Solvent reservoir. HPLC

One of the major practical problems to the installation of HPLC as a permanent process monitor is the need to replace solvent. A large solvent reservoir may present problems both in terms of size and safety. One solution is the use of packed capillary columns, which consume much less solvent than conventional columns, as the comparison (at constant linear velocity) in Table 1 shows. 

A modern solvent delivery system consists of one or more pumps, solvent reservoirs, and a degassing system. HPLC pumps can be categorized in several ways by flow range, driving mechanism, or blending method. A typical analytical pump has a flow range of 0.001-10 mL/min, which handles comfortably the flow rates required for most analytical work (e.g., 0.5-3 mL/min). Preparative pumps can have a flow range from 30 mL/min up to L/m. 

As the name implies, capillary electrophoresis is electrophoresis that is made to occur inside a piece (50 to 100 cm) of small-diameter capillary tubing, similar to the tubing used for capillary GC columns. The tubing contains the electrolyte medium, and the ends of the tube are dipped into solvent reservoirs, as is the paper in paper electrophoresis. Electrodes in these reservoirs create the potential difference across the capillary tube. An electronic detector, such as those described for HPLC (Chapter 13), is on-line and allows detection and quantitative analysis of mixture components. 

Detail the path of the mobile phase through the HPLC system from the solvent reservoir to the waste receptacle, giving brief explanatory descriptions of instrument components along the way. 

High Performance Liquid Chromatography (HPLC) (Chapter 30) gives an elaborate discussion of theoretical aspects. Instrumentation encompasses the various important components e.g., solvent reservoir and degassing system pressure, flow and temperature pumps and sample injection system ... 

There are typically six components in an HPLC system (1) solvent reservoirs (2) a pumping or solvent management system (3) an injector, which can be either manual or automated (4) a column (5) a detector (6) a data recorder, which can be an integrator or a computer system. 

A schematic diagram of a typical high-pressure liquid chromatograph is shown in Figure 3.12. The basic components are a solvent reservoir, high-pressure pump, packed column, detector, and recorder. A computer is used to control the process and to collect and analyze data. The similarities between a gas chromatograph and an HPLC are obvious. The tank of carrier gas in GC is replaced by the solvent reservoir and high-pressure pump in HPLC. 

An HPLC chromatograph was used together with a mass detector (when required, a stream splitter (app. 10 1) was inserted between the column and the detector). A column (250 X 4.6-mm ID) of NUCLEOSIL 5SA was flushed with 1% aqueous ammonium nitrate solution at a flow rate of 0.5 ml/min for 1 h, then with distilled water at 1 ml/min for 1 h. Silver nitrate (0.2 g) in water (1 ml) was injected onto the column in 50-/zl aliquots at 1-min intervals silver began to elute from the column after about 10 min. Twenty minutes after the last injection, the column was washed with methanol for 1 h, then with 1,2-dichloroethane-dichloromethane (1 1) for another hour. The three solvent reservoirs contained the following (A) 1,2-dichloroethane-dichloromethane (1 1) (B) acetone and (C) acetone-acetonitrile (9 1). For linoleic acid-rich seed oils, gradients of A were employed to 50% A-50% B over 15 min, then to 50% B-50% C over a further 25 min and held there for 5 min. For linolenic acid-rich seed oils, C was changed to acetone-acetonitrile (4 1), and the flow rate was increased to 1 ml/min gradients of A were utilized to 50% A-50% B over 10 min, then to 70% B-30% C over 20 more min, and finally to 100% C over another 30 min. 

The first step in the wetted path in the HPLC is the solvent reservoir holding freshly filtered (and possibly degassed or deoxygenated) solvent. Most systems use a porous fritted stone (5-30-/an filter) as a solvent line sinker. The tubing to the solvent inlet is wide-diameter Teflon . In the solvent inlet line we may have another frit and a sapphire ball/stainless steel check valve. The wetted surfaces in the pumping chamber are all stainless steel except for the plunger... 

Check your HPLC system manual to make sure the system is compatible with nitric acid washing. Make sure the reservoir sinker is made of stainless steel and not monel metal. Replace the water in the solvent reservoir with 20% nitric acid. Stop Note Make sure the column has been replaced with the column bridge. Do not pump nitric acid through a bonded-phase column. Wash the system for 15 min at 2 mL/min with 6 N (20%) nitric acid. Discard the wash carefully. 

