## Supplementary Data

Basic model structure common for all the life stages during gestation and lactation To simulate life stage specific processes, additional compartment(s) were added to the six existing compartments of lung, nose, brain, bone, liver, and a lumped body compartment (rest of body) in the adult male model (Figure 2). The mammary gland was added for the adult female and lactating female and was described using tissue binding parameters same as the rest of body compartment. In the pregnant fema le, the placenta was additionally modeled to simulate the gestation . The placenta doesn’t have Mn binding capacity. In the fetus, the lung, brain, bone, liver, and rest of body compartment were modeled (Figure 2). The neonate and child were modeled using t he same tissue compartment structure as the adult male (Figure 2). Three representative brain regions along with the brain blood were explicitly described to simulate Mn kinetics in the brains of th e pregnant and lactating mother , fetus, neonate, and chil d as in the adult male. The globus pallidus, cerebellum, and olfactory bulb were included based on the criteria of being a target for potential Mn neurotoxicity, a representative of general brain tissue, or a region with expected contributions from direct transport from olfactory epithelium, respectively. The pituitary gland was not i ncluded in the present model due to limited information on tissue Mn kinetics in this brain region in early life in humans. Mn exposure for the mother and neonate/child was ei ther through diet only or simultaneously through diet and inhalation. Only a small fraction of dietary Mn is taken into the liver (FdietUp). The dietary intake for the suckling neonate was through breast

- milk. The FdietUp was appropriately varied from the adult male model to account for the known gender difference in gut absorption. The impacts of early age and/or the molecular forms of Mn in infant food sources on the gut absorption of Mn were also modeled by changing neonatal FDietUP for each type of milk , either breast

- milk or infant formulas , accordingly.In the current model, a ce rtain portion of unabsorbed Mn (Fent) is transiently stored in gut

epithelial cells. They are not absorbed to the systemic circulation due to the rapid turnover rate of gut
ep ithelium in adults, while they do in neonates as a result of slower turnover of the enterocytes in their gut compared to the adults. This delayed absorption in neonates was dependent on an absorption rate constant delGI in the current model . The sloughing rate of gut epithelial cells is reflected by a rate constant of kent, which describes the slow loss of Mn from the gut epithelium to feces (Figure 2)

. For simulating inhalation exposure, the absorption of deposited Mn in the lung and nasal cavity to the s ystemic circulation was rapid both in the mother and neonate/child as the adult male model. A direct transport from olfactory epithelium to the olfactory bulb region in the brain was included for all the life stages except the fetus. After enter ing the sy stemic circulation, Mn is then subsequently cleared into the tissues. This clearance was described as an asymmetric bi

- directional diffusion for the brain. The free Mn in systemic blood rapidly equilibrates with free Mn in the brain tissue blood. The flux between brain tissue

- blood and brain tissue are diffusion limited, the rate of which are determined by kin and kout, respectively ( Figure 2; Table 2). These values are region specific and are responsible for the preferential increase in free Mn in the mid

- brain region seen in excess Mn exposures. The diffusion limitation in the brain was considered reflecting the role of blood

- brain barrier in controlling Mn flux . The feature of asymmetrical diffusion was brain region specific, which was rationalized by reg ion

- specific expression of transporter proteins for essential element in the brain and the underlying mechanisms for the reported preferential influx of Mn into the mid

- brain in over

- exposure conditions . The se brain diffusion rate constants were varied dur ing perinatal period to reflect maturational changes both in the blood

- brain barrier integrity and in the types/ levels of Mn transporter expression that are likely in parallel with the needs for this element during the active neuro development. For other tissues, a rapid and symmetrical flux of Mn between the systemic blood and tissue compartment was used and thus the tissue Mn is determined by the tissue binding characteristics.Once distributed into the tissue, Mn is bound t o the cellular compo nents that require Mn for a

variety of biological functions and thereby it is not readily crossing cell membranes. This incorporation of
Mn into cellular proteins or tissue constituents and subsequent degradation from those ‘binding sites’ are governed by tissue spe cific association and dissociation rate constants ka and kd, respectively. Total Mn in any tissue is therefore, the sum of free and bound forms within the tissue, the ratio between the two governed by the dissociation equilibrium constant (kd/ka) as well a s the maximum binding capacity (Bmax). Once the binding sites are saturated, free Mn will rise swiftly, consequently a rapid increase in total tissue Mn occurs . The Bmax and binding rate constants are likely determined not only by the degree of needs for M n in a specific tissue, but also by the nature of those needs, i.e., biological functions requiring Mn in that tissue. In this reagard, tissue binding constants were subject to vary during per

- and postnatal growth in the currenet model. Free Mn is the form that circulates in blood and is available for excretion into bile and for placental or lactational transfer. The bound form stays in the tissue and slowly dissociates from the binding sites with a rate constant kd , which probably corresponds to biological degradation rates of cell materials such as Mn

- containing enzymes . At all times, the fraction bound in a specific tissue was kept over 70% to be consistent with our assumption on the nature of Mn tissue binding. We assumed that most of Mn in the tissue is associated with cellular constituents. The fraction bound can be changed depending on the Mn content in diet. With a sufficient Mn diet, the fraction of Mn bound in a tissue would be lower, while with a marginal Mn diet, it should become higher. It should be noted that the fraction bound does not represent the actual occupancy of binding sites, though. The description for the free Mn dependent influx control in the globus pallidus region used i n the adult male model was kept similar when simulating all th e life stages in the current model. The kin M ax , the maximal fold of induction , was slightly reduced in the fetus and neonate . The kin50 values were retained the same as the adult male due to limited data on this additional layer of brain Mn control, especi ally during gestation and lactation (Table 2).

