Analysis of secondary metabolites from eschscholtzia californica by high‐performance liquid chromatography

Abstract

A rapid and precise analytical HPLC method has been developed for screening the major benzophenanthridine alkaloids produced by cell cultures of Eschscholtzia califomica, namely, sanguinarine, chelirubine, macarpine, chelerythrine and chelilutine. Separation was achieved on a C18, reversed-phase column with gradient elution using acetonitrile and 50 mM phosphoric acid. Detection was performed by both fluorescence (lambda(ex) 330 nm, lambda(em) 570 nm) and photodiode array, leading to good selectivity and precision in determining peak purity. A simple and quick sample preparation protocol was elaborated involving a methanolic extraction for the measurement of intracellular concentrations of the alkaloids and a solid phase extraction for their quantification in culture medium. Owing to the non-availability of commercially standards, a method for the purification of chelirubine, macar pine and chelilutine by semi-preparative HPLC was developed. Coupled together, the isolation method and the analytical method were highly reliable for screening the alkaloids of interest produced by E. califomica.

HPLC in Natural Product Analysis: The Detection Issue

“…It combines sample cleanup with immunoaffinity columns followed by HPLC-FD, and it enabled the detection of aflatoxin content ranging from 7 to 20 μg/kg in the plant material. HPLC-FD also has been used for the screening of the major benzophenanthridine alkaloids produced by cell cultures of Eschscholtzia californica [37]. Thanks to the selectivity of the method (excitation 330 nm, emission 570 nm), the sample preparation was notably simplified compared with previous methods.…”

Metabolism of sanguinarine in human and in rat: Characterization of oxidative metabolites produced by human CYP1A1 and CYP1A2 and rat liver microsomes using liquid chromatography–tandem mass spectrometry

“…Once the enzymes known, prediction may be made concerning drug-drug interactions potentially resulting in clinical alteration of pharmacokinetics. Very sensitive HPLC methods coupled to fluorescence detection [13,14] or electrospray ionization-mass spectrometry (ESI-MS) [7] are required for quantifying SA in culture medium or in rat plasma or urine. Thus, the biotransformation of SA and chelerythrine (CHEL, Fig.…”

A high-rate perfusion bioreactor for plant cells

“…Extracts from cells, medium, and resins were analyzed for alkaloid content using the following chromatographic method previously described (Klvana et al, 2004(Klvana et al, , 2006. The HPLC apparatus used consisted of a model 126 Beckman Coulter pump module and a model 508 Beckman Coulter auto-sampler, coupled with a model 821-FP Jasco fluorescence detector and a model 168 Beckman Coulter photo diode array absorbance detector.…”

“…Peak purity was verified by UV spectra symmetry. Calibration curves were obtained with standards that were purchased: sanguinarine and chelerythrine (Sigma) and standards obtained from a semi-preparative method (Klvana et al, 2006).…”