A NMR method was implemented to assess 'in vivo' oxygenation levels by a quantitative determination of the (1)H-NMR resonances of Mb: the proximal His-F8 N(delta)H at 70-90 ppm and Val-E11 gammaCH(3) resonance at -2.8 ppm, reflecting deoxygenated (deoxy-Mb) and oxygenated (met-Mb) states, were alternately recorded. The method was developed 'in vitro' choosing a couple of NMR sequences that could each maximize the SNR, while avoiding baseline rolling and suppressing the water signal. Two quantitative calibration methods were implemented for deoxy- and met-Mb samples (0.1 -1mM), respectively. LOD and LOQ resulted 0.015 mM and 0.05 mM for MetMb and 0.013 mM and 0.042 mM for deoxy-Mb. Sequences and calibration curves were employed 'in vivo' in Arenicola Marina, to obtain, for the first time, an accurate measurement of oxy and deoxy-Mb actual concentrations. In Arenicola, the peaks at about 87 ppm and -2.7 ppm, reflecting the deoxy and oxy-Mb states, respectively, were alternately recorded during increasing hypoxia. The deoxy-Mb concentrations were obtained from the calibration curve. The oxy-Mb concentrations were calculated from the calibration of met-Mb, as it was proved that oxy and met-Mb gave the same NMR molar response. From oxy and deoxy-Mb concentrations, the PiO(2) trend was determined
A quantitative method to assess muscle tissue oxygenation 'in vivo' by monitoring (1)H-NMR myoglobin resonances.
Vezzoli A;
2010
Abstract
A NMR method was implemented to assess 'in vivo' oxygenation levels by a quantitative determination of the (1)H-NMR resonances of Mb: the proximal His-F8 N(delta)H at 70-90 ppm and Val-E11 gammaCH(3) resonance at -2.8 ppm, reflecting deoxygenated (deoxy-Mb) and oxygenated (met-Mb) states, were alternately recorded. The method was developed 'in vitro' choosing a couple of NMR sequences that could each maximize the SNR, while avoiding baseline rolling and suppressing the water signal. Two quantitative calibration methods were implemented for deoxy- and met-Mb samples (0.1 -1mM), respectively. LOD and LOQ resulted 0.015 mM and 0.05 mM for MetMb and 0.013 mM and 0.042 mM for deoxy-Mb. Sequences and calibration curves were employed 'in vivo' in Arenicola Marina, to obtain, for the first time, an accurate measurement of oxy and deoxy-Mb actual concentrations. In Arenicola, the peaks at about 87 ppm and -2.7 ppm, reflecting the deoxy and oxy-Mb states, respectively, were alternately recorded during increasing hypoxia. The deoxy-Mb concentrations were obtained from the calibration curve. The oxy-Mb concentrations were calculated from the calibration of met-Mb, as it was proved that oxy and met-Mb gave the same NMR molar response. From oxy and deoxy-Mb concentrations, the PiO(2) trend was determinedI documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
