Journal of Toxicology public health oﬃcials, environmental legislators, advocacy groups, and fuel manufacturers. USEPA health eﬀects testing program for MMT emerged from this challenging climate. 3. Overview of the Regulatory History of MMT Manganese is listed as an air toxic by the USEPA. When the USEPA ordered the phasing out of leaded gasoline, MMT and other alternative octane enhancers were used in unleaded gasoline. In 1977, a Congressional amendment to the CAA banned the use of all fuel additives not “substantially similar” to gasoline, including MMT, unless the USEPA granted a waiver [21]. The prohibition on the use of MMT in gasoline was largely based on concerns that MMT use could aﬀect the ﬁrst generation of automotive emissions- control systems. Ethyl Corporation ﬁrst applied for this waiver in 1978. This and several subsequent waiver petitions submitted by Ethyl Corporation were denied because of USEPA concerns regarding potential increases in exhaust hydrocarbon emissions resulting from MMT use [21]. As part of a later waiver application for MMT, the USEPA conducted a risk assessment of exposure to inhaled manganese in 1993, with the publication of an inhalation RfC of 0.05 µg/m3 [6]. The USEPA used standard RfC devel- opment methodologies for a noncancer endpoint and based the RfC on the Roels et al. study [10] that evaluated neu- robehavioral and motor movement impairments observed in workers exposed to manganese dioxide (MnO2). The exposure concentration reported by Roels was considered by the USEPA to be a lowest-observable-adverse-eﬀect level (LOAEL). The LOAEL value was adjusted for continuous exposure durations, and several uncertainty factors were applied. These uncertainty factors included use of a LOAEL instead of a no-observable-adverse-eﬀect level (NOAEL), extrapolation from subchronic to chronic, protection of potential sensitive members of the human population, and a factor reﬂecting other uncertainties in the database, such as less-than chronic periods of exposure, inadequate informa- tion regarding developmental and reproductive toxicity, and uncertainty about the toxicity of various forms of manganese [22]. Although the nutritional essentiality of manganese in the diet was discussed in the documentation of the RfC, it played no practical role in the calculation of an RfC [21]. The USEPA also published a risk characterization that included exposure and dose-response assessments for MMT in gasoline [23, 24]. After completing this evaluation, EPA determined that use of MMT would not “cause or contribute” to the failure of vehicle emission control systems. EPA was unable to determine, however, whether a risk to the public health occurred from use in gasoline [25]. The agency stated “[a]lthough it is not possible based on the present information to conclude whether speciﬁc adverse health eﬀects will be associated with manganese exposures in the vicinity of or exceeding the (estimated safe level over a lifetime of exposure), neither is it possible to conclude that adverse health eﬀects will not be associated with such exposures. . .Given the information that is available at present and the uncertainties discussed here, a reasonable basis exists 3 for concern regarding potential public health risks, especially for sensitive subpopulations, if MMT were to be widely used in unleaded gasoline [25].” The USEPA also concluded that long-term animal testing and exposure research were needed to more accurately deﬁne the risk. Coincidental to these activities, in July 1994, the USEPA Administrator denied Ethyl Corporation newest waiver petition speciﬁcally because of concerns about potential risks to public health [24] and refused to register MMT for use in the USA In 1995, the Ethyl Corporation successfully challenged the denial of its petition based on public health concerns (Ethyl Corporation v. Browner, 51 F.3d 1053 (D.C. Cir. 1995)), as well as USEPA decision not to register MMT (Ethyl Corporation v. Browner, 67 F.3d 941 (D.C. Cir. 1995)). As a result, the USEPA formally approved the use of MMT in conventional unleaded gasoline and registered it as a fuel additive under the CAA, allowing for its domestic sale. Provisions of the CAA provide the USEPA with the authority to require testing of fuels and fuel additives used in motor vehicles, including MMT, to help ﬁll data gaps and provide information that potentially would result in a more deﬁnitive risk evaluation. These health testing requirements are addressed in Sections 211(b)(2) and 211(e) of the CAA. Section 211(b)(2) states “For the purpose of registration of fuels and fuel additives, the Administrator shall, on a regular basis, require the manufacturer of any fuel or fuel additive” to conduct “Tier 2” tests to determine potential public health and environmental eﬀects of the fuel or additive (including carcinogenic, teratogenic, or mutagenic eﬀects). These stud- ies would be conducted using test procedures and protocols established by the USEPA. Moreover, the CAA also provides USEPA with the discretion to modify the standard Tier 2 health eﬀects testing requirements for a fuel or fuel additive by substituting, adding, or deleting testing requirements or changing the underlying vehicle/engine speciﬁcations (40 CFR 79.58(c)). Health eﬀects testing for MMT fell under this so-called “Alternative Tier 2” requirement, as the concern and subsequent testing was focused around the inorganic exhaust products of MMT and not MMT itself. 4. Establishment of the Alternative Tier 2 Testing Program for MMT In 2001, the Ethyl Corporation was notiﬁed by the USEPA of the Alternative Tier 2 provisions for MMT which fell within two general categories: pharmacokinetic testing of manganese compounds and characterization of manganese emissions from vehicles utilizing fuels containing MMT. A central objective of the MMT Alternative Tier 2 program was to generate data to support development of physiologically based pharmacokinetic (PBPK) models for manganese. As a result, PBPK model development became the subject of a series of studies funded by Afton Chemical Corp. and handled in a way similar to other facets of the Alternative Tier 2 program. Multiple pharmacokinetic studies were performed in response to the USEPA mandate (Table 1). Studies that were required by the USEPA were performed in accordance with