SPE Method Development

SPE is Chromatography
Keep in mind that solid-phase extraction has the same fundamental basis as HPLC. Any knowledge
of the chromatographic behavior of the analytes of interest, and of other matrix components, can
help in choosing the proper sorbent and eluents. If, for example, you know that certain
chromatographic conditions provide excellent separation of your analyte from interferences, then
you may choose a similar SPE sorbent and solvent combination. Similarly, if you are trying to
remove an interference that coelutes in HPLC, then you know a priori that similar SPE conditions
will not be successful.

General Elution Protocols

There are two general strategies for isolating and cleaning up sample components of interest:
adsorb matrix interferences while components of interest pass through the cartridge

unretained.
adsorb components of interest while matrix interferences pass through the cartridge

unretained.
The first strategy is usually chosen when the desired sample component is present in high
concentration. When components of interest are present at low levels, or multiple components of
widely differing polarities need to be isolated, then the second strategy is generally employed.
Trace enrichment of compounds present at extremely low levels and concentration of dilute
samples are also achieved by the second strategy.

Steps of a Solid-Phase Extraction Procedure

The following section describes the steps involved in a complete solid-phase extraction procedure.
In many applications, one or more of the steps, listed below and subsequently described by
general examples, can be omitted, thereby simplifying the procedure. The procedures illustrated
here use samples containing dyes so that separations may be easily visualized. Keep in mind that
most samples contain colorless components that require some type of detector or test to locate
them in the collected fractions. Use the following information as a guideline in the development of
your own procedure or when modifying procedures published in the literature.
1.

Pretreatment of the sample

2.

Conditioning of the cartridge

3.

Loading the sample

4.

Elution of the fractions

Principal Separation Modes in Solid-Phase Extraction [SPE]

Normal-Phase Chromatography
This mode is classically used to separate neutral organic compounds whose chemical nature ranges
from hydrophobic to moderately polar.
To perform normal-phase chromatography with SPE cartridges, use a step gradient of nonpolar
solvents with a polar packing material.

1.

Condition the cartridge with six to ten hold-up volumes of non-polar solvent, usually the
same solvent in which the sample is dissolved.

2.

Load the sample solution onto the cartridge bed.

3.

Elute unwanted components with a non-polar solvent.

4.

Elute the first component of interest with a more polar solvent.

5.

Elute remaining components of interest with progressively more polar [stronger] solvents.

6.

When you recover all of your components, discard the used cartridge in a safe and
appropriate manner.
This procedure is illustrated in the figure below for a sample containing a mixture of three neutral,
relatively non-polar organic dyes [yellow, red, and blue] that appears black when initially loaded
onto the cartridge bed.
Illustration of a General Elution Protocol for Normal-Phase Chromatography on SPE Cartridges
(Silica, Florisil, Alumina, Diol, CN, NH2)

Reversed-Phase Chromatography
Because of the multiplicity of aqueous samples spanning a breadth of applications from
environmental water to fruits and vegetables, from beverages to biological fluids, reversed-phase
chromatography has become the predominant mode of SPE.
To perform reversed-phase chromatography with SPE cartridges, use a gradient of strongly to
weakly polar solvents [from weak to strong solvent elution strength] with a non-polar packing
material.
1.

Solvate the silica-bonded phase or polymer packing with six to ten hold-up volumes of
methanol or acetonitrile. Flush the cartridge with six to ten hold-up volumes of water or buffer. Do
not allow the cartridge to dry out [unless using HLB].

2.

Load the sample dissolved in a strongly polar [weak] solvent [typically water].

3.

Elute unwanted components with a strongly polar solvent.

4.

Elute weakly retained components of interest with a less polar solvent.

5.

Elute more tightly bound components with progressively more non-polar [stronger]
solvents.

6.

When you recover all the components of interest, discard the used cartridge in a safe and
appropriate manner.
This procedure is illustrated in the figure below for a sample of an aqueous grape drink containing
two polar food dyes [red and blue], as well as sugar and artificial flavor [but no real grape juice!].
As prepared, this drink appears light purple in a glass, since the dye concentration is dilute. When
a portion is loaded onto a prepared SPE cartridge, the strongly retained dyes become concentrated
near the inlet in a dark purple band.
Illustration of a General Elution Protocol for Reversed-Phase Chromatography on SPE Cartridges
(C18, tC18, C8, CN, Diol, HLB, Porapak RDX, NH2)

Ion-Exchange Chromatography
Compounds that are ionic or ionizable are often best isolated using some form of ion-exchange
chromatography. This separation mode is orthogonal to the more widely used normal-phase and
reversed-phase modes and provides a powerful, selective second dimension to sample preparation
protocols.
Illustration of the Two Major Types of PhasesAnion and Cation Exchange
and How They Selectively Attract and Retain Molecules of Opposite Charge

To perform ion-exchange chromatography with SPE cartridges, use a gradient of pH or ionic

strength with an ion exchange packing material.
1.

