An in vivo 31P NMR comparison of freely suspended and immobilized Catharanthus roseus plant cells

Abstract

31P NMR spectra obtained for Catharanthus roseus plant cells entrapped in agarose or alginate were compared to those obtained for freely suspended cells. Oxygenated buffer was circulated through the NMR tube in all three instances. Essentially no differences were observed in the levels of the major metabolites: ATP, NAD(H), UDPG, cytoplasmic P i, and sugar phosphates. Furthermore, the saturation of ATP by Mg 2+, the energy status as determined from the ADP/ATP ratio, as well as the intracellular cytoplasmic and vacuolar pH, were not affected by the entrapment in the polymers. Since the cytoplasmic pH for freely suspended cells drops about 0.5 pH units upon shifting from aerobic to anaerobic conditions, these results indicate that the majority of the entrapped cells are properly oxygenated and that products such as CO 2 can diffuse away. In addition, we found that the uptake rate of P ifrom the medium was lower in the polymer entrapped cells, and that the utilization rate of P i from the vacuolar pool was also considerably reduced. However, the same pattern of P i uptake, storage and utilization was observed in all cases. Thus, all the results obtained with the noninvasive 31P NMR technique suggest that the entrapment in agarose or alginate does not adversely affect cell metabolism since the phosphate metabolism and the cytoplasmic pH appear unaltered.