The importance of absorption, elimination and feeding pattern: using - PowerPoint PPT Presentation

The importance of absorption, elimination and feeding pattern: using toxicokinetics modelling to refine the risk assessment of pesticides to wildlife Agnieszka Bednarska (1) , Peter Edwards (1) , Richard Sibly (2) , Pernille Thorbek (1) (1) Syngenta, Jealott’s Hill International Research Centre, Bracknell (2) School of Biological Sciences, University of Reading, Reading, UK University of Reading

Content ● The current risk assessment for birds and mammals ● TK model: what, why and how ? ● What do regulators say about importance of feeding patterns and avoidance? ● Case study: pros and cons of body burden model 2

Background ● The current risk assessment for birds and mammals* • Based on external exposure measurements • Toxicity endpoints calculated from the gavage dose (acute) or the dietary toxicity (chronic) do not represent field exposure • No account taken of the physiological processes (e.g., absorption, elimination), ecological factors (e.g., feeding rate, avoidance), duration of exposure ● TK model • Absorption, Distribution, Metabolism, Excretion (ADME) * EFSA Journal 2009, 7(12), 1438 3

TK models in EFSA Guidance 6.3. Metabolism & avoidance – application of body-burden models and dietary toxicity data “ Within the registration process of PPP under Directive 91/414/ECC, often data from metabolism studies (ADME) within rat, live-stock or hen are available . ” “ Where risk-refinement is necessary based on results from lower tier assessment, ‘metabolism’ data should be evaluated by the risk assessor for options to reduce the uncertainty associated with the risk assessment. ” EFSA Journal 2009, 7(12), 1438 4

Case study: metabolism data for an insecticide Data from rats metabolism study with radiolabelled insecticide k a =0.36 h -1 k a =0.30 h -1 k e =0.37 h -1 k e =0.30 h -1 Compartmental analysis of data on Concentration [µg ml -1 ] insecticide concentrations in blood One-compartment model fits data best k a =2.56 h -1 k a =1.01 h -1 k e =0.27 h -1 k e =0.36 h -1 Insecticide concentrations in blood highly correlated with concentrations in different tissues Up to 90% eliminated as parent k a =3.45 h -1 k a =2.21 h -1 compound through the urine k e =0.13 h -1 k e =0.30 h -1 Time [h] FIG. The concentration of an insecticide in blood of three male rats administered 0.5 mg kg -1 bw [Thiazol-2- 14 C] (left-hand column) or 0.5 mg kg -1 bw [Oxadiazin-4- 14 C] (right-hand column). Lines are the one-compartment models fit to the experimental blood data. Note different scale on y axis. WinNonlin software; comparison between different models based on residual plots and AIC. 5 Model parameters estimated using Marquardt method.

Case study: TK model I k out gut a.i. in food     k a C I k C k C gut out gut a gut    central C k C F k C compartment int a gut e int (bloodstream) k e Δ C change in the gut gut or internal int concentration of pesticide in given time interval, here one min. intake rate [mg a.i. kg -1 bw min -1 ] I F bioavailability, here F =1 k out the rate of excretion of toxicant not absorbed into the system from the gut [min -1 ], here k out =0 the rate of toxicant absorption from the gut into the system [min -1 ] k a the rate of toxicant elimination from the system [min -1 ] k e 6

Case study: using metabolism data for parameterization of TK model Kinetics parameters estimated from rat study based on radiolabeled test substance intravenous bolus gavage exposure exposure Parameters [Thiazol-2- 14 C] [Thiazol-2- 14 C] [Oxadiazin-4- 14 C] [Thiazol-2- 14 C] [Oxadiazin-4- 14 C] 0.5 mg kg -1 bw 0.5 mg kg -1 bw 0.5 mg kg -1 bw 100 mg kg -1 bw 100 mg kg -1 bw male female male female male female male female male female 2.2 1.3 k a (h -1 ) 2.1 ± 1.62 2.3 ± 1.93 1.20 ± 0.93 3.2 ± 0.29 0.78 ± 0.32 1.6 ± 0.71 0.71 ± 0.73 2.00 ± 1.37 - - 0.40 0.25 k e (h -1 ) 0.26 ± 0.02 0.50 ± 0.11 0.23 ± 0.09 0.23 ± 0.13 0.34 ± 0.034 0.25 ± 0.06 0.28 ± 0.12 0.18 ± 0.06 0.25 ± 0.11 0.19 ± 0.06 AUC ( h ug -1 ml -1 ) 2.30 ± 0.19 1.63 ± 0.38 1.49 ± 0.15 1.56 ± 0.53 1.30 ± 0.015 1.03 ± 0.13 342 ± 80 278 ± 59 359 ± 50 294 ± 24 0.65 ± 0.07 0.96 ± 0.33 0.56 ± 0.007 0.63 ± 0.08 0.74 ± 0.17 0.85 ± 0.18 0.78 ± 0.11 0.90 ± 0.07 F - - 0.5 mg a.i. kg -1 bw 100 mg a.i. kg -1 bw Concentration [mg/kg] Concentration [mg/kg] measured k a = 2.2 h -1 k e = 0.25 h -1 k a = 1.3 h -1 k e = 0.40 h -1 Time [min] Time [min] FIG. The concentration of an insecticide in blood of rats (points) after administration of 0.5 or 100 mg a.i. kg -1 bw and model simulations (lines) for two extreme combinations of k a and k e . 7

