Analysis of Chronic Toxicity Data 

and Acute Chronic Ratios (ACRs) in 

Support of Deriving Chronic HC5s: 

Acetylcholinesterase Inhibitors

David R Mount and Russell J Erickson

Office of Research and Development, Duluth, MN

1

Presentation to the FIFRA Scientific Advisory Panel 

January 31, 2012
Acute Chronic Ratios (ACRs)

• Ratio of acute effect concentration to chronic 

effect concentration

• Acute effect conc / chronic (threshold) effect conc

• Acute effect concentration typically the 96-h 

LC50 (48-h for some species)

• Chronic effect concentration may be

• Chronic NOEC

• Geomean of chronic NOEC and LOEC

• Chronic effect point estimate such as EC20

• Will use subscripts to differentiate

• ACRNOEC , ACRGM , ACREC20 , respectively 2
Distributions of Toxicity Data

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99.8

Acute

Effect Concentration (g/L) 3

0.03 0.1 0.3 1 3 10 30 100 300 1000 3000

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0.5

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70

Acute

HC5
Distributions of Toxicity Data

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Chronic

Acute

Effect Concentration (g/L) 4

0.03 0.1 0.3 1 3 10 30 100 300 1000 3000

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0.5

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50

70

Acute

HC5

Chronic

HC5
Distributions of Toxicity Data

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Chronic

Acute

Effect Concentration (g/L) 5

0.03 0.1 0.3 1 3 10 30 100 300 1000 3000

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0.5

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Acute

HC5

Chronic

HC5

ACRHC5
Distributions of Toxicity Data

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Chronic

Acute

ACR

Large

Effect Concentration (g/L) 6

0.03 0.1 0.3 1 3 10 30 100 300 1000 3000

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0.5

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Acute

HC5

Chronic

HC5

ACRHC5
Distributions of Toxicity Data

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Chronic

Acute

ACR

Large

Effect Concentration (g/L) 7

0.03 0.1 0.3 1 3 10 30 100 300 1000 3000

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0.5

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Acute

HC5

Chronic

HC5

ACRHC5

ACR

Small
Distributions of Toxicity Data

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Chronic

Acute

ACR

Large

ACRs vary widely within and across chemicals; how do we 

estimate ACRHC5 when chronic data are limited or unavailable?

Effect Concentration (g/L) 8

0.03 0.1 0.3 1 3 10 30 100 300 1000 3000

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Acute

HC5

Chronic

HC5

ACRHC5

ACR

Small
Scope of Current ACR Analysis

• Presume that ACRs reflect underlying 

toxicokinetic and toxicodynamic

characteristics of chemicals

• Hypothesize considering mode of action 

(MOA) can help reduce, or at least parse, 

uncertainty in estimation of ACRHC5

• Begin by focusing on chronic toxicity data for 

AChE inhibiting chemicals

9
Data Compilation

• 119 chronic tests for AChE inhibitors compiled from:

• Existing ORD data compilations

• FIFRA DERs

• ECOTOX searches

• Treatment level data for each endpoint reviewed and 

screened (details in Appendix)

• 73 tests passed screening and had defined NOEC 

and LOEC values

• NOEC/LOEC for individual tests based on most 

sensitive endpoint relating to survival, growth, or 

reproduction

10
Data Distribution

Americamysis 

bahia

Ceriodaphnia 

dubia

Daphnia 

magna

Cyprinodon 

variegatus

Jordanella 

floridae

Menidia 

beryllina

Menidia 

peninsulae

Oncorhynchus 

mykiss

Opsanus 

beta

Pimephales 

promelas

Aminocarb 1

Carbaryl (Sevin) 1

Carbofuran 2 1

Methomyl 1 1 1 1

Propoxur (Baygon) 1

Azinphos-methyl 1

Carbophenothion 1

Chlorpyrifos (Dursban) 2 1 6 1 1 1 6

Diazinon 1 1 1 3

11

Diazinon 1 1 1 3

Dichlorvos (DDVP) 1 1 1

Disulfoton 1 1 1 2

Ethoprop 1 1 1

Fensulfothion 1

Fenthion 1

Fonofos 1 1 1 1

Malathion 1 1 1 1

Methidathion 1 2

Methyl parathion 1 2 1

Naled 1 1 1

Parathion 1

Phosmet 1 1

Profenofos 1 1

Terbufos 1
Acute Data

• Acute value for ACR selected based on 

following priorities:

