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PBPK MODELS

An overview of research conducted at the Chemical Industry Institute of Toxicology. Most of the results have been published in the peer review literature beginning in 1985. Early research was largely devoted to characterizing the prenatal toxicity of 2-methoxyethanol (2-ME) and its potential mode of action. In later stages, PBPK models were developed to to improve the scientific foundation for extrapolation of data obtained in pregnanat laboratory animals to pregnant women in occupational settings. PBPK simulations indicate that at the 5 ppm TLV exposure level (8 hr/day, 5 days/wk, with weekend breaks) maternal blood concentrations of 2-MAA would be in the range of ~60 uM (human) and 70 – 90 uM (mice and rats). The concentrations causing developmental toxicity in rodents are much higher (in the low mM range in both mice and rats) than those ever anticipated to be in the human occupational setting.

 

PBPK models for 2-methoxyethanol (2-ME) in pregnant rats and humans were used to calculate estimated human-equivalent no adverse effect levels based on internal dosimetry at the NOEL for developmental toxicity in rats (10 ppm). Uncertainties in the modeled estimates were estimated from the distribution of internal dose estimates obtained by varying key model parameter values (identified by sensitivity analysis) over expected ranges and probability distributions. Distributions of the values of these parameters were sampled using techniques and appropriate dose metrics calculated for 1600 parameter sets. The 95thpercentile values were used to calculate interindividual pharmacokinetic uncertainty factors to account for variability among humans. The calculated interindividual pharmacokinetic uncertainty factor for 2-ME was 1.7, which is less than the default value of 3.0 commonly used for this area of uncertainty.  The estimated human equivalent no adverse effect level derived using average values for physiological, thermodynamic and metabolic parameters in the PBPK model was 12 ppm (based on a NOAEL in rats of 10ppm). This value was divided by the calculated interindividual pharmacokinetic uncertainty factor of 1.7 and default uncertainty factors of 2.5 for interspecies pharmacodynamic variability, and 3.16 for intraspecies pharmacodynamic variability, to calculate proposed occupational exposure limits. The author concluded that this methodology indicates that an occupational exposure limit (8-hr time-weighted average) of 0.9 ppm (3 mg/m3 - based on an animal NOAEL of 10ppm) should protect workers from the most sensitive adverse effects of 2-ME, assuming that no or minimal dermal exposure occurs. Short-term exposure scenarios (15 min) to elevated concentrations were also simulated. Once daily 15 min expsosures (inhalation only) to 29 ppm 2-ME produced the same average daily blood AUC levels of 2-MAA as the 8-hr TWA exposure to 0.9 ppm.

 

In a study to expand previous work on a PBPK model for 2 -methoxyethanol (2 -ME), pregnant rats were exposed for 5 days (gestation days 11-15), 6 h/day, to 2-ME vapor at 10 and 50 ppm. Validation consisted of comparing model output to maternal blood and fetal 2-ME and 2-MAA concentrations during and following 5 days of exposure (gestation days 11-15). These concentrations correspond to a known no observed effect level (NOEL) and a lowest observed effect level (LOEL) for developmental effects in rats. The rat PBPK model for 2-ME/2-MAA was scaled to humans and the model (without the pregnancy component) used to predict data collected by other investigators on the kinetics of 2-MAA excretion in urine following exposures to 2-ME in human volunteers. The partially validated human model (with the pregnancy component) was used to predict equivalent human exposure concentrations based on 2-MAA dose measures (maximum blood concentration, C(max), and average daily area under the 2-MAA blood concentration curve, AUC, during pregnancy) that correspond to the concentrations measured at the rat NOEL and LOEL exposure concentrations. The human equivalent exposure concentrations that were estimated in this work were 20% higher than the corresponding NOEL and LOEL values established in the rat. This indicates that pregnant women exposed to inhaled concentrations of 2-ME for 8 hr/day, 5 days/week for the duration of pregnancy do not reach blood 2-MAA concentrations that are known to be developmentally toxic to mice and rats until the exposure concentration reaches 60 ppm or higher (12ppm for the NOAEL in rats).