Condition the cartridge with six to ten hold-up volumes of deionized water or weak buffer.

2.

Load the sample dissolved in a solution of deionized water or buffer.

3.

Elute unwanted, weakly bound components with a weak buffer.

4.

Elute the first component of interest with a stronger buffer (change the pH or ionic
strength).

5.

Elute other components with progressively stronger buffers.

6.

When you recover all of your components, discard the used cartridge in an appropriate
manner.
This procedure is illustrated in the figure below for a sample of an aqueous mixture of two ionic
dyes with different pKa values. When loaded onto the cartridge, both are strongly retained, and the
combination of blue and yellow components appears as a green band near the inlet.
Illustration of General Elution Protocol for Ion-Exchange Chromatography on SPE Cartridges
(NH2, Accell Plus QMA, Accell Plus CM, SCX, SAX, WCX, WAX)

Cation and anion exchangers are further categorized as either weak or strong exchangers,
depending upon the type of ionic group on their surface. Strong cation exchangers possess an
acidic surface moiety such as a sulfonic acid that is always ionized [negatively charged] over the
whole pH range. Weak cation exchangers possess an acidic surface moiety such as a carboxylic
acid that is negatively charged at high pH but neutral at low pH. Similarly, strong anion exchangers
typically bear quaternary ammonium groups that are always positively charged, while weak anion
exchangers possess primary, secondary, or tertiary amine groups that may be positively charged
at low pH but neutral at high pH.
Use the following table as a guideline to choose the appropriate SPE ion-exchange cartridge type
for your particular analyte.

Mixed-mode ion exchange chromatography combines the use of reversed-phase and ion-exchange
modes into a single protocol on a single SPE cartridge. It can be used to isolate and separate
neutral, acidic, and basic compounds from a single complex matrix. An ideal mixed-mode SPE
sorbent substrate remains water-wettable while exhibiting strong reversed-phase retention of
hydrophobic compounds. On its surface are ion-exchange functionalities of one of the four general
types just described above. Intermediate washes with organic solvent mixtures of appropriate
elution strength may be used to isolate neutral compounds [including ionizable analytes in their
neutral state]. Selective elution of ionically bound analytes may be attained by manipulating the
charge of either the analyte [when bound to strong ion exchangers] or of the sorbent [for analytes
bound to weak ion exchangers].

SPE Method Development Summary

The following table summarizes the foregoing discussion of the modes of SPE:
Summary of Utility and Practice of Principal LC Modes for Solid-Phase Extraction [SPE]

Analyte

Reversed Phase

Normal Phase

Ion Exchange

Moderate to low

Low to high

Charged or Ionizable

polarity

polarity/neutral

Separation

Separation based

Separation based on

Mechanism

on hydrophobicity

polarity

Sample Matrix

Aqueous

Non-polar organic

Separation based on charge

Aqueous/ Low ionic strength

solvent

Condition/
Equilibrate SPE
Sorbent

Preliminary Wash

1. Solvate with

Non-polar organic

Low ionic strength buffer

Aqueous/buffer

Non-polar organic

Low ionic strength buffer

Increase polar

Increase eluotropic

Stronger buffers - ionic strength or

organic content

strength of organic

pH to neutralize the charge

polar organic
2. Water

Step

Elution Steps

solvent mixture

Sorbent
Functionality

CX
[Cation

Exchange]

Exchange]

