5. Advantages of cell, tissue and organ cultures as sources of secondary metabolites

5.1. Plant cell cultures

Once interesting bioactive compounds have been were identified from plant extracts, the first part of the work consisted in collecting the largest genetic pool of plant individuals that produce the corresponding bioactive substances. However, a major characteristic of secondary compounds is that their synthesis is highly inducible, therefore, it is not certain, if a given extract is a good indicator of the plant potential for producing the compounds. The ability of plant cell cultures to produce secondary metabolites came quite late in the history of in vitro techniques. For a long time, it was believed that undifferentiated cells, such as callus or cell suspension cultures were not able to produce secondary compounds, unlike differentiated cells or specialized organs.

5.2. Callus culture

Callus is a mass of undifferentiated cells derived from plant tissues for use in biological research and biotechnology. In plant biology, callus cells are those cells that cover a plant wound. To induce callus development, plant tissues are surface sterilized and then plated onto in vitro tissue culture medium. Different plant growth regulators, such as auxins, cytokinins, and gibberellins, are supplemented into the medium to initiate callus formation. It is well known that callus can undergo somaclonal variations, usually during several subculture cycles. This is a critical period where, due to in vitro variations, production of secondary metabolite often varies from one subculture cycle to another. When genetic stability is reached, it is necessary to screen the different cell (callus) lines according to their aptitudes to provide an efficient secondary metabolite production. Hence, each callus must be assessed separately for its growth rate as well as intracellular and extracellular metabolite concentrations. This allows an evaluation of the productivity of each cell line so that only the best ones will be taken for further studies, for example, for production of the desired compound in suspensions cultures.

5.3. Cell suspension cultures

Cell suspension cultures represent a good biological material for studying biosynthetic pathways. They allow the recovery of a large amount of cells from which enzymes can be easily separated. Compared to cell growth kinetics, which is usually an exponential curve, most secondary metabolites are often produced during the stationary phase. This lack of production of compounds during the early stages can be explained by carbon allocation mainly distributed for primary metabolism when growth is very active. On the other hand, when growth stops, carbon is no longer required in large quantities for primary metabolism and secondary compounds are more actively synthesized. However, some of the secondary plant products are known to be growth-associated with undifferentiated cells, such as betalains and carotenoids.