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Measures of Bone Mineral Carbonate Content and Mineral Maturity/Crystallinity for FT-IR and Raman Spectroscopic Imaging Differentially Relate to Physical–Chemical Properties of Carbonate-Substituted Hydroxyapatite

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Abstract

Bone mineral carbonate content assessed by vibrational spectroscopy relates to fracture incidence, and mineral maturity/ crystallinity (MMC) relates to tissue age. As FT-IR and Raman spectroscopy become more widely used to characterize the chemical composition of bone in pre-clinical and translational studies, their bone mineral outcomes require improved validation to inform interpretation of spectroscopic data. In this study, our objectives were (1) to relate Raman and FT-IR carbonate:phosphate ratios calculated through direct integration of peaks to gold-standard analytical measures of carbonate content and underlying subband ratios; (2) to relate Raman and FT-IR MMC measures to gold-standard analytical measures of crystal size in chemical standards and native bone powders. Raman and FT-IR direct integration carbonate:phosphate ratios increased with carbonate content (Raman: p < 0.01, R2 = 0.87; FT-IR: p < 0.01, R2 = 0.96) and Raman was more sensitive to carbonate content than the FT-IR (Raman slope + 95% vs FT-IR slope, p < 0.01). MMC increased with crystal size for both Raman and FT-IR (Raman: p < 0.01, R2 = 0.76; FT-IR p < 0.01, R2 = 0.73) and FT-IR was more sensitive to crystal size than Raman (c-axis length: slope FT-IR MMC + 111% vs Raman MMC, p < 0.01). Additionally, FT-IR but not Raman spectroscopy detected differences in the relationship between MMC and crystal size of carbonated hydroxyapatite (CHA) vs poorly crystalline hydroxyapatites (HA) (slope CHA + 87% vs HA, p < 0.01). Combined, these results contribute to the ability of future studies to elucidate the relationships between carbonate content and fracture and provide insight to the strengths and limitations of FT-IR and Raman spectroscopy of native bone mineral.

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References

  1. Taylor EA, Donnelly E (2020) Raman and Fourier transform infrared imaging for characterization of bone material properties. Bone 139:115490. https://doi.org/10.1016/j.bone.2020.115490

    Article  CAS  PubMed  Google Scholar 

  2. Boskey A, Mendelsohn R (2005) Infrared analysis of bone in health and disease. J Biomed Opt 10:31102. https://doi.org/10.1117/1.1922927

    Article  CAS  Google Scholar 

  3. Paschalis EP, Mendelsohn R, Boskey AL (2011) Infrared assessment of bone quality: A review. ClinOrthopRelat Res 469:2170–2178. https://doi.org/10.1007/s11999-010-1751-4

    Article  Google Scholar 

  4. Boskey AL, Donnelly E, Boskey E et al (2016) Examining the relationships between bone tissue composition, compositional heterogeneity, and fragility fracture: a matched case-controlled FTIRI study. J Bone Miner Res 31:1070–1081. https://doi.org/10.1002/jbmr.2759

    Article  CAS  PubMed  Google Scholar 

  5. Hunt HB, Donnelly E (2016) Bone quality assessment techniques: geometric, compositional, and mechanical characterization from macroscale to nanoscale. Clin Rev Bone Miner Metab 14:1–17. https://doi.org/10.1007/s12018-016-9222-4

    Article  Google Scholar 

  6. Mandair GS, Morris MD (2015) Contributions of Raman spectroscopy to the understanding of bone strength. BoneKEy 4:1–8. https://doi.org/10.1038/bonekey.2014.115

    Article  CAS  Google Scholar 

  7. Boskey A, Mendelsohn R (2005) Infrared analysis of bone in health and disease. J Biomed Opt 10:031102. https://doi.org/10.1117/1.1922927

    Article  CAS  PubMed  Google Scholar 

  8. Donnelly E, Boskey AL, Baker SP, van der Meulen MCH (2010) Effects of tissue age on bone tissue material composition and nanomechanical properties in the rat cortex. J Biomed Mater Res A 92:1048–1056. https://doi.org/10.1002/jbm.a.32442

