No abstract

Fenugreek seeds are known as a source of various compounds, the most common of which are steroidal saponins. However, despite the growing interest in this plant material as a healing agent, spice and dietary supplement ingredient, the composition of Polish fenugreek seeds remains unknown. Therefore, the steroidal saponin complex in the seeds of T. foenum-graecum cultivated in Poland was qualitatively and quantitatively analyzed by the HPLC-ELSDESI-MS method. Two C-18 columns connected in series were used for the first time in analysis of fenugreek saponins and ELS detector parameters were optimized. A total of 26 furostanol saponins were revealed, of which 24 were tentatively identified. The HPLC-ELSD method developed for quantitative analysis was preliminarily validated and the determined amount of steroidal saponins in Polish fenugreek seeds was 0.14 %.

Context Trigonella foenum-graecum L. (Fabaceae) has many therapeutic properties and anticancer potential. Objective The cytotoxic activities of standardized extracts and a fraction from fenugreek seeds and their compounds (sapogenins, flavone C-glycosides, alkaloid trigonelline) against human cancer SKOV-3, HeLa and MOLT-4 cells were evaluated. Materials and methods Fenugreek seeds were extracted with 70% methanol (A) or water (B). Furthermore, the seeds were purified with petroleum ether and chloroform and next extracted with methanol to obtain fraction (C). The quantitative analysis of saponins and flavonoids in the extracts was done with HPLC methods. The extracts (5–120 µg/mL) and compounds (1–50 µg/mL) were tested on the cells by MTT assay and RTCA system. The effect of a fraction on ROS production, mitochondrial membrane potential and caspase-3/7 activity in HeLa and SKOV-3 cells was also evaluated by flow cytometry. Results The strongest cytotoxic activity on cancer cells showed the fraction C (IC 50 was 3.91 ± 0.03 for HeLa, 3.97 ± 0.07 for SKOV-3, and 7.75 ± 0.37 for MOLT-4) with the highest content of steroidal saponins (163.18 ± 11.03 μg/mg) and flavone C-glycosides (820.18 ± 0.05 μg/mg). The fraction significantly increased ROS production (up to four times higher than in keratinocytes as control) and caspases activity in the cells. The examined flavonoids did not exhibit the cytotoxic activity in contrast to yamogenin, tigogenin, and diosgenin. Conclusions The obtained results complement the data on the cytotoxic activity of Foenugraeci Semen and synergistic effect of flavonoids and saponins complex contained in the plant.

A new HPTLC-densitometric method for diosgenin determination in fenugreek seeds was established after optimization of the conditions for efficient saponin extraction and acid hydrolysis. Several procedures were tested, the best of which was a three-step Soxhlet extraction, followed by hydrolysis of the obtained methanolic extract with 2 mol L-1 H2SO4. Best diosgenin separation from other hydrolysis products was obtained on HPTLC Si60F254 plates u sing a mixture of n-heptane/ethyl acetate (7:3, V/V) and modified anisaldehyde as a spraying reagent. The method was preliminarily validated and the determined amounts of diosgenin in fenugreek seeds of Polish and African origin were found to be similar and ranged from 0.12-0.18 %.

The on-line heart-cutting two-dimensional liquid chromatography method with the use of diode array detector and mass spectrometer (LC-LC-DAD-ESI-MS) was established and validated for quantitation of C-glycosylflavones in fenugreek seeds (Foenugraeci...