High-performance liquid chromatography data systems

The LC system consisted of a Jasco 880 PU pump (JASCO, Tokyo, Japan) for the mobile phase, a Rheodyne Model 7125 injector with a 25 pL loop, and a 0.2 x 15 cm Mightysil RP-18 GP (5 pm) column (Kanto Chemical, Tokyo, Japan). LC was performed using an aqueous solution containing 80% MeCN as the mobile phase at a flow rate of 0.2 ml/min at 26°C. The column was connected to L-7455 diode array detector (Hitachi, Tokyo, Japan) with a flow cell (17.7 pL, 10 mm), and Hitachi D-7000 advanced high performance liquid chromatography (HPLC) system manager was utilized for data acquisition. 

These incubations are often carried out at 37 °C for 1-2 h. At different time points, 20-200 /aL of incubation mixture is withdrawn from each incubation and mixed with equal volume of ice-cold acetonitrile by vortexing. For preparation of acyl glucuronide, ice-cold acetonitrile containing 1% of formic acid is used to minimize acyl-migration [3,14]. After centrifugation at 13 000 rpm for 5-15 min, the supernatant (10-30 /aU) is analyzed by high-performance liquid chromatography (HPUC)-UV-MS. The metabolite of interest is identified based on HPLC retention time, UV spectrum and MS/MS data. Conversion yield is estimated based on UV absorption peak areas. A suitable in vitro enzyme system for scale-up is then... 

High-performance liquid chromatography (HPLC) is one of the premier analytical techniques widely used in analytical laboratories. Numerous analytical HPLC analyses have been developed for pharmaceutical, chemical, food, cosmetic, and environmental applications. The popularity of HPLC analysis can be attributed to its powerful combination of separation and quantitation capabilities. HPLC instrumentation has reached a state of maturity. The majority of vendors can provide very sophisticated and highly automated systems to meet users needs. To provide a high level of assurance that the data generated from the HPLC analysis are reliable, the performance of the HPLC system should be monitored at regular intervals. In this chapter some of the key performance attributes for a typical HPLC system (consisting of a quaternary pump, an autoinjector, a UV-Vis detector, and a temperature-controlled column compartment) are discussed [1-8]. 

The following table provides some comparative data for the selection and operation of the more common detectors applied to high-performance liquid chromatography.15 In general, the operational parameters provided are for optimized systems and represent the maximum obtainable in terms of sensitivity and linearity. In this table, the molar extinction coefficient is represented by s. 

The resolution of a multicomponent system involves the description of the variation of measurements as an additive model of the contributions of their pure constituents [1-10]. To do so, relevant and sufficiently informative experimental data are needed. These data can be obtained by analyzing a sample with a hyphenated technique (e.g., HPLC-DAD [diode array detection], high-performance liquid chromatography-DAD) or by monitoring a process in a multivariate fashion. In these and similar examples, all of the measurements performed can be organized in a table or data matrix where one direction (the elution or the process direction) is related to the compositional variation of the system, and the other direction refers to the variation in the response collected. The existence of these two directions of variation helps to differentiate among components (Figure 11.1). 

Based on the quantity of data and the complexity of the process, an enterprise operational informatics system must be robust and scalable. It should also integrate with analytical instrumentation such as high performance liquid chromatography (HPLC) MS to verify compound identity. A recent review of HTS informatics provides a compact overview of the key informatics elements of HTS operations and data handling (Ling, 2008). 

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