30 jan 2006 page 1 focus kinetics training workshop chapter 7 recommended procedures to derive...

30 Jan 2006

FOCUS Kinetics training workshop

Chapter 7

Recommended Procedures to Derive Endpoints for Parent Compounds

Ralph L. Warren, Ph.D.DuPont Crop Protection

Delaware, USA

Presentation


Objectives of this part of the training:

• Description of the procedures to follow for a parent compound to derive kinetic fitting endpoints

a) “best fit” values (compared to triggers for additional work in EU)

b) inputs for environmental exposure models

• Assessment of kinetic model fits to the observed data using visual and statistical techniques.

• Selection of the appropriate kinetic model and endpoints for the case of triggers and exposure modeling in the EU.

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Hands-on exercise


Why the distinction between fitting for trigger endpoints versus exposure modeling endpoints?

• Current EU regulatory environmental exposure models are based on SFO kinetics. Therefore, an endpoint (i.e. DT50) calculated using a non-SFO kinetic model will not appropriately represent the observed behavior when input into a SFO-based exposure model. A SFO endpoint, if appropriate, or a conservative estimate or a ‘work around’ must be used.

• EU regulatory triggers are based on DT50 and DT90 values which are not constrained to any kinetic model form. The model that most appropriately describes the observed data should be used to generate the endpoint values.

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The same DT50 does not mean the same pattern of decline when calculated using different kinetic models

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time (days)

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SFO

FOMCDFOS

DFOP

M0 = 100% and DT50 = 5 days in each case

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EU regulatory trigger examples

Annex II to Directive 91/414/EEC

• 7.1.1.2.2. Field dissipation studies are required when DT50lab > 60 days at 20C or 90 days at 10 C

Annex III to Directive 91/414/EEC

• 10.7.1 Testing for effects on soil micro-organisms required when DT90field > 100 days

Draft Guidance Doc. Terrestrial Ecotoxicology (SANCO/10329/2002 rev. 2 final)

• Sub-lethal earthworm tests required depending on number of applications and DT90field

Guidance Doc. Aquatic Ecotoxicology (SANCO/3268/2001 rev. 4 final)

• Chronic study on daphnids required when DT50 in water > 2 days

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Modeling endpointsTriggers for additional work Modeling endpointsTriggers for additional work


So what’s involved in the fitting procedure?

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• Run SFO and FOMC as a first step

• Check visual fit and calculate error percentage at which 2 test passed

• Check parameter uncertainty

• If FOMC better than SFO, test other bi-phasic models

• Use best fit model

• Run SFO as a first step

• Check visual fit and calculate error percentage at which 2 test passed

• Check parameter uncertainty

• If error % < 15% and visual fit acceptable, use SFO DT50

• If error % > 15% and visual fit not acceptable, run bi-phasic model

• If 10% of initial reached in study period then calculate DT50 as FOMC DT90/3.32

• If 10% of initial not reached in study period then use longer DT50 from slow phase of HS or DFOP


Chi-square (2) test statistic – test of association

whereC = calculated valueO = observed value

= mean of observed (element of scale)err = measurement error (element of proportionality)O

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2

22

)O x 100/err(

)OC(

If calculated 2 > tabulated 2 then the model is not appropriate at the chosen level of significance (5%)

Error percentage unknown Calculate error level at which 2 test is passed

2

2

2tabulated O

OC

χ

1100err


Visual Assessment

• Subjective, yet powerful tool for assessing goodness of fit.

• Keeps common sense in the assessment process.

• Two recommended plots

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t (days)

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t (days)

resi

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Observed and predicted through time

Residuals(predicted - observed)


Parameter uncertainty

• Confidence intervals or t-tests may be used.

• The t-test is shown below, which assumes normally distributed parameters.

i

iatˆ

iai

where = estimate of parameter i = standard error of parameter i

• The probability (p-value) for the calculated t-value can be read from statistical tables or calculated with Excel TDIST(tcaclulated,df,1)

• If P is < 0.05 then the parameter is considered significantly different than zero. If P is between 0.05 and 0.1 then weight of evidence should be considered.

