Hydrophobic Extraction Columns

This sorbent is composed of a silica backbone bonded with hydrocarbon chains. It is used to extract compounds which exhibit non-polar or neutral characteristics out of complex matrices. The C18 phase is the most widely used for non-polar interactions because of its nonselective nature; C18 will extract a large number of compounds with differing chemical properties. To enchance selectivity, UCT offers a wide range of hydrophobic sorbents, from C2 to C20. Multiple chain configurations are available for some sorbents. Endcapped or unendcapped sorbents are available for all chain lengths.

Mechanism of Hydrophobic Bonding

Compounds are retained by non-polar interactions from polar solvents or matrix environments. They are bound by dispersion forces / van der Waals forces. Elution, or disruption of the non-polar interactions is achieved by solvents or solvent mixtures with sufficient non-polar character. Some polar solvents, such as acetonitrile have enough non-polar characteristics to disrupt non-polar binding to cause elution of a compound from the sorbent. Methanol can be used as well, although it should be noted that it will take off both polar & non-polar analytes of interest & interferences.

Example of a hydrophobic Phase           

                Silica Backbone      

                Hydrocarbon Chain

Analytes*

alkanes

alkenes

aromatics

neutral compounds

Washes

aqueous,

usually with

some polar

organic solvent

Elutions

non-polar

to

polar organic

*typical compounds which can be extracted using hydrophobic columns

Example of hydrophobic bonding

Hydrophobic Sorbents & Structures

Sorbent

C2 ethyl

C3 propyl

C4 n-butyl

Ci4 isobutyl

Ct4 tertiary butyl

C5 pentyl

C6 hexyl

C7 heptyl

C8 octyl

C10 decyl

C12 dodecyl

C18 octadecyl

C20 eicosyl

C30 tricontyl

Cyclohexyl

Phenyl

Structure

-SiCH2CH3

-Si-(CH2)2CH3

-Si-(CH2)3CH3

-Si-CH2CH(CH3)2

-Si-C(CH3)3

-Si-(CH2)4CH3

-Si-(CH2)5CH3

-Si-(CH2)6CH3

-Si-(CH2)7CH3

-Si-(CH2)9CH3

-Si-(CH2)11CH3

-Si-(CH2)17CH3

-Si-(CH2)19CH3

-Si-(CH2)29CH3

Unendcapped vs. Endcapped

Bonded phases are manufactured by the reaction of organosilanes with activated silica.  During the polymerization reaction of carbon chains to the silica backbone, a very stable silyl ether linkage forms. Our unendcapped columns allow hydroxyl sites to remain, thus making these columns slightly hydrophilic. In order to decrease this slight polarity, these hydroxyl sites are deactivated. Proprietary bonding techniques ensure that these sites are 100% reacted, leading to a complete endcapping.  Because there are no hydroxyl sites left, our endcapped columns are more hydrophobic than our unendcapped columns.

Hydrophilic Normal Phase Columns

This sorbent is composed of a silica backbone bonded with carbon chains containing polar functional groups. Groups which will possess such polarity include amines, hydroxyls and carbonyls.

Mechanism of Hydrophilic Bonding

Compounds are retained on hydrophilic sorbents through polar interactions including hydrogen bonding, pi-pi or dipole-dipole interaction. These types of interactions occur when a distribution of electrons between individual atoms in functional groups is unequal, causing negative and positive polarity. Compounds typically extracted on a hydrophilic column include analytes which have polar groups, including amines, hydroxyls and carbonyls. Elution is performed by strong polar solvents.

Example of a hydrophilic Phase           

                Silica Backbone      

                Hydrocarbon Chain

Analytes*

R-OH, R-SH

R-NH2,

R2-NH,

R3-N

Washes

non-polar organic

solvents

ie: hexane/ethyl acetate (80:20)

methylene chloride

Elutions

polar organic

solvent

usually with

some aqueous

*typical compounds which can be extracted using hydrophilic columns

Example of hydrophilic bonding

Hydrophilic Sorbents & Structures

Sorbent

Silica

Diol

Cyanopropyl

Structure

-SiOH

-Si-(CH2)3OCH3CHOHCH2OH

-Si-(CH2)3CN

Note:

If un-ionized, ion exchange sorbents can

be used as hydrophilic (polar) sorbents.

Ion Exchange Extraction Columns

This sorbent is composed of a silica backbone bonded with a carbon chain terminated by a negatively or positively charged functional group. Ion exchange interactions occur between a sorbent that carries a charge and a compound of opposite charge.

This electrostatic interaction is reversible by neutralizing the sorbent and /or analyte. Ion exchange bonds can also be disrupted by introduction of a “counter ion” to compete with the analyte for binding sites on the sorbent.

