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TERMINOLOGY
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Pure Chromatography
The Chromatography
and Sample Preparation
Terminology Guide
The LCGC Editorial Team
W e are pleased to pres- tions and that nomenclature will be

ent the 2020 edition of covered here as well.
The Chromatography and The Chromatography and Sample
Sample Preparation Guide, written Preparation Guide is not intended
by Ron Majors and John Hinshaw. to be an in-depth listing or highly
Since 2002, we have provided our theoretical coverage. For example,
readers with various glossaries to we have elected not to cover many
organize the myriad terms used in of the myriad terms used in instru-
gas chromatography (1), and high ment at ion, detec t ion, d at a ha n-
performance liquid chromatogra- d ling, a nd va lidation a ssociated
phy (HPLC) columns and column with chromatographic analysis but
technology (2). Because there are a have chosen to use terms that may
number of terms common to both be encountered in everyday labora-
GC and LC, we decided to combine tor y work around columns, injec-
the glossaries into one large listing tion techniques, pha ses, method
in 2013, which also included termi- development, sample preparation
nology specific to sample prepara- tasks, and general usage. The list-
tion and ion chromatography (IC) ing should be helpful to those just
(3). These glossaries have been very sta rting in chromatography, but
popular. We hope you find it use- it can also serve as a refresher for
ful, too. long-time users in the field.
In this guide, we stick to the con-
ventions of the International Union
References
of Pu re a nd Applied Chem ist r y (1) J.V. H i n sh aw, LC G C 2 0 (11), 103 4 –
(IUPAC) in their “Nomenclature 1040 (2002).
for Chromatography” that provides (2) R .E . M ajor s a nd P.W. C a r r, LC G C
26(2),118–168 (2008).
guidance and changes in some of (3) R.E. Majors and Hinshaw, LCGC: The
the more commonly accepted terms Chromatography and Sample Preparation
(4). Still, there are many terms in Terminolog y Guide 31(s10), (2013).
(4) L .S. Ettre, “Nomenclature for Chro-
c om mon u s a g e t h at a re not i n matography” in Pure and Appl. Chem.
alignment with the IUPAC defini- 65(4), 819–872 (1993).
www.chromatographyonline.com TERMINOLOGY GUIDE FEBRUARY 2020 7
Absorption: The process of retention in

96-well collection plate: A fixed-size which the solute partitions into a liquid-
polyethylene rectangular plate (127.8 like coating.
mm × 85.5 mm): consisting of an array Accelerated solvent extraction (ASE):
of 8 × 12 (96) small “test tubes” called Trade name for a pressurized fluid ex-
wells; volumes of wells range from 0.5 traction system introduced by Dionex
to 2 mL. and now sold by Thermo Fisher Scien-
96-well filtration plate: A fixed-size tific; see pressurized fluid extraction
polyethylene rectangular plate (127.8 for details of technique.
mm × 85.5 mm) consisting of an array Active flow technology: A concept
of 8 × 12 (96) of small filter tubes (vol- that incorporates two types of column
umes range from 0.5 to 2 mL); a mem- designs: curtain flow technology means
brane filter placed at the bottom of the segmenting the flow at the injection end
well is used to filter liquid samples; of the column to ensure the analyte sees
sometimes a prefilter is placed above the middle of the packed bed where it is
the membrane filter to prevent clogging not disturbed by wall effects; parallel seg-
with particulate samples. mented flow at the column outlet selects
96-well plate: A small rectangular just the middle portion of the flow profile
plastic plate consisting of 96 individual resulting in improved efficiency without
wells that are basically small-volume test the presence of wall effects, giving the
tubes arranged in an 8 × 12 well pat- best overall column efficiency; a special
tern; used for liquid handling and other endfitting design is used to sample the
such requirements. center of the parabolic flow profile.
96-well solid-phase extraction plate: Active sampling: In active gas sam-
A small rectangular plastic plate con- pling, a pump is used to push the sample
sisting of 96 individual flow-through through a mass flow controller and into
SPE wells arranged in an 8 × 12 array the canister. Additional sample can be
that have top and bottom frits to con- collected, relative to the amount that
tain solid particles of sorbent or resin to can be collected by passive sampling,
perform SPE on a miniaturized scale; by pressurizing the canister with sample.
generally 1 mg to 0.2 g of packing is Commonly the sample is pressurized to
placed into the well, which can have 103 kPa (15 psig), effectively doubling
a volume of up to 2 mL; used for au- the sample volume.
tomated SPE with xyz liquid handling Active site: A reactive or strongly at-
systems or customized workstations. tracting site on the surface of a chro-
matographic packing that may bind
A analytes or cause peak tailing; some-
A solvent: Usually the weaker solvent in times mobile phase additives (such as a
a binary eluent or gradient elution sepa- competing base) can negate the effects
ration. In reversed-phase liquid chroma- of active sites.
tography (LC), the A solvent typically is Activity: In adsorption chromatography,
water or a water-rich mixture. the relative strength of the surface of the
A term: The first term in the van Deem- packing. For silica gel, the more avail-
ter equation. See eddy dispersion term able the silanol groups, the more active
and van Deemter equation. the surface. Activity can be controlled
by the addition of water or other polar Adsorption isotherm: In adsorption, a

modifier that hydrogen-bonds to the plot of the equilibrium concentration of
active sites thereby reducing the surface sample in the mobile phase per unit vol-
activity; can also refer to biological ac- ume versus the concentration in the sta-
tivity of a biomolecule. tionary phase per unit weight. The shape
Additive: A substance added to the of the adsorption isotherm can determine
mobile phase to improve the separation the chromatographic behavior of the sol-
or detection characteristics; examples ute such as tailing, fronting, or overload.
would be a competing base to negate Aerogel: A packing prepared when the
the effects of silanols, a chelating agent dispersing agent is removed from a gel
to block metal sites, or addition of a system without collapsing the gel struc-
UV-absorbing compound to perform ture. Silica gels and glass beads used for
indirect photometric detection. SEC are examples of aerogels that can
Adsorbent: Packing used in adsorption retain their structures even at the high
chromatography. Silica gel and alumina pressure used in HPLC. See xerogels.
are the most frequently used adsorbents in Affinity chromatography: A tech-
chromatography and sample preparation. nique in which a biospecific adsorbent
Adsorption: A process of retention in is prepared by coupling a specific li-
which the interactions between the gand (such as an enzyme, antigen, or
solute and the surface of an adsorbent hormone) for the macromolecule of
dominate. The forces can be strong interest to a solid support (or carrier).
forces (for example, hydrogen bonds) or This immobilized ligand will interact
weak (van der Waals forces). For silica only with molecules that can selectively
gel, the silanol group is the driving force bind to it. Molecules that will not bind
for adsorption and any solute functional are eluted unretained. The retained
group which can interact with this compound can later be released in a
group can be retained on silica. The purified state. Affinity chromatogra-
term adsorption places emphasis on the phy is normally practiced as an “on–off”
surface versus penetration or embedding separation technique.
in the stationary phase coated or bonded Agarose: High-molecular-weight poly-
to a surface. saccharide used as a separation medium
Adsorption chromatography: One of in biochromatography. It is used in bead
the basic separation and SPE modes that form and often used in gel filtration chro-
relies on the adsorption process to ef- matography using aqueous mobile phases.
fect a separation. Silica gel and alumina Alkoxysilane: A reactant used for the
are the most frequently used normal- preparation of chemically bonded
phase adsorbents in LC. Molecules phases. It will react with silica gel as fol-
are retained by the interaction of their lows : R3SiOR + ≡SiOH → ≡Si–OSiR3
polar function groups with the surface + ROH where R is an alkyl group.
functional groups (for example, silanols Alumina: A normal-phase adsorbent
of silica). Carbon is also used as an ad- used in adsorption chromatography.
sorbent in a reversed-phase LC mode. Aluminium oxide (Al2O3) is a porous
Porous polymer, carbonaceous, and adsorbent which is available with a
molecular sieve packings in GC exhibit slightly basic surface; neutral and acidic
adsorptive properties as well. modifications can also be made. Basic
alumina can have advantages over silica, added to a sample before injection to cut
which is considered to have an acidic down on interactions between analytes
surface; alumina is seldom used as an that are unstable or behave poorly in
HPLC column packing in practice. In the GC flow path on active sites; the
GC applications, alumina will separate protectorants are chosen so that they
low molecular-weight gases. do not interfere with the analysis of the
Amino phase: A propylamino phase compounds of interest yet prevent these
used in normal-phase chromatography. compounds from interacting with the
It is a somewhat reactive phase for any active sites in the flow path; these pro-
solute molecule (for example, aldehydes) tectorants are not generally required for
or mobile phase additive that can react LC and LC–MS.
with amines. The amino phase has found Analytical column: A chromatogra-
some applications as a weak anion ex- phy column used for qualitative and
changer and for the separation of carbo- quantitative analysis; a typical analyti-
hydrates using a water–acetonitrile mo- cal column for LC will be 50–250 cm
bile phase. It is a relatively unstable phase. × 4.6 mm, but columns with smaller
Amperes full-scale (AFS): Extent of the diameters (down to 0.05 mm i.d.) can
maximum detector output, for detectors also be considered as analytical col-
utilizing an electrometer. umns. GC analytical columns range
Amperometric detection: Electro- in length from 1 m to as much as 60
chemical detection applying a constant m, with inner diameters ranging from
potential to the working electrode. Mea- less than 100 µm up to 2 mm. Station-
sured current from oxidation or reduc- ary phases can be coated or bonded
tion is proportional to the sample con- onto the interior of the tubing; packed
centration. Very selective and sensitive GC columns are generally wider and
method. Works with electrode reactions shorter and are less frequently used
not changing the electrode surface (for nowadays. Chromatography columns
example, cyanide, nitrite, thiosulfate, can be constructed of stainless steel,
phenols). Only approximately 10% of glass, glass-lined stainless steel, PEEK,
the analyte is oxidized or reduced. May fused silica, and other metallic and
be used standalone as well as in series or nonmetallic materials.
parallel to other detectors. Anion exchange: The ion-exchange
Ampholyte: A substance that carries procedure used for the separation of an-
both positive and negative charges (they ions. Synthetic resins, bonded phase sili-
are amphoteric). Examples: amino acids, cas, and other metal oxides are available
proteins. for this mode. A typical anion-exchange
Amphoteric ion-exchange resin: Ion- functional group is tetraalkylammo-
exchange resins that have both positive nium, making a strong anion exchanger.
and negative ionic groups. These resins are An amino group on a bonded stationary
most useful for ion retardation where all phase would be an example of a weak
ionic materials can be removed from solu- anion exchanger.
tion since the anionic and cationic func- Argentation SPE: The incorporation
tionalities coexist on the same material. of a silver salt into the SPE stationary
Analyte protectorant: In GC, a chemi- phase will help in retaining compounds
cal compound or compounds that are with olefinic bonds. Normally used in
organic solvents to maximize charge-

transfer interactions.
Normalized peak height

Array 96-well plate: A 96-well SPE 1.0
plate where the 96 individual wells are
removable from the base plate; such a wa = A + B
0.5 hp
setup allows users to place different
types and amounts of SPE sorbents B A
0.1
0.0
into various configurations in each of 32
t1 t
36 p 40
t2
44 48
the 96-wells. This type of 96-well plate
Time (s)
has also been referred to as a flexible 96-
Figure 1: Example of a tailing peak. (Mod-
well plate configuration. ified with permission from reference 3.)
Asymmetry: Factor describing the
shape of a chromatographic peak. The- Average particle size (d p): The aver-
ory assumes a Gaussian shape and that age particle size of the packing in the
peaks are symmetrical. A quantitative column. A 5-µm LC column would be
measure is the peak asymmetry factor, packed with particles with a definite
which is the ratio of the distance from particle size distribution because pack-
the peak apex to the back side of the ings are never monodisperse. Particle
chromatography curve over the distance sizes in GC usually are expressed in
from the peak apex to the front side of terms of mesh size distribution; 80–100
the chromatography curve at 10% of the mesh is a commonly used particle range.
peak height. Various other measures of See particle size distribution.
asymmetry are in common use, espe-
cially the USP method. See also Foley– B
Dorsey equation. B solvent: Usually the stronger solvent in
Asymmetry factor: A factor that de- a binary eluent or gradient separation. In
notes band shape; calculated from the reversed-phase LC, typically the organic
chromatographic peak by dropping a modifier or modifier-rich binary mixture
perpendicular at the peak apex and with water.
a horizontal line at 10% of the peak B term: The second term of the van
height; at the intersection the distance Deemter equation; the first term of the
to the tail of the peak along the hori- Golay equation. See longitudinal dif-
zontal line (distance B) divided by the fusion, molecular diffusion term, van
distance along the horizontal line to Deemter equation, Golay equation.
the front of the peak (distance A); Back extraction: Used in liquid–liquid
this ratio is the peak asymmetry fac- extraction to perform an additional ex-
tor; for a symmetrical peak the value traction to further purify a sample; ini-
is one; for a fronting peak the value is tially the extraction may take place with
less than one; for a tailing peak, the an aqueous solvent buffered at a high pH
value is greater than one; the higher and an immiscible organic solvent; after
the value the less symmetrical the peak the initial extraction takes place and in-
is; values greater than 2 are generally terferences are removed, then by having
unacceptable. another aqueous solution at a low pH,
Atmosphere (atm): A unit of pressure. one can back-extract the analyte into the
1 atm = 101.325 kPa or 14.6959 psi. organic layer based on the analyte now
being in a neutral form. An example tion of the chromatographic band as it

would be for the cleanup of an acidic moves down the column. The peak is
substance containing –COOH groups; injected as a narrow slug and ideally
at high pH the carboxyl would be ionized each separated component would elute
and prefer the aqueous layer and impu- as a narrow slug of pure compound if
rities may migrate to the organic phase not for the process of band broadening.
and discarded; then the pH of the aque- The measure of band broadening is the
ous layer can be adjusted to a low value. peak dispersion, σ, or more correctly N,
Now the carboxyl group is in an union- the number of theoretical plates in the
ized form and readily extracted into the column. Sometimes called band disper-
organic layer as a purified substance. sion or band spreading.
Backflushing: Useful in chromatogra- Band width: See peak width at base and
phy to remove compounds that are held peak width at half-height.
strongly at the head of a column. By re- Baseline: The baseline is the line drawn
versing the flow at the conclusion of a by the recording device representing the
run, analytes trapped at the head of the signal from the detector when only mo-
column can be flushed from the column bile phase is passing through, in the ab-
entrance because they have a shorter dis- sence of any solutes. It also represents the
tance to travel; sometimes a strong solvent point from which calculations are often
in LC or elevated temperatures in GC made on peaks to determine peak area
will be needed to move them along. A or peak height.
valve or fluidic device is used to effect the Baseline drift: Term for any regular
change of mobile-phase flow direction. change occurring in baseline signal
Backflushing can be used for analysis of from an LC or GC detector; it may
these compounds or merely to remove arise from changes in flow rate of the
them from the column. mobile phase or from stationary phase
Back-pressure regulator: In LC, a de- bleed and may trend in a positive or
vice placed on-line after the detector to negative direction. Baseline drift oc-
maintain a positive pressure on the flow curs over a longer period of time than
cell minimizing solvent outgassing prob- baseline noise.
lems in the detector. In GC, the term Baseline noise: Irregular variations
usually refers to a carrier-gas regulator in (short term) in the chromatographic
the split vent line that maintains a con- baseline as a result of electrical noise or
stant pressure at the inlet as split flows temperature fluctuations, outgassing in
change. the flow cell, or poorly mixed mobile-
Bakeout: The process of removing con- phase solvents.
taminants from a column by operation Bed volume: See column volume.
at elevated temperatures, which should BEH: Bridged ethyl hybrid; an inorganic–
not exceed the maximum column tem- organic HPLC particle; has higher pH
perature. limits than silica gel.
Band: Refers to the chromatographic BET method: A method for measuring
peak as it moves along and is eluted from surface area developed by Bruner, Em-
the column. mett, and Teller (BET) that uses nitro-
Band broadening: The process of in- gen adsorption–condensation in pores at
crease in width and concomitant dilu- liquid nitrogen temperature. Pore volume
Tangents drawn to
the in ection points
1.000
σ
0.882
In ection points
Normalized peak height
wi
0.607 wi = 2σ
wh wh = 2.355σ
0.500
3σ
0.324
0.134 4σ
0.044 5σ
wb = 4σ
Figure 2: Widths of a Gaussian peak at various heights as a function of the standard

deviation σs) of the peak. (Modified with permission from reference 2.)
and pore size distribution can also be ob- Binary mobile phase: Mobile phase con-
tained from BET calculations. sisting of two solvents or buffers (or one
Bidentate silane: A specific type of of each).
bonded phase in which a short hydrocar- Bind–elute : In SPE, the normal mode of
bon bridge connects two silicon atoms in operation where upon loading the sample
a silane that is bounded to the surface onto a conditioned sorbent or resin, the
through two siloxane groups. analytes of interest are retained (bound)
Bimodal: In SEC, can be a porous while interferences and perhaps some of
packing material that has two distinct the matrix is not retained by the pack-
pore sizes or pore size distributions; in ing; after a wash step to remove some of
ion-exchange or HILIC chromatog- the undesired sample components, the
raphy or sample preparation can be a elution step uses a strong solvent to elute
packing material that has two types the analytes of interest in a small volume.
of functionalities (for example, cation Biocompatible: A term to indicate that
exchange and reversed phase; cation the column or instrument component
and anion) on one packing; in some will not irreversibly or strongly adsorb
cases, mixed beds consisting of two or deactivate biomolecules, such as pro-
different packings in one column can teins. Frequently means metal-free or
be bimodal. ceramic surfaces and components.
Blank: More correctly named method Breakthrough volume: The volume at

blank. A blank prepared to represent which a particular solute pumped con-
the matrix as closely as possible. The tinuously through a column will begin
method blank is prepared and analyzed to be eluted. It is related to the column
exactly like the field samples. Purpose: volume plus the retention factor of the
Assess contamination introduced dur- solute. It is useful to determine the
ing sample preparation activities. total sample capacity of the column
Bleed: Loss of material from a column for a particular solute.
or septum due to high-temperature BTEX: Refers to benzene, toluene, eth-
operation. May result in ghost peaks ylbenzene, and xylenes analysis.
plus increased detector baseline offset Buffer: A solution that maintains con-
and noise; in extreme cases, bleeding stant pH by resisting changes in pH as
chemicals from the stationary phase a result of dilution or addition of small
may build up on detector surfaces. amounts of acids and bases.
Blending: Refers to the process of mak- Buffer capacity: A quantitative mea-
ing a heterogeneous sample into a more sure of the potential of a buffer solu-
consistent and uniform sample by some tion (defined as the number of equiva-
type of blending operation; the most lents of strong acid or base to cause a
popular type of blender is the mechani- one unit change in the pH of 1 L of
cal blender that chops a semisoft mate- a buffer solution) or simply the abil-
rial into smaller parts. ity of a buffer to withstand injections
Bonded phase: A stationary phase that of a buffered sample solution without
has been chemically bonded to the a change in mobile-phase pH; capac-
inner wall of an open-tubular (capil- ity determined by pH, buffer pKa, and
lary) column or to the support particles. buffer concentration.
In LC, the substrate is usually a silica Buffer strength: See ionic strength.
gel particle or other base material.
Bonded-phase chromatography: The C
most popular mode in LC, in which a C4, C8, C18: Refers to the alkyl chain
phase chemically bonded to a support length of a reversed bonded phase.
is used for the separation. The most C term: The interphase mass transfer
popular support for bonded-phase term of the van Deemter and Golay
chromatography is microparticulate equations.
silica gel and the most popular type of Canister collection: A stainless steel
bonded phase is the organosilane, such vessel designed to hold vacuum to
as octadecyl (for reversed-phase chro- less than 1.3 Pa (10 mTorr) or pres-
matography). Approximately 70% of sure to 275 kPa (40 psig). Canisters
all HPLC is carried out on chemically are available in a range of volumes:
bonded phases. 400 mL, 1.0 L, 3.0 L, 6.0 L, and 15
Bonded-phase concentration: See L. The size of canister used usually
coverage. depends on the concentration of the
Boxcar chromatography: See column analytes in the sample, the sampling
switching; alternate name. time, the f low rate, and the sample
Breakthrough capacity: See break- volume required for the sampling pe-
through volume. riod. Typically, smaller canisters are
used for more concentrated samples, Capillary tubing: Tubing to connect

such as soil gas collection, 3-L and various parts of the chromatograph in
6-L canisters are used to obtain in- order to direct flow to the proper place.
tegrated (TWA) ambient air samples Most capillary tubing used in HPLC
at sampling times of up to 24 h, and is less than 0.020 in. for internal di-
large 15-L canisters are used for refer- ameter. The smallest useful internal
ence standards. Sampling time will be diameter is about 0.004 in.
limited by the combination of canis- Capping: Same as endcapping.
ter size and the flow rate at which the Carbon load: For a bonded-phase
sample is to be collected. silica, term usually used to describe
Capacity: See sample capacity. the surface coverage or the degree to
Capacity factor (k’): Deprecated name which the available silanols on the
for retention factor. column packing’s surface have reacted
Capillary column: Refers to chro- and been replaced with the bonded
matography columns of small inner phase; the higher the carbon load, the
diameter, ostensibly small enough to lower number of residual silanols. The
display a capillary effect with liquids. carbon load is normally expressed as
The diameter below which a column is % carbon (for example, 12% carbon).
considered “capillary” is poorly defined. In reversed-phase LC, the higher the
See open-tubular column, capillary carbon load, the greater the analyte
LC. retention.
Capillary column, packed: A capillary Carrier: A term most often used in af-
column that is packed with stationary- finity chromatography; refers to the
phase particles. In GC, 1/16-in. o.d. by support that is used to attach the ac-
1-mm i.d. columns are common. tive ligand, usually by a covalent bond.
Capillary electrochromatography Can also refer to the support in other
(CEC): A hybrid technique where chromatography modes, such as LLC.
capillary columns are packed with Carrier gas: Term for the gaseous mo-
chromatographic sorbents and elec- bile phase in GC.
troosmotic flow moves mobile phase Cartridge: Generally refers to the con-
through the column rather than pres- tainer used in SPE or filtration; a car-
sure; the technique has the surface- tridge may be as simple as a medical-
mediated selectivity potential of HPLC grade syringe barrel that is filled with
and the high efficiency of CE. packing contained at both ends by frits;
Capillary GC: See open-tubular col- it can also be a molded device or even
umn. a stainless steel device that contains
Capillary LC: Generally refers to HPLC similar sorts of packing material. In
carried out in a fused-silica or other SPE, the device is also referred to as
type of capillary column; most of the an SPE tube.
time the dimensions are in the sub-0.5- Cartridge column: A column type
mm i.d. range. Has also been called that has no endfittings and is held in a
micro LC. cartridge holder. The column consists
Capillary micellar electrochromatog- of a tube and the packing is contained
raphy (CMEC): The CEC version of by frits in each end of the tube. Car-
MEKC. tridges are easy to change and are less
expensive and more convenient than ample, C8, C18). Specifically excludes
conventional columns with endfittings. the short chains typical methyl, isopro-
Cation-exchange chromatography: pyl, and sec-butyl groups also attached
The form of ion-exchange chroma- to the silane.
tography that uses resins or packings Channeling: Occurs when voids cre-
with functional groups that can sepa- ated in the packing material cause mo-
rate cations. An example of a strong bile phase and accompanying solutes
cation functional group would be a to move more rapidly than the average
sulfonic acid; a weak cation-exchange flow velocity allowing band broadening
functional group would be a carbox- to occur. The voids are created by poor
ylic acid. packing or erosion of the packed bed.
Centrifugation: Centrifugation is Charged aerosol detection (CAD):
a process that involves the use of the The effluent from the LC column is
centrifugal force for the sedimenta- nebulized and then vaporized in a
tion of mixtures with a centrifuge (see heated drift tube, which results in a
centrifuge). This process is used to cloud of analyte particles; these par-
separate two immiscible liquids. More- ticles are charged and then the current
dense components of the mixture mi- from the charged particle flux is mea-
grate away from the axis of the centri- sured. The ELSD technique measures
fuge, while less-dense components of the light scattering properties of the
the mixture migrate toward the axis. aerosol particles. CAD is more sensi-
Chemists and biologists may increase tive and gives a more linear response
the effective gravitational force on a test than ELSD; it is also a universal detec-
tube so as to more rapidly and com- tion method.
pletely cause the precipitate (“pellet”) Check valve: A device inserted into a
to gather on the bottom of the tube. moving fluid stream that allows flow of
The remaining solution is properly the stream in only one direction; most
called the “supernate” or “supernatant often used on the inlet and outlet sides
liquid.” The supernatant liquid is then of an HPLC pump.
either quickly decanted from the tube Chelating resin: Chelating resin con-
without disturbing the precipitate, or tains functional groups that will inter-
withdrawn with a Pasteur pipette. act with cationic species (for example,
Centrifuge: A centrifuge is a piece metals such as copper, iron, heavy
of equipment, generally driven by an metal ions); useful for concentrating
electric motor (some older models were trace quantities or for separation.
spun by hand), that puts an object in Chemical filtration: A liquid sample
rotation around a fixed axis, applying is passed through a packing material
a force perpendicular to the axis (see (for example, adsorbent, ion exchange,
centrifugation). and so forth) that selectively interacts
Certify: Specific SPE products for with one or more compounds within
drugs of abuse isolation and analysis. the sample and acts as a chemical way
Chain length: The length of carbon to remove and purify the liquid sample.
chain in the hydrocarbon portion of a Regular filtration does not involve any
reversed-phase packing. It is expressed chemical interaction but merely re-
as the number of carbon atoms (for ex- moves particulates.
Chemical suppression: Remove back- Chiral stationary phase (CSP): A sta-

