114 THE ROYAL SOCIETY OF CANADA 



It will ])e ol)scrved that we are at present unal)le to give very exact 

 determinations for the minimal temperature which a frog can survive. 

 As will be explained later, this is due to the fact that cooling for pro- 

 longed periods at fairly constant temperatures is essential, and up to 

 the present time we have been unable to devise a cheap self-regulating 

 thermostat for the requisite range of temperature. Further, until 

 such a constant-temperature apparatus is available, it will be impos- 

 sible to test frogs of the same species for slight individual variations. 



It is interesting to attempt to conceive the conditions of the tissue 

 cells when the body is completely frozen, and while it is yet unkilled. 

 Experiment 1 gives an idea as to the gross condition of the frog at this 

 stage. His abdomen is full of ice particles. His limbs are frozen 

 stiff, and the muscular tissue is full of ice, so that it cuts like cheese, 

 or like hard ice cream. 



Both physical theory and our actual experiments support the view 

 that the blood and lymph contain large amounts of ice crystals. Unless 

 these separate simultaneously in the tissue cell and the lymph bathing 

 it, it seems possible that none separates in the cell before it is killed. 

 Should ice separate out first in the 13'mph, then the increased 

 osmotic pressure of the remaining solution would result in passage of 

 water outwards across the cell membrane. This would prevent freez- 

 ing at a temperature of -0-44°C. The process would continue until 

 much of the water of the lymph was frozen, i.e. until the concentration 

 of the lymph and cell fluid was high. At a corresponding low temper- 

 ature — perhaps in the neighbourhood of 3°C. (corresponding to a con- 

 centration of 5 per cent. NaCl) for muscular tissue — ice commences 

 to separate in the cell, but the cell solution is so concentrated that the 

 separation irretrievably damages the cell-protoplasm and death re- 

 sults. 



Consideration of the effects of cold on certain plant species — also 

 poikilothcrmic organisms — lends some support to this hypothesis. 

 In most plants, and in potatoes, beet-roots, and apples, the formation 

 of ice is followed by death. The ice is usually formed in the inter- 

 cellular spaces and not within the cells themselves. "The mode of 

 freezing is due to the fact that the thin film of water which covers 

 the outer walls of the cells lining intercellular spaces freezes first. This 

 causes more water to be withdrawn, and this again freezes. As water 

 is withdrawn from the cell, the cell-sap concentrates, for the salts are 

 kept back by the protoplasm, so that the ice formed in the intercellular 

 space is nearly pure."* According to Miiller-Thurgau f and Molisch J 



*Pfeffcr'.s "Physiology of Plants," trans. Ewart, 1903, vol. II, p. 240. 



tLandw. Jalirb., 1886, 15, 534. 



JMolisch, "Das Erfrieren der Pflanzen," 1897, p. 534. 