C18, tC18, C8, tC2,

Silica, Alumina,

Accell Plus QMA,

Accell Plus CM,

CN, NH2, HLB,

Florisil, Diol, CN, NH2

NH2, SAX, MAX,

SCX, MCX, WCX,

WAX

Rxn CX

RDX, Rxn RP

Sorbent Surface

AX
[Anion

Low to Medium

High to Medium

High

High

High to Medium

Low to Medium

High

High

Water, low strength

Hexane, chloroform,

Water, low

Water, low

Polarity

Typical Solvent
Polarity Range

Typical Sample

Loading Solvent

buffer

methylene chloride

strength buffer

strength buffer

Typical Elution

CH3OH/water,

Ethyl acetate,

Buffers, salts

Buffers, salts

Solvent

CH3CN/water

acetone, CH3CN

with high ionic

with high ionic

strength,

strength,

increase pH

decrease pH

Most weakly

Most weakly

Sample Elution

Most polar sample

Least polar sample

Order

components first

components first

Mobile Phase
Solvent Change

ionized sample

ionized sample

component first

component first

Decrease solvent

Increase solvent

Increase ionic

Increase ionic

polarity

polarity

strength or

strength, or

increase pH

lower pH

Required to Elute
Compounds

This has been a brief introduction to sample enrichment and purification using solid-phase
extraction [SPE]. The best way to start using SPE is to first learn what others have done with
analytes and/or matrices similar to those of interest to you. You will find > 7,700 references to the
use of SPE in the Resource Library on waters.com. Fill in the blank with a partial compound or
matrix name in the following search phrase:
Sep-Pak OR Oasis AND ______*
NOTE: Rather than risk a spelling error, use an asterisk [*] with a root name for best results. Using
this same search string, even more references [> 60,000] may be found on GOOGLE Scholar.
Further reading:
J.C. Arsenault and P.D. McDonald, Beginners Guide to Liquid Chromatography, Waters [2007];
Order P/N 715001531 on waters.com
P.D. McDonald and E.S.P. Bouvier, A Sample Preparation Primer and Guide to Solid-Phase
Extraction Methods Development, Waters [2001] Search for WA20300 on waters.com
Waters, Purity by SPE [2008]; Search for 720001692en on waters.com
U.D. Neue, P.D. McDonald, Topics in Solid-Phase Extraction. Part 1. Ion Suppression in LC/MS
Analysis: A Review. Strategies for its elimination by well-designed, multidimensional solid-phase
extraction [SPE] protocols and methods for its quantitative assessment [2005]; Search for
720001273en on waters.com

Beginner's Guide to SPE [Solid-Phase Extraction]

A Powerful Tool for Improved Sample Preparation
As an analytical scientist, you are faced with many challenges when determining what tools you
can best use to achieve the desired result. Determining which sample preparation tools and
approaches are important considerations that can significantly impact your success.
Ideally, you would be happy if you did not have to do any sample preparation. In reality, however,
sample preparation is often necessary. You may need to optimize a method for an existing sample
to improve throughput or lower the cost per analysis. Or, you may be asked to analyze a wide
variety of different types of samples to report on new compounds of interest. Each new sample
type can present different analytical challenges. In addition, scientists today are faced with the
significant challenge of reporting values at lower concentration levels than ever before, without
compromising accuracy and precision.
This book is designed to help you explore and understand a very powerful tool in sample
preparation technology: solid-phase extraction [SPE]. You will see how this technology, which uses
devices with chromatographic packing material, can help meet your analytical challenges.

Definiton of Solid-Phase Extraction

SPE is a sample preparation technology that uses solid particle, chromatographic packing material,
usually contained in a cartridge type device, to chemically separate the different components of a
sample. Samples are nearly always in the liquid state [although specialty applications may be run
with some samples in the gas phase]. Figure 1 shows a sample, which appears black, being
processed on a SPE device so that the individual dye compounds, which make up the sample, are
chromatographically separated.

Figure 1: Examples of an SPE Method

The chromatographic bed can be used to separate the different compounds in a sample, to make
subsequent analytical testing more successful. For example, SPE is often used for the selective
removal of interferences.
The technically correct name for this technology is Liquid-Solid Phase Extraction, since the
chromatographic particles are solid and the sample is in the liquid state. The same basic
chromatographic principles of liquid chromatography that are used in HPLC are also used here, but
in a different format and for a different reason. Here, chromatography is used to better prepare a
sample before it is submitted for analytical testing.
In sample preparation, samples can come from a wide range of sources. They can be biological
fluids such as plasma, saliva, or urine; environmental samples such as water, air, or soil; food
products such as grains, meat, and seafood; pharmaceuticals; nutraceuticals; beverages; or
industrial products. Even mosquito heads can be the sample! When a scientist needed to analyze
neuropeptides extracted from the brains of mosquitoes, SPE was the sample preparation method
of choice [Waters Applications Database, 1983].