ground conductivity by ion exchange. tionary phase that is designed to sepa-
Converts the eluent into a low- or non- rate enantiomeric compounds. The
conducting component (for example, phase can be coated or bonded to solid
carbonate into carbonic acid, hydrox- supports, created in situ on the surface
ide into water, or nitric acid into water). of the solid support, or can include sur-
Anion analysis: the counter cation face cavities that allow specific interac-
(for example, sodium) is replaced by tions with one enantiomeric form.
the proton (H+). Cation analysis: the Chlorosilane: A chemical reagent used
counter anion (for example, nitrate) is to prepare siloxane bonded phases; re-
replaced by hydroxide (OH-). The mea- activity changes from a monochlorosi-
sured signal is the corresponding acid lane < dichlorosilane < trichlorosilane;
or base of the anion or cation respec- the alkyl portion (for example, octa-
tively. The dissociation of these acids decyl, octyl, and so forth) will dictate
and bases influences the signal. All sup- the hydrophobicity of the resultant
pressor devices work on this principle. bonded phase; alkoxysilanes can be
The typical background conductivity used but are less reactive.
after suppression is <20 µS/cm. Chopping: The process of mechanically
Chemisorption: Sorption due to a cutting a sample into smaller parts.
chemical reaction with the packing. Chromatogram: A plot of detector
Most such interactions are irreversible. signal output versus time or elution
Usually occurs on packings with reac- volume during the chromatographic
tive functional groups such as silanol or process.
bonded amino phases. Chemisorption Chromatograph: (n) A device used to
is common with metal oxide phases implement a chromatographic separa-
that have strong Lewis acid sites. tion; (v) the act of separation by chro-
Chip format: A miniaturization tech- matography.
nique where small channels on a glass, Chromatographic conditions: Those
polymer or other type of matrix are chromatographic method experimen-
used instead of large-bore columns, tal parameters that describe how an
capillaries, and so forth. The result- analysis was performed. Sufficient
ing format is greatly reduced in size information must be presented so that
compared to conventional chromato- the analysis can be duplicated for veri-
graphic instruments; advantages are fication purposes.
the reduction in sample, solvent, and Classification: The process of sizing
so forth; can be more easily coupled to column packing particles; generally, in
detection techniques (for example, MS HPLC a small particle size distribution
and MS/MS) where smaller amounts provides better efficiency and a greater
of mobile phase can lead to sensitivity permeability because of the absence of
enhancements and reduction in ion fines. Classification can be performed
suppression. by sedimentation, elutriation, and
Chiral recognition: The ability of a using centrifugal air classifiers.
chiral stationary phase to interact dif- Co-ion: An ion of the same sign of
ferently with two enantiomers leading charge as the ionic groups making up
to their chromatographic separation. the stationary phase.
Coating efficiency (CE, UTE, UTE%): Column outlet flow rate, corrected
A metric for evaluating column quality. (Fa): In GC, the column outlet flow
The minimum theoretical plate height rate corrected from column tempera-
divided by the observed plate height: ture and outlet pressure to room tem-
CE = Hmin/H perature and pressure, for example,
Cold injection: An injection that takes the f low rate as measured by a f low
place at temperatures below the final meter. Difficult to measure directly
oven temperature, usually at or below for narrow-bore open-tubular col-
the solvent boiling point. umns, the f low rate can be calcu-
Column: The tube and stationary lated from the average carrier-gas
phase through which mobile phase linear velocity, pressure drop, temper-
flows resulting in a chromatographic atures, and the column dimensions:
separation. F a = (u– πd c2T0)/(4jTc). Such calcula-
C o l u m n c h r o m a t o g r a p h y : A ny tions are the basis of electronic pres-
form of chromatography that uses sure control.
a column, tube, or plate to hold the Column overload: If one exceeds the
stationary phase. Open-column chro- sample capacity (or loading capacity) of
matography, HPLC, and open-tubu- a column, peaks will become distorted
lar capillary gas chromatography are and may be difficult to measure and
all forms of column chromatography. to achieve reproducible chromatogra-
Most often refers to open-column phy from run to run. One can measure
chromatography used for prepara- column capacity by running a break-
tive work. through study (see breakthrough vol-
Column dead time: See hold-up time. ume).
Column equilibration: To provide re- Column packing: The solid material,
producible results, a column should be usually a porous solid with or without
equilibrated with the surrounding envi- a chemically interactive surface, placed
ronment be it a temperature condition, inside or on the walls of the column
mobile phase equilibrium, pressure used to differentially retain analytes;
condition, and so forth; in GC, it is also referred to as the stationary
important that the temperature of the phase; common packings include un-
column be stabilized after a tempera- bonded and bonded silica, resins, in-
ture programmed run and in LC, the organic–organic hybrids, graphitized
column must be returned to its origi- carbon, porous polymers, and molecu-
nal conditions before another gradient lar sieves.
is run. Column performance: Denotes the
Column inner diameter (d c ): The column efficiency. See theoretical
inner diameter of an uncoated chro- plate.
matography column. Column plate number: Denotes the
Column length (L): The length of the column efficiency. See theoretical
analytical chromatography column plate.
used to perform the chromatographic Column suppressor: Initial setup.
separation. Distinct from the length Packed ion-exchanger columns were
of a precolumn (LC) or retention gap used for chemical suppression. Draw-
(GC) connected in series. backs: require regeneration, changes
of the selectivity throughout the of two columns connected in series by

usage. a retention modulation device, thereby
Column switching: The use of mul- generating much higher resolution than
tiple columns connected by switching with any single column.
valves to effect better chromatographic Comprehensive two-dimensional
separations or for sample cleanup. Frac- chromatography: Two-dimensional
tions from a primary column can be chromatography applied to every frac-
switched to two or more secondary tion. See two-dimensional chroma-
columns which in turn can be further tography.
diverted to additional columns or to Compressibility correction factor (j):
the detector (or detectors); sometimes Due to gas compressibility, the carrier
referred to as multidimensional chro- gas expands and its velocity increases
matography. as it proceeds along a GC column from
Column temperature (Tc ): Tempera- the inlet pressure pi to the outlet pres-
ture of the column. A uniform tem- sure po. The carrier gas compressibility
perature across the column usually is correction factor corrects the carrier
desirable; however, GC separations gas velocity at the outlet of a GC col-
also may be performed with a moving umn to the average carrier gas velocity:
temperature gradient along the column j = 3(P 2 – 1)/2(P 3 – 1), where P is the
length. column pressure drop: P = pi/po
Column volume (Vc): The volume of Concentration: The process of increas-
the unpacked, uncoated column: Vc ing the strength or density of a diluted
= AcL = πrc2L, where Ac and L are the sample; a more concentrated sample
cross-sectional area of the tube and the will be easier to measure; concentration
tube length, respectively. can be accomplished by a wide variety
Cool-down time: Length of time re- of sample preparation techniques such
quired to cool a GC oven from the final as evaporation, adsorption, diffusion,
oven temperature to the initial oven and so forth.
temperature. Shorter cool-down times Conditioning (SPE): This is gener-
allow a greater number of analyses to ally considered to be the first step in
be performed in a given time period. SPE; the stationary phase must first
Competing base: In reversed-phase be put into a chemical or physical
LC, addition of a small basic com- state that it can accept the sample
pound such as triethylamine or di- solution loaded in the second SPE
methyloct ylamine at 25 –50 mM step; a conditioning solvent is passed
concentration to the mobile phase to through the SPE stationary phase
inhibit basic analytes from interact- where it will solvate the phase so that
ing with residual silanols; works by it will more easily sorb the sample of
law of mass action because the con- interest; for a reversed-phase SPE car-
centration of competing base is much tridge, methanol or acetonitrile serves
greater than that of the analyte. See as a conditioning solvent; sometimes
also additive. the excess conditioning solvent must
Comprehensive GC (GC×GC): Two- be removed but the packing shouldn’t
dimensional technique in which all be allowed to dry out because that
compounds experience the selectivity may affect the “conditioned” phase.
Conductivity: Conductivity is the in- method used by analytical chemists

herent parameter of all ions. Therefore to reduce the sample size of a powder
it is the signal for measuring the chro- without creating a systematic bias. The
matogram. It may be measured directly technique involves pouring the sample
or after chemical and sequential sup- so that it takes on a conical shape, and
pression respectively. As conductivity is then flattening it out into a cake. The
strongly dependent on the temperature cake is then divided into quarters;
(around 2 %/°C), a thorough isolation the two quarters that sit opposite one
and thermal stabilization of the detec- another are discarded, and the other
tor block is recommended. The mea- two are combined and constitute the
sured conductivity of a solution is given reduced sample. The same process is
by the sum of the “single ion conduc- continued until an appropriate sample
tivities.” The single ion conductivity is size remains. Analyses are made with
a linear function of the concentration respect to the sample left behind.
of the ion and its equivalent conduc- Continuous liquid–liquid extraction:
tivity. At very low concentrations or at Useful when the K D value is very low
constant ion strength the equivalent or the required sample volume is very
conductivity is constant. The mea- large when multiple extractions are im-
sured signal in nonsuppressed IC is practical; also if the extraction is slow,
proportional to the concentration and a long time may be required for equi-
the difference of eluent and sample ion: librium to be established; in continu-
As the ionic strength of the solution is ous LLE, fresh solvent is continually
constant (stoichiometric ion exchange) recycled through the aqueous sample;
also the equivalent conductivities are continuous extractors are available for
constant. This leads to very linear cali- heavier-than-water and lighter-than-
bration curves. The measured signal water solvents.
in suppressed IC is proportional to the Controlled surface porosity support:
sum of the equivalent conductivities Same as porous layer bead and pel-
of the analyte ion and the counterion licular stationary phase.
(which has been introduced by suppres- Cool on-column injection: Cool on-
sion). As only the sample ion and its column injection is a technique of in-
counterion is adding to the measured troducing a sample as a liquid directly
conductivity after suppression, the total into a GC column; this lack of prior
concentration is changing during the vaporization offers the following advan-
peak. Therefore the equivalent conduc- tages: It eliminates sample discrimina-
tivities are no longer constant. This is tion; it eliminates sample alteration;
one reason for the inherent nonlinear- and it provides high analytical preci-
ity of the calibration curves in sup- sion. However, there are some special
pressed IC. requirements: It requires relatively
Coning and quartering: A sample size clean samples; real samples are often
reduction technique where a portion of too concentrated for on-column injec-
free-flowing solid material (powder) is tion and must be diluted; and peak
systematically divided into quadrants splitting or peak distortion can occur
to achieve a statistically representative due to differing polarities of solvent,
sample. Coning and quartering is a stationary phase, and solutes.
Core–shell: See superficially porous HPLC and CE (MEKC) by a partition-

particles (SPPs). ing mechanism.
Coulometric detector: Same as am- Cross-linked phase: A stationary
perometric detection, but with a con- phase that includes cross-linked poly-
version rate of 100%. All the analyte mer chains. Usually it is also bonded
is oxidized or reduced at the working to the column inner wall. See bonded
electrode. The response is larger than phase.
with amperometric detection. But on Crosslinking: For resins, during the
the other hand also the baseline noise process of copolymerization to form
is larger. Therefore the detection limits a three-dimensional matrix a difunc-
are almost the same. tional monomer is added to form cross-
Counterion: In an ion-exchange pro- linkages between adjacent polymer
cess, the ion in solution used to displace chains. The degree of cross-linking
the ion of interest from the ionic site. is determined by the amount of this
In ion pairing, it is the ion of opposite monomer added to the reaction. For
charge added to the mobile phase to example, divinylbenzene is a typical
form a neutral ion pair in solution. cross-linking agent for the production
Coupled columns: A form of column of polystyrene ion-exchange resins. The
switching that uses a primary column swelling and diffusion characteristics
connected to two secondary columns of a resin are governed by its degree of
via a selector valve. Fractions from col- cross-linking.
umn one can be selectively transferred Crushing: Tungsten carbide variable
to columns two and three for addi- jaw crushers for reducing the size of
tional separation to occur. The term is large, extremely hard, brittle samples.
also used to describe two or more col- Curtain flow technology: Curtain
umns connected in series to provide an flow technology refers to the process
increased number of plates. of injection of sample across a radial
Coverage: Refers to the amount of cross section of an HPLC column to
bonded phase on a silica support in ensure the analyte sees the middle por-
bonded phase chromatography. Cov- tion of the packed bed and not the wall
erage is usually described in µmol/m2 where flow effects may be different; the
or in terms of %C (w/w). technique is coupled with a parallel seg-
Crash plate: Refers to the process of mented flow fitting at the column out-
precipitating protein from plasma by let to select just the middle portion of
the addition of a miscible organic sol- the flow profile resulting in improved
vent such as acetonitrile; when a 96- efficiency without the presence of wall
well flow-through or fixed-well plate is effects.
used for this process, it is referred to as Cutting: Cutting mills can reduce soft-
crashing and the plate a crash plate. to-medium hard materials (diameter <
Critical micelle concentration (CMC): 100 mm).
The concentration of an ionic surfac- Cyano phase: A chemically bonded
tant above which a micelle is formed phase that terminates with the -CN
by aggregation; micelles added to the functional group; it can be used in
mobile phase are used to improve the normal-phase chromatography as
separation of nonionic substances in a moderate polarity sorbent and in
reversed-phase chromatography as a Degassing: The process of removing

short chain bonded phase. dissolved gas from the mobile phase
Cyclodextrins: Cyclic oligomers of prior or during use. Dissolved gas that
several D -(+)-glucopyranose units used may come out of solution in the detec-
in chiral HPLC and CE separations; tor cell can cause baseline spikes and
popular ones are named α-, β-, and noise. Dissolved air can affect certain
γ-CDs; they have a truncated cone detectors, such as electrochemical (by
shape, a relatively hydrophobic cavity, reaction) or fluorescence (by quench-
and primary and secondary hydroxyl ing). Dissolved gases can also cause
groups at their end; separate on basis pumps to lose prime. Degassing is car-
of differential inclusion of enantio- ried out by heating the solvent or by
mers; modified CDs with derivatized vacuum (in a vacuum flask), or on-line
hydroxyl groups are also used for se- using evacuation of a tube made from a
lectivity modification. gas-permeable substance such as PTFE,
or by helium sparging.
D Denaturing HPLC: Use of reversed-
Data acquisition rate: A term refer- phase HPLC to investigate genetic
ring to the rate of sampling of a de- mutations by the investigation of DNA
tector output. To characterize a chro- base pairs.
matographic peak at least 20–30 data Derivatization: A technique used in
points must be collected. The data chemistry that transforms a chemical
acquisition rate, usually measured in compound into a product (the reac-
hertz, defines how many data points tion’s derivate) of similar chemical
per second are collected while the peak structure, called a derivative. Gener-
is moving through the detector. For fast ally, a specific functional group of
chromatography, the data acquisition the compound participates in the de-
rate must be sufficiently rapid to char- rivatization reaction and transforms
acterize a narrow peak. Modern detec- the compound into one with a differ-
tors have data rates up to 200 Hz; also ent reactivity, solubility, boiling point
known as data rate and sampling rate. (volatility), melting point, aggregate
See detector time constant. state, or chemical composition. The
Dead volume: Dead volume is extra resulting new chemical properties can
volume experienced by solutes as they be used for quantification (for example,
pass through a chromatographic sys- UV or fluorescence detection) or better
tem, in particular any unswept vol- separation properties.
ume exposed to the mobile phase flow. Desalting: Technique where low-
Excessive dead volume causes addi- molecular-weight salts and other com-
tional peak broadening. Related to the pounds can be removed from nonionic
hold-up volume, which is the volume a nd high-molecular-weight com-
of mobile phase necessary to elute an pounds. The use of a reversed-phase
unretained compound. See hold-up packing to retain sample compounds
volume. by hydrophobic effects yet allow salts
Deep-well plate: A 96-well plate ca- to pass through unretained would be
pable of handling up to 2 mL of liquid an example of desalting. The use of an
volume per well. SEC column to exclude large molecules
and retain lower-molecular-weight salts layer of material that contains holes

is another example. Desalting using of various sizes, or pores. Smaller
dialysis is commonly used in protein solutes and f luid pass through the
purification. m e m br a ne , but t he m e m br a ne
Desorption: The process in chroma- block s the passage of larger sub-
tography where a molecule residing on stances (for example, red blood cells,
the surface of a packing material or on large proteins). It is a technique used
another solid surface (for example, col- in biological sample prep to desalt
umn wall or frit) or stationary phase biological f luids.
moves from the surface into the mobile Diatomaceous earth: Also known as
phase. diatomite or kieselguhr, it is a natu-
Detector response time: Time for a rally occurring, soft, siliceous sedi-
detector to respond to ~90% of the in- mentary rock that is easily crumbled
coming solute amount. The response into a fine white to off-white powder.
time is generally taken as 2–4 times the Diatomaceous earth consists of fos-
time constant. See also detector time silized remains of diatoms, a type of
constant. hard-shelled algae. It has a particle
Detector time constant (τ): The time size ranging from less than 3 µm to
for a detector to respond to 1/e = 63.2% more than 1 mm, but typically 10–
of an instantaneous change in solute 200 µm. Depending on the granular-
amount. In general the detector time ity, this powder can have an abrasive
constant should be less than 10% of feel, similar to pumice powder, and is
the peak width at half-height. Excessive very light as a result of its high poros-
detector dead volume, slowly respond- ity. The typical chemical composition
ing electronics, digital data acquisition of oven-dried diatomaceous earth is
speeds, and signal filtering strongly in- 80–90% silica, with 2–4% alumina
fluence detector response times. Too- (attributed mostly to clay minerals)
slow detector response times cause peak and 0.5–2% iron oxide; highly puri-
tailing, loss of peak height and detect- fied diatomaceous earth is used as a
ability, plus loss of peak resolution for support for chromatography and for
closely adjacent peaks. supported liquid–liquid extraction.
Dextran: Polydextran-based packing Diethylaminoethyl (DEAE): A popu-
material primarily used for low pres- lar weak anion-exchange functionality
sure biochromatography; an example typically attached to cellulose or Sep-
would be Sephadex (Amersham Phar- harose (GE Healthcare); used for the
macia Biotech). separation of biomolecules.
Dialysis: Dialysis works on the prin- Diethylene glycol succinate (DEGS):
ciples of the diffusion of solutes and A GC stationary phase.
ultrafiltration of f luid across a semi- Diffusion coefficient (D M or D G ; D S
permeable membrane. Diffusion is or D L ): A fundamental parameter of
a property of substances in water; a molecule in the liquid or gaseous
substances in water tend to move mobile phase (D M or D G) or in the
from an area of high concentration liquid stationary phase (D S or D L) that
to an area of low concentration. A expresses the degree of free mobility
semipermeable membrane is a thin of the molecule in solution. Expressed
in cm 2/s, the diffusion coefficient is Dimethylchlorosilane (DMCS): Some-

dependent on molecular weight of the times used for silanizing glass GC parts
solute, temperature, solvent viscosity, such as inlet liners and endcapping sil-
and molar volume of the solute. A ica-based HPLC bonded phases. Dis-
typical value of a small molecule (100 posable presilanized inlet liners are a
Da) in a liquid phase at room tem- preferable alternative that avoid use and
perature is on the order of 10 -5 cm 2/s. storage of this hazardous reagent.
Gaseous solutes in helium carrier gas Diode-array detection (DAD): Each
at 120 °C have diffusion coefficients wavelength of the UV and visible range
that are several orders of magnitude of the light is measured with an indi-
higher, around 0.4 cm 2/s. vidual diode. The optical resolution of
Digestion: The process of treating an the detector is defined by the number
insoluble chemical compound with a of diodes used (for example, 844 di-
reactive substance (for example, for odes: optical resolution = 1.4 nm).
inorganic compounds it might be a Diol phase: A hydrophilic phase use-
strong acid; for a biological compound ful in both normal and reversed phase.
it might be an enzyme) that will break It consists of a diol structure (two -OH
it down or disintegrate the compound groups on adjacent carbon atoms in
into a more soluble form that can be an aliphatic chain). In normal-phase
further treated or analyzed. work, it is less polar than silica and in
Digital chromatography: The pro- reversed phase work has been used for
cess of solid-phase extraction (SPE) is the separation of proteins and poly-
sometimes referred to as digital chro- peptides.
matography. A substance is either on Direct injection: Sample enters the
the SPE stationary phase or is off the inlet and is swept into the column by
stationary phase during its retention carrier gas flow. No sample splitting
and elution; in chromatography we or venting occurs during or after the
are often trying to resolve closely re- injection.
lated substances by exploiting subtle Direct sampling: A method of sample
differences in retention in more of an collection where a sample is taken di-
analog separation mode; in terms of k rectly from the source. For example, a
values, ideally the solute in SPE has a cannister may be used to collect a gas
value of infinity when on the sorbent sample exactly where the scientist desires.
and zero when eluted into solution. A river water sample can be obtained
Dilute and shoot: A simple sample by lowering a collection vessel directly
preparation procedure where one into the water. A thermal desorption
merely dilutes the sample with solvent, tube can be used to concentrate volatile
mobile phase, or a compatible liquid and semivolatile analytes by passing a
and then injects that diluted sample gas stream through the adsorbent con-
into a chromatograph without any fur- tained within the tube. This would be
ther sample preparation. an example of direct sampling.
Dilution: Reducing the concentration Direct thermal sampling: Refers to
of a chemical by adding an inert sub- the process of using temperature as a
stance; the substance can be a liquid, variable in sample volatile and semi-
solid, or gas. volatile substances; static headspace at
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a given temperature is an example of of solvent containing extracted ana-

thermal sampling; by selecting a cer- lytes (tetrachloroethylene) is removed
tain temperature certain sample com- by a microsyringe for direct injection.
ponents can be ruled out because they Extraction is almost instantaneous and
may have extremely low volatility at a enrichment values are quite high.
selected temperature; thermal sampling Dispersive SPE (dSPE): In dSPE,
can occur in stages all the way up to loose SPE packing material is added
pyrolysis where chemical bonds in the directly to a solution rather than pass-
sample are purposely broken to access ing it through the packed material in a
the structure of the material. cartridge or tube; dSPE is most often
Disk: A number of sample prepara- used as the second step in QuEChERS
tion and separation media take the where matrix compounds are removed
form of a disk; the most popular disks from the organic solvent salting out ex-
are used in filtration and may consist traction of the first step.
of any number of porous polymeric Displacement c hromatography:
materials; the most popular types Chromatographic process where the
of SPE disks would have embedded sample is placed onto the head of the
particles in a disk made of PTFE or column and then is displaced by a
other inert polymeric material or a compound that is more strongly sorbed
fiberglass matrix with interdispersed than the compounds of the original
sorbent particles; some biological mixture. Sample molecules are then
purification media employ the disk displaced by each other and by the
format. The stationary phases may more strongly sorbed compound. The
contain ion exchange groups or other result is that the eluted sample solute
functionality that attracts solutes of zones may be sharpened; displacement
interest or impurity that one may techniques have been used mainly in
want to get rid of. preparative HPLC applications.
Disk cartridge (SPE): See disk. Disposable filter: See syringe filter.
Dispersion: See system dispersion. Dissolution: The process of having
Dispersive liquid–liquid microextrac- a sample dissolve in an appropriate
tion (DLLME): The technique is based solvent.
on a three-component solvent system. Distillation: A method of separating
The container is usually a centrifuge mixtures based on differences in vola-
tube and the appropriate mixture of tility of components in a boiling liquid
immiscible organic extraction solvent mixture. Distillation is a unit operation,
(usually a few microliters, such as 8 µL or a physical separation process, and
of tetrachloroethylene) and a dispersive not a chemical reaction; it can be used
solvent (for example, ~1 mL of acetone) to purify organic compounds or to re-
is rapidly injected with a syringe into move solvent; fractional distillation is
an aqueous solution (~5 mL) contain- used to separate compounds with close
ing the analyte of interest. When the boiling points; azeotropic distillation is
three solvents are rapidly mixed, a using an azeotrope to remove a solvent
cloudy solution is formed consisting of that has a boiling point too close or
droplets of extraction solvent; the entire equal to another compound that can-
mixture is centrifuged and the droplet not be separated.
Distribution constant (coefficient) nal fluid, saliva) as well as other non-