• The t-test is most applicable to degradation rates (k), not necessarily other parameters such as or for FOMC.

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NO

YES see text

YES

RUN SFO, FOMC

Data entry M0 free, use all data, no weighting

SFO more appropriate than FOMC and gives

acceptable fit?

RUN DFOP (unmodified &

modified fitting routine)

Does the best-fit model give an acceptable description

of the data?

STEP 1: SFO appropriate?

STEP 2: Identify best model other than SFO

Deviation from SFO due to experimental

artifact/decline in microbial activity?

NO

Case-by-case decision (see text)

Determine which of the models (FOMC, DFOP)

is best

NO

YES STOP

STEP 3: Evaluate goodness of fit

NO

Modify fitting routine stepwise: 1. Exclude outliers 2. Constrain M0 3. Weighting

RUN modified fitting

SFO more appropriate than FOMC & fit acceptable?

(modified fitting)

YES STOP

STOP

Parent only flow chart for deriving trigger endpoints

(zoom to view)

Triggers flowchart

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NO YES

RUN SFO

Data entry M0 free, use all data, no weighting

SFO statistically and visually acceptable? Modify fitting routine for

SFO stepwise: 1. Exclude outliers 2. Constrain M0 3. Weighting until best SFO fit achieved

STEP 1: SFO appropriate?

RUN modified SFO

Use SFO DT50 for fate modelling

Aim: modelling fate of parent only?

YES

YES 10% initially measured concentration reached

within experimental period?

NO RUN FOMC

RUN HS or DFOP

Use DT50 from slow phase of HS of DFOP

model for fate modelling


NO

HS or DFOP statistically and

visually acceptable?

YES

FOMC statistically and visually acceptable?

YES

Back-calculate DT50 from DT90 for FOMC (DT50 = DT90 / 3.32)


NO

YES

Use SFO DT50 (modified fitting routines) for fate modelling

NO

Bi-phasic pattern? (assess experimental

artefacts!)

SFO statistically and visually acceptable?

YES


NO

STEP 2:Correction procedure

Aim: modelling metabolite fate linked to

parent?

see text

YES

YES

Parent only flow chartfor deriving exposure modeling endpoints

(zoom to view)

Modeling flowchart

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Let’s look at an example for the triggers flowchart…

Time (days)

% of applied radioactivity

003377

14143030454562629090

120120

93.199.772.983.860.360.341.737.423.326.020.917.118.818.817.918.516.715.9

Laboratory degradation of a compound in aerobic soil

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2 error (%) = 19.0DT50 (d) = 18.1DT90 (d) = 60.1

2 error (%) = 6.69DT50(d) = 10.6DT90 (d) = 158

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Measured & Predicted vs. Time

Parent

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Residual Plot

Parent

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Parent

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Residual Plot

Parent

SFO FOMC


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2 error (%) = 6.69DT50(d) = 10.6DT90 (d) = 158

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Parent

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Residual Plot

Parent

FOMC

2 error (%) = 1.36DT50(d) = 10.6 dDT90 (d) = 481 d

DFOP

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Parent

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Residual Plot

Parent



Model Parameter Optimized value

Standard error

Different than zero?

(P<0.05)

SFO M0 (%)k (d-1)

87.01630.0383

5.39280.0060

--Yes

FOMC M0 (%)

98.17690.71006.4023

3.02890.09711.8756

------

DFOP M0 (%)g

k1 (d-1)k2 (d-1)

96.74970.79240.09310.0015

1.76890.03270.00850.0020

----

YesNo (P=0.225)

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-- = not applicable


• Use DFOP with associated endpoints > DT50 = 10.6 d, DT90 = 481 d > Relax t-test criteria for k2 based on visual fit and 2. > Check if other aerobic soil deg and fate studies support this DT90.