Four Major Benefits of SPE

There are many benefits to using SPE, but four major benefits deserve special attention.
1. Simplification of Complex Sample Matrix along with Compound Purification

One of the most difficult problems for an analytical chemist is when compounds of interest are
contained in a complex sample matrix, such as mycotoxins in grains, antibiotic residues in shrimp,
or drug metabolites in plasma, serum, or urine. T he large number of interfering constituents or
substances in the sample matrix along with the compounds of interest makes analysis extremely
difficult.
The first problem to solve is the resulting complexity of the analysis itself due to the presence of
so many entities which must be separated in order to identify and quantitate the compound[s] of
interest. See Figure 2.

Figure 2: Example of a Complex Sample

The robustness of the assay may not be adequate, because any slight change could impact the
resolution of the separation of a critical pair of analytes.
Another consideration is that the presence of all the interferences in the original sample matrix can
result in instrument downtime due to a buildup of contamination with each injection. If the
interferences were removed as part of the sample preparation, then the compounds of interest

could be analyzed with a simpler, more robust method. This can be seen in Figure 3, which
compares the original sample on top to the new SPE-prepared sample on the bottom.

Figure 3: Comparison of Sample Matrix Complexities

An additional benefit of simplifying the sample matrix is improved quantitation accuracy. T he top
blue trace for compound 1 in Figure 4, initially appears to be acceptable. However, it really has
some contamination from the sample matrix when compared to the blank sample matrix trace in
red shown just beneath it. With a proper SPE protocol, the lower traces show the same compounds
with no problems with interference, making quantitation much more accurate.

Figure 4: Improved Quantitation with Better Sample Preparation

Another example is shown in Figure 5. T he upper trace shows significant interference from the
sample matrix on both compounds 1 and 2. T he lower trace shows much improved results [clean

base line] due to proper sample preparation with SPE. Notice a much cleaner baseline improves
the accuracy of the analytical results. Also, a much purer extract can be obtained if the sample
requires isolation and purification of that compound.

Figure 5: Significant Improvement in Baseline Using SPE Technology

2. Reduce Ion Suppression or Enhancement in MS Applications
The second problem with complex sample matrices can be seen when we look at mass
spectrometer output [LC/MS or LC/MS/MS]. For proper MS signal response [sensitivity], the
compound ion must be allowed to form properly. In cases where the formation of the compound
ion is suppressed by interferences in the sample matrix, the signal strength is greatly diminished.
We can see this effect in Figure 6. The upper output is the signal for our compounds of interest
when injected in a saline solution. The lower trace shows significant reduction in response [> 90%
suppression] of these same compounds when they were analyzed in human plasma. For the lower
trace, only a common protein precipitation step was performed. This technique does not clean up
the matrix interferences that cause the ion suppression, resulting in poor signal response.

Figure 6: Example of Ion Suppression Due to Sample Matrix

Another good example of this suppression effect can be seen in Figure 7. In the upper trace of the
MS output, where the plasma sample was prepared with just a protein precipitation step, we can
see that the terfenadine peak is suppressed by 80%. In the lower trace, where the same sample
was prepared with a SPE method, we can see minimal ion suppression. Because the interferences
from the sample matrix were removed, this allowed the compound ion to form properly, creating a
better signal.

Figure 7: Reduced Ion Suppression with Proper SPE

In some instances, interferences from the sample matrix can artificially increase the signal
reported for a compound. This is called ion enhancement, resulting in an inaccurately high
reported value. A proper SPE method will minimize this effect by cleaning away the interferences
from the compound, resulting in a more accurate reported value.
3. Capability to Fractionate Sample Matrix to Analyze Compounds by Class
An analyst may be faced with a sample that contains many compounds, with a need to separate
them by class so that further analysis can be carried out much more efficiently. For example, a soft
drink beverage contains a wide range of compounds in its formulation. An SPE method could be
developed to separate the different classes of compounds, for example by their polarity. The polar
compounds could be collected, as a separated fraction, from the more non-polar compounds.
These two fractions could then be separately analyzed in a much more efficient way because their
compounds would be more similar.
An example of the power of fractionation by SPE is shown in Figure 8. Here, a complex sample of a
dry powder [purple grape drink mix] is easily separated into four fractions: a fraction of just the
polar compounds, a purified red compound, a purified blue compound, and a fraction containing all
the remaining very non-polar compounds. You will see elsewhere in this book how very powerful
this capability can be.

