetone
reductions on growth would be to overestimate increased growth and underestimate
growth reductions. No significant effect was found on sheepshead minnow hatch-
ability or larval survival at test levels as high as 620 ug/1.
Reproductive effects on daphnids were obscured by high variability in the
test results and non-dose-related effects. One pair of replicate controls
produced 52 percent fewer offspring compared to the other control. The lowest
test level (2.7 ug/1) produced 44 percent fewer offspring compared to acetone
controls, while 5.0 ug/1 produced only 16.4 percent fewer offspring and the
highest concentration (8.9 ug/1) produced 66 percent fewer. While none of the
>
results are statistically significant (P £ 0.05), differences between treatment
groups are apparent. The effect at the highest test level appears to be at
least partially due to the toxicant, but the degree of effect can not be
ascertained. Differences in parental length followed the same erratic effect
pattern seen in the production of offspring. The description of the test
vessel positions and similarity between results for each repetition, especially
between the two sets of controls, provide strong evidence to support the hypo-
thesis that the erratic results are due to the position of the test vessels
and not primarily due to the toxicant. In the flow-through chronic study, the
insipient daphnid LC50 value of greater than 8.9 ug/1 and less than 16.3 ug/1
,s
was attained by Day 6 and remained unchanged through Day 21. The semi-static
chronic test produced similar mortality levels with 50 percent mortality at
12.0 ug/1 from Day 7 to Day 14 and no deaths at 6.3 ug/1. No mortality
occurred in the static test before Day 5.
Chronic effects on mysid shrimp include chronic mortality, which may be
sex-related, and a possible reduction in reproduction. First generation deaths
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began on Day 2, crested on Day 21, and continued throughout the 28-day test.
Parental mortalities seemed to be sex-related with 77 percent of the deaths
being male. Unfortunately, the chronic mortality among parents"was not dose-
related. While some chronic mortality reported in the mysid study might have
been due to the test material, the effect levels on reproduction were obscured
apparently by husbandry problems. These husbandry problems also appeared to
affect the number of young per female which were too variable to identify
significant effects. The reduction in young per female at 7.3 ppb, compared to
acetone controls (7.46 versus 11.07 young/female, respectively), would appear
to be due to chlorinated paraffin toxicity. The resultant MATC for mysid
shrimp probably should be greater than 3.8 ug/1 and less than 7.3 ug/1.
Chronic effects in chironomid midges exposed to 58% chlorinated short
chain length (Cio-12^ n-paraffins include reduced hatching in Gj parental egg
masses, no emergence of adults, and reduction in the number of eggs per egg
mass. Hatching of midge larve from parental egg masses was reduced 60 percent
at test concentrations of 78 ug/1 compared to controls. At concentrations of
121 and 162 ug/1, there was no emergence of first generation adult midges. At
78 ug/1, the highest test level yeilding young, the number of second generation
eggs per mass was reduced 10 percent, but that was not significantly (P _< 0.05)
different than controls.
Behavioral effects were reported in a number of the Consortium studies on
mussels and rainbow trout. Abnormal behavioral effects in mussels were reported
in test concentrations for all formulations tested in Phase I. The unquantified
behavioral observations reported were reduced filtration (feeding) activity.
The frequency and intensity of the effect corresponded directly to the test
concentration and generally corresponded conversely to carbon chain length and
the degree of chlorination (Table 2). Behavioral effects were also evident as
reduced growth in all of the mussel studies tested with 58% chlorinated short
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chain paraffins. In the Phase I study, the reduced filter-feeding activity was
observed on a number of occasions at 18 and 56 ug/1, but caused no nortality.
Significant mortality (over 50 percent) followed abnormal behavior at 71, 130,
and 930 ug/1. No behavioral observations were reported in the growth or
bioconcentration studies, but adverse effects were indicated in both studies.
In the growth study, a significant reduction in shell and tissue growth was
reported at 9.3 ug/1, which would be expected from reduced feeding activity.
In the bioconcentration study, 33 percent mortality was reported at 10.1 ug/1
(23% during the exposure phase and 10% during depuration). Deaths at 2.35
ug/1 were not significantly higher than controls.
In the 60-day Phase I test on rainbow trout exposed to 58% chlorinated
short chain paraffins, abnormal behavior began on Days 2 and 3 for all tests
levels and showed a dose-related response. The lowest test level (33 ug/1)
affected occasional individuals and deaths began on Day 11 (33 percent by the
end of the study). The highest test level (3,050 ug/1) consistently affected
behavior in over half of the fish after Day 14 with the first deaths occurring
on Day 9. The smaller fish appeared to be affected first in any test group
and subsequently their symptoms were usually more severe. Symptoms proceeding
death followed a regular pattern which began with a slow response to food and
eventually to not feeding at all, next came lethargic behavior, an apparent
inability to change position within a depth in the aquarium, long periods on
the bottom, darkening of skin pigmentation, occasional periods of activity
**
without direction, infrequent tetanic spasms, and death within 5 to 15 days
after establishment of symptoms in an individual. Starvation was suggested as
possibly contributing to eventual death. While fish behavior appeared normal
in the growth study and no significant mortality occurred, the bioconcentration
study indicated significant dose-related mortality. While only three deaths
occurred about two-thirds of the way through the exposure phase, a total of
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thirteen fish died beginning on Day 64 through'Day 69 of the depuration phase.
All fish which had been exposed to the higher test level, 14.3 ug/1, died 36
days before the final sampling date. Only two fish survived afthe 3.1 ug/1
test level. The authors indicated that these deaths were proceeded by symptoms *
of abnomal behavior reminiscent of those described in the 60-day toxicty study.
No behavioral observations were made in the sheepshead minnow studies and no
significant larval mortality occurred.
Chronic Wildlife Toxicity - American Manufacturers
The mallard reproduction study indicate no significant effects from diets
of 28 and 166 ppm 58% chlorinated short chain length (Cio-12^ n-paraffins. The
statistically significant effects identified in mallards fed 1000 ppm include:
1) a decrease in eggshell thickness of 0.02 mm (mean 0.355 mm versus 0.375 mm
in controls), and 2) slight reduction in the percent of viable embryos per egg
set (85% versus 95% in controls), which were statistically significant only for
Weeks 3 and 6. While eggshell thinning was statistically significantyreduced
\-f
in the group fed 1000 ppm, the thickness falls within levels which the protocol
considers acceptable for for controls. The statistically significant decrease
in adult food consumption at 28 ppm during Week 17 should be considered to be
spurious and unrelated to toxicological effects. All other reproductive
parameters analyzed statistically showed no significant differences from the
controls and control reproductive parameters were within acceptable levels.
Chronic Toxicity - Published Literature
Many publications have reported chronic effects on test organisms from
exposure to chlorinated paraffins. Johnson was the first to find neurotoxic
effects on fish behavior following chronic exposure to chlorinated paraffins.
The results of the flow-though tests on rainbow trout were later reported in
Johnson and Finley (1980). Sublethal effects were seen at concentrations as
low as 40 ppb and included progressive loss of motor function to the point of
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immobilization after 15 and 20 days. Death, when it occurred, resulted from
debilitation and other secondary effects. These sublethal effects were not
seen or only slightly evident in bluegill and channel catfish. No significant
differences in susceptibility were found between yolk-sac fry, swim-up fry, and
fingerlings in. tests on rainbow trout.