 

A study was performed to adapt an existing physiologically based pharmacokinetic (PBPK) model for the kinetics of 2 -methoxyethanol (2 -ME) and the proximate toxicant, the metabolite 2-methoxyacetic acid (2-MAA) during midorganogenesis in mice to rats on gestation days (GD) 13 and 15. Blood and tissue data were analyzed using the extrapolated PBPK model that was modified to simulate 2-ME and 2-MAA kinetics in maternal plasma and total embryo tissues in pregnant rats. The original mouse model was simplified by combining the embryos and placenta with the richly perfused tissue compartment. The model includes a description of the growth of the developing embryo and changes in the physiology of the dam during pregnancy. Biotransformation pathways of 2-ME to either ethylene glycol (EG) or to 2-MAA were described as first-order processes and tissue partition coefficients (PCs) were determined for a variety of maternal tissues and the embryos. Model simulations closely reflected the biological measurement of 2-ME and 2-MAA concentrations in blood and embryo tissue following gavage or iv administration of 2-ME or 2-MAA. The authors concluded that the PBPK model for rats was suitable for extrapolation to pregnant women and for assessment of 2-MAA dosimetry under various conditions of possible human exposure to 2-M. They also concluded that since the concentration profiles of 2-MAA in rat embryos and maternal plasma were similar for all doses of 2-ME tested, the maternal plasma concentration of 2-MAA appears to be a reasonable indicator of and surrogate for embryo exposure to 2-MAA.

 

A PBPK model was been developed to describe the disposition of methoxyethanol and its metabolite methoxyacetic acid in pregnant mice, rats and humans. The model was validated by comparison against independently generated data sets. The conclusion of the authors of the work was that the model demonstrates that it is not necessary to use an assessment factor for interspecies toxicokinetic differences when extrapolating animal data to humans.

 

ABSORPTION

 

A pharmacokinetic study using dermal exposure showed that dermal uptake can be significant even from unoccluded exposure sites. The amounts absorbed and the route of excretion was independent of dose. However, the pattern of urinary metabolites varied with dose; methoxyacetic acid predominated at low dose, with increasing amounts of an unknown metabolite being present as dose increased. The pattern of metabolites (reduced CO2 and ethylene glycol, increased level of an unknown) was different from the dermal route of exposure compared to dosing via drinking water.

An in vitro dermal absorption study using human skin showed that methoxyethanol readily passes through the stratum corneum and causes slight but detectable damage to the skin.

 

DISPOSITION

 

An autoradiographic study of methoxyethanol disposition in mice following oral and i.v. dosing showed that widespread disposition throughout the animals. High levels were detected in the liver, bladder, kidney and GI tract. Significant levels were also detected in bone marrow and epididymides, known target organs for the toxicity of the substance. Oral exposure resulted in higher tissue concentrations than the i.v. route.

 

A pharmacokinetic and autoradiography study using oral exposure in pregnant female mice showed that methoxyethanol is very rapidly distributed throughout the body. Peak concentrations of it and/or its metabolites persist for about 2 hours before elimination proceeds but concentrations of the substance and/or metabolite remain relatively high in the liver and conceptus compartment and at levels 2 -4x blood concentrations. Radio labelled material was detectable in sensitive areas of the developing embryo.

 

METABOLISM AND ELIMINATION

 

A toxicokinetic study using radiolabelled 2-methoxyethanol showed that approximately 95% of orally ingested material is excreted within 48 hours and that the main route of metabolism accounting for more than 50% of ingested material is to 2-methoxyacetic acid, which is excreted in urine. A smaller but significant amount is metabolized to CO2 and exhaled in air.