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Bi X, Patil CA, Lynch CC et al (2011) Raman and mechanical properties correlate at whole bone- and tissue-levels in a genetic mouse model. J Biomech 44:297–303. https://doi.org/10.1016/j.jbiomech.2010.10.009

    Article  PubMed  Google Scholar 

  10. Lloyd AA, Gludovatz B, Riedel C et al (2017) Atypical fracture with long-term bisphosphonate therapy is associated with altered cortical composition and reduced fracture resistance. ProcNatlAcadSci 114:201704460. https://doi.org/10.1073/pnas.1704460114

    Article  CAS  Google Scholar 

  11. Burket J, Gourion-arsiquaud S, Havill LM et al (2011) Microstructure and nanomechanical properties in osteons relate to tissue and animal age. J Biomech 44:277–284. https://doi.org/10.1016/j.jbiomech.2010.10.018

    Article  PubMed  Google Scholar 

  12. Gourion-Arsiquaud S, Burket JC, Havill LM et al (2009) Spatial variation in osteonal bone properties relative to tissue and animal age. J Bone Miner Res 24:1271–1281. https://doi.org/10.1359/jbmr.090201

    Article  PubMed  PubMed Central  Google Scholar 

  13. Miller LM, Vairavamurthy V, Chance MR et al (2001) In situ analysis of mineral content and crystallinity in bone using infrared micro-spectroscopy of the nu(4) PO(4)(3-) vibration. BiochimBiophysActa 1527:11–19. https://doi.org/10.1016/S0304-4165(01)00093-9

    Article  CAS  Google Scholar 

  14. Mendelsohn R, Paschalis EP, Boskey AL (1999) Infrared spectroscopy, microscopy, and microscopic imaging of mineralizing tissues: spectra-structure correlations from human iliac crest biopsies. J Biomed Opt 4:14–21

    Article  CAS  Google Scholar 

  15. Boskey AL, DiCarlo E, Paschalis E et al (2005) Comparison of mineral quality and quantity in iliac crest biopsies from high- and low-turnover osteoporosis: An FT-IR microspectroscopic investigation. OsteoporosInt 16:2031–2038. https://doi.org/10.1007/s00198-005-1992-3

    Article  CAS  Google Scholar 

  16. Ou-Yang H, Paschalis EP, Mayo WE et al (2001) Infrared microscopic imaging of bone: spatial distribution of CO3(2-). J Bone Miner Res 16:893–900. https://doi.org/10.1359/jbmr.2001.16.5.893

    Article  CAS  PubMed  Google Scholar 

  17. Imbert L, Gourion-Arsiquaud S, Villarreal-Ramirez E et al (2018) Dynamic structure and composition of bone investigated by nanoscale infrared spectroscopy. PLoS ONE 13:1–15. https://doi.org/10.1371/journal.pone.0202833

    Article  CAS  Google Scholar 

  18. Cowin SC, Cardoso L (2015) Blood and Interstitial flow in the hierarchical pore space architecture of bone tissue. J Biomech 48:842–854. https://doi.org/10.1016/j.jbiomech.2014.12.013.Blood

    Article  PubMed  Google Scholar 

  19. Vahidi G, Rux C, Sherk VD, Heveran CM (2020) Lacunar-canalicular bone remodeling: Impacts on bone quality and tools for assessment. Bone. https://doi.org/10.1016/j.bone.2020.115663

    Article  PubMed  Google Scholar 

  20. Väänänen HK, Laitala-Leinonen T (2008) Osteoclast lineage and function. Arch BiochemBiophys 473:132–138. https://doi.org/10.1016/j.abb.2008.03.037

    Article  CAS  Google Scholar 

  21. Rey C, Combes C, Drouet C, Glimcher M (2010) Bone mineral: update on chemical composition and structure. OsteoporosInt 20:1013–1021. https://doi.org/10.1007/s00198-009-0860-y.Bone

    Article  Google Scholar 

  22. Franco W (1994) Mineral, Synthetic and Biological Carbonate Apatites. Stud InorgChem 18:191–304. https://doi.org/10.1016/B978-0-444-81582-8.50009-2

