Rodzaj Panax – systematyka, skład chemiczny, działanie i zastosowanie oraz analiza fitochemiczna nadziemnych i podziemnych organów żeń-szenia amerykańskiego – Panax quinquefolium L. Metody ekstrakcji i analizy ginsenozydów. Cz. II.

*Tadeusz Wolski, Agnieszka Ludwiczuk, Tomasz Baj, Kazimierz Głowniak

Summary
In this article literature review regarding extraction and identification methods of ginsenosides occurring in Panax genus was presented. Own data which concern analysis of ginsenosides fraction from Panax quinquefolium L. was also described. The occurrence of these compounds in plant material collected in different periods of ginseng vegetation as well in pharmaceutical preparations received from Panax ginseng was compared. This article contain data concerning preliminary investigations of ginseng e.g. the loss of drying and also quantitative analysis of ginsenosides by spectrophotometric method, optimization of extraction method and chromatographic separation for TLC, HPTLC, OPLC and HPLC methods. The quantitative analysis of the main compounds using TLC and HPLC methods in optimized chromatographic conditions was also performed.
Preliminary investigations concerning the loss of drying showed that underground parts of ginseng are characterized by higher content of moisture in comparison to aerial parts. The content of ginsenosides in roots as well in aerial parts of P. quinquefolium increased along with the age of the plants. The highest yield of ginsenosides was obtained in 4-year-old leaves and roots of P. quinquefolium. Ginseng leaves in comparison to ginseng roots are characterized by highest concentration of ginsenosides. Percentage of ginsenosides in roots is ranged from 6.5 to 12.5% while in ginseng leaves from 24.8 to 37.5%. Stems and fruits of Panax quinquefolium are characterized by much lower content of saponins.
Studies on isolation of ginsenosides were concerned selection of extraction solvent and extraction method. The highest extraction efficiency of ginsenosides in roots and leaves of American ginseng was observed during mechanical shaking with 50% aqueous methanol. Separation of ginsenosides conducted in 26°C and by use of chloroform – methanol – ethyl acetate – water, 15+22+40+9 (v/v) as the mobile phase was characterized by good selectivity and resolution of saponins. The best results for ginsenosides separation, using OPLC method, gave mobile phase: chloroform – methanol – ethyl acetate – water – hexane, 20+22+60+8+4 (v/v). Results from TLC, HPTLC and OPLC analyses of ginsenosides indicated that use of the different development methods resulted in significant differences between the resolutions of the compounds. Forced-flow OPLC is more sensitive method for analysis of ginsenosides.
Quantitative analysis of the main ginsenosides occurring in leaves and roots of American ginseng was executed using TLC and HPLC methods in optimized chromatographic conditions. The linear calibration curves for 6 ginsenosides, concentration ranges, limits of detection and limits of quantification for both TLC and HPLC was determined. Recovery for all investigated ginsenosides was between 88.8 and 101.2%. TLC and HPLC analyses showed that the main compound of American ginseng roots is ginsenoside Rb₁. The amount of this compound was ranged from 1.36 to 3% according to extraction method and extraction solvent used. Ginsenosides Rd, Rg₂ and Rb₂ are the main compounds occurring in ginseng leaves. They content in plant material after mechanical shaking was 3.8, 2.4 and 1.4% respectively. The amount of another ginsenosides in ginseng leaves not exceeded 1%. Ginseng leaves, in comparison to ginseng roots are characterized by higher concentration of ginsenosides. Therefore, based on the concentration of major saponins, leaves can by alternative to root source of ginsenosides used in herbal preparations. The lack of ginsenoside Rf in aerial and undergrounds parts of Panax quinquefolium confirmed its status as a phytochemical marker differentiating American and Asian ginseng.

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