What is Solid Phase Extraction (SPE)?
Solid Phase Extraction is used in the chemical analysis of fluid samples by removing unwanted constituents and keeping only the analytes of interest. It is a crucial sample preparation step for liquid and gas chromatography. Typical applications include the analysis of pesticide and various pollutants in water, drug in food and blood, as well as oil and grease in environmental samples.
Original Sample ===❯ Solid Phase Extraction ===❯ Analysis
The working principle behind SPE is the adsorption of analytes to the solid phase (sorbent material in SPE cartridge), thus removing them from the original liquid phase (sample fluid). The type of SPE cartridge has to be selected based on the chemical and physical properties of the analytes. Refer to our columns/cartridges section for more information.
Why automate SPE?
Solid phase extraction is a time consuming procedure that requires careful and controlled handling. Performing SPE on large sample batches can be strenuous and prone to human error. In fact, majority of the time spent and errors during chemical analysis come from sample preparation. PromoChrom's automated SPE equipment can alleviate this problem by executing the steps unattended and with high repeatability and reproducibility. The open design makes it easy to place samples, solvents, cartridges and collection vials. Once that is done, all the user has to do is input the desired SPE steps and hit start. The simple interface allows users to choose which sample and solvent to use, as well as the flow rate and volume. To see all features, take a look at our automated offline SPE and automated online SPE pages.
What is different between conventional column chromatography and solid phase extraction?
Column chromatography is a very old technique for separation. It uses silica gel, alumina, or other sorbent to separate mixtures. The mode is like the normal phase HPLC. Around 40 years ago, silica gel bonded with C18 is packed into small columns (sometimes called cartridges) and is used to extract organic compounds from water, as an alternative of liquid/liquid partition. This practice is called solid phase extraction. It is actually one type of column chromatography, which is like the reverse phase HPLC. Nowadays, Silica gel, Alumina are also packed into small cartridges and referred to as SPE columns. So now it seems as long as the sorbent is packed into small cartridges, the practice is called solid phase extraction.
What are normal procedures in solid phase extraction?
SPE normally involves 5 steps: 1) activate to make the sorbent wet well; 2) equilibrate to adjust the extraction strength; 3) load sample; 4) remove interference; 5) wash down and collect targeted fraction.
How to select solid phase extraction columns?
SPE columns may be classified, using the practice in HPLC, as reverse phase type, normal phase type, ion exchange type, and mixed mode type. If the sample is aqueous (such as water, urine, plasma, juice), C18 columns (reverse phase type) are most commonly used. If the sample is in organic solvents, a normal phase column (Florisil, Silica gel, Alumina etc.) should be used.
How to select solvents for activation, equilibration and collection in solid phase extraction?
Activation solvent should be able to wet the SPE column sorbent well and compatible with the solvent for equilibration. For columns of reverse phase type, methanol, iso propanol or other polar solvents can be used. For columns of normal phase type, hexane, toluene, or other low polarity solvents can be used. Solvents for equilibration should be compatible with the activation solvent, wet the sorbent well, and of low elution power. If the elution power is high, targeted compound will not be trapped well on the column. For reverse phase column, water or buffer solution is often used for equilibration. For normal phase column, a low polarity solvent is often used for equilibration. Solvents for collection should be compatible with previously used elution solvent, can remove the targeted compounds with small volume and easy to evaporate.
How to decide flow rate in solid phase extraction?
For 3-mL column the flow rate is normally 3-5 mL/min. For 6-mL column, the flow rate is normally 6-10 mL/min. If the column is of ion exchange nature, the flow rate is normally at lower end. For reverse phase column, flow rate ma be set at the higher end.
When do I need to adjust the pH of sample and elution solvent in solid phase extraction?
When the pH can affect extraction of targeted compounds (normally ionizable compounds, such as amines and acids) or the sample matrix that can affect analysis.
Is air bubble a problem in solid phase extraction?
When elution is by gravity, air in the sorbent can change the flow path of solvents and reduce the extraction efficiency. As a standard practice in old type column chromatography, the sorbent must be covered with solvent all the time to avoid air bubble. However in the new SPE practice, the sorbent particle is much smaller and the elution is by a pump or pressure. Air bubble in sorbent can be pushed out quickly. So it is not a problem anymore. Actually the column is often purged with air for solvent exchange.
Do I need to keep the sorbent bed wet all the time during solid phase extraction?
As mentioned in the above question, for SPE using a solvent pump, sorbent bed does need to be wet all the time. For example after a C18 column is activated with methanol, the column can be purged with air and then load an aqueous sample.
When do I need to dry the solid phase extraction column before fraction collection?
When an aqueous sample is loaded or aqueous solvents are used, solvent for collection may have problem to interact with the column properly. Beside the water from column may end up in the collected fraction and affect further evaporation. The SPE column can be dried by purging nitrogen or air. Flow rate of gas cannot be too high or some targeted compounds may be lost. It may be controlled at 300-500 mL/min.
When my recovery result is low in solid phase extraction, what factors should I check?
If the targeted compounds are polar, major cause of low recovery would be the extraction efficiency. Relevant factors are strength of elution solvents, amount of sorbent in the column, volume of sample, and pH of the sample solution. If the targeted compounds are of low polarity (such as PAH, PCB, hydrocarbon), major causes of low recovery would be adsorption by the container and tubing and evaporation loss. Relevant factors are inertness of the wetting components and the column and compatibility of the solvents with the targeted compounds.
Can I regenerate used solid phase extraction columns for repeated use?
A used SPE column will trap components from sample matrix and particles or large molecules (such as humic acids and proteins) may also block the column. A SPE column may be re used in two cases: 1) the sample matrix is simple so the column capacity is not fully used; or 2) the trapped interference from sample can be removed. In the second case, only C18 column is easy to remove trapped sample matrics. For normal phase column and ion exchange column, removing the trapped components is not easy. In general, if you are using a reverse phase column and your sample is clean, you may regenerate the column for repeated use.