Many plants, fungi, and bacterial species have cytotoxic agents called Ribosome Inactivating Proteins (RIPs). Due to the fact that the RIPs have a broad spectrum of antiviral activity, they have become an intricate part of making advancements in the study of antiviral agents. One such protein, Pokeweed Antiviral Protein (PAP), is a type I RIP which inhibits protein synthesis by depurinating the conserved Sarcin/ricin loop (SRL) of the large ribosomal RNA (rRNA). Our lab used 16S+23S and 18S+28S rRNA for protocol refinement and in order to confirm that our PAP from locally harvested plants is depurinating rRNA. Our modified protocol converts the free adenine into the fluorescent etheno-adenine derivative using chloroacetaldehyde. We then monitor the emission intensity using steady-state fluorescence spectroscopy. Here we report our PAP purification protocol, our modified fluorescence protocol for detection of adenine release, and depurination studies using an SRL oligoribonucleotide with three known depurination sites. This oligo can also be transcribed as a capped variant. This data will be correlated with binding data performed in our lab to explore the relationship between mRNA structure and depurination by PAP. In analyzing this relationship, it can lead to improving the application of RIPs to anti-pathogenic studies (plant and mammalian) as well as bioterrorism toxins.