Simultaneous Determination of Podophyllotoxin, Quercetin and Kaempferol in Podophyllin by Liquid Chromatography Tandem Mass Spectrometry

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Journal of Food and Drug Analysis, Vol. 16, No. 6, 2008, Pages 29-40

Simultaneous Determination of Podophyllotoxin, Quercetin and Kaempferol in Podophyllin by Liquid Chromatography

Tandem Mass Spectrometry

MEI-CHIH LIN1,2, JER-HUEI LIN2, SHU-KONG CHEN2, YU-WEN CHENG1* AND HUEI-WEN CHENG1*

1. College of Pharmacy, Taipei Medical University, Taipei, Taiwan (R.O.C.)

2. Bureau of Food and Drug Analysis, Department of Health, Executive Yuan, Taipei, Taiwan (R.O.C.)

(Received: March 26, 2008; Accepted: September 5, 2008)

ABSTRACT

A quantitative high performance liquid chromatography-tandem mass spectrometry (LC/MS/MS) was developed to analyze

the constituents of podophyllin. Chromatographic separation was performed on a Cosmosil 5C18-MS (25 cm 4.6 mm I.D., 5 m)

reverse phase column using a gradient of mobile phase (0.25% formic acid-methanol). The column effluent was split 2: 3 into the

photodiode detector and tandem mass spectrometer. Podophyllotoxin, quercetin and kaempferol in podophyllin were identified by

daughter ion scan mode and then determined their contents by multiple-reaction monitoring (MRM) mode. The limits of detection

and quantitation for podophyllotoxin, quercetin and kaempferol were 2.40, 8.01; 2.94, 9.87 and 3.10, 10.2 ng/mL, respectively. The

relative standard deviations of intraday and interday analyses for podophyllotoxin, quercetin and kaempferol ranged from 0.52-

6.01% and 2.48-9.88%, respectively. The mean recoveries for podophyllotoxin, quercetin and kaempferol were 101.4%, 98.3% and

98.7%, respectively. The developed LC/MS/MS method was suitable for the simultaneous determinations of podophyllotoxin,

quercetin and kaempferol in podophyllin.

Key words: LC/MS/MS, podophyllin, podophyllotoxin, quercetin, kaempferol

INTRODUCTION

Podophyllin is an alcoholic plant extract from the dried rhizomes and roots of Podophyllum emodi (Indian Podophyllum) and P. peltatum (Mayapple or Mandrake). It possessed various biological activities, and had been used to treat constipation, hepatic disorder, and rheumatic arthritis. In 1942, podophyllin was suggested as a treatment of condylomata acuminata, one type of venereal warts, by Kaplan(1). Though severe systematic toxici-ties from ingestion or tropical application of podophyllin had been reported, the side effects were usually reversible and fatal(2). Hence, so far it was used as a tropical treatment for genital warts and required to apply a thin layer to warts to minimize its side effects(3).

The chemical constituents of podophyllin had been studied since the nineteenth century. Podophyllotoxin (Figure 1), the major constituent of podophyllin, was rstly separated and identied in 1880, and subsequently, a...