title: Determination of natamycin in rabbit cornea by high-performance liquid chromatography-tandem mass spectrometry with protective soaking extraction technology.
abstract: A new selective and sensitive high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed for the quantification of natamycin (NAT) in rabbit corneas with amphotericin B as the internal standard (IS). The cornea samples were processed by a simple and protective methanol soaking extraction technology. The NAT could be extracted completely from rabbit cornea after 24h of soaking with methanol under a mild condition. Chromatographic separation was performed on a C18 column (2.1mm×50mm, 3.5μm) using mobile phase with ammonium acetate buffer (pH 4.5; 4.0mM):acetonitrile (40:60, v/v) at a flow rate of 0.25ml/min. Quantification was performed using the transitions 666.2→503.2 m/z for NAT and 924.5→906.6 m/z for IS by positive ion electrospray ionization in multiple reaction monitoring mode. The assay was validated over a concentration range of 8.64ng/ml to 843ng/ml with lower limit of detection of 4.32ng/ml. The method was validated with respect to linearity, accuracy, precision, recovery, stability and extracting efficiency. The extraction recovery of NAT from cornea samples was approximately 100% with the new methanol soaking extraction procedure. The method has been successfully applied to the ocular pharmacokinetic studies of NAT eye drops in the cornea of Japanese white rabbit.
authors.affiliation: Henan Eye Institute, Henan Eye Hospital, No. 7 Weiwu Road, Zhengzhou 450003, China.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Tianyang
authors.affiliation: School of Pharmaceutical Science, Zhengzhou University, No. 100 Science Road, Zhengzhou 450001, China.
authors.forename: Zhu
authors.initials: Z
authors.lastname: Ling
authors.affiliation: Henan Eye Institute, Henan Eye Hospital, No. 7 Weiwu Road, Zhengzhou 450003, China.
authors.forename: Xia
authors.initials: X
authors.lastname: Huiyun
authors.affiliation: Henan Eye Institute, Henan Eye Hospital, No. 7 Weiwu Road, Zhengzhou 450003, China.
authors.forename: He
authors.initials: H
authors.lastname: Jijun
authors.affiliation: Henan Eye Institute, Henan Eye Hospital, No. 7 Weiwu Road, Zhengzhou 450003, China. Electronic address: zhangjj66@126.com.
authors.forename: Zhang
authors.initials: Z
authors.lastname: Junjie
mesh_terms: D060433:Administration, Ophthalmic; D000818:Animals; D005591:Chemical Fractionation; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003315:Cornea / Q000737:chemistry* / Q000378:metabolism; D004355:Drug Stability; D057230:Limit of Detection; D008297:Male; D010866:Natamycin / Q000032:analysis* / Q000493:pharmacokinetics; D011817:Rabbits; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D010866:Natamycin