Cell volume, detector, narrow-bore

Although somewhat specialized, integrated flow-cells are normally commercially available. Ideally, they should be short (0.2- 1.5 mm) and narrow-bore in order to avoid problems arising from inadequate detector capacity and sensitivity, respectively. Ideally, they should also have small inner volumes in order to boost sensitivity and sample throughput. 

Narrow-bore columns of between 1.0 and 2.5 mm ID are available for use in specially designed liquid chromatographs having an extremely low extracolumn dispersion. For a concentration-sensitive detector such as the absorbance detector, the signal is proportional to the instantaneous concentration of the analytes in the flow cell. Peaks elute from narrow-bore columns in much smaller volumes compared to those from standard-bore columns. Consequently, because of the higher analyte concentrations in the flow cell, the use of narrow-bore columns enhances detector sensitivity. The minimum detectable mass is directly proportional to the square of the column radius (107) therefore, in theory, a 2.1-mm-ID column will provide a mass sensitivity about five times greater than that of a 4.6-mm-ID column of the same length. 

Because of the interest in narrow-bore and microbore columns, instrument manufacturers now have developed solvent delivery systems that are capable of accurately pumping at the low flow rates typically required for microbore applications (>10 / L/min). In addition, injectors have been designed that are capable of introducing the smaller sample volumes, and detector cells are available that are small enough to monitor the reduced sample volumes passing through the detector. Thus narrow-bore and microbore applications are possible with readily available instrumentation, and reports may be found in the literature. 

Because of the small elution peak volume obtained from narrow-bore packed columns, conventional concentration dependent detectors such as UV-vlslble absorbance, fluorometrlc, and electrochemical detectors must be purged with makeup solvent or miniaturized to allow minimum extra-column contribution to peak spreading. A cell volume 0.1 i Is desirable for narrow-bore packed columns of I.D. Ishll et. al. reported the reduction of an UV detector cell volume to OAiiZ by using a quartz tube of... 

With narrow bore columns the flow-cell volume must be < 3 pi in order that the resolution achieved on the column is not lost due to analyte spreading in the detector cell. 

HPLC sample flow-cells have internal volumes of less than 20 pi, with a typical path length of 1mm and window area of 2-3 mm (Figure 7.13c). NaCl or KBr window materials may be used with many organic solvents and PTFE and polyethylene windows are available for both organic and aqueous based solvents. Narrow bore or microcolumns (2-3 mm i.d.) have lower flow volumes than normal columns, typically 0.3-0.5mlmin and are therefore ideally suited for infrared detectors. Micro-HPLC-FTIR techniques may employ a direct flow cell or solvent elimination techniques [16 18]. Considerable care is required to match the chromatographic system with the sample cell to avoid loss of resolution. 

LC-NMR One information-rich spectral technique that is more suited to the liquid mobile phase of HPLC than to the vapor phase of GC is NMR. LC-NMR has been implemented, but it has significant limitations. To obtain interpretable spectra of unknowns, concentrations in the measurement cell must be higher than with other detectors. The cell must be smaller than the usual NMR tube, so for any but the very highest concentrations of analytes, FT-NMR acquisition is preferred, with each eluted peak being retained in the measurement cell by a stopped-flow procedure similar to that employed to increase sensitivity in GC-IR (Section 12.8.2). Expensive deuterated mobile phase solvents are required for proton NMR, which mandates the use of low mobile phase volume flow columns narrow bore or even capillary HPLC. LC-NMR is expensive to implement and not readily available from commercial vendors at this time. 

The flow analyzer involves simple apparatus such as samplers, liquid drivers (peristaltic pumps, piston pumps, solenoid pumps), injection devices (rotary valves, injector-commutators), reactors and flow lines (usually narrow bore tubing), mixing chambers, and flow-through detectors. As a rule, these devices are readily available in most laboratories devoted to chemical analysis. Regarding detection, almost all analytical techniques have been used in flow analysis a small flow cell volume and a short response time that is compatible with system dynamics are important detector parameters. 

With the next example shown in Figure 2.20, the speed-up of a gradient separation performed on modern UHPLC-type columns and instrumentation is discussed. The attempted use of a narrow bore (2.1 mm) column packed with 2.2 pm particles requires an instrument with significantly reduced extra-colunm volume. This also includes the adaptation of the flow cell volume and the requirement of faster data handhng by the detector electronics (which was already a prerequisite in the example in Figure 2.19). Moreover, the instrument must be able to handle precise injection of 2 pi sample volume and withstand a column pressure of almost 600 bar. 

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