Split injection/flow system

FIGURE 16.5 Schematic of instrumental setup for 2D micro-RPLC-CZE. A split injection/ flow system is used to deliver a nanoliter per second flow rate to the micro-RP-HPLC column from the gradient LC pump. The HPLC microcolumn has 50 pm i.d. and 76 cm length, and the electrophoresis capillary has 17 pm i.d., L — 25 cm, and/= 15 cm. The valve is air-actuated and controls the flow of flush buffer (reprinted with permission from Analytical Chemistry). 

Two-phase flow in parallel pipes, fed from a common manifold, displays interesting phenomena, as two phases may split unevenly when entering the parallel piping. Ozawa et al. (1979, 1989) performed experimental smdies on two-phase flow systems in parallel pipes of 3.1 mm diameter. They simulated the flow in boiling channels by injection of air and water into the pipes. 

Thus, the column diameters chosen for the two dimensions are determined by the amount of sample available and will dictate the flow rate ranges available to use. In split-flow systems, where only a portion of the first-dimension effluent is injected into the second dimension, the choice of column size is unlimited and the two methods can be developed independently. In comprehensive systems where the entire sample from the first dimension is injected into the second dimension, the flow rates are generally lower in the first dimension to accommodate the lower injection volumes into the second dimension. For example, for a 1-mm ID column in the first dimension with a flow rate of 50 (tL/min and a sampling rate of 1 min, 50 pL could be injected onto the second dimension. A 50-(lL injection onto a4.6-mm ID column flowing at 1 mL/min should be accommodated fairly well based upon its composition. In Chapter 6, the first dimension column diameters are estimated based upon the injection volume and sampling rate into the second dimension. 

In both systems the flow of helium carrier gas through the columns was 0.7-0.8 ml min-1, with a septum purge of 0.5 ml min-1 and a split valve flow of 4-4.5 ml min-1. The injection ports were maintained at 260°C and the detector ovens at 240° C. The detector employed was either a flame ionisation or a nitrogen-specific NPD-40 thermionic detector (Erba Science (UK) Ltd) and the output was recorded on a HP 3390 integrator (Hewlett Packard Ltd, Wokingham, UK). 

One of the key developments in the development of thermal desorption devices was the possibility for cryofocusing systems that have the advantage of injection-like samples. A short section of capillary tubing at liquid nitrogen temperatures (i.e., -160°C) traps the volatiles. When capillary columns replaced packed columns as the standard, complete flow from the desorption trap (5 ml/min minimum) to the capillary columns ( 1 ml/min) was possible through the use of cryofocusing. The split injection interface was another development that splits the flow so that only a part of the desorbed volatiles entered the column. While this allowed the need for cryofocusing to be circumvented, sensitivity was lost due to the split. 

Ideally, a sample is introduced into a chromatograph as a perfect plug. In practice, this is not the case, and diffusion occurs because of the injector. For narrow-bore and microbore applications, injectors capable of introducing the required sample volumes are commercially available and optimized to reduce dispersion. This is not the case for capillary LC, and homemade injection systems include the sample tube technique, in-column injection, stopped-flow injection, pressure pulse-driven stopped-flow injection (PSI), groove injection, split injection, heart-cut injection, and the moving injection technique (MIT). Of the injection techniques, only the split injector, MIT and PSI approaches can introduce subnanoliter sample volumes accu-... 

Yao et al. reported a flow injection analytical system for the simultaneous determination of acetylcholine and choline that made use of immobilized enzyme reactors and enzyme electrodes [25]. Acetylcholineesterase-choline oxidase and choline oxidase were separately immobilized by reaction with glutaraldehyde onto alkylamino-bonded silica, and incorporated in parallel as the enzyme reactors in a flow injection system. The sample containing acetylcholine and choline in 0.1 M phosphate buffer (pH 8.3) carrier solution was injected into the system. The flow was split to pass through the two reactors, recombined, and mixed with 0.3 mM K4Fe(CN)6 reagent solution before reaching a peroxidase immobilized electrode. Because each channel had a different residence time, two peaks were obtained for choline and total acetylcholine and choline. Response was linear for 5 pM-0.5 mM choline, and for 5 pM 1 mM acetylcholine plus choline. The detection limits were 0.4 pM for choline and 2 pM for acetylcholine. 

Figure 6-11 Flow diagram of a GC inlet system for split injection.The head pressure and total flow are adjusted to achieve a desired flow rate in the column and a fractional spilt between the column and the spilt vent. GC, Gas chromatography.

