ID Number		1793
Study Type		In Vitro
Model		RF heating and conventional heating of chemical reactions and enzymatic digests of proteins (as used in proteomics protocols) demonstrate that previously reported non-thermal effects could not be confirmed.
Details		Chemical reactions evaluated included the following: Diels-Alder cycloaddition of 5-methoxycarbonyl-2 pyrone and phenylacetylene (prior reports of 64% conversion with 3 hr non-thermal RF exposure vs 19% without RF);nucleophilic substitution of benzyl chloride with triphenylphosphine (prior reports of 78% conversion with non-thermal RF vs 24% without RF); alkylation of triazole with 2.2.4-trichloroacetophenone (prior reports of different end-products with non-thermal RF vs % without RF); direct amidation of methacrylic acid with (R)-1-phenylethylamine (prior reports of different end-products with non-thermal RF vs % without RF). The authors developed a well controlled exposure system with stirring to insure evenly distributed temperature and no local hotspots. When the stirrer was on, they observed no effects and suggested prior reports were due to inhomogeneous thermal gradients. AUTHORS' ABSTRACT: Damm et al. 2012 (IEEE #5185): This study reevaluates the putative advantages of microwave-assisted tryptic digests compared to conventionally heated protocols performed at the same temperature. An initial investigation of enzyme stability in a temperature range of 3780 °C demonstrated that trypsin activity declines sharply at temperatures above 60 °C, regardless if microwave dielectric heating or conventional heating is employed. Tryptic digests of three proteins of different size (bovine serum albumin, cytochrome c and ²-casein) were thus performed at 37 °C and 50 °C using both microwave and conventional heating applying accurate internal fiber-optic probe reaction temperature measurements. The impact of the heating method on protein degradation and peptide fragment generation was analyzed by SDS-PAGE and MALDI-TOF-MS. Time-dependent tryptic digestion of the three proteins and subsequent analysis of the corresponding cleavage products by MALDI-TOF provided virtually identical results for both microwave and conventional heating. In addition, the impact of electro-magnetic field strength on the tertiary structure of trypsin and BSA was evaluated by molecular mechanics calculations. These simulations revealed that the applied field in a typical laboratory microwave reactor is 34 orders of magnitude too low to induce conformational changes in proteins or enzymes.
Findings		Effects (only at thermal levels)
Status		Completed With Publication
Principal Investigator
Funding Agency		Private/Instit.
Country		AUSTRIA
References		Herrero, MA et al. J Org Chem, (2008) 73:36-47 Damm , M et al. J. Proteomics., (2012) http://dx.doi.org/10.1016/j.jprot.2012.07.043:11 pages-
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