Svanberg et ad. (1978) reported obvious signs of neurotoxic effects in
adult bleaks which first became apparent after exposure for 14 days at 1 mg/1,
the high dose, and later at the low dose (0.1 mg/1) of Chlorparaffin Huls 70 C
(short chain, 70% chlorination). Symptoms were similar to other studies and
included sluggish movements, disturbed orientation in the aquarium, tetanic
spasms, and death. The three deaths (7 fish/level) occurred on Days 15, 28,
and 27 at exposure levels of 0.1, 0.1, and 1.0 mg/1, respectively. Residues
in the three dead fish were 20.3, 32.8, and 47.5 ug/g, respectively. Live
fish residues reported on Day 29 in the same test were 26.2 and 28.6 ugCl/g at
exposures of 0.1 and 1.0 mg/1, respectively. While data reported by Svanberg
et^ al. indicate that the residues in dead fish were generally higher than in
live fish, insufficient data were available to confirm this observation or to
establish lethal body residue levels.
Bengtsson et cd. (1979) reported sluggish movements, absence of shoaling
behavior, and abnormal vertical positions after 7 days exposed to 125 ug/1 of
Witaclor 149, 159, and 171P in decreasing order of pronounced effects. No
abnormal behavioral effects were reported for Witaclor 350, 549, and the PCB,
s
Clophen A50, during the 14-day exposure.
Several published studies indicate similar neurotoxic symptoms in fish
following dietary exposures to chlorinated paraffins. Bengtsson and Ofstad
(1982) fed several chlorinated paraffins to bleaks and reported the following
behavioral effects. The first changed behavior was noted in the Witaclor 149
high dose group (5800 ug/g of 49% chlorinated Cio-13 paraffins) after 5 weeks
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exposure. Symptoms included sluggish swiinming closer to the bottom than usual.
After 7 and 12 weeks, the same behavior was seen in the Witaclor 149 medium
dose group (2500 ug/g) and the Witaclor 171P group (3180 ug/g of 70% chlorin-
ated Cio-13 paraffins), respectively. This behavior was generally accompanied
by folded dorsal fins and minor balance problems. The effects gradually
disappeared within a couple of weeks during the subsequent depuration period.
Evidence of chronic lethality of chlorinated paraffins had previously been
reported by Zitko (1974) in contaminated food uptake tests on juvenile Atlantic
salmon. While the differences in mean time to lethality (LT50) are evident
between the control and both chlorinated paraffins test levels, an inversion
in the LT50 data is apparent between 10 and 100 ug/g Cereclor 42 (Table 8).
Insufficient information was given to identify the source of the LT50 inversion,
but reduced feeding at the 100 ug/g concentration which was neither measured
nor reported could easily be responsible for such a toxicity inversion.
In tests where fingerling rainbow trout were fed a diet fortified with 10
ppm Chlorowax 500C (X C\2 an^ 60% chlorination) for 82 days, growth was reduced
in treated fish, but they showed no gross pathological effects (Lombardo et_
al.f 1975).
Significant chronic effects reported on 58% chlorinated short chain length
n-paraffins include chronic mortality in most species, dose-related increases
and reduction in rainbow trout and sheepshead minnow growth, growth reduction
in mussels, reduced adult chironomid emergence, reduced reproduction in mysid
shrimp and daphnids, and possibly sex-related lethality in mysid shrimp. While
chronic exposures in Phase I testing on other formulations failed to produce
significant mortality, abnormal behavioral effects were reported in mussels
and rainbow trout. These behavioral effects are especially evident for all
formulations in the form of reduced filtration (feeding) activity in mussels.
Adverse effects, other than mortality, were most apparent in sensitive test
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species which were tested only with the 58% short chain length paraffins.
Mortality and neurotoxic effects have been reported in fish studies following
both aqueous and dietary exposures to several chlorinated paraffins. The
appearance of synpterns are sequential and are similar over a wide range of
formulations.
Comparative Toxicity
Significant growth effects present in rainbow trout and sheepshead minnow
show a similar pattern of significant increases at low test levels and signifi-
cant reductions at high test concentrations. The test concentrations at which
the growth effects are found are nearly the same for both species (Figure 2).
Similarity in the growth curves for the two species strengthens the validity
of the unusual dose-response to a test substance. The mechanism responsible
for increased growth at low test levels is not understood, pifferenoes in
rainbow trout mortality in the bioconcentration and growth studies tested at
similar concentrations and exposed for 168 days each are not clear. Two
possible sources for the mortality difference are duration of observation
period and higher levels of chlorinated paraffin contamination in the fish
food (2.2 ppm versus 0.8 ppm) in the bioconcentration study. An estimate of
the residue contribution from water and from food indicates that at the higher
test levels, about 15 ug/1, the food contribution would have been only about 2
percent of the total body residue and therefore, insignificant compared to the
exposure from water. At the lower test level, about 3.0 ug/1, contaminated
s
food would have contributed about 30 percent. Analysis of the mortality data
showed that the significant mortality occurred during the depuration phase
64 to 69 days after the 168 day exposure had ceased. The timing of the deaths
and the fact that 80 percent of the whole body residues had been eliminated at
the time of death, indicate that the duration of the observation period might
best account for the discrepancy in the mortalities. Absence of toxicant-
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related mortality in sheepshead minnow larvae exposed 32 days at test levels
through 620 ug/1, a concentration level greater than which killed most rainbow
trout in the 60-day toxicity test, also supports the concept that mortality
differences might be time related. Significant differences in susceptibility
are not found either among various earlylife stages in rainbow trout or sheeps-
head minnow. Some species differences in sensitivity to chlorinated paraffins
are indicated by the occurrence of only slight or no sublethal effects in
bluegills and channel catfish compared to effects reported on rainbow trout.
Growth studies on mussels exposed to the same short-chained length chlor-
inated paraffin indicate reduced growth rates at concentrations greater than
2.3 ug/1 and less than 9.8 ug/1 (53 percent reduction in both tissue and shell
length). Toxicant levels reducing mussel growth are less than the concentra-
tions reported to reduce growth in sheepshead minnow (greater than 280 ug/1 and
less than 620 ug/1) and in rainbow trout (greater than 350 ug/1 and less than
1,070 ug/1).
Daphnids and mysid shrimp are both affected by the 58% chlorinated short
chain length paraffin at similar concentrations. The 96-hour LC50 values are
18 ug/1 and less than 14 ug/1, respectively. The number of daphnid offspring
per female is reduced 44 percent at 2.7 ug/1, the lowest test concentration,
versus a 33 percent reduction in offspring/female in mysid shrimp at 7.3 ug/1.
Chironomid midges, another aquatic invertebrate, are not as sensitive as the
above two invertebrates, but adverse reproductive effects on midge larvae were
^
reported for hatching, emergence, and eggs per mass at concentrations of either
78 or 121 ug/1.
The two species of algae reacted very differently when acutely exposed to
58% chlorinated short chain length n-paraffins. The marine alga, Skeletonema
costatum, was the more sensitive species with a 96-hour EC50 of 42.3 (27.3 -
93.1) ug/1 for growth (cell count). The effect of the test material on growth
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*
rate of the marine alga was however transitional and by Day 10 no difference in
growth rates were apparent when compared to controls (Figure 3). The highest
reduction in growth rate occurred during the first two days and produced 48-hr
EC50 of 31.6 (20.7 - 57.6) ug/1. Toxicant effects on the freshwater green
alga, Selenastrum capricornutum, differed from the marine alga in that growth
rate reduction was produced by higher test concentrations and the greatest
effect occurred at the end of the 10-day study rather than in the first few
days (Figure 4). The lowest reported EC50 for the green alga was 1,310 (880 -
4,060) ug/1 at 10 days, which was derived by extrapolation from the 45 percent
reduction found at the highest test level, 1,200 ug/1. Increasing differences
in growth rates compared to controls in the latter days of the study indicate
that longer exposure would probably produce lower effect levels for green alga.
