Ovenden Simon P B, Fredriksson Sten-Ake, Bagas Christina K, Bergström Tomas, Thomson Stuart A, Nilsson Calle, Bourne David J
Defence Science and Technology Organisation, 506 Lorimer Street Fishermans Bend, Victoria 3207, Australia.
Anal Chem. 2009 May 15;81(10):3986-96. doi: 10.1021/ac900371y.
Ricinus communis (also know as the castor bean plant) whose forbears escaped from suburban gardens or commercial cultivation grow wild in many countries. In temperate and tropical climates seeds will develop to maturity, and plants may be perennial. In Australia these plants have become widespread and are regarded as noxious weeds in many localities. The seeds of R. communis contain ricin, a protein toxin which can easily be extracted into an aqueous solution. Ricin is toxic by ingestion, inhalation, and injection. The history of terrorist and anarchist interest in the use of seeds from R. communis has driven the development of strategies for determination of cultivar and geographic location of the source of an extract of wild-grown castor bean seed. This forensic information is of considerable interest to law enforcement and intelligence organizations. During forensic studies of both the metabolome and proteome of extracts from eight specimens of six different cultivars of R. communis ("zanzibariensis" collected from Kenya and Tanzania, "gibsonii", "impala", "dehradun", "carmencita", and "sanguineus" collected from Spain and Tanzania), three peptide biomarkers (designated Ricinus communis biomarkers, or RCB) were identified in both the MALDI and electrospray LC-MS spectra. Two of these peptides (RCB-1 and RCB-2) were present in varying amounts in all cultivars, while RCB-3 was present only in the "carmencita" cultivar. The amino acid sequences of RCB-1 to -3 were determined using LC-MS(n) fragmentation and de novo sequencing on both the intact and the carbamidomethyl modified peptides. The connectivity of the two disulfide bonds that were present in all three RCB were determined using a strategy of partial reduction and differential alkylation using tris-(2-carboxyethyl)phosphine with N-ethylmaleimide to reduce and alkylate the most accessible disulfide bond, followed by reduction and alkylation of the remaining disulfide bond with dithiolthreitol and iodoacetamide. The possible functional role of RCB-1 to -3 in R. communis seeds is also discussed.
蓖麻（也被称为蓖麻子植物），其祖先从郊区花园或商业种植中逸出，在许多国家野生生长。在温带和热带气候下，种子会发育成熟，植株可能为多年生。在澳大利亚，这些植物已广泛分布，在许多地区被视为有害杂草。蓖麻种子含有蓖麻毒素，一种蛋白质毒素，它能很容易地被提取到水溶液中。蓖麻毒素通过摄入、吸入和注射均具有毒性。恐怖分子和无政府主义者对使用蓖麻种子的兴趣推动了确定野生蓖麻种子提取物来源的品种和地理位置策略的发展。这些法医信息对执法和情报组织具有相当大的吸引力。在对六个不同品种的八个蓖麻样本（从肯尼亚和坦桑尼亚收集的“桑给巴尔ensis”、从西班牙和坦桑尼亚收集的“吉布森ii”、“黑斑羚”、“德拉敦”、“卡门西塔”和“血色”）提取物的代谢组和蛋白质组进行法医研究期间，在基质辅助激光解吸电离（MALDI）和电喷雾液相色谱 - 质谱（LC - MS）光谱中鉴定出三种肽生物标志物（称为蓖麻生物标志物，或RCB）。其中两种肽（RCB - 1和RCB - 2）在所有品种中含量各不相同，而RCB - 3仅存在于“卡门西塔”品种中。使用LC - MS(n)碎片化和从头测序对完整肽和氨基甲酰甲基修饰肽确定了RCB - 1至 - 3的氨基酸序列。使用三（2 - 羧乙基）膦与N - 乙基马来酰亚胺进行部分还原和差异烷基化的策略确定了所有三种RCB中存在的两个二硫键的连接性，以还原和烷基化最易接近的二硫键，然后用二硫苏糖醇和碘乙酰胺还原和烷基化剩余的二硫键。还讨论了RCB - 1至 - 3在蓖麻种子中可能的功能作用。