Simulating placental transfer and the fetusAll the characteristics of the placental transfer of Mn were retained the same as described in the

rat gestation model (Yoon et al. , 2009a)

. Transfer of Mn from placenta to fetal venous blood was mediated by two pathways, one is an active process described by the maximum rate of transport VmaxC ( μ g/hr/kg BW F 0.75 ) and the affinity constant Km ( μ g/L) ; and the ot her is a diffusion process governed by a diffusional clearance rate constant ktrans1C (L/hr/kg BW F 0.75 ). A minor transfer of Mn from fetal arterial blood to placenta also occurs, which is driven by diffusion governed by another clearance rate constant ktra ns2C (L/hr/kg BW F 0.75 ). Both the active and diffusion processes were assumed to be correlated with fetal growth and were scaled to fetal BW 0.75 . The placenta to maternal blood partition coefficient , PPla , reflects the high placenta to blood ratio and the e xperimentally observed Mn trapping in the placenta

. The fetus acquires Mn solely from the placenta. The excretion of Mn from the fetus is to the placenta only (Figure 2)

. The movements of Mn between maternal and fetal circulations were modeled assuming m ajority of transfer being maternal to fetal direction through active transport(s) following the same approach applied in the rat model (Yoon , et al. , 2009a) . P arameters for this placental transfer of Mn were then estimated based on the observed placental and fetal tissue Mn concentrations in late gestation and/or at delivery in humans (Casey and Robinson, 1978; Osman et al. , 2000; Takser et al. , 2004)

. T he

es timation criteria was t o allow the active process to work near at its maximum capacity at normal steady state for the pregnant mother on adequate Mn intake as we did for the rat model (Table 2). To achieve this, model parameters for maternal to fetal flux, both for active transport, i.e., VmaxC and Km, and for diffusion, i.e., ktrans1C were varied until the active transfer becomes more than 95% of the flux to the fetal side with 5% or less contribution from simple diffusion. The active process was assumed t o work almost at its saturation, at about 75% of its maximum, by adjusting the affinity constant Km to an appropriate value (Table 2). The diffusion clearance parameter from the fetal arterial blood to the maternal placenta, ktrans2C, was set the same as k trans1C. With t he se feto

- placental parameter s, we

were able to successfully recapitulate the reported Mn concentrations in the placenta and in several fetal tissues at late or at the end of gestation without inhalation exposure in the mother in humans (Fi gure 3 ).The placenta to blood partition coefficient for Mn, PPla , was estimated to reproduce the

concentration ratio between the placenta and maternal blood seen at the end of pregnancy and the resulting value was 10 (Takser , et al. , 2004) (Tabl e 2). A kinetic assessment of Mn using the dually perfus ed human placenta in vitro supported this high PPla . A rapid accumulation of substantial portion of
Mn was observed within the placental tissue after Mn was added in the perfusate (Miller et al., 1987 ).

Simulating lactational transfer and the suckling neonate The free Mn in mammary gland is secreted into milk with a clearance rate constant CLmilkC (L/hr/kg BW M 0.75 ; Table 2) as described in the rat lactation model (Yoon et al. , 2009b) . The clearance of Mn into milk is assumed to be proportional to maternal BW 0.75

. T he rate of milk production (k lac , L/hr)

was assumed to be the same as the milk consumption rate by infants (Casey et al ., 1985). The concentration of milk Mn (C milk , μg/L) is defined as:
(2)

where CL milk indicates the lactational clearance of Mn (μg/hr) and C f_ Mam is the free Mn concentration in the mammary gland tissue The impacts of early age and/or the molecular forms of Mn in infant food sources on gut absorption of Mn were modeled by adjusting neonatal FDietUP for each type of milk , either breast

- milk or infant formulas , accordingly. E nhanced absorption for breast milk Mn was incorporat ed using a constant FDP so that the actual fraction absorbed in neonate was FDP times FdietUp (Table 2). In addition to the enhanced absorption, the description for Mn movement in gastrointestinal (GI) tract in suckling neonate was modeled differently fro m adult s . In the neonate, some of the Mn transiently trapped in enterocytes ( Fent ) is absorbed to the systemic circulation due to slower turnover of the enterocytes ( kent ) compared to the adults. This delayed absorption was described with an absorption ra te
constant delGI in the current model . Maturation of GI epithelium was assumed to occur at aroundweaning and the sloughing rate reaches the adult level. Otherwise, the suckling neonate was modeled

following the same Mn kinetic description as adults.

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- 58. Yoon, M., Nong, A., Clewell, H. J., Taylor, M. D., Dorman, D. C. and Andersen, M. E. (2009b). Lactational transfer of manganese in rats: Predicting manganese tissue concentration in the dam and pups from inhalation exposure with a pharmacokinetic model. Toxicological Sciences 112 , 23

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