X-Message-Number: 30337 Date: Sun, 20 Jan 2008 23:57:02 -0800 (PST) From: Subject: The lowest toxicity cryoprotectant is... (part 1/2) [It may be 1,3-propanediol. (also called trimethylene glycol) 1,3-propanediol has been tested just three times as a cryopreservative, in 2006 (PMID: 17256068), 1989 (2708506), and 1980 (7431324). The reason it has been largely over-looked in the field of cryopreservation appears to be because it is a poor glass former, while the most effective cryoprotectants tend to good glass formers. However good glass formers have a strong tendancy to exhibit increased toxicity in vitrification solutions (see Fahy et al in 14969679). As a consequence vitrification solutions tend to contain small amounts of good glass formers, supplemented by larger amounts of relatively poor glass formers to eliminate ice formation, while incurring a reduced toxicity penalty. An extract from table 1 of (10788314) implies that 1,3-propanediol is the lowest toxicity cryoprotectant. Fahy found that substituting ethylene glycol for 1,2-propanediol reduced vitrification toxicity. If Fahy is correct then further substituting 1,3-propanediol for ethylene glycol should further lower toxicity. Critical Cooling Rates to Avoid Any Crystalization 45% solute C/minute) L2,3-butanediol <2.5 1,3-butanediol 7 1,2-propanediol 10 1,2,3- butanetriol 11 dimethylsulfoxide 14 dimethylformamide 23 diethylformamide 48 1,4-butanediol 71 ethylene glycol 260 glycerol 380 1,3-propanediol 1100 There exists independent data confirming that 1,3-propanediol is the lowest toxicity cryoprotectant. In (7431324) the effect of various concentrations of cryoprotectants on rabbit spermatozoa motility at 20 C for 30 minutes was documented in table 1 below. At the highest dosage tested 1,3-propanediol showed the highest motility, indicating that it was the least toxic. Cryoprotectant 0.31M 0.93M 1.55M 2.17M 3.10M 1,3-propanediol 50 52 54 54 52 Dimethylsulfoxide 62 63 61 60 38 Lactamide 61 62 57 51 35 Acetamide 60 61 54 45 30 Ethylene glycol 48 50 50 39 27 Glycerol 47 23 7 3 2 Dimethylformamide 63 55 33 18 0 Propionamide 44 46 19 6 0 Malomide 61 33 7 5 0 Formamide 53 39 16 4 0 Butyramide 30 17 0 0 0 The effect on motility after freezing and thawing was also measured, as shown below. Here 2.0M solutions were always significantly more toxic than 1.0 M solutions, except those for 1,3-propanediol. However 1,3-propanediol still did not have the best results even at the most concentrated 2.0M solution. This highlights the difference between most effective, and least toxic. If one were developing a vitrification solution based solely on the results shown here, one would base it on the most effective cryoprotectant, which was 1.0M lactamide. The rest of the vitrification solution would then consist of 3.1M plus of 1,3-propanediol. Cryoprotectant 0.125M 0.25M 0.5M 0.75M 1.0M 1.25M 1.5M 2.0M 1,3-propanediol 19 11 11 13 17 23 16 15 Dimethylsulfoxide 25 31 31 38 44 36 28 17 Lactamide 5 5 29 42 53 - 47 31 Acetamide 9 12 24 37 40 33 25 2 Ethylene glycol 12 17 24 22 22 20 22 15 Glycerol 12 10 8 4 2 <1 <1 <1 Dimethylformamide 14 27 32 30 19 - - - Propionamide 6 4 6 22 12 2 0 0 Malomide 12 5 <1 - - - - - Formamide 4 6 6 3 <1 <1 <1 <1 Butyramide 2 0 0 - - - - - Without(control) <1 Below are some relevant abstracts.] Rate This Message: http://www.cryonet.org/cgi-bin/rate.cgi?msg=30337