(Kc): The equilibrium concentration biological media have been investigated.
of a component in or on the station- Drying: Drying of sample extracts can
ary phase divided by the equilibrium be accomplished by heating (evapora-
concentration of the component in the tion), vacuum dessication, and other
mobile phase; also called the distribution means; water can be removed (dried)
coefficient or in partition chromatogra- from organic solvents by using anhy-
phy the partition coefficient; in partition drous sodium sulfate.
chromatography Kc is used and refers to Dwell time: The time equivalent to
the case where the concentration in the dwell volume; determined by the prod-
stationary phase is expressed per unit uct of flow rate times the dwell volume.
volume of the phase: VR = VM + KcVS; Dwell volume: In LC, refers to the
in the case of a solid stationary phase, volume between the point of mixing of
Kg is used and is expressed as per mass solvents (usually in the mixing chamber
(weight) of the dry solid phase; in the or at the proportioning valves in the liq-
case of adsorption chromatography with uid chromatograph) and the head of the
a well characterized adsorbent of known chromatographic column; important in
surface area, the concentration in the gradient elution or when changes in sol-
stationary phase is expressed as per unit vent composition are made in isocratic
surface area. elution so that the column experiences
Dividers: A mechanical device used in the composition change in the shortest
subdividing solid powder samples into possible time. Low-pressure mixing sys-
smaller units; they can be manual or au- tems generally have larger dwell volumes
tomated; sample dividers will subdivide than high-pressure mixing systems.
material samples into two smaller por- Dynamic coating: The formation of in-
tions by a single pass or further subdivi- situ coatings on the packing in HPLC
sions can be attained by multiple passes. or on capillary walls in CE by addition
The important feature of sample divid- of a substance to the mobile phase that
ers is that each subdivision retains the adsorbs (or absorbs) onto the pack-
characteristics of the original sample. ing or at the wall surface; the purpose
Dried blood spot (DBS) analysis: A of a dynamic coating is to generate a
newer method for sampling and trans- new stationary phase or to deactivate
porting blood samples; a small (~15 µL) the packing material or capillary wall
sample of whole blood is placed on a cel- to prevent unwanted interactions; one
lulose or other paper-like material and simple example is the adjustment of
is dried for 2 h; the dried blood spot the mobile phase or running buffer to
can be extracted to remove analytes of a pH < 3 to protonate silanols and ne-
interest for further workup; has poten- gate their effect; another is coating the
tial to replace drawing large quantities phase with a hydrophilic polymeric ma-
for blood analysis; used in conjunction terial to prevent adsorption of proteins.
with LC–MS/MS for high sensitivity In GC, dynamic coating applies a con-
and specificity. trolled-thickness coating to the inside
Dried media spot (DMS) analysis: In of open-tubular columns as a solution
addition to DBS, other biological fluids flows through the column and leaves
(for example, plasma, serum, cerebrospi- a thin coating behind, which is then
evaporated to yield a thin coating of any expression is misleading as the suppres-

non-volatile material from the solution. sion is chemical. Only the supply of the
Dynamic headspace: See purge-and- respective ion is done electrochemically.
trap sampling. Electrodialysis: Used to transport salt
ions from one solution through ion-ex-
E change membranes to another solution
Eddy dispersion (diffusion) term (A under the influence of an applied elec-
term): The A term in the van Deemter tric potential difference. This is done in
equation. It is the contribution to plate a configuration called an electrodialysis
height that arises from the heterogeneity cell. The cell consists of a feed (diluate)
in axial velocities as a result of the par- compartment and a concentrate (brine)
ticle size and geometry of the packing compartment formed by an anion-
as well as wall effects: a = 2λdp. Typi- exchange membrane and a cation-ex-
cal values of λ for well-packed columns change membrane placed between two
are 0.8–1.0. Some theories of chroma- electrodes; can provide good enrichment
tography indicate a velocity dependent factors.
contribution to the HETP from this Electrolytic suppression: Synonym to
process. Also known as eddy diffusion, electrochemical suppression.
flow-heterogeneity induced broadening, Electrolytic conductivity detection
and the multipath term. The A term is (ELCD): An electrolytic conductivity de-
zero in open-tubular GC columns. See tector catalytically reacts halogen-con-
van Deemter equation. taining solute with hydrogen (reductive
Efficiency: The ability of a column mode) to produce strong acid by-prod-
to produce sharp, well-defined peaks. ucts that are dissolved in a working fluid.
More efficient columns have more theo- The acids dissociate and the increased
retical plates (N) and smaller theoretical electrolytic conductivity is measured.
plate heights (H). For asymmetric peaks Other operating modes modify the
it is better determined from the peak chemistry for response to nitrogen- or
centroid and variance by mathematical sulfur-containing substances.
analysis of the peak shape. See Foley- Electron-capture detection (ECD): An
Dorsey equation. electron-capture detector ionizes solutes
Effluent: The mobile phase leaving the by collision with metastable carrier gas
column; the same as eluate. molecules produced by β-emission from
Electrochemical detector: Global term a radioactive source such as 63Ni. ECD
for all detection modes recording elec- is one of the most sensitive detection
trical potential or current (conductivity, methods and responds strongly to halo-
amperometric, pulsed amperometric, genated solutes and others with a high
coulometric, and potentiometric detec- electron-capture cross-section.
tion). More often used for amperometric, Electronic or programmed pressure
pulsed amperometric, and coulometric control (EPC or PPC): Any of a number
detection only. of pressure and flow control devices that
Electrochemical suppression: Con- incorporate electronic pressure or flow
tinuously working chemical suppressor sensing and can be programmed from
where H+ or OH- are electrochemically the GC microcontroller. Such devices
produced by the electrolysis of water. The enable method control of flow, velocity,
and pressure for GC columns, as well ment chromatography and frontal

as providing a convenient means of in- analysis.
corporating gas-related parameters into Elution step: This is generally consid-
electronic methods. ered to be the fourth step in SPE and
Eluate: Combination of mobile phase occurs after the washing (rinsing) step;
and solute exiting the column; also in the elution step, analytes are removed
called effluent. from the SPE stationary phase by elu-
Eluent: The mobile phase used to carry tion with a strong solvent so that the
out a separation. analytes are now in a concentrated state;
Eluite: The species being eluted: the often, the strong solvent is removed by
analyte, or the sample. evaporation and reconstituted in a sol-
Eluotropic series: A series of solvents vent more compatible with the chro-
(eluents) with an increasing degree matographic system.
of solvent strength generally used in Elution volume (VR): Refers to the vol-
liquid–solid or adsorption chroma- ume of mobile phase required to elute a
tography. In normal-phase chroma- solute from the column. For a symmet-
tography, a nonpolar solvent such as ric peak, it is the volume from the point
pentane would be at one end of the of injection to the volume at maximum
scale, an intermediate solvent such concentration (apex): VR = FtR where F
as dichloromethane would be in the is the flow rate and tR is the retention
middle of the scale, and a strongly time of the peak of interest.
polar solvent such as methanol would Elutriation: A technique used to frac-
be near the upper end of the scale. In tionate packing particles by size based on
normal-phase chromatography, the the difference in their Stokes terminal
reverse order of strength would be velocities. It is most often used for the
observed; water would be weak and separation of ion-exchange resins that
hexane strong. Thus, when developing need to have a particularly narrow size
a method or running a gradient, an range, such as amino acid resins. The
eluotropic series is useful for selecting technique involves the upward flow of
solvents. water into a large tube. The unsized
Elute: To chromatograph by elution beads are added to the moving water
chromatography. The term elute is pre- and the particles seek their own level, de-
ferred to the term develop used in older pending on their density and particle size.
nomenclature. They are then removed at certain levels
Elution: The process of passing mobile in the tube. High purity spherical silica
phase through the column to transport gels are sometimes sized by elutriation.
solutes down the column. Emulsion: A mixture of two or more
Elution chromatography: The most liquids that are normally immiscible
commonly used chromatographic (nonmixable or unblendable). Emul-
method where the sample is applied to sion is usually referred to when both
the head of the column as a narrow zone the dispersed and the continuous phase
and individual molecules are separated are liquids. In an emulsion, one liquid
and eluted at the end of the column (the dispersed phase) is dispersed in the
under the influence of a directed flow other (the continuous phase). Emulsions
of mobile phase. Compare to displace- are bothersome in LLE because they
sometimes are hard to break so that isolating a compound of interest; sol-

the layers can be separated and further vent evaporation is the most often used
treated, if necessary. There are numer- sample preparation technique.
ous ways to break emulsions. Evaporative light scattering detec-
Enantiomeric compound: Chemical tion (ELSD): The effluent from the LC
compounds that display chiral activity; column is nebulized and then vapor-
such compounds will require a separa- ized in a heated drift tube. This process
tion mechanism that can differentiate results in a cloud of analyte particles
between the R- or S-enantiomer. Spe- that passes through a beam of light; the
cialty columns are available for this particles scatter the light and generate
purpose. a signal at a photodiode or photomulti-
Endcapping: A technique used to re- plier; it is a universal detection method
move silanol groups on silica gel that where molecules are not required to
may remain after reaction with a large have a chromophore, be fluorescent, or
silylating agent such as octadecyltri- be electrochemically active.
chlorosilane. The column is said to Exchange capacity: See ion-exchange
be endcapped when a small silylating capacity.
reagent (for example, trimethylchlo- Excluded volume: See interstitial
rosilane and dichlorodimethylsilane) volume.
is used to bond residual silanol groups Exclusion chromatography: See ste-
on a silica gel-based packing surface. ric-exclusion chromatography and
Most often used with reversed-phase ion exclusion.
packings to minimize undesirable Exclusion limit: In SEC, the upper limit
adsorption of basic, ionizable, and of molecular weight (or size) beyond
ionic compounds. For polymeric which molecules will be eluted at the
phases with terminal silanol groups, same retention volume, called the exclu-
endcapping reactions are also used to sion volume. Many SEC packings are
remove them. referred to by their exclusion limit. For
Endfitting: The fitting at the end of example, a 105 column of porous silica
the column that permits connection to gel will exclude any compounds with a
the injector or detector. Most HPLC molecular weight over 100,000, based
endfittings contain a frit to hold the on a polystryene calibration standard.
packing and have a low dead volume Exclusion volume (Vo): The minimum
for minimum band spreading. They retention volume of a molecule on an
are usually constructed of stainless steel SEC packing where all molecules larger
but PEEK and other polymeric materi- than the size of the largest pore are to-
als are also used. tally excluded. These molecules are inca-
Equilibration: See column equilibration. pable of penetrating the pores and elute
Equivalent conductivity: The equiva- at the interstitial (interparticle) volume
lent conductivity is a function of the of the column.
concentration and the limiting conduc- Exponentially modified Gaussian
tivity (equivalent conductivity at infinite peak: A theoretical model for peak
dilution). asymmetry arising from the column,
Evaporation: The process of removing inlet, and detector. The basis for the
a volatile compound for the purposes of Foley-Dorsey equations.
External calibration: Calibration mode (d p = 1.5–5 µm). Separation times in

using one or more standard solutions to the range of minutes, sometimes sec-
establish the calibration function (cali- onds, are common; sometimes referred
bration curve). With this function the to as ultrafast LC.
concentration is calculated out of the Fast protein liquid chromatography
measured peak area (or height) of the (FPLC): A term coined to cover the spe-
sample chromatogram. Linear or qua- cific use of HPLC for the separation of
dratic regressions are mainly applied but
proteins. Generally, glass columns, mod-
other regression modes are possible as erate pressure, and spherical microbeads
well (for example, polynomial, point- are used for FPLC.
to-point). FFAP: Free fatty-acid phase.
Extracolumn effects: The total band- Fiberglass disks: A format where SPE
broadening effects of all parts of the particles are embedded in a fiberglass
chromatographic system outside of matrix; the disk format is especially
the column itself. Extracolumn effects useful for processing large volumes of
must be minimized to maintain the ef- sample (for example, water) given that
ficiency of the column. Areas of band the larger cross-sectional area allows for
broadening can include the injector, a higher flow rate than can be used for
injection volume, connecting tubing, a typical cartridge.
endfittings, frits, detector cell volume,
Filter funnel: A filter funnel is a
and internal detector tubing. The vari-laboratory funnel used for separating
ances of all of these contributions aresolids from liquids via the laboratory
additive. process of filtering. To achieve this,
Extracolumn volume: The volume be- a disk-shaped piece of filter paper is
tween the effective injection point andusually folded into a cone and placed
the effective detection point, exclud- within the funnel. The suspension
ing the part of the column containing of solid and liquid is then poured
the stationary phase. It is composed ofthrough the funnel. The solid par-
the volumes of the injector, connectingticles are too large to pass through
lines and frits, and the detector; it de-
the filter paper and are left on the
termines the extra column effects. See paper, while the much smaller liquid
dead volume. molecules pass through the paper to
Extraction: The general term for remov-a vessel positioned below the funnel,
ing analytes of interest from a matrix.producing a filtrate. The filter paper is
used only once. If only the liquid is of
F interest, the paper is discarded; if the
FAME: Fatty acid methyl ester. suspension is of interest, both the solid
Fast GC: Gas chromatography per- residue on the paper and the filtrate
formed on short, narrow-bore open- are kept for further analysis.
tubular columns, or on conventional Filter holder: Membrane filters or
columns at elevated linear velocities. membrane disks are sometimes fur-
Fast LC: The use in HPLC of short nished loose and can be placed in a
columns (1.5–7 cm in length) with holder (usually of stainless steel con-
conventional internal diameters ( struction) for processing samples; after
2–6 mm) packed with small particles the filter is used the holder is opened
and the used filter is replaced with a heteroatomic solutes in a hydrogen-air

fresh one. flame. The visible-range atomic emis-
Filter porosity: Pore size relates to the sion spectrum is filtered through an
filter’s ability to filter out particles of interference filter and detected with a
a certain size. For example, a 0.5-µm photomultiplier tube. Different inter-
membrane will filter out particles with ference filters may be selected for sul-
a diameter of 0.5 µm or larger from a fur, tin, or phosphorus emission lines.
filtration stream. Filter porosity is typi- FPD is a sensitive and selective detec-
cally not related to, nor controlled by, tion method.
pore size. These two parameters are es- Flash chromatography: A very fast
sentially independent. Porosity is also form of classical liquid chromatography
unrelated to thickness. Rather, it is a used by synthetic organic chemists to ef-
function of the polymer and casting fect rapid purification. Done primarily
process used in the manufacture of in the normal-phase mode, sometimes
the filter. with reversed-phase LC. See column
Filter vial: A filter vial is a membrane chromatography.
filter unit that consists of two pieces. Flexible well plate: See array 96-well
One is the filtration plunger, which con- plate.
tains a membrane filter suitable for the Flow injection extraction: An on-line
solvent being filtered. The second part extraction technique where a sample is
of the filter vial is the vial body itself; injected as a plug into an aqueous flow
once the sample is loaded and filtered, stream that is divided into small seg-
the filter vial can be placed directly in ments by an organic phase; the aqueous
the autosampler without transferring the and organic segments are mixed during
filtered sample to another vial. their passage down a coil, and eventu-
Filtration: The process of passing a liq- ally the phases are separated at the end
uid through a paper, membrane, glass, by a special fitting and the amount of
or other type of filter for the purposes of extract compound can be measured in
removing particulates that could cause the organic phase.
problems downstream during a chro- Flow programming: Used to decrease
matographic analysis; a chemical filter the retention time of slow-moving com-
also removes certain chemical species. pounds. Flow programming is occasion-
See chemical filtration. ally used in concert with temperature
Fixed-well plate: A 96-well plate with programming in GC.
fixed (nonremovable) wells, not an array. Flow rate (F, Fc): The volumetric rate of
See 96-well plate and array 96-well flow of mobile phase through a column.
plate. For a conventional 4.6-mm i.d. HPLC
Flame ionization detection (FID): A column, typical flow rates are 1–2 mL/
flame ionization detector ionizes hydro- min. GC packed column flow rates
carbon-containing solutes in a hydro- typically range from 10 to 40 mL/min,
gen–air flame. FID is a nearly universal and open-tubular column flows range
detection method that responds strongly from less than 1 mL/min up to about
to most classes of organic compounds. 10 mL/min.
Flame-photometric detection (FPD): Flow resistance parameter (Φ): Φ =
A flame-photometric detector burns dp2/Bo See permeability.
Fluoro phase: One of a family of Freeze drying: The process of removing

aliphatic and aromatic reversed-phase water, mainly from biological samples,
materials in which a substantial frac- by using vacuum sublimation.
tion of the bonded phase is fluorinated. Frictional heating: Viscous heating of
Sometimes called fluorous phases or solvent molecules passing through very
perfluoro phases. Typically have dif- small-diameter (micrometer) particles;
ferent selectivities than hydrocarbon causes a rise in temperature over the
phases. length of the column; reduced by using
Fluted filter paper: Filter paper that smaller-diameter columns (smaller heat
is folded in a systematic manner to dissipation) and superficially porous
allow more air space in the filter fun- particles, which also have improved heat
nel; this allows liquid to f low faster dissipation.
through the filter paper. Frit: The porous element at either end
Foley-Dorsey equation: A correc- of a column that serves to contain the
tion of the plate count and retention column or SPE packing. It is placed at
time to correct for peak tailing from the very ends of the column tube or in
extracolumn sources of broadening. the end fitting. Frits can be stainless
(See J.P. Foley and J.G. Dorsey, Anal. steel or another inert metal or plastic
Chem. 55, 730–737 [1983].) such as porous PTFE or polypropylene.
Forced-flow leaching: Solid mate- The frit porosity must be less than the
rial is packed into stainless steel col- smallest particle in the column; other-
umn and toluene (or other extraction wise particles will pass through the frit
solvent) is pumped into the column and packing will be lost.
under pressure and with heating; hot Frontal analysis: A chromatographic
solvent leaches (extracts) out extract- technique that involves continuous ad-
able compounds which are collected at dition of sample to the column with
the exit of the column. the result that only the least sorbed
Fraction of analyte extraction (E): compound, which moves at the fast-
The fraction of analyte extracted: E est rate, is obtained in a pure state.
= (C oVo)/(C oVo + C aqVaq) = (K DV )/(1 + The second-least-sorbed compound
K DV ) where Vo is the volume of or- is eluted with first eluted compound,
ganic phase, Vaq the volume of aque- the third-least-sorbed compound with
ous phase, and V is the phase ratio the first and second compound, and so
Vo/Vaq. forth until the original sample passes
Fractionation range: In SEC, refers through the column exit. Frontal
to the operating range of a gel or pack- analysis is seldom used and is mainly
ing. This is the range in which the a preparative technique.
packing can separate molecules based Frontal chromatography: The same as
on their size. At one end of the range, frontal analysis.
molecules that are too large to diffuse Fronting: Peak shape in which the front
into the pores are excluded. At the part of the peak (before the apex) in a
other end of the range, molecules that chromatogram tapers in advance of the
can diffuse into all of the pores totally remainder of the peak (that is, the front
permeate the packing are eluted (un- is less steep than the rear). There is an
separated) at the permeation volume. asymmetric distribution with a leading
edge. The asymmetry factor for a front- Gaussian peak: The equation of a
ing peak has a value less than 1. The Gaussian distribution can be written in
opposite effect is tailing. Fronting can terms of chromatography parameters as
result at high sample loads because of C = Cmaxexp(–(t – tR )2/2σ2)
positive curvature in the isotherm and Gel: The solid packing used in gel
from use of poorly packed columns. See chromatography or GPC. An actual gel
asymmetry factor. consists of two parts: the dispersed me-
Fused-core packing: See superficially dium (solid portion) and the dispersing
porous particles (SPPs). medium (the solvent). Also defined as
Fused silica: Open-tubular GC col- a colloidal dispersion of a solid and liq-
umns and LC nanocolumns are com- uid in which the solid is the continuous
monly manufactured from fused-silica phase.
tubing that is coated externally with a G e l f i l t ra t i o n c h r o m a to g ra p hy
protective polymeric material such as (GFC): Also called aqueous size-ex-
polyimide. clusion chromatography; carried out
Fused-silica open-tubular column with aqueous mobile phases. Gener-
(FSOT): Open-tubular GC columns ally refers to molecular size separa-
made of fused silica. See open-tubular tions performance on soft gels such as
column. polydextrans but highly crosslinked
polymers, silica gels, and other po-
G rous media may also be used. Most
Gas–liquid (phase) chromatography gel filtration separations involve bio-
(GLC, GLPC): Solutes partition between polymers and water-soluble polymers
a gaseous mobile phase and a liquid such as polyacrylic acid.
stationary phase. Selective interactions Gel permeation chromatography
between the solutes and the liquid phase (GPC): Size-exclusion chromatography
give rise to different retention times in (SEC) carried out with organic mobile
the column. phases used for the separation and
Gas-phase extraction: See direct ther- characterization of polymers. SEC with
mal sampling. aqueous mobile phases is referred to as
Gas–solid chromatography (GSC): Sol- gel filtration chromatography.
utes partition between a mobile gaseous Ghost peaks: Peaks not present in the
phase and a solid stationary phase. Se- original sample. Ghost peaks can be
lective interactions between the solutes caused by septum bleed, solute decom-
and the solid phase give rise to different position, or carrier-gas contamination.
retention times in the column. Gigapores: See perfusion chroma-
Gaussian curve: A standard error curve, tography.
based on a mathematical function, that Golay equation: M.J.E Golay formu-
is a symmetrical, bell-shaped band or lated an equation for the theoretical
peak. Most chromatographic theory plate height versus the average linear
assumes a Gaussian peak. Use of the velocity of open-tubular (capillary) col-
peak maximum position as a measure umns. The Golay equation is similar to
of retention and equations mentioned the van Deemter equation, except that
above for efficiency assume a Gaussian the A term is dropped because there is
peak shape. no column packing, and the B and C
terms are modified accordingly as well: heat treatment; this packing material
h = (B/u–) + u– (C M + C S) has a strong affinity for polar com-
Grab sampling: In gas sampling, an pounds in aqueous samples and water
evacuated sample canister can be miscible organic extracts. Commonly
opened and sample rapidly collected used in pesticide analysis of food sam-
at an uncontrolled rate, usually over ples; also known as graphitized carbon
several seconds, until the container black (GCB). Also used as a GC station-
reaches equilibrium with atmospheric ary phase.
pressure. Generally this qualitative ap- Graphitized carbon packing: A re-
proach is used when unknown analytes versed-phase packing material which
must be identified, when the air con- consists of pure graphitic carbon;
tains high concentrations of analytes possesses interesting sorbent proper-
at certain (short) times, or when an ties such as preferential separation of
odor is noticed and a sample must be geometric isomers such as o-, m- and
obtained quickly. Paired grab samples p-aromatics and cis–trans isomers.
(before–after or smell–no smell) often Grinding: Both manual and automated
are used to qualitatively diagnose a per- mortar and pestles are available; grind-
ceived problem. ing can be performed under wet or dry
Gradient: A process to change solvent conditions; by this process particles of
strength as a function of time (normally approximately 10 µm can be achieved.
solvent strength increases) thereby elut- Guard column: A small column placed
ing progressively more highly retained between the injector and the analyti-
analytes; typically gradients can be bi- cal column. It protects the analytical
nary, ternary, and quaternary solvent column against contamination by sam-
mixtures where solvents are blended ple particulates and strongly retained
to achieve the proper strength. In GC, species. The guard column is usually
an older term that refers to column packed with the same material as is in
temperature programming to achieve the analytical column and is often of
a similar effect. the same inner diameter. Its length is
Gradient delay volume: See dwell much shorter, it costs less, and is usu-
volume. ally discarded when it becomes con-
Gradient elution: Technique for de- taminated. Integrated guard–analytical
creasing separation time by increasing column systems are often preferred to
the mobile-phase strength over time minimize extracolumn effects caused
during the chromatographic separa- by the use of connecting tubing with
tion. Also known as solvent program- separate guard and analytical columns.
ming. Gradients can be continuous or For GC, see also retention gap.
stepwise. Binary, ternary, and quater-
nary solvent gradients have been used H
routinely in HPLC. Headspace sampling: Gas-phase sam-
Graphitized carbon: Graphitized car- pling technique in which solute is re-
bon is a graphitic carbon with more moved from an enclosed space above a
or less perfect three-dimensional hex- solid or liquid sample. The headspace
agonal crystalline order prepared from refers to the vapors that form above
non-graphitic carbon by graphitization liquids and solids; if the sample is in
thermodynamic equilibrium with the that ionizes solute molecules, which are
gas phase in a closed thermostated measured with an electrometer. Meta-
vessel, the method is called static stable He may also be harnessed in an
headspace sampling; if an inert gas electron-capture mode for halogen-
passes over or through the sample and specific response. See also pulsed dis-
stripped sample volatiles accumulate charge detection.
in an adsorbent or cryogenic trap, the Helium sparging: See degassing. He-
method is dynamic headspace or purge- lium has a very low solubility in most
and-trap sampling. common liquids.
Headspace single-drop microextrac- High performance capillary electro-
tion (HS-SDME): A single drop of sol- phoresis (HPCE): A technique where
vent (1–2 µL) suspended in the head- small diameter capillaries, buffered
space can partition volatile analytes conducting solutions, and high voltage
into the solvent; the drop can be with- (up to 30,000 V) are used to separate
drawn into the syringe and injected ionic molecules based on their differ-
into a GC system. ential electrophoretic mobilities; non-
Headspace solid-phase microextrac- ionic (neutral) molecules can be sepa-
tion (HS-SPME): Instead of a drop of rated by MEKC.
solvent, a polymer-coated fiber can be High performance liquid chromatog-
placed in the headspace and once the raphy (HPLC): Modern liquid-phase
analytes adsorb on the polymer coat, chromatography technique using small
the fiber can be transferred to a GC particles and high pressures.
inlet and the sorbed analytes volatilized High-pressure mixing: A configura-
by thermal desorption. tion of a gradient HPLC system where
Heart cutting: In preparative LC, re- the solvents are mixed on the high-
fers to collection of the center of the pressure side of multiple pumps (usu-
peak where purity should be maximum. ally two, binary); such a system offers
The term is also used for analytical col- a lower gradient delay volume than
umn switching for LC or GC in which low pressure mixing systems where the
two or more partially resolved peaks solvents are mixed by proportioning
eluted from one column are directed valves before a single pump.
onto another column of different po- High-abundance protein depletion:
larity, or at a different temperature, for By using antibody columns specific
improved resolution. for the highest abundance proteins,
Helium ionization detection (HID): they can be selectively removed from
An ionization detection method for plasma. This process enables investiga-
GC. Helium is ionized by a radioac- tion of the lower abundance proteins,
tive or other energetic source; the re- which may be biomarkers or other com-
sulting He2+ ions interact with solutes pounds of interest.
to produce ions that measured with a Hold-up time (t M , t 0 ): The time re-
sensitive electrometer. quired for an unretained compound
Helium plasma or microwave-in- to be eluted, or the time required for
duced detection (HPD): A helium one column volume (V M ) of mobile
plasma is created in a microwave RF phase to pass through the column. In
field. The plasma emits UV radiation reversed-phase LC, uracil is often used
to measure hold-up volume and hold- Homogenization: The process of mak-

up times. In GC, methane or another ing a sample more uniform by size re-
unretained compound is used to mea- duction and blending; homogenizers
sured the column hold-up time. Also with high speed blades are available to
called the unretained peak time or dead do the job.
time. Hybrid silica: Silica gel consisting of
Hold-up volume (V M or V 0 ): The both organic and inorganic moieties
total volume of mobile phase in the with hybrid properties of polymeric
column regardless of where it exists. packings and silica packings; synthe-
In LC: The hold-up volume consists sized from silanes containing organic
of the entire space accessible to a functionality; different selectivity but
small molecule able to fully permeate higher high pH stability than bare or
all the pores of the packing material. uncoated silica gel.
It comprises the interstitial volume Hydrodynamic volume: The molecu-
and the unoccupied pore volume. It lar volume defined by the effective di-
is denoted as Vo or V M . The system ameter of a molecule in free solution
hold-up volume includes the ad- where the hydrodynamic sphere would
ditional volume in the tubing used be a sphere defined by the molecule
to connect the injector and detector as it revolves around its central axis in
to the column. The hold-up volume solution; termed used in size-exclusion
is usually approximated by inject- chromatography to define molecular
ing a small essentially unretained shape and to explain why molecules
species. In reversed-phase LC, ura- with the same molecular weight often
cil, acetone, and thiourea are most have totally different elution volumes;
commonly used: V M = t M F c . In GC: measured by determining the Stokes
The gas-phase volume of the column radius.
that corresponds to the hold-up time. Hydrophilic: “Water loving”; refers
Measured by injecting an unretained both to stationary phases that are fully
species such as methane that fits in compatible with water or to water-sol-
all the pores. See hold-up time. uble molecules in general. Many col-
Hold-up volume, corrected (V M0 ): umns used to separate proteins (ion ex-
The hold-up volume corrected for change, SEC, affinity) are hydrophilic
carrier-gas expansion along the col- in nature and should not irreversibly
umn: V M0 = V j sorb or denature protein in the aque-
Hollow-fiber liquid-phase microex- ous environment.
traction (HF-LPME): A hollow-fiber Hydrophilic interaction liquid chro-
(HF) membrane technique where an matography (HILIC): An alternative
HF membrane separates two extrac- technique to reversed-phase HPLC
tion phases; the membrane serves as a for the separation of highly polar
barrier and can be impregnated with analytes that may be only slightly re-
solvent to permit liquid–liquid or liq- tained or unretained by reversed-phase
uid–liquid–liquid extractions to take LC, HILIC requires a high percent-
place; the membrane can be selected to age of a nonpolar mobile phase and
allow certain analytes to pass through a polar stationary phase, similar to
but not others. the requirements in normal-phase
chromatography. However, unlike chromatography; usually little or no