 

A pharmacokinetic study showed coadministration of ethanol by i.p. significantly inhibited the metabolism of methoxyethanol when administered by inhalation,suggesting that alcohol dehydrogenase metabolism is an important pathway for the metabolism of methoxyethanol.

 

A pharmacokinetic study in humans using inhalation exposure showed that the rate of metabolism of methoxyethanol is relatively slow, with a half life of 77 hours being calculated.

 

A study which dosed pregnant mice orally with methoxyethanol with and without other substances such as 4-methyl pyrazole and/or ethanol which can block/saturate the alcohol dehydrogenase enzyme showed that metabolism is essential to produce the typical pattern of developmental toxicity seen and that this enzyme plays a key role in such metabolism.

 

A metabolism study in the pregnant mouse showed that, following oral exposure, methoxyethanol is eliminated primarily in the urine as methoxyacetic acid in pure form or conjugated with glycine. Most product is eliminated within 48hrs. 

 

A metabolism study in the pregnant mouse showed that, following oral exposure, methoxyethanol is eliminated in the urine as primarily methoxyacetic acid in pure form or conjugated with glycine. A number of other direct metabolites, including ethylene glycol, sulphate and other conjugates were detected in urine along with metabolites of methoxyacetic acid, such as the glycine conjugate and derivatives of methoxyacetyl CoA.

 

A pharmacokinetic study using oral exposure in pregnant female mice showed thatmethoxyethanol is predominantly eliminated in the urine. Very little radioactivity was detected in the conceptus.

 

A developmental toxicity study in the monkey with pharmacokinetic observations showed that the mainmetabolite of methoxyethanol (methoxyacetic acid - MAA) and the metabolite that causes developmental toxicity) has a relatively long half life (22hrs) which can lead to accumulation of the toxicant when daily dosing occurs. The metabolite was found to readily crosses the placenta and distribute evenly between embryo and dam in pregnant monkeys. The yolk sac showed signs of accumulating MAA, but the toxicological significance of this is unknown.

Discussion on bioaccumulation potential result:

PBPK MODELS

An overview of research conducted at the Chemical Industry Institute of Toxicology. Most of the results have been published in the peer review literature beginning in 1985. Early research was largely devoted to characterizing the prenatal toxicity of 2-methoxyethanol (2-ME) and its potential mode of action. In later stages, PBPK models were developed to to improve the scientific foundation for extrapolation of data obtained in pregnanat laboratory animals to pregnant women in occupational settings. PBPK simulations indicate that at the 5 ppm TLV exposure level (8 hr/day, 5 days/wk, with weekend breaks) maternal blood concentrations of 2-MAA would be in the range of ~60 uM (human) and 70 – 90 uM (mice and rats). The concentrations causing developmental toxicity in rodents are much higher (in the low mM range in both mice and rats) than those ever anticipated to be in the human occupational setting.

 

PBPK models for 2-methoxyethanol (2-ME) in pregnant rats and humans were used to calculate estimated human-equivalent no adverse effect levels based on internal dosimetry at the NOEL for developmental toxicity in rats (10 ppm). Uncertainties in the modeled estimates were estimated from the distribution of internal dose estimates obtained by varying key model parameter values (identified by sensitivity analysis) over expected ranges and probability distributions. Distributions of the values of these parameters were sampled using techniques and appropriate dose metrics calculated for 1600 parameter sets. The 95thpercentile values were used to calculate interindividual pharmacokinetic uncertainty factors to account for variability among humans. The calculated interindividual pharmacokinetic uncertainty factor for 2-ME was 1.7, which is less than the default value of 3.0 commonly used for this area of uncertainty.  The estimated human equivalent no adverse effect level derived using average values for physiological, thermodynamic and metabolic parameters in the PBPK model was 12 ppm (based on a NOAEL in rats of 10ppm). This value was divided by the calculated interindividual pharmacokinetic uncertainty factor of 1.7 and default uncertainty factors of 2.5 for interspecies pharmacodynamic variability, and 3.16 for intraspecies pharmacodynamic variability, to calculate proposed occupational exposure limits. The author concluded that this methodology indicates that an occupational exposure limit (8-hr time-weighted average) of 0.9 ppm (3 mg/m3 - based on an animal NOAEL of 10ppm) should protect workers from the most sensitive adverse effects of 2-ME, assuming that no or minimal dermal exposure occurs. Short-term exposure scenarios (15 min) to elevated concentrations were also simulated. Once daily 15 min expsosures (inhalation only) to 29 ppm 2-ME produced the same average daily blood AUC levels of 2-MAA as the 8-hr TWA exposure to 0.9 ppm.

