[NMR paper] A novel microfluidic rapid freeze-quench device for trapping reactions intermediates for high field EPR analysis

[nmrlearner]

A novel microfluidic rapid freeze-quench device for trapping reactions intermediates for high field EPR analysis

Available online 5 March 2013
Publication year: 2013
Source:Journal of Magnetic Resonance



Rapid freeze quench electron paramagnetic resonance (RFQ)-EPR is a method for trapping short lived intermediates in chemical reactions and subjecting them to EPR spectroscopy investigation for their characterization. Two (or more) reacting components are mixed at room temperature and after some delay the mixture is sprayed into a cold trap and transferred into the EPR tube. A major caveat in using commercial RFQ-EPR for high field EPR applications is the relatively large amount of sample needed for each time point, a major part of which is wasted as the dead volume of the instrument. The small sample volume (~2?l) needed for high field EPR spectrometers, such as W-band (~3.5T, 95GHz), that use cavities calls for the development of a microfluidic based RFQ-EPR apparatus. This is particularly important for biological applications because of the difficulties often encountered in producing large amounts of intrinsically paramagnetic proteins and spin labeled nucleic acid and proteins. Here we describe a dedicated microfluidic based RFQ-EPR apparatus suitable for small volume samples in the range of a few ?l. The device is based on a previously published microfluidic mixer and features a new ejection mechanism and a novel cold trap that allows collection of a series of different time points in one continuous experiment. The reduction of a nitroxide radical with dithionite, employing the signal of Mn2+ as an internal standard was used to demonstrate the performance of the microfluidic RFQ apparatus.
Graphical abstract

Highlights

? A microfluidic rapid freeze quench apparatus was developed for W-band EPR. ? It features new ejection and freezing approaches. ? All samples are collected continuously. ? 100–150?l are needed for collecting 7 time points in triplicates. ? The estimated deadtime is below 5ms.





More...