Narrowbore high-performance liquid chromatography for the simultaneous determination of sildenafil and its metabolite UK-103,320 in human plasma using column switching.
A fully automated narrowbore high-performance liquid chromatography method with column switching was developed for the simultaneous determination of sildenafil and its active metabolite UK-103,320 in human plasma samples without pre-purification. Diluted plasma sample (100 microl) was directly introduced onto a Capcell Pak MF Ph-1 column (20x4 mm I.D.) where primary separation occurred to remove proteins and concentrate target substances using 15% acetonitrile in 20 mM phosphate solution (pH 7). The drug molecules eluted from the MF Ph-1 column were focused in an intermediate column (35x2 mm I.D.) by a valve switching step. The substances enriched in the intermediate column were eluted and separated on a phenyl-hexyl column (100x2 mm I.D.) using 36% acetonitrile in 10 mM phosphate solution (pH 4.5) when the valve status was switched back. The method showed excellent sensitivity (detection limit of 10 ng/ml), good precision (RSD < or = 2.3%) and accuracy (bias: +/-2.0%) and speed (total analysis time 17 min). The response was linear (r2 > or = 0.999) over the concentration range 10-1000 ng/ml.
Bioanalysis Laboratory, College of Pharmacy, Wonkwang University, Shinyongdong, Iksan, South Korea.
C K
CK
Jeong
H Y
HY
Lee
M S
MS
Jang
W B
WB
Kim
H S
HS
Lee
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D008297:Male; D010879:Piperazines / Q000097:blood* / Q000493:pharmacokinetics; D011687:Purines; D011744:Pyrimidinones / Q000097:blood* / Q000493:pharmacokinetics; D012016:Reference Values; D015203:Reproducibility of Results; D000068677:Sildenafil Citrate; D013450:Sulfones
D010879:Piperazines; D011687:Purines; D011744:Pyrimidinones; D013450:Sulfones; C109995:UK 103320; D000068677:Sildenafil Citrate