Many specialized injection systems are used with the GC in the modern laboratory to deliver the complete sample to the column without alteration. The conventional injection method is simply to inject a small volume (about 1 /d) of a dilution of the sample using a syringe with a fine needle, which pierces a silicon rubber septum and delivers the sample into a heated chamber where it is vaporized and carried on to the column in the stream of carrier gas. Some of the carrier gas containing vaporized sample may be split from the main column flow and vented to reduce the amount of sample delivered to the column. This is known as a split injection and is suitable for almost all liquid or... 

S.M.B. Brienza, E.A.G. Zagatto, O.M. Matsumoto, A simplified split zone flow injection system, Lab. Robot. Autom. 3 (1991) 115. 

The samples were injected into a Varian gas chromatograph (CP-3380 model), equipped with a flame ionization detector (FID) and a CP WAX 52 CB capillary column 30 mxO.25 mm X 0.25 xm, and a split injection system with a 1 20 ratio. Injector and detector temperatures were kept at 250°C. The oven was initially maintained at 200°C for 4.5 min, then heated up to 210°C, and kept constant at this temperature for 0.5 min. After that, it was heated to 220°C for 0.5 min. The oven was heated again to 250°C and maintained at this temperature for 1.5 min. Hydrogen was used as the carrier gas at a 1.8 ml/min flow rate column pressure was set at 12 psi. A computer loaded with the Star Workstation 6.2 software was connected to the GC by a Star 800 Module Interface to automatically integrate the peaks obtained. Methyl heptadecanoate was the internal standard used. 

Small diameter capillary columns require very small samples, often only a fraction of microgram in size and as this sample size is too small for practical injection syringes, a split-flow system must be used. In effect, the sample is vaporized into a gas stream and a fraction of the gas stream (and consequently a fraction of the sample) is allowed to pass through the column. A diagram of a split-flow injector is shown in figure 4.4. 

The split-flow injector is very similar to the packed column injector except that only part of the carrier flow passes to the column, the rest exits to waste. By varying the exit flow-impedance, the split-ratio can be adjusted over a wide range. Without this type of split injection system, the small bore capillary columns would be virtually impossible to use. However, because of the waste of sample and the relatively small mass range obtainable from small bore columns, the large bore capillary column was introduced. 

A similar approach was used by Ferreira et al. (1996) for the simultaneous assay of nitrite and nitrate in meat products. The system was based on splitting the flow after injection and the subsequent confluence of it before reaching the detector, allowing the reduction of nitrate to nitrite online in part of the sample plug. Each channel had a different residence time, therefore separate peaks where obtained for nitrite and nitrite + nitrate. The same system was later used by Pinho et al. (1998) for the evaluation of nitrite and nitrate contents in 15 different brands of pate. 

Split injection is used for volatile to semi-volatile compounds, and is one of the easiest injection techniques. With this technique, the flow of carrier gas is split between the capillary column and the atmosphere. This split does not occur in case of splitless injection. Splitless injection is used in case residual solvents remain in the sample at low eon-centrations, due to increased sensitivity compared to split injection. Other possibilities of direct injection are on-column injection and Programmed Temperature Vaporisation (PTV) injection. Both techniques allow detection limits to achieve put levels but are large volxune injection techniques. In on-column injection systems, the sample is injected on a pre-colmnn and then the solvent is vented, leaving only the analytes to be injected on the coltunn. In PTV, after the sample injection, the solvent is evaporated at a low temperature in a packed chamber and then removed. This leaves the solutes on the packing. When the injection port is heated the analytes are transferred to the analytical column. ... 

Principles and Characteristics As mentioned already (Section 3.5.2) solid-phase microextraction involves the use of a micro-fibre which is exposed to the analyte(s) for a prespecified time. GC-MS is an ideal detector after SPME extraction/injection for both qualitative and quantitative analysis. For SPME-GC analysis, the fibre is forced into the chromatography capillary injector, where the entire extraction is desorbed. A high linear flow-rate of the carrier gas along the fibre is essential to ensure complete desorption of the analytes. Because no solvent is injected, and the analytes are rapidly desorbed on to the column, minimum detection limits are improved and resolution is maintained. Online coupling of conventional fibre-based SPME coupled with GC is now becoming routine. Automated SPME takes the sample directly from bottle to gas chromatograph. Split/splitless, on-column and PTV injection are compatible with SPME. SPME can also be used very effectively for sample introduction to fast GC systems, provided that a dedicated injector is used for this purpose [69,70],... 

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