Bioanalytical applications

The demands on the analyst in the pharmaceutical environment are somewhat different in the drug discovery 

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Other bioanalytical applications of systems in which the eluate of a first LC column is sampled in continuous and repetitive intervals and subjected to a second LC dimension are, for example, described by Wheatly et a/. (11) and Matsuoka et al. (12). Wheatly coupled gradient affinity LC with RPLC for the determination of the isoenzymatic- and subunit composition of glutathione 5-transferses in cytosol... 

The first bioanalytical application of LC-GC was presented by Grob et al. (119). These authors proposed this coupled system for the determination of diethylstilbe-strol in urine as a replacement for GC-MS. After hydrolysis, clean-up by solid-phase extraction and derivatization by pentafluorobenzyl bromide, the extract was separated with normal-phase LC by using cyclohexane/1 % tetrahydrofuran (THE) at a flow-rate of 260 p.l/min as the mobile phase. The result of LC-UV analysis of a urine sample and GC with electron-capture detection (ECD) of the LC fraction are shown in Ligures 11.8(a) and (b), respectively. The practical detection limits varied between about 0.1 and 0.3 ppb, depending on the urine being analysed. By use of... 

Presently, the on-line coupling of NPLC and GC via heart-cutting is an established procedure which has been used successfully for several bioanalytical applications. Obviously, dfrect analysis of aqueous samples is not possible by NPLC, and therefore, a solvent switch by a sample pretreatment step (e.g. liquid-liquid extraction or SPE) is always requfred when biological samples are analysed by NPLC-GC. 

T. A. G. Noctor, Bioanalytical applications of enantioselective high-performance liquid cliromatography in A Practical Approach to Chiral Separations by Liquid Chromatography, Subramanian G (Ed.), VCH, Weinheim, Ch. 12, pp. 357-396 (1994). 

Particularly attractive for numerous bioanalytical applications are colloidal metal (e.g., gold) and semiconductor quantum dot nanoparticles. The conductivity and catalytic properties of such systems have been employed for developing electrochemical gas sensors, electrochemical sensors based on molecular- or polymer-functionalized nanoparticle sensing interfaces, and for the construction of different biosensors including enzyme-based electrodes, immunosensors, and DNA sensors. Advances in the application of molecular and biomolecular functionalized metal, semiconductor, and magnetic particles for electroanalytical and bio-electroanalytical applications have been reviewed by Katz et al. [142]. 

The focus of the examples given in this chapter is clearly on micro reactors for chemical processing in contrast to p-TAS or Lab-Chip systems for bioanalytical applications. In the latter microfluidic systems, the fluidic requirements are somehow different from those in micro reactors. Typically, throughput plays only a minor role in p-TAS systems, in contrast to micro reactors, where often the goal is to achieve a maximum molar flux per unit volume of a specific product. Moreover, flow control plays a much greater role in p-TAS systems than in micro reactors. In... 

Kennedy R.T., Bioanalytical applications of fast capillary electrophoresis, Anal. Chim. Acta, 400, 163, 1999. 

Agiamamioti and co-workers [72] synthesized a novel biotinylated fluorophore, 10-(2-biotinyloxyethyl)-9-acridone 25 with favorable properties for bioanalytical applications. In aqueous solutions, it displayed high fluorescence ([Pg.37]

Optical read out will compete with micro electrode arrays. New developments in array detectors will open new perspectives. Direct optical detection techniques will add new possibilities to bioanalytical applications in addition to fluorescence measurements presently preferred in biochips read out. 

Hemmila I, Stahlberg T, Mottram P (1995) Bioanalytical applications of labelling technologies, 2nd edn. Wallac Oy, Turku, Finland... 

Bright, F.V. (1988) Bioanalytical applications of fluorescence spectroscopy. Anal. Ghent. 60,1031A-1039A. 

Another important bioanalytical application of voltammetric ISEs is the detection of polyions (see also above). A technique using cyclic voltammetry on micropipette electrodes filled with the organic electrolyte solutions in 1,2-dichloroethane was successfully applied for the detection of protamine [65] in saline solution and heparin in undiluted sheep plasma samples [66]. Protamine transport was facilitated with dino-nylnaphthalenesulfonic acid (DNNS). As a heparin-selective component the tetrakis-(4-chlorophenyl)borate salt of trimethyloctadecyl ammonium was used. 

J.H. Yun, V.C. Yang, and M.E. Meyerhoff, Protamine-sensitive polymer membrane electrode characterization and bioanalytical applications. Anal. Biochem. 224, 212-220 (1995). 

The applications of nanoparticles in biosensors can be classified into two categories according to their functions (1) nanoparticle-modified transducers for bioanalytical applications and (2) biomolecule-nanoparticle conjugates as labels for biosensing and bioassays. We intend to review some of the major advances and milestones in biosensor development based upon nanoparticle labels and their roles in biosensors and bioassays for nucleic acids and proteins. Moreover, we focus on some of the key fundamental properties of certain nanoparticles that make them ideal for different biosensing applications. 

For certain bioanalytical applications such as those described in the previous section, it is desirable to collect analytes of interest while separation in the tPLC system takes place (e.g., for follow-up by MS). To collect the analytes, the tPLC system may be equipped with a time-triggered fraction collection mechanism. This system allows the selection of time intervals in which samples will be diverted (after passing through the detectors) to the selected wells of microtiter plates. Figure 6.11 shows an overall view of a system with time-triggered fraction collection capabilities. Figure 6.12 depicts the system along with a detailed view of the collection sampler employed. 

These systems have been used in many bioanalytical applications. A Prospekt system coupled with MS quantitated eserine N-oxide, a cholinesterase inhibitor, in human plasma for low level (4.5 mg) oral administration pharmacokinetic studies (Pruvost et al. 2000). After conditioning of the SPE cartridge (PLRP-S, Spark) with methanol (5 mL/min, 0.5 min) and water (5 mL/min, 0.5 min), a volume of 250 jj.L plasma plus internal standard was injected and washed (water, 1 mL/min, 3 min). The analytes were flushed out with 80 20 ammonium acetate (20 mM, pH 3.5 adjusted with formic acid) and acetonitrile (0.3 mL/min) and separated on a Zobax SB-CN column (150 x 2.1 mm inner diameter, 5 jim). A calibration range of 25 pg/mL to 12.5 ng/mL was achieved with a run time of 10.5 min. 

Online SPE LC/MS/MS is commonly used for bioanalytical applications in the pharmaceutical industry. Column switching systems and TFC systems are easy to build and control. Sophisticated commercial systems and SPE cartridges are readily available. Compared to offline sample preparation, the online approach can save time and labor. However, the development of online SPE bioanalytical assays remains analyte-dependent. Generic methods can be applied to many analytes. For extremely hydrophobic, hydrophilic, and ionic analytes at normal pH range and analytes with a variety of hydrophobicity and pKa values, analyte-specific methods must be developed. An understanding of the chemistry of the analytes and SPE is critical. 

Mizukami S, Nagano T, Urano Y et al (2002) A fluorescent anion sensor that works in neutral aqueous solution for bioanalytical application. J Am Chem Soc 124 3920-3925... 

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