Abstract
Mitochondria from green leaves of spinach have been prepared using a three-step procedure involving differential centrifugation, partition in an aqueous dextran polyethylene glycol two-phase system and Percoll gradient centrifugation. The mitochondrial fractions after the different steps of purification were compared. The final mitochondrial preparation was totally free from chloroplast material measured as chlorophyll content. The enrichment of mitochondria in relation to peroxisomes and microsomes was approximately 12 and 33 times, respectively, based on NAD:isocitrate dehydrogenase activity, glycolate oxidase activity, and NADPH:cytochrome c oxidoreductase activity. The apparent intactness of the inner and the outer mitochondrial membranes was higher than 90% as measured by latency of enzyme activities. The mitochondria showed high respiratory rates with respiratory control and the ADP/O ratios approached the theoretical limits.
Full text
PDF
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- BRUINSMA J. A comment on the spectrophotometric determination of chlorophyll. Biochim Biophys Acta. 1961 Sep 30;52:576–578. doi: 10.1016/0006-3002(61)90418-8. [DOI] [PubMed] [Google Scholar]
- Douce R., Christensen E. L., Bonner W. D., Jr Preparation of intaintact plant mitochondria. Biochim Biophys Acta. 1972 Aug 17;275(2):148–160. doi: 10.1016/0005-2728(72)90035-7. [DOI] [PubMed] [Google Scholar]
- Douce R., Moore A. L., Neuburger M. Isolation and oxidative properties of intact mitochondria isolated from spinach leaves. Plant Physiol. 1977 Oct;60(4):625–628. doi: 10.1104/pp.60.4.625. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jackson C., Dench J. E., Hall D. O., Moore A. L. Separation of mitochondria from contaminating subcellular structures utilizing silica sol gradient centrifugation. Plant Physiol. 1979 Jul;64(1):150–153. doi: 10.1104/pp.64.1.150. [DOI] [PMC free article] [PubMed] [Google Scholar]
- LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
- Nishimura M., Graham D., Akazawa T. Isolation of intact chloroplasts and other cell organelles from spinach leaf protoplasts. Plant Physiol. 1976 Sep;58(3):309–314. doi: 10.1104/pp.58.3.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Siegenthaler P. A., Depéry F. Influence of unsaturated fatty acids in chloroplasts. Shift of the pH optimum of electron flow and relations to deltapH, thylakoid internal pH and proton uptake. Eur J Biochem. 1976 Jan 15;61(2):573–580. doi: 10.1111/j.1432-1033.1976.tb10052.x. [DOI] [PubMed] [Google Scholar]