Alkaloids from regenerated plants

Miura and co-workers 121) have successfully induced multiple shoot cultures of C. roseus from seedlings in the presence of 1.0 mg/liter of the cytokinin benzyladenine. Vindoline (3) and catharanthine (4) were predominating alkaloids in the MSC-B-1 line, showing levels of 1.8 and 0.37 mg/g dry weight, respectively in the leaf tissue. In the case of catharanthine (4) this represented a 10-fold increase over the parent plant tissue, and such levels were sustained in the regenerated plants. When the benzyladenine was eliminated, overall growth was reduced, but vindoline (3) and catharanthine (4) concentrations increased to 3.2 and 1.1 mg/g dry weight, respectively. 

Table 9. Alkaloid contents in 1-year old regenerated plants from internodal segments...

Table 11. Alkaloid content in the regenerated plants from the cultured root segments and shoot meristem culture...

Hortiamine, which can lower blood pressure, is a red alkaloid and the major hypotensive component of the bark of the Brazilian plant Hortia arborea (Rutaceae). Chemically, it is 10-methoxy-14-methyl-5-oxo-5,7,8,14-tetrahydroindolo[2, 3 3,4]pyrido[2,l-6]quinazoline.43 When boiled with moist benzene it forms a yellow product, hydrated as shown in 44, and Amaxfalls from 411 nm to 375 nm. The anhydrous form is regenerated on gentle drying.71... 

The winning of the alkaloids is generally achieved by the usual procedures modified to suit the plant source and the properties of the bases. Pais el al. 17), when working with Panda oleosa, moistened the dried and ground plant material with 10% aqueous ammonia and 1% aqueous sodium carbonate followed by extraction with ether. The concentrated extract was shaken with aqueous amidosulfonic acid and the bases regenerated from the aqueous phase by ammonia. An ether solution of the liberated bases frequently yields crystals upon standing but these are virtually always a mixture of bases. 

Scutianine-A was obtained from a methanol extract of the ground plant. The somewhat concentrated extract was diluted with water, basified with ammonia to pH 8, and exhausted with benzene. The alkaloids were removed from the benzene extract by means of 5% aqueous citric acid. The bases, regenerated from the aqueous phase, were recovered with chloroform or methylene chloride (18). This procedure is recommended when large amounts of plant material are available. 

Fifteen shoots thus obtained were transferred to HF solid medium for rooting. The method described in section 1.1 declares auxin as essential for rooting, whereas the shoots here could be rooted easily without auxin and could be transplanted to pots and cultivated in a greenhouse. The plants regenerated from the intemodal segments exhibited quite normal features, Fig. (22), as well as accumulated alkaloids in same amounts as the parental plants (Tables 4, 9). 

The cell and tissue culture of the major tropane alkaloid-producing species does not apparently offer any special problems. The regeneration of plantlets from callus and tissue cultures seems to be routine (286,307,309,323,325,332,350-352). Plants have also been regenerated from protoplasts of Atropa belladonna (353), Duboisia myoporoides (354), and Hyoscyamus muticus (355,356). Cryopreservation has been reported for Anisodus and Datura species (349,357). 

An alkaloid, jaborandine, has been shown to be responsible for the local anaesthetic properties of this plant 186). From the purified hydrochloride, Ci8H2802N2-HCl (mp 134°) the free base was regenerated. On alkaline hydrolysis it gave p-aminobenzoic acid and a base, CioHigNMeOH, whose benzoyl derivative formed a crystalline picro-lonate (mp 105°). The hydrolytic base on selenium dehydrogenation gave a base which is probably a derivative of pyridine 187). This alkaloid should not be confused with the jaborandine of uncertain composition isolated from a Piper species not further identified (Vol. Ill, p. 207). 

Callus and cell suspension cultures of opium poppy have, so far, not been seen to develop typical laticifers as displayed, for instance, in micrographs of shoots from seedlings and regenerants by Thureson-Klein (1970) and Nessler and Mahlberg (1977, 1979). Laticifer-like cells were demonstrated foxP. bracteatum cell cultures by Kut-chan et al. (1983). Absence of typical laticifers was often cited as the reason for failure to show occurrence of thebaine or codeine in poppy cell cultures. Today one may propose that accumulation of low amounts of morphinan alkaloids do not require latex and respective vessels. Indeed, other parts of the plant besides latex have... 

Yoshikawa T, Furuya T (1983) Regeneration and in vitro flowering of plants derived from callus cultures of opium poppy Papaver somniferum). Experientia 39 1031-1033 Zenk MH, El-Shagi H, Arens H, Stockigt J, Weiler EW, Deus B (1977) Formation of the indole alkaloids serpentine and ajmalicine in cell suspension cultures of Catharanthus roseus. In Barz W, Reinhard E, Zenk MH (eds) Plant tissue culture and its bio-technological application. Springer, Berlin Heidelberg New York, pp 27-43... 

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