An HPLC Method for the Quantification of Colchicine and Colchicine Derivatives in Gloriosa superba seeds

Abstract

An 80% ethanolic extract of Gloriosa superba L. seeds (glory lily, Colchicaceae), as well as a colchicine-poor/colchicoside-rich extract, were shown to exhibit antitumor activity in a murine model for pancreatic cancer. Phytochemical investigations of the 80% ethanolic extract led to the identification of colchicine, 3- O-demethylcolchicine, and colchicoside. The objective of this work was to develop and validate a high performance liquid chromatographic analytical method according to the ICH guidelines for the quantification of these constituents. The calibration model appeared to be linear, ranging from 2.1 μg/mL to 41.9 μg/mL. The method was shown to be precise with respect to time (RSD% of 3.1% for colchicine, 2.9% for 3- O-demethylcolchicine, and 4.7% for colchicoside, 3 days, n = 6) and with respect to the concentration (RSD% of 2.9% for colchicine, 3.0% for 3- O-demethylcolchicine and 4.1% for colchicoside, 3 levels, n = 6). The recovery of colchicine resulted in a mean recovery of 100.02% with a RSD% of 2.1%. The correction factors for colchicoside and 3- O-demethylcolchicine were determined as 1.94 and 1.20, respectively. The total amount of colchicine and colchicine derivatives found in the crude extract of G. superba was 4.6% (m/m) expressed as colchicine and the overall mean of colchicine found in the crude extract was 2.8% (m/m). By using the correction factors, the other constituents of the crude extract could also be quantified, and it was found to contain 1.5% (m/m) colchicoside and 1.3% (m/m) 3- O-demethylcolchicine.