Mechanism of Ion Exchange Bonding

Compounds are retained on the sorbent through ionic bonds. Therefore, it is essential that the sorbent and the analyte to be extracted are charged. Generally, the number of molecules with charged cationic groups increases at pH values below the molecules pKa value. The number of molecules with charged anionic groups decreases at pH values below the molecule’s pKa value. To ensure 99% or more ionization, the pH should be at least two pH units below the pKa of the cation and two pH units above the pKa of the anion. Elution occurs by using a solvent to raise the pH above the pKa of the cationic group or to lower the pH below the pKa of the anion to disrupt retention. At this point, the sorbent or compound will be neutralized.

Analytes*

Anions

Cations

Washes

Organic solvent or aqueous buffer at pH that allows the ion to remain charged AND/OR at a low ionic strength AND/OR at a weak concentration.

Elutions

Organic solvent or aqueous buffer at pH that would neutralize the ion AND/OR at a high ionic strength AND/OR at a strong concentration.

Example of a Cation Exchange Phase  

                Silica Backbone      

                Hydrocarbon Chain

Example of a Anion Exchange Phase           

                Silica Backbone      

                Hydrocarbon Chain

Ion Exchange Sorbents & Structures

Note: Neutralization can occur on either the sorbent or the analyte of interest.

Either will disrupt the bond of the desired compound.

Copolymeric Extraction Columns

(Ion Exchange with Hydrophobic Character)

This sorbent is composed of a silica backbone with two types of functional chains attached - an ion exchanger or polar chain and a hydrophobic carbon chain. Our copolymeric phases are produced in a way to allow for equal parts of each functional group to attach to the silica backbone. This copolymerization technique yields reproducible bonded phases and unique copolymeric chemistries which allow the controlled use of mixed mode separation mechanisms.

This type of dual chemistry is beneficial especially when one is looking for both a neutral & charged compound. This is common when a neutral parent drug metabolizes & becomes a charged compound.

Example of a Copolymeric Phase

Analytes*

Cations/Anions

Alkanes

Alkenes

Aromatics

Washes

1) Aqueous to disrupt hydrophilic interactions.

2) Methanol to disrupt residual hydrophobic and

    hydrophilic interferences.

Elutions

1) Organic, possibly with some aqueous to elute hydrophobically bound analytes.

2) Aqueous buffer with a pH that would neutralize ionically bound analytes or an aqueous with high ionic strength or a solvent with a counter ion that would bind to sorbent.

*Typical compounds which can be extracted using copolymeric columns

Example of Copolymeric Bonding

Mechanism of Copolymeric Bonding

Using a sample composed of a theoretical neutral parent drug and its charged (acidic) metabolite, it is applied at a pH of 6 (figure 1). At this pH, many amine groups are positively charged. Since the column is also positively charged, compounds with this chemistry (cations) are repelled. Depending on the pKa of the metabolite, carboxylic acid groups may be negatively charged, allowing the metabolite to bond to the positively charged sorbent. Since the column also possesses a hydrophobic chain, the neutral parent drug also bonds to the column.

Water or a weak aqueous buffer (pH6) washes away hydrophilically bound interferences (figure 2). The column is then dried, careful to free the column of any residual aqueous phase that would interfere with elution.

Sample Application

Column Wash

Elution 1

The hydrophobically bound neutral parent drug is eluted with a solvent of minimal polarity, such as hexane/ ethyl acetate - 80:20.

Elution 2

The final elution employs an acid to neutralize the charge of acidic analytes. Ionic interaction is released, and analytes are eluted in an appropriate solvent mixture.

CLEAN SCREEN®

Copolymeric Bonded Phases for Drug Abuse Testing

Analytical demand for more efficient, robust and clean extraction of drugs from biological matrices led to the development of WORLDWIDE MONITORING® CLEAN SCREEN® sorbents. Since 1986, CLEAN SCREEN® has led the industry with dependable and reproducible Solid Phase Extraction products and applications. CLEAN SCREEN® phases are true copolymeric sorbents that contain hydrophobic and ion exchange functional groups uniquely polymerized to a silica substrate. The design and quality of CLEAN SCREEN® provides superior sample clean up, recovery and reproducibility.

Mixed mode separations allow maximum selectivity for extraction of acids, neutrals and bases. This selectivity makes CLEAN SCREEN® ideal for both screening and confirmation analysis for virtually all drug categories. CLEAN SCREEN® DAU and THC columns are used extensively by forensic and clinical chemists including:

• Post Mortem Investigations

• Criminal Investigations

• Urine Drug Testing

• Athletic Drug Testing

• Racing Laboratories

• Therapeutic Drug Monitoring

• Medical Drug Screening

Note:

If performing extractions out of viscous matrices such as tissue or horse urine, turn to our XtrackT® section, where high-flow/gravity flow columns are found. The DAU CLEAN SCREEN® sorbent as well as other phases are available in this larger particle size.

