4.3. Saponin and Inulin Contents

The method determines the sum of saponins in E1, E2, and E3 based on the method in the dry water extract from the root of the balloon flower (PG) using the precipitation-weight method after previous extraction with methanol [54,55]. Briefly, a dry extract of about 2.0 g was placed in a thimble in a Soxhlet apparatus, diluted with 50 mL of methanol and left for 15 h. Next, 50 mL of methanol was added and the mixture was extracted for 6 h. The filtrate was evaporated under reduced pressure to a volume of 15–20 mL. After cooling, 50 mL of diethyl ether was added, then the solution was mixed thoroughly and allowed to clarify. The clear supernatant solution was discarded. The precipitate was purified by adding methanol in the amount of 20, 10, and 5 mL several times, further heated and filtered. The obtained filtrates were combined and evaporated under reduced pressure to a volume of 15–20 mL. After cooling, 50 mL of ethyl ether was added and the procedure was repeated as described above. After re-purification, the methanol filtrates were placed in a pre-weighed, round-bottomed flask, then the solvent was completely stripped in a vacuum and dried to a constant weight at 105 °C.

The sum of saponins content was calculated using the formula:

where:

A—percentage of the sum of saponins in the extract

M—mass of dry extract in grams

m—mass of the residue after drying to a constant mass in grams

The extraction method of inulin by Gibson et al. [56] and Kumari et al. [57] was adapted to the research. Approximately 0.1 g of powered sample of the extract was placed in a 10 mL flask. 10 mL of hot water was added, mixed, and after 5 min of incubation, the sample was filtered. Next, 2 mL of supernatant was transferred to a volumetric flask (10 mL). Afterwards, 2 mL of concentrated hydrochloric acid was added and the sample was incubated for 10 min in a boiling water bath. The sample was cooled down, then 0.5 mL of 0.5% resorcinol in 20% hydrochloric acid was added. The solution was heated for 1 min in a water bath, cooled down, and filled up with water to a volume of 10.0 mL. The absorbance of the test solution was measured at λ = 520 nm in comparison with water.

Calibration curve—an inulin standard stock solution (1 mg/mL) was prepared and was used to prepare solutions with different concentrations. The procedure with the standard solution was the same as with the sample.

All reagents used in the experiment were of analytical grade and obtained from approved sellers.