Extraction of clenbuterol from calf urine using a molecularly imprinted polymer followed by quantitation by high-performance liquid chromatography with UV detection.
A method for the extraction of clenbuterol from calf urine samples using a molecularly imprinted polymer (MIP) has been developed. The aim was that the final extracts from the MIP should allow quantitation of clenbuterol down to 0.5 ng/mL urine using HPLC with UV detection. The MIP was produced using brombuterol as a template and the selectivity of the MIP, for clenbuterol, was tested against a non-imprinted polymer (produced without template) and was found to be high. After loading of 5 mL diluted centrifuged urine, selective binding was established in acetonitrile-acetic acid (98:2). For further elution of interferences, 0.5 M ammonium acetate buffer pH 5 and 70% acetonitrile in water was used. Clenbuterol was eluted using 1% trifluoroacetic acid in methanol, which was evaporated and reconstituted in buffer. Results from the HPLC analyses showed that the extraction of clenbuterol using MIP is linear in the range 0.5-100 ng/mL with good precision (4.3% for 0.6 ng/mL and 2.1% for 6.0 ng/mL) and accuracy (96.7% for 0.6 ng/mL and 96.7% for 6.0 ng/mL). The recoveries were 75%. The results show that the method offers a selectivity and sensitivity that make the quantitation of 0.5 ng clenbuterol/mL urine by HPLC-UV possible and a competitive alternative to state-of-the-art routine analytical methods.
Blomgren
Anders
A
anders.blomgren@mip.ideon.se
Berggren
Christine
C
Holmberg
Anna
A
Larsson
Fredrik
F
Sellergren
Börje
B
Ensing
Kees
K
D000316:Adrenergic alpha-Agonists / Q000652:urine*; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002976:Clenbuterol / Q000652:urine*; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet / Q000379:methods*
D000316:Adrenergic alpha-Agonists; D002976:Clenbuterol