Limits...
Proton-Detected Solid-State NMR Spectroscopy of Bone with Ultrafast Magic Angle Spinning.

Mroue KH, Nishiyama Y, Kumar Pandey M, Gong B, McNerny E, Kohn DH, Morris MD, Ramamoorthy A - Sci Rep (2015)

Bottom Line: Our investigations demonstrate that two-dimensional (1)H/(1)H chemical shift correlation spectra for bone are obtainable using fp-RFDR (finite-pulse radio-frequency-driven dipolar recoupling) pulse sequence under ultrafast MAS.Our results infer that water exhibits distinct (1)H-(1)H dipolar coupling networks with the backbone and side-chain regions in collagen.These results show the promising potential of proton-detected ultrafast MAS NMR for monitoring structural and dynamic changes caused by mechanical loading and disease in bone.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biophysics, University of Michigan, Ann Arbor, Michigan, 48109-1055, United States [2] Department of Chemistry, University of Michigan, Ann Arbor, Michigan, 48109-1055, United States.

ABSTRACT
While obtaining high-resolution structural details from bone is highly important to better understand its mechanical strength and the effects of aging and disease on bone ultrastructure, it has been a major challenge to do so with existing biophysical techniques. Though solid-state NMR spectroscopy has the potential to reveal the structural details of bone, it suffers from poor spectral resolution and sensitivity. Nonetheless, recent developments in magic angle spinning (MAS) NMR technology have made it possible to spin solid samples up to 110 kHz frequency. With such remarkable capabilities, (1)H-detected NMR experiments that have traditionally been challenging on rigid solids can now be implemented. Here, we report the first application of multidimensional (1)H-detected NMR measurements on bone under ultrafast MAS conditions to provide atomistic-level elucidation of the complex heterogeneous structure of bone. Our investigations demonstrate that two-dimensional (1)H/(1)H chemical shift correlation spectra for bone are obtainable using fp-RFDR (finite-pulse radio-frequency-driven dipolar recoupling) pulse sequence under ultrafast MAS. Our results infer that water exhibits distinct (1)H-(1)H dipolar coupling networks with the backbone and side-chain regions in collagen. These results show the promising potential of proton-detected ultrafast MAS NMR for monitoring structural and dynamic changes caused by mechanical loading and disease in bone.

No MeSH data available.


Related in: MedlinePlus

1H MAS NMR spectra recorded at 14.1 T from bovine cortical bone under multiple MAS rates.The spectra were acquired by co-adding 4 transients with a rotor-synchronized spin- echo pulse sequence using a 0.75-mm MAS HX probe with a 0.72 μs 1H 90° pulse and a 3 s recycle delay.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4495383&req=5

f1: 1H MAS NMR spectra recorded at 14.1 T from bovine cortical bone under multiple MAS rates.The spectra were acquired by co-adding 4 transients with a rotor-synchronized spin- echo pulse sequence using a 0.75-mm MAS HX probe with a 0.72 μs 1H 90° pulse and a 3 s recycle delay.

Mentions: In this study, we systematically explore the benefits of employing ultrafast MAS frequencies in 1H-detected one- and two-dimensional NMR experiments for the purpose of enhancing the resolution and sensitivity of 1H MAS NMR spectra for bone. Experimental results obtained from powder bovine cortical bone samples are presented and interpreted, taking into account the complex heterogeneous structure and composition of bone, which result in complications arising from overlapping 1H NMR signals of the various organic and mineral constituents of bone. The 1H MAS spectra of bone recorded at multiple MAS frequencies up to 110 kHz MAS using a rotor-synchronized spin-echo sequence on a 600-MHz NMR spectrometer are presented in Fig. 1. Overall, we observe that increasing the MAS frequency produces a progressively better resolved and more sensitive spectral line shapes. At moderate-to-fast MAS rates (20–50 kHz), the spectrum exhibits only two peaks, with the water peak at ~5 ppm dominating the spectra. It is believed that water, the third major component in bone, plays a pivotal role in its biomechanical behavior, including stabilizing the matrix and contributing to its ductility. Water accounts for about 10% of fresh bone weight and occupies ~15–25% of its volume40. Nevertheless, studies have shown that the amount of water in bone decreases with age and with skeletal growth4142. Bone water is found in different forms with various binding conditions: free (or mobile) water filling the bulk of the microscopic pores in the calcified matrix, water bound to organic matrix mainly in the collagen network and organic-mineral interface, and water associated with the mineral phase434445. In addition to the strong water signal, the other broad peak centered at ~1.2 ppm originates from multiple overlapping signals that correspond to aliphatic side-chain protons in the bone organic matrix as well as to the structural OH groups in bone mineral. The line-narrowing advantage of MAS is clearly manifested in these spectra upon increasing the spinning frequency, where better resolution is obtained in the ultrafast MAS regime (νR ≥ 60 kHz). Here, a gradual resolution of the aliphatic region (0.5–3 ppm) is observed, where peaks from protons in the side chains of the amino acid residues in the organic matrix can now be observed. Notably, the broad peak centered at ~7.5 ppm in the (6–8 ppm) region, which comes from amide NH protons associated with the peptide bonds in collagen and which is almost invisible at lower MAS rates, becomes narrow and clearly more visible as the MAS frequency reaches its highest limits. Similarly, the peak at ~3.5 ppm, which is most likely due to α-protons of glycine (the most abundant amino acid residue in collagen), can only be resolved at ultrafast MAS rates. At 100 kHz MAS and beyond, the spectrum displays fairly resolved peaks for the structural hydroxyl group from the bone mineral, and for the side-chain protons from the organic matrix, as well as for the amide NH groups.


