Quantification of the major metabolites of bromhexine in human plasma using RRLC-MS/MS and its application to pharmacokinetics.
(E)-4-hydroxydemethylbromhexine (E-4-HDMB) and (E)-3-hydroxydemethylbromhexine (E-3-HDMB) were found as major metabolites, while (Z)-4-hydroxydemethylbromhexine and (Z)-3-hydroxydemethylbromhexine as minor metabolites of bromhexine in human plasma. These compounds were identified in comparison with synthetic authentic samples. A sensitive and selective rapid resolution liquid chromatography tandem mass spectrometry (RRLC-MS/MS) method was developed to quantify the concentration of bromhexine and its two major metabolites (E-4-HDMB and E-3-HDMB) in human plasma. Following solid phase extraction, the analytes were separated on a Zorbax 1.8microm particle size reversed-phase C(18) column, using a gradient elution program with solvents consisting of 0.1% formic acid in acetonitrile and 0.1% formic acid in 5mM ammonium acetate at a flow rate of 0.7mL/min. Detection was carried out with an Agilent 6460 triple-quadrupole mass spectrometer operated with an electrospray ionization source mode operated in the positive ion mode. The recovery of bromhexine, E-4-HDMB, E-3-HDMB, and internal standard (IS) was 63.1-70.9%, 60.5-68.4%, 57.0-63.5%, and 87.8%, respectively. The matrix factors of bromhexine, E-4-HDMB, E-3-HDMB, and IS were 89.9-96.7%, 89.6-94.8%, 90.4-91.4%, and 103%, respectively. After an oral administration of 8.0mg bromhexine to five healthy male subjects, AUC(0-24h) values of bromhexine, E-4-HDMB, and E-3-HDMB were found to be 93.5+/-31.9, 34.0+/-14.5, and 15.8+/-6.89ngh/mL, respectively; while C(max) values were 24.6+/-5.16, 3.11+/-1.13, and 5.36+/-2.55ng/mL, respectively. Plasma concentration of bromhexine, E-4-HDMB, and E-3-HDMB declined with t(1/2) which gave 3.6+/-0.5, 8.4+/-2.7, and 6.4+/-2.5h, respectively.
Liu
Jia
J
Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, PR China.
Chen
Xiaoyan
X
Hu
Youhong
Y
Cheng
Gang
G
Zhong
Dafang
D
D000284:Administration, Oral; D001711:Biotransformation; D001964:Bromhexine / Q000008:administration & dosage / Q000097:blood / Q000493:pharmacokinetics*; D002853:Chromatography, Liquid / Q000592:standards; D003640:Dealkylation; D004355:Drug Stability; D005100:Expectorants / Q000008:administration & dosage / Q000493:pharmacokinetics*; D006207:Half-Life; D006801:Humans; D006900:Hydroxylation; D008297:Male; D012015:Reference Standards; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000592:standards; D053719:Tandem Mass Spectrometry / Q000592:standards; D055815:Young Adult
D005100:Expectorants; D001964:Bromhexine