normal-phase chromatography, which organic solvent is used in the mobile
uses nonpolar solvents such as hexane phase in HIC.
and methylene chloride and tries to Hydrophobic subtraction model:
exclude water from the mobile phase, Developed by Lloyd Snyder and John
HILIC requires some water in the Dolan, this model is used to charac-
mobile phase to maintain a stagnant terize reversed-phase columns; using
enriched water layer on the surface five types of probes, based on their
into which analytes may selectively equations, they can predict if a cer-
partition. In addition, water-miscible tain column will be close in selectiv-
organic solvents are used instead of ity characteristics to another column
the water-immiscible solvents used in or totally different (orthogonal); over
normal-phase chromatography. With 300 columns have been characterized
HILIC, sorbents such as bare silica, using this model. (See L. Snyder and
bonded diol, and polyhydroxyethylas- J. Dolan, LCGC 20[11], 1016–1026
partamide are used. Polar analytes are [2002].)
well retained and are eluted in order Hydroxyapatite: A porous calcium
of increasing hydrophilicity, just the hydroxyphosphate solid that chemi-
inverse of reversed-phase LC. cally resembles bone and tooth used
Hydrophobic: “Water hating”; re- as a packing material used in bio-
fers both to stationary phases that chromatography for nucleic acid con-
are not compatible with water or to stituents, monoclonal antibodies, and
molecules in general that have little proteins.
a f f init y for water. Hydrophobic Hypercrosslinking: Mainly refers to
molecules have few polar functional a new way to synthesize a polymeric
groups; most have a high content of monolith; hypercrosslinked monolithic
hydrocarbon (aliphatic and aromatic) capillary columns contain an array of
functionality. small pores and have very high surface
Hydrophobic interaction chroma- areas.
tography (HIC): A technique in which Hyphenated techniques: Refers to
weakly polar (nonhydrocarboneous) the family of techniques best known
packings are used to separate molecules by their abbreviations, including LC–
by virtue of the interactions of their hy- MS, LC–FTIR, and MS/MS.
drophobic moieties and the hydropho-
bic sites on the packing surface. High I
concentrations of salt solutions are used Immobilized liquid extraction: Simi-
in the mobile phases and separations lar to SPE but a polymeric stationary
are affected by changing the salt con- phase is bonded to the inside of a glass
centration. The technique is analogous vial; analytes partition into polymeric
to “salting out” molecules from solu- phase and loading, washing, and elu-
tion. Gradients are run by decreasing tion steps are performed by addition of
the salt concentration; often used for various solvents.
the separation of proteins that are sen- Immobilized metal affinity chroma-
sitive to denaturization by the organic tography (IMAC): See metal affinity
solvents used in regular reversed-phase chromatography.
Impinger: Impingers are glass bubble analytes; a UV-absorbing or f luores-

tubes designed for the collection of cent compound is added to the mo-
airborne hazards into a liquid medium. bile phase that maintains a high back-
When using a personal air sampler, a ground signal; when a nonabsorbing
known volume of air bubbles is pumped or nonf luorescing analyte is eluted,
through the glass tube that contains a the background is diluted and a nega-
liquid specified in the method. The liq- tive peak is observed for that analyte;
uid is then analyzed to determine air- when an analyte acts to increase the
borne concentrations. An impinger may concentration of the indicating species,
be mounted on the side of an air sample a positive peak is observed. When a
pump or put into a holster and placed negative signal is detected, the detec-
near a worker’s breathing zone. tor signals are reversed to the output
Imprinted phases: Polymer and silica device.
phases generated in the presence of a Infinite diameter column effect
“template” or “printing” molecule. Such (IDE): Name given by John Knox to
phases have enhanced selectivity for the the following phenomenon: At a cer-
templating molecule; also called molec- tain column length, a sample injected
ularly imprinted phases (MIPs). into center of a packed bed spreads by
In-line filter: A device that prevents radial diffusion but never reaches col-
particulate matter from damaging the umn wall, where wall effects can cause
column. Modern low-volume, in-line band broadening. Knox showed that
filters can be placed between the injec- a sample peak collected in the exact
tor and the column without major con- center of the column exit displayed a
tributions to band broadening. A filter higher efficiency than a sample peak
in this position is used to prevent sample collected near the wall. The infinite
particles from entering the packed bed diameter effect depends on column
or column inlet frit. length, internal diameter, particle size,
In situ derivatization: The act of de- and mobile-phase properties. Very sel-
rivatizing a compound of interest in dom applied in HPLC.
its native environment; for example, Injection solvent: Solvent used to inject
for a tenaciously held analyte on a soil sample into an HPLC column; solvent
sample changing its chemical nature should be of equal or lower strength
by performing an in situ derivatization, than the mobile phase to prevent pre-
the compound may be more easily re- mature movement down the column
leased and isolated for further workup due to the presence of a stronger solvent.
or analysis; derivatization may also be Inlet: In LC, the initial part of the col-
formed in solution and the derivatized umn where the solvent and sample enter.
compound extracted by LLE. There is usually an inlet frit that holds
Included volume: Also known as totally the packing in place and, in some cases,
included volume. The volume at which protects the packed bed. In GC, a de-
a small molecule that explores the en- vice between the carrier gas source and
tire pore space of a column is eluted. See the column inlet that transfers sample
steric-exclusion chromatography. from outside the chromatograph into
Indirect detection: Used for non-ul- the column, often by vaporizing the
traviolet-absorbing or nonfluorescing sample. See split injection, splitless
injection, on-column injection, pro- stance being quantitatively determined

grammed temperature vaporizer. and should have a retention time fairly
Inlet filter: Filtration devices attached close to it.
to the inlet lines of the pump that re- Interparticle porosity (ε): The inter-
moves particulate matter from the mo- particle volume of a packed column per
bile phase before the solvent reaches the unit column volume: ε = Vo/Vc. See also
pump; reservoir filters are an inlet filter interstitial porosity.
that resides in the solvent bottle. Interstitial porosity (εe): The fraction of
Inlet liner: Deactivated glass tube in an the volume in the column located in the
inlet system into which liquid sample is interparticle (interstitial) space: εe = Ve/Vc
injected. An inlet liner may be open or Interstitial volume (Ve): The volume
packed with deactivated glass wool, and between the particles. Does not include
it may have various internal structures. the volume in the pores of the particles.
The purpose is to vaporize and disperse Also called the excluded volume (see
evaporating sample into the carrier gas steric-exclusion chromatography)
stream as uniformly as possible while and interparticle volume. Measured by
not causing significant sample break- injecting a molecule which does not per-
down, adsorption, or discrimination. meate any pores and does not interact
Inlet/outlet check valves: The check with the surface of the particles. In SEC
valve (or valves) on an LC pump that this volume is denoted Vo.
allows mobile phase to flow in one di- Intraparticle porosity (εi): The fraction
rection but not in the reverse direction. of the particle volume which lies in the
The inlet check valve allows flow from pores: εi = Vpore/Vparticle
the reservoir into the pump, and the Intraparticle volume (V): The volume
outlet check valve allows mobile phase inside the pores of the particles. Also
to flow to the column from the pump. called the internal volume and included
Instrumental bandwidth: The contri- volume. Can be measured by BET or
bution of the analytical instrument to mercury intrusion porosimetry.
peak broadening; see extracolumn ef- Ion chromatography (IC): An ion-ex-
fects for explanation. change technique in which low concen-
Instrumental dispersion: See extracol- trations of organic and inorganic anions
umn effects. or cations are determined using ion ex-
Internal standard (IS): In quantitative changers of low ion-exchange capacity
analysis, precision and accuracy are with dilute buffers. Conductivity detec-
greatly improved by use of internal stan- tors are often used. Ion chromatography
dards (IS). The procedure involves the is practiced in two forms. In suppressed
addition of a fixed amount of internal IC, a second column or a membrane sep-
standard to a series of increasing con- arator is used to simultaneously remove
centrations of reference sample and to the buffer counterion to the analyte and
the unknown concentration. The ratio replace it with hydrogen or hydroxide
of the areas of the reference concentra- ion, which concomitantly converts the
tions and the areas of the IS is plotted buffer to an uncharged species, thereby
against the known concentration of the suppressing background and enhancing
reference samples. The internal standard sensitivity. In nonsuppressed IC, weakly
should be chemically similar to the sub- conducting buffers at low concentration
are carefully selected and the entire ef- take place in the exchange process. The
fluent is passed through the detector— exchange capacity is expressed in milli-
ions are detected above the background equivalents per gram. A typical styrene-
signal. divinylbenzene strong anion exchange
Ion exclusion: The process in which resin may have 3–5 meq/g capacity.
ionized solutes can be separated from Exchangers for IC have very low capac-
unionized or partially ionized solutes ity. Capacity of weak anion and cation
using ion-exchange resins. Separation exchangers varies dramatically with pH.
results from Donnan potential where Ion-exchange chromatography: A
ionic solutes exist at a higher concen- mode of chromatography in which ionic
tration in solution than in the station- substances are separated on cationic or
ary phase whereas nonionic solutes are anionic sites of the packing. The sample
evenly distributed between the mobile ion (and usually a counterion) will ex-
phase and resin. Therefore, ionic sol- change with ions already on the iono-
utes will move faster down the column genic group of the packing. Retention is
than non-ionic solutes. Ion exclusion is based on the affinity of different ions for
known to take place in reversed-phase the site and a number of other solution
LC when anions are separated at pH val- parameters (pH, ionic strength, coun-
ues where the silanol groups are ionized. terion type, and so forth). Ion chroma-
Ion moderated partitioning chromatography is basically an ion-exchange
tography: A technique used for the technique.
separation of carbohydrates using strong Ion-pair chromatography: Form of
cation-exchange packings that are in chromatography in which ions in solu-
specific cationic form (for example, cal- tion can be “paired” or neutralized and
cium, hydrogen, silver); the separation separated as an ion pair on a reversed-
mechanism is complexation rather than phase column. Ion-pairing agents are
ion exchange. usually ionic compounds that contain
Ion retardation: Refers to the use of a hydrocarbon chain that imparts a cer-
amphoteric ion-exchange resins, which tain hydrophobicity so that the ion pair
are used to retard ionic molecules and can be retained on a reversed-phase col-
allow nonionic molecules or nonelectro- umn. Retention is proportional to the
lytes to be eluted preferentially. length of the hydrophobic chain and the
Ion suppression: Buffering in an aque- concentration of the ion-pair additive.
ous mobile phase at a particular pH to Ion pairing can also occur in normal-
suppress solute ionization. For example, phase chromatography when one part
weak carboxylic acids can have their ion- of the pair is dynamically loaded onto a
ization suppressed by the adjustment of sorbent, but this technique is not as pop-
the pH below their pKa. Useful for im- ular as reversed-phase LC. Also known
proving peak shape of weak acids and as ion-interaction chromatography or
bases in reversed-phase LC. dynamic ion-exchange chromatogra-
Ion-trap detector: Mass spectrometric phy, stressing the fact that the precise
detector that uses an ion-trap device to mechanistic details of how the additive
generate mass spectra. controls retention are not always known.
Ion-exchange capacity: The number Ionic strength: Ionic strength is a
of ionic sites on the packing that can characteristic of an electrolyte solution.
It is typically expressed as the average Isotope-coded affinity tags (ICAT):

electrostatic interactions among an Isotope-coded affinity tags (ICATs)
electrolyte’s ions. It is related to elec- are a gel-free method for quantitative
trolyte concentration, but the main proteomics that relies on chemical la-
difference between ionic strength beling reagents. These chemical probes
and electrolyte concentration is that consist of three general elements: a reac-
the ionic strength is higher if some tive group capable of labeling a defined
of the ions are more highly charged. amino acid side chain (for example,
The higher the ionic strength of a mo- iodoacetamide to modify cysteine resi-
bile phase the more the mobile phase dues), an isotopically coded linker, and
competes with the analyte for ionic or a tag (for example, biotin) for the af-
adsorptive sites. finity isolation of labeled proteins and
Irregular packing: Refers to the shape peptides. For the quantitative compari-
of a column packing. These irregular son of two proteomes, one sample is
packings are available in micropartic- labeled with the isotopically light (d 0)
ulate sizes. The packings are obtained probe and the other with the isotopi-
from grinding solid materials into cally heavy (d 8) version. To minimize
small particles and then sizing them error, both samples are then combined,
into narrow fractions using classifica- digested with a protease (such as tryp-
tion machinery. Spherical packings are sin), and subjected to avidin affinity
now used more than irregular pack- chromatography to isolate peptides
ings in analytical HPLC but the less- labeled with isotope-coded tagging
expensive irregular packings are still reagents. These peptides are then ana-
widely used in preparative LC. lyzed by liquid chromatography-mass
Irreversible adsorption: When a spectrometry (LC–MS). The ratios of
compound that has a very strong af- signal intensities of differentially mass-
finity for an adsorbent is injected tagged peptide pairs are quantified to
onto a column, it can be adsorbed so determine the relative levels of proteins
strongly that it cannot be eluted from in the two samples. The original tags
the column. A chemical reaction be- were developed using deuterium, but
tween the sample and the surface of later the same group redesigned the tags
the adsorbent is an example of irre- using 13C instead to circumvent issues
versible adsorption. of peak separation during LC because
Isocratic: Use of a time-invariant eluent of the deuterium interacting with the
composition in LC. stationary phase of the column.
Isolate: Analyte to be isolated from ma-
trix background and then analyzed. K
Isotherm: See adsorption isotherm. Kieselguhr: A diatomaceous earth used
Isothermal chromatography: Use of both in column chromatography and as
conditions of constant column tem- a sample cleanup media. Only weakly
perature. The vast preponderance of adsorptive, it is also used as a support
all LC is done under isothermal condi- in liquid–liquid chromatography and
tions, while most GC separations are in supported liquid extraction, see sup-
performed with column temperature ported liquid extraction (SLE). Rarely
programming. used in HPLC.
Kilopascal (kPa): A unit of pressure. normal volumes of sample in a solvent

100.0 kPa = 1.0 bar = 0.9869 atm. into a capillary GC column; in this
101.325 kPa = 1.0 atm. approach, the bulk of the solvent is
Kinetic plot: Kinetic plots are methods evaporated before the sample transfers
to characterize the practical limits of to the analytical column; there are two
column performance, where theoretical popular LVI techniques: programmed
plates (H) and separation impedance (E) temperature vaporization and cool on-
are plotted as a function of the pressure- column injection with solvent vapor exit;
drop limited plate number (N). The ki- both are approaches to lowering detec-
netic plot retains the information shown tion limits.
in van Deemter plots but completes it Ligand: In ligand-exchange chromatog-
with the information on the bed perme- raphy, refers to the analyte that under-
ability. See Poppe plot. goes ligand exchange with the stationary
Knox equation: A modification of the phase. In affinity chromatography, re-
van Deemter equation developed by fers to the biospecific material (enzyme,
John Knox that uses reduced terms, in antigen, or hormone) coupled to the
which the A term that represents eddy support (carrier) to form the affinity col-
dispersion is replaced with Aν 1⁄3 where ν umn. In bonded-phase chromatography
is the reduced interstitial eluent velocity. this term denotes the moiety covalently
bound to the surface.
L Ligand-exchange chromatography:
Laminar flow: The smooth time-in- A technique in which chelating li-
variant flow that develops when a liq- gands are added to the mobile phase
uid is moving under conditions where and undergo sorption onto a pack-
viscous forces dominant over inertial ing. These sorbed molecules can act
forces. Laminar flow is characterized as chelating agents with certain sol-
by a low Reynolds number. For flow utes. An example would be the use of
in a cylindrical tube fluid stream-lines copper salt added to the mobile phase
near the center move faster than those for the chelatation and separation of
at the tube wall which are static result- amino acids. Chelating resins function
ing in a radially parabolic distribution in in a similar manner, where chelating
axial fluid velocity. This nonuniformity groups are chemically bonded to the
of axial velocities in the interstices in a polystyrene backbone.
packed bed also leads to substantial peak Limit of detection (LOD): The concen-
broadening in packed columns. tration of the analyte at which the re-
Langmuir adsorption isotherm: A spe- sulting peak can be distinguished from
cific form of an isotherm CS = (N0CM)/ baseline noise. Literature and norms
(K d + C M ) where C S and C M are the describe different ways of determining
equilibrium stationary and mobile the LOD.
phase concentrations of the solute, N0 Limit of quantitation (LOQ): The
the total number of surface sites avail- minimum concentration of the ana-
able for sorption and Kd the sorption lyte at which the resulting peak can
binding constant. be quantified with a defined level of
Large-volume injection (LVI): A tech- certainty. Typically 3–5 times higher
nique for introduction of larger than than the LOD.
Linear elution adsorption chromatog- found in the organic solvent; by the

raphy (LEAC): A term coined by Lloyd use of additives (for example, buffers
Snyder; refers to adsorption chromatog- or ion-pair reagents), equilibria can be
raphy carried out in the linear portion shifted to “force” analytes or matrix
of an adsorption isotherm; sometimes compounds into one or other of the
referred to as linear chromatography. two layers.
Linear range (LR): Also, linear dynamic Liquid-phase microextraction (LPME):
range. The range of solute concentration A liquid extraction technique where
or amount over which detector response there is a great reduction in the accep-
per solute amount is constant within a tor-to-donor phase ratio; a hollow fiber
specified percentage. is impregnated with an organic solvent
Linearity: In quantitative analysis, it is used to accommodate or protect micro-
essential for the detector to yield a linear volumes of acceptor solution. This novel
response with respect to solute concen- methodology proved to be an extremely
tration; some detectors may show non- simple, low-cost, and virtually solvent-
linear performance in certain concentra- free sample-preparation technique that
tion ranges, especially on the high end provided a high degree of selectivity and
but also on the low end. enrichment by additionally eliminating
Liquid chromatography: A separation the possibility of carryover between runs.
technique in which the mobile phase Liquid–solid chromatography (LSC):
is a liquid; most often carried out in a Same as adsorption chromatography.
column. Liquid–solid extraction: The general
Liquid phase: In GC, a stationary liq- expression for extraction techniques
uid layer coated on the inner column that uses an organic solvent to extract
wall (WCOT) or on a support (packed, analytes from a solid material. In its
PLOT, SCOT) that selectively interacts simplest form, the “shake flask” extrac-
with different solutes to produce differ- tion takes place at room temperature
ent retention times. Also refers to the and works well for the case where the
stationary phase in LLC. matrix is porous and the analytes are
Liquid–liquid chromatography (LLC): easily extractable.
Same as partition chromatography. Loadability: The maximum amount of
One of the earliest separation modes analyte that can be injected onto a col-
of HPLC; it gave way to chemically umn above which it no longer permits
bonded phases in the early 1970s. the isolation of product at the desired
Liquid–liquid diffusion coefficient level of purity or recovery level; impor-
(DL): See diffusion coefficient. tant in preparative chromatography.
Liquid–liquid extraction (LLE): LLE is Loading (phase loading versus sam-
an extraction technique for separating ple loading): The amount of stationary
interferences from the analytes by par- phase coated or bonded onto a solid sup-
titioning the analytes between two im- port. In liquid–liquid chromatography,
miscible liquids (or phases); one phase is the mass of liquid phase per gram of
usually aqueous and the second phase an packing. In bonded-phase chromatog-
organic solvent; more hydrophilic com- raphy, the loading may be expressed in
pounds prefer the aqueous phase while µmol/m2 or %C (w/w); also called cov-
more hydrophobic compounds will be erage or surface coverage. An alternate
(unrelated) meaning is the amount of the polymer matrix. Bioreactive mol-

sample mass that is injected on an ana- ecules can be adsorbed or coupled to
lytical or preparative column; prepara- their surface, and used to separate
tive columns are often operated in an biological materials such as cells, pro-
overloaded condition for throughput teins, or nucleic acids; by the use of
reasons. magnets or magnetic fields, the beads
Loading step (SPE): The second step can be easily manipulated in test tubes
in SPE (after conditioning) where the or 96-well plates. These microbeads
sample is loaded onto the SPE phase are used for isolation and handling of
(cartridge). specific material or molecules, as well
Log KW: In reversed-phase LC, the ex- as for analyzing sensitive molecules or
trapolated intercept of a plot of log k ver- those that are in low abundance (for
sus volume fraction of organic modifier example, in miniaturized and auto-
in reversed-phase LC. See also solvent mated settings).
strength (S). Make-up gas: Extra carrier gas or
Longitudinal diffusion: Same as mo- other gas added to the carrier gas as
lecular diffusion term; B term in van it f lows into or through a detector.
Deemter equation; see van Deemter Make-up gas serves to improve peak
equation. shapes for open-tubular columns with
Low pressure mixing: See high pres- detectors not necessarily designed for
sure mixing. them exclusively by reducing the ef-
Lyophilization: The process of dehy- fects of detector dead volume. Also,
drating a sample, often biological, con- make-up gas may play an active role
taining water by using vacuum sublima- in detector operation, as for example
tion; also referred to as freeze drying. when hydrogen serves as both make-
up and combustion gas in a flame ion-
M ization detector.
Maceration: The process of breaking Mass spectrometric (MS) detector:
down a soft material into smaller parts Chromatography detector that records
by tearing, chopping, cutting, and so mass spectra of solutes as they are eluted
forth. from the column.
Macroporous resin (macroreticular): Mass transfer (inter-phase) (C term):
Cross-linked ion-exchange resins that The process of solute movement be-
have both micropores of molecular di- tween the moving and stationary zones.
mensions but also macropores of sev- The C term of the van Deemter equa-
eral hundred angstroms wide. These tion is referred to as the interphase
are highly porous resins with large mass transfer term. The faster the
internal surface area accessible to large process of mass transfer the better the
molecules. efficiency of column. In HPLC, slow
Magnetic-bead technology: Micro mass transfer is the most important fac-
magnetic beads are uniform polymer tor affecting column efficiency. Its rate
particles, typically 0.5–500 µm in di- can be increased by the use of small-
ameter, that have iron oxide particles particle packings, thin layers of station-
(or other particles that may be at- ary phase, low viscosity mobile phases,
tracted to magnets) contained within and high temperatures.
Matrix: In sample preparation, the ma- pane, and pyridine are taken together
trix normally refers to the substance in to express the overall phase polarity,
which the analyst is attempting to mea- or separately to express the station-
sure a solute or series of solutes; often the ary-phase behavior toward individual
matrix is of no interest, and its concen- compound classes.
tration must be reduced or eliminated Mean pore diameter: The average
for a separation and measurement to pore diameter of the pore in a porous
take place; the matrix can be organic, packing. It is most commonly deter-
inorganic, or biological. mined by BET analysis and is reported
Matrix adsorption mode (SPE): A lesser as four times the specific pore volume
used mode of SPE where the sorbent is divided by the specific surface area
chosen to maximize retention of the based on the assumption of uniform
matrix and other interferences while the cylindrical pores. The pore diameter is
analyte of interest is unretained; the op- important in that it must allow free dif-
posite of the normal process (bind–elute) fusion of solute molecules into and out
of SPE; there is no concentration of the of the pore so that the solute can inter-
analytes. act with the stationary phase. Addition-
M a t r i x s o l i d p h a s e d i s p e r s i o n ally, the pores must be well-connected,
(MSPD): Technique uses bonded phase with a minimum of dead ends, so that
solid supports as abrasives to produce there are many paths that can allow a
disruption of sample architecture and molecule to access any part of the pore
as a bound solvent to aid complete space. In SEC, the packings have dif-
sample disruption during the sample ferent pore diameters and therefore
blending process; the finely divided molecules of different sizes can be
sample is gently blended with a mor- separated. For a typical substrate such
tar and pestle, transferred to a column, as silica gel, 60- and 100-Å pore diam-
and the analytes eluted with appropri- eters are most popular. For packings
ate solvents. used for the separation of biomolecules,
Matrix-solid phase extraction (MASE): pore diameters > 300 Å are used.
See matrix solid phase dispersion. Megapascal (MPa): A unit of pressure; 1
Maximum allowable operating tem- MPa = 10 bar, 10.133 atm, or 145.0 psi.
perature (MAOT): Highest continuous Megapores: See perfusion chroma-
column operating temperature that will tography.
not damage a column, if the carrier gas Membrane extraction with sorbent
is free of oxygen and other contami- interface (MESI): A version of dy-
nants. Slightly higher temperatures may namic headspace where a silicone hol-
be permissible for short periods of time low fiber membrane is placed in the
during column bake-out. headspace about the sample; an inert
McReynolds constants: System for gas is passed through the membrane
stationary-phase characterization. and analytes that are permeable to
McReyonolds expanded on the ear- the membrane pass from the head-
lier Rohrschneider polarity probes. space and are swept to an adsorbent
The retention indices of a series of trap; after a period of concentration,
test probes such as benzene, 1-buta- the trapped analytes are thermally de-
nol, methyl-n-propylketone, nitropro- sorbed to the GC column.
Membrane filtration: Membrane filter; to the desired limits of precision and

