Crab-shell chitin has been shown to effectively reduce SO 4 2-, enhance alkalinity, and remove metals from mine impacted water (MIW) at the bench scale and in field trials. To date, this research has been conducted using inexpensive, raw crab shell (ChitoRem" SC-20), which in addition to chitin, contains CaCO 3 and residual protein. All three of these components contribute to MIW treatment simultaneously, and are therefore difficult to uncouple. In this experiment, three different purities of crab shell (SC-20, SC-40, and SC-80) and limestone were tested for their ability to remediate natural MIW from an abandoned coal-mine in central Pennsylvania. The goals of this project were: 1) to compare the efficiency of metals and sulfate removal between different purities of chitin; and, 2) to begin to uncouple the contributions of chitin, protein, and calcium carbonate when raw crab shell chitin is used for the treatment of acid mine drainage. Sacrificial batch microcosms containing natural MIW, stream sediment, and either SC-20, SC-40, SC-80, or limestone were established in duplicate and incubated at room temperature for up to 117 days. The most complete and rapid metals removal was observed with SC-20, followed by SC-40, SC-80, and limestone. SC-20 removed more than 99% of Al, Fe, and Zn and more than 98% of dissolved Mn. SC-40 exhibited similar metals removal, but at slower rates. SC-80 and limestone were not effective at removing Mn. Alkalinity production followed similar trends, with SC-20 surpassing the other substrates with a total alkalinity of 1175 mg/L as CaCO 3 after 117 days. Elevated NH 4 production was observed at early times only with SC-20, indicating that it is residual protein, not chitin, releasing this nutrient. It is likely that the rapid dissolution of CaCO 3 from the crab shell, coupled with NH 4 release and biological SO 4 2- reduction all contributed to elevated alkalinity values and consequently superior metals removal with SC-20. Preliminary geochemical modeling suggests that the probable mechanisms for metals removal with SC-20 include precipitation of Al and Fe oxides/hydroxides and manganese carbonates, as well as physical adsorption onto components of the crab shell.