Feeding pattern in EFSA Guidance 6.2. Avoidance “ What rates of feeding occur in the field? “ “ Do the feeding rates achieved in laboratory studies or assumed in model correspond to the maximum rates occurring in the field? “ EFSA Journal 2009, 7(12), 1438 8

Case study: different exposure simulations Intake rate for rat of uncontaminated food [g diet kg -1 bw min -1 ] at 15-min time intervals over 2h Intake rate Intake rate of LD 50 Time from Control food [g diet kg -1 bw] [mg a.i. kg -1 bw min -1 ] [g kg -1 bw min -1 ] start of feeding (min) C7 C15 C21 C17 ..... mean mean mean 0 0 0 0 0 0.0 0.00 0.0 15 10.9 7.8 6.3 5.6 9.2 0.61 35.3 30 14.1 11.3 6.3 9.4 14.2 0.33 19.1 45 20.3 16.9 6.8 13.8 17.5 0.22 12.7 60 23.5 24.5 10.3 16.4 20.5 0.20 11.6 75 26.7 28.1 15.5 18.9 23.6 0.21 12.2 90 28.8 28.1 19.8 19.7 25.1 0.10 5.8 105 28.8 31.9 20.7 19.7 26.8 0.11 6.4 120 28.8 31.9 20.7 20.6 27.1 0.02 1.2 LD 50 eaten with constant 13.0 intake rate over 120 min.: Concentration [mg kg -1 bw] bolus gavage exposure maximum feeding rate in control over 2 h constant ingestion rate over 2h The concentration of an insecticide in the body after eating LD 50 dose according to different intake rates; scenarios for k a = 2.2 and k e = 0.25. Time [min] 9

Case study: LD 50 eaten with different feeding patterns Continuous feeding 4h Continuous feeding 2h Concentration [mg kg -1 bw] ΔC int ΔC gut C max bolus gavage cumulative a.i. intake Time [min] Time [min] 1h feeding +2h break +1h feeding 1h feeding +4h break +1h feeding Concentration [mg kg -1 bw] FIG. The concentration of an insecticide in the body, in the gut, and cumulative intake over time after exposure to LD 50 dose according to Time [min] Time [min] different feeding pattern. The slower animals eat the lower internal maximum concentrations are reached. Feeding pattern influences internal concentration of pesticide, especially if there are breaks of low/no feeding activity after short feeding bouts. 10

Avoidance in EFSA Guidance 6.2. Avoidance “ A degree of avoidance of food contaminated with pesticides , commonly seen in dietary studies with captive animals, has the potential to reduce exposure and hence risk in the field . It can be combination of several different responses including (a) a reduction in the rate of feeding due to novel or unpleasant characteristics of the contaminated food, and (b) temporary cessation of feeding due to sublethal intoxication. It is hard to determine the precise mechanism(s) of avoidance for a given pesticide. “ EFSA Journal 2009, 7(12), 1438 11

Avoidance To be useful for TK modelling the strength of avoidance must be determined quantitatively I k out ‘... (b) temporary cessation of feeding due to a.i. in food gut sublethal intoxication ...’ k a Avoidance experiment on rats AVOIDANCE central determine the highest dose (mg kg -1 bw) and Aim: : compartment intake rate [mg a.i. kg -1 bw min -1 ] of an insecticide (bloodstream) Methods: acclimatisation to maximise the feeding k e 2h access to contaminated food without access to alternative food, then 10h access to untreated food video-recording of intake of food (a balance reading) during 1 st day of exposure exposure for a further 2 days with a 2h deprivation period, but without video recording direct measurement of eaten diet 12

Avoidance: results from study on rats Food consumption at Day -2 Food consumption at Day 0 Food consumption [g kg -1 bw] Food consumption [g kg -1 bw] 50 50 45 45 40 40 35 35 30 30 25 25 20 20 15 15 10 10 5 5 0 0 0 3126 6252 15630 31260 46900 96300 0 3126 6252 15630 31260 46900 96300 Treatment [mg a.i. kg -1 food] Treatment [mg a.i. kg -1 food] Treatment Treatment FIG. Food consumption for rats before exposure (Day-2) and on the first day of exposure (Day 0) at different treatments. Intake dose [mg a.i. kg -1 bw] Results of Multifactor ANOVA: Treatment p<0.0001 Day p<0.0001 Day*Treatment p< 0.001 FIG. Potential intake dose (mean based on food consumption at Day-2 and measured intake dose (mean) at different treatment. Treatment [mg a.i. kg -1 food] Clear evidence of avoidance in terms of eaten dose but not stable over time 13