Catharanthus roseus L. (Periwinkle)

The periwinkle, Catharanthus roseus, is a member of the family Apocynaceae and belongs to the subfamily Plumerioideae. This plant is native to Madagascar, but is cultivated as an ornamental plant throughout tropical and subtropical areas, and has been abundantly naturalized in many parts of the world. The plant has many uses in folk medicine in tropical areas. For example, gargling with an infusion of the plant is considered to relieve pain from a sore throat, laryngitis, and chest complaints in Central America. Juice squeezed from the leaves is applied to wasp stings in India.

All parts of the plant contain alkaloids, and the leaves, in particular, contain some antineoplastic bisindole alkaloids, among which are vinblastine and vincristine, which have been widely employed as chemotherapeutic agents against cancers such as Hodgkin’s lymphoma and acute leukemia. Bisindole alkaloids are composed of vindoline and catharanthine-like moieties with minor various molecular modification. The isolation of these alkaloids is costly because of their very low levels in the plant. In the past 30 years, many attempts have been made to produce these alkaloids in cultured cells; however, no bisindole alkaloids have been isolated from such cells to date. Recently, there have been many investigations of the potential synthesis in vitro of these alkaloids via chemical or enzymatic coupling of vindoline and catharanthine. In the intact plant, vindoline is localized in the leaves, and catharanthine is found in both the leaves and roots. The mature plant is a rich source of vindoline, but its catharanthine content is much lower and varies widely, depending on the age of the plant and the environmental conditions. Catharanthine can also be produced by cultured cells, and there have been many reports of this phenomenon. High levels of catharanthine have been obtained by improvements in cultivation conditions, by using immobilization techniques, and by treatment with various elicitors, such as homogenates of fungal mycelia and vanadium. Furthermore, a scale-up of cell cultures for the production of catharanthine has been achieved with 14-1 and 170-1 jar fermentors. It can be concluded from the results of these investigations that cell cultures are a more useful source of catharanthine than the intact plant. In contrast, vindoline has not been produced by cell culture because its biosynthesis requires the structural organization associated with intact leaves and light irradiation. Therefore, the supply of vindoline and catharanthine for synthesis in vitro of bisindole alkaloids must depend on two different sources.

Vindoline and catharanthine can be produced simultaneously in shoot cultures. In multiple shoot cultures (MSC-B-1 line) established in our laboratory, vindoline was the most abundant alkaloid, and its level was mostly equal to that in the parent plant. However, the level of catharanthine was much higher than that in the plant, but was still only about 20% that of vindoline in the cultures. Vinblastine was also detected in the cultures. Bisindole alkaloids are synthesized from equimolar amounts of vindoline and catharanthine by a chemical or enzymatic coupling reaction. Therefore, it is necessary, if multiple shoot cultures are to be a useful source of vindoline and catharanthine, that the levels of the alkaloids be much higher than those in the plant leaves, and their stoichiometric ratio be about 1:1.

In this study, we selected a new line of multiple shoot cultures capable of producing similar levels of vindoline and catharanthine and the cultivation conditions to improve the productivity of these alkaloids in the cultures were optimized.

Catharanthus roseus L. (Periwinkle): Conclusions and Prospects

Multiple shoot cultures of Catharanthus roseus stably produced vindoline and catharanthine, which are the biosynthetic precursors of bisindole alkaloids and important substrates for chemical and enzymatic synthesis of alkaloids, such as vinblastine. The production of vindoline and catharanthine in the cultures was affected by phytohormones and light in different ways. The rates of production of vindoline and catharanthine were markedly increased by the two-step cultivation procedure, in which the first step was a growth-stimulation phase and the second step an alkaloid-production phase. However, the production of vinblastine and leurosine from vindoline and catharanthine in the cultures was specifically stimulated by near-ultraviolet light with a peak at 370 nm. The action of the near-ultraviolet light would be applied to the efficient conversion of vindoline and catharanthine, accumulated in the cultures by our two-step cultivation, to bisindole alkaloids.

Shoot cultures offer industrially attractive material for the production of useful metabolites that are not synthesized in cell cultures and root cultures. However, large-scale cultivation of shoot cultures is much more difficult than that of cell and root cultures because the cultures require light irradiation and gas exchange on the leaves. Unique methods suitable for large-scale shoot cultures must now be developed.

Selections from the book: “Medicinal and Aromatic Plants VI”, 1994.