In vivo comprehensive multiphase NMR

Abstract

Traditionally, due to different hardware requirements, nuclear magnetic resonance (NMR) has developed as two separate fields: one dealing with solids, and one with solutions. Comprehensive multiphase (CMP) NMR combines all electronics and hardware (magic angle spinning [MAS], gradients, high power Radio Frequency (RF) handling, lock, susceptibility matching) into a universal probe that permits a comprehensive study of all phases (i.e., liquid, gel‐like, semisolid, and solid), in intact samples. When applied in vivo, it provides unique insight into the wide array of bonds in a living system from the most mobile liquids (blood, fluids) through gels (muscle, tissues) to the most rigid (exoskeleton, shell). In this tutorial, the practical aspects of in vivo CMP NMR are discussed including: handling the organisms, rotor preparation, sample spinning, water suppression, editing experiments, and finishes with a brief look at the potential of other heteronuclei (2H, 15N, 19F, 31P) for in vivo research. The tutorial is aimed as a general resource for researchers interested in developing and applying MAS‐based approaches to living organisms. Although the focus here is CMP NMR, many of the approaches can be adapted (or directly applied) using conventional high‐resolution magic angle spinning, and in some cases, even standard solid‐state NMR probes.

Comprehensive Multiphase NMR Probehead with Reduced Radiofrequency Heating Improves the Analysis of Living Organisms and Heat-Sensitive Samples

“…In this study, all samples were actively cooled to 5 °C. Such a temperature has shown to increase the survivability of living organisms in previous studies ,, and is commonly used for the analysis of biological samples, in general, to reduce sample degradation during analysis. , Table shows an ∼11 °C temperature increase with the E-free design, resulting in ∼16 °C (5 °C + 11 °C) in the freshwater shrimp tissue. This is within the ranges of the freshwater environments where the shrimp reside in nature. , Conversely, the heating from the solenoid probe increases the temperature such that the shrimp tissue will reach ∼39 °C (5 °C + 34 °C) clearly above the level at which the organisms can normally survive …”

“…CMP-NMR was recently applied to living organisms and provides unrivaled information on real-time responses and structural information in vivo . − ,, It is of utmost importance that the organisms under investigation survive the duration of all experiments. As CMP-NMR involves the use of editing experiments to differentiate and identify components in different phases, selecting solid-phase components (i.e., via CP with high-power decoupling) is a requirement.…”

“…In this study, all samples were actively cooled to 5 °C. Such a temperature has shown to increase the survivability of living organisms in previous studies 12,14,15 and is commonly used for the analysis of biological samples, in general, to reduce sample degradation during analysis. 33,34 Table 2 shows an ∼11 °C temperature increase with the E-free design, resulting in ∼16 °C (5 °C + 11 °C) in the freshwater shrimp tissue.…”

“…CP spectra were collected on individual in vivo shrimps ( 13 C-enriched) on each probe (Figure B). The experimental parameters were identical to previous in vivo CMP studies except for decoupling bandwidth, which was increased from 50 to 100 kHz. , Each CP experiment collected 30 720 scans (∼4 h), and a total of six shrimps were used for each probe. The MAS rate was 2500 Hz, which is the maximum speed a living organism can be spun without detrimental impacts …”

“…Because of its unique capabilities, CMP-NMR has found applications in a range of fields, including solving the structure of swollen soil, understanding oil spill contamination, following contaminant sequestration, engine part swelling, biofuel production, mineral–organic material interactions, seed germination, , and plant growth . It has even been shown that when applied to living organismsDaphnia magna (freshwater fleas) andHyalella azteca (freshwater shrimp), all components in all phases (liquids, gels, and solids) can be detected in vivo , giving a unique and unprecedented insight into the living molecular world. − …”