Abstract
Ricin is a protein toxin produced by the castor bean plant (Ricinus communis) together with a related protein known as R. communis agglutinin (RCA120). Mass spectrometric (MS) assays have the capacity to unambiguously identify ricin and to detect ricin’s activity in samples with complex matrices. These qualitative and quantitative assays enable detection and differentiation of ricin from the less toxic RCA120 through determination of the amino acid sequence of the protein in question, and active ricin can be monitored by MS as the release of adenine from the depurination of a nucleic acid substrate. In this work, we describe the application of MS-based methods to detect, differentiate and quantify ricin and RCA120 in nine blinded samples supplied as part of the EQuATox proficiency test. Overall, MS-based assays successfully identified all samples containing ricin or RCA120 with the exception of the sample spiked with the lowest concentration (0.414 ng/mL). In fact, mass spectrometry was the most successful method for differentiation of ricin and RCA120 based on amino acid determination. Mass spectrometric methods were also successful at ranking the functional activities of the samples, successfully yielding semi-quantitative results. These results indicate that MS-based assays are excellent techniques to detect, differentiate, and quantify ricin and RCA120 in complex matrices.
Highlights
Ricin is a protein toxin produced by the castor bean plant, Ricinus communis
Through the EQuATox study, it has been demonstrated that mass spectrometric measurements of ricin play a vital role in correct identification of ricin in complex matrices
In analyses with limited sample volume and the desire to perform multiple mass spectrometric measurements, a valid approach would be to divide each sample into separate aliquots for enzymatic activity measurements and structural measurements, perhaps putting more emphasis on the enzymatic activity measurements for public health purposes or highlighting the structural measurements for forensic purposes
Summary
Ricin is a protein toxin produced by the castor bean plant, Ricinus communis. It is composed of two polypeptide chains, known as the A- and B-chains, which are each approximately 32 kDa, making intact ricin approximately 64 kDa with glycosylation [1]. The simplest technique for analysis of ricin by mass spectrometry is to detect the intact ricin protein with an approximate molecular weight of 64 kDa. the detection of a signal with a mass close to 64 kDa could indicate the presence of ricin. Analysis of intact ricin can provide useful information including differentiation between ricin and RCA120, it is not a definitive analysis of ricin, as there are many proteins, which have the same approximate molecular weight as ricin. This method suffers from the high limit of detection of concentrations greater than 1 μg/mL [17]. In combination with other detection methods with higher specificity, this analysis can provide useful information
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