Wr. Cullen et Di. Hettipathirana, APPLICATION OF WHOLE-CELL NMR TECHNIQUES TO STUDY THE INTERACTION OF ARSENIC COMPOUNDS WITH CATHARANTHUS-ROSEUS CELL-SUSPENSION CULTURES, Applied organometallic chemistry, 8(5), 1994, pp. 463-471
H-1 spin-echo NMR spectroscopy of intact cells of C. roseus facilitate
s monitoring changes inside the cells on treatment with arsenicals. Th
is in situ detection method is non-invasive and nondestructive in comp
arison to other available biochemical methods. Short term uptake of th
e arsinicals, methylarsinate MMA and dimethylarsenate DMA, by C. roseu
s cells that have reached stationary phase in 1-B5 medium, is followed
by using NMR spectroscopy, and in particular, the Carr-Purcell-Meiboo
m-Gill pulse sequence. An increase in the peak height of the methylars
enic resonance over a period of 11 h is indicative of uptake of each a
rsenical. However, there is no evidence of any biotransformation produ
cts in the H-1 NMR spectra. The accumulation site of DMA is probably t
he vacuole as is seen from the change in the chemical shift of DMA as
it moves into a compartment of lower pH. Biochemical changes associate
d with the presence of arsenicals are evident in the H-1 NMR spectra o
f C. roseus cells isolated at different stages in the growth cycle. Al
though uptake has been demonstrated by other analytical techniques, th
e resonances corresponding to both MMA and DMA are not observed in the
H-1 NMR spectra of cells growing in media containing each arsenical.
The association of these arsenicals with large biomolecules in the cel
l may account for these absences. In this event, the spin-spin relaxat
ion time of the arsenic species would shorten and the signals would no
t be seen in the spin-echo NMR spectrum. In cells growing in the prese
nce of MMA, a new resonance is observed at a chemical shift position 2
.2 ppm after 15 days of growth. The shift in position of the resonance
, from 1.75 ppm expected at physiological pH, may indicate an altered
environment around the arsenic species such as high intracellular acid
ity.