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freezing point of biological fluids with glycerol

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What is the degree to which the freezing point of blood and other biological fluids is lowered with the help of glycerol. What is the ideal concentration range of glycerol required to achieve this?

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A freezing point of biological fluids with glycerol

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Freezing point of biological fluids with glycerol
What is the degree to which the freezing point of blood and other biological fluids is lowered with the help of glycerol. What is the ideal concentration range of glycerol required to achieve this?
The advantage of including glycerol for freezing point depression in biological samples is that it is non-toxic, soluble in water and allows storage of biological samples at low temperature (<0C) without freezing the sample. This is particularly important for biological samples containing cells (i.e. blood or primary cell cultures). If such a sample is allowed to freeze , water crystals formed can be very damaging and can cause cells to lyse (rupture). In this respect glycerol (or another commonly used "polyol" - polyethylene glycol) acts as a cryoprotectant. The graph below, taken from the Dow Corning site provides a freezing point graph for glycerol /water mixtures as a function of glycerol concentration.
You can see that at a concentration of ~67% glycerol/water (w/w%) the freezing point is lowered to about - 47C.

Reference for above graph -

A possible concern in using too high a concentration of an organic cryoprotectant (including glycerol) is that there is a potential to denature DNA and proteins and disrupt cell membranes and kill cells.
Information below on toxicity of glycerol (http://www.cryonet.org/cgi-bin/dsp.cgi?msg=30125 ) is extensive and suggests that glycerol is the least cytotoxic of a number of cryoprotectants evaluated. Results also indicate that glycerol does not lead to denaturation of DNA ( Pharmazie. 2001 Oct;56(10):808-9) even up to 99% glycerol (Biotechnol Bioeng. 2000 May 5;68(3):339-44 ). Proteins including Lysozyme are denatured in many commonly used cryoprotectants, with the single exception of glycerol (Proc Natl Acad Sci U S A. 1999 Feb 16;96(4):1262-7).
In considering an ideal concentration of glycerol, due to its low toxicity, it can be used at a concentration resulting in the maximum depression of freezing point with minimal risk of denaturation or cytotoxicity. The graph above indicates a concentration of 67% (w/w ) glycerol can be used which will lower the freezing point of blood or other biological fluid by ~47C.

Information and references on glycerol cytotoxicity -

Information below (including literature citations) from http://www.cryonet.org/cgi-bin/dsp.cgi?msg=30125 ) is

X-Message-Number: 30125
Date: Wed, 12 Dec 2007 00:07:47 -0800 (PST)
From: [email protected]
Subject: no denaturation damage from glycerol

[Not much progress has been made in elucidating the nature of cryoprotectant
toxicity since Dr Fahy produced the following paper in 1986. It has been
proposed that the intrinsic toxicity of cryoprotectants is due largely to
their hydrophobicity, and consequence ability to denature DNA and proteins.
However the rank order of hydrophobicity is a poor predictor of overall
cryoprotectant toxicity. In particular, it does not explain glycerol
toxicity, since this solvent ...

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