Capillary Electrophoresis and MicrochipMass Spectrometry Interfaces and Their Utilization in Bisphosphonate Analysis

Miniaturization of analytical instruments is being driven by the growing demand for fast and automated analytical methods. Lower sample and reagent consumption, reduced chemical waste production, and low cost of the instruments are also highly relevant. Capillary electrophoresis (CE) is already a step toward decreasing the scale of analyses, providing efficient analytical separations with small sample and reagent volumes. A much more dramatic step is to apply microfabrication technology to analytical instruments, to manufacture "micro-total analysis systems (μ-TAS)" and "Lab-on-a-chip" devices. Mass spectrometry (MS) is a key technique for the highly specific and reliable detection of analytes in multicomponent samples. To couple CE and microfabricated devices with MS requires the use of efficient and robust interfacing methods. Miniaturized analytical techniques were developed utilizing CE, microchip technology, and MS. A series of forensic drugs and amino acids were used as test compounds in the construction and evaluation of the techniques and the techniques were applied for the analysis of bisphosphonate compounds. Bisphosphonates are an important class of drugs for the treatment of calcium metabolic disorders and of various bone diseases such as bone resorption and dissolution. Direct and specific methods are needed for their identification in bulk drug materials. Two coaxial sheath liquid interfaces were constructed and evaluated for the direct coupling of CE to electrospray ionization (ESI)-MS using triple quadrupole and ion trap mass analyzers. For the proper performance of the coaxial sheath liquid CE/ESI-MS system, various operation parameters were optimized; among these the sheath liquid composition and flow rate and the nebulizing and drying gas flows were found to be the most crucial. In addition, a sheathless nanospray CE/ESI-MS interface was constructed. Although higher sensitivity for CE/ESI-MS was provided by the sheathless nanospray technique, the sheath liquid interface was more robust and it worked in both positive and negative ion ESI-MS. Properties of porous silicon were optimized for a new, matrix-free laser desorption/ionization technique, desorption/ionization on silicon (DIOS). DIOS-MS provided fast and reliable identification of low-molecular-weight compounds. The sensitivity varied significantly for the various analytes examined (100 fmol–60 pmol). The highest response was obtained for the basic compounds analyzed in positive ion mode and lowest for the bisphosphonate and phosphonate compounds in negative ion mode. A microchip was also fabricated for ESI-MS. The electrospray was generated directly from a microchannel orifice. Use of poly(dimethylsiloxane) (PDMS) as microchip material aided the formation of a small Taylor cone in the ESI process. The fabrication process for the PDMS microchip was optimized to facilitate the ESI-MS coupling and to allow a straightforward and low-cost production of chips for ESI-MS. The performance of the PDMS microchip was demonstrated in directinfusion ESI-MS analysis. Fragmentation pathways of bisphosphonates were examined using multistage ESI tandem mass spectrometry with multistage precursor selection (ESI-MS, n=1–6). The DIOS-MS behavior of the bisphosphonates was also explored. The new CE/ESI-MS method was suitable for the selective identification and screening of bisphosphonates. Furthermore, a CE-