HPLC-DAD and HPLC-ESI-MS separation, determination and identification of the spin-labeled diastereoisomers of podophyllotoxin.
Spin-labeled nitroxide derivatives of podophyllotoxin had better antitumor activity and less toxicity than that of the parent compounds. However, the 2-H configurations of these spin-labeled derivatives cannot be determined by nuclear magnetic resonance (NMR) methods. In the present paper, a high-performance liquid chromatography-diode array detection (HPLC-DAD) and a high-performance liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI/MS/MS) method were developed and validated for the separation, identification of four pairs of diastereoisomers of spin-labeled derivatives of podophyllotoxin at C-2 position. In the HPLC-ESI/MS spectra, each pair of diastereoisomers of the spin-labeled derivatives in the mixture was directly confirmed and identified by [M+H](+) ions and ion ratios of relative abundance of [M-ROH+H](+) (ion 397) to [M+H](+). When the [M-ROH+H](+) ions (at m/z 397) were selected as the precursor ions to perform the MS/MS product ion scan. The product ions at m/z 313, 282, and 229 were the common diagnostic ions. The ion ratios of relative abundance of the [M-ROH+H](+) (ion 397) to [M+H](+), [A+H](+) (ion 313) to [M-ROH+H](+), [A+H-OCH(3)](+) (ion 282) to [M-ROH+H](+) and [M-ROH-ArH+H](+) (ion 229) to [M-ROH+H](+) of each pair of diastereoisomers of the derivatives specifically exhibited a stereochemical effect. Thus, by using identical chromatographic conditions, the combination of DAD and MS/MS data permitted the separation and identification of the four pairs of diastereoisomers of spin-labeled derivatives of podophyllotoxin at C-2 in the mixture.
Zhao
Lei
L
State Key Laboratory of Applied Organic Chemistry, Lanzhou Universitry, Lanzhou, China.
Liu
Zhen-Ling
ZL
Fan
Peng-Cheng
PC
Zhang
Zhi-Wei
ZW
Liu
Xiong
X
Zhan
Yun-Jing
YJ
Tian
Xuan
X
D000972:Antineoplastic Agents, Phytogenic / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D003497:Cyclic N-Oxides / Q000737:chemistry; D015394:Molecular Structure; D011034:Podophyllotoxin / Q000031:analogs & derivatives / Q000032:analysis* / Q000138:chemical synthesis / Q000737:chemistry / Q000302:isolation & purification*; D021241:Spectrometry, Mass, Electrospray Ionization; D013113:Spin Labels; D013237:Stereoisomerism; D053719:Tandem Mass Spectrometry
D000972:Antineoplastic Agents, Phytogenic; D003497:Cyclic N-Oxides; D013113:Spin Labels; C415032:picropodophyllin; D011034:Podophyllotoxin