Void VolumeDefinition - What doesVoid Volumemean?Void volume is the volume of the pores or space between particles in: Ion exchanger Filter media Other granular materialThis is often expressed as a percentage of the total volume occupied by the material.Void volume refers specifically to the volume of liquid phase contained inside a column. The same term is sometimes also used informally to refer to the volume of a cavity in the column/tubing or fittings.Void volume is also known as dead volume.Corrosionpedia explainsVoid VolumeVoid volume is the volume of mobile phase (Vmor V0) in a column. In an ideal case, it is equal to the mobile phase hold-up volume. For example, if the stationary phase occupies 40% of the total column volume, the void volume would be 60% of the total column volume. Consider a column that is 25 cm long with an inner diameter of 1 cm. The total column volume is 19.6 mL. If the mobile phase occupies 60% of the column volume, the void volume is 11.8 mL.In column chromatography, the volume of the mobile phase is the total bed volume of the column minus the volume occupied by the support particles.In gel chromatography or exclusion volume, it is the volume of the mobile phase passing through the gel required to elute a molecule that never entered the stationary phase.Various methods to estimate the total void volume include: Pycnometry Minor disturbance method Tracer pulse method Hold-up volume based on unretained compounds Accessible volume based on Martin's rule and its descendantsIn post-tensioned ducts, where voids are discovered or suspected, a simple pressure test using compressed air can be used to determine the approximate void volume. It is the presence of corrosion, which may occur in the void, which is a cause for concern.Void volume is an important carbon black structure property. A profile of void volume as a function of applied pressure provides a means to assess carbon black structure at varying levels of density and aggregate reduction.

Void VolumeThe void volume is the volume of mobile phase (VmorV0) in a column. For example, if the stationary phase occupies 40% of the total column volume, the void volume would be 60% of the total column volume.Consider a column that is 25 cm long with an inner diameter of 1 cm. The total column volume is 19.6 mL (V =r2L = 3.14 * 0.52* 25 cm). If the mobile phase occupies 60% of the column volume, the void volume is 11.8 mL.

PUMP SYSTEM

In the last module you appreciated the crucial role played by a detector in the HPLC system. In this module you will be introduced to the pump which provides continuous and consistent flow of mobile phase through the HPLC system. A pump can be compared to the human heart which continuously pumps blood throughout the body but though the human heart can withstand changes in blood pressure within specified limit due to stress and strain the HPLC pump is required to deliver flow of mobile phase at constant pressure and flow rate. Changes in both these parameters can lead to errors in the results. In simple language the HPLC pump has to have ruggedness and at the same time should be able to provide reproducible flow characteristics run after run. The operational pressure limits have a vast range depending upon analysis requirements. In normal analytical operation the pressure can vary between 2000 5000 psi but in applications covered under UHPLC mode operating pressure can be as high as 15000 18000 psi.An ideal pump should have the following desirable characteristics Solvent compatibility and resistance to corrosion Constant flow delivery independent of back pressure Convenience of replacement of worn out parts Low dead volume for minimum problems on solvent changeoverThree commonly used pump types are Syringe type pumps, Constant pressure pumps and Reciprocating piston pumps.Constant pressure pumpsprovide consistent continuous flow rate through the column with the use of pressure from a gas cylinder. Valving arrangement allows rapid refill of solvent chamber. A low pressure gas source is needed to generate high liquid pressures.Syringe Type Pumpsare suitable for small bore columns. Constant flow rate is delivered to column by a motorized screw arrangement. Solvent delivery rate is set by changing voltage on the motor. These pumps deliver pulseless flow independent of column backpressure and changes in viscosity but major disadvantages are limited solvent capacity and limitation on gradient operationReciprocating Piston pumpsdeliversolvent(s) through reciprocating motion of a piston in a hydraulic chamber. On the back stroke the solvent is sucked in and gets delivered to the column in the forward stroke. Flow rates can be set by adjusting piston displacement in each stroke. Dual and triple head pistons consist of identical piston chamber units which operate at 1800or 1200phase difference. The solvent delivery of reciprocating pump systems is smooth because while one pump is in filling cycle the other is in the delivery cycle. High pressure output is possible at constant flow rate and gradient operation is possible. However, pulse dampening is required for further elimination of pressure pulses.We now have a fair understanding of different pumps and their operation principles and our next post will deal with injectors for injection of samples into the flowing mobilephase stream.