• 1) value from concurrent study

• 2) value from same laboratory but from a different 

time

• 3) value (or geomean) from ICE database, with 

preference for

• Meets ICE minimum modeling requirements

• Measured chemistry

• Flow-through methods

• Following EPA/OPP methods

12
Aggregate ACR Distribution

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Acute Chronic Ratio (LC50/GM)

0.1 0.3 1 3 10 30 100 300 1000 3000 10000

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0.2

0.5

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50
Aggregate ACR Distribution

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Acute Chronic Ratio (LC50/GM)

0.1 0.3 1 3 10 30 100 300 1000 3000 10000

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0.2

0.5

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5

10

20

30

50

Acute Chronic Ratio (LC50/GM)

0.1 0.3 1 3 10 30 100 300 1000 3000 10000

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0.2

0.5

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50

Invertebrates

Fish
Fish ACRs > Invertebrate ACRs

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Acute Chronic Ratio (LC50/GM)

0.1 0.3 1 3 10 30 100 300 1000 3000 10000

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50

Invertebrates

Fish
OP ACRs ≈ Carbamate ACRs

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Acute Chronic Ratio (LC50/GM)

0.1 1 10 100 1000 10000

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50

Invert Carbamate

Invert OP

Fish Carbamate

Fish OP
Acute Sensitivity to AChE Inhibitors

Planktonic

Invertebrates

(Daphnia)

Fish

F

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y

0.8

1.0

1.2

Fish roughly 100x less 

acutely sensitive than 

daphnids to AChE 

inhibitors

17

Relative Acute LC50 for AChE Inhibitors

1 10 100 1000 10000

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0.0

0.2

0.4

0.6

Extracted from Erickson (Appendix D)
Fish GMs > D. magna GMs

Fis

h

G

M

(

u

g

/

L

)

10

100

1000

Fish chronic

less sensitive

D. magna GM = Fish GM

D. magna GM (g/L) 18

0.01 0.1 1 10 100 1000

Fis

h

G

M

(

u

g

/

L

)

0.01

0.1

1

Fish chronic

more sensitive

D. magna GM = Fish GM
Large Fish ACRs ≠ High Sensitivity

N

O

E

C

f

r

o

m

Fis

h

C

h

r

o

nic

T

e

s

t (

g

/

L)

100

1000 Fish NOEC

Fish Unbounded NOEC

Fish chronic

less sensitive

1:1 Line

Daphnia magna 48-h LC50 (g/L) 19

0.1 1 10 100

N

O

E

C

f

r

o

m

Fis

h

C

h

r

o

nic

T

e

s

t (

0.1

1

10

Fish chronic

more sensitive
Large Fish ACRs ≠ High Sensitivity

N

O

E

C

f

r

o

m

Fis

h

C

h

r

o

nic

T

e

s

t (

g

/

L)

100

1000 Fish NOEC

Fish Unbounded NOEC

Fish chronic

less sensitive

Estimated

Acute HC5

(Mean EF)

Daphnia magna 48-h LC50 (g/L) 20

0.1 1 10 100

N

O

E

C

f

r

o

m

Fis

h

C

h

r

o

nic

T

e

s

t (

0.1

1

10

Fish chronic

more sensitive

(Mean EF)
Large Fish ACRs ≠ High Sensitivity

N

O

E

C

f

r

o

m

Fis

h

C

h

r

o

nic

T

e

s

t (

g

/

L)

100

1000 Fish NOEC

Fish Unbounded NOEC

Fish chronic

less sensitive

Estimated

Daphnia magna 48-h LC50 (g/L) 21

0.1 1 10 100

N

O

E

C

f

r

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m

Fis

h

C

h

r

o

nic

T

e

s

t (

0.1

1

10

Fish chronic

more sensitive

Estimated

Acute HC5

(90th %tile)
Saltwater and Freshwater ACRs

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Acute-Chronic Ratio (LC50/GM)

