Uptake of C-13-glucose by cell suspensions of carrot (Daucus carota) measured by in vivo NMR: Cycling of triose-, pentose- and hexose-phosphates

After a lag phase of 2 days, batch-grown cells of carrot (Daucus carota L,) cv, Flakkese entered the exponential growth phase and started to accumulat e sucrose and hexoses, Short-term feeding C-13-glucose in this period resul ted in only minor labelling of sucrose or fructose. CO2 production from [1- C-13]- and [6-C-13]-glucose revealed, that at least 40% of the added glucos e passed through the oxidative pentose phosphate pathway (OPPP), up to 40% through glycolysis leaving only minor C-13-glucose for incorporation in var ious cell components in the exponential growth phase. After about 11 days o f culture, the medium sugars were exhausted, cells entered the stationary g rowth phase and consumed stored sugar. Both neutral and acid invertase (EC 3.2.1.26) and sucrose synthase (EC 2.4.1.13) increased 50% from day 0 to da ys 11-13; thereafter their levels decreased again, Labelling with C-13-gluc ose resulted in the accumulation of labelled sucrose and fructose during th e stationary growth phase. Sucrose labelling was transient, i.e, after 6 h its level started to decrease again, Labelled fructose, however, evolved sl ower and increased even after 8 h. In sucrose and fructose up to 20% of the C-13-label was exchanged from C-1 to C-6 carbons, indicating intensive cyc ling of at least 40% of the carbon between hexoses and triose phosphates. I n the stationary phase only 10% of the labelled glucose passed through the OPPP and about 30% passed through the respiratory pathway; the remaining 60 % was incorporated in cell constituents and sugars, Comparing the various c ycles revealed that the regulation-of the OPPP operated relatively independ ently from the cytosolic cycling of hexose phosphates through sucrose and f rom the cycling between hexose phosphates and triose phosphates.