Development and Validation Method for Identification of PDE-5 inhibitors In Seized Materials
Identification of PDE-5 inhibitors in Seized Materials using TLC and HPLC methods was developed and validated. TLC method, the stationary phase was siliga gel 60 F 254, the mobile phase comprised two systems. System 1 was the mixture of ethyl acetate, acetonitrile and ammonia solution and system 2 was the mixture of ethyl acetate, ethanol and ammonia solution. TLC of isolated sample was firstly detected by UV radiation at 254 nm, spectrum scanning and then confirmed by color reaction with ethanolic-sulfuric acid and acidified iodoplatinate reagent. In system 1, Rf were found to be 0.49, 0.71, 0.50, 0.57, and 0.56 for sildenafil, tadalafil, vardenafil, aminotadalafil, methisosildenafil, respectively. In system 2, the Rf were found to be 0.31, 0.78, 0.45, 0.72, and 0.43 for sildenafil, tadalafil, vardenafil, aminotadalafil, methisosildenafil, respectively. A purple color after spraying with ethanolic-sulfuric acid reagent indicated the presence of tadalafil and aminotadalafil. When acidified iodoplatinate solution was used, a brown to purple color developed indicated the presence of sildenafil, tadalafil, vardenafil and aminotadalafil. With TLC techniques, limit of detections of sildenafil, tadalafil and vardenafil were 100, 1000 and 250 μg/ml respectively. HPLC method, the separation was performed on a C18 column by isocratic elution with a mobile phase of 0.01 M ammonium acetate buffer and methanol and PDA detection at 210-350 nm. All sildenafil, tadalafil, vardenafil, aminotadalafil and methisosildenafil peaks were completely resolved as shown by resolution values greater than 1.8, with no interference from matrix components. With HPLC techniques, limit of detections of sildenafil, tadalafil, vardenafil, aminotadalafil and methisosildenafil were 3.8, 1.0, 2.5, 0.8 and 2.3μg/ml respectively. The validated method was applied to the identification of illicit drugs containing PDE-5 inhibitors in seized materials in fiscal year 2015. It was found that among 230 samples tested, 213 samples contained sildenafil, 11 samples contained a mixture of sildenafil, vardenafil, aminotadalafil, tadalafil and one sample contained a mixture of sildenafil and vardenafil.