 

In a study to expand previous work on a PBPK model for 2 -methoxyethanol (2 -ME), pregnant rats were exposed for 5 days (gestation days 11-15), 6 h/day, to 2-ME vapor at 10 and 50 ppm. Validation consisted of comparing model output to maternal blood and fetal 2-ME and 2-MAA concentrations during and following 5 days of exposure (gestation days 11-15). These concentrations correspond to a known no observed effect level (NOEL) and a lowest observed effect level (LOEL) for developmental effects in rats. The rat PBPK model for 2-ME/2-MAA was scaled to humans and the model (without the pregnancy component) used to predict data collected by other investigators on the kinetics of 2-MAA excretion in urine following exposures to 2-ME in human volunteers. The partially validated human model (with the pregnancy component) was used to predict equivalent human exposure concentrations based on 2-MAA dose measures (maximum blood concentration, C(max), and average daily area under the 2-MAA blood concentration curve, AUC, during pregnancy) that correspond to the concentrations measured at the rat NOEL and LOEL exposure concentrations. The human equivalent exposure concentrations that were estimated in this work were 20% higher than the corresponding NOEL and LOEL values established in the rat. This indicates that pregnant women exposed to inhaled concentrations of 2-ME for 8 hr/day, 5 days/week for the duration of pregnancy do not reach blood 2-MAA concentrations that are known to be developmentally toxic to mice and rats until the exposure concentration reaches 60 ppm or higher (12ppm for the NOAEL in rats).

 

A study was performed to adapt an existing physiologically based pharmacokinetic (PBPK) model for the kinetics of 2 -methoxyethanol (2 -ME) and the proximate toxicant, the metabolite 2-methoxyacetic acid (2-MAA) during midorganogenesis in mice to rats on gestation days (GD) 13 and 15. Blood and tissue data were analyzed using the extrapolated PBPK model that was modified to simulate 2-ME and 2-MAA kinetics in maternal plasma and total embryo tissues in pregnant rats. The original mouse model was simplified by combining the embryos and placenta with the richly perfused tissue compartment. The model includes a description of the growth of the developing embryo and changes in the physiology of the dam during pregnancy. Biotransformation pathways of 2-ME to either ethylene glycol (EG) or to 2-MAA were described as first-order processes and tissue partition coefficients (PCs) were determined for a variety of maternal tissues and the embryos. Model simulations closely reflected the biological measurement of 2-ME and 2-MAA concentrations in blood and embryo tissue following gavage or iv administration of 2-ME or 2-MAA. The authors concluded that the PBPK model for rats was suitable for extrapolation to pregnant women and for assessment of 2-MAA dosimetry under various conditions of possible human exposure to 2-M. They also concluded that since the concentration profiles of 2-MAA in rat embryos and maternal plasma were similar for all doses of 2-ME tested, the maternal plasma concentration of 2-MAA appears to be a reasonable indicator of and surrogate for embryo exposure to 2-MAA.

 

A PBPK model was been developed to describe the disposition of methoxyethanol and its metabolite methoxyacetic acid in pregnant mice, rats and humans. The model was validated by comparison against independently generated data sets. The conclusion of the authors of the work was that the model demonstrates that it is not necessary to use an assessment factor for interspecies toxicokinetic differences when extrapolating animal data to humans.