How much lower is unknown.
Physical Test Factors Affecting Toxicity
Adverse effects due to458% chlorinated short chain length paraffins are
evident across a wide array of taxonomic groups tested. Significant (P <_ 0.05)
effects occurred in a range of about 2.4 to 20 ug/1 for most species including
both fish species, the mussel, daphnids, mysid shrimp, and one of the two algae
species tested. No toxicity pattern could be found between species found in
marine or freshwater or between species tested at high or low temperatures.
The two major factors affecting the toxicity of chlorinated paraffins appear
to be time and concentration. While the influence of the test concentration is
^
an obvious factor, the duration of the test period to the extent necessary to
show toxicity as is the case in the chlorinated paraffins is unusual. As seen
above, the duration of the exposure (168 days) and an additional 64 to 69 days
of post-exposure observation passed before significant mortality occurred in
the bioconcentration study. Very few chemicals show such prolonged developing,
chronic effects, and even fewer demonstrate delayed mortality so late into the
-------
16
*
depuration phase. While some fish species are iriore sensitive to chlorinated
paraffins than others, no toxicological pattern could be identified.
Little information is available from which the extent of chronic effects
may be determined for the other chlorinated paraffin formulations. The 60-day
toxicity studies reveal no significant chronic mortality, and the sublethal
chronic observations were not quantified. The occurrence of sublethal effects
is indicated by some reported, but unquantified, abnormal behavior, especially
the reduction of filtration (feeding) activity seen in the mussels. Neurotoxic
effects reported for rainbow trout and bleaks include a similar reduction in
feeding activity described as a slowed response to food. The results of growth
studies on mussels, rainbow trout, sheepshead minnow, and bleaks indicate
reductions in growth, which most likely result from reduced feeding. Abnormal
behavior has been reported in bleaks for other formulations of chlorinated
paraffins, especially for the shorter carbon chain lengths of all levels of
chlorination. Abnormal behavioral effects are greater for the 49% chlorinated
formulation than for 59% chlorination, which in turn is greater than for 71%
chlorination.
Aquatic Bioconcentration and Bioaccumulation
Long-term bioconcentration studies on mussels and rainbow trout exposed to
58% chlorinated short chain length paraffins demonstrated high BCF levels in
whole organisms ranging from 24,800 to 40,900 and 3,550 to 5,260, respectively.
While the data for some organs were erratic and never stabilized, equilibrium
^
between water concentrations and whole organism residue levels were reached in
about 45 to 80 days in the mussel and by about Day 90 in the rainbow trout.
Depuration half-life rates for the whole organisms were reported as 9.2 to 19.8
days in the mussel and 18.7 to 19.8 days in the rainbow trout. Of the tissues
measured the highest residues occurred in the digestive organs of both species.
BCF levels in the mussel's digestive gland/stomach ranged from 104,000 to
-------
17
226,000. In the rainbow trout, initial residue levels were highest in the
liver and viscera with BCF values of 11,430 to 15,970, but the levels in the
liver declined in the latter half of the study to 2,770 to 3,930. BCF values
found in flesh or carcass were considerably lower (1,330 to 5,040). Declining
residues in the trout liver give the impression that active elimination of Cl4,
possibly occurs via metabolic breakdown of the chlorinated paraffin.
The bioconcentration study on mussels exposed to nominal concentrations of
2.35 and 10.1 ug/1 of 58% chlorinated short chain paraffins indicate BCF values
of 40,900 and 24,800, respectively for the whole animal. Compared to the gonad
and residual tissues, the digestive gland had the highest residue levels with
BCF values of 104,000 and 226,400 at levels of 2.35 and 10.1 ug/1, respectively.
Whole animal residues attained equilibrium at the highest exposure level at
about Day 42, which also corresponded to the onset of low level mortality that
persisted throughout the 91-day exposure and through Day 125 (34 days into the
depuration period). As discussed earlier, mortality also occurred in the
rainbow trout bioconcentration study during the depuration phase. However the
trout deaths began after 64 days of elimination and ceased on Day 69, leaving
only two surviving fish at the lowest test level. As indicated above, the
contamination of the fish food source at 2.2 ppm is not considered responsible
for this late mortality during the depuration phase.
Additional data on bioconcentration are available from the Phase I, 60-day
toxicity tests, but the results must be considered preliminary, because the
^
studies did not follow OTS-recommended protocols. The two major deficiencies
in the 60-day studies are that only one sample per treatment level was made and
the test organisms were fed a maintainenoe diet rather than a diet adequate for
growth and deposition of body fat. The BCF value reported for each exposure
level provides a single data point at 60 days with no indication of either
sample variability or the maximum bioconcentration level. For most chemicals,
-------
18
60-days exposure would be expected to be sufficient to reach equilibrium, but
for the chlorinated paraffin formulations that duration has not been shown to
be adequate. The effect of a minimal diet level necessary to maintain body
functions on bioconcentration is uncertain. The preliminary BCF levels
reported in all mussel studies may be underestimated, because the mussels were
fed a maintenance diet which limited growth and the deposition of additional
lipids where chlorinated paraffins would be stored. The growth rate in the
60-day mussel tests was only 0.2 to 2 percent compared to growth of about 30
percent for the same time period in the mussel growth study. Frequently,
chemicals are bioconcentrated and deposited in fatty tissue and lipids more
effectively when the organism is actively growing. As a result of the above
deficiencies, the BCF values reported in the 60-day tests may be used, but must
be considered preliminary and are an indiction of the minimum BCF level for
each tested formulation.
The BCF values reported in the two bioconcentration studies agree well
with the results reported on the same test material in the 60-day toxicity
tests submitted on the mussel and rainbow trout. The preliminary BCF values
for the other chlorinated paraffin formulations with different carbon chain
lengths and degrees of chlorination reported in the 60-day toxicity tests also
indicate a propensity for bioconcentration. The preliminary bioconcentration
rates in both species appear to decrease with increasing chain length (Table
2). The differences in the preliminary BCF between the low (43 percent) and
s
high (70 percent) chlorinated long chain paraffins are too small to draw any
conclusion about the effect of chlorination levels on BCF. The BCF values
reported in the mussel and rainbow trout bioconcentration tests are consider-
ably higher than chlorinated paraffin values previously reported in the liter-
ature (Table 9), but in close agreement with BCF levels reported for the same
mussel species exposed to DDT (4,550-49,600) and PCB (7,200-26,600) by Geyer et
-------
19
al. (1982).
Svanberg et al. (1978) exceed bleaks to 0.1 and 1.0 ppm of Huls Chlor-
paraffin 70C (X GH.S and 70% CD which resulted in BCF levels"of 28.5 - 328 X
after 29 days exposure. Bengtssen et al. (1979) exposed fish to several chlor-
inated paraffins for 14 days followed by an elimination period of 7 days.
While their BCF values ranged from 32 to 760 X, the pattern of decreasing
bioconcentration rates with increasing chain length and increasing percent
chlorination is evident (Table 6). Residue levels after 7-days depuration
indicate residue losses of less than 50 percent for all formulations, except
the PCB formulation.