• Use DFOP. Fix k2 to a conservative value (e.g. 1000 d) > 2 and visual fits equivalent to above. > DT50 = 10.7 d, DT90 = 962 d > Check if other aerobic soil deg and fate studies support this DT90.

Possible conclusions for this data set for the triggers flowchart

• For comparison with EU regulatory DT50 triggers, the result is the same.

• For comparison with EU regulatory DT90 triggers, the result is the same.

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Continuing with the same data, now let’s look at it using the modeling flowchart…

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2 error (%) = 19.0DT50 (d) = 18.1DT90 (d) = 60.1

2 error (%) = 6.69DT50(d) = 10.6DT90 (d) = 158

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Parent

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Parent

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SFO FOMC


• Assuming no artifacts, the data is clearly bi-phasic. FOMC fit to the data is superior based on visual assessments and 2 error.

• If aim of modeling is to link parent with metabolites, then the guidance in Chapter 8 should be followed (covered later).

• If the aim is to model parent fate only then check to see if 10% of the initially measured value was reached during the study period.

> If yes, then use FOMC DT90/3.32 to derive a conservative estimate of SFO DT50 for modeling (i.e. 158 d/3.32 = 47.6 d).

> If no, then use slower k from DFOS (HS) or slower k from DFOP to derive a conservative estimate of DT50 for modeling.

We did not reach 10% of initial in this example so further analysis is required.

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SFO DT50 = 18.1 d DT90 = 60.1 d

FOMC DT50 = 10.6 dDT90 = 158 d

FOMC DT90/3.32 = 47.6 d (SFO)

FOMC DT90/3.32 is a conservative option where parent only exposure modeling is desired (can’t link to metabolites!)

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FOMC DT90/3.32

FOMC DT90/3.32 example (assume last point did reach 10%)

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2 error (%) = 2.59DT50(d) = 10.7DT90 (d) = 244

DFOS (HS)

2 error (%) = 1.36DT50(d) = 10.6 dDT90 (d) = 481 d

DFOP

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Parent

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Model Parameter Optimized value

Standard error

Different than zero?(t-test)

DFOP M0 (%)g

k1 (d-1)k2 (d-1)

96.74970.79240.09310.0015

1.76890.03270.00850.0020

----

YesNo (P=0.225)

DFOS (HS) M0 (%)tb (d)

k1 (d-1)

k2 (d-1)

95.811921.91500.06460.0040

1.82591.63650.00380.0016

----

YesYes

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-- = not applicable


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DFOPfast phase, k1, DT50 = ln(2)/0.0931 = 7.45 d slow phase, k2, DT50 = ln(2)/0.0015 = 462 d

DFOS (HS)fast phase, k1, DT50 = ln(2)/0.0646 = 10.7 d slow phase, k2, DT50 = ln(2)/0.0040 = 173 d


• Use longest phase of DFOS (HS) to derive conservative value of DT50

> 10% of initial not reached, so DFOS (HS) and DFOP were assessed. > Longest k from DFOP is not different than zero so it is unreliable.

Possible conclusions for this data set for the modeling flowchart

• Conduct higher-tier modeling using conservative value for DFOP slow phase DT50 (e.g. 1000 d).

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Summary

• Standardized procedures (flow charts) can be readily followed for deriving parent only endpoints

• Two flow charts are provided, one for determination of “best fit” kinetic parameters, the other for deriving inputs for use with SFO environmental exposure models

• Statistical and visual methods described provide a consistent way to assess kinetic model fits

• There is still room for judgment and discussion in the fitting and endpoint selection process, but the procedures described here should lead to greater consistency and transparency

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Questions?

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Now it’s your turn to work through the flowcharts using some other real data sets…

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If you finish the exercise and have additional time, you might try duplicating the fitting (SFO, FOMC, DFOS, DFOP) of the example data given in this presentation.

30 jan 2006 page 1 focus kinetics training workshop chapter 7 recommended procedures to derive...

Documents

sfo dt

sfo kinetics

fomc dt

longer dt

usa slide

dfop slide

sfofomc slide

visual fit