Department of Preventive Medicine and Public Health, Faculty of Pharmacy, University of Valencia, Av. Vicent Andrés Estellés s/n, 46100 Burjassot, Spain.
Anal Bioanal Chem. 2011 May;400(5):1525-31. doi: 10.1007/s00216-011-4906-6. Epub 2011 Mar 27.
Staphylococcal enterotoxin A (SEA) is an exotoxin excreted mainly by Staphylococcus aureus and nowadays is the most prevalent compound in staphylococcal food poisoning worldwide. SEA is highly heat-resistant, and usual cooking times and temperatures are unlikely to completely inactivate it. A procedure for extraction of this toxin based on protein precipitation with a mixture of dichloromethane and acidified water was used before SDS-PAGE separation of soluble proteins. Finally, bands of interest were excised from the gel and in-gel enzymatic digestion was done. SEA from pasteurized milk was detected with matrix-assisted laser-desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. Nineteen peptides (range 800-2400 Da) were identified as products of trypsin cleavage of the SEA standard with a score of 204 and 73% coverage of the protein sequence, whereas thirteen peptides were revealed for SEA extracted from milk with a score of 148 and 58% sequence coverage obtained. This procedure has been applied successfully for identification of SEA in milk.
金黄色葡萄球菌肠毒素 A（SEA）是一种主要由金黄色葡萄球菌分泌的外毒素，如今是全球范围内导致葡萄球菌食物中毒的最常见化合物。SEA 具有很强的耐热性，通常的烹饪时间和温度不太可能将其完全灭活。在 SDS-PAGE 分离可溶性蛋白之前，曾使用一种基于二氯甲烷和酸化水混合物的蛋白沉淀方法来提取这种毒素。最后，从凝胶中切下感兴趣的条带，并进行胶内酶解。采用基质辅助激光解吸/电离飞行时间（MALDI-TOF）质谱法检测巴氏奶中的 SEA。鉴定出 19 个肽段（范围为 800-2400 Da），它们是 SEA 标准品经胰蛋白酶切割的产物，得分 204，蛋白质序列覆盖率为 73%，而从牛奶中提取的 SEA 则显示出 13 个肽段，得分 148，序列覆盖率为 58%。该方法已成功应用于牛奶中 SEA 的鉴定。
