Tissue Cultures, Microbial Transformations

Tissue Cultures, Microbial Transformations.—Little success has rewarded the search for cell cultures that effectively biosynthesize monoterpenes de novo. The most impressive studies utilize cultures from a variety of Mentha spp. yields of oil were some 60 % (w/v) of those in the parent plants, but the monoterpene products were generally more oxidized (i.e. ketones extra C=C bonds predominated). In vitro, oxidation at C-3 of the menthane skeleton was also restricted, apparently owing to an inhibition of the enzymic reduction of the 4(8) double bond in the intermediates formed.925 926 Colchicine stimulated synthesis of essential oil by Mentha cultures.927 Iridoid glucosides have been produced by cultured cells of Gardenia spp.673 Menthone was biotransformed to neomenthol by Mentha suspension cultures,928 and Nicotiana lines oxidized linalool and its derivatives at C-10 to aldehydes and alcohols,929 and also foreign substrates such as a-terpineol (at C-6 and C-7) and /raw.s-/ -menthan-9-en-l-ol (at C-4 and C-10).930 

Monoterpenols were esterified by lipases from various micro-organisms (especially Aspergillus spp.),931 and ( )-carvyl acetates were hydrolysed by other species to give chiral carveols together with (unreacted) acetates of the enantiomer.932 The metabolic pathways for the conversions of (—)-carvone into (—)- 

Kyonaas, C. Martinkus-Taylor, and R. Croteau, Plant Physiol., 1980, 65, Suppl., 96. 

Paupardin, Prod. Subst. Nat. Cult. In Vitro Tissue Cell. Veg., J. Etud., 1979, 119. 

Okumura, and Y. Tsujisaka, Agric. Biol. Chem., 1980, 44, 2731. 

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All of the principles and ideas covered in the previous section may be translated directly to the use of microorganisms as tools in the production of compounds of plant biosynthetic or biodegradative importance. Just as one finds microbial systems to be of value in preparing metabolites in mammalian systems, it may be possible to use microbial transformations to prepare derivatives of alkaloids that might be found rarely or only in very small quantities in plants. In this way, abundant prototype alkaloids may be used as microbial transformation substrates to provide a range of metabolites. As in the mammalian case, metabolism studies using plant tissues, tissue cultures, or cell-free extracts may be conducted in parallel with microbial metabolic systems. Metabolites common to both would be prepared in quantity by relatively simple fermentation scale-up methods. 

For example, vanillin can be obtained via at least five different ways (i) by isolation from the orchid (Vanilla planifolia), which is a very expensive method (ii) by tissue culture followed by extraction (iii) by microbial transformation of eugenol, the main compound of clove (iv) from lignine by synthesis, and (v) from guaiacol, a natural aroma compound, with comparable molecular structure. Only the vanillin obtained via the first three methods is natural. The other routes afford a nature-identical vanillin. 

Biotransformations in tissue culture like microbial transformation are mostly focused on steroid biosynthesis 97>. By the addition of organic compounds to the medium, substrates of new structures with new chemical and biological properties can be synthesized. 

In fermentation reactors, cell growth is promoted or maintained to produce metabolite, biomass, transformed substrate, or purified solvent. Systems based on macro-organism cultures are usually referred as tissue cultures. Those based on dispersed non-tissue forming cultures of micro-organisms are loosely referred as microbial reactors. In enzyme reactors, substrate transformation is promoted without the life-support system of whole cells. Frequently, these reactors employ immobilized enzymes, where an enzyme is supported on inert solids so that it can be reused in the process. Virtually all bioreactors of technological importance deal with a heterogeneous system involving more than two phases. 

The importance of bioconversions in the industrial production of steroids has been reviewed ° and other reviews on the applications of microbial transformations have appeared.Biotransformations by plant tissue cultures and the application of mathematical models to optimization of fermentation have been reviewed. 

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