**A New Method for Extraction and Analysis of Ricin Samples through MALDI-TOF-MS**/**MS**

#### **Roberto B. Sousa 1, Keila S. C. Lima 2, Caleb G. M. Santos 2, Tanos C. C. França 3,4, Eugenie Nepovimova 4, Kamil Kuca 4,\*, Marcos R. Dornelas 2 and Antonio L. S. Lima 5,\***


Received: 16 February 2019; Accepted: 2 April 2019; Published: 3 April 2019

**Abstract:** We report for the first time the efficient use of accelerated solvent extraction (ASE) for extraction of ricin to analytical purposes, followed by the combined use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and MALDI-TOF MS/MS method. That has provided a fast and unambiguous method of ricin identification for in real cases of forensic investigation of suspected samples. Additionally, MALDI-TOF MS was applied to characterize the presence and the toxic activity of ricin in irradiated samples. Samples containing ricin were subjected to ASE, irradiated with different dosages of gamma radiation, and analyzed by MALDI-TOF MS/MS for verification of the intact protein signal. For identification purposes, samples were previously subjected to SDS-PAGE, for purification and separation of the chains, followed by digestion with trypsin, and analysis by MALDI-TOF MS/MS. The results were confirmed by verification of the amino acid sequences of some selected peptides by MALDI-TOF MS/MS. The samples residual toxic activity was evaluated through incubation with a DNA substrate, to simulate the attack by ricin, followed by MALDI-TOF MS/MS analyses.

**Keywords:** ricin; MALDI-TOF MS; chemical weapons; biological weapons; CBRN defense

**Key Contribution:** ASE; SDS-PAGE; MALDI-TOF MS and MALDI-TOF MS/MS combined provide an efficient method for identification of ricin.
