Recovery Enhancement Techniques for
Automated Solid Phase Extraction
Susie Petitti, Horizon Technology, Inc.
Key Words
SPE, Solid Phase Extraction, Automation., Recovery Optimization
Introduction
There are two critical steps that impact recoveries in solid phase extractions (SPE) after the extraction step: 1) Removal of
residual water from the final extract using sodium sulfate (Na 2SO4), and 2) Concentration or evaporation procedure to
reduce the extract volume to 1 mL. Liquid-liquid extractions (LLE) involve the use of one solvent (the extraction solvent) to
extract the compounds from aqueous samples. The final extract consists of the extraction solvent with the compounds of
interest and residual water. The drying and concentration steps involve the drying and evaporation of one solvent. On the
other hand, SPE uses a water-soluble solvent to remove residual water from the disk, followed by an extraction solvent to
remove the organic compounds off the dried disk. Therefore, the final extract from SPE is composed of two solvents that
have different boiling points and greater water solubility. The use of a water soluble solvent such as ethyl acetate or
acetone, makes drying with sodium sulfate more difficult. This means that potentially, issues can arise in both the drying
and concentration/evaporation procedures. It is therefore, important to begin at the end of the analytical process (GC,
evaporation, drying, SPE) and determine the impact on recoveries of each step in the process. Three steps are performed
to measure the loss in recoveries from the drying, evaporation and extraction processes.
Instrumentation





Horizon Technology SPE-DEX® Series Automated Extractor System
Evaporation Unit
Drying Set-up
GC Instrument
SPE Disks (packing is method-dependent)
Method Summary and Procedure
Step 1: Concentration/Evaporation Techniques
1.
2.
3.
Run the desired method with an empty bottle on the SPE- DEX Automated Extractor System. No disk is required.
After the extraction has been completed, spike the extract with a known quantity of the analytes of interest. The
spiking solution must be a water-soluble solvent like acetone or methanol, not methylene chloride.
Concentrate the extract using the laboratory’s present technique. The drying step is not necessary since no water
sample was used.
Page 2
Technical Note
4.
5.
Analyze using the appropriate instrumentation.
Report the % recovery for the compounds of interest. This represents the % loss due to the evaporation
technique.
Step 2: Drying and Concentration Technique
1.
2.
3.
4.
5.
6.
7.
Run the desired method using reagent water as a sample. Acidify the water sample to the pH specified by the
method.
After the extraction has been completed, spike the extract (eluate) with known quantity of analytes. The
spiking solution must be a water-soluble solvent like acetone or methanol, not methylene chloride.
Remove residual water from the extract by using sodium sulfate.
Concentrate the extract as done in Step 1.
Analyze using instrumentation.
Report the % recovery for the compounds of interest.
Subtract the values from Step 2 from Step 1 to obtain the loss due to the drying technique. This represents the
% loss due to the drying technique.
Step 3: The Entire SPE Process
1.
2.
3.
4.
5.
6.
7.
8.
Adjust the pH of reagent water as per the method.
Spike the reagent water with known quantity of analytes.
The spiking solution must be a water-soluble solvent like acetone or methanol, not methylene chloride.
Run the desired method.
Remove residual water from the extract by using sodium sulfate as in Step 2.
Concentrate/evaporate the extract using the technique from Step 1.
Analyze using instrumentation.
Report the % recovery for the compounds of interest. This represents the loss due to the entire SPE process.
Results
Tables 1 & 2 summarize the results obtained by two independent labs switching from liquid-liquid extractions to
automated SPE using Horizon Technology Extractors. The three steps in the method summary were performed
and recoveries determined for each.
The data for Step 1 show the loss attributed to the concentration/evaporation technique; Step 2 data show the
loss attributed to the sodium sulfate drying and concentration/evaporation techniques; and Step 3 data show the
loss attributed to the entire extraction process. For the compound hexachlorocyclopentadiene in Table 1, 21 %
was lost from the concentration/evaporation technique alone, 25 % was lost from both the sodium sulfate drying
and concentration techniques, and 28 % was lost from the entire extraction process. This represents a significant
loss attributed to the backend drying and concentration techniques. The best results that can be obtain, with the
drying and concentration techniques being the limiting factors, is the recoveries obtained for Step 2. To improve
recoveries, the drying and concentration/evaporation techniques need to be optimized to reduce loss. This is
especially critical for the more volatile compounds. The following are some suggestions for improvement.