0.1 0.3 1 3 10 30 100 300 1000 3000 10000

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50

SW Invertebrates

FW Invertebrates

SW Fish

FW Fish
Invert ACRs by Species

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98

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Acute Chronic Ratio (LC50/EC20)

0.1 0.2 0.5 1 2 5 10 20 50 100 200 500 1000

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50

Americamysis bahia

Daphnia magna

Ceriodaphnia dubia
Paired Comparison of Mysid/magna

Chemical Mysid ACR D. magna ACR Mysid/magna

Log 

Mysid/magna

Chlorpyrifos (Dursban) 1.75 3.84 0.456 -0.341

Diazinon 2.64 4.97 0.532 -0.274

Ethoprop 6.13 1.56 3.941 0.596

Fonofos 7.33 2.75 2.664 0.426

Methidathion 26.71 9.19 2.907 0.463

Methomyl 8.84 3.67 2.411 0.382

24

Methomyl 8.84 3.67 2.411 0.382

Naled 6.73 2.38 2.832 0.452

Profenofos 8.42 6.45 1.306 0.116

mean = 0.228

SD = 0.357

2-tailed t p=0.05 df=7 2.365 t = 1.803

Not significantly different under 2-tailed test at p=0.05
Selecting an Invertebrate ACR

• Uncertain whether mysid ACRs are larger 

than daphnid ACRs

• Even if they are, is this a FW v SW difference 

or a species difference?

• Is it likely that there are other FW invertebrates 

that would have mysid-like ACRs?

• Under 1985 guidelines, final ACR for AChE 

inhibitor would average daphnid and mysid 

ACRs

• Seems reasonable to select AChE inhibitor 

ACRHC5 from combined invert distribution 25
Invertebrate ACRs by Endpoint

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ACRNOEC

ACRGM

ACREC20

ACR 26

0.1 0.2 0.5 1 2 5 10 20 50 100 200 500

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50
Comparison of ACRs

ACR Distribution Source/Type

Percentile

50th 80th 90th

Present Analysis:

AChE inhibitors, all species ACRGM 13.9 61.1 124

AChE inhibitors, fish only ACRGM 29.2 79.0 247

AChE inhibitors, invertebrates only ACRNOEC 7.98 17.9 30.6

AChE inhibitors, invertebrates only ACRGM 5.91 13.6 18.8

AChE inhibitors, invertebrates only ACREC20 3.88 8.63 18.0

27
Comparison to “Generalized” ACRs

• Raimondo et al. (2007)

• Evaluated distributions of ACRs in light of several 

variables, including MOA

• Comparisons were univariate

• Host et al (1995)

• Assessed distribution of final ACRs (ACRHC5) from 

WQC documents (80th percentile used for GLI)

• TenBrook et al. (2009)

• Subset of final ACRs selecting only chemicals 

used as pesticides (multiple MOA)

28
Comparison of ACRs

ACR Distribution Source/Type

Percentile

50th 80th 90th

Present Analysis:

AChE inhibitors, all species ACRGM 13.9 61.1 124

AChE inhibitors, fish only ACRGM 29.2 79.0 247

AChE inhibitors, invertebrates only ACRNOEC 7.98 17.9 30.6

AChE inhibitors, invertebrates only ACRGM 5.91 13.6 18.8

AChE inhibitors, invertebrates only ACREC20 3.88 8.63 18.0

29

Raimondo et al. (2007):

Combined chemicals, all species ACRGM 8.3 NR 79.5

AChE inhibitors, all species ACRGM 6.4 NR 60.2
Comparison of ACRs

ACR Distribution Source/Type

Percentile

50th 80th 90th

Present Analysis:

