Alkaloid production in cultured tissues and organs of Lycopersicum esculentum Mill

Roddick, James George (1971) Alkaloid production in cultured tissues and organs of Lycopersicum esculentum Mill. PhD thesis, University of Glasgow.

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Excised root cultures, initiated from seedling radicles of tomato (Lycopersicum esculentum Mill. var. Buttons Best of All), have been found to synthesise the steroidal alkaloid tomatine, although alkaloid levels were lower than in intact radicles. There was no evidence of the aglycone, tomatidine, or modified forms of tomatine in cultured root extracts, and the alkaloid was not excreted into the culture medium. A procedure has been devised for purifying extracted tomatine by thin-layer chromatography (TLC). The identity of extracted tomatine has been confirmed by TLC, colour reactions, visible/ultra-violet spectroscopy, mass-spectroscopy and sugar analyses. A positive correlation has been found between growth of cultured roots, as measured by weight, and production of tomatine. This suggests that actively growing cells, and particularly root apices are the sites of tomatine synthesis. Indirect evidence which supports this hypothesis has been presented. However, tomatine does not appear to accumulate in apical regions, but is present in equal concentrations in all parts of the root. The implications of this finding in the mode of distribution of tomatine in the root are discussed. Changes in the concentration of tomatine in cultured roots during growth have been discussed on the basis of differential rates of tomatine synthesis (and possibly degradation) and dry-matter accumulation. Growth of cultured roots was unaffected by the presence of 'authentic' tomatine (up to 100 ppm) in the culture medium. Nor was such alkaloid absorbed to any significant degree or chemically transformed. Radicle growth in tomato seedlings was likewise unaffected by 'authentic' tomatine, but that in lettuce seedlings was inhibited. The question has been raised of whether response of tissues to exogenous tomatine is influenced by the presence of endogenous tomatine. The optimum temperature for tomatine synthesis appears to be higher than that for cultured root growth. There was also evidence that the former process is less adversely affected than the latter by deviations from the optimum temperature. Addition of nitrogenous compounds or steroid precursors to the culture medium did not result in significant increases in tomatine levels in cultured roots. Nevertheless, these experiments, on the whole, confirmed the previously observed correlation between growth of cultured roots, as measured by weight, and tomatine production. No definite correlations could be established between tomatine production and other growth parameters. Small amounts of tomatine have been found in 'spontaneous' hypocotyl callus grown on a medium supplemented with coconut-'milk', but when the tissue developed chlorophyll, no alkaloid could be detected 'Spontaneous' root callus, grown on the same medium, was also devoid of tomatine. Hypocotyl callus was established on two different chemically defined media. On one medium, tomatine was absent from the callus, but on the other it was present. Tomatine was not detected in root callus grown on either of these media. The changes in tomatine- synthesising capacity of hypocotyl callus are thought to be due to differences in certain culture medium components, especially growth substances. New callus cultures have been initated on a chemically-defined medium from hypocotyl, radicle and cotyledon of tomato seedlings. All cultures produced roots during the early stages of growth and tomatine was present in both callus tissue and roots. Alkaloid levels in roots were comparable with those in cultured excised roots of the same age. Both number of roots and tomatine levels in the callus tissues decreased with time, until eventually, neither roots nor alkaloid was produced by any of the cultures. There was evidence that the reduction in alkaloid levels was due to both decrease in numbers of roots and loss, by the callus cells, of the capacity to synthesise tomatine. Liquid suspension cultures were initiated from 'spontaneous' hypocotyl callus using both 'coconut-milk' medium and chemically- defined medium but tomatine was not found in the cells or 'spent' medium of either culture. It is concluded that tomatine biosynthesis is associated with the degree of morphological organisation pertaining within a cell system. Possible reasons for, and implications of, the reduced capacities of cultured tissues and organs to synthesise tomatine, compared with those of the corresponding structures in the intact plant, are discussed.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: D N Butcher
Keywords: Plant sciences
Date of Award: 1971
Depositing User: Enlighten Team
Unique ID: glathesis:1971-73873
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 14 Jun 2019 08:56
Last Modified: 14 Jun 2019 08:56

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