Introduction .................................................................................................................................................................. 513 Developments in Excised Root Culture Technique ...................................................... 514Inorganic Nutrition ............................................................................................................................................. 518 Nitrogen Metabolism .......................................................................................................................................... 521 Carbohydrate Uptake and Metabolism ....................................................................................... 528 The B Vitamins ...................................................................................................................................................... 533 Release of Metabolites .................................................................................................................................... 533 Growth-Regulating Substances ............................................................................................................... 536 Differentiation ............................................................................................................................................................ 542 General Discussion ............................................................................................................................................. 547 Literature Cited ........................................................................ : ............................................................................ 574

Excised root culture

Summary

1
Detailed knowledge of the growth requirements and growth responses of excised roots is available for only a few species, of which tomato (Lycopersicum esculentum) is the outstanding example.

2
Excised tomato roots have absolute requirements for thiamine and pyridoxine. The growth of some strains is further enhanced by niacin. The view is advanced that these requirements arise because the vitamins are synthesized in the roots at an inadequate rate rather than from a complete loss of synthetic ability. By use of possible precursors and analogues knowledge has been obtained relating to pathways of vitamin biosynthesis, the specificity of the vitamins and the ‘sparing action’ on vitamin requirements of certain metabolites.

3
Excised tomato roots can assimilate nitrate nitrogen. Recently the development of techniques for the adjustment and stabilization of the pH of the culture medium and the use of a stable ferric ethylenediamine tetra-acetate have made possible growth on ammonium nitrogen.





Absolute requirements for the micro-nutrient elements iron, copper, molybdenum and manganese have been demonstrated. Present evidence indicates that the micro-nutrient requirements of excised roots are qualitatively similar to those of whole plants.



The importance of certain iodine compounds in maintaining the healthy growth of excised tomato roots suggests that the status of iodine as a plant nutrient should be further studied.



4
Sucrose is the only sugar capable of supporting a high rate of excised tomato-root growth. Fructose and glucose, singly or together, only support a very low level of growth. Other sugars and all sugar alcohols tested do not support growth. Preliminary work suggests that organic acids will not prove to be satisfactory carbon sources.





The specificity of sucrose as an effective carbon source does not apply to those monocotyledonous roots so far cultured.



5
Studies have been made of liquid compared with solid medium and of the effects on excised tomato-root growth of temperature and light.

6
The standard technique developed for the culture of excised tomato roots does not make possible the culture of the excised roots of angiosperms in general. In the present state of our knowledge of the growth requirements of excised roots, those of many species and varieties remain intractable to culture.

7
Excised root culture exposes variation from the inter-specific down to the intra-clonal level. Interesting variations between species and between strains within species, in the responses of their excised roots to vitamins, amino-acids and auxins, have been studied.

8
Excised root culture has shed light on the nutritive interrelationships between shoot and root and made possible a new approach to the study of certain aspects of general physiology. Study of the physiological differences underlying variation in culture is laying the foundations of a comparative physiology of excised root systems. Recent work has indicated the potential value of excised root cultures for studies in the biochemical genetics of higher plant cells.

9
The present status of the excised root-culture technique should, in the immediate future, permit of a rapid expansion of our knowledge of the physiological basis and genetic control of the natural variation expressed in the growth requirements and physiological responses of excised roots.

SUMMARY The rate of respiration of tomato stem slices varied considerably, the highest values (0Oi. 2_3) being obtained for plants in a good nutritional state, and the lowest (0o,. I) in starved plants. The respiratory quotient of i-o remained constant. Glucose fermentation was found to follow both glycolytic and alcohol fer mentation pathways, the ratio of ethyl alcohol : lactic acid being 6-6: i. Fermenta tion seems to take place according to the Embden-Meyerhof scheme, as shown by the presence of some of these enzymes operative in this scheme and by inhibition experiments. In the presence of oxygen there was no formation of alcohol or lactic acid. Pyruvate added to tomato stem slices was metabolized by direct oxidation to acetic acid and by dismutation to lactic and acetic acids and C02. The metabolism of acetic acid was demonstrated by its condensation with oxaloacetic acid to form citrate, this being the second time that synthesis of citric acid by this mechanism has been found in plants. The presence of aconitase, of «ocitric dehydrogenase, of succinic dehydrogenase, and of malic dehydrogenase, as well as the inhibition of respiration by malonic acid, favour the hypothesis that oxidation of carbohydrate in tomato stem slices proceeds via the citric acid cycle. The possibility of an auxiliary route, the malic acid oxidation pathway, also was demonstrated. Tomato stem tissue anaerobically split malic acid into glycolic acid. The further oxidation of glycolic, glyoxylic, and formic acid was demonstrated. In experiments with C14-labelled acetate and butyrate a dilution of the C14 labelled acids was found after incubation indicating new formation of these acids and of active participation of fatty acid metabolism in the metabolic activities of the tissue. With the exception of alanine, added amino acids produced a definite increase in 02 uptake without extra formation of ammonia. Experimental demonstration of the possibility of electron transport from sub strate to molecular oxygen in respiration via polyphenol oxidase was provided by the attainment in a tomato tissue homogenate of a coupled oxidation-reduction between a-ketoglutarate and catechol with DPN and tyrosinase as the catalysts. . The presence of cytochrome oxidase was also demonstrated. Thus both systems possibly may take part in the respiration of tomato stem slices.

Metabolism of Slices of the Tomato Stem

Since Quastel and Whetham's discovery in 19251, biochemists have come to regard the malonate ion as one of the very few specific enzyme inhibitors—as a competitive inhibitor of succinic dehydrogenase2 Malonate has therefore been widely used with animal tissue, in work designed to elucidate the Krebs cycle. Recently it has also been applied to plant tissues, but with discordant results. Machlis3 showed that malonate inhibited part of the respiration of barley roots; but several other workers have independently stated that malonate does not inhibit and may even stimulate plant respiration4–7.

James H. M. Henderson

Papers

The influence of some respiratory inhibitors and intermediates on growth and respiration of excised tomato roots