Methylation of the Cryopreservative N-Methylacetamide into N,N-Dimethylacetamide in the Living Body

The polar molecule dimethyl sulfoxide (DMSO) protects against freezing-induced damage by interfering with the polarised water arrangement during ice nucleation inside both nucleated and anucleated cells. Although DMSO is a useful cell cryopreservative, the oxidation and lone pair electrons due to the d-orbital effect of the sulfinyl group cause cytotoxicity. Replacement of the sulfinyl group in DMSO with an amide group yields N-methylacetamide (NMA). DMSO and NMA have similar dipole moments and association properties. Because NMA retains the two methyl groups present in DMSO, which are known to confer permeation ability to DMSO, using NMA rather than DMSO attenuates cell death due to cryopreservation, both by ensuring sufficient cryopreservation action and by reducing cytotoxicity. Because the molar concentrations of the cryopreservatives, which are much higher than those of saline, can injure cells and cause cytotoxicity, the fact that the minimum concentration that can provide similar cryopreservative activity of NMA is lower than that of DMSO is helpful, even if complete removal of the cryopreservative is not achieved after thawing, which can occur in the clinical setting. Moreover, it is possible NMA can be methylated, producing N,N-dimethylacetamide (DMA), which exhibits hepatotoxicity via methyltransferase and methane, in the living body. However, because no DMA was detected in rat tail surface gas within 80 min after injection of NMA solution into the abdomen, NMA was found not to be metabolised into DMA in the body. Therefore, it was determined that there was no potential for NMA was not methylated and did not produce hepatotoxic DMA in the living body