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An 348:extraction of anthelmintic drugs from a veterinary formulation using accelerated solvent extraction (ase)
Application Note 348
Extraction of Anthelmintic Drugs from a
Veterinary Formulation Using Accelerated
Solvent Extraction (ASE®)
Isolation of the active drug from some veterinary
Dionex ASE 200 Accelerated Extractor with Solvent
formulations can be difficult because the matrix is often
complex. These difficulties were traditionally solved by
11-mL stainless steel extraction cells (P/N 055422)
extracting with a wrist-shaker method, sonication, or
Soxhlet. Although these techniques produce adequate
Dionex collection vials, 40 mL (P/N 048783)
results, they are very labor intensive and use large
Analytical balance (accurate nearest 0.0001 g or better)
amounts of solvent. The time required for these methodscan cause bottlenecks in the sample preparation area
Sand (Ottawa Standard, Fisher Scientific, Cat. No. S23-3)
and solvent use can cost the laboratory thousands of
dollars in unwarranted expenses per year.
PTFE syringe Filter, 0.2 µm (Fisher Scientific)
Accelerated Solvent Extraction (ASE) is an auto-
mated extraction technique that uses the same solventsas traditional extraction methods but in significantly
smaller amounts and with minimal analyst exposure.
ASE achieves equivalent or better results in a fraction ofthe time by using increased temperature and pressure toenhance the kinetics of the extraction process. Thisentire process is fully automated and allows the unat-
tended extraction of up to 24 samples.
This application note describes the extraction of the
two active species of ivermectin (H2B1a and H2B1b), an
anthelmintic drug, from a veterinary formulation contain-
ing dried meat products. This formulation is used to treat
household cats and dogs for heartworm disease. Figure 1
shows the chemical structure of ivermcetin.
R = CH2CH3
Figure 1. Chemical structure of ivermectin.
Application Note 348
Table 1. ASE Recovery of Ivermectin from Spiked
(All solvents are pesticide-grade or equivalent and
* Shows a range of 50–150% of the target concentration (0.9 µg/mL)
Table 1 shows the results of extracting placebo
preparations spiked with a varying range of ivermectinconcentrations (50–150%). The average recovery for all
The tablet should be finely ground using a food
Table 2 shows the precision of the ASE method. Six
processor or coffee grinder to a powder that can pass
preparations from one lot of HEARTGARD® Chewables
through a Tyler 10 sieve. Accurately weigh out approxi-
for Cats were extracted and analyzed. The average
mately 0.5–1.5 g of the powder and blend with 1.5 g of
percent recovery for these six preparations was 102.3%
Hydromatrix using a mortar and pestle. Transfer the
with an RSD of 0.90%. These recovery and precision
mixture to an 11-mL stainless steel extraction cell
values are as good or better than observed with tradi-
containing a cellulose filter. Top off any dead space in
the cell with Ottawa sand. Prepare any other tabletsamples and load them into extraction cells.
Table 2. ASE Method Precision Summary: Extraction
of Ivermectin from HEARTGARD Chewables for Cats
Place the cells onto the ASE 200. Label the appro-
priate number of collection vials and place these into the
carousel. Set up the method suggested above and begin
the extraction. When the extraction is complete, theextract can then be diluted to the desired volume and
passed through a 5-mL alumina cartridge. Finally, filter
the extract into an HPLC vial through a 0.2-µm PFTEfilter and analyze using HPLC.1
RESULTS AND DISCUSSION
Sample preparation is critical to good recoveries.
Grind the samples to a uniform particle size to ensure
proper permeation of the solvent into the matrix. Thesample extracts may be somewhat cloudy due to the
extractions of fats and other coextractables, so it is impor-tant to pass the extracts through a short column of alumina.
Extraction of Anthelmintic Drugs from Veterinary
Formulation Using Accelerated Solvent Extraction (ASE)
These results confirm that ASE is comparable to
We would like to acknowledge the work of Andreas
traditional extraction methods for the difficult extrac-
Abend and his colleagues at Merck & Co. Inc.
tions of active drugs from veterinary formulations.
Traditional extraction methods usually take from one to
several hours for each sample and require large amounts
of solvent. With ASE, the extraction time is cut to
Wuelfing, W. P. “Development and Validation of an
approximately 15 min per sample and uses only 25–30
Automated Extraction Method (Accelerated Solvent
mL of solvent. In addition, the ASE 200 can extract up
Extraction®) and a Reverse-Phase HPLC Analysis
to 24 samples, sequentially, without user intervention.
Method for Assay of Ivermectin in a Meat-Based
Chewable Formulation.” Pharm and Bio Med
Analysis 2003, 31,
HEARTGARD is a registered trademark of Merial.
ASE is a registered trademarks of Dionex Corporation.
Dionex U.S. Regional Offices
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France 01 39 30 01 10 Germany 06126-991-0 Italy (06) 66 51 50 52 Japan (06) 6885-1213 Application Note 348
The Netherlands (0161) 43 43 03 Switzerland (062) 205 99 66 United Kingdom (01276) 691722
* Designed, developed, and manufactured under an NSAI registered ISO 9001 Quality System.
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