membrane disk. accuracy in retention, resolution, and
Membrane suppressor: Continuous quantitation of the sample components
chemical suppression. Ion exchange of interest.
through ion-exchange membranes. H+ Micellar chromatography: The addi-
or OH- are supplied by the respective tion of micelles to the mobile phase
regeneration solution (for example, sul- to effect separations. The micelles
furic acid). may act as displacing or partitioning
Metal affinity chromatography: A agents and provide another parameter
special form of ligand exchange chro- which may be used to change selec-
matography used for the separation of tivity. Surfactants above their critical
biopolymers with a particular affin- micelle concentration are used in mi-
ity for a specific metal cation typically cellar chromatography and in MEKC
copper(II), zinc(II), and iron(II). form of CE.
Metalophile: A compound that has Micro LC: Refers collectively to tech-
high affinity for active (acidic) silanol niques where a column of smaller than
groups on silicas surface. Usually a conventional internal diameter is used
strongly basic amine. for separation. The term micro LC is
Method detection limit (MDL): The most often used for HPLC in <0.5-mm
minimum amount of solute that can be i.d. columns; micro LC is used in high
analyzed within specified statistical lim- sensitivity analysis when the sample
its of precision and accuracy, including amount is limited and with certain ion-
sample preparation. ization techniques in LC–MS where the
Method development: A process of volume of solvent flowing into the ion-
optimizing the separation including ization source must be minimized.
the sample pretreatment so as to obtain Microbore: Refers to the use of col-
a reproducible and robust separation. umns with smaller-than-usual internal
Usually it emphasizes the search for the diameters in HPLC. Columns with
stationary phase, eluent, and column internal diameters of 2 mm and below
temperature combination that provides are considered to be microbore columns;
an adequate separation. columns with internal diameters below
Method translation: Several math- 0.5 mm are referred to as micro LC col-
ematical techniques for adjusting GC umns. In GC, microbore may signify
method parameters for variations in the columns with inner diameters less than
carrier gas type or column dimensions, 200 µm.
with the objective of maintaining either Microchip devices: Microdevices based
the same or ratiometric retention times. on silicon, glass, and other types of mi-
Useful when changing from helium to crofabricated chips where experiments
hydrogen carrier gas, or when increas- can be miniaturized into single- or mul-
ing speed of analysis or resolution by tichannel microfluidic circuits; these
adjusting column dimensions. Not use- devices can be used for CE and CEC,
ful if changing the stationary phase to a and should be low cost and disposable.
chemically different type. Microchip-based GC devices have been
Method validation: A process of test- available since approximately 1995. The
ing a method to show that it performs use of these devices for separations is
currently in its infancy, and applications Microporous resin: Same as microre-

should expand with time. ticular resin.
Microdialysis: Microdialysis is a mini- Microreticular resin: Cross-linked
mally invasive sampling technique that synthetic ion-exchange resins that
is used for continuous measurement of have pores with openings correspond-
free, unbound analyte concentrations ing to molecular sizes. Diffusion into
in the extracellular fluid of virtually the narrow pores can be impaired and
any tissue. Analytes may include en- low exchange rates can occur, as well as
dogenous molecules (for example, neu- poor performance, especially for large
rotransmitters, hormones, and glucose) molecules.
to assess their biochemical functions in M ic rowave -a s s i sted ex t rac t ion
the body, or exogenous compounds (for (MAE): The use of microwave energy
example, pharmaceuticals) to determine to heat samples in the presence of a
their distribution within the body. The solvent allowing for rapid extraction;
microdialysis technique requires the in- MAE can be performed in open ves-
sertion of a small microdialysis catheter sels. A nonmicrowave-absorbing sol-
(also referred to as microdialysis probe) vent is used and the sample contain-
into the tissue of interest. After the ing a substance with a high dielectric
probe is inserted into the tissue or (body) constant (for example, water) is rapidly
fluid of interest, small solutes can cross heated, with the extracted analytes
the semipermeable membrane by passive passing into the extraction solvent. A
diffusion. The microdialysis probe is de- variation of this technique involves the
signed to mimic a blood capillary and addition of an inert microwave-ab-
consists of a shaft with a semipermeable sorbing solid substance that transfers
hollow fiber membrane at its tip, which the heated energy to the surrounding
is connected to inlet and outlet tubing. solvent. MAE can also take place in
Microextraction: The general process closed vessels that are non-microwave-
of liquid extraction using small amounts absorbing containers.
of organic solvent where the phase ratio Migration time (tM): The time it takes
Vo/Vaq is quite low; other techniques for a charged molecule to move from the
using hollow microfibers as a barrier point of injection to the point of detec-
are also referred to as microextraction. tion in a CE capillary.
Microparticulate: Refers to the small Milling: Devices for reducing the par-
particles used in HPLC. Generally ticle sizes of solid materials. Disk mills
packings with a particle diameter of less pulverize <20-mm diameter hard sam-
than 10 µm and that are totally porous ples by feeding between stationary and
are considered microparticle packings. rotating disks with adjustable gap set-
Micropipette tip: A form of SPE in tings; samples are generally reduced to
which the packing material is embed- a 100-µm diameter; rotor speed mills
ded or adsorbed on the inner walls combine impact and shearing processes
of a pipette tip; useful for the SPE of to grind soft-to-medium hard and fi-
very small amounts of liquid sample; brous materials to 80 µm; ball mills
often used with xyz liquid handling grind material to submicrometer size
systems for automation purposes. See by developing high-grinding energy via
pipette tip. centrifugal or planetary actions using
agate, tungsten carbide, or PTFE- Mobile-phase modifier: Modifiers are

coated stainless steel balls. materials (usually organic or inorganic
Mincing: The process of breaking down compounds ) added to the mobile phase
a meat or vegetable product into smaller to alter its elution properties.
parts by tearing, chopping, cutting, dic- Mobile-phase strength: See solvent
ing, and so forth. strength.
Minimum detectable quantity (MDQ): Modifier: An additive that changes
The amount of solute that produces a the character of the mobile phase. In
signal twice the noise level. reversed-phase LC, for example, water
Mixed-bed column: Combination of is the weak solvent and methanol, the
two or more stationary phases in the strong solvent, is sometimes called the
same column, used most often for ex- modifier; sometimes other additives
change separations (IEC mixed anion such as competing bases like triethyl-
and cation resins) and SEC (mixture of amine or ion pair reagents are referred to
different pore size packings). The ad- as modifiers but they should more cor-
vantage in IEC is the total removal of rectly be called additives. See additive.
both cationic and anionic compounds; Molecular diffusion term (B term):
the technique is useful in SEC because Refers to the B term (second term) of
a wider molecular weight range can be the van Deemter and Golay equations.
accommodated by the same column. Also called longitudinal or axial diffu-
Mixed-mode separation: A separation sion term. It dominates band broaden-
that takes place in a single column as ing only at very low flow rates below the
a result of retention and selectivity pro- minimum plate height where the diffu-
vided by a dual retention mechanism. sion of individual solutes can occur in a
For example, at intermediate-to-high longitudinal (lengthwise) direction on
pH values, a reversed-phase column the column. See van Deemter equation,
with residual silanols can separate by Golay equation.
hydrophobic interactions as well as Molecular sieve: GC column packing
ionic interactions by virtue of the ion- that retains solute by combined mo-
ized silanols; sometimes mixed-mode lecular size and adsorptive interactions.
separations can be quite beneficial to Molecular sieves can separate light gases
the selectivity (band spacing) but can and hydrocarbons.
cause peak asymmetry; the precise bal- Molecular weight distribution: The
ance of interactions may be difficult to distribution of the molecular weight of
reproduce with subsequent batches of molecules in a polymer sample. Distri-
packing. bution can be defined as weight average
Mobile phase: The fluid that moves and number average.
solutes through the column. In LC, the Molecularly imprinted phases (MIPs):
mobile phase interacts with both the sol- See imprinted phases.
ute and the stationary phase and there- Monolith: A monolithic HPLC col-
fore can have a powerful influence on umn is a special type of column used
the separation. In GC, the mobile phase, in HPLC with porous channels rather
as an inert gas, has little interaction with than beads; monoliths, in chromato-
stationary phase and analytes and serves graphic terms, are porous rod struc-
to move the sample through the column. tures characterized by mesopores
and macropores. These pores provide tryptic peptides to analyze an entire

monoliths with high permeability, a proteome of a cell or tissue type pro-
large number of channels, and a high tein extract. The approach uses a dual
surface area available for reactivity. enzymatic digestion (Lys-C followed
The backbone of a monolithic column by trypsin) to increase the number
is composed of either an organic (poly- of peptides observed. The peptides
meric) or inorganic (silica) substrate, are separated using strong cation ex-
and the column can be chemically al- change and are identified by MS/MS
tered for specific applications. detection.
Monomeric phase: Refers to a bonded Multimodal SPE: The practice of SPE
phase where single molecules are bonded where two different phases or modes are
to a support. For silica gel, monomeric used to clean up a sample; the process
phases are prepared by the reaction of an can consist of two separate cartridges
alkyl- or arylmonochloro- or alkoxysi- placed in series with the analytes sepa-
lane. Polymeric phases generally are pre- rated on the two different cartridges; a
pared from a di- or trichlorosilane or an second process is where two different
alkoxysilane reactant. phases are present in the same cartridge
Moving zone: The moving zone is that or even on the same packing; sometimes
fraction of the mobile phase in the col- referred to as mixed-mode SPE.
umn that occupies the interstitial spaces.
Multidimensional chromatography: N
The use of two or more columns or Nano LC: LC practiced with columns
chromatographic techniques to ef- that have internal diameters less than
fect a better separation. It is useful for 100 µm; usually requires specialized in-
sample cleanup, increased resolution, strumentation; often used in proteomic
and increased throughput. Separa- studies where sample is limited and sen-
tion is carried out with two or more sitivity is required.
columns in which peaks are selectively Narrow-bore column: Columns of less
directed onto or removed from at least than 2 mm i.d. used in HPLC, and less
one of the columns by use of a timed than 320 µm i.d. in GC; also referred to
valve system. In GC, a Deans fluidic as microbore.
switch is often used. It also can be Nitrogen – phosphorus detection
used off-line by collecting fractions (NPD): The nitrogen–phosphorus detec-
and reinjecting onto a second column. tor catalytically ionizes N- or P-contain-
Also called coupled column chromatog- ing solutes on a heated rubidium or ce-
raphy, column switching, multicolumn sium surface in a reductive atmosphere.
chromatography, and boxcar chromatog- NPD is highly selective with sensitivity
raphy. See backf lushing, heart cut- somewhat better than FID.
ting, precut. Noise: See baseline noise.
M ult idimen sional protein iden - Non-aqueous reversed phase chroma-
tification technology (MudPIT): tography (NARP): Refers to reversed-
Multidimensional protein identifi- phase chromatography performed with-
cation technology combines both a out water as a component of the eluent
cation-exchange prefractionation and on a reversed-phase packing. Used for
reversed-phase HPLC separation of compounds that are very nonpolar that
either cannot be eluted or are poorly Nonporous packing (NPR, NPS, NPZ):
eluted from a reversed-phase column Particles similar to porous-layer bead
with 100% methanol or acetonitrile; in but with particle diameters in the sub-
these cases, solvent A would be aceto- 5-µm range, often particles are in the
nitrile and solvent B would be a stron- sub-2-µm range; used for high-speed
ger solvent such as tetrahydrofuran; for separations in short columns; NPS refers
NARP, reversed-phase rules apply (that to nonporous silica, NPR to nonporous
is, the more nonpolar the analyte, the resins, and NPZ to nonporous zirconia.
greater the retention). Normal-phase chromatography: A
Nonpolar: A nonpolar molecule is one mode of chromatography in which the
in which the electrons are distributed stationary phase is more polar than the
more symmetrically and thus does not mobile phase. Adsorption chromatogra-
have an abundance of charges at the phy on silica gel or alumina using mix-
opposite sides. The charges all cancel tures of less polar eluents (for example,
out each other. Nonpolar compounds, hexane–diethethyl ether) as a mobile
solvents, or bonded phases readily dis- phase would be a typical normal-phase
solve in organic solvents, such as hexane, system. Also refers to the use of polar
or prefer such solvents in place of water. bonded phases, such as -CN or NH2.
Nonpolar substances do not readily dis- Sometimes referred to as straight phase
solve in water. chromatography.
Nonporous particle: Refers to a solid
particle used as a support for a porous O
coated or bonded phase; pellicular par- Octadecylsilane (ODS, C18): The most
ticles are nonporous particles with large popular reversed phase in HPLC. Oc-
particle diameters (approximately 40 µm) tadecylsilane phases are bonded to silica
and nonporous silicas and resins with or polymeric packings. Both monomeric
small particle diameters of less than 3 µm and polymeric phases are available.
usually consist of a microbead with a thin Octylsilane (C8): A popular stationary
porous outer coating of silica gel, bonded phase in reversed-phase chromatogra-
silica gel, or polymeric phase. phy; usually has slightly less retention
Nonsuppressed ion chromatography: than the more popular C18; both mo-
Direct ion chromatography. After the nomeric and polymeric phases available.
separation column the eluent is directly Off-line SPE: The normal practice of
fed into the conductivity detection with- SPE where SPE cartridges, disks, pipette
out prior chemical suppression. The tips, and so forth are handled using
conductivity measurement takes place manual processes (for example, vacuum
on the high background conductivity. manifolds or pipette transfer); opposite
This requires a very high quality detec- to on-line SPE.
tor with perfectly stable temperature. On-column detection: The column it-
The nonsuppressed approach allows de- self serves as the flow cell in HPLC, CE,
tection of weak acids or bases that are or CEC. Generally the term used when
undissociated after suppression—for fused-silica capillaries are employed; the
example, silicate or borate. For cation outer polyimide layer is removed and op-
analysis the peaks for the components of tical beam is directed through the capil-
interest are larger than with suppression. lary; a measuring device (for example, a
photomultiplier tube) is located on the 750 µm. In HPLC, SFC, and capillary
opposite side of the capillary. electrophoresis, small internal diameter
On-column injection (OCI): In GC, re- (less than 100 µm) columns are used.
fers to the process of injecting the entire The most frequently used column ma-
liquid sample directly onto the head of terial is fused-silica tubing. Used very
the column using a fine needle that will little in routine HPLC or SFC but
fit inside the capillary. Usually carried routinely in CE and GC. Also termed
out at an initial column temperature capillary column.
less than the solvent boiling point, also Open-tubular column, packed: A cap-
termed cold on-column injection. illary-dimension column that is packed
On-line column switching: See mul- with stationary phase particles. Also
tidimensional chromatography, on- called micropacked columns, particularly
line SPE. in GC.
On-line preconcentration: A precol- Optically active resin: Incorporation
umn is placed in front of the separation of optically active groups into an ion-
column to concentrate analytes before exchange resin to allow separation of
their separation; different mechanisms optically active isomers. Not many are
may be used (for example, hydrophobic commercially available in HPLC.
interaction, adsorption, or enzymatic re- Organic modifier: Water-miscible
action) to retain analytes as a function of organic solvent added to an aqueous
time and then by a displacement process mobile phase to effect separations in re-
(such as solvent elution or pH change) versed-phase HPLC. Common organic
concentrated analytes are transferred to modifiers are acetonitrile, methanol, iso-
the separation column. propanol, and tetrahydrofuran.
On-line SPE: Refers to the use of small Orthogonality: Refers to two separation
stainless steel cartridges packed with dimensions for which the elution times
SPE packings placed across two ports in the two dimensions can be treated
of a 6- or 10-port injection or column- as statistically independent; ideally, the
switching valve. The SPE trap is loaded two dimensions should have totally dif-
with sample by an external pump or ferent retention mechanisms (for exam-
syringe transfer and then the valve is ple, reversed phase and normal phase;
switched so that the SPE trap becomes ion exchange and reversed phase; polar
part of the HPLC flow stream and ana- and nonpolar)
lytes can be swept into the column based Overload: In preparative chromatog-
on the solvent being used for displace- raphy, the overload is defined as the
ment. On-line SPE columns are usually sample mass injected onto the column
used multiple times whereas off-line SPE where efficiency and resolution begins
cartridges are generally used once. to be affected if the sample size is further
Open-tubular column: Also termed increased. See sample capacity.
capillary columns, open-tubular col-
umns for GC have the stationary phase P
coated or chemically bonded on the Packing: The adsorbent, gel, or solid sup-
inner walls or have support particles port used in the chromatography column.
deposited on the inner walls. Internal Most modern analytical HPLC packings
diameters range from ~100 µm up to are less than 10 µm in average diameter
with 5 µm currently the favorite. GC col- escape from the column exit may cause
umn packings range in size from 60–80 detector noise.
mesh down to 100–120 mesh. Partition chromatography: Separa-
Paired ion chromatography: The same tion process where one of two phases is
as ion-pair chromatography. held stationary on a solid support or the
Paper filtration: Using porous filter column inner wall (stationary phase or
paper (mainly cellulose) to remove liquid phase) while the other is allowed
particulates from liquid samples; pa- to flow freely down the column (mo-
pers with different porosities are avail- bile phase or carrier gas). Solutes parti-
able. Low porosity filters will remove tion themselves between the two phases
very fine particulates but may have a based on their individual partition co-
lower f low rate while high porosity efficients. LLC is an example; modern
filters filter out larger particulates at bonded-phase LC can be considered to
a higher flow rate; paper filtration is be a form of partition chromatography
often used in wet chemistry to filter, where one of the liquid phases is actu-
combust, and then weigh insoluble ally bonded to the solid support. Mech-
materials; ashless filter paper is used anistically, partition chromatography
for this purpose. implies that the solute becomes at least
Particle diameter (dp): Average diam- partially embedded within the station-
eter of the column packing particles. ary phase, which is impregnated, coated,
Particle size distribution: A measure of or bonded to the substrate, in contrast
the distribution of the particles used to to an adsorption process in which the
pack the LC column. In HPLC, a nar- solute does not penetrate into the reten-
row particle size distribution is desirable. tive surface or interphase.
A particle size distribution of dp ± 10% Partition coefficient (K): The ratio of
would mean that 90% of the particles the equilibrium concentration of solute
fall between 9 and 11 µm for a 10-µm in the stationary phase relative to the
average dp packing. equilibrium concentration of solute in
Particle size reduction: The general the mobile phase. In GC, the relative
process of reducing larger particles down concentration of solute in the mobile
to a size that can be more conveniently and stationary phases is a function of
extracted; the smaller the particle the k and β: K = βk. Also called distribu-
more quickly it will dissolve or if in- tion coefficient, K D, and distribution
soluble the more quickly analytes can constant, Kc.
be extracted for further sample cleanup. Passive sampling: In passive gas sam-
Typical methods for reducing particle pling, an air sample is pulled through a
size include pulverizing, milling, ho- flow controller into an evacuated canis-
mogenizing, chopping, blending, and ter over a chosen period of time, rang-
so on. ing from 5 min to 24 h. The sampling
Particulates: Generally refers to a period and the flow rate determine the
small particles found in the mobile canister volume required.
phase that can cause back pressure Peak: The profile of an analyte com-
problems by lodging in frits; it can pound as it is eluted from a column
also refer to the small particles packed through a detector; usually depicted
into HPLC columns. Particulates that on a visual output on a recorder or
printer based on the detector’s electri- other column degradation or drift in the
cal response. chromatographic conditions.
Peak area (A p): The area measured Peak variance (σ2): The second central
under a chromatographic peak; usu- moment of the peak about the retention
ally measured by an integrator or data time. For a Gaussian peak the variance
system; the peak area is related to the is the fundamental parameter control-
amount of substance eluted in a peak. ling peak width. See Gaussian peak.
Peak capacity (n): The number of Peak volume (Vp): The volume occu-
equally well resolved peaks that can be fit pied by a chromatographic peak from
in a chromatogram between the hold-up starting basepoint to ending basepoint
volume and some upper limit in retention as it passes through the detector: Vp =
kn. For R = 1, n can be ex—ressed by the
p Fcwb
approximation n = 1 + √N /4 · ln(1 + kn) Peak width at base (w b): The width
where N is the plate number and kn is the of the chromatographic peak at the
retention factor of peak n. baseline as eluted from the column. It
Peak dispersion: See band broadening. is measured at the baseline by drawing
Peak doublet: A split peak generally tangents to the inflection points on the
caused by column void, poor injection sides of the Gaussian curve representing
technique, or solvent flooding in GC. the peak. Smaller peak widths usually
Split peaks also could be closely eluted represent efficient separations; also re-
compounds. ferred to as band width. It is sometimes
Peak height (hp): The maximum height convenient to estimate the peak width
of a chromatographic peak as measured at base from the peak area and height:
from the baseline to the peak apex; the wb = 1.596 Ap/hp (see Figure 2).
peak height is related to the amount of Peak width at half-height (w h): The
substance eluted in a peak. width of the chromatographic peak
Peak overload: When too much of any at half of the peak height above the
one solute is injected its peak may be baseline. Smaller peak widths usually
distorted into a triangular shape. represent efficient separations; also re-
Peak shape: Describes the profile of a ferred to as band width. It is sometimes
chromatography peak. Theory assumes convenient to estimate the peak width
a Gaussian peak shape (perfectly sym- at half-height from the peak area and
metrical); peak asymmetry factor de- height: wb = 0.94 Ap/hp (see Figure 2).
scribes shape as a ratio. See asymmetry. PEEK: Polyether ether ketone (PEEK) is
Peak tracking: A method of matching a colorless organic polymer. It is used as
of peaks that contain the same com- a material for inert capillaries and fit-
pound between different experimental tings in HPLC and IC systems
runs during method development; relies Pellicular: See porous layer bead.
upon detection parameters of each pure Percent B (%B): Refers to the stronger
analyte; diode-array detectors and mass solvent in a binary solvent mixture; %A
spectrometers are among the best detec- would be the weaker solvent analog.
tors for peak tracking because of their Per fu sion c hromatog ra phy : Re-
specificity. Also refers to data-system fers to chromatography performed
tracking of gradual changes in retention u si ng pa r t ic le s w it h ver y la rge
times caused by stationary-phase loss or pores (for example, 4000–8000 Å)
called throughpores (megapores or Phenol extraction: A sample prepara-

gigapores). Eluent f lows through tion technique used for the isolation of
the particle as well as smaller in- DNA from biological samples.
terconnecting pores (for example, Phenyl phase: A popular nonpolar
300–1000 Å) called diffusive pores bonded phase prepared by the reac-
between the large pores. Best suited tion of dimethylphenylchlorosilane or
for the preparative separation of mac- alkoxysilane with silica gel for LC, or
romolecules. as the components of a cross-linked
Permeability (Bo): Also referred to as or bonded phase for GC. Claimed to
column permeability and specific perme- have affinity for aromatic-containing
ability; a term expressing the resistance compounds and does impart a different
of the packed column to the flow of selectivity compared to alkyl bonded
mobile phase. For a packed column: Bo phases.
= (dp2/180) · ε3/(1 - ε)2 ≈ (dp2)/1012.For Photoionization detection (PID): The
an open-tubular column: Bo = dc2/32. A photoionization detector ionizes solute
column with high permeability gives a molecules with photons in the UV en-
low pressure drop. ergy range. PID is a selective detection
Permeation: In SEC, refers to the pro- method that responds to aromatics and
cess where a solute can enter a mobile olefins when operated in the 10.2-eV
phase filled pore of the packing. photon range. It can respond to other
Phase collapse: See phase dewetting. materials with a more energetic light
Phase dewetting: A term used in source.
reversed-phase LC where very dense PIONA: Refers to the analysis of paraf-
bonded-phase coverage and a high per- fins, isoparaffins, olefins, naphthenes,
centage of aqueous content in mobile and aromatics.
phase can lead to expulsion of water Pipette tip: Replaceable tips used in
from the pores, which prevents the automation of liquid handling chores;
normal partitioning process from tak- used once and discarded to avoid con-
ing place. Phase dewetting may occur tamination.
with as high as 10% organic content Pirkle column: Chiral “brush type” sta-
in the mobile phase but can occur at tionary phases based on 3,5-dinitroben-
lower %B values; results in earlier than zoyl-phenylglycine silica that are used in
normal elution of analytes, poor peak the separation of a wide variety of en-
shape, and nonreproducible retention antiomers. Named after the developer,
times. Dr. William Pirkle, University of Illnois.
Phase ratio (β): The relative amount pKa : An acid dissociation constant,
of stationary to mobile phase in the Ka, (also known as acidity constant, or
column. In partition chromatography: acid-ionization constant) is a quantita-
β = VS/V M where VS and V M are the tive measure of the strength of an acid
volume of stationary and mobile phase in solution. It is the equilibrium con-
in the column respectively. For open- stant for a chemical reaction known
tubular columns: β ≈ rc /2d f. Thicker as dissociation in the context of acid-
stationary phase films or higher phase base reactions. The equilibrium can be
loading gives longer retention and written symbolically as: HA ←→ H+ +
higher peak capacity. A - where HA is a generic acid that
dissociates by splitting into A−, known solvent and is useful for identifying suit-
as the conjugate base of the acid, and able mobile phase solvents or extraction
the hydrogen ion or proton, H+, which, solvents. The polarity index increases
in the case of aqueous solutions, ex- with polarity; examples: hexane, P′ =
ists as the hydronium ion — in other 0.0; isopropanol, P′ = 3.9; tetrahydrofu-
words, a solvated proton. The disso- ran, P′ = 4.0; methanol, P′ = 5.1; aceto-
ciation constant is usually written as a nitrile, P′ = 5.8; water, P′ = 9.0
quotient of the equilibrium concentra- Polyacrylamide gel: Neutral hydro-
tions (in mol/L), denoted by [HA], [A−], philic polymeric packings used in
and [H+]: Ka = ([H+] [A−])/[HA]; due to aqueous SEC. They are prepared by the
the many orders of magnitude spanned copolymerization of acrylamide with
by Ka values, a logarithmic measure of N,N′-methylene-bis-acrylamide.
the acid dissociation constant is more Polyaromatic hydrocarbon (PAH):
commonly used in practice. The loga- Members of a class of hydrocarbon
rithmic constant, pKa, which is equal to molecules characterized by one or more
−log10 Ka, is sometimes also (but incor- fused aromatic rings.
rectly) referred to as an acid dissociation Polychlorinated biphenyl (PCB): Bi-
constant. phenyl molecule with two or more
Planar chromatography: A separation chlorine substitutions.
technique in which the stationary phase Polyethylene glycol (PEG): Polymeric
is present as or on a plane (IUPAC). hydrocarbon used as a GC stationary
Typical forms are paper and thin layer phase; possesses moderately polar reten-
chromatography. tion characteristics.
Plate height (H): See theoretical plate Polyethyleneimine (PEI): Polyethyl-
height. eneimine, an anionic polymeric phase
Plate height, effective (Heff): The used to coat or bond onto silica or a
column length divided by the number polymeric packing. Most often used for
of effective theoretical plates: Heff = L/ the separation of proteins and peptides.
Neff Polymeric packings: Packings based
Plate number (N): See theoretical on polymeric materials, usually in the
plate number. form of spherical beads. Typical poly-
Polar: A polar molecule may be polar mers used in LC as well as GC are
as a result of polar bonds or as a result polystyrene–divinylbenzene (PS-DVB),
of an asymmetric arrangement of non- polydivinybenzene, polyacrylamide,
polar bonds and nonbonding pairs of polymethylacrylate, polyethyleneoxide,
electrons; polar molecules are generally polydextran, or polysaccharide.
able to dissolve in water (H2O) because Polymeric phase: Refers to chemically
of the polar nature of water; polar mol- bonded phase where a polymer species
ecules do not prefer nonpolar organic is bonded to silica-based particles or to
solvents such as hexane. Polar molecules the wall of an open-tubular column.
have slightly positive and slightly nega- Polymeric SPE: The use of a polymeric
tively charged ends; we often refer to a base material (for example, PS-DVB or
compound’s polarity. methacrylate) rather than an inorganic
Polarity index (P’): The polarity index material (for example, silica or alu-
is a measure of the relative polarity of a mina); polymers generally have a wider
pH range and higher sample capacity a submicrometer-thick shell; wide pore