Zitko (1974) compared the level of accumulation of Cereclor 42 (42% CD,
Chlorez 700 (70% CD, and PCB, Aroclor 1254 (54% CD in juvenile Atlantic
salmon (Salmo salar) exposed via two sources, uptake of chlorinated paraffins
in suspended solids (1 g/1 of contaminated silica) for six days (Table 9)
or over 181 days in dry fish food at levels of 10 and 100 ug/g (Table 10). The
BCF level could not be calculated in either case, because data on the level of
exposure is missing for both exposures. Although the test period was consider-
ably shorter, residue levels were higher from exposure to the suspended contam-
inated silica particles as opposed to consumption in the food.
Several other authors also reported residues in fish following dietary
exposures to various formulations. Lombardo et al. (1975) fed rainbow trout a
diet with 10 ppm of Chlorowax 500C (X Ci2 and 60 % CD for 82 days. In the
f>
samples taken approximately every two weeks, chlorinated paraffin residues in
body tissues (i.e., less head, tail, and viscera) measured only as high as 1.1
ppm. Analysis of the data indicate that a residue plateau was never attained
and the rate of uptake never slowed even after 82 days.
Bengtsson and Ofstad (1982) fed several chlorinated paraffin formulations
mixed in food pellets to adult bleaks for a period of 91 days. Whole body,
-------
20
residue levels were measured periodically during the exposure and depuration
periods (Table 10). Residues of short chain length, low chlorinated paraffin
accumulated faster than the residues of the other two formulations. As might
be expected, the residue levels in the fish correlate directly to the concen-
tration in food, but the BCF values are inversely related to the dose level.
Depuration is rapid, about a 92 percent residue loss in 7 days. The short
chain, highly chlorinated and long chain length, low chlorinated paraffins
accumulate residues in bleaks more slowly than the short chain, low chlorinated
paraffins. Their depuration half-lives could not be calculated from the avail-
able figure, but they were longer than the 316-day elimination period. A
similar pattern of bioconcentration and depuration rates were reported in an
earlier study, where bleaks were tested for shorter exposure periods in water
(Bengtsson et al., 1979).
Comparable data from the Consortium studies and from published literature
indicate that bioconcentration and depuration rates for chlorinated paraffins
are inversely related to carbon chain length and the degree of chlorination
(Table 11). Comparison of residue bioconcentration rates from water and
dietary exposures indicate that chlorinated paraffins are generally taken up
more quickly from water than from food by bleaks and rainbow trout. Even when
dietary levels are over 1000 times the water concentration, uptake rates from
water are equal to or greater than from the diet (Figure 5). Uptake rates for
long chain length or highly chlorinated paraffins from water and the diet are
^
similar, despite the 1000-fold difference in their concentrations.
High levels of bioconcentration such as indicated by chlorinated paraffins
is a concern to the well-being of the organism which stores it and to any other
animal that would consume that organism. The stored chemical might adversely
affect the organism during periods of stress, such as starvation, migration,
and reproduction, or affect the developing offspring as a result of chemical
-------
21
residues stored in lipids of the egg. Bioconcentration is especially signifi-
cant, because the residues usually also accumulate in those species highest on
the food web and any adverse effect on them is accentuated by their typically
low reporductive potential. Furthermore, mortality data in the mussel and
rainbow trout bioconcentration studies indicate that adverse effects do not
cease when exposure ends. Both species experienced mortality during the depur-
ation phase. The mussel mortality at the higher test level began during the
exposure period about Day 43 and occurred persistently 34 days after the depur-
ation period started. Rainbow trout deaths began 64 days into the depuration
period and within a week all died except two fish at the lower test level. The
combination of high BCF values, slow depuration, high toxicity, persistence,
and widespread distribution of these chlorinated paraffins in the environment
are of considerable concern with respect to fish and wildlife safety.
Terrestrial Bioconcentration and Bioaccumulation
No bioconcentration data, per se, are available on chlorinated paraffins.
£
Samples from muscle, fat, and eggs of mallards were frozen during the avian
reproduction study, but no residue analyses on these tissues were made. In the
literature, the distribution of chloroparaffin residues in quail and mice was
investigated using autoradiography. Biessmann et al. (1982) gavaged Japanese
quail with two chloroparaffins (Ci2 ~ 55.9% chlorination and Cig - 34.1%
chlorination). Distribution of both chloroparaffins were nearly identical at
each sampling. Shortly after dosing the highest radioactivity was found in
s
tissues with high metabolic activity and high cell turnover rates, and in the
bile and urine. Tissues with strong labelling were: liver, intestinal mucosa,
spleen, bone marrow, oviduct, gall bladder, kidney, yolk, eggshell, and, to a
lesser, extent in the albumen. Later, the radioactivity was most evident in
follicle yolk, uropygial gland, and fat. Residue levels of C^g were about
twice that of Cj2 in the first 10 eggs laid after dosing.
-------
22
Darnerud and Brant (1982) found similar residue patterns in active tissues
in mice following intravenous and oral doses with 34.1% chlorinated hexadecane
(Cig). Marked uptake of radioactivity occurred in the brown fat, intestinal
mucosa, bone marrow, and exocrine glands. All of these tissues have high rates
of turnover and/or high metabolic capacity. No differences were found in the
distribution of residues for oral or intravenous dosing with the exception of
higher radiolabelling of the stomach and intestines in the orally-dosed mice.
Biessmann et al. (1983) repeated the above tests on quail and mice using
the same methods of administration with Cjg ~ 69% chlorination for comparison.
Distributions of residues were similar to the results above, except that little
l^COj was released indicating a different metabolic pathways. The formation of
^CC>2 and incorporation of radioactivity into metabolically active tissues are
inversely related to the degree of chlorination.
Darnerud et al. (1983) also studied residue distribution patterns in two
fish species, carp and bleaks. The distribution of 34% chlorinated hexadecane
(Cig) injected arterially into the fish was similar for the two species. Like
the quail and mice, residues were found to concentrate in active tissues, such
as kidneys, liver, nasal mucosa, and fat. Large amounts of radioactivity were
also excreted in bile for both fish. Samples taken five days after injection
also showed marked uptake in the gills, testis, and brain. After 13 days, the
residue levels were generally lower, except for the comparably high residues
seen in the liver. They concluded that the residues were incorporated in the
^
tissues and subsequently organically bound chlorine are retained for long time.
Adequacy of Test Data
Several major problem areas were recognized as pertinent to all or nearly
all of the submitted studies. The reoccurring problems identified in these
studies are of varying seriousness and have been submitted to the researchers
for clarification, comment, and improved testing in future tests.
-------
23
f
Additional Testing Needed
Several of the tests were incomplete or inconclusive including all those
tests submitted for which the test levels were too low to produce an observable
effect. If the predicted environmental concentrations (PECs) used in the risk
assessment are in the range of 0.1 to 100 ppb, some aquatic tests might have to
be repeated in order to clearly quantify the adverse effects. The studies
which may need to be retested on 58% chlorinated short chain length n-paraff ins
include:
1) Daphnia magna life-cycle test
2) Mysid shrimp life-cycle test
3) Sheepshead minnow embryo-larvae test
Other testing may be required, especially since the testing matrix has
failed to provide adequate data points from which to interpolate the toxicity
of all chloroparaff in formlations in the matrix based on percent chlorination
and chain length. These toxicity data do not permit extrapolation to other
chlorinated paraffin formulations. Phase II tests for each of the matrix
group may be needed, unless an adequate new matrix can be constructed.