High Performance Liquid Chromatography Methods, Benefits and ApplicationsByG.P. ThomasTopics CoveredWhat is HPLC?How Does HPLC Work?Manufacturers of HPLC SystemsBenefits of HPLCApplications of HPLCReferencesWhat is HPLC?High-performance liquid chromatography or high-pressure liquid chromatography (HPLC) is a chromatographic method that is used to separate a mixture of compounds in analytical chemistry and biochemistry so as to identify, quantify or purify the individual components of the mixture.Reversed phase HPLC/UHPLC chromatography is a commonly used separation mode. It provides dynamic retention of compounds possessing hydrophobic and organic functionality. A combination of hydrophobic and van der Waals type interactions between all the target compound and both the stationary and mobile phases enables the retention of these compounds by reversed phase.How Does HPLC Work?In very small amounts, the sample mixture to be separated and tested is sent into a stream of mobile phase percolating via the column. There are different types of columns available with sorbents of varying particle sizes and surfaces.The mixture moves through the column at varying velocities and interact with the sorbent, also known as the stationary phase. The velocity of each component in the mixture depends on its chemical nature, the nature of the column and the composition of the mobile phase. The time at which a specific analyte emerges from the column is termed as its retention time. The retention time is measured under specific conditions and considered as the identifying characteristic of a given analyte.Sorbent particles might be hydrophobic or polar in nature. The commonly used mobile phases include any miscible combination of water and organic solvents such as acetonitrile and methanol. Water-free mobile phases can also be used.The aqueous component of the mobile phase might contain acids like formic, phosphoric or trifluoroacetic acid or salts to enable the separation of the sample components. The composition of the mobile phase is either maintained as a constant or as varied during the chromatographic analysis. The constant approach is effective for the separation of the sample components that are not very dissimilar in their affinity for the stationary phase. In the varied approach, the composition of the mobile phase differs from low to high eluting strength. The eluting strength of the mobile phase is reflected by analyte retention times where high eluting strength produces fast elution.The composition of the mobile phase is chosen based on the intensity of interactions between several sample components and the stationary phase.The HPLC partitioning process is quite similar to the liquid-liquid extraction process except that the former is a continuous process unlike the latter which is a step-wise process. It is recommended that trial partitioning processes be performed to determine the exact HPLC method that would provide adequate separation.Manufacturers of HPLC SystemsThere are a wide range of HPLC options in the market today. The following is a list of various HPLC system producers with a brief introduction of their products: Dionex, a company based on the science of chromatography, manufactures the UltiMate3000 Rapid Separation LC system, the UltiMate 3000 RSLCnano system, and the Coronafamily of universal charged aerosol detectors. Jasco Analytical Instruments provide a range of HPLC systems. The LC-2000 Plus Series HPLC Systems is uniquely configurable. With high performance and low cost, these HPLC systems can be adapted to virtually any requirement from simple isocratic QA to advanced multi-solvent/multi-column method development. The Isocratic LC-2000plus-Iso is provided with a pump, degasser, autosampler, and UV/Vis detector. The pump is designed with flow rates from 1L to 10mL/min at pressures up to 500 bar for use with 2, 3, 4, 6 and up to 10mm ID columns. The Binary Gradient LC-2000plus-HPG is similar to the isocratic system but is provided with a two-solvent gradient pump configuration. The pumps flow rates are compatible with 1mm columns. The Quaternary Gradient LC-2000plus-LPG is also similar to the isocratic system; however, this model is provided with a quaternary gradient pump configuration for maximum solvent flexibility. The Preparative HPLC System from Gilson has a wide flow range that can be adapted for both semi-preparative and preparative separations. Its high pumping power allows a wide range of preparative column sizes. The system is designed with a new rinse station system comprising a flowing jet wash, and allows the use of up to two different rinse solvents. Sharp HPLC Systems from AAPPTec are provided with pumps that are microprocessor controlled, thus enabling highly precise and highly accurate flow rates, which can help obtain excellent analytical reproducibility and highest measurement accuracy.Benefits of HPLCThe key benefits of HPLC systems are as follows: Controls and automates chromatography instrumentation Provides data management, security features, and reporting and instrument validation. Powerful and adaptable Increases productivity by managing all the areas of analysis - from sample to instrument, and from separation to reporting results. AffordableApplications of HPLCHPLCs can be used in the following applications: Water purification Preconcentration of trace components Ligand-exchange chromatography Ion-exchange chromatography of proteins High-pH anion-exchange chromatography of carbohydrates and oligosaccharides