AChE inhibitors, all species ACRGM 13.9 61.1 124

AChE inhibitors, fish only ACRGM 29.2 79.0 247

AChE inhibitors, invertebrates only ACRNOEC 7.98 17.9 30.6

AChE inhibitors, invertebrates only ACRGM 5.91 13.6 18.8

AChE inhibitors, invertebrates only ACREC20 3.88 8.63 18.0

30

Raimondo et al. (2007):

Combined chemicals, all species ACRGM 8.3 NR 79.5

AChE inhibitors, all species ACRGM 6.4 NR 60.2
Comparison of ACRs

ACR Distribution Source/Type

Percentile

50th 80th 90th

Present Analysis:

AChE inhibitors, all species ACRGM 13.9 61.1 124

AChE inhibitors, fish only ACRGM 29.2 79.0 247

AChE inhibitors, invertebrates only ACRNOEC 7.98 17.9 30.6

AChE inhibitors, invertebrates only ACRGM 5.91 13.6 18.8

AChE inhibitors, invertebrates only ACREC20 3.88 8.63 18.0

31

Raimondo et al. (2007):

Combined chemicals, all species ACRGM 8.3 NR 79.5

AChE inhibitors, all species ACRGM 6.4 NR 60.2

Host et al. (1995):

Combined chemicals, freshwater FACR 7.1 17.9 29.1

Combined chemicals, saltwater FACR 4.4 13.5 24.2

TenBrook et al. (2010)

Combined chemicals, freshwater FACR NR 12.4 NR
Sources of Variability in ACRs

• ACR distributions contain variability from 

multiple sources:

• True differences among species

• True differences among chemicals

• Random variation in LC50 and chronic value 

(variation multiplicative in ratio)

32
T

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80

100

120

Survival Normalized Reproduction

Daphnia magna tests measuring total young or young/female

Chronic

LC50

Exposure Relative to Chronic LC50

0.003 0.01 0.03 0.1 0.3 1 3 10

T

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0

20

40

60

Test 1

Test 2

Test 3

Test 4

Test 5

Test 6

EC50 = 1.02

EC20 = 0.77

EC0 = 0.48

33
Y

o

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n

g

P

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R

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p

r

o

D

a

y

a

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80

100

120

Survival Normalized Reproduction

Daphnia magna tests measuring young per reproductive day

Chronic

LC50

Exposure Relative to Chronic LC50

Y

0.003 0.01 0.03 0.1 0.3 1 3 10

o

u

n

g

P

e

r

R

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o

D

a

y

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0

20

40

60

Test 1

Test 2

Test 3

Test 4

Test 5

Test 6

Test 7

EC50 = 1.24

EC20 = 0.73

EC0 = 0.29

34
Effect of Random Variation

• Monte Carlo simulation

ACR Distribution Source/Type

Percentile

50th 80th 90th

Present Analysis:

AChE inhibitors, invertebrates only ACRGM 5.91 13.6 18.8

• Mean acute value = 5.91, mean chronic value = 1

• Appropriate variability uncertain; for example 

purposes, used “factor of 2” rule of thumb

• Log-normal distributions with mean ± 2 SD equal 

to 0.5x and 2x of true value

35
Monte Carlo Results

ACR Distribution Source/Type

Percentile

50th 80th 90th

Present Analysis:

AChE inhibitors, invertebrates only ACRGM 5.91 13.6 18.8

Monte Carlo Simulation, true mean 5.91

Log-normal, mean ± 2 SD = 0.5x to 2x 5.91 11.0 15.6

36

• Observed variability in ACRs for AChE inhibitors 

comparable to variability expected if assumed 2x 

variability is realistic

• May affect risk management decisions regarding 

percentile selection if upper tails of distribution result 

primarily from random variability rather than true 

differences in response
Conclusions

• For AChE inhibitors, ACR distribution for fish 

higher than for invertebrates

• Despite this, an acute HC5 protective of 

invertebrates would also protect fish from 

chronic effects

• Acute HC5 and chronic HC5 driven by 

invertebrate data, so ACRHC5 should be 

selected from invertebrate ACRs

• For AChE inhibitors, MOA-specific ACRs up 

to 2-fold lower than generalized ACRs

37
Questions?

38