High-resolution magic angle spinning (1)H NMR spectroscopy of intact liver and kidney: optimization of sample preparation procedures and biochemical stability of tissue during spectral acquisition.
High-resolution magic angle spinning (MAS) (1)H NMR spectroscopy has been used to investigate the biochemical composition of whole rat renal cortex and liver tissue samples. The effects of a number of sample preparation procedures and experimental variables have been investigated systematically in order to optimize spectral quality and maximize information recovery. These variables include the effects of changing the sample volume in the MAS rotor, snap-freezing the samples, and the effect of organ perfusion with deuterated saline solution prior to MAS NMR analysis. Also, the overall biochemical stability of liver and kidney tissue MAS NMR spectra was investigated under different temperature conditions. We demonstrate improved resolution and line shape of MAS NMR spectra obtained from small spherical tissue volume (12 microl) rotor inserts compared to 65 microl cylindrical samples directly inserted into the MAS rotors. D(2)O saline perfusion of the in situ afferent vascular tree of the tissue immediately postmortem also improves line shape in MAS NMR spectra. Snap-freezing resulted in increased signal intensities from alpha-amino acids (e.g., valine) in tissue together with decreases in renal osmolytes, such as myo-inositol. A decrease in triglyceride levels was observed in renal cortex following stasis on ice and in the MAS rotor (303 K for 4 h). This work indicates that different tissues have differential metabolic stabilities in (1)H MAS NMR experiments and that careful attention to sample preparation is required to minimize artifacts and maintain spectral quality.
Waters
N J
NJ
Biological Chemistry, Imperial College, London, SW7 2AZ, United Kingdom.
Garrod
S
S
Farrant
R D
RD
Haselden
J N
JN
Connor
S C
SC
Connelly
J
J
Lindon
J C
JC
Holmes
E
E
Nicholson
J K
JK
D000818:Animals; D003903:Deuterium / Q000737:chemistry; D005615:Freezing; D007668:Kidney / Q000033:anatomy & histology* / Q000737:chemistry; D007672:Kidney Cortex / Q000033:anatomy & histology / Q000737:chemistry; D008099:Liver / Q000033:anatomy & histology* / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000295:instrumentation* / Q000379:methods*; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D014020:Tissue Extracts / Q000737:chemistry
D014020:Tissue Extracts; D003903:Deuterium