A
␣:See paration facto r.
A solvent:Usually the weaker solvent in a binary eluent or gradient elution paration. In reverd-pha liquid chromatography (LC), the A solvent is typically water or a water-rich mixture.
A term:The first term in the van Deemter equation.See eddy dispersion term and van Deemter equation.
Absorption:The process of retention in which the solute partitions into a liquid-like coating.Activity:The relative strength of the surface of the packing in adsorption chromatography. For silica gel, the more available the silanol groups, the more active the surface. Activity can be controlled by adding water or another polar modifier that hydrogen bonds to the active sites, thereby reducing the surface activity.Additive:A substance added to the mobile pha to improve the paration or detection characteristics;for example, a competing ba to negate the effects of silanols, a chelating agent to block metal sites or a UV-absorbing compound to perform indirect photometric detection.
Adjusted retention time (t R Ј):A measure of the retention time adjusted for the hold-up time; t R Јϭt R
Ϫt M , where t R is the retention time and t M is the hold-up time (the time it takes for a small,unretained compound that completely permeates the pores to be eluted from the chromatographic column).
Adjusted retention volume (V R Ј):Adjusts the retention volume for the holdup volume; V R ЈϭV R ϪV M ,where V R is the retention volume of the peak of interest and V M is the hold-up volume (the volume corresponding to the total volume of mobile pha
2
GLOSSARY OF LIQUID-PHASE SEPARATION TERMS
The first glossary of common and not-so-common terms and buzzwords for reference to high performance liquid chromatography (HPLC) columns and column technology was published in 1988 (1). It is time for an update becau
•many new terms have arin or, in some instances, their original meanings have expanded or changed
•the various techniques of capillary electrophoresis (CE) have become well developed and are ud
in many laboratories throughout the world
•the International Union of Pure and Applied Chemistry (IUPAC) published its massive
undertaking titled “Nomenclature for Chromatography,” which provides guidance and changes in some of the more commonly accepted terms (2).
This booklet updates the earlier glossary and will expand coverage into techniques beyond HPLC. This glossary is not intended to be an in-depth or highly theoretical treatment. For example,we have elected not to cover the myriad terms ud in instrumentation, detection, data handling,quantitative analysis and validation associated with liquid-pha analysis but instead have chon to u terms that analysts may encounter in everyday laboratory work with columns, phas and method development. The listing should be helpful to tho just starting in HPLC, CE and related techniques. It may also rve as a refresher for long-time urs.
LIQUID-PHASE SEPARATION TERMS
GLOSSARY OF
in the column). See also dead volume and hold-up volume.
Adsorbent:Packing ud in adsorption chromatography.
Silica gel and alumina are the most frequently ud adsorbents in high performance liquid
chromatography (HPLC).
Adsorption:The process of retention in which the interactions between the solute and the surface of an adsorbent dominate. The forces can be strong
forces (hydrogen bonds) or weak (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 on silica. The term adsorption places
emphasis on the surface versus penetration or
embedding in the stationary pha coated or
bonded to a surface.
Adsorption chromatography:One of the basic LC modes that relies upon adsorption to the surface of an active solid to effect the paration. Silica gel
and alumina are the most frequently ud normal-pha adsorbents, and molecules are retained by the interaction of their polar function groups with the surface functional groups; for example, silanols of silica. Carbon is also ud as an adsorbent in a
reverd-pha mode.
Adsorption isotherm:A plot of the equilibrium concentration of sample in the mobile pha per
unit volume versus the concentration in the
stationary pha per unit weight in adsorption
chromatography. The shape of the adsorption
isotherm can determine the chromatographic
behaviour of the solute; for example, peak tailing, peak fronting and column overload.
Aerogel:A packing prepared when the dispersing agent is removed from a gel system without
collapsing the gel structure. Silica gels and glass
beads ud for size-exclusion chromatography (SEC) are examples of aerogels that can retain their
structures even at the high pressures ud in HPLC.
See also xerogels.
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 lectively bind to it. Molecules that will not bind will be
eluted unretained. The retained compound can later be relead in a purified state. Affinity
chromatography is normally practid as an on–off paration technique.
Agaro:High molecular weight polysaccharide ud as a paration medium in biochromatography. It is ud in bead form, often in gel-filtration
chromatography, with aqueous mobile phas. Alkoxysilane:A reactant ud for the preparation of chemically bonded phas. It will react with silica gel as follows:
R3SiOR ϩϵSiOH ¡ϵSi–OSiR3ϩROH
where R is an alkyl group.
Alumina:A normal-pha adsorbent ud in adsorption chromatography. Aluminium oxide is a porous
adsorbent that is available with a slightly basic
surface; neutral and acidic modifications can also be made. Basic alumina can have advantages over
silica, which is considered to have an acidic surface. Amino pha:A propylamino pha ud in normal bonded-pha chromatography. It is somewhat
reactive for solute molecules such as aldehydes or
mobile-pha additives that can react with amines.
