Reference dose

A reference dose is the United States Environmental Protection Agency's maximum acceptable oral dose of a toxic substance. Reference doses are most commonly determined for pesticides. The EPA defines an oral reference dose (abbreviated RfD) as: "[A]n estimate, with uncertainty spanning perhaps an order of magnitude, of a daily oral exposure to the human population (including sensitive subgroups) that is likely to be without an appreciable risk of deleterious effects during a lifetime."

Regulatory Status
Unlike National Ambient Air Quality Standards, RfDs are not enforceable standards. Instead, the EPA uses RfDs as risk assessment benchmarks and tries to set other regulations so that people are not exposed to chemicals in amounts that exceed RfDs. According to the EPA, "[a]n aggregate daily exposure to a [chemical] at or below the RfD (expressed as 100 percent or less of the RfD) is generally considered acceptable by EPA." For example, the EPA set an acute RfD for children of 0.0015mg/kg/day for the organochlorine insecticide endosulfan, based on neurological effects observed in test animals. The EPA then looked at dietary exposure to endosulfan, and found that for the most exposed 0.1% of children age 1-6, their daily consumption of the endosulfan exceeded this RfD. To remedy this, the EPA revoked the use of endosulfan on the crops that contributed the most to exposure of children: certain beans, peas, spinach, and grapes.

Types of RfDs
Reference doses are chemical specific, i.e. the EPA determines a unique reference dose for every substance it evaluates. Often separate acute and chronic RfDs are determined for the same substance. Reference doses are specific to dietary exposure. When assessing inhalation exposure, EPA uses "Reference Concentrations," (RfCs), instead of RfDs. Note that RfDs apply only to non-cancer effects. When evaluating carinogenic effects, the EPA uses the Q1* method.

Determination
RfDs are usually derived from from animal studies. Animals (typically rats) are dosed with varying amounts of the substance in question, and the largest dose at which no effects are observed is identified. This dose level is called the "No observable effect level," or NOEL. To account for the fact that humans may be more or less sensitive than the test animal, a 10-fold uncertainty factor is usually applied to the NOAEL. This uncertainty factor is called the "interspecies uncertainty factor" or Ufinter. An additional 10-fold uncertainty factor, the "intraspecies uncertainty factor" or Ufintra, is usually applied to account for the fact that some humans may be substantually more sensitive to the effects of substances than others. Additional uncertainty factors may also be applied. In general:


 * $$RfD (mg/kg/day) = {NOEL (mg/kg/day) \over Uf_{inter} * Uf_{intra} * Uf_{other}}$$

Frequently, a "No observable adverse effect level" or NOAEL is used in place of a NOEL. If adverse effects are observed at all dose levels tested, then the smallest dose tested, the "Lowest observed adverse effect level" or LOAEL, is used to calculate the RfD. An additional uncertainty factor usually applied in these cases, since the NOAEL, by definition, would be lower than the LOAEL had it been observed. If studies using human subjects are used to determine a RfD, then the interspecies uncertainty factor can be reduced to 1, but generally the 10-fold intraspecies uncertainty factor is retained. Such studies are rare.

An Example
As an example, consider the following determination of the RfD for the insecticide chlorpyrifos, adapted from the EPA's Interim Reregistration Eligibility Decision for chlorpyrifos.

The EPA determined the actue RfD to be 0.005 mg/kg/day based on a study in which male rats were administered a one time dose of chlorpyrifos and blood cholinesterase activity was monitored. Cholinesterase inhibition was observed at all dose levels tested, the lowest of which was 1.5 mg/kg. This level was thus identified at the lowest observed adverse effect level (LOAEL). A NOAEL of 0.5 mg/kg was estimated by dividing the LOAEL by a 3-fold uncertainty factor. The NOAEL was then divided by the standard 10-fold inter- and 10-fold intraspecies unscertainty factors to arrive at the RfD of 0.005 mg/kg/day. Other studies showed that fetuses and children are even more sensitive to chlorpyrifos than adults, so the EPA applies an additional 10-fold uncertainty factor to protect this subpopulation. A RfD that has been divided by an additional uncertainty factor that only applies to certain populations is called a "populaiton adjusted dose" or PAD. For chlorpyrifos, the acute PAD (or "aPAD") is thus 5*10-4 mg/kg/day, and it applies to infants, children, and women who are breast feeding.

The EPA also determined a chronic RfD for chlorpyrifos exposure based on studies in which animals were administered low doses of the pesticide for 2 years. Cholinesterase inhibition was observed at all dose levels tested, and a NOAEL of 0.03 mg/kg/day estimated by dividing a LOAEL of 0.3 mg/kg/day by an uncertainty facor of 10. As with the acute RfD, the chronic RfD of 3*10-4 mg/kg/day was determined by dividing this NOAEL by the inter- and intraspecies uncertainty factors. The chronic PAD ("cPAD") of 3*10-5 mg/kg/day was determined by applying an additional 10-fold uncertainty factor to account for the increased susceptibility of infants and children. Like the aPAD, this cPAD applies to infants, children, and breast feeding women.