Abstract
A simple isolation method and rapid, specific high performance liquid chromatographic method was developed and validated for filixic acid PBP in Dryopteris filix-mas extract. HPLC analysis was performed on C18 column using a 90:10 (v/v) mixtures of acetonitrile and methanol as isocratic mobile phase at a flow rate 1.0 ml/min. UV detection was at 254 nm for filixic acid PBP. Filixic acid shown retention time at 4.02 min. Method was validated for accuracy, precision, linearity, specificity and sensitivity in accordance with International Conference on Harmonisation guidelines. Validation studies revealed that the method is specific, accurate, precise, reliable and reproducible. Good linear correlation coefficient (r 2 > 0.993) was obtained for calibration plots in the range tested. The limit of detection was 2.25 µg/ml and limit of quantification was 7.53 µg/ml for filixic acid PBP. Intra and inter-day relative standard deviation of precision was less than 1.50 %. Recovery was between 94.57 and 101.05 % filixic acid. The method can be successfully used for quantitative analysis of filixic acid PBP in D. filix-mas for day-to-day studies. Keyword: Dryopteris filix-mas; male fern; filixic acid PBP; HPLC; isolation.
Highlights
Male fern consist of dried rhizomes and fond bases of Dryopteris filix-mas and other species of Dryopteris, belonging to family Polypodiaceae[1]
Thin layer chromatography (TLC) and High Performance Thin Layer Chromatography (HPTLC) method for qualitative analysis of male fern is available in literature[9]
3.2 Validation of chromatographic method In HPLC study filixic acid PBP was resolved with retention time (Rt) at 4.03
Summary
Male fern consist of dried rhizomes and fond bases of Dryopteris filix-mas and other species of Dryopteris, belonging to family Polypodiaceae[1]. Indian male fern yield 8-10% ether extractive value of which 30 % is filicin. It is the mixture of six homologous compound filixic acid, i.e. filixic acid BBB, ABB and PBP Filixic acid PBP is pale yellow plates with melting point in the range 184-186°C5-7. Pamttila and Sundman have done isolation work for filixic acids[8]. Thin layer chromatography (TLC) and High Performance Thin Layer Chromatography (HPTLC) method for qualitative analysis of male fern is available in literature[9]. In present study attempt was made to isolate the filixic acid PBP from the resin. The specific, reliable and reproducible HPLC method was developed and validated according to International Conference on Harmonization (ICH) guidelines[10]
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