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Cytoplasmic pH of Dunaliella parva and Dunaliella acidophila as monitored by in vivo 31 P‐NMR spectroscopy and the DMO method
Author(s) -
Gimmler H.,
Kugel H.,
Leibfritz D.,
Mayer A.
Publication year - 1988
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1111/j.1399-3054.1988.tb02013.x
Subject(s) - intracellular ph , dunaliella , cytoplasm , nuclear magnetic resonance spectroscopy , in vivo , biochemistry , biology , atpase , biophysics , chemistry , algae , intracellular , stereochemistry , botany , enzyme , microbiology and biotechnology
In connection with investigations of the pH‐stat mechanism of the unicellular, halotolerant green algae Dunaliella parva and the acid‐resistant species D. acidophila , a comparative study on the in vivo measurement of the cytoplasmic pH of Dunaliella cells was carried out, using the so‐called dimethyloxazolidine‐2,4‐dione (DMO) method (distribution of a weak acid between the cells and the medium) and 31 P‐NMR spectroscopy. As judged by the effects of the external pH, light, anaerobiosis, nitrogen nutrition, permeable weak acids and bases and of ATPase inhibitors, both methods yield reproducible and in principle similar results, although absolute pH values obtained by the NMR method were always slightly higher than those obtained by the DMO method: 1) D. parva is able to maintain a cytoplasmic pH close to 7.0 at external pH 5–8, whereas D. acidophila maintains a neutral pH even at an external pH of 1. 2) The pH‐homeostasis requires ATP. 3) During illumination the cytoplasm and the stroma is alkalized. 4) Anaerobiosis induces reversible acidifications of the cells. The physiological importance of these effects is discussed. Advantages of the NMR method for the assessment of the cytoplasmic pH of D. parva are: 1) Better resolution of kinetic effects. 2) Independence of volume determinations. 3) Information about phosphate, phosphomonoester, nucleotide and polyphosphate contents of the cells. However, the ability of 31 P‐NMR spectroscopy to monitor pH values in different compartments of cells could not be exploited with Dunaliella cells: Although a light‐induced alkalization of the cells was observed, differences in the pH of the cytoplasm and the stroma of illuminated cells could not be detected. The disadvantages of the 31 P‐NMR method are the low sensitivity in comparison to the DMO method and the requirement for an appropriate calibration matrix. Advantages of the DMO method are the high sensitivity and the possibility of measuring many samples in parallel. Disadvantages of the DMO method are the poor resolution of kinetic effects and the need to know the osmotic volume of the cells.