DEVELOPMENT OF LC/MS METHODS FOR OPIOID PEPTIDES

我们致力于开发和完善在线高性能液体 色谱/质谱（HPLC/MS）技术用于灵敏和 生物基质中阿片肽的特异性测定。 宽 已知阿片肽的分子量范围，极低的 这些肽在复杂的生物材料中的浓度， 不断增加的单个肽挑战当前分析 方法论 该方案将通过调查解决这些问题： (1)电离技术，如热喷雾和快速原子轰击 (FAB)，其特别适用于高分子量化合物， 用于阿片肽的在线HPLC/MS测定，和（2）新质量 质谱方法，如MS/MS和高分辨率/MS结合 HPLC/MS，有助于减少化学噪音，提高灵敏度， 提供更多的结构信息。 使用FAB和热喷雾电离技术，样品分子不需要 在气相中发生电离，因此具有高分子量 可以分析肽（分子量约10，000）。 准确 高分子量范围内的分子量测定应 可以使用这些软电离技术与 仪器有效传输高质量离子的能力。 我们将研究使用高分辨率/MS检测，MS/MS（在 质量分析离子动能谱法[MIKES]的形式），以及位移 试剂 这些技术应提供降低的化学噪声， 从而增加灵敏度，并且还应该提供额外的 用于确定结构的结构信息。 所有HPLC/MS开发将使用标准品溶液进行。 各种阿片肽 注意样品处理和 清理，主要依靠以前发布的程序。 验证 将使用加标组织进行开发的方法学， 脑脊液和尿液 在该方案下开发的方法， 用于在各种生物学中分析阿片肽的程序 组织和液体应该证明是有价值的，有助于研究的， 阿片类药物谱与某些精神和代谢的关系 紊乱

We seek to develop and perfect on-line high performance liquid chromatography/mass spectrometry (HPLC/MS) techniques for the sensitive and specific determination of opioid peptides in biological matrices. The wide range of molecular weights of the known opioid peptides, the extremely low concentrations of these peptides in complex biological materials, and the ever-increasing number of individual peptides challenge current analytical methodology. This program will address these problems by investigating: (1) ionization techniques such as thermospray and fast atom bombardment (FAB), which are particularly useful with high molecular weight compounds, for on-line HPLC/MS determination of opioid peptides, and (2) new mass spectrometric methods such as MS/MS and high resolution/MS combined with HPLC/MS, which help to reduce chemical noise, improve sensitivity, and provide additional structural information. Using FAB and thermospray ionization techniques, sample molecules need not be in the gas phase for ionization to occur, thus high molecular weight peptides (molecular weight approximately 10,000) can be analyzed. Accurate molecular weight determinations in the high molecular weight ranges should be possible using these soft ionization techniques together with the instruments' ability to efficiently transmit high mass ions. We will investigate the use of high resolution/MS detection, MS/MS (in the form of mass analyzed ion kinetic energy spectrometry [MIKES]), and shift reagents. These techniques should provide reduced chemical noise and thereby increased sensitivity, and should also provide additional structural information for definitive structure determination. All HPLC/MS development will be performed using standard solutions of the various opioid peptides. Attention will be given to sample processing and cleanup, relying primarily on previously published procedures. Validation of the developed methodologies will be performed using spiked tissues, cerebrospinal fluid, and urine. Methods developed under this program, procedures for the profiling of the opioid peptides in various biological tissues and fluids should prove valuable in aiding the study of the relationships between opioid profiles and certain psychiatric and metabolic disorders.