HPLC Terms(转载)

Activity

In adsorption chromatography, this is the relative strength of the surface of the packing. For silica gel, the more exposed the silanol groups, the more active the surface. Activity can be controlled by adding water or another polar modifier, which is hydrogen bonded to the active sites, thereby reducing the surface activity.

Adsorbent

Packing used in adsorption chromatography. Silica gel and alumina are the most frequently used adsorbents in HPLC.

Adsorption

The process of interaction between the solute and the surface of an adsorbent. The forces involved can be strong such as hydrogen bonds, or weak such as van der Waals forces. For silica gel, the silanol group is the driving force for adsorption, and any solute functional group that can interact with this group can be retained by liquid-solid chromatography on silica.

Adsorption chromatography

One of the basic LC modes which relies on the adsorption process to effect the separation. Silica gel and alumina are the most frequently used supports. The molecules are retained by the interaction of their polar functional groups with the surface functional groups such as silanols of silica.

Adsorption isotherm

In adsorption chromatography, this is a plot of the equilibrium concentration of sample in the mobile phase per unit volume verses the concentration in the stationary phase per unit weight. The shape of the adsorption isotherm can determine the chromatographic behavior of the solute such as tailing, fronting, and sample overload.

Affinity chromatography

A technique in which a biospecific adsorbent is prepared by coupling a specific ligand (such as an enzyme, antigen, or hormone) for the macromolecule of interest to a solid support (or carrier). This immobilized ligand will interact only with molecules that can selectively bind to it. Molecules that will not bind elute unretained. The retained compound can later be released in a purified state. Affinity chromatography is not a chromatographic technique but selective filtration.

Alumina

An adsorbent sometimes used in adsorption chromatography. Aluminum oxide (AI203) is a porous adsorbent that is available with a slightly basic surface. For this reason, it can have advantages over silica, which is considered to have an acidic surface.

Amino phase

A propylamino stationary phase used mostly in normal bonded phase chromatography. It is somewhat reactive for any solute molecule or mobile phase additive that can react with amines. The amino phase has found some applications as a weak anion exchanger.

Asymmetry

A factor describing the shape of a chromatographic peak. Theory assumes a Gaussian shape peak that is symmetrical. The peak asymmetry factor is the ratio (at 10 percent of the peak height) of the distance between the peak apex and the back side of the chromatographic curve to the distance between the peak apex and the front side of the chromatographic curve. A value >1 is a tailing peak, while a value <1 is a fronting peak.



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Backflushing

A column switching technique in which a four-way valve placed between the injector and the column allows the mobile phase to flow in either direction. Backflushing is used to elute strongly held compounds at the head of the column. It can be used to analyze these compounds or merely to remove them from the column.

Backpressure

The pressure above gravity at the head of the column. Expressed in psig, bar, atm., or MPa.

Bandspreading

The dilution of the chromatographic band as it moves down the column. The peak is injected as a slug, and, if not for the process of band broadening, each separated component would elute as a narrow slug of pure compound. The measure of band broadening is band width, tw, or more correctly, the number of theoretical plates in the column, N.

Baseline

The baseline is the line drawn by the data system when the only signal from the detector is from the mobile phase.

Baseline noise

Interferences to the detector and data system caused by electrical noise or environmental effects. This interference keeps the baseline from being perfectly flat.

Baseline-resolved peaks

When both sides of a peak reach the baseline without interfering with other peaks.

BET method

A method developed by Bruner, Emmett, and Teller for measuring surface area by using nitrogen adsorption condensation in pores at liquid nitrogen temperature. Pore volume and pore size distribution can also be obtained by this method.

Bonded-phase chromatography (BPC)

A stationary phase chemically bonded to a support that is used for the separation. It is the most commonly used LC mode. The most popular support used is microparticulate silica gel. An organosilane, such as octadecyl (for reversed-phase chromatography), is the most accepted type of bonded phase. Approximately 70 percent of all HPLC is carried out on chemically bonded phases.

Breakthrough volume

The volume at which a particular solute pumped continuously through a column begins to elute. The breakthrough volume is useful in determining the total sample capacity of the column for a particular solute.



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Calibration standards

These are standards of known quantities and substances which assist in peak identification or quantitation. Calibration standards can be external standards or internal standards.

Capacity factor

A chromatographic parameter that measures the degree of retention. See k’ for calculation method.

Capping

Same as Endcapping.

Carrier

A term used most often in affinity chromatography, which refers to the support that is used to carry the active ligand, usually by a covalent bond. It can also refer to the support in other chromatographic modes.

Cartridge column

A type of column with no endfittings which is held in a cartridge holder. The column is an open tube with the packing contained by frits in each end. Cartridges are easy to change, less expensive, and more convenient than conventional columns with endfittings.

Chain length

The length of carbon chain in the hydrocarbon portion of a reversed-phase packing. It is expressed as the number of carbon atoms such as C8 and C18.

Channeling

This occurs when voids are created in the packing material of a column. It results in the mobile phase and accompanying solutes moving more rapidly than the average flow velocity producing band broadening. These voids are created by poor packing or by erosion of the packed bed.

Chemisorption

This is sorption caused by a chemical reaction with the packing. Most of these interactions are irreversible. They usually occur on packings with reactive functional groups such as silanol or bonded amino phases.

Chiral stationary phases

Stationary phases that are designed to separate enantiomeric compounds. They can be bonded to solid supports, created in situ on the surface of the solid support, or they can be surface cavities that allow specific interactions with one enantiomeric form.

Chromatography

A chemical separation technique based on the differential distribution of the constituents of a mixture between two phases, one of which moves relative to the other.

Chromatographic methods

A record of the parameters used in a separation that yields a particular result. It allows another analyst following the method and conditions to reproduce the separation, and achieve the same results.

Chromatogram

The electronic result of a chromatographic separation which is a plot of detector signal output versus time or elution volume. It is represented as a series of peaks from the data system.

Chromatographic conditions

Parameters used in an analysis such as column type, mobile phase, and wavelength.

Column

A tube which contains the stationary phase. The stationary phase differentially interacts with the sample’s constituent compounds as they are carried along in the mobile phase.

Column backpressure

See Backpressure.

Column chromatography

Any form of chromatography that uses a column to hold the stationary phase. Open column chromatography, HPLC and open tubular capillary chromatography are all examples.

Column-dependent chemical factors

Chemical factors associated with column packing materials including the nature of the base material, surface activity of the bonded phase, and degree of interference from silanol groups.

Column performance

The efficiency of a column which is measured as the number of theoretical plates for a given test compound.

Column switching

The use of multiple columns connected by switching valves. Fractions from a primary column can be switched to two or more secondary columns, which in turn can be further diverted to additional columns or to the detector. This process is used for better chromatographic separations or for sample cleanup.

Conductivity detectors

These detectors identify changes in the conductivity of the mobile phase as it passes through a flow cell. They are used to detect a wide variety of ionized species separated by ion chromatography.

Connecting tube

A tube that connects the column to the injector and detector. Diffusion within connecting tubing broadens the peaks, but does not contribute to the separation.

Counterion

In an ion-exchange process, this is the ion in solution which is used to displace the ion of interest from the ionic site. In ion pairing, it is the ion of opposite charge added to the mobile phase to form a neutral ion pair in solution.

Coupled columns

A form of column switching. This uses a primary column connected to two secondary columns via a selector valve. Fractions from the first column can be selectively transferred to the other two columns for additional separation. This term is also used to describe two or more columns connected in series to provide increased plate numbers.