 

ABSORPTION

 

A pharmacokinetic study using dermal exposure showed that dermal uptake can be significant even from unoccluded exposure sites. The amounts absorbed and the route of excretion was independent of dose. However, the pattern of urinary metabolites varied with dose; methoxyacetic acid predominated at low dose, with increasing amounts of an unknown metabolite being present as dose increased. The pattern of metabolites (reduced CO2 and ethylene glycol, increased level of an unknown) was different from the dermal route of exposure compared to dosing via drinking water.

 

DISPOSITION

 

An autoradiographic study of methoxyethanol disposition in mice following oral and i.v. dosing showed that widespread disposition throughout the animals. High levels were detected in the liver, bladder, kidney and GI tract. Significant levels were also detected in bone marrow and epididymides, known target organs for the toxicity of the substance. Oral exposure resulted in higher tissue concentrations than the i.v. route.

 

A pharmacokinetic and autoradiography study using oral exposure in pregnant female mice showed that methoxyethanol is very rapidly distributed throughout the body. Peak concentrations of it and/or its metabolites persist for about 2 hours before elimination proceeds but concentrations of the substance and/or metabolite remain relatively high in the liver and conceptus compartment and at levels 2 -4x blood concentrations. Radio labelled material was detectable in sensitive areas of the developing embryo.

 

METABOLISM AND ELIMINATION

 

A toxicokinetic study using radiolabelled 2-methoxyethanol showed that approximately 95% of orally ingested material is excreted within 48 hours and that the main route of metabolism accounting for more than 50% of ingested material is to 2-methoxyacetic acid, which is excreted in urine. A smaller but significant amount is metabolized to CO2 and exhaled in air.

 

A pharmacokinetic study showed coadministration of ethanol by i.p. significantly inhibited the metabolism of methoxyethanol when administered by inhalation,suggesting that alcohol dehydrogenase metabolism is an important pathway for the metabolism of methoxyethanol.

 

A pharmacokinetic study in humans using inhalation exposure showed that the rate of metabolism of methoxyethanol is relatively slow, with a half life of 77 hours being calculated.

 

A study which dosed pregnant mice orally with methoxyethanol with and without other substances such as 4-methyl pyrazole and/or ethanol which can block/saturate the alcohol dehydrogenase enzyme showed that metabolism is essential to produce the typical pattern of developmental toxicity seen and that this enzyme plays a key role in such metabolism.

 

A metabolism study in the pregnant mouse showed that, following oral exposure, methoxyethanol is eliminated primarily in the urine as methoxyacetic acid in pure form or conjugated with glycine. Most product is eliminated within 48hrs. 

 

A metabolism study in the pregnant mouse showed that, following oral exposure, methoxyethanol is eliminated in the urine as primarily methoxyacetic acid in pure form or conjugated with glycine. A number of other direct metabolites, including ethylene glycol, sulphate and other conjugates were detected in urine along with metabolites of methoxyacetic acid, such as the glycine conjugate and derivatives of methoxyacetyl CoA.

 

A pharmacokinetic study using oral exposure in pregnant female mice showed thatmethoxyethanol is predominantly eliminated in the urine. Very little radioactivity was detected in the conceptus.

 

A developmental toxicity study in the monkey with pharmacokinetic observations showed that the mainmetabolite of methoxyethanol (methoxyacetic acid - MAA) and the metabolite that causes developmental toxicity) has a relatively long half life (22hrs) which can lead to accumulation of the toxicant when daily dosing occurs. The metabolite was found to readily crosses the placenta and distribute evenly between embryo and dam in pregnant monkeys. The yolk sac showed signs of accumulating MAA, but the toxicological significance of this is unknown.

Discussion on absorption rate:

An in vitro dermal absorption study using human skin showed that methoxyethanol readily passes through the stratum corneum and causes slight but detectable damage to the skin.