I N preparing nuclei by means of non-aqueous solvents, there are several technical factors that must be considered with respect to the study of enzymes. These include the effect of freezing, the effect of lyophilization, and the effect of solvent treatment. Siebert [lo] has presented evidence that the solvents used had little effect on the activity of many of the enzymes studied. The effects of freezing and lyophilization do not appear to have been defined however and could very well have a profound effect upon the results and their interpretation. Berenbom et al. [3] studied the effect of freeze-drying on the activities of several hydrolytic and oxidative enzymes in mouse liver about ten years ago. They found that the hydrolytic enzymes suffered only slight losses in activity as a result of this treatment. Cytochrome oxidase and succinoxidase, however, were inactivated 50 per cent or more by freezing alone. The possibility must thus be considered that selective effects are being observed in the study of lyophilized tissues. If one assumes for the purposes of discussion that freezing and lyophilization also did not affect the enzyples studied by Siebert [lo], the results indicate that with one notable exception (DPN-pyrophosphorylase), most of the enzymes studied were present in the isolated nuclei in about the same concentration as in the original tissue. It is the interpretation of the latter findings that is the main point at issue. This is an important point because if the results are accepted at face value, one would have to conclude that the nucleus is able to carry out glycolysis, certain oxidations and hydrolytic functions. In short, it would almost appear as if the nucleus is able to conduct all of the functions that the cell is able to perform. The late Dr George H. Hogeboom and the author have always felt however that it is very difficult to interpret such data [9]. The nucleus, for example, accounts for 10 per cent or less of the mass of the liver cell. Consequently, if the concentration of enzyme in the nucleus is the same as in the original tissue, less than 10 per cent of the total tissue enzyme would actually be localized in the nucleus. The remaining 90 per cent would have to be localized in the cytoplasm. In order to accept the association of such small amounts of
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