Mechanism of CLEAN SCREEN®

When a sample is loaded onto the column at pH 6, many carboxylic acid functionalities present in the sample are ionized. This creates a repulsion between the column and many sample borne interferences, thereby reducing the likelihood of their adsorbing onto the column. At this pH, ibuprofen & barbiturates are not ionized and are hydrophobically adsorbed onto the column (figure 1). At the same time, drugs with amine functionalities such as cocaine and phencyclidine adsorb onto the column via both hydrophobic and ionic attraction (figure 1).

The column can then be washed with water or weak aqueous buffers at or below pH 6 without risking loss of the analytes. After drying the column, it is possible to elute the hydrophobically bound analytes using solvents of minimal polarity such as methylene chloride or a hexane/ethyl acetate mixture (figure 2). Cationic analytes will remain bound to the column. Many compounds of intermediate polarity and potential interferences will also remain on the column. The majority of these potential interferences can be removed by using a methanol wash.

Cationic analytes bound to the column can be eluted after another drying step. The drying steps are necessary to remove water which would have prevented the water-immiscible elution solvents from optimally interacting with the analytes (figure 3).

To elute the cationic analytes, an organic solution with a high pH (between 11 & 12) should be used. A methylene chlorideisopropanol- ammonium hydroxide mixture will simultaneously disrupt these ionic interactions and successfully elute the desired compound (figure 4).

CLEAN SCREEN^® - RSV

Reduced Solvent Volume Extraction Columns

Reduced Solvent Volume extraction columns are micro bed packed columns which offer the advantages of disc technology yet maintain the proven track record of our conventional SPE columns. Reduced Solvent Volume columns use 75% less solvent than traditional packed columns. Less solvent means faster extractions, higher throughput and less waste disposal, which translates into significant savings in both time and money. Results demonstrate that therapeutic and abused drugs in urine and blood matrices can be extracted with cleanliness, high recoveries and consistent reproducibility by using the Reduced Solvent Volume extraction column.

RSV columns are available in 1 mL, 3 mL and 10 mL configurations. These columns can be used with vacuum manifolds or positive pressure, as well as conventional automated extraction equipment.

Advantages  of CLEAN SCREEN® RSV:

• 75% Reduction in total liquid volumes
• Lower cost per extraction
• Faster extraction times
• Less disposal cost
• Increased automated throughput

• 50% Reduction in eluate volume
• Faster dry down times
• Reduced exposure to organic solvents

• Superior flow characteristics
• Less flow restriction from matrix proteins or particulates
• More reliable for automated processing

• High capacity
• Greater linear range

XtrackT®

High-Flow Bonded Phases

Viscous sample matrices are frequently resistant to flow through standard solid phase columns. Increased particle size enhances flow characteristics allowing ease of sample application and analysis. XtrackT® columns are designed to give uniform flow for even the most viscous samples including equine urine, post mortem blood and tissues, meconium, amniotic fluid, milk, etc.

XtrackT® also functions as a gravity flow column for most blood and urine samples. A single column provides extraction for a broad spectrum of compounds with selective elution of acid neutrals, steroids and bases. XtrackT® yields very clean extractions and excellent recoveries without need for additional liquid clean up steps. XtrackT® is available in  hydrophobic, hydrophilic, ion exchange, and copolymeric phases, including the CLEAN SCREEN® DAU sorbent. XtrackT® is recommended for any chemist challenged by viscous sample matrices, or those desiring gravity flow capacity.

Upon request, we can also provide any of our CLEAN-UP® sorbents in this large particle size.

Advantages of XtrackT® :

• Resists plugging
• Improved flow of all sample types
• Gravity flow of most samples
• Broad spectrum extractions
• Very clean extractions
• High recoveries
• Reproducibility

CLEAN-UP®

Covalent Phase Columns

Covalent sorbent has aldehyde functional groups that are bound to the silica backbone by a hydrocarbon chain. The aldehyde group will react selectively with compounds containing a primary amine. A formal bond is created between the stationary support and the primary amine containing material.

Mechanism of Covalent Bonding

The primary amine in the sample performs a nucleophilic attack on the aldehyde functional group attached to the support. This results in a Schiff base, with the amine immobilized on the stationary support. This chemistry can be utilized to bind proteins, such as antibodies, to the support, allowing highly specific extractions.