than some of the inorganic materials. versions (>300 Å) allow rapid diffusion
Polystyrene – divinylbenzene (PS- of macromolecules and smaller pore
DVB) resin: The most common base versions (90–120 Å) are for small mol-
polymer for ion-exchange chromatog- ecules; give similar efficiency to sub-
raphy. Ionic groups are incorporated 2-µm particles but at much lower pres-
by various chemical reactions. Neutral sure because of their larger particle size.
PS-DVB beads are used in reversed- Porosity: For a porous substrate, the
phase LC. Porosity and mechanical ratio of the volume of the pores in a
stability can be altered by variation ofparticle to volume occupied by the par-
the crosslinking through the variation ticle. The pore volume is a measure of
of the DVB content. In GC, a number the porosity and is expressed in mL/g.
of porous polymer stationary phases are Porous layer bead: A small glass bead
available for gas and light-compound coated with a thin layer of stationary
separations. phase. The thin layer can be an ad-
PONA: Refers to the analysis of paraf- sorbent, resin, or a phase chemically
fins, olefins, naphthenes, and aromatics.
bonded onto the adsorbent. These pack-
Poppe plot: A kinetic plot named after ings were among the first to be used in
Professor Hans Poppe ( J. Chromatogr. HPLC. They were of larger particle size
A 778, 3 [1997]), University of Amster- (20–40 µm) than the microparticulate
dam, the Netherlands, where the plate packings of today but were easy to pack
time log (t 0/N ) is depicted as a func-and gave adequate efficiency. Also re-
tion of the number of theoretical platesferred to as controlled surface porosity
to assess the limits of column perfor- supports and pellicular materials.
mances as a function of particle size, Porous particle: Refers to column
column pressure drop, and so forth. packing particles possessing intercon-
Pore diameter: Same as mean pore necting pores of specified diameter and
diameter. pore volume; generally in HPLC porous
Pore size: The average size of a pore inparticles with diameters below 10 µm
a porous packing. Its value is expressedare the most popular, and in prepara-
in angstroms or nanometers. The pore tive chromatography larger particles
size determines whether a molecule can are used because of their lower cost and
diffuse into and out of the packing. Seehigher column permeability.
mean pore diameter. Porous polymer: A packing material,
Pore volume (Vi): The total volume of generally spherical, based on organic
the pores in a porous packing, usually polymers or copolymers; popular ex-
expressed in mL/g. Better termed the amples would be polystyrene–divinyl-
specific pore volume. It is measured by benzene, polyacrylates, polydextrans,
the BET method of nitrogen adsorp- polyacrylamides, and polybutadienes.
tion or by mercury intrusion where Hg Retains solutes by selective adsorption
is pumped into the pores under high or molecular size interaction.
pressure. Porous-layer open-tubular (PLOT) col-
Poroshell: Similar to nonporous par- umn: An open-tubular column used in
ticles and porous-layer beads; particles GC or HPLC that has particles coated
are generally in the 2–5 µm range with or uncoated with stationary phase
attached to the inner walls, which al- Precolumn reaction: Analytes are con-
lows more rapid mass transfer. In GC, verted into components with better
small porous particles such as polymer, detectability (for example, UV–vis ab-
alumina, silica, and so forth are attached sorbance) by a chemical reaction before
to the walls or the wall may be modified injection. The analytes are separated and
by etching or other treatment to increase detected by UV–vis detection. Complex-
the inner surface area and provide gas– ing agents such as EDTA, NTA, and so
solid chromatographic retention behav- forth are used as their Fe(III) complexes.
ior. In LC, porous polymers have been Precolumn: A short section of similar
used occasionally. In GC, the technol- column placed before the analytical col-
ogy is more developed. umn; used to physically retain undesired
Postcolumn derivatization: See postcol- compounds or to saturate the mobile
umn reaction. phase with stationary phase that may be
Postcolumn reaction: In LC and IC, packed into the precolumn (for example,
after the analytical column a UV-trans- a silica precolumn saturates the mobile
parent ion or molecule is converted into phase with dissolved silica and prevents
a component with better detectability mobile phase from dissolving silica in the
(that is, UV–vis absorbance, fluores- analytical column).
cence) by adding a specific reagent. This Precolumn filter: A filter used between
product is then detected with UV–vis the injector and the column (or guard
or fluorescence detection. The reaction column) to keep unwanted sample
of the analyte and the reagent is usually components from reaching the column;
very selective and yields often in a col- sometimes called in-line filter, occasion-
ored product (visible detection), that is, ally inlet filter.
chromate + diphenylcarbazide complex Preconcentration: See also trace en-
(540 nm); bromate + iodide → triiodide richment.
(352 nm). Parameters that will influence Precut: Peaks in the beginning of a chro-
the sensitivity are the reaction time (flow matogram are removed to vent or are di-
rate/length of reaction coil), the reaction rected onto another column of different
temperature, pH, concentration of cata- polarity, or at a different temperature, for
lysts. In GC, postcolumn methanization improved resolution. See heart cutting,
may be used to convert CO and CO2 to multidimensional chromatography.
CH4 with hydrogen and a heated nickel Prefilter (SPE): In cases where samples
catalyst to achieve flame ionization de- contain a large amount of particulates,
tection, more sensitive than thermal regular SPE cartridges and disks may
conductivity detection. become clogged and flow is reduced.
Potentiometric detection: Ion selec- Prefilters are filter devices that have
tive electrodes in a flow-through cell higher porosity that will filter out
are used to detect the analyte ions. Not large particles and allow the SPE bed
a very common type of detection. to operate more efficiently. Some SPE
Pounds per square inch (psi): A unit of devices have prefilters built in; in oth-
pressure: 14.6959 psi = 1 atm = 101.325 ers one can add a prefilter. In some
kPa = 1.013 bar. cases, the use of an inert packing such
Precolumn derivatization: See precol- as glass beads serves the same purpose
umn reaction. as an actual filter.
Preparative chromatography: Refers proximated as Δp ≈ (1012 L η u–)/(dp2 ), where

to the process of using chromatogra- u– is the average linear velocity. In open-
phy as a technique for the isolation of a tubular column GC, the pressure drop is
sufficient amount of material for other Δp ≈ (8 L η u–)/(rc2). The equations for GC
experimental or functional purposes. will overestimate the required pressure drops
For pharmaceutical or biotechnological by more than 10% at inlet pressures above
purifications, large columns of several 400 kPa (58 psig) because of gas compress-
feet in diameter can be used for puri- ibility effects. Pressure drop can be expressed
fying multiple grams of material. For in pressure units of psig, bar, atm, kPa, or
isolating a few micrograms of valuable MPa. The above equations will yield pres-
natural product, a 4.6-mm i.d. analytical sures in pascals if the dimensions are ex-
column can be used. Both of these sepa- pressed in centimeters, times in seconds, and
rations can be considered preparative viscosities in pascal-seconds.
chromatographic approaches. Prepara- Pressure, relative (P): Relative pressure
tive LC is often employed; preparative across the column: P = pi /po
GC is seldom used. Pressurized-fluid extraction (PFE):
Pressing: The general process of squeez- Pressured f luid extraction is a liq-
ing liquid from a semisolid material uid–solid extraction process where
(such as plants, fruit, or meat). the sample and solvent are placed in a
Pressure drop (Δp): The pressure drop closed container and heated well above
across a column: Δp = pi – po, where pi the solvent’s normal boiling point. The
and po are pressure at the column inlet combination of increased temperature
and outlet, respectively. and resultant pressure extracts analytes
Pressure injection (CE): Pressure-in- and matrix compounds into the super-
duced injection; the use of pressure or heated fluid, often in a few minutes.
vacuum to inject small (nanoliter) vol- Because the technique extracts a wide
umes of sample into a capillary column; variety of soluble compounds, addi-
best for narrow-bore capillaries (<10 µm tional cleanup steps may be required
i.d.); a version of hydrostatic injection. after PFE is completed; method de-
Pressure, absolute inlet (pi): The col- velopment involves selecting the best
umn inlet pressure expressed relative to solvent for analytes and the poorest
a vacuum. solvent for the matrix and other in-
Pressure, absolute outlet (po): Pressure terferences that may be present. The
at the column outlet, relative to vacuum. technique has been approved for vari-
Pressure, back: Same as head pressure, ous environmental samples by the U.S.
column pressure. EPA under the generic name of PFE or
Pressure, head (Δp): The pressure dif- pressurized-solvent extraction. See
ference between the inlet and outlet of accelerated solvent extraction.
the column. In LC, governed by the Pressurized-solvent extraction (PSE):
following approximate equation for a See accelerated solvent extraction,
column packed with spherical particles: pressurized-fluid extraction.
Δp ≈ (3000 L η)/(t0dp2) where η is the mobile Primary sampling: The collection of
phase viscosity, t 0 the column holdup time, one or more increments or units ini-
and dp is the particle diameter. In packed- tially taken from a population; the
column GC the pressure drop can be ap- primary sample is that taken from the
primary source; proper statistical sam- the temperature of the inlet is heated
pling protocols are recommended. very rapidly to transfer the sample into
Process-scale chromatography: Re- the column; using PTV there is the
fers to the use of liquid chromatography potential for less sample discrimination
at the industrial scale level outside the and less thermal degradation of sensi-
laboratory; generally requires specially tive compounds compared to hot inlet
designed columns (usually with di- injections.
ameters > 5 cm), recoverable solvents, Programmed temperature vaporizer
lower-cost packings (with larger and ir- (PTV): An inlet system designed to per-
regular-shaped particles), and overloaded form programmed-temperature injection.
operating conditions compared to those Protein crashing: The term used in
of laboratory-scale HPLC. removing or reducing the protein con-
Programmed elution: A procedure in centration in a biological fluid such as
which the conditions of separation are plasma. After slight dilution, an organic
changed in a programmed manner. Un- solvent such as acetonitrile is added to
like LC, in GC and SFC both tempera- the plasma and the proteins, which are
ture and pressure can be programmed, insoluble, precipitate (crash). Centrifuga-
separately or simultaneously. tion or filtration is used to remove the
Programmed temperature chroma- protein, and the supernatant liquid is in-
tography: Use of conditions in which jected into an HPLC system or worked
the temperature is varied during the run up further.
in a controlled manner. Widely used in Protein precipitation: See protein
GC; seldom seen in LC. crashing.
Programmed temperature injection Pulsating flow: Flow originating from
(PTI): A cold injection technique in a reciprocating pump. Normally the
which the inlet temperature is specifi- pulses are dampened out by a pulse
cally programmed from the GC. damper, an electronic pressure feedback
Programmed temperature rate: The circuit, or an active damper pump head.
rate, in °C/min, at which the GC oven Some detectors (for example, electro-
temperature is increased during a con- chemical, refractive index) are greatly
trolled temperature program ramp. The affected by flow pulsations.
program may consist of multiple ramps Pulsed amperometric detection:
with variable hold times before and after Electrochemical detection applying dif-
each. Typical GC programming rates ferent potentials (pulses) to the working
range from <0.5 °C/min up to 40 °C/ electrode. Components that can be an-
min. Programming rates up to 200 °C/ alyzed include those that are oxidized
min and higher have been applied to or reduced at the electrode and those
high-speed gas chromatography. that react with the electrode surface or
Programmed temperature vaporiza- cover it. To remove reaction products
tion (PTV): In PTV, the sample is intro- that could foul the electrode, highly
duced into the inlet liner at a tempera- oxidative and reductive potentials are
ture slightly below the boiling point of applied to the working electrode after
the solvent; the solvent is continually the measuring potential. This removes
evaporated and vented through the the reaction products from the previ-
inlet split line; after the solvent is gone, ous measuring cycle and renews the
electrode surface. Typical applications lyzed by GC. Often applied to polymer

are carbohydrates and amino acids. characterization.
Pulsed discharge detection (PDD):
Several ionization detectors use a pulsed- Q
discharge ion source to improve detec- Quaternary methyl amine (QMA): A
tivity compared to constant-discharge strong anion-exchange functionality
detectors. popular in resin-based packings; usually
Pulsed-splitless injection: A form of supplied in chloride form.
GC injection recommended for large Quaternary mobile phase: See quater-
volumes (up to 5 µL) of sample where nary-solvent mobile phase.
a short-term high pressure pulse is im- Quaternary-solvent mobile phase: A
posed on the inlet such that there is not mobile phase consisting of four separate
a large volume of solvent vapor gener- solvents that allow the mobile-phase
ated and most or all of the sample is di- composition to be fine-tuned; most
rected to the column; after the sample often this mobile phase is delivered by a
is transferred, then normal pressure is low-pressure quaternary pump.
resumed. Using this technique, highly QuEChERS: A technique initially used
volatile compounds are less likely to be for the extraction of pesticides from
lost through the split vent line and ther- fruits and vegetables. It consists of
mally unstable compounds spend less two steps: salting out extraction using
time in the hot injection port so there is buffered or unbuffered solvent (usually
less degradation. acetonitrile), and dispersive SPE where
Pulverizing: Electromechanically a solid adsorbent is used to treat an
driven rod or vibrating base devised to aliquot from the first step to remove
reduce the particle size of solid samples. interferences and matrix compounds.
A freezer mill can be used with liquid QuEChERS (which stands for quick,
nitrogen to treat malleable polymers easy, cheap, effective, rugged, and safe)
or those with low glass transition tem- is mostly used with GC–MS and LC–
peratures. MS (or MS/MS) to more selectively an-
Purge-and-trap sampling: Dynamic alyze pesticide extracts. More recently,
headspace technique where the head- QuEChERS has expanded to matrices
space vapors over a liquid or solid such as cooking oil, meat, fish, and
sample are continuously removed by a biological fluids, and to other analytes,
flow of gas over the sample (purging) or such as pharmaceuticals, antioxidants,
through the sample (sparging); volatil- and toxins.
ized analytes are usually concentrated by
trapping on an adsorbent or by cryogenic R
means. The trap is then heated to desorb Radial compression: The use of radial
trapped components into a GC column. pressure applied to a flexible wall col-
Most often used for volatile trace ana- umn to cut down on wall effects.
lytes where concentration is needed. Radial diffusion or dispersion: Diffu-
Pyrolysis gas chromatography: The sion or dispersion across the column in a
process of heating a sample enough to radial direction. If the sample is injected
break its chemical bonds, thereby form- into the exact center of a column, it will
ing smaller molecules that can be ana- spread not only in a longitudinal
direction as it moves down the column Refractive index peak: A pseudo peak
but radially as well, allowing the solute normally found near the hold-up volume
to reach the wall region where the eluent resulting from the refractive index sensi-
velocity is different than in the center of tivity of absorbance and other detectors.
the column. Regeneration: Regeneration of the
Recovery: The amount of solute (sam- packing in the column to its initial
ple) that is eluted from a column rela- state after a gradient elution. Mobile
tive to the amount injected. Excellent phase is passed through the column
recovery is important for good quanti- stepwise or in a gradient. The sta-
tation, for preparative separations, es- tionary phase is restored (solvated) to
pecially for biomolecules, and for good its initial condition. In ion exchange,
peak shape and resolution. Reasons for regeneration involves replacing ions
inadequate recovery can be solute inter- taken up in the exchange process with
action with active sites on the packing, the original ions which occupied the
with column frits, and with column exchange sites. Regeneration can also
tubing. Compound decomposition refer to bringing back any column to its
during the separation process can also original state (for example, the removal
effect recovery. of impurities with a strong solvent).
Recycling chromatography: A tech- Relative retention (r): Retention rela-
nique where the column effluent is re- tive to a standard peak: r = t′R/t′R(st) where
circulated onto the head of the column t′R is the retention time of the compo-
in an attempt to gain the advantage of nent of interest and t′R(st) is the retention
extended column length. Can be car- time of the standard peak. Also: r = ki/
ried out on a single column by passing kst where ki and kst are the correspond-
the effluent back through the pump. An ing retention factors. For two adjacent
alternative technique uses two columns peaks, the separation factor, α, expresses
connected by a switching valve where the relative retention. See separation
the effluent of one column is directed factor, resolution.
onto the head of the other column. Very Relative standard deviation (RSD,
seldom used in HPLC, and then only in %RSD): In probability theory and
exclusion chromatography. statistics, the relative standard devia-
Reduced plate height (h): The plate tion (RSD or %RSD) is the absolute
height expressed in terms of the average value of the coefficient of variation. It
particle diameter for packed columns: is often expressed as a percentage. A
h = H/dp where dp is the particle diam- similar term that is sometimes used is
eter, or in terms of the column inner di- the relative variance which is the square
ameter for open-tubular columns: h = H/ of the coefficient of variation. Also, the
dc where dc is the column inner diameter. relative standard error is a measure of a
Refractive index detection (RI detec- statistical estimate’s reliability obtained
tion): Based on differential refractive by dividing the standard error by the
index between the mobile-phase sol- estimate; then multiplied by 100 to
vent and the eluted analyte in mobile- be expressed as a percentage. The rela-
phase background; not useful in gradi- tive standard deviation is widely used
ent work; often used in size-exclusion in analytical chemistry to express the
chromatography. precision and repeatability of an assay.
Removable well plates: See array 96- They contain an outer hydrophilic sur-
well plate. face that provides minimal interaction
Representative sample: A sample re- with proteins and when combined with
sulting from a statistically worked out small pores on the sorbent exclude the
sampling plan; it can be expected to ad- proteins. The inner surface is hydropho-
equately reflect the properties of interest bic, and when small molecules diffuse
of the parent population. into the pores they interact by reversed-
Residual silanols: The silanol (-Si–OH) phase mechanisms and are retained. The
groups that remain on the surface of a small molecules such as drugs and their
packing after chemically bonding a metabolites can be removed by rinsing
phase onto its surface. These silanol with an organic solvent. RAMs are most
groups, that may be present in very successfully used in a column switching
small pores, may not be accessible to setup where the secondary column is
the reacting bulky organosilane (such as used to resolve the small molecules and
octadecyldimethylchlorosilane) but may the proteins are directed to waste so as
be accessible to small polar compounds. not to foul the secondary column.
Often they are removed by endcapping Retention factor (k): The measure
with a small organosilane such as tri- of time the sample component resides
methylchlorosilane. See endcapping. in the stationary phase relative to the
Resin: A solid polymeric packing used time it resides in the mobile phase:
in ion exchange separations. The most k = (tR – tM)/tM. Formerly, kʹ was used
popular resins are polystyrene–divinyl- and it was called the capacity factor or
benzene copolymers of small particle size capacity ratio.
(<10 µm). Ionic functionality is incorpo- Retention gap: A short piece of deac-
rated into the resin. tivated but uncoated column placed
Resolution (Rs): Peak resolution; incor- between the inlet and the analytical
porates both efficiency and separation. A column. A retention gap often helps
resolution of 1.5 is said to be “baseline” relieve solvent flooding. It also entrains
resolution, and a minimum resolution of nonvolatile sample contaminants from
1.7–2.0 is considered essential for robust- on-column injection.
ness. For two closely eluted peaks: R = Retention index (I): A uniform system
(tR,2– tR,1)/wb where the subscripts 1 and of retention classification according to a
2 refer to the first and second
— peaks. solute’s relative location between a pair
From N, k2 and α: Rs= (√N /4)((α – 1)/α) of homologous reference compounds on
(k2/(k2 + 1)) (k2 is the retention factor of a specific column under specific condi-
the second peak). tions. A series of normal straight-chain
Resolution equation: See resolution. hydrocarbons, fatty acid esters, or mul-
Response factor (RF): Defines the rela- tiring polycyclic aromatic hydrocarbons
tionship between the measured peak area have been used for the reference com-
or height and the quantity of substance pounds. For a solute i that is eluted at tʹRi
represented by a peak. between two hydrocarbons with chain
Restricted access media (RAM): RAM length z and z + 1: I = 100[z + (logt′Ri –
are sorbents are used for the direct injec- logt′Rz)/(logt′Rz1 – logt′Rz)].
tion of biological fluids such as plasma Retention time (tR): The time between
or serum into an HPLC flow stream. injection and the appearance of the peak
maximum. It is usually measured from lipophilicity) of the solutes; the more

the point of injection to the apex of the hydrophobic, the stronger the retention.
peak. For asymmetric peaks it should be The greater the water solubility of the
measured to the center of the mass of the analyte, the less it is retained. There are
peak. Also called the total retention time many variations of reversed-phase LC
(IUPAC). See retention volume; reten- where various mobile phase additives
tion time, adjusted. are used to impart a different selectivity.
Retention time, adjusted (t’R): A mea- For example, for the reversed-phase LC
sure of the retention time adjusted for of anions, the addition of a buffer and
the void time or unretained peak time: a tetraalkylammonium salt would allow
t′R = tR – tM where tR is the retention timeion pairing to occur and effect separa-
and tM (or t 0) is the hold-up time, void tions that rival ion-exchange chromatog-
time, or unretained peak time (that is, raphy. More than 90% of HPLC users
the time it takes for a small, unretained employ reversed-phase LC.
compound that completely permeates Reynolds number (Re): For flow in a
the pores to be eluted from the chro- smooth unpacked pipe where u– is the
matographic column). average velocity (cm/s), d c is the pipe
Retention volume (V R): The volume diameter, η is the viscosity (Pa·s) and
of mobile phase required to elute a sub- ρ is the density (g/cm3): Re = (u– dcρ)/η.
stance from the column: VR = Fct R or The Reynolds number is the ratio of
VR = VM + K DVS where VM is the void viscous to inertial energy of the mov-
volume, KD the distribution coefficient, ing fluid. At low R e viscous friction
and VS the stationary phase volume. Also dominates and controls fluid motion,
termed the total retention volume. See re- making it slow and steady. In an un-
tention time. packed tube flow becomes fully turbu-
Retention volume, adjusted (V′R): Ad- lent when Re exceeds 4200. In a packed
justs the retention volume for the holdup bed u– is replaced with the average inter-
volume (or V0) where VR is the retention stitial velocity and dc with the average
volume of the peak of interest and VM particle diameter. Flow becomes turbu-
is the hold-up or void volume, the vol- lent in a packed bed at Re above about
ume corresponding to the holdup time: 10 but is not fully turbulent until R e
V′R = VR – VM exceeds 100–200.
Retention volume, corrected (V R0 ): Riffler: A mechanical device used in
Corrects the retention volume for the subdividing solid powder samples into
effect of carrier-gas expansion along the smaller units. Rifflers can be manual or
column: VR0 = VR j automated. Rifflers will subdivide mate-
Reversed-phase chromatography: The rial samples into two smaller portions by
most frequently used mode in HPLC. It a single pass or further subdivisions can
uses low polarity packings such as octa- be attained by multiple passes.
decylsilane or octylsilane phases bonded Rinse step: In SPE, the rinse (wash)
to silica or neutral polymeric beads. The step is the third step in the process.
mobile phase is usually water or water- After the sample is loaded, the rinse step
miscible organic solvents such as metha- is designed to eliminate interferences
nol or acetonitrile. Elution usually occurs including various matrix compounds.
based on the relative hydrophobicity (or A solvent (or solvents) or buffer is selected
to remove interferences but not the ana- onto a column without overload and
lytes of interest. loss of column efficiency. Often ex-
Room temperature (To): The room or pressed as grams of sample per gram
laboratory temperature can be used as a of packing. Overload is defined as the
reference temperature for gas measure- sample mass injected that will cause
ments, for example 20 °C or 25 °C. the column efficiency to decrease by
Round-well plates : 96-well plates that 10% from its normal value. Sometimes
have round-shaped wells resembling 96 referred to as sample loading.
small test tubes. See 96-well plate. Sample discrimination: The charac-
teristic of systematic change in sample
S composition according to a specific
Salting-out effect: The use of a high sample property. For example, a GC
concentration of salt buffer in the mo- inlet may exhibit mass discrimination
bile phase to cause a low polarity ana- and admit relatively higher amounts of
lyte to have a decreased solubility in low-boiling sample components than
water and therefore precipitate or come high-boiling components in the same
out of solution; most often used for the sample or injection.
hydrophobic interaction chromatog- Sample division: The process of sam-
raphy of proteins where proteins are ple reduction to divide the sample into
first precipitated at high salt con- smaller portions while retaining repre-
centrations then eluted by gradual sentative characteristics of the primary
dilution using reverse gradient elu- sample. See sample size reduction.
tion. Sa lting-out is a lso used in Sample loop: Part of an injection
head space sa mpling to increa se valve that delivers an accurate volume
the ionic strength of the sample of liquid or gas to the column, giv-
solution and thereby decrease the ing a “slug” injection; loops come in
solubility of dissolved analytes and in- different volumes depending on the
crease their headspace concentrations. needs of the analysis and the size of
Can also be used in liquid–liquid the column.
extraction; see salting-out extraction. Sample pretreatment: Often syn-
Salting-out extraction: In this extrac- onymous with sample preparation; the
tion approach, high concentrations of process of manipulating the sample to
salt in the aqueous phase will cause make it easier to analyze.
certain compounds to migrate into an Sample size reduction: The process of
organic phase or perhaps vice-versa; sample reduction to divide the sample
high concentrations of salt also will into smaller portions while retaining
force normally miscible solvents (such representative characteristics of the
as water and acetonitrile) to become primary sample. See sample division.
immiscible and be used for further Sample tracking: The process of track-
partitioning more polar analytes than ing primary, secondary, laboratory, and
could be achieved by an extraction further samples through the analyti-
using a non-polar organic solvent. See cal cycle; it is important for chain of
QuEChERS or salting-out effect. custody reasons to be able to ensure
Sa m ple c a pac it y : Refers to t he that the sample analyzed in the instru-
amount of sample that can be injected ment was the original sample collected
at the source; sample tracking can be as reproducibility of results on columns of