Conclusions
Phase I test results indicate that the 58% chlorinated short chain
length n-paraff ins are more toxic than the other three tested chlorinated
paraffin formulations. The matrix fails to indicate if it is the most toxic
of all chlorinated paraffin formulations. While the short chain length paraf-
^
fins are more toxic, the other formulations are not without observed chronic
effects. Unquantified abnormal behavior reported, especially upon mussel
filtration (feeding) activity, indicates that chronic effects are probable in
all formulations. Preliminary bioconcentration factors indicate that all four
«
formulations will accumulate. The extent of bioconcentration in the Phase I
tests could not be asertained due to the inadequacy of the sampling methods,
-------
24
(i.e., tissue levels were measured at only one point in time with no assurance
that these slowly accumulated chemicals had reach' equilibrium with levels in
the water). Consequently, the BCF values reported for the tested formulations
in Phase I tests must be considered preliminary and minimal values. Bioconcen-
tration factors were determined in Phase II testing only for the 58% chlorinated
short chain length paraffins. In rainbow trout and mussel studies, the BCF
values were reported as 3,550-5,250 and 24,800-40,900, respectively. These
levels of bioconoentration are of considerable concern, especially when combined
with the persistence such as has been indicated for chlorinated paraffins.
Residues may be expected to enter the aquatic environment and be found in
most, if not all, organisms, especially those species at the top of the food
web.
Phase II chronic tests on 58% chlorinated short chain length n-paraffins
indicate significant (P = 0.05) chronic adverse effects in the range of 2.4 to
20 ug/1 for rainbow trout, sheepshead minnow embryo-larvae, mussels, daphnids,
mysid shrimp, and the marine alga. These effects generally include chronic
lethality, altered growth, and reduced reproduction. Shortcomings identified
in most of the studies preclude identifying the lowest effect level concentra-
tion as well as the percent of the adverse effect. Analysis of the aquatic
data indicate that adverse effects occur at the lowest concentration tested
(2.4 ug/1) and that testing at lower levels may produce significant adverse
effects below 1 ug/1.
s
Reproductive effects of 58% chlorinated short chain length n-paraffins on
mallard ducks include statistically significant effects on eggshell thickness
and percent viable embryos per egg set at 1000 ppm. The no observed effect
level found in the avian reproductive test was 166 ppm.
Distribution of chloroparaffin residues in tissues appear to be similar
for species as diverse as mussels, fish, quail, and mice. Residues tend to
-------
25
accumulate in tissues with high cell turnover 'rates and/or a high metabolic
capacity. Rates of C£>2 formation and levels of residue incorporation into
metabolically active tissues appear to be inversely related to the degree of
chlorination.
Summary
The chronic studies on the 58% chlorinated short chain paraffins indicate
a concern for the well-being of aquatic organisms across a broad spectrum of
taxonotiic groups at environmental levels of 2.4 to 20 ug/1. Adverse effects
occur below that level into the parts per trillion range are indicated, but
they can not be assessed due to the inadequacies in the studies identified
above. Also, available data do not demonstrate that the most toxic chlorinated
paraffin has been identified. The 60-day toxicity data on the other paraffin
formulations are also insufficient to conclude that they have no adverse
chronic effects. In fact, the unquantified observations of reduced filtration
(feeding) activity reported in mussels indicate that effects on mussel growth
are probable for all tested formulations. The breadth of toxic effects on a
wide variety of species from various environments indicate that chlorinated
paraffins pose a potential threat to a wide variety of aquatic species in
freshwater, estuarine, and marine environments.
Chloroparaffins are much less toxic to mammals and birds than to aquatic
species. Acute toxicity to these two groups is virtually non-existent.
Moderate chronic effects occur in avian reproduction. Avian reproductive
s
effects include statistically significant reductions in egg shell thickness
and the percent of viable embyros per egg set at 1000 ppm (NOEL 166 ppm).
Organically-bound chlorine residues appear to concentrate in tissues with
high cell turnover rates and/or a high metabolic capacity.
-------
Literature Cited
Bengtsson, B.-E.f O. Svanberg, and E. Linden. 1979. Structure related uptake
of chlorinated paraffins in bleaks (Alburnus alburnus). Ambio 8(2-3):
121-122.
Bengtsson, B.-E. and E. B. Ofstad. 1982. Long-term studies on uptake and
elimination of some chlorinated paraffins in the bleak, (Alburnus
alburnus). Ambio 11(1):38-40.
Biessman, A., I. Brandt, and P. 0. Darnerud. 1982. Comparative distribution
and metabolism of two l^oiabeHecl chlorinated paraffins in Japanese quail
Cotumix coturnix japonica. Environ. Poll. (Ser. A) 28(2):109-120.
Biessman, A., P. O. Darnerud, and I. Brandt. 1982. Chlorinated paraffins:
Deposition of a highly chlorinated polychlorohexadecane in mice and quail.
Arch. Toxicol. 53(l):79-86.
Birtley, R. D. N., D. M. Conning, J. W. Daniel, D. M. Ferguson, E. Longstaff,
and A. A. B. Swan. 1980. The toxicological effects of chlorinated
paraffins in mammals. Toxicol. Appl. Pharmacol. 54(3):514-525.
Campbell, I. and G. McConnell. 1980. Chlorinated paraffins in the environment.
1. Environmental occurrence. Environ. Sci. Technol. 14(10):1209-1214.
Darnerud, P. O., B.-E. Bengtsson, A. Bergman, and I. Brandt. 1983. Chlorinated
paraffins: Disposition of a polychloro-[l-14c]-hexadecane in carp
(Cyprinus carpio) and bleak (Alburnus alburnus).
Toxicol. Lett. 19(3):345-351.
Darnerud, P. O., A. Biessmann, and I. Brandt. 1982. Metabolic fate of chlorin-
ated paraffins: Degree of chlorination of [l-l^cj-chlorododecanes in rela-
tion to degradation and excretion in mice. Arch. Toxicol. 50(2).:217-226.
Darnerud, P. O. and I. Brandt. 1982. Studies on the distribution and metabolism
of a 14c-labelled chlorinated alkane in mice. Environ. Poll. (Ser. A)
27(l):45-56.
Geyer, H., P. Sheehan, D. Kotzias, D. Freitag, and F. Korte. 1982. Prediction
of ecotoxicological behaviour of chemicals: Relationship between physico-
chemical properties and bioaccumulation of organic chemicals in the mussel
Mytilus edulis. Chemosphere 11(11):1121-1134.
Howard, P. H., J. Santodonato, and J. Saxena. 1975. Investigation-of selected
potential environmental contaminants: Chlorinated paraffins. Final
Report. Contract No. 68-01-3101.' Project L1259-05. U.S. EPA, Document
EPA-560/2-75-007. 109 p.
Linden, E., B.-E. Bengtsson, O. Svanberg, and G. Sundstrom. 1979. The acute
toxicity of 78 chemicals and pesticide formulations against two brackish
water organisms, the bleak (Alburnus alburnus) and the harpacticoid
Nitocra spinipes. Chemosphere 8(11/12):843-851.
Lcmbardo, P., J. L. Dennison, and W. W. Johnson. 1975. Bioaccumulation of
chlorinated paraffin residues in fish fed Chlorowax 500C. J. Assoc. Off.
-------
Anal. Chem. 58(4):707-710.
Madeley, J. R. and R. D. N. Birtley. 1980. Chlorinated paraffins and the
environment. 2. Aquatic and avian toxicology. Environ. Sci..Technol.