The amino pha has found some applications as a
weak anion exchanger, and it is also ud for the
paration of carbohydrates with a water–acetonitrile mobile pha. It is a relatively unstable pha. Amphoteric ion-exchange resin:Ion-exchange resins that have both positive and negative ionic groups.
The resins are most uful for ion retardation in which all ionic materials can be removed from
solution becau the anionic and cationic
functionalities coexist on the same material. Analyte:The compound of interest to be analyd by injection into and elution from an HPLC column. Anion exchange:The ion-exchange procedure ud for the paration of anions. Synthetic resins, bonded-pha silicas and other metal oxides can be analyd in this mode. A typical anion-exchange functional
group is the tetraalkylammonium, which makes a
strong anion exchanger. An amino group on a
bonded stationary pha is an example of a weak
anion exchanger.
Asymmetry:Factor describing the shape of a chromatographic peak. Chromatographic theory
assumes a Gaussian shape and that peaks are
symmetrical. A quantitative measure is the peak
asymmetry factor, which is the ratio of the distance
LC•GC Europe3
C8:See octylsilane.
C18:See octadecylsilane.
C4, C8, C18etc.:Refer to the alkyl-chain length of a reverd bonded pha.
C S:See Langmuir isotherm.
Capacity:See sample capacity.
Capacity factor (kЈ):Old term for a chromatographic parameter that measures the degree of retention.
Now defined as the retention factor(k) by the
International Union of Pure and Applied Chemistry (IUPAC). See also retention factor for method of
calculation.
Capillary column:Refers to columns with inner diameters less than 0.5 mm.
Capillary electrochromatography (CEC):A hybrid technique in which capillary columns are packed
with chromatographic sorbents, and electroosmotic flow rather than pressure moves mobile pha
through the column; technique has the surface-
mediated lectivity potential of HPLC and the high efficiency of capillary electrophoresis (CE).
Capillary gel electrophoresis (CGE):A technique in which a capillary is filled with, or the walls coated or covalently bonded with, cross-linked polyacrylamide to simulate slab gel electrophoresis; this polymer
network us a sieving mechanism; ud for protein, carbohydrate and DNA parations such as
fingerprinting and quencing.
Capillary isoelectric focusing:Separation is bad on isoelectric points of proteins; the capillary is filled
with solution; the sample is introduced into the
capillary in the prence of ampholytes; under the application of an electric field, the protein migrates until it reaches a pH at which it is neutralized and maintains that position in the capillary.
Capillary LC:Generally refers to HPLC performed in a fud-silica or other type of capillary column; the
inner diameters are typically less than 0.5 mm; has also been called micro-LC.
Capillary micellar electrochromatography:The CEC version of micellar electrokinetic capillary
chromatography (MEKC).
Capillary tubing:Tubing to connect various parts of a chromatograph and to direct flow to the proper
places. Most capillary tubing ud in HPLC is less
than 0.020 in. in inner diameter. The smallest uful inner diameter is approximately 0.004 in.
Capillary zone electrophoresis (CZE):CE performed in
an open fud-silica capillary tube with and without various additives and capillary coatings; also called open-tube capillary zone electrophoresis. Capping:Same as endcapping.
Carrier:A term most often ud in affinity
chromatography; refers to the support that binds
the active ligand, usually by a covalent bond; can
also refer to the support in other chromatography modes such as liquid–liquid chromatography. Carrier gas:The mobile pha in gas chromatography (GC).
Cartridge column:A column type that has no endfittings and is held in a cartridge holder. The
column compris a tube and packing contained by frits in each end of the tube. Cartridges are easy to change and are less expensive and more convenient than conventional columns with endfittings.
Cation-exchange chromatography:The form of ion-exchange chromatography that us resins or
packings with functional groups that can parate cations. An example of a strong cation functional
group would be a sulfonic acid; a weak cation-
exchange functional group would be a carboxylic
acid.
CE:Capillary electrophoresis.
CEC:See capillary electrochromatography.
CGE:See capillary gel electrophoresis.
Chain length:The length of carbon chain in the hydrocarbon portion of a reverd-pha packing. It is expresd as the number of carbon atoms (C8, C18 etc.). It specifically excludes the short chains —
typically methyl, isopropyl and c-butyl groups —that are also attached to the silane.
Channelling:Occurs when voids created in the packing material cau mobile pha and accompanying
solutes to move more rapidly than the average flow velocity, which in turn allows band broadening to occur. The voids are created by poor packing or
erosion of the packed bed.
Chemisorption:Sorption caud by a chemical reaction with the packing. Most of the interactions are
irreversible and usually occur on packings with
reactive functional groups such as silanol or bonded amino phas. Chemisorption is common with metal oxide phas that have strong Lewis acid sites. Chiral recognition:The ability of a chiral stationary pha to interact differently with two enantiomers thereby leading to their HPLC paration.
6GLOSSARY OF LIQUID-PHASE SEPARATION TERMS
