X-Message-Number: 7581
Date: 28 Jan 97 15:28:02 EST
From: Michael Darwin <>
Subject: Cryonics, cryoprotectants, conflict

Jan Cotzee writes:

>How toxic cryo protectants are is directly related to the quality and 
>purity of these chemicals. Apparently Visser's critics do their 
>experiments with 75c per liter chemicals. This is why she can 
>demonstrate that her heart and HIV methodes work and theirs fail! .

Jan, where did you get such information?  Who told you could buy DMF at $0.75 a

liter from _any_ source?  Which of "Visser's critics" are using DMF purchased at
such a price?

Since 21st Century Medicine (21CM) and BioPreservation (BPI) have worked with

DMF and attempted to reduplicate Visser'swork to some extent (and your statement
is without qualifications), I take these allegations _very_ seriously as they
are both untrue and create a impression of unprofessionalism and incompetence
about 
21CM and BPI.  This is unfortunate.

THE FACTS:  We do a lot of cryoprotective agent sceening.  We have from the
start known that purity and storage procedures used for evaluating putative
cryoprotectants are critical to success.  We generally use high pressure liquid
chromotography (HPLC) grade materials when we can get them in that purity.
Otherwise we use reagent grade at a minimum.  I can think of only one putative
cryoprotectant we have evaluated that was "technical grade" and that was
triethylene glycol diacetate (TEGDA).  We were constrained to do this because
TEGDA is not available as a reagent grade or HPLC grade chemical (at least we
could not find it). Incidentally, TEGDA failed as CPA for reasons we are
reasonably confident were not related to purity.


The dimethylformamide (DMF) we have used has come from several different sources
and the _minimal_ purity has always been reagent grade.  Our sources were:

Sigma Chemical Company (reagent and HPLC)
Spectrum Chemicals (reagent)
BASF Wyandotte (reagent)

We have found no significant differences between suppliers in how this agent
behaves.

We _have_ found very significant differences in the behavior of other
cryoprotectants we have evaluated in terms of supplier and purity.  Generally,

freezing point depression, toxicity and glass forming ability are better in HPLC
grade chemicals _if_ they are agents with the potential for containing toxic
quantities of unreacted starting materials used to synthesize them.  Technical
grade TEGDA might, for instance, contain acetic acid.  Water as an impurity is
another critical consideration.

Further, some years ago Greg Fahy had a costly experience with DMSO which bears
repeating.  The difference between survival and nonsurvival of kidney slices

exposed to VS4 can hinge on as little as a difference of 1% in the concentration
of the VS4.  They had a successful run of experiments with a slightly modified
version of VS4 which they were subsequently unable to reproduce.  The one
variable (other than the one they _knew_ about) was found to be the bottle of
DMSO that was used to prepare the solution.  Spectrophotometic analysis of this

bottle subsequently demonstrated that it contained water (thus diluting it) from
being stored with a loose cap in the refrigerator.

At 21CM and BPI we are acutely sensitive to such complicating factors.

We do not use technical grade or industrial grade chemicals for cryobiological
research. 

Mike Darwin
21st Century Medicine, Inc.
10743 Civic Center Drive
Rancho Cucamonga, CA 91730
(909)987-3883


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