14(10):1215-1221. - *
Renberg, L., G. Sundstrom, and K. Sundh-Nygard. 1980. Partition coeffients of
organic chemicals derived from reversed phase thin layer chromatography:
Evaluation of methods and application on phosphate esters, polychlorinated
paraffins and some PCB-substitutes. Chemosphere 9(11):683-691.
Stetten, N. D., Jr. 1943. Metabolism of a paraffin. J. Biol. Chem. 147( ):
327-332.
Svanberg, 0. 1978. Chlorinated paraffins — A case of accumulation and
toxicity to fish. Ambio 7(2):64-65.
Svanberg, O. and E. Linden. 1979. Chlorinated paraffins — An environmental
hazard? Ambio 8(5):206-209.
Tarkpea, M., E. Linden, B.-E. Bengtsson, A. Larson, and 0. Svanberg. 1981.
Nat. Swed. Environ. Prot. Bd., Brackish Water Tox. Lab., Rep. NBL Rapp.
111. 22 p.
Zitko, V. 1974. Uptake of chlorinated paraffins and PCB from suspended solids
and food by juvenile Atlantic salmon. Bull. Environ. Contain. Toxicol.
12(4):406-412.
-------
Table 1. Tiered testing scheme agreement published in the Federal Register on
January 8, 1982 between the Chlorinated Paraffins Consortium and the
Environmental Protection Agency.
Phase I Testing *
Percent Chlorination by Weight
Carbon Chain
Length:
c 10-12
C 14-19
C 20-30
40 to 50 % Cl | 50 to 60 % Cl | 60 to 70 % Cl
** liquid - 58 % Cl
liquid - 52 % Cl
liquid - 43 % Cl solid - 70 % Cl
**
Each compound was tested for 60-day lethal and sublethal effects on mussel
(Mytilus edulis) and rainbow trout (SaLno gairdneri ) .
According to chlorine content (58 percent), the short chain
length paraffins belong in this column and not the column indicated in
the Federal Register.
Phase II Testing
Additional testing was to be conducted on the 58 % chlorinated short chain
(^10-12) length paraffins, which were already known to be the most toxic from
Phase I. The tests included:
Growth (rainbow trout and mussel)
Bioconcentration (rainbow trout and mussel)-v
Life Cycle (Daphnia) -
Life Cycle (mysid shrimp)- ^>
Embryo-juvenile (sheepshead minnow)
14-Day Bioassay (freshwater alga)
14-Day Bioassay (marine alga)
Chronic (partial life-cycle) (midge) w^'*
Solubility
Biodegradation (aerobic and anaerobic)
Avian Study (test substance to be selected)***
Reproductive Study (duck)****
Received and Validated
X X
X X
X X
X X
X X
X X
X X
X X
X not validated
X not validated
X X
X . X
*** Remarks made by the Chlorinated Paraffins Corsortium indicate that this
study refers to the duck reproduction study and is not a separate study.
**** This study is not a part of the proposal by the International Chlorinated
Paraffins Manufacturers Consortium, but it was submitted by the American
members of the Consortium.
-------
Table 2. Results of Phase I and Phase II testing on rainbow trout and mussels.
Rainbow Trout Salmo gairdneri
60-Day LC50 Abnormal Growth Whole Animal
Behavior MATC BCF
Overall MATC
Short-chain (Cio-12)
58 % Cl
Intermediate-chain
52 % Cl
Long-chain (C20-30>
43 % Cl
70 % Cl
200 ug/1 < 33 ug/1
(340 ug/D*
(continuing deaths)
(33 ug/1 - 33% dead)
> 3.4 -
< 17.2 ug/1
(26% increase
in weight
growth rate)
> 4,500 ug/1
(3 % dead;
no control
deaths)
» 4,000 ug/1
(0 deaths)
> 3,800 ug/1
(3 % dead;
3-6%
control
deaths)
> 1,050 -
< 4,500 ug/1
(transient on
Days 5, 8, & 9)
> 4,000 ug/1
(all normal)
abnormal
behavior in
all groups
(highest
effect in
controls)
3550-5260
> 3.4 < 17.2 ug/1
(increased weight)
45 - 67**
1,050 < 4,500 ug/1
(abnormal behavior)
18 - 38**
6 - 54**
> 4,000 ug/1
(no effect)
> 3,800 ug/1
' (no sign.
effect)-
* This reported value has been replaced by a lower estimated toxicity level.
** These bioconcentration factors are preliminary and have not been confirmed by acceptable studies.
-------
Table 2. (Cont.)
Short-chain (Cio-12)
58 % Cl
Intermediate-chain
52 % Cl
Long-chain (C20-30>
43 % Cl
70 % Cl
Mussel Mytilus edulis
60-Day LC50
Abnormal
Behavior
Growth
MATC
Whole Animal
BCF
Overall MATC
74 ug/1
(68 - 81)
(chronic
mortality)
» 3,800 ug/1
(2 % death
at 220 ug/1;
0 - 4 %
control
deaths)
» 2,180 ug/1
(0 deaths)
»1,330 ug/1
(0 deaths)
< 13 ug/1
(occasional
reduction
in feeding)
> 2.3 -
< 9.3 ug/1
(53 % red.
in tissue and
shell growth
rates)
24,800-40,900
> 2.3 < 9.3 ug/1
(growth reduction)
< 220 ug/1
(transient
reduction
in feeding)
< 3,800 ug/1
(consistent
red. in feeding)
> 120 -
< 2180 ug/1
(consistent
reduction
in feeding)
> 460 -
< 1330 ug/1
(marginally
reduced feeding)
430- 2,860*
< 220 ug/1
(abnormal behavior)
260- 1,160*
> 120 < 2,180 ug/1
(abnormal behavior)
220- 340*
> 460 < 1,330 ug/1
(abnormal behavior)
These bioconcentration factors are preliminary and have not been confirmed by acceptable studies.
-------
Table 3. Acute toxicity values for Chlorowax reported by Johnson and Finley
(1980).
96-Hour LC50 (ng/1)
Bluegill Channel Fathead Rainbow
Sunfish Catfish Minnow Trout
Chlorinated Carbon %
Paraffin Length Cl
Chlorowax 500C 10-13 59 > 300
Chlorowax 40 20-30 40 > 300
Chlorowax LV 20-30 46 > 300
Chlorowax 50 20-30 50 > 300
Chlorowax 70 20-30 70 > 300
> 300 > 100 > 300
> 300
> 300
> 300
> 300
"Note: Variations in test temperature between 5° and 25°C did not reduce the
static toxicity of Chlorowax 500C below a 96-h LC50 value of 300 mg/1.
Sublethal effects were noted in flow-througfh tests with rainbow trout
in concentrations as low as 40 ug/1."
Table 4. Acute toxicity values for bleaks exposed to chlorinated paraffins
(Linden et al., 1979).
Chlorinated
Paraffin
Witaclor 49
Witaclor 55EN
Witaclor 63
Witaclor 71P
Chlorparaffin huls 70C
Chlorparaff in huls 40G
Witaclor 50
Cereclor S52
Cereclor 42
Carbon
Length
10-13
10-13
10-13
10-13
11.5
15.5
14-17
14-17
22-26
%
Cl
49
56
63
56
70
40
50
52
42
96-Hour LC50 (ng/1)
Bleaks
(Alburnus alburnus)
> 5,000
> 10,000
> 5,000
> 5,000
> 10,000
> 5,000
> 5,000
> 10,000
> 5,000
-------
Table 5. Acute mammalian toxicity values summarized by Howard et'al_. (1975)
for various chlorinated paraffins.