An HPLC system, shown schematically in Figure 2.1, consists of a solvent reservoir, which contains the eluent or mobile phse a pump, often called a solvent delivery system an injector through which the sample is introduced into the system without a drop in pressure or change in flow rate the analytical column, which is usually stainless steel and contains the solid packing or stationary phase and a suitable detector to monitor the eluent. 

The basic equipment required for an HPLC system includes a solvent reservoir, a pump, an injector, an analytical column, a detector, and a recorder. The analysis of the sample is displayed as a chromatogram, with detector deflection presented usually as a function of time after loading the sample. By virtue of the shape of the curves, the distance between them, and their... 

Solvent Reservoirs. Storage of sufficient amount of HPLC solvents for continuous operation of the system. Could be equipped with an online degassing system and special filters to isolate the solvent from the influence of the environment. 

Figure 4-16. Block diagram of HPLC equipment. A typical HPLC set-up consists of a column linked via a pump and gradient former to the solvent reservoirs. The sample application port is located in a separate spur. The column is...

Chandler and McNair (Hercules Inc.) [278] discussed the requirements for the basic parts of a high pressure liquid chromatographic system. These include a solvent reservoir, pump, injection system, column detector and recorder. They listed some of the different models available and described their major features. The developments in HPLC apparatus have also been reviewed by Martin and Guiochon [279] and the basic requirements also described by Bombaugh (Chromatec Inc.) [280]. The pumps and injectors available have been reviewed [281]. Some detectors currently used in HPLC have also been reviewed [282]. 

Many manufacturers have used the capabilities of on-board processing power to provide a user friendly interface for the operator. This results in a host of useful (and some not so useful) features for the analyst, and allows the pump to be remotely controlled by a computer, or another component of the HPLC system. This can facilitate method development for instance, as the pump can be programmed to change automatically the composition of the mobile phase after a number of runs. It does this by changing the proportion of time that it draws from each of a number (up to four) of solvent reservoirs, that can contain diflferent modifiers or buffers. Computerisation of pumps can also allow unattended runs to be performed with a greatly increased measure of confidence. In a networked... 

A modification of this system, which is depicted in Fig. 6.17, has been used for the selective hydrogenation of a number of substrates. 4 This is composed of a stainless steel high pressure bomb, A, which is used as the solvent reservoir, an HPLC pump, B, a stainless steel reactor tube with temperature control capabilities, C, to hold the catalyst and flow restrictor back pressure regulators, D, at the end of the flow system. This is a liquid phase flow system in which the only hydrogen available for reaction is that which is dissolved in the solvent in the reservoir. 

Gradient elution in HPLC is achieved using two pumps, two different solvent reservoirs, and a solvent mixer. In capillary electrophoresis (CE), electro-osmotic flow controls the flow of the mobile phase, which is, in most cases, an aqueous buffer and is used in place of a mechanical pump. 

The HPLC pump is operated in either an isocratic or gradient mode (Figure 6-16). In the isocratic mode, the mobile phase composition remains constant throughout the chromatographic run. This mode is usually used for simpler separations and separations of those compounds with similar structures and/or retention times. An isocratic mobile phase is a single solvent (e.g., methanol) or a prepared mixture of several solvents (e.g., methanol, acetonitrile, and water) delivered from a single solvent reservoir. Alternatively a multisolvent mobile phase can be metered and proportioned from two or more reservoirs. Most HPLC separations are performed under isocratic conditions. 

It is most convenient if a HPLC system with two solvent reservoirs can be used. Use water and acetonitrile both solvents need to be filtered (filter with < 1 p,m pores) and degassed. Flush the system with pure acetonitrile, then connect a so-called reversed-phase column (octadecyl ODS or Cig, but an octyl or Cg column can be used as well)... 

Figure 1.3 Schematic diagram of an HPLC unit. 1 = Solvent reservoir 2 = transfer line with frit 3 = pump (with manometer) 4 = sample injection 5 = column (with thermostat) 6 — detector 7 = waste 8 = data acquisition.

The equipment necessary for HPLC analysis includes column, pump, solvent reservoirs, a gradient former, and a suitable detector. It is beyond the scope of this review to cover the rapid, recent developments of HPLC instrumentation. 

A modern HPLC solvent delivery system consists of one or more pumps, solvent reservoirs, and a degassing system. Typical requirements of an analytical HPLC pumps are ... 

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