Peptide drugs, metabolites

The coupling of HPLC with NMR represents a powerful method for the high-throughput screening of peptides in mixtures run in stop-flow and continuous-flow modes. It is possible to obtain routine high-quality HPLC/NMR ID NMR data with as little as 5 pg of compound in a chromatogram peak. However, the HPLC/NMR technique cannot be favorably compared to mass spectrometry techniques (HPLC/MS) in terms of sensitivity and speed of analysis. To date, the majority of reports of the use of HPLC/NMR have been for drug metabolites.1 ... 

NSI Nanospray ionization (NSI) is a low-flow (10-500-nL/min) ESI technique with many advantages over conventional-flow ESI ( 200 xL/min) for the analysis of drugs, metabolites, peptides, proteins, and other macromolecules. Advantages of NSI over ESI include decreased sample consumption and increased sensitivity (Wilm and Mann, 1996 Corkery et al., 2005). NSI can be used for LC-MS or direct-infusion MS analysis of molecules (Wickremsinhe et al., 2006 Ramanathan et al., 2007c). 

One of the first applications of HPLC in the clinical field was the quantitation of theophylline in asthmatic infants. This highly accurate measurement was an important test because of the very low amount of sample required and the accuracy of the determination (see Fig. 1-10). More recent clinically related HPLC separations include drugs and drug metabolites, neurochemicals and their metabolites, histamines, thyroid hormones, and enkephalins. The earliest bioresearch applications of HPLC included the determination of peptides, proteins, and amino acids. Application of HPLC to the analysis of these compounds remains important, as indicated by the rapid growth in references (Fig. 1-11). Bioresearch remains one of the most rapidly expanding growth areas of LC. 

Trace enrichment of polar pesticides from water Isolation of polar drug metabolites Reversed-phase application - hydrophobic phase Isolation of hydrophobic species from aqueous solution Drugs from serum, urine, and plasma Peptides in serum and plasma Normal-phase application—polar neutral phase Isolation of low to moderate polarity species from non-... 

Applications of Dns-Q range from the determination of small amounts of amines, amino adds, phenols and catechols in tissues and body fluids to the quantitative assay of drugs and drug metabolites from the evaluation of peptide sequences to the characterization of active centres of enzymes. The selected applications in Table 5 may help to identify appropriate procedures for particular analytical problems. 

IMS is commonly coupled with a Unear time-of-flight (TOP) analyzer for the molecule separation and detection of proteins and reflector TOP analyzers for peptides. Ion trap quadrupole, TOP/TOP, and quadrupole TOP analyzers are typically used for drugs, metabolite, and other low MW analytes because of their MS/MS capabilities, providing high sensitivity in the presence of a complex matrix background (58). 

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