Acute Oral LD50 (g/kg)
Chlorinated
Paraffin
Chlorowax 500C
Chlorowax 40
Chlorowax 70
Chlorez 700
Khp 470
Carbon
Length
10-13
20-30
20-30
20-30
%
Cl
59
40
70
70
48
Rat
> 21.5
» 10
» 50
» 50
26.1
Mouse Guinea
Pig
» 25
» 25
21.85
» Indicates no mortality at or below that dose level.
Table 6. Acute toxicity values reported by Madeley and Birtley (1980) for two
chlorinated paraffins.
Acute Oral Subacute Dietary 96-Hour
LD50 (gAg) LC50 (ppm) LC50 (ug/1)
Ring-necked Mallard Ring-necked Mallard Rainbow
Pheasant Duck Pheasant Duck Trout
Chlorinated Carbon %
Paraffin Length Cl
Cereclor S52 14-17 52 » 24.606 » 10.280 » 24,063 » 24,063
(loss of (reduced^
weight) feeding)
Cereclor 42 20-30 42 » 770*
» Indicates no mortality at or below that dose level.
* Toxicity value too high (water solubility was exceeded as indicated by
the reported presence of an opaque emulsion).
-------
Table 7. Growth effects on weight and length caused by 58% chlorinated short
(CIQ-IS) chain length n-paraffins in rainbow trout and sheepshead
minnow.
Percent Difference Compared to Solvent Controls
Weight Length
Test
Concentrat ion
(ug/1)
2.4
3.4
4.1
6.4
17.2
22.1
33
36.2
54.8
71.0
100
161.8
279.7
350
620.5
1,070
3,050
Rainbow Sheepshead
Trout Minnow
+ 3.8
+ 0.02
+ 14.9*
+ 31.3*
+ 25.4*
+ 27.5*
+ 37.9**
+ 21.3*
+ 31.7*
+ 15.1*
+ 13.5**
+ 13.0
+ 1.9
- 3.1**
- 30.9*
- 57.6**
- 74.7**
Rainbow Sheepshead
Trout Minnow
+ 4.0*
+ 0.7
+ 3.7*
+ 5.5*
+ 6.2
+ 7.2*
+ 34.2**
+ 7.4*
+ 6.4*
+ 5.6*
+ 6.7**
+ 3.4*
+ 1.9
- 1.9**
- 9.2*
- 28.7**
- 32.6**
* Statistically significant (P £ 0.05) differences conpared to the acetone
control.
** Statistical analyses were not reported for these values. '
-------
Table 8. Time to lethality (LT50) in juvenile Atlantic saliton (zitko, 1974).
Control
Cereclor 42
Chlorez 700
Carbon
Length
% Cl
20-30
42
20-30 70
Feeding Cone.
(ug/g)
10
100
10
100
LT50
(days)
138
47
80
71
39
-------
Table 9. Bioconcentration studies and BCF values resulting from the uptake of
residues from the water column as reported in Consortium generated
studies and in published literature.
Chlorinated
Paraffin Carbon
(Data Source) Length
Short Chain 10-13
(Consortium,
unpublished)
Short Chain 10-13
-
Intermediate Chain 14-17
Long Chain 20-30
Long Chain 20-30
Short Chain 10-13
(Consortium,
unpublished)
Short Chain 10-13
Intermediate Chain 14-17
Long Chain 20-30
Long Chain 20-30
Exposure
% Test Species Time Route
Cl (days)
59 Mussels
59 Mussels
52 Mussels
43 Mussels
70 Mussels
59 Rainbow
Trout
59 Rainbow
Trout
52 Rainbow
Trout
43 Rainbow
Trout
70 Rainbow
Trout
147
91
60
60
60
60
60
60
60
60
60
60
168
168
60
60
60
60
60
60
60
60
60
60
60
60
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water BCF
Cone, (whole
(ug/1) animal)
2.35
10.1
13
44
71
130
220
3,800
120
2,180
460
1,330
3.1
14.3
33
100
350
1,070
3,050
1,050
4,500
s
970
4,000
840
1,900
3,800
40,900
24,800
329
723
411
12,177
2,856
429
1,158
261
341 _
223
3,550
5,260
7,155
7,816
3,723
2,642
1,173
44.9
66.7
17.9
37.6
53.8
5.7
32.5
7
7
-------
Table 9. (cont.)
Chlorinated
Paraffin Carbon
(Data Source) Length
Huls chlorparaff in 10-13
70C (Svanberg et
al.r 1978)
Witaclor 149 10-13
(Bengtsson et
- al., 1979)
Witaclor 159 10-13
Witaclor 171P 10-13
Witaclor 350 14-17
Witaclor 549 18-26
PCB (Clophen A50)
Cereclor 42 20-30
(Zitko, 1974)
Chlorez 700 20-30
PCB (Aroclor 1254)
%
Cl
70
49
59
70
50
49
50
42
70
54
Test Species
Bleaks
Bleaks
Bleaks
Bleaks
Bleaks
Bleaks
Bleaks
Atlantic
salmon
Atlantic
salmon
Atlantic
salmon
Exposure
Time Route
(days)
15
28
29
27
29
14
14
14
14
14
14
2
6
2
6
1
2
6
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Water
Suspended
solids
Suspended
solids
Suspended
solids
Water BCF
" Cone, (whole
(ug/1) animal)
0.1
0.1
0.1
1.0
1.0
125
125
125
125
125
125
ib
lb
Ib
2033
328*
262
47.5a
28.6
760
720
160
40
20
1920
0.44C
0.75C
0.22C
0.46C
19. 9C
28. 3^
134C
a These Bcf values were derived from single fish that died during- the study.
b since the concentration of suspended solids in the water was not reported,
the value indicated is the concentration of chlorinated paraffins that was
adsorbed onto suspended solids in the water column.
c These BCF values are reported as chlorine (ug/g wet wt.) in fish, because
the final concentration of test material present in the water column is
unreported.
-------
Table 10. Bioconcentration studies and BCF values reported in the literature
for the uptake of chlorinated paraffin residues in food.
Chlorinated
Paraffin Carbon
(Data Source) Length
Chlorowax 500C 10-13
(Lombardo et
al., 1975)
Witaclor 149 10-13
(Bengtsson and
Ofstad, 1982)
Witaclor 149 10-13
Witaclor 149 10-13
Witaclor 171P 10-13
Witaclor 549 18-26
Cereclor 42 20-30
(Zitko, 1974)
Chlorez 700 20-30
PCB Aroclor 1254
Test
% Species
Cl
60 Rainbow
trout
49 Bleaks
49 Bleaks
49 Bleaks
71 Bleaks
49 Bleaks
42 Atlantic
salmon
70 Atlantic
salmon
54 Atlantic
salmon
Exposure
Time Route
(days)
82
91
91
91
91
91
33
109
181
33
109
181
33
109
181
33
109
181
33
109
181
33
109
181
Food
Food
Food
Food
Food
Food
Food
Food
Food
Food
Food
Food
Cone.
(ug CP/
g food)
10
590
2500
5800
3180
3400
10
100
10
100
10
100
'^
BCF Uptake
Efficiem
2.759 3
41a 45
9a 10
4.6a 5
5.53 6
2a 2
O.llb
nd
nd
O.Slb
nd
nd
0.29b
nd
nd
0.49b
nd
nd
3.86b
3.80b
3T.88b
13. 9b
24. Ob
30. Ob
a These BCF values were calculated as using the total daily consumption level
of chlorinated paraffins as the exposure level.
t> These values are reported as chlorine (ug/g wet wt.) in fish, because BCF
values could not be calculated without knowing the amount of toxicant eaten.