simple as writing a sample number on different internal diameters when using
a container but can be more complex, the same particle size and bonded phase;
such as using bar-coded vials or radio normally a larger diameter column is
frequency identification (RFID) tags to used to increase capacity; a linear scale-
automatically keep track of sample flow. up process minimizes time required to
Sampling: The process of collecting optimize preparative separations.
a representative sample at the source. Scavenger: Special type of solid-phase
Sampling can also refer to further sam- particle that uses chemical reactions (un-
ple division as it more closely approaches like SPE, which uses molecular interac-
the laboratory analysis; it is important tions) to remove undesired species such
to make sure that the final sample ana- as undesired reaction products or excess
lyzed represents a subsample of the origi- starting material from an organic syn-
nal sample without any imposed bias or thesis. Scavengers mostly operate on the
discrimination. basis of covalent bonding. Packing ma-
Sampling error: In statistics, sampling terials contain reactive groups that can
error is incurred when the statistical be used for organic or inorganic species
characteristics of a population are esti- such as catalysts.
mated from a subset, or sample, of that Secondary sampling: Refers to the pro-
population. Because the sample does not cess of taking a representative portion of
include all members of the population, the primary sample to further reduce its
statistics on the sample, such as means particle size or to prepare a laboratory
and quantiles, generally differ from sample for eventual analysis.
parameters on the entire population. Sedimentation: A technique used for
Because sampling typically is done to the sizing of resins for ion-exchange
determine the characteristics of a whole chromatography; a broad distribution of
population, the difference between the beads is placed in a solvent, often water,
sample and population values is consid- in a container that is affixed to a station-
ered a sampling error. Exact measure- ary surface. Based on particle size and
ment of sampling error generally is not particle density the beads will settle at
feasible because the true population different velocities into a gradient of sizes
values are unknown; however, sampling and the fraction of interest is removed.
error can often be estimated by probabi- Very narrow cuts of particle size can be
listic modeling of the sample. obtained by sedimentation.
Sampling rate: See data acquisition Selectivity: The fundamental ability
rate. of a stationary phase to selectively re-
Sandwich technique: Injection tech- tain substances based on their chemical
nique in which a sample plug is placed characteristics, including vapor pres-
between two solvent plugs in the syringe sure and polarity. In LC, selectivity
so as to wash the syringe needle with sol- is strongly influenced by the mobile-
vent and obtain better sample transfer phase composition. In GC, carrier gas
into the inlet. has less, if any, impact on chromato-
Saturator column: See precolumn. graphic selectivity.
Scaleability: In going from analytical to Selectivity (α): Term replaced by the
preparative chromatography, refers to the separation factor. See separation factor.
Selectivity coefficient (kA/B): In ion-ex- (LOD). For a concentration-sensitive

change chromatography, the equilibrium detector such as a thermal conductivity
coefficient obtained by application of the detector or UV–vis detector: S = mmax/c
law of mass action to an ion exchanger where mmax is the peak height and c is
and characterizing the ability of an ion the solute concentration in the detec-
exchanger to select one of two ions pres- tor; units of sensitivity for a concentra-
ent in the same solution. For example, tion-sensitive detector that responds in
the exchange of Na+ for H+ in: KNa/H = millivolts are mV·mL/g. For mass-flow
[Na]S*[H]S/[Na]M[H]M sensitive detectors such as the flame-
Selectivity triangle: An approach to ionization detector: S = mmax/Wt where
classify the properties of stationary Wt is the mass of solute passing through
phases in reversed-phase LC. Results the detector per unit time; the units of
can be represented in a “selectivity S are then expressed as A•s/g or C/g.
triangle” in which the apices of the Separation: The degree of separation of
triangle represent the relative con- two peaks in time. See separation fac-
tributions of steric hindrance (χS), tor (α), relative retention, resolution.
hydrogen bonding basicity (χB) and Separation factor (α): The separa-
cation exchange capacity (χC) to se- tion factor α expresses the relative
lectivity. A graphical visualization of retention of two adjacent peaks: α =
the column selectivity allows three- t′R2/t′R1 = k 2/k1 where t′R2 is the reten-
dimensional data to be presented in tion time of the second peak and t′R1
a two-dimensional space. Provides an is the retention time of the first peak;
informative yet universal approach for k 2 and k1 are the corresponding reten-
phase classification compared to other tion factors.
models. With this model, selection of Separation impedance (E): A figure
columns of either equivalent or dif- of merit developed by John Knox to
ferent selectivity is readily achievable, compare the efficiency of two chro-
which should further facilitate the ap- matographic systems by normalizing for
plication of reversed-phase LC. both analysis time and pressure drop:
Selectivity tuning: Several techniques E = tR Δp/N 2 ν(1 + k) where tR is the re-
for adjusting the selectivity of separa- tention time, Δp is the pressure drop, N
tions involving more than one column or is the theoretical plate number, ν is the
stationary-phase type. Serially coupled reduced velocity, and k the retention fac-
columns and mixed-phase columns can tor. The lower the value of E, the better
be selectivity-tuned. the system.
Semipreparative chromatography: Re- Separation number (SN): Separation
fers to preparative LC carried out on an number, or Trennzahl (TZ ). A mea-
analytical size (4–5 mm i.d.) or a slightly sure of the number of peaks that could
larger (6–10 mm i.d.) column. Normal be placed with baseline resolution be-
injection size would be in milligram to tween two sequential peaks, z and z+1,
low gram amounts. in a homologous series, such as two
Sensitivity (S): Degree of detector hydrocarbons: SN = (t R(z+1) – t R(z))/
response to a specified solute amount (wh(z+1) + wh(z))
per unit time or per unit volume, often Septum: Silicone or other elastiomeric
defined by lower limit of detection material that isolates inlet carrier flow
from the atmosphere and permits sy- each other and based on the metal con-
ringe penetration for injection. tent of the silica. The strongest silanols
Septum purge: Carrier gas is swept are acidic and often lead to undesirable
across the septum face and out to a interactions with basic compounds dur-
separate vent so that material emitted ing chromatography.
from the septum does not enter the Silanophile: A compound that has high
column. affinity for active (acidic) silanol groups
Sequential suppression: Combina- on the silica surface. Usually a strongly
tion of chemical suppression (MSM) basic amine.
and CO 2 suppression (MCS). The Silica gel: The most widely used HPLC
background conductivity of carbonate packing. It has an amorphous structure,
and hydrogencarbonate eluents after is porous, and consists of siloxane and
suppression is approximately 10–20 µS/ silanol groups. It is used in all modes of
cm. This is a result of the dissolved LC as a bare packing for adsorption, as
carbonic acid that partially dissociates. the support for LLC or for chemically
The MCS removes the CO2 from the bonded phases, and, with various pore
suppressed eluent and therefore reduces sizes, as an SEC packing. Microparticu-
the background even further (typically late silicas of 3-, 5-, and 10-µm average
>1 µS/cm). particle diameter are used in HPLC.
Shell particle: See superficially porous Compared to irregular silicas, in mod-
particles. ern analytical HPLC columns, spheri-
Sieving: Process of passing a sample of cal silicas are preferred because of their
solid particles through a metal or plastic packing reproducibility and because they
mesh of a uniform cross-sectional area have lower pressure drops; sometimes re-
(square opening from 3 µm to 123 mm) ferred to as silica. Also used as a gas-solid
to separate particles into uniform sizes; adsorbent in GC.
can be performed under wet and dry Siloxane: The Si-O-Si bond. A principal
conditions. bond found in silica gel or for a silylated
Signal-to-noise ratio (S/N): The ratio compound or bonded phase. Stable ex-
of the peak height to the noise level. A cept at high pH values; has little effect
detector gives a valid signal if there is on the HPLC separation.
some measurable response above the Silylation: The process of reaction of
normal background noise; both detec- an organochlorosilane or organoalk-
tor sensitivity and limit of detection are oxysilane with a compound containing
dependent on the level at which the sig- an reactive group. In LC it refers to the
nal can be distinguished. A minimum process of derivatizing the solute before
S/N is equivalent to 2 but for quantita- chromatography to make the solute de-
tive methods sometimes a higher value is tectable or to prevent unwanted station-
chosen (such as S/N = 6), meaning that ary phase interactions. It can also refer
the signal is 6 times that of the baseline to the process of adding a chemically
noise. bonded phase to a solid support or to
Silanol: The Si-OH group found on deactivating the packing to cut down
the surface of silica gel. There are dif- on surface activity.
ferent strengths of silanols depending Simulated distillation (SIMDIS):
on their location and relationship to Boiling-point separation technique
that simulates physical distillation of Solid-phase extraction (SPE): A tech-

petroleum products. nique for sample preparation using a
Simulated moving bed : A chromato- solid phase packing (dp of 20–40 µm)
graphic system involving a series of contained in a small plastic cartridge
columns and valves set up to simulate or disk or in a well of a 96-well flow-
the countercurrent movement of the through plate. The solid stationary
mobile and stationary phases to allow phases used are no different than
for the continuous removal of product HPLC packings. However, although
and reapplication of sample. A complex related to chromatography, the prin-
form of recycle chromatography used in ciple is different and is sometimes
preparative-scale chromatography. referred to as digital chromatography.
Single drop microextraction (SDME): The process as most often practiced
A single drop of solvent (1–2 µL) sus- requires four steps: conditioning the
pended in the headspace can partition sorbent; adding the sample; washing
volatile analytes into the solvent; the away the impurities; and eluting the
drop can be withdrawn into the syringe sample in as small a volume as possible
and injected into a GC instrument. with a strong solvent. SPE can be per-
Single-ion conductivity (K i ): The formed in a variety of formats, such as
single-ion conductivity is proportional cartridges, disks, pipette tips, and 96-
to the concentration and the equivalent well plates, and in a variety of modes
conductivity of the respective ion. such as reversed phase, ion exchange,
Size-exclusion chromatography (SEC): and normal phase. It is a widely used
Same as steric exclusion chromatog- sample preparation technique.
raphy. Solid support: The same as support.
Slurry packing: The technique most Solid-core packing: See superficially
often used to pack HPLC columns porous particles.
with microparticles. The packing is Solid-phase microextraction (SPME):
suspended in a slurry (~10% w/v) and A technique in which a small polymer-
rapidly pumped into the empty column. coated solid fiber is placed into a solu-
Special high pressure pumps are used. tion or above the headspace of a solid or
Snyder solvent strength parameter liquid sample; analytes will diffuse into
(E0): Solvent strength parameter in ad- the coating until equilibrium is estab-
sorption chromatography; the energy of lished; for GC, the fiber containing the
solvent adsorption per unit surface area sorbed sample is transferred to the GC
occupied by the solvent. and the trapped analytes are thermally
Soap chromatography: The earlier desorbed into the column. In HPLC,
name for ion-pair chromatography. Long- solvent is used to rinse the sorbed ana-
chain soaps or detergents were used as lytes for eventual injection into the LC
the mobile phase additives. column; less popularly used in LC than
Sol gel: Silica gel formed by the aggrega- in GC.
tion of silica sol; results in type B silica Solid-phase trapping: The use of an
gel with lower surface acidity, lower trace SPE cartridge or packed column to trap
metal, lower surface area and porosity, specific analytes that flow through the
and higher high pH stability than older device; the packing material is chosen to
type A silica gels. selectively retain the analytes of interest
and let other compounds pass through tile sample residue and partially restore
unretained. column performance. Some stationary
Solute: The dissolved component of a phases may be damaged by solvent rins-
mixture that is to be separated in the ing or flushing.
chromatographic column. May be re- Solvent selectivity: Ability of a solvent
ferred to as the analyte. to influence selectivity; there is solvent
Solvent: The liquid used to dissolve a strength selectivity where a change in
sample for injection into a chromatogra- solvent strength (say from 5% B to
phy column or CE capillary; sometimes 10% B) will change band spacing or
refers to the mobile phase used in LC. solvent-type selectivity where change
Solvent demixing: Occurs when two from methanol to acetonitrile as a
solvents with very different strengths reversed-phase organic modifier will
(A = weak solvent and B = strong sol- change band spacing.
vent) are used with unmodified silica Solvent selectivity triangle: A useful
or alumina; the strong solvent (B) will guide for choosing among different sol-
be preferentially adsorbed by the active vents for the purposes of changing band
surface of the stationary phase until it spacing; solvent selectivity is dependent
is saturated; until this occurs, the weak on dipole moment, acidity, and basicity
solvent (A) will be enriched (demixed) as of the solvent molecule. For details, see
it travels down the column; eventually L.R. Snyder, P.W. Carr, and S.C. Rutan,
when the entire column is saturated with J. Chromatogr. A 656, 537–547 (1993).
B, this solvent will elute mixed with A at Solvent strength (S): Refers to the abil-
the initial strength and sample compo- ity of a solvent to elute a particular solute
nents are eluted with the sudden change or compound from a column. Described
in solvent strength. for HPLC by Lloyd Snyder for linear
Solvent effect: A solute-profile sharp- elution adsorption chromatography on
ening technique used with splitless and alumina, solvents were quantitatively
on-column injection. Condensed solvent rated in an eluotropic series; S varies
in the column during and shortly after with modifier type, stationary phase,
injection traps volatile solutes into a nar- and temperature. Less extensive data are
row band. See also retention gap. available for silica and carbon adsorbents.
Solvent exchange: The process of ex- Sonication: The use of ultrasound to
changing one solvent that may not be create vigorous agitation at the surface of
compatible with the analysis method a finely divided solid material. The direct
for a solvent that is more compatible. In method uses a specially designed inert
some cases, evaporation is used to re- acoustical tool (a horn or probe, called a
move a volatile solvent and the sample is sonotrode) placed in sample–solvent mix-
reconstituted in a different solvent. ture. In the indirect method, a sample
Solvent flooding: A source of peak- container is immersed in an ultrasonic
shape distortion caused by excessive bath with solvent and subjected to ul-
solvent condensation inside the column trasonic radiation. Dissolution is aided
during and after splitless or on-column by the ultrasonic process. Heat can be
injection. added to increase the rate of extraction.
Solvent flushing: A column rinsing The method is safe and rapid and is best
technique that may remove nonvola- for coarse, granular material. With the
indirect method, multiple samples can Spin filter: Similar to a spin column but
be done simultaneously. instead of the column packing a mem-
Sorb: The process of being retained by brane filter is used; the purpose of the
a stationary phase when the retention filter is to remove particulates.
mechanism—adsorption, absorption. Spin tube: See spin column.
partitioning—is not clear. Split injection: An injection technique
Sorbent: Refers to a packing used in ad- for GC where only a portion of the
sorptive chromatography LC. Common sample is directed to the column. The
sorbents are polymers, silica gel, alumina, ratio of the vented volume to the injected
titania, and zirconia and chemically volume is called the split ratio, which has
modified materials. typical values of 100:1, 50:1, and so on.
Soxhlet extraction: A well accepted Split injection tries to avoid overloading
technique for the extraction of com- the column; it ensures a representative
pounds from a solid sample; the sample sample reaches the column. The tech-
is placed in a disposable porous container nique is simple and rugged and protects
(thimble); constantly refluxing fresh the column. However, sample discrimi-
condensed solvent flows through the nation is possible; splitless injections are
thimble and dissolves analytes that are usually performed automatically.
continuously collected in a boiling flask; Split ratio (s): The ratio of the sample
special glassware called a Soxhlet extrac- amount vented to the sample amount
tor is designed to perform the extraction entering the column in split injection.
unattended. Higher split ratios place less sample on
Specific surface area: The surface area the column. Usually measured as the
of an LC packing based on measure- ratio of total inlet flow to column flow:
ment by an accepted technique such s = (Fs + Fc)/Fc
as the BET method using nitrogen Split vent flow rate (Fs): Carrier gas
adsorption. flow rate from the split vent, measured
Spherical packing: Refers to spherical at room temperature and pressure.
solid packing materials. In analytical Splitless injection: Derivative of split in-
HPLC spherical packings generally are jection. During the first 0.5 to 4 min of
preferred over irregular particles but in sampling the sample is not split, and en-
preparative work irregular particles are ters only the column. Splitting is restored
often used because of their lower cost. afterwards to purge sample remaining in
Spin column: A small column that the inlet. Up to 99% of sample enters the
usually contains a packing material for column. Ensures higher sensitivity than
sample cleanup or isolation; the sample split injections but flashback can occur
is added to the column, which has a se- and a higher possibility of sample deg-
lective packing material; the column, in radation is possible as a result of longer
turn, fits into a small collector tube that residence time in the hot injection port.
is placed in a centrifuge, and the liquids Square-well plates: 96-well plates that
in the tube are separated by centrifu- have square-shaped wells instead of the
gation. Spin tubes are very popular in normal round-bottom wells.
handling biological samples for isolating Stagnant mobile phase: The fraction
DNA, RNA, and other biocompounds of the mobile phase contained with the
of interest. pores of the particle.
Standard addition: Process used to im- evaporates, a thin uniform film of sta-
prove quantitation; necessary to have a tionary phase is left behind.
pure standard of known concentration. Stationary phase: The immobile phase
An unknown concentration of sample is involved in the chromatographic process.
first injected to give a peak area; then to The stationary phase in LC can be a
the unknown concentration a measured solid, a bonded, immobilized, or coated
amount of pure compound is added. As phase on a solid support, or a wall-coated
a result of the new peak area, one can phase. The stationary phase used often
determine the original concentration. An characterizes the separation LC mode.
alternative procedure is to add a constant For example, in LC, silica gel is used in
amount of unknown concentration to a adsorption chromatography, whereas an
series of standards of pure substances octadecylsilane bonded phase is used in
and to plot the peak areas obtained reversed-phase chromatography. In GC,
against the known concentrations of the liquid or polymeric stationary phases
original standards. The slope of the line are used for liquid–liquid partitioning
obtained gives the concentration of the separations, and porous-polymer, silica,
unknown. alumina, or molecular sieve packings are
Standards: A sample that contains used for adsorption and molecular size-
known quantities of the compounds based separations.
of interest. Standards are used to help Stationary phase film thickness (df ):
identify sample peaks by comparing The average thickness of the stationary-
the time in which they are eluted to the phase film coated on the walls of an
retention times obtained through the open-tubular (capillary) GC column.
injection of the sample under the same Most open-tubular GC columns have
conditions. For quantitation, external film thicknesses of 0.1–5 µm.
standards are compounds that are used Stationary zone: To be distinguished
to construct calibration curves of detec- from the stationary phase. The station-
tor output (peak area or peak height) ary zone includes the stagnant mobile
versus concentration; the concentration phase and the chromatographically ac-
of unknowns is determined by fitting tive stationary phase.
the detector output to the calibration Step gradient: See stepwise elution.
curve. Internal standards are com- Stepwise elution: Use of eluents of
pounds of known concentration with different compositions during the
different retention times that are added chromatographic run. These eluents
to the sample and relative detector re- are added in the stepwise manner with
sponses between the internal standard a pump or by a selector valve. Gradi-
and the unknown are compared in ent elution is the continuous version of
order to quantitatively measure un- changing of solvent composition.
known compounds. Steric exclusion chromatography
Static coating: A technique for station- (SEC): A major mode of LC in which
ary-phase deposition in open-tubular samples are separated by virtue of their
columns. The column is filled with a size in solution. Also known as size exclu-
solution of stationary phase and one sion chromatography, gel permeation chro-
end sealed. A vacuum, heat, or both are matography, gel filtration chromatography,
applied to the open end. As the solvent or gel chromatography. SEC is most often
used for polymer separation and charac- average particle diameter; current prod-
terization, the separation of proteins, and ucts vary from 1.5 to 2.0 µm.
the desalting of biological samples. Sulfonyl cation exchanger: A strong
Sterically protected bonded phase: cation-exchange functionality found
Bonded phase that has sterically pro- in resin-based packings; usually propyl-
tecting bulky functional groups such SO3H; may come in other cationic forms
as isopropyl and isobutyl surrounding such as sodium, ammonium, silver, and
siloxane covalent surface bond; prevents calcium.
attack on siloxane bond and prevents Sulfur chemiluminescence detection
catalyzed hydrolysis and loss of bonded (SCD): Detection method that responds
phase at low pH (< 3). to sulfur-containing compounds by gen-
Stir-bar sorbent extraction (SBSE): erating and measuring light from chemi-
Principle similar to solid-phase mi- luminescence.
croextraction (SPME) but instead of a Supercritical fluid: The defined state
coated fiber a polymer-coated stir bar of a compound, mixture, or element
is used, greatly increasing the surface above its critical pressure and critical
area, thus providing higher capacity temperature.
and greater mass sensitivity. Similar Supercritical fluid chromatography
to SPME, equilibration requires tens (SFC): A technique that uses a supercriti-
of minutes. For GC, a special thermal cal fluid as the mobile phase. The tech-
desorption unit is needed to handle the nique has been applied to the separation
stir bar; in LC, the stir bar is usually of substances which cannot be handled
rinsed off-line. effectively by LC (because of detection
Straight phase chromatography: Same problems) or GC (because of the lack
as normal-phase chromatography. of volatility). Examples are separations
Strong anion exchanger: Anion- of triglycerides, hydrocarbons, and fatty
exchange packing with strongly basic acids. GC detectors and HPLC pumps
ionogenic groups (for example, tetraal- have been used together in SFC.
kylammonium). Supercritical fluid extraction (SFE):
Strong cation exchanger: Cation- Uses supercritical f luid, most often
exchange packing with strongly acidic carbon dioxide alone or containing a
ionogenic groups (for example, sulfonic). small percent of organic modifier for
Strong solvent: In general, refers to more polar analytes, to extract ana-
a solvent which is a good solvent for a lytes from solid materials; supercritical
chemical compound; in chromatography, fluid has the diffusivity of a gas and
refers to the mobile phase constituent the solvent power of a liquid; requires a
that provides a higher solvent strength special SFE unit where the pressure and
that causes an analyte to elute more temperature can be precisely controlled;
quickly from the column; in a water- analytes are collected in a cold trap, on
acetonitrile binary solvent system for an adsorbent or in a liquid; a “green”
reversed-phase liquid chromatograhy, extraction technique.
acetonitrile would be considered to be Superficially porous particles (SPPs):
the strong solvent. Same as porous-layer bead. Recently
Sub-2-µm packing: A term that refers to there has been a revival of superficially
the use of porous packings below 2 µm porous particles based on smaller
particles (1.3–5.0 µm) with thicker po- Suppression: Method to reduce the
rous shells (0.3–0.6 µm); such particles background signal before detection (see
give similar or better performance than electrochemical suppression, chemi-
sub-2-µm particles. cal suppression, and sequential sup-
Superheated water extraction: Water pression). Typically used together with
is heated well above its boiling point conductivity detection.
in a closed pressurized system; heating Suppressor column: In ion chromatog-
changes dielectric constant and increases raphy, refers to the column placed after
the solvating power such that it becomes the ion-exchange column. Its purpose is
“organic-like.” It is a “green” method for to remove or suppress the ionization of
extracting organic analytes from solid buffer ions so that sample ions can be
matrices. observed in a weakly conducting back-
Support: Refers to solid particles. Sup- ground with a conductivity detector;
port can be naked, coated, or have a sometimes rather than a column, mem-
chemically bonded phase. The solid brane suppressors are used.
support doesn’t contribute to the liquid– Surface area: In an adsorbent, refers to
liquid chromatographic process but is the total area of the solid surface as de-
active for adsorptive processes. termined by an accepted measurement
Support-coated open-tubular column technique such as the BET method,
(SCOT): A capillary column in which which uses nitrogen adsorption. The
stationary phase is coated onto a sup- surface area of a typical porous adsor-
port material that is distributed over the bent such as silica gel can vary from 100
column inner wall. A SCOT column to 600 m2/g.
generally has a higher peak capacity Surface coverage: Usually refers to the
than a wall-coated open tubular col- mass of stationary phase per unit area
umn (WCOT) with the same average bonded to an LC support. Often ex-
film thickness. See wall-coated open pressed in micromoles per square meter
tubular column WCOT. of surface. Sometimes %C is given as an
Supported liquid extraction (SLE): A indicator of surface coverage.
technique based on the principles of liq- Surrogate samples: A pure analyte that
uid–liquid extraction where the aqueous is extremely unlikely to be found in any
phase is supported on a bed of highly sample, and which is added to a sample
purified, high surface area diatomaceous aliquot in known amounts before ex-
earth (in a tube, cartridge, or 96-well traction and is measured with the same
format); this aqueous phase may be procedures used to measure other sample
buffered and may contain the sample components. A surrogate behaves simi-
to be partitioned; the organic phase is larly to the target analyte and is most
then percolated through the packed bed often used with organic analytical pro-
allowing for intimate contact with the cedures. The purpose of a surrogate ana-
dispersed aqueous phase. The effluent lyte is to monitor method performance
collected at the exit of the column con- with each sample.
tains the extracted analytes; compared Swelling or shrinking: Process where
to LLE, the SLE technique is miniatur- resins and gels increase (or decrease)
ized, easily automated, and provides ex- their volume because of their solvent en-
cellent extraction efficiency. vironment. Swelling is dependent on the
degree of crosslinking; low cross-linking A typical example of a tailing phenom-

