*4.3. Castor Bean Seeds*

The castor bean seed used belonged to the species *R. communis* L., cultivar IAC Guarani, harvest 2012/2012, category S2, lot 05/2012, with 96.80% of purity. They were received from the Company "BR Seeds Production and Commerce of Seeds Ltda" based in the city of Araçatuba in the São Paulo State, Brazil.

#### *4.4. Production of the Ricin Samples*

Samples containing ricin were produced from the seeds of castor bean (*R. communis* L.) adapting the extraction method with acetone described in the literature [34,35,44], for ASE. This was performed in a Dionex extractor (Thermo Fischer Scientific, Waltham, MA, USA), model ASE100, with a 100 mL cylindrical extraction cell made of stainless steel. Firstly, the seeds were peeled with the help of tweezers and a spatula, until exposing their whitish inner part. After, they were milled in an 80 mL Ika® stainless steel knives mill, model A11 from Ika manufacturer. The oiled mass (4 g) was transferred to the extraction cell that was inserted in the oven of the extractor ASE100. The temperature was programmed to stay constant at 40 ◦C. Extraction was performed with a mixture 8:2 of the solvents n-hexane 99% UV/HPLC-Spectroscopic from Vetec (Rio de Janeiro, Brazil), and acetone 99.8% HPLC from J. T. Baker. This mixture was pumped into the cell, filling the whole volume, and raising the pressure to 1650 psi. After 5 min under this pressure, the extract obtained was filtered and collected into a flask. After removal of the extract, the cell was purged with nitrogen for 1 min and the extraction procedure repeated. After four rounds of extraction and evaporation of the solvent, around 1.2 g of a white powder containing ricin was separated from the extract for each extraction step.

All white powder produced was homogenized and separated in fractions of 1 g. Each fraction was packed into a 15 mL conic tube for centrifugation. After centrifugation, each tube was placed in a transparent plastic bag with a double closing system, identified externally and sent for irradiation. The samples were prepared in triplicate and named according to the irradiation dosages to be received (0 kGy, 10 kGy, 20 kGy and 30 kGy).