Proton-Detected Solid-State NMR Spectroscopy of Bone with Ultrafast Magic Angle Spinning.

Mroue KH, Nishiyama Y, Kumar Pandey M, Gong B, McNerny E, Kohn DH, Morris MD, Ramamoorthy A - Sci Rep (2015)

1H MAS NMR spectra recorded at 14.1 T from bovine cortical bone under multiple MAS rates.The spectra were acquired by co-adding 4 transients with a rotor-synchronized spin- echo pulse sequence using a 0.75-mm MAS HX probe with a 0.72 μs 1H 90° pulse and a 3 s recycle delay.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4495383&req=5

f1: 1H MAS NMR spectra recorded at 14.1 T from bovine cortical bone under multiple MAS rates.The spectra were acquired by co-adding 4 transients with a rotor-synchronized spin- echo pulse sequence using a 0.75-mm MAS HX probe with a 0.72 μs 1H 90° pulse and a 3 s recycle delay.
Mentions: In this study, we systematically explore the benefits of employing ultrafast MAS frequencies in 1H-detected one- and two-dimensional NMR experiments for the purpose of enhancing the resolution and sensitivity of 1H MAS NMR spectra for bone. Experimental results obtained from powder bovine cortical bone samples are presented and interpreted, taking into account the complex heterogeneous structure and composition of bone, which result in complications arising from overlapping 1H NMR signals of the various organic and mineral constituents of bone. The 1H MAS spectra of bone recorded at multiple MAS frequencies up to 110 kHz MAS using a rotor-synchronized spin-echo sequence on a 600-MHz NMR spectrometer are presented in Fig. 1. Overall, we observe that increasing the MAS frequency produces a progressively better resolved and more sensitive spectral line shapes. At moderate-to-fast MAS rates (20–50 kHz), the spectrum exhibits only two peaks, with the water peak at ~5 ppm dominating the spectra. It is believed that water, the third major component in bone, plays a pivotal role in its biomechanical behavior, including stabilizing the matrix and contributing to its ductility. Water accounts for about 10% of fresh bone weight and occupies ~15–25% of its volume40. Nevertheless, studies have shown that the amount of water in bone decreases with age and with skeletal growth4142. Bone water is found in different forms with various binding conditions: free (or mobile) water filling the bulk of the microscopic pores in the calcified matrix, water bound to organic matrix mainly in the collagen network and organic-mineral interface, and water associated with the mineral phase434445. In addition to the strong water signal, the other broad peak centered at ~1.2 ppm originates from multiple overlapping signals that correspond to aliphatic side-chain protons in the bone organic matrix as well as to the structural OH groups in bone mineral. The line-narrowing advantage of MAS is clearly manifested in these spectra upon increasing the spinning frequency, where better resolution is obtained in the ultrafast MAS regime (νR ≥ 60 kHz). Here, a gradual resolution of the aliphatic region (0.5–3 ppm) is observed, where peaks from protons in the side chains of the amino acid residues in the organic matrix can now be observed. Notably, the broad peak centered at ~7.5 ppm in the (6–8 ppm) region, which comes from amide NH protons associated with the peptide bonds in collagen and which is almost invisible at lower MAS rates, becomes narrow and clearly more visible as the MAS frequency reaches its highest limits. Similarly, the peak at ~3.5 ppm, which is most likely due to α-protons of glycine (the most abundant amino acid residue in collagen), can only be resolved at ultrafast MAS rates. At 100 kHz MAS and beyond, the spectrum displays fairly resolved peaks for the structural hydroxyl group from the bone mineral, and for the side-chain protons from the organic matrix, as well as for the amide NH groups.

Bottom Line: Our investigations demonstrate that two-dimensional (1)H/(1)H chemical shift correlation spectra for bone are obtainable using fp-RFDR (finite-pulse radio-frequency-driven dipolar recoupling) pulse sequence under ultrafast MAS.Our results infer that water exhibits distinct (1)H-(1)H dipolar coupling networks with the backbone and side-chain regions in collagen.These results show the promising potential of proton-detected ultrafast MAS NMR for monitoring structural and dynamic changes caused by mechanical loading and disease in bone.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biophysics, University of Michigan, Ann Arbor, Michigan, 48109-1055, United States [2] Department of Chemistry, University of Michigan, Ann Arbor, Michigan, 48109-1055, United States.

ABSTRACT
While obtaining high-resolution structural details from bone is highly important to better understand its mechanical strength and the effects of aging and disease on bone ultrastructure, it has been a major challenge to do so with existing biophysical techniques. Though solid-state NMR spectroscopy has the potential to reveal the structural details of bone, it suffers from poor spectral resolution and sensitivity. Nonetheless, recent developments in magic angle spinning (MAS) NMR technology have made it possible to spin solid samples up to 110 kHz frequency. With such remarkable capabilities, (1)H-detected NMR experiments that have traditionally been challenging on rigid solids can now be implemented. Here, we report the first application of multidimensional (1)H-detected NMR measurements on bone under ultrafast MAS conditions to provide atomistic-level elucidation of the complex heterogeneous structure of bone. Our investigations demonstrate that two-dimensional (1)H/(1)H chemical shift correlation spectra for bone are obtainable using fp-RFDR (finite-pulse radio-frequency-driven dipolar recoupling) pulse sequence under ultrafast MAS. Our results infer that water exhibits distinct (1)H-(1)H dipolar coupling networks with the backbone and side-chain regions in collagen. These results show the promising potential of proton-detected ultrafast MAS NMR for monitoring structural and dynamic changes caused by mechanical loading and disease in bone.

No MeSH data available.


Related in: MedlinePlus