Figure 8: Sample Preparation by SPE

For a more detailed discussion of sample fraction by SPE see page 125 in the Method Development
section.
4. Trace Concentration [Enrichment] of Very Low Level Compounds

Analysts today often need to report on compounds at far lower concentration levels than ever
before, as little as parts per trillion [ppt] and even lower. Typically these levels are lower in the
neat sample than the sensitivity capability of the analytical instruments.
A good example of this is the analysis for trace contaminants in environmental samples or
metabolite development over time in biological fluids. The upper trace in Figure 9 shows the poor
response of the original neat sample for the compound of interest. Using the same analytical
conditions but with the sample prepared with SPE used in a trace concentration strategy, the lower
trace shows a dramatic increase in signal strength for this compound. With this result, an accurate
calculation of the original compound concentration in the neat sample can be made.

Figure 9: Example of Trace Concentration

Without the retention capability of chromatographic packing materials in SPE, the ability to trace
concentrate a specific compound[s] would be very difficult, if not impossible, with other sample
preparation approaches.

Summary
As weve seen, an SPE device with a chromatographic bed can perform four critical functions to
make the analysis of the sample more successful. See Figure 10.

Figure 10: The Power of SPE

In this book, we have endeavored to provide all of the SPE fundamentals and success techniques
derived from scientists from all over the world who have counted on this technology in the past
thirty years. Today, scientists are finding SPE more useful than ever in solving difficult sample
preparation and analytical problems.
We hope this book will enable you to understand and master the capabilities of SPE, so that you
too can put the power of this technology to use in your laboratory.

SPE - Sample Enrichment and Purification using Solid-Phase

Extraction
Three Reasons To Use Solid-Phase Extraction (SPE)
1. You need to remove specific interferences from your sample so that they do not cause problems
for the detection and quantitation of analytes of interest. In the example shown here, an
inadequate sample preparation protocol failed to remove interferences, as seen by the residual
yellow color of the extract and many peaks overlapping the analytes of interest in the
chromatogram.

2. You need to increase the concentration of the analyte of interest in the original sample so that it
can be more readily detected and more accurately quantitated by your analytical technique. A
large sample volume may be loaded onto an SPE column if the analyte of interest is strongly
retained. Then the analyte may be eluted in a very small volume, thereby increasing its
concentration in the sample aliquot presented to your chosen analytical tool.

3. You need to remove interferences in your sample that, though invisible, suppress the signal for
the analyte of interest as detected by mass spectrometry. In the example shown here, protein
precipitation failed to remove the phospholipids from a plasma extract, causing severe ion
suppression. An optimized mixed-mode SPE protocol provides the cleanest extract and minimizes
ion suppression.

Goals and Benefits of SPE

What is Solid-Phase Extraction (SPE)?
Don't be confused by the term solid-phase extraction [SPE]. A typical SPE device has 50 times
more separation power than a simple, single liquid-liquid extraction. SPE is actually column liquidsolid chromatography. Since SPE is liquid chromatography [LC], its practice is governed by LC
principles. A sample is introduced into a column or a cartridge device containing a bed of
appropriate particles, or other form, of a chromatographic packing material [stationary phase].
Solvent [mobile phase] flows through the bed. By choosing an appropriate combination of mobile
and stationary phases, sample components may pass directly through the column bed, or they
may be selectively retained.
Individual compounds in the sample each typically appear to travel at different speeds through the
device. Using a weaker solvent causes them to move slowly and/or be strongly retained. A
stronger solvent speeds up their passage through the bed and elutes the analyte(s) in a more
concentrated volume. Elution from an SPE device is usually done by increasing the strength of the
mobile phase in a series of discrete, rather than continuous, steps during which selected analytes
or interferences are either fully retained or rapidly eluted-this variation of gradient elution called a
step gradient.
Most commonly, SPE is practiced using miniature column or cartridge devices. An example is
shown here. A mixture of three dyes is loaded onto the cartridge in a weak solvent, causing strong
sample retention in a narrow band that appears black at the column inlet. Subsequent gradient
steps, each with a successively stronger solvent, are used to elute the dyes individually [yellow,
red, then blue].

Typical SPE cartridges are low-pressure devices-constructed of solvent-resistant plastic or glassfilled with particles 30 m in diameter. Suitable flow rates may be achieved by gravity or with the
assistance of vacuum or low positive pressure. [The latter requires putting a cap on the open inlet
of a column or using a sealed device with inlet and outlet fittings.]

