Biomedical subjects

Jun Kameoka

Publications and source records attributed to Jun Kameoka.

4 recordsLinked to original sources

PubMed2005

A polymeric microchip with integrated tips and in situ polymerized monolith for electrospray mass spectrometry.

We describe the integration of a cyclo-olefin polymer based microchip with a sheathless capillary tip for electrospray ionization-mass spectrometry (ESI-MS). The microchip was fabricated by hot embossing and thermal bonding. Its design includes a side channel for adjusting the composition of the electrospray solution so that analytes in 100% water can be analyzed. The fused silica capillaries, used for sample introduction, and the electrospray tips for MS coupling were directly inserted into the microchannel before thermal bonding of the device. A microfabricated on-chip gold microelectrode was used to apply the electrospray voltage. Annealing the device after thermal bonding increased the pressure resistance of the microchip. The cross section of the microchannel was imaged by scanning electron microscopy to estimate the effects of the annealing step. The relationship between the applied electrospray voltages and MS signal was measured at different flow rates by coupling the device to an ion trap mass spectrometer. The performance of the microchip was evaluated by MS analysis of imipramine in ammonium acetate buffer solution by direct infusion. An alkylacrylate based monolith polymer bed for on-chip sample pretreatment and separation was polymerized in the microchannel and tested for ESI-MS applications.

Journal Article↗

PubMed2004

Quantitative mass spectrometric determination of methylphenidate concentration in urine using an electrospray ionization source integrated with a polymer microchip.

We have demonstrated the use of a simple microfabricated electrospray ionization source for coupling microfluidic chips to mass spectrometry (MS). A polymer-based microchip, coupled to a triple quadrupole mass spectrometer, has been employed for direct infusion quantitative bioanalysis of methylphenidate (Ritalin) extracted from human urine samples. The approach used a microfabricated polymer electrospray emitter to couple a microfluidic channel to a stable electrospray ionization source. The microchip was fabricated from cycloolefin plastic plate by hot embossing and thermal bonding. This microfluidic chip contained two independent microfluidic channels, integrated with two corresponding electrospray emitters and an internal gold electrode. Liquid-liquid extraction was used to prepare urine samples, spiked with methylphenidate. A trideuterated analogue of methylphenidate (methylphenidate-d(3)) was used as the internal standard for the analysis. The system showed good electrospray stability and reproducibility with different spray tips. Four different electrospray tips were used to analyze the same sample, and the results showed very small variation with a relative standard deviation of 1.4%. A standard curve prepared for methylphenidate in urine (R(2) = 0.999) was linear over the range of 0.4-800 ng/mL. The precision of the quality control samples for three different concentrations ranged from 19.1% at 20 ng/mL, 3.2% at 200 ng/mL, to 3.5% at 667 ng/mL while the accuracy was 96.3% at 20 ng/mL, 101.2% at 200 ng/mL, and 101.6% at 667 ng/mL. No system carryover was detected even when the same device was used for sequential analysis. These results suggest the potential of this microdevice for MS-based quantitative analysis in drug discovery and development.

Electrophoresis, Microchip↗

PubMed2003

Chip-based P450 drug metabolism coupled to electrospray ionization-mass spectrometry detection.

A chip-based P450 in vitro metabolism assay coupled with ESI-MS and ESI-MS/MS detection is described in this paper. The chips were made of a cyclic olefin polymer using a hot embossing process. The introduction of reagent solutions into the chip was carried out using fused-silica capillaries coupled to two syringes with the flow rate controlled by a syringe pump. Initial experiments described here employed a small commercial guard column in an off-chip format to desalt and concentrate the products of the enzymatic reaction prior to ESI-MS analysis. The system was used both to yield the Michaelis constant (K(m)) of the P450 biotransformation of imipramine into desipramine and to determine the IC50 value of a chemical inhibitor (tranylcypromine) for this CYP2C19-mediated reaction. The results demonstrated that the kinetics of the reaction inside the 4-microL volume within the channels of the cyclic olefin polymer chip provided results in agreement with those reported in the literature using conventional assays. The above reactions were carried out using human liver microsomes, and the metabolites were detected by ESI-MS showing the potential of the chip-based P450 reaction for metabolite screening studies as well as for P450 inhibition assays. A porous monolithic column was subsequently integrated into the chip to perform the reaction mixture cleanup process in an integrated fashion on the chip that is necessary for ESI-MS detection. The miniature monolithic SPE column was prepared in situ inside the chip via UV-initiated polymerization. The results obtained using the integrated system demonstrated the possibility of performing P450 enzymatic reactions in a microvolume reaction chamber coupled directly to ESI-MS detection and required less than 4 microg of HLM protein.

Alkenes↗

PubMed2002

An electrospray ionization source for integration with microfluidics.

We have demonstrated a new electrospray ionization (ESI) device incorporating a tip made from a shaped thin film, bonded to a microfluidic channel, and interfaced to a time-of-flight mass spectrometer (TOFMS). A triangular-shaped thin polymer tip was formed by lithography and etching. A microfluidic channel, 20 microm wide and 10 microm deep, was embossed in a cyclo olefin substrate using a silicon master. The triangular tip was aligned with the channel and bonded between the channel plate and a flat plate to create a microfluidic channel with a wicking tip protruding from the end. This structure aided the formation of a stable Taylor cone at the apex of the tip, forming an electrospray ionization source. This source was tested by spraying several solutions for mass spectrometric analysis. Because the components are all made by lithographic approaches with high geometrical fidelity, an integrated array system with multiple channels can be formed with the same method and ease as a single channel. We tested a multichannel system in a multiplexed manner and showed reliable operation with no significant cross contamination between closely spaced channels.

Cytochrome c Group↗