Plant Cell Reports (1989) 7:652-654 Plant Cell Reports © Springer-Verlag 1989 Growth of Dioscorea deltoidea at high sugar concentrations E. Battat, J. S. Rokem, and I. Goldberg Department of Applied Microbiology, Institute of Microbiology, The Hebrew University-Hadassah Medical School, P.O. Box 1172, Jerusalem 91010, Israel Received July 21, 1988/Revised version received December 2, 1988 - Communicated by F. Constabel Abstract Dioscorea deltoidea cell suspension cultures were grown at initial sucrose concentrations of 35 to 200 $/L. The growth rates were similar (about 0.50 day--) with all of the initial sugar concentrations examined. The ratio of fresh weight to dry weight of cells was dependent on the initial sugar concentra2 tion, however, it remained fairly constant as long as the sugar was present in the growth medium. These results are different from results recently published, claiming that the growth rate of D. deltoidea cells is dependent on sugar concentration and the fresh weight to dry weight ratio increases throughout growth. Introduction There is considerable commercial interest in growing suspensions of plant cells on large scale for the pro- duction of biomass and plant metabolites (Fowler,1983; Rokem & Goldberg,1984; Tanaka, 1987). The carbon source concentration is a major determinant for cellular yield and it has also been shown to affect metabolite formation (Knobloch et al., 1982; Berlin et al., 1983). We have earlier reported that for initial sugar (sucrose) concentration of 15 to 60 g/L, there was no effect on the growth rate and yield of Dioscorea deltoidea cells (Tal et al., 1982). Recently, Drapeau et al. (1986)claimed that the sugar concentration strongly decreased the growth rate and the fresh weight to dry weight ratio of Dioscorea deltoidea cells. If high sugar concentrations would have decreased growth rate, the industrial realization of use of plant cells would be in jeopardy, because in order to obtain high productivity, it is important to be able to grow cells as quickly as possible to high cell densities. In this communication we present results which indi~ cate that at initial sugar concentrations of 35 to 200 g/L the concentration does not affect the growth rate of Dioscorea deltoidea. As was shown earlier (Tal et al., 1982), for initial sugar concentrations up to 60 g sucrose/L, the cellular yield of 0.4 g dry weight/g sugar utilized was constant. The ratio of fresh weight to dry weight was dependent on the initial sugar concentration. However, it remained fairly constant as long as the sugar was present in the growth medium. The weight ratio of cells increa- sed only at the end of the experiment when sugar was almost depleted from the medium. Materials and Methods With Dioscorea deltoidea A-51 callus tissue obtained from E.J. Staba (University of Minnesota, Minneapolis) cell suspension cultures were prepared as previously descrzbed (Tal & Goldberg, i~82). Cell cultures have been maintained for 8 years both as " suspensions and callus cultures. The growth medium contained Murashige and Skoog's (1962) inorganic nutrient mixture (MS) with 140 mg/L inositol, 14 mg/L thiamine, 1,4 mg/L pyridoxine hydrochloride, 1.4 mg/L nicotinic acid and 0.I mg/L 2.4-dichlorophenoxyacetic acid. The KH^PO. concentration was increased to 480 4 mg/L. Prior 4o growth rate and fresh weight to dry weight ratio determinations, the inoculum for each culture was transferred at least twice onto medium with the sugar concentration under investigation. For each transfer (at sugar concentrations up to I00 g sucrose/L) the cells were grown twice to constant weight. The incubation time depended on the initial sugar concentration. For sugar concentrations above 1O0 g/L the cells were transferred to fresh medium after 72 h. Cell suspensions corresponding to I0 to 20% of the final culttrre volume were added as inoculum, so that a constant initial dry weight of cells was used in all experiments. Cells were grown at ]50 r!~m at 28°C. Dry weight and fresh weight were determined as described previously (Tal ~ Goldberg, 1982; Tal et al~, 1983). Sugar concentration was de- termined according to Roe (1949) and to the techni- cal bulletin of Sigma Chemical Co. (St. Louis, Mo.). ReSults As shown in Table i, Dioscorea deltoidea cells were grown at various sucrose concentrations. After an ini- tial short lag phas% the cells entered the exponen- tial growth phase, where growth rates were similar with all of the initial sucrose concentrations exa- mined. The length of the active growth phase was de- pendent on sugar concentration; i.e. the higher the initial concentration the longer it took before the cultures reached the stationary phase. For sugar con- centrations up to 100 g/L the cellular yield was 0.4 g cell dry weight/g suqar utilized. The final pH was de- pendent on initial sugar concentration (Table 2). For sugar concentrations up to I00 g/L the pH decreased to 4.5, a pH which still allowed for growth of the cells. Offprint requests to: J. S. Rokem