Lillard S J, Chiu D T, Scheller R H, Zare R N, Rodríguez-Cruz S E, Williams E R, Orwar O, Sandberg M, Lundqvist J A
Department of Chemistry, Howard Hughes Medical Institute, Stanford University, California 94305, USA.
Anal Chem. 1998 Sep 1;70(17):3517-24. doi: 10.1021/ac980462j.
Several amine-containing components of individual vesicles from the atrial gland of Aplysia californica were identified with capillary electrophoresis (CE). On-line derivatization with naphthalene-2,3-dicarboxaldehyde was performed, and the derivatized amine-containing components were detected with laser-induced fluorescence (LIF). Amino acids, including taurine, that had not been determined previously in atrial gland vesicles were observed by using CE-LIF, and their identities were confirmed with CE, HPLC, NMR, and electrospray ionization mass spectrometry. The finding that taurine is packaged and stored into secretory vesicles supports the hypothesis that taurine may exhibit neuromodulatory activity. The bioactive peptides, well-known to be in atrial gland vesicles, were detected in lysed vesicle samples fractionated with HPLC and analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. These peptides were also observed in single-vesicle runs with CE-LIF. The atrial gland vesicles (ranging from 0.5 to 2 microns diameter and 65 aL to 4 fL volume, respectively) studied in this work represent the smallest biological entities to be analyzed chemically on an individual basis.
利用毛细管电泳(CE)鉴定了加州海兔心房腺单个囊泡中的几种含胺成分。采用萘-2,3-二甲醛进行在线衍生化,并用激光诱导荧光(LIF)检测衍生化的含胺成分。通过CE-LIF观察到了此前未在心房腺囊泡中测定的包括牛磺酸在内的氨基酸,并通过CE、HPLC、NMR和电喷雾电离质谱法确认了它们的身份。牛磺酸被包装并储存到分泌囊泡中的这一发现支持了牛磺酸可能具有神经调节活性的假说。在通过HPLC分级分离并用基质辅助激光解吸/电离飞行时间质谱法分析的裂解囊泡样品中,检测到了心房腺囊泡中众所周知的生物活性肽。在CE-LIF的单囊泡分析中也观察到了这些肽。在这项研究中所研究的心房腺囊泡(直径分别为0.5至2微米,体积为65飞升至4飞升)代表了能够在个体基础上进行化学分析的最小生物实体。