Flow splitters

S ample loop volumes The volumes of the sample loops that store eluent from the first dimension and inject eluent into the second-dimension column system must be determined. The loop volume divided by the second-dimension elution time range determines the first-dimension flow rate in comprehensive 2DLC. If the dilution factor is small in the second column, a flow splitter can maintain a small loop volume even with a substantial flow rate from the first-dimension column. 

NaCl or KC1 (Peng et al., 2003 Ballif et al., 2004 Beausoleil et al., 2004 Wilmarth et al., 2004 DeSouza et al., 2005 Vitali et al., 2005) may be used for the SCX fractionation, in spite of the incompatibility of these salts with mass spectrometers. When using KC1, for example, the sample must be desalted off-line (Ballif et al., 2004 Beausoleil et al., 2004), on the RP column before MS/MS acquisition (DeSouza et al., 2005 Vitali et al., 2005), with a vented column (Peng et al., 2003), or with a RP-trap (Vollmer et al., 2004 Wilmarth et al., 2004). The configuration with a RP-trap is shown in Fig. 11.1, and in this case, a flow splitter is used to reduce the flow rate from hundreds of microliters per minute to hundreds of nanoliters per minute. However, HPLC pumps of lower flow rate are now available and could eliminate the need for a flow splitter. 

Maximum flow-rates compatible with DLI interfaces are in the range of 50 to 100 pL/min. Microbore columns (<1.0mm i.d. column) operating at 5 to lOOpL/min are ideally suited for DLI LC/MS. A flow splitter is required to couple conventional LC with a DLI interface so that only a fraction of the total eluent is introduced into the mass spectrometer. Splitting the flow outside the mass spectrometer results in loss of sensitivity, an undesirable consequence. Henion and co-workers have reported an approach in which the splitter is incorporated into the desolvation chamber of the mass spectrometer. The removal of solvent is achieved by diverting the vapor generated by the solvent without loss of sample and, therefore, sensitivity. Excess pressure inside the mass spectrometer is therefore avoided while higher flow-rates can be accommodated. 

There are many variations on setting up the inlets to the MUX interface, depending on the needs of the laboratory. In one setup, one pump is used to deliver the flow through a flow splitter and multiple probe injector to four LC columns.The flow from each column is then fed into the MUX interface. In another variation, on-line SPE is coupled with LC. One pump, followed by a flow splitter, and one four-injector autosampler are used to feed samples into four extraction columns. A second pump is used (again with a flow splitter) to run gradients on the four LC columns into the MUX interface. In yet another system, an eight-channel UV MUX system is utilized. ... 

Flow splitters are used to divide the flow of total fluids to several desired treating vessels They are sometimes provided with coalescing and knockout sections. Here again, excessive agitation can be detrimental resulting in water carry-over. Corrosion of the weir boxes will result in excessive carry-over... 

A reversed-phase HPLC Cl8 or C30 narrow-bore column is typically used for LC/MS with APCI. Details about chromatography columns used for carotenoids are contained in unit F2.3. For most APCI systems, the optimum flow rate into a mass spectrometer or tandem mass spectrometer equipped with APCI, as controlled by a syringe pump or HPLC pump, is usually between 100 and 300 pl/min, which is ideal for narrow-bore HPLC columns. Larger diameter columns should be used with a flow splitter postcolumn to reduce the solvent flow into the mass spectrometer. For example, if a 4.6 mm i.d. column was used at a flow rate of 1.0 ml/min, then the flow must be split postcolumn 5 1 so that only 200 pl/min enters the mass spectrometer. 

Due to the high sensitivity it is favorably to couple a nanoHPLC to an ESI-source. As mass spectrometers are concentration dependent detectors, the sensitivity of an instrumental setup is mostly determined by the peptide concentration of the eluate but not by the peptide amount. Thus a nanocolumn with a flow rate of 300 nL/min provides an about thousand times higher sensitivity than a microbore column with a flow rate of 300 (xL/min. As an alternative to buying a nanoHPLC system it is also possible to use a relatively inexpensive flow splitter after the pump and before the injection valve and the column. Thereby the flow rate can be reduced to use a capillary column (flow rate 4 (xL/min) on an analytical HPLC system or a nanocolumn on a capillary HPLC system. Instead of a flow-splitter it is preferred to couple a nanoHPLC to an ESI-source. Thereby, the flow rate is split according to the column backpressure, i.e., mostly the column volumes if the same packing materials are used. However, these low-cost setups are less reliable than a nanoHPLC and the reproducibility is worse. 

The upper position allowed isocratic and gradient micro-LC with the possibility to assist the separation with an electric field. The capillary was introduced into the electrode and the solvent was delivered to the capillary via the inner channel. The solvent overflow was liberated through the outlet capillary, which had a higher flow resistance in this case. In this position, the special vial functions as a flow splitter. 

Splitters and Separators Flow Splitter Substream Splitter Component Splitter Two Product Separator Substream Attribute Separator... 

For optimum separation efficiency, reflux holdup should be minimized by eliminating surge drums and using flow splitters that retain little or no liquid. 

Ultraviolet (UV) spectroscopy, mass spectrometry (MS), refractive index (RI) detection, and electrochemical detection (ECD) are common online monitoring techniques for analytical chromatography. UV and RI are regularly used for monitoring preparative operations as well. To employ MS or ECD in a high-flow scheme, usually a side stream must be diverted from the main eluate line via a flow splitter so that what passes through the detector has a flow rate that is acceptable for an analytical-scale system. 

Besides using a lot of mobile phase, analyses run at these high flow rates are not compatible with an MS detector without some sort of split to divert flow. For many LC analyses, flow splitters are a fact of life when using MS detection. However, Fig. 2B illustrates that separation of the chromatographic test sample can be further scaled to a 2.1 -mm i.d. column, now running at an equivalent linear velocity (1.0 mL/min). Under the conditions used in... 

The system as diagrammed in Fig. 14 used two-column regeneration to improve throughput. The system has both preparative and analytical capability so that samples can be run on the same system for rapid screening of original samples, or an initial purity assessment, as well as fraction collection capability. In addition to the two 6-port column-switching valves, two flow splitters... 

FIGURE 1.4 Schematic of a typical nanoscale LC-MS interface system. A precolumn flow splitter is used to reduce to flow rate of a conventional pumping system to the 100-500 nL/min. The voltage for ESI is applied on the high-pressure distal side of the column using a junction electrical contact. 

---