Importance of Sample Preparation

In the last two decades, dramatic advances in analytical instrumentation and laboratory
information management systems shifted the analyst's predominant tasks from assay
measurements to sample preparation and data processing. As the stringency of requirements for
higher sensitivity, selectivity, accuracy, precision, and number of samples to be processed has
escalated, the corresponding increases in speed and sophistication of analysis and data collection
have outpaced improvements in the many traditional techniques of sample collection and
preparation. By some estimates, 75 to 80% of the work activity and operating cost in a
contemporary analytical lab is spent processing and preparing samples for introduction or injection
into an analytical separation and/or measurement device. Clearly, efforts directed and products
designed to streamline sample preparation protocols are essential to future progress in analytical
science.

Goals of Sample Preparation

Successful sample preparation for most analytical techniques [HPLC, GC, spectrophotometry, RIA,
etc.] has a threefold objective: namely, to provide the sample component of interest

in solution

free from interfering matrix elements

at a concentration appropriate for detection or measurement.

To accomplish these goals, a sample, or a representative portion thereof [not always easy to
obtain], is prepared via traditional methods of dissolution, homogenization, extraction [liquid- or
solid-phase], filtration, concentration, evaporation, separation, chemical derivatization,
standardization [internal or external], etc.
Usually such methods are used in combinations of multiple steps, which form a sample prep
protocol. The fewer steps and methods used in any given protocol, the simpler, more convenient,
cost effective, and less time consuming it is. Simpler protocols lend themselves more readily to
automation and also lead to increased accuracy, reliability, reproducibility, and safety.

Innovation in Sample Preparation Methods

There are many ways to combine standard tools and techniques to accomplish the goals of sample
prep. However, it is best to seek innovative means to streamline sample prep protocols:

to combine the functions of several steps, if possible, into one operation;

to eliminate needless sample transfers and manipulations;

to reduce the scale as much as practicable [gaining economies of time, labor, and cost];

to use new tools in creative ways.

Benefits of Solid-Phase Extraction [SPE] Cartridges

When compared to other sample preparation processes, solid-phase extraction using SPE
cartridges offers:
Lower Cost

lower solvent consumption

lower reagent consumption
less apparatus

Greater Recoveries

minimal sample transfer

Faster Protocol

fewer steps

Greater Safety

less exposure to toxic agents

Greater Accuracy

no cross contamination

No Emulsion Problems

less sample handling

fewer steps

No Transporting of Samples to Lab

direct field sampling

Reduced Harm to Labile Samples

minimal evaporation

Minimal Glass Breakage

less glassware used, less to wash

Achieving Sample Preparation Objectives with Solid-Phase Extraction [SPE]

To remove sample constituents that elute after the analytes of interest or are strongly

adsorbed:

use solid-phase extraction with sorbent surface chemistry that is the same as that

in the analytical HPLC column.

tailor the gradient steps to elute analytes selectively.

To remove sample constituents that coelute with an analyte of interest:

use solid-phase extraction with sorbent surface chemistry and/or separation mode

different from that in the analytical column.

tailor the gradient steps to elute analytes selectively.

To enrich sample components present in low concentration:

tailor the gradient steps to elute analytes selectively.

use "large" sample volumes in adsorption-promoting solvent.

use "small" collection volume in desorption-promoting solvent.

use sorbent chemistry tailored to the analyte, independent of that in analytical

column.
carefully choose chemistry of solid-phase extraction column so further sample prep

will be unnecessary.

To desalt samples:
first, adsorb analytes on reversed-phase sorbent while salt breaks through

unretained.

then, after using water to wash away residual salt, desorb analytes using watermiscible organic solvent.

To exchange solvents:
adsorb the sample completely onto a strongly retentive sorbent and flush away the
original solvent with a weaker eluent.

elute the analyte with the desired solvent.

To fractionate classes of compounds:
use a step-gradient sequence to divide a sample-on the basis of hydrophobicity,
polarity, or charge-into fractions containing groups of analytes that share common properties.

To derivatize analytes using solid-phase reagents:

adsorb a derivatization reagent on the surface of the sorbent; then, collect the
sample (usually a gas) under conditions that favor complete adsorption of the analyte; wait for the
reaction to occur and then selectively elute the derivative.