resins will swell and shrink more than enon would be the strong adsorption of
highly cross-linked resins. If swelling amines on the residual silanol groups of
occurs in a packed column blockage or a low coverage reversed-phase packing at
increased back pressure can occur. In ad- intermediate pH values. Tailing can also
dition, column efficiency can be affected. result from injecting an excessive mass
Syringe filter: A small plastic holder or sample, from badly packed columns,
containing a membrane filter that has from excessive extracolumn volume, poor
Luer-lock fittings at both the top and the fittings, and excessive detector volume,
bottom so that it can be affixed to a sy- or slow detector response. Tailing peaks
ringe (which also has a Luer-lock fitting) show an asymmetry factor greater than
to pass a sample through the filtration 1.0; see asymmetry factor.
media. Syringe filter diameters can range Tailing factor: U.S. Pharmacopeia
up to 90 mm. measure of peak asymmetry defined as
System dispersion: The contribu- the ratio of the peak width at 5% of the
tion to band broadening outside of the apex to two times the distance from the
column itself; it generally refers to the apex to the 5% height on the short time
instrumental contributions as well as side of the peak. Greater than unity for
other extra column contributions. With tailed peaks. See Figure 1 and asym-
newer high efficiency columns, decreas- metry factor.
ing the system dispersion contributions TCDD: Tetrachlorodibenzo-p-dioxin.
will result in better chromatographic TCEP: Stationary phase for GC: tris-
performance. cyanoethoxypropane.
System peak: The system peak is the Tedlar bags: Used for grab sampling
peak of the eluent ion. There is no pos- of air or other gases; Tedlar (Dupont)
sibility of quantifying that peak. It is ansampling bags are a whole-air sampling
unwanted peak in the chromatogram. A device for high-level volatile organic com-
possible explanation of the system peak: pounds (VOCs) and permanent gases.
Because of the sample injection, the ion Several EPA, NIOSH, and OSHA meth-
exchange equilibrium of the eluent ions ods exist for bag sampling for a variety of
get disturbed. The re-equilibration pro- applications: stationary sources emissions,
cess yields in this additional peak. It ap-workplace atmospheres, ambient, indoor
pears in suppressed and nonsuppressed air quality, and breath analysis. The
IC, but it is pretty small with suppression.
unique design of these sample bags in-
Using sequential suppression minimizes corporates the sampling septum directly
the system peak. in the valve (polypropylene or stainless
steel construction), providing easier use
T and lighter weight than other styles.
Tailing: The phenomenon where the Ternary mobile phase: Mobile phase
normal Gaussian peak has an asymme- consisting of a mixture of three individ-
try factor greater than one. The peak will ual solvents or buffers or both.
have an extended trailing edge. Tailing is Theoretical plate: A hypothetical entity
caused by sites on the packing that have inside a column that exists by analogy to
both a stronger-than-normal retention for a multiplate distillation column. As sol-
the solute and slower desorption kinetics. utes migrate through the column they
partition between the stationary phase inner column diameter and, ignoring
and the carrier gas. Although this process stationary-phase contributions to band
is continuous, a stepwise model is often broadening: Hmin = (dc/2)((1 + 6k + 11k2)/
visualized. One step roughly corresponds (3(1 + k)2 ))1/2
to a theoretical plate. Theoretical plate height, reduced (h):
Theoretical plate height (H): The dis- Used to compare efficiencies of different
tance along a chromatographic column columns. A reduced plate-height value
that corresponds to a single theoretical of 2 or less at the optimum velocity is
plate. H = L ⁄N where L is column length considered to be a well-packed column.
and N is the number of theoretical plates. For packed columns: h = H/dp. For open-
A carryover from distillation theory; a tubular columns: h = H/dc
measure of efficiency of a column. For Theoretical plate number (N): The
a typical well packed HPLC column, H number of theoretical plates measured in
should be about 2–3 dp for 5-µm particles, a column. A concept described by Mar-
usually in the range of 0.01–0.03 mm; tin and Synge. Relates chromatographic
modern superficially porous and sub- separation to the theory of distillation.
2-µm particles sometimes show plate The length of column that corresponds
heights of less than 2 dp. In open-tubular to a single theoretical plate relating to
column GC, H should be between 0.5–2 this concept is called the plate height or
times the column inner diameter. HETP height equivalent to a theoretical plate.
is a deprecated term for the plate height. The larger the plate number, the more
The combined van Deemter–Golay equa- theoretical plates the column possesses.
tion gives the theoretical plate height for a A typical well-packed HPLC column
chromatography column: H = A + B/u– +u– with a 5-µm porous packing in a 15-cm
(CM + CS) where A is the contribution due column of 4.6-mm i.d. should show
to eddy diffusion and multipath flow and 10,000–12,000 plates, which is the same
B is the contribution from longitudinal number of plates for a 5-cm column of
solute diffusion in the mobile phase. The the same internal diameter packed with
C terms are related to the effects of diffu- sub-2-µm particles or superficially po-
sion on mass transfer; CM in the mobile rous particles. A typical 25-m, 0.25-mm
phase and CS in the stationary phase. See i.d. open-tubular GC column with a thin
A term, B term, C term, Golay equation, stationary-phase film of 0.25 µm or less
and van Deemter equation. should exhibit 50,000 theoretical plates
Theoretical plate height, minimum or more. The theoretical plate number is
(Hmin): The minimum of the van Deem- calculated from a chromatogram as fol-
ter curve that results from a plot of H lows: N = 16(tR/wb)2 = 5.54(tR/wh)2 where
versus u (LC) or H versus u– (GC). This wb is the width at the peak base and wh is
value represents the most theoretical the peak width at half-height. See theo-
plates that can be obtained for a certain retical plate height.
column and mobile phase system. Usu- Theoretical plates, effective (Neff): The
ally occurs at excessively slow flow rates. true number of theoretical plates in a col-
Also known as the optimum plate height. umn. The number of effective theoretical
For well-packed columns it is typically plates corrects theoretical plates (N) for
2–3 times the particle diameter; for hold-up volume: Neff = 16(t′R /wb)2 where
open-tubular columns 0.5–2 times the t′R is the adjusted retention time and wb
is the peak width at base. It is a better Time-integrated sampling: In gas

figure of merit than simple plate number sampling, to obtain a more represen-
when comparing devices of very different tative sample requires time-integrated
geometries and phase ratios; sometimes sampling. A flow restrictor is used to
referred to as effective plate number. spread the sample collection flow over a
Theoretical plates, required (N req ): specific time period to ensure an “aver-
Number of theoretical plates required age” composited or time-weighted aver-
to yield a particular resolution (R) at a age (TWA) sample. A TWA sample will
specific peak separation (α) and reten- accurately reflect the mean conditions
tion factor (k): Nreq = 16R 2 (α/(α – 1))2 of the ambient air in the environment
((k + 1)/k)2 and is preferred when, for regulatory
Thermal desorption: The use of heat to or health reasons, a typical exposure
desorb analytes from SPME fibers, an concentration is required for a situation
SBSE bar, or solid matrices placed in a that may have high variability, as in an
thermal desorption tube. occupational setting.
Thermal extraction: Uses high tem- Titania: TiO2, is an uncommon adsor-
peratures (below pyrolysis tempera- bent used in adsorption chromatography;
tures) to extract stable analytes from also used as an SPE sorbent primarily for
porous solid matrices; samples are removal of phosphorous-containing com-
placed in thermal desorption tubes just pounds such as phospholipids.
as in thermal desorption. TMS: Trimethylsilyl (a chemical de-
Thermal- conductivit y detection rivative). In LC, the TMS group is fre-
(TCD): A thermal-conductivity detec- quently found on endcapped silica gel-
tor measures the differential thermal based columns.
conductivity of carrier gas and refer- Tortuosity (tortuousity factor) (ω): A
ence gas flows. Solutes emerging from a property of a packed column that con-
column change the carrier-gas thermal trols the inhibition of longitudinal dif-
conductivity and produce a response. fusion of the solute as it diffuses along
TCD is a universal detection method the column axis. The B term in the van
with moderate sensitivity. Deemter equation is proportional to the
Thermally tuned tandem column chro- tortuousity.
matography (T3C): A form of LC in Total mobile-phase volume (Vt): In
which two columns with distinctly dif- SEC the total volume of mobile phase
ferent selectivities are placed in tandem in the column. The same as VM. Also
and operated at two different tempera- known as the totally included volume.
tures so as to optimize the resolution and/ Total permeation volume (Vp): The
or speed of analysis. A common eluent retention volume on an SEC packing
is used in both columns and the entire where all molecules smaller than the
sample passes through both columns smallest pore will elute. In other words,
and is detected with a single detector. It at Vp all molecules totally permeate all of
is not a two-dimensional technique in the pores and are eluted as a single peak.
that each sample component gives only Total porosity (εT ): The ratio of the total
a single peak. volume of mobile phase in the column to
Thermionic specific detection (TSD): the total column volume: εT = VM/Vc =
See nitrogen–phosphorus detection. εe + εi(1 – εe)
Totally porous packing: A stationary characterized by chaotic and stochastic

phase that is a porous matrix. Solutes property changes. Flow in which the ki-
penetrate the porous to interact with the netic energy dies out as a result of the ac-
stationary phase. tion of fluid molecular viscosity is called
Trace enrichment: Technique where laminar flow. Although there is no theo-
trace amounts of compounds are re- rem relating the nondimensional Reyn-
tained on an HPLC or precolumn olds number (Re) to turbulence, flows at
packing out of a weak mobile phase Reynolds numbers larger than 4200 are
or solution and then are eluted by the typically (but not necessarily) turbulent,
addition of a stronger mobile phase in whereas those at low Reynolds numbers
a concentrated form. The technique usually remain laminar.
has been most successfully applied in Turbulent flow: A form of fluid motion
the concentration of trace amounts of in which the flow ceases to be smooth
hydrophobic compounds (for example, and steady, and becomes chaotic and
polynuclear aromatic hydrocarbons) out fluctuates with time. It is characterized
of water using a reversed-phase packing. by a pressure drop significantly higher
A strong solvent such as acetonitrile than that which would be extrapolated
serves to elute the enriched compounds. from the laminar region to achieve the
Trapping: Process of using a solid mate- same volumetric flow rate.
rial (such as silica gel, polymer, or in- Turbulent flow chromatography:
organic sorbent) or liquid solution to Chromatography performed at very
physically or chemically retain solutes of high linear velocities with large particles,
interest from a diluted stream of liquid if present, under conditions using high
or gas. Frequently used to concentrate Reynolds numbers. At these conditions
analytes for more sensitive analysis. the H versus u curves show a decrease
Tre n n z a h l (T Z ): See se paration in H with increase in u. Turbulent flow
number. chromatography can be used for separa-
Triethyl amine: A very common addition or sample preparation.
tive used to block silanol groups in re- Two-dimensional chromatography:
versed-phase LC when separating basic A procedure in which part or all of
analytes. the separated sample components are
Trifluoroacetic acid: A very common subjected to additional separation steps.
mobile phase additive in reversed-phase This can be done by conducting a par-
LC for peptides and proteins. Also a ticular fraction eluted from the first
derivatization reagent for amines and column into a second column or sys-
carboxylic acids. tem having a different separation char-
Tryptic digestion: A method of selec- acteristic. It includes techniques such as
tively and reproducibly dissecting pep- two-dimensional thin-layer chromatog-
tide chains of proteins to yield a char- raphy using two eluent systems, where
acteristic pattern of smaller units that the second eluent is applied after ro-
allows analysis of the parent protein by tating the plate through 90°. This also
gradient elution reversed-phase liquid includes LC or GC followed by GC, or
chromatography. one LC mode followed by a different
Turbulence: In fluid dynamics, turbu- mode—for example, reversed-phase
lence or turbulent flow is a flow regime LC followed by SEC.
Two-dimensional electrophoresis: cess is used for purifying and concen-

Two-dimensional gel electrophoresis, trating macromolecular (103 –106 Da)
abbreviated as 2DE or 2D electropho- solutions, especially protein solutions.
resis, is a form of gel electrophoresis Ultrafiltration is not fundamentally
commonly used to analyze proteins. different from microfiltration, nano-
Mixtures of proteins are separated filtration or gas separation, except
by two properties in two dimensions in terms of the size of the molecules
on 2D gels. 2D electrophoresis be- it retains. Ultrafiltration is applied
gins with 1D electrophoresis but then in cross-flow or dead-end mode and
separates the molecules by a second separation in ultrafiltration undergoes
property in a direction 90° from the concentration polarization.
first. In 1D electrophoresis, proteins Ultrasonic sieving: Used for the accel-
(or other molecules) are separated in eration of sieving processes alternatively
one dimension, so that all the proteins or complementary to the classical low
will lie along a lane; the molecules arefrequency vibrators. Especially useful for
spread out across a 2D gel. Because very fine powders where ultrasound is
it is unlikely that two molecules will often the only possibility to enable the
be similar in two distinct properties, sieving process at all.
molecules are more effectively sepa- Ultrasonication: The irradiation
rated in 2D electrophoresis than in of a liquid sample with ultrasonic
1D electrophoresis. (>20 kHz) waves resulting in agita-
Type A silica: Silica gel formed by gell-
tion. Sound waves propagate into
ing soluble silicates; generally higher the liquid media result in alternating
acidity, higher surface area and poros- high-pressure (compression) and low-
ity, more trace metals, poorer high-pH pressure (rarefaction) cycles. Dur-
stability than Type B silicas. ing rarefaction, high-intensity sonic
Type B silica: See sol gel. waves create small vacuum bubbles
or voids in the liquid, which then
U collapse violently (cavitation) dur-
Ultrahigh-pressure liquid chroma- ing compression, creating very high
tography (UHPLC): Ultrahigh-pressure local temperatures; several regulatory
liquid chromatography is often used methods for environmental samples
loosely for any separation performed (for example, soils or solid waste)
at pressures greater than provided by specify ultrasonication.
conventional pumps (400 bar); origi- USP categories for chromatographic
nal meaning was for separations in the columns: United States Pharmacopeia
20,000 psi+ range. characterizes columns for use in their
Ultrafiltration: Variety of membrane HPLC methods by an “L” designation:
filtration in which hydrostatic pres- L1 = octadecylsilane, L7 = octylsilane,
sure forces a liquid against a semiper- L8 = aminopropyl, and so on. For GC
meable membrane. Suspended solids columns, a “G” designation is used:
and high-molecular-weight solutes G1 and G2 are dimethylpolysiloxane
are retained, and water and low-mo- columns, G3 is a 50% phenylmethyl–
lecular-weight solutes pass through polysiloxane column, G16 is a polyeth-
the membrane. This separation pro- ylene glycol column, and so on.
UV–Vis detection: The absorbance of van Deemter equation: An equation

light is the signal for measuring the used to explain the band broadening
chromatogram. There are four differ- in chromatography. The equation rep-
ent ways of applying UV–Vis detection resents the height of a theoretical plate
in IC: direct UV–Vis, indirect UV–Vis, (H) and has three terms. The A term is
UV–Vis after postcolumn reaction, and used to describe eddy dispersion (dif-
UV–Vis after precolumn reaction. fusion) that results from axial velocity
heterogeneity. The B term is for the
V contribution from molecular diffusion
Vacancy chromatography: Technique or longitudinal diffusion for the solute
where a mobile-phase additive causes a while passing through the column. The
positive detector signal output. When C term is the contribution from inter-
a solute is eluted from the column, it phase mass transfer and allows for the
dilutes the signal and yields a negative finite rate of transfer of the solute be-
peak (“a vacancy”). The technique has tween the stationary phase and mobile
been mostly been applied to single col- phase. In its simplest representation it
umn ion chromatography where mo- is expressed as follows: H = A + B/u +
bile phases such as citrate and phthal- C u. The van Deemter equation applies
ate buffers absorb in the UV. When a to packed columns both for LC and for
nonabsorbing anion is eluted it dilutes GC. The related Golay equation ap-
the UV-absorbing background and plies to open-tubular or capillary GC
causes a negative peak; the detector columns. See Golay equation.
output leads are usually reversed so that Velocity (u): The same as velocity,
the chromatogram looks normal. The linear.
technique has also been used in CE for Velocity, average linear (u – ): The aver-
detection. age speed at which a molecule of GC
Vacuum compensation: Method of carrier gas or LC liquid mobile phase
carrier-gas pressure control in GC with passes through a column: u– = L/t M
the column exit at mass-selective de- where L is the column length and t M
tector vacuum levels. Enabling vacuum is the hold-up or unretained peak time.
compensation adjusts the column inlet Velocity, column outlet (uo): In GC,
pressure to maintain a set flow or ve- the carrier gas velocity at the column
locity when the column exit is not at outlet. Equal to the average carrier gas
room pressure. velocity divided by the compressibility
Vacuum filtration: Using a vacuum to correction factor: uo = u– /j. The carrier
help pull liquids through a filter; espe- gas expands as it pass through the col-
cially useful for viscous liquids or very umn from the inlet to the outlet pres-
fine, low porosity filters. sure, which causes the local carrier-gas
Vacuum manifold: A manifold de- velocity to increase along the column.
signed for SPE cartridges and SPE The outlet velocity is always greater
disks that uses a vacuum to pull liquids than the average velocity. See com-
through the packed beds; pressurized pressibility correction factor.
manifolds are also available. Vacuum Velocity, interstitial (u e ): The ac-
manifolds can process multiple samples tual velocity of the eluent as it moves
from ranging from 8 to 24 at a time. through the column flowing around
the particles: ue = F/(Ac εc). The inter- operated unpacked but with the same
stitial velocity is the basis for computa- flow rate: us = F/Ac
tion of the reduced velocity. Velocity, zone (u z ): The velocity of
Velocity, linear (u): The velocity of travel of the solute zone: uz = u/(1 + k)
the mobile phase moving through the = L/tR
column. Expressed in cm/s. In LC it is Viscosity (η): Also referred to as mo-
directly related to column flow rate by bile phase viscosity. The viscosity of
the cross-sectional area of the column the mobile phase varies with the tem-
and is determined by dividing the col- perature of the column. Column back
umn length (L) by the retention time pressure is directly proportional to
of an unretained compound: u = L/tM. solvent viscosity. Low-viscosity mobile
In GC, the speed at which carrier gas phases generally give better efficiency
moves through the column usually is than less viscous ones because diffu-
expressed as the average linear velocity sion coefficients are inversely related
to account for carrier-gas compressibil- to solvent viscosity. For example, in
ity. See hold-up time; velocity, aver- reversed-phase LC, column efficiency
age linear; velocity, column outlet. is higher with acetonitrile as an organic
Velocity, mobile phase (u M): The ve- modifier than with isopropanol which
locity at which the liquid mobile phase is more viscous. In GC, the viscosity
percolates through the bed of particles of the gaseous mobile phase increases
in an LC column: uM = L/tM. See veloc- with temperature, which causes the
ity, linear; velocity, average linear. carrier-gas flow rate and linear veloc-
Velocity, optimum linear (Uopt): The ity to decrease during temperature pro-
mobile-phase velocity corresponding to gramming if the inlet pressure is held
the minimum theoretical plate height, constant. Different GC carrier gases
ignoring stationary-phase contributions such as nitrogen, helium, or hydrogen
to band broadening. In open-tubular have different viscosities.
GC: u opt = 8 (D G/d c) ((3(1 + k)2)/(1 + Void: The formation of a space or gap,
6K + 11k2))1/2 usually at the head of the column,
Velocity, reduced (ν): Along with the caused by a settling or dissolution of
reduced plate height, the reduced veloc- the column packing. A void in the col-
ity is used to compare different chro- umn leads to decreased efficiency and
matographic columns. It relates the loss of resolution. Even a small void
solute diffusion coefficient (D M ) in can be disastrous for small particle
the mobile phase to the particle size of microparticulate columns. The void
the column packing (dp): ν = u dp/Dm can be removed sometimes by filling
where u is the interstitial mobile-phase it with glass beads or the same porous
linear velocity in packed columns, or packing used in the remainder of the
the average carrier-gas linear velocity column.
in GC. For open-tubular columns, the Void time: See hold-up time.
column internal diameter is used in- Void volume: See hold-up volume.
stead of the particle diameter. Volume, liquid phase: See volume,
Velocity, superficial (u s ): The hy- stationary phase.
pothetical velocity the mobile phase Volume, mobile phase (VG or V M ):
would have if the same column were For wall-coated open-tubular columns
(WCOT), ignoring the stationary phase Weak solvent: In general, refers to a

film thickness (df = 0): VG ≈ L(πdc2)/4 solvent that is a poor solvent for a par-
Volume, stationary phase (VL or VS): ticular chemical compound; in chro-
Volume of the liquid stationary phase matography, refers to the mobile phase
contained in the column. The ratio of constituent that provides a low solvent
the mobile-phase volume to the station- strength and causes an analyte to be
ary liquid-phase volume is the phase eluted more slowly from the column. In
ratio of a GC column. See phase ratio. a water–acetonitrile binary solvent sys-
tem for reversed-phase LC, water would
W be considered the weak solvent; in a bi-
Wall effect: The consequence of a looser nary solvent eluent would normally be
packing density near the walls of a rigid the “A” solvent.
HPLC column. The mobile phase has Wide-bore open-tubular column
a tendency to flow slightly faster near (WBOT): Open-tubular (capillary) GC
the wall because of the increased local column with a nominal inner diameter
permeability. The solute molecules that dc of ≥530 μm.
happen to be near the wall are carried Wilke-Chang equation: A semiempiri-
along faster than the average of the cal equation used to estimate diffusion
solute band and, consequently, band coefficients in liquids as a function of
spreading results and there is a loss of molecular size of solute and solvent
column efficiency. viscosity.
Wall-coated open-tubular column
(WCOT): An open-tubular (capillary) GC XYZ
column in which a uniform stationary Xerogels: Gels used in SEC that will
phase film is coated directly onto the col- swell and shrink in different solvents;
umn wall. See also porous-layer open tu- also refers to silica-based packings that
bular (PLOT) column, support-coated are prepared from acidification of solu-
open-tubular column (SCOT). ble silicates to give a amorphous, high
Wash step: See rinse step. surface, high porosity, rigid particle.
Water dip: Indicates the hold-up time in Zero dead volume: Any fitting or com-
suppressed IC. Usually a negative peak ponent in which all of the volume is
that corresponds to the volume of sample swept by the eluent. See dead volume.
water. The area depends on the difference Zirconia: Porous zirconium oxide; used
in concentration of the eluent anions be- as a chromatographic sorbent usually
tween eluent and sample. The water dip coated or bonded with polymeric or-
is large if the sample is almost ultrapure ganic phase.
water. If the sample is diluted with eluent Zone: See band.
there will be almost no water dip. Zwitterionic packing: A packing ma-
Wea k a n ion e xc ha n ge r : A nion- terial for HPLC that carries both posi-
exchange packing with weakly basic tive and negative charges on its surface;
ionogenic groups (for example, amino zwitterionic packings are useful in the
or diethylaminoethyl). HILIC mode.
Weak cation exchanger: Cation-ex- Zwitterions: Compounds that carry
change packing with weakly acidic iono- both positive and negative charges in
genic groups (for example, carboxyl). solution.
A New Paradigm in Light Scattering Technology
500
A) 500 ng → 250 ng → 50 ng
490
Right Angle (90 °) LS Monomer

480
• Direct Molecular Weight by Low Angle

470
Aggregates - Dimer
• Rg Below 10 nm for the First Time

mAb Fragment
RALS (mV)
460
• HPLC/UHPLC Compatibility
450
• Powerful and Intuitive Software
440
mAb Monomer, Fragment, Oligomers and Aggregate 430

Detection with
2 2.5 3 3.5 Sensitivity
4 4.5 5 5.5 6 6.5Down
7 7.5 to
8 Only
8.5 9 2
9.5 ng!
10
Retention time (minutes)
B) 10 ng → 4 ng → 2 ng 6
Monomer
Right Angle (90 °) LS 5
Aggregates
Fragment
4
RALS (mV)
3
10 ng
2
4 ng
1
2 ng
0
2 2.5 3 3.5 4 4.5 5 5.5 6 6.5 7 7.5 8 8.5 9 9.5 10

Retention time (minutes)
• Herceptin® biosimilar
• TSKgel UP-SW3000, 2 µm, 4.6 mm ID × 15 cm column
• LenS3 MALS detector
Tosoh Bioscience and TSKgel are registered trademarks of Tosoh Corporation. LenS is a trademark of Tosoh Bioscience LLC.
Herceptin is a registered trademark of Genentech, Inc.
www.tosohbioscience.com

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Volume 38 | Number s2 | February 2020

e in nnects ounde group b e a wo dis ions; i tog

Learn how to end you dependence on reference

Senior Vice President,

Associate Editor I.T. & Enterprise Systems

• Technical Articles • Education & Instruction

Sign up today to access Restek’s

W e are pleased to pres- tions and that nomenclature will be

Absorption: The process of retention in

by the addition of water or other polar Adsorption isotherm: In adsorption, a

organic solvents to maximize charge-

Normalized peak height

being in a neutral form. An example tion of the chromatographic band as it

Figure 2: Widths of a Gaussian peak at various heights as a function of the standard

Blank: More correctly named method Breakthrough volume: The volume at

used for more concentrated samples, Capillary tubing: Tubing to connect

Chemical suppression: Remove back- Chiral stationary phase (CSP): A sta-

of the selectivity throughout the of two columns connected in series by

Conductivity: Conductivity is the in- method used by analytical chemists

Core–shell: See superficially porous HPLC and CE (MEKC) by a partition-

reversed-phase chromatography as a Degassing: The process of removing

and retain lower-molecular-weight salts layer of material that contains holes

in cm 2/s, the diffusion coefficient is Dimethylchlorosilane (DMCS): Some-

48 Position Positive Pressure Manifold

Contact your UCT sales representative

a given temperature is an example of of solvent containing extracted ana-

Distribution constant (coefficient) nal fluid, saliva) as well as other non-

evaporated to yield a thin coating of any expression is misleading as the suppres-

and pressure for GC columns, as well ment chromatography and frontal

sometimes are hard to break so that isolating a compound of interest; sol-

External calibration: Calibration mode (d p = 1.5–5 µm). Separation times in

and the used filter is replaced with a heteroatomic solutes in a hydrogen-air

Fluoro phase: One of a family of Freeze drying: The process of removing

to measure hold-up volume and hold- Homogenization: The process of mak-

chromatography. However, unlike chromatography; usually little or no

Impinger: Impingers are glass bubble analytes; a UV-absorbing or f luores-

injection, on-column injection, pro- stance being quantitatively determined

It is typically expressed as the average Isotope-coded affinity tags (ICAT):

Kilopascal (kPa): A unit of pressure. normal volumes of sample in a solvent

Linear elution adsorption chromatog- found in the organic solvent; by the

(unrelated) meaning is the amount of the polymer matrix. Bioreactive mol-

Membrane filtration: Membrane filter; to the desired limits of precision and

currently in its infancy, and applications Microporous resin: Same as microre-

agate, tungsten carbide, or PTFE- Mobile-phase modifier: Modifiers are

and macropores. These pores provide tryptic peptides to analyze an entire

called throughpores (megapores or Phenol extraction: A sample prepara-

pH range and higher sample capacity a submicrometer-thick shell; wide pore

Preparative chromatography: Refers proximated as Δp ≈ (1012 L η u–)/(dp2 ), where

electrode surface. Typical applications lyzed by GC. Often applied to polymer

maximum. It is usually measured from lipophilicity) of the solutes; the more

at the source; sample tracking can be as reproducibility of results on columns of

Selectivity coefficient (kA/B): In ion-ex- (LOD). For a concentration-sensitive

that simulates physical distillation of Solid-phase extraction (SPE): A tech-

degree of crosslinking; low cross-linking A typical example of a tailing phenom-

is the peak width at base. It is a better Time-integrated sampling: In gas

Totally porous packing: A stationary characterized by chaotic and stochastic

Two-dimensional electrophoresis: cess is used for purifying and concen-

UV–Vis detection: The absorbance of van Deemter equation: An equation

(WCOT), ignoring the stationary phase Weak solvent: In general, refers to a

Right Angle (90 °) LS Monomer

• Direct Molecular Weight by Low Angle

• Rg Below 10 nm for the First Time