Q&A for SPE Column

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Solid phase extraction (SPE): also known as liquid-solid extraction. It refers to the process in which the analyte in the liquid sample is retained on the adsorbent by adsorption and then eluted by a certain solvent.

SPE method is a separation technology which combines selectively retention and selectively elution, mainly used in separation, purification and concentration of samples. Compared with conventional liquid-liquid method, SPE improves the recovery rate of analytes and effectively separates analytes and interfering components to simplify the pretreatment process, indicating its advantages of time-saving, convenient and low-cost.

According to some frequently asked problems during the use of SPE column, they can be summarized in several aspects including low recovery rate, poor reproducibility and purification effect. So how to address these problems?

Low Recovery Rate

Possible reasons and solution

1. Insufficient retention of object substances on packing materials

Judgement method: Blank spiked solvent, recycle effluent and flushing liquid, if the content of object substances in effluent or flushing liquid exceeds 5%, that means insufficient retention.  

Possible ReasonsSolution
Inappropriate activation condition of column or with activationActivate SPE column according to the type of stationary phase accurately
Column is dried up in the operating processReactivate or equilibrate the column to ensure the column is not dried before finishing flushing(except SPE columns based on polymers)
Inappropriate selection of SPE columnChoose the column with strong retention to object substances
Elution strength of sample solvent or flushing liquid is too strongReduce the elution strength of sample solvent and flushing liquid
Overloading and permeation caused by large solution volume of high concentrationReduce the sample loading or concentration, increase the packing materials

2. Incomplete elution of object substance

Judgement method: blank spiked solvent, recycle effluent and flushing liquid, if object compounds don’t appear in effluent and flushing liquid of sample or they exist in eluent of a small amount volume, that means insufficient retention.

Possible ReasonsSolution
Stationary phase has too strong selectivity to object compoundsChoose column which has weak retention to object compounds; Adjust the pH of eluent; Eliminate the selectivity of stationary phase to object components
Too weak elution abilityChoose stronger elution solvent
Incomplete elution led by eluent solventIncrease the elution solvent volume
Incomplete elution led by too fast flow rateLet the eluent permeates column, vacuumize the column or add pressure, then set the flow rate at 1-2ml/min

3. Other reasons

Judgement method: blank spiked solvent, recycle effluent, eluent, flushing liquid, if object compounds don’t appear in above liquids or they exist in eluent of a large amount volume with insufficient recovery rate, that means insufficient retention.

Possible ReasonsSolution
Extration efficiency of SPE column is insufficientChoose the appropriate extraction solvent
Unstable object substances dissemble or decomposeChange extraction way or transferring way; Decrease heating temperature
Too many steps of pretreatment or loss of object substanceSimplify pretreatment process
Object substances combine with impuritiesRemove impurities in an appropriate method
Interfering impurities impact too muchChange pretreatment method or compare with spiked substances
Too many impurities or exceed the highest retentionChange pretreatment method;
Reduce sample loading or increase the packing materials of column

Poor Reproducibity

Possible ReasonsSolution
Complicate operation steps and operative errorOptimize pretreatment method or make SOP method
Complex sample matrices, poor effect of cleaning impuritiesOptimize or change the whole pretreatment method;
Add internal standard or contrast with spiked data of matrices
Dried columnReactivate or equilibrate the column to ensure the column is not dried before finishing flushing (except SPE columns based on polymers)  
Column blockage or too fast flow rateThe former changes the pretreatment method, the latter uses flow control valves to change flow rate.

Poor Purification Effect

Solution

  1. Change the pretreatment method
  2. Change the strength of flushing and eluent
  3. Choose a suitable SPE column
  4. Operate SPE experiment correctly

Inappropriate Flow Fate

Reason of Slow Flow RateSolution
Too small particle sizeChoose SPE column with large particle size
After pretreatment of sample, suspension insoluble substances still exist, which leads to blockage of fritFreeze and centrifuge before sample loading to make sure the supernatant is clear.
Sample viscosity is too highDilute the sample
The SPE resistance is excessive, liquids can not drop naturallyUse manual SPE device; Control valve by regulating flow; Adjust flow rate by giving positive or reducing negative pressure

Reason of Fast Flow RateSolution
Too large particle sizeChoose SPE column with small particle size
Flow rate is too fast when gravity worksUse manual SPE device to adjust controlling valve; Control valve by regulating flow 
Flow rate is too fast when reducing negative pressureRegulating flow control valves and valves; Control vacuum gauge pressure to be constant within a certain range

Welchrom SPE Column

If you have any problem or require further information, please contact info@welchmat.com.

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