Distribution and Importance
Gentiana species belong to the family Gentianaceae, order Gentianales, superorder Gentiananae, subclass Asteridae, class Magnoliopsida (). The species are divided into several sections according to the morphology of the above-ground organs (). The subgenera Eugentiana Kusnezow and Gentianella Kusnezow () are entered in Flora Europaea as separate genera: Gentiana L. and Gentianella Moench ().
The genus Gentiana comprises about 400 species distributed chiefly in mountain regions, especially in the Alps, the Carpathians, the Central Asia mountains, and the Andes in South America. Due to their impressive and colorful flowers, gentians decorate mountain meadows. Some species are also found in the monsoon zone of India, in New Zealand, and in southern Australia. More rarely, gentians are found in the temperate zone lowlands of the northern hemisphere (). The yellow gentian root was already mentioned as a remedium stomachicum by Galen and Dioscorides (). Apart from Gentiana Iutea L., there are other medicinal species included in many pharmacopoeias and plant registers of the world (). According to most European pharmacopoeias, the official drug may also contain material from Gentiana pannonica Scop., Gentiana punctata L., and Gentiana purpurea L.
The active constituents of the drug are bitter tasting secoiridoid glucosides which are applied in the treatment of gastrointestinal tract diseases (). A beneficial effect of these compounds on the immunological system of the entrails has also been observed (). Gentian roots are applied as a decoction, an extract, and a tincture. According to the Rote Liste (), Radix Gentianae is a component of about 100 drugs belonging to the groups amara, tonica, cholagoga, and laxantia. Once the gentian root was a component of Pilulae stomachicae cum Fello (). Because of their bitter taste, extracts are used in the production of wines and liquors ().
Conventional Methods for Propagation and Demand in the World Market
The difficulties encountered in the harvest of valuable gentian roots from wild populations prompted distributors to attempt the cultivation of these plants. So far, only Gentiana lutea has been an object of interest ().Gentiana lutea cultivation was carried out in the presence of symbiotic fungi as well (). However, the results of cultivation are not very encouraging and primarily they are uneconomical. Plants require good soil conditions and laborious cultivation practices. Moreover, roots suitable for harvest are developed during the flowering period which takes place after some years of growth (). In many countries gentians do not occur or are under protection. For example, in Poland, the entire yellow gentian root supply is imported. The yearly demand for the dry drug for medicinal use only is about 3000 kg. The drug is exported from Europe to Japan, where cultivation is also attempted (). As reported earlier by Franz and Fritz (), a yearly production amounting to 10001 of the fresh gentian root did not satisfy the demand for the drug.
Secondary Metabolites in Gentiana Species
Secoiridoid glucosides are the main compounds with medicinal properties in roots of Gentiana species. Acyl derivatives of glucosides – amarogentin and amaroswerin () – were found to be the most bitter compounds among natural products. Two other compounds, amaropanin and gentiopicroside, are also of biological importance. Gentiopicroside has the lowest bitter coefficient but its content in the drug is higher than that of acyl derivatives (). During enzymatic hydrolysis, gentiopicroside liberates a metabolite with fungistatic properties () called gentiopicral (), previously gentiogenal (). El-Sedawy et al. () characterized the activity of gentiopicroside metabolites on several bacterial tribes and Isiguro et al. () reported on their anticancer properties.
For a long time the bitter coefficient of gentian roots and other drugs are based on the sensorial method. After the structure of biologically active secoiridoids had been established (), spectrophotometric methods were introduced for quantitative analyses (). For these purposes HPLC, FD-MS, DCI-MS, and TLC densitometry methods are equally used (). Bakuridze et al. () made a qualitative analysis of secoiridoids in 17 species (including also pharmaco-poeial ones) of the genera Gentiana and Gentianella and found that all of them contained gentiopicroside, while amarogentin was present in only three species. A secoiridoid glucoside-trifloroside, found for the first time by Inouye et al. () in Gentiana triflora Pall, var. japonica (Kusnez.) Hara, was also detected in Gentiana scabra Bge. and Gentiana pneumonanthe L.
Yellow gentian roots also contain pharmacologically active xanthones (). The occurrence of some known compounds in G. asclepiadea L. and G. cruciata L. was investigated and compared with that of G. lutea and G. punctata (). Flavonoids, xanthones, and L-( + )-bornesitol were found in G. lutea leaves (). The above-mentioned publications initiated a series of papers entitled Contribution a la phytochimie du genera Gentiana, dealing with studies on secondary metabolites, especially flavonoids and xanthones, and their meaning for the chemotaxonomy of the family Gentianaceae (). Similarly, Ghosal et al. () published the series Chemical constituents of Gentianaceae. The isolation by these authors of 6-aryl-2-py rones with adaptogenic and antistress properties from G. pedicellata Wall, was the first report on the occurrence of these compounds in the family Gentianaceae (). In the same species an iridoid glucoside loganin, and its aryl derivative were also found for the first time (). New iridoid glucosides were found in G. verne L. and G. campestris L. () and a new secoiridoid in Gentiana alpina (). Moreover, some Gentiana species contain triterpenoids, phenolic acids, and oils ().
Conclusion and Prospects
Conditions for the micropropagation and callus culture of some species of the genus Gentiana were established. Seedlings were obtained from seeds or isolated embryos placed on a B5 medium with the addition of GA3 and kinetin. It was also found that the stratification of seeds for a few months was not necessary, contrary to earlier suggestions (). Seeds collected in the given year developed into seedlings after they had been placed on the medium for several days at a temperature of 0-4 °C. The rooting of shoots was most effective when they were put in a liquid medium with IAA for 24 h, and then placed on an MS medium without growth regulators. Rooted plantlets were transferred to pots and then to the ground.
In vitro cultures of Gentiana lutea and Gentiana pneumonanthe were reported by Lamproye et al. (), who initiated them also from seeds by placing them on an MS medium without phytohormones. Seedling fragments fed with MS medium with the addition of auxins and cytokinins formed buds which developed into shoots that rooted in the presence of PBA and IBA. These conditions, however facilitated only the growth of Gentiana pneumonanthe. With Gentiana lutea the germination of seeds was initiated by the addition of GA3 (100 mg/1) to an MS medium. In turn, the development of shoots and the induction of roots were only facilitated after the culture was kept for 2 months at 5 °C without light and without phytohormones in the medium.
The micropropagation of Gentiana lutea from apical meristems was carried out on an MS medium with BAP (). Buds appeared when a medium containing kinetin or 2iP was employed. A high rooting index was observed in the presence of IBA or NAA.
The research carried out to date leads to the conclusion that species of Gentiana can be propagated via tissue cultures. Regenerated plants synthesize biologically active secoiridoids and hence they can be of pharmaceutical and economic significance. The presence of these compounds also in the above-ground parts of gentians grown in the ground and in in vitro cultures is striking. Suggestion for the utilization of not only the root but also the herb of G. lutea as a medicinal material () seems to be justified.
Selections from the book: “Medicinal and Aromatic Plants IV”, 1993.