Page 3
Technical Note
Table 1. Recoveries for selected EPA 525.2 Compounds—Lab A
Analytes
Hexachlorocyclopentadiene
Propachlor
Hexachlorobenzene
Simazine
Atrazine
Lindane
Heptachlor
Alachlor
Metribuzin
Aldrin
Metolachlor
Heptachl Epoxide
Butachlor
Dieldrin
Endrin
Bis(2-Ethylhexyl)adipate
Methoxychlor
Benzo(a)pyrene
Step 1
79
81
89
87
97
87
84
93
101
79
103
84
95
84
94
113
103
108
Step 2
75
89
92
86
90
89
87
96
99
78
99
85
96
84
93
92
87
88
Step 3
72
79
85
35
90
88
79
95
22
71
103
82
96
82
93
93
99
103
Table 2. Recoveries for selected EPA 608 compounds—Lab B
Step 1: Concentration Technique
Compound
g-BHC
Spike Conc.
0.08
Result
0.097
%Rec
121
Heptachlor
0.08
0.092
115
Aldrin
Dieldrin
Endrin
4,4’-DDT
0.08
0.16
0.16
0.16
0.09
0.174
0.17
0.18
113
109
106
113
Spk. Conc.
0.08
0.08
0.08
0.16
0.16
0.16
Result
0.032
0.033
0.035
0.065
0.073
0.068
%Rec
40
41
44
41
46
43
Step 2: Drying and Concentration Technique
Compound
g-BHC
Heptachlor
Aldrin
Dieldrin
Endrin
4,4’-DDT
Page 4
Technical Note
Table 2. Recoveries for selected EPA 608 compounds—Lab B (Continued)
Step 3: The Entire SPE Process
Compound
g-BHC
Spk Conc.
0.08
Result
0.024
%Rec
29
Heptachlor
0.08
0.021
26
Aldrin
Dieldrin
Endrin
4,4’-DDT
0.08
0.16
0.16
0.16
0.017
0.045
0.052
0.046
21
28
32
29
Optimizing the Na2SO4 drying procedure:
a.
b.
c.
d.
e.
Place about 10 g of sodium sulfate in a glass column (approximately 11 inches tall with and 1-inch diameter) with a
glass frit bottom.
Ensure that sufficient solvent is poured through the column so that it is thoroughly wetted.
Place a 40 mL collection tube under the column and pour the extracted sample (12-16 mL) through the sodium
sulfate column.
Rinse the original collection vessel three times with 4 mL of methylene chloride and pour the rinses through the
sodium sulfate column.
Ensure that sufficient solvent is run through the column so that all compounds are washed off the sodium sulfate.
An alternative and better option is to use the DryDisk®, designed to replace conventional sodium sulfate as a drying
technique. The DryDisk Separation Membrane uses a physical separation of the water and solvent. The hydrophobic
property of the membrane allows the extracting solvent to pass through it, while retaining any water in the sample
extract, on the membrane’s surface.
Optimizing the concentration technique:
a.
b.
c.
d.
e.
f.
If using heat as part of this process, a maximum of 35 ºC is recommended.
Use a nitrogen blow down technique such as an N-Evap device with about 8 psi of pressure.
Place the cannula approximately ¼ inch above the liquid level in the 40 mL vessel. Adjust the nitrogen flow rate just
to the point before you get bubbles in the sample. You will see a depression on the liquid surface but no bubbles
are generated.
Re-adjust the cannula level as described above as the liquid level evaporates.
When the liquid level drops to about 1 inch, transfer the sample to a 10 mL KD tube. Rinse the sample tube three
times with 1 mL of methylene chloride.
Repeat the blow down procedure for the KD tube until the sample volume reaches the desired volume.
Alternatively, an XcelVap® water bath, nitrogen blow down system or a DryVap® closed in-line drying and evaporation
system can be used for the drying/evaporation/concentration step.
Technical Note
Page 5
Results
The data in Tables 1 & 2 emphasize the importance of optimizing the drying and concentration techniques to minimize loss in
recoveries for SPE. The presence of two solvents, with different boiling points, requires special attention because of the longer
time required to concentrate and evaporate the two solvents to the desired volume. To successfully implement a method using
SPE, the drying and concentration steps must also be optimized. This note describes how to do evaluate the contribution of
these two steps to loss in recoveries. The SPE-DEX system is used in this example, but the SmartPrep cartridge extractor
methodology can also be evaluated with a similar procedure.
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