-------
Table 11. Characteristics indicative of bioconcentration potential in fish for
various groupings of chlorinated paraffins.
Chain Length
ClO-13
H20 Sol.(ug/l)
log P
BCF fron:
H20 - max.
(60 days)
(14 days)
Food - max.
(91 days)
Rates of:
Accumulation
Depuration
C14-17
H20 Sol.(ug/l)
log P
BCF from:
H2O - max.
(60 days)
(14 days)
Food
Rates of:
Accumulation
Depuration
C18-26
H20 Sol.(ug/l)
log P
BCF from:
H2O - max.
(14 days)
Food
(91 days)
Rates of:
Accumulation
Depuration
C20-30
H20 Sol.(ug/l)
log P
BCF fron:
H20 - max.
(60 days)
Food
Rates of:
Accumulation
Depuration
Percent Chlorination
42 - 50
59 - 60
-70 - 71
4.39 - 6.93
760 ( 125 ppb)
33 ( 590 ppm)
10 (2500 ppm)
95 - 470 (Cjj)
4.48 - 7.38
3,550 - 5,260
1,173 - 7,816
720 ( 125 ppb)
fast and constant fast and constant
92% loss in 7 days half-lives of
9 to 21 days
5-27 (C15)
5.47 - 8.21
64 ( 125 ppb)
slow
little in 7 days
7.46 - 12.83
32 ( 125 ppb)
2 (3400 ppm)
slow and constant
less than 50% loss
in 316 days
3.6 - 6.6 (C25)
8.69 ->12.83
17.9 - 37.6
44.9 - 66.7
5.37 - 8.69
160 ( 125 ppb)
6 (3180 ppm)
fast and constant
less than 50% loss
in 316 days
6 (C25)
'5.7'- 32.5
-------
Figure 1. EC50 chronicity in the waterflea Daphnia magna following exposure to
58 % chlorinated short chain (Cio_i3) length n-paraffins.
600
500
C
o
n
c
e
n 400
t
r
a
t
o
n 300
i
n
g
/ 200
1
100
456
Time (days)
10
* The acute 48-hour EC50 value reported in the daphnid study.
-------
Figure 2. Growth effects on weight caused by 58% chlorinated short chain
length n-paraffins in rainbow trout and sheepshead minnows.
P
e
r
c
e
n
t
o
f
50
40
30
20
10
S
o
i q
V
e
n -10
t
X
0 0
« o
x o
c
o
h
t
r
o
1
G
r
o
w
t
h
-20
-30
-40
-50
-60
-70
•
-80
X
\ I T
10 100 1,000
log Test Concentration (ug/1)
10,000
(X) - Rainbow Trout
(0) - Sheepshead Minnow
-------
Figure 3. Growth of the marine alga, Skeletonema costatum, cultured in a 58 %
chlorinated short chain (^0-13) length n-paraffin (particle count).
CO
LU
-J
CJ
cc
0=
Q.
10
DflYS
-------
Figure 4. Growth of the freshwater green alga, Selenastrum capricornutum,
cultured in 58 % chlorinated short chain (Cio_i3) length n-paraffin
(particle count).
cn
LJ
u
CE
L9
KEY f
O QJNTHUL
——SOL. CONTRJL
O % 0.18 MCA.
* B.32 HC/,
t 1.55 MCA.
l.J MC/L
1.8 MC/L
—X- 3.2 MCA.
DflYS
-------
APPENDIX. Results of Phase II testing of 58 % Chlorinated Short-Chain Length (Cio-12) n-Paraffins on
additional species.
Test Species
Test Type
LC50
Overall MATC
MATC MATC
Hatchability Survival
(percent) (percent)
MATC
% Growth Rate
Length Weight
Sheepshead Minnow Embryo-larvae
Cyprinodon variegatus (Study # 1)
(Study I 2)
> 2.4 < 4.1-55 ug/1
(increased growth)
> 55 ug/1 > 55 ug/1 > 2.4 - > 2.4
(77-95) (68-90, 55 ug/1 <4.1-55 ug/1
88 -100) (4 - 7 % (14 - 31 %
increase) increase)
< 36-71 < 162 ug/1 > 620 ug/1 > 620 ug/1 < 36- 71
(increased growth) (80 -
> 280 < 620 ug/1
(reduced growth)
95) (65.8- 90.7, < 162 ug/1
75.8-100) (5 - 7 %
increase)
> 280 -
< 620 ug/1
(9 % red.)
Waterflea
Daphnia magna
Life-cycle
Mysid Shrimp Life-cycle
Mysidopsis bahia
530 ug/1*
46 ppb
(48-hr EC50)
12 ug/1*
8.9 < 16.3 ug/1
(6-21 day EC50)
14.1 ug/1*
< 14.1 ug/1
(96-hr LC50)
< 2.7 ug/1* < 2.7 ug/1*
(reduced young (44 % red.
per female) offspring
< 8.9 ug/1* /female)
(66 % red. in
total reprod.)
> 7.3 < 13.7 ug/1* > 5.0 -
> 0.6 < 1.2 ug/1 < 7.3 ug/1
(sign, parental (33 % red.
mortality) offspring
/female)
Midge Life-cycle
Chironomus tentans
> 162 ug/1
(48-hr LC50
no deaths)
> 60 < 78 ug/1
(red. hatching)
> 60 -
< 78 ug/1
(60 % red.
hatching)
< 36- 71
< 162 ug/1
(15 - 21 %
increase)
> 280 -
< 620 ug/1
(31% red.)
> 5.0 - > 8.9 ug/1*
< 8.9 ug/1* (1 % red.)
(37 % dead
offspring
not sign.)
> 0.6 - > 7.3 ug/1 > 7.3 ug/1
< 1.2 ug/1 ( 1 % ( 0.4 %
(40-50 % Increase) reduction)
parental
deaths)
> 78 - < 78 ug/1 < 78 ug/1
< 121 ug/1 (10 % (1 %
(no red. in red. in
emergence) eggs/mass) hatch)
Data value can not be used with confidence.
-------
APPENDIX (Cont.)
Test Species
Test Type
LC50
MATC
Cell Growth
(particle count)
Green Alga
Selenastrum
capricomatum
Acute
3,690 ug/1*
> 1,200 ug/1
(96-hr EC50)
1,310 ug/1*
> 1,200 ug/1
(10-day EC50)
> 390 < 570 ug/1
(35 % reduction
in growth)
> 390 < 570 ug/1
(35 % reduction
in cell growth)
Marine Alga
Skeletoneroa
costatum
Acute
31.6 ug/1
(48-hr EC50)
42.3 ug/1
(96-hr EC50)
> 69.8 ug/1
(10-day EC50)
> 12.1 < 19.6 ug/1
(44 % reduction
in growth on Day 2)
> 19.6 < 43.1 ug/1
(Day 4 - 34 % red.)
> 69.8 ug/1
(Day 10 - no sign.)
> 12.1 < 19.6 ug/1
(44 % reduction
in growth on Day 2)
> 19.6 < 43.1 ug/1
(Day 4 - 34 % red.)
> 69.8 ug/1
(Day 10 - no sign.)
* Data can not be used with confidence.
------- |