X-Message-Number: 3535 Date: 26 Dec 94 00:12:20 EST From: Mike Darwin <> Subject: CRYONICS autolysis The recent postings by Mr. Cotzee on the only source(s) of damage during high temperature preservation being microorganisms or their byprpducts, and further his assertion that natural inhibitors of enzyme activity would block biological degradation of the stored specimin are, quite simply, wrong. While there is truth to the notion that some enzymes have sharp (relatively) temperature ranges where they become inactivated, this is by no means true for ALL enzymes. As long as diffusion can take place these enzymes will continue to operate. I have charted the activity of catalase (from raw liver in 72% glycerol) all the way down to -79xC; it decomposes peroxide and behaves (not surprizingly) much as the Arrhenius equation predicts. Unless it has been deliberately dried before storage, cryoprotectant treated organisms will have active autolytic enzymes in them. In fact, it is probably no small part of hibernators' success that they manage to retool their enzyme activity vs. temp. curves and probably use inhibitors as well. For instance, hibernating woodchucks' serum, if given to rats, will cause a profound drop in metabolic activity, torpor, and hypothermia. This serum factor has even been used to extend high temperature storage time (in the above 0xC range) for solid organs such as the liver and kidney from nonhibernators. To the best of my knowledge this factor has not been purified and characterized and so is referred to as HIT (hibernation induction trigger). Too bad too, because it DOES seem to extend organ storage time significantly. Now, there's a project for Genentech... I would also point out that I have personally observed molds growing in 30% glycerol solutions at -10xC on pieces of chicken aorta. In the typical clinically (and legally) dead cryonics patient autolysis is already underway hours before legal death is pronounced. The immune/inflammatory cascade has been activated by shock, and the serum enzymes for almost every tissue in the body show the impact of premortem hypoperfusion by marked elevation even before legal death. Finally, storage limits for meat at -20xC are usually based not on the risk of bacterial contamination, but rather on autolytic changes in the meat and dehydration. Have you ever tried to eat a steak that's been in your home freezer for several years? Furthermore, the character of the change at such high temperatures in stored meat and produce is so gross that you can smell it and thus taste it as having an "off flavor." As someone who grew up where our family raised, processed and froze corn and other vegetables for winter storage, I can also tell you (again speaking from experience) that you must first blanch the produce, by briefly heating to high but subcooking temperatures, in order to inactivate native enzymes before freezing it; otherwise you will get inferior stored product. The major food processors do the same thing. Similarly, storage times for cooked meat and produce are usually longer than for fresh. The point of all this is that mammals and even vegetables that were not designed to be frozen will NOT behave in the same way that those which are do. Mike Darwin Rate This Message: http://www.cryonet.org/cgi-bin/rate.cgi?msg=3535