    Article  Google Scholar 

  23. Penel G, Leroy G, Rey C, Bres E (1998) MicroRaman spectral study of the PO4 and CO3 vibrational modes in synthetic and biological apatites. Calcif Tissue Int 63:475–481. https://doi.org/10.1007/s002239900561

    Article  CAS  PubMed  Google Scholar 

  24. Awonusi A, Morris MD, Tecklenburg MMJ (2007) Carbonate assignment and calibration in the Raman spectrum of apatite. Calcif Tissue Int 81:46–52. https://doi.org/10.1007/s00223-007-9034-0

    Article  CAS  PubMed  Google Scholar 

  25. Barth A, Zscherp C (2002) What vibrations tell us about proteins. Q Rev Biophys 35:369–430. https://doi.org/10.1017/S0033583502003815

    Article  CAS  PubMed  Google Scholar 

  26. Baldassarre M, Li C, Eremina N et al (2015) Simultaneous fitting of absorption spectra and their second derivatives for an improved analysis of protein infrared spectra. Molecules 20:12599–12622. https://doi.org/10.3390/molecules200712599

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Grunenwald A, Keyser C, Sautereau AM et al (2014) Revisiting carbonate quantification in apatite (bio)minerals: A validated FTIR methodology. J ArchaeolSci 49:134–141. https://doi.org/10.1016/j.jas.2014.05.004

    Article  CAS  Google Scholar 

  28. Yerramshetty JS, Lind C, Akkus O (2006) The compositional and physicochemical homogeneity of male femoral cortex increases after the sixth decade. Bone 39:1236–1243. https://doi.org/10.1016/j.bone.2006.06.002

    Article  CAS  PubMed  Google Scholar 

  29. Akkus O, Polyakova-Akkus A, Adar F, Schaffler MB (2003) Aging of microstructural compartments in human compact bone. J Bone Miner Res 18:1012–1019. https://doi.org/10.1359/jbmr.2003.18.6.1012

    Article  CAS  PubMed  Google Scholar 

  30. Kavukcuoglu NB, Arteaga-Solis E, Lee-Arteaga S et al (2007) Nanomechanics and Raman spectroscopy of fibrillin 2 knock-out mouse bones. J Mater Sci 42:8788–8794. https://doi.org/10.1007/s10853-007-1918-x

    Article  CAS  Google Scholar 

  31. Donnelly E, Meredith DS, Nguyen JT et al (2012) Reduced cortical bone compositional heterogeneity with bisphosphonate treatment in postmenopausal women with intertrochanteric and subtrochanteric fractures. J Bone Miner Res 27:672–678. https://doi.org/10.1002/jbmr.560

    Article  CAS  PubMed  Google Scholar 

  32. Gourion-Arsiquaud S, Faibish D, Myers E et al (2009) Use of FTIR spectroscopic imaging to identify parameters associated with fragility fracture. J Bone Miner Res 24:1565–1571. https://doi.org/10.1359/JBMR.090414

    Article  PubMed  PubMed Central  Google Scholar 

  33. Schmidt FN, Zimmermann EA, Campbell GM et al (2017) Assessment of collagen quality associated with non-enzymatic cross-links in human bone using Fourier-transform infrared imaging. Bone 97:243–251. https://doi.org/10.1016/j.bone.2017.01.015

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Faibish D, Gomes A, Boivin G et al (2005) Infrared imaging of calcified tissue in bone biopsies from adults with osteomalacia. Bone 36:6–12. https://doi.org/10.1016/j.bone.2004.08.019

    Article  CAS  PubMed  Google Scholar 

  35. Farlay D, Panczer G, Rey C et al (2010) Mineral maturity and crystallinity index are distinct characteristics of bone mineral. J Bone Miner Metab 28:433–445. https://doi.org/10.1007/s00774-009-0146-7

    Article  PubMed  PubMed Central  Google Scholar 

  36. Gadaleta SJ, Paschalis EP, Betts F et al (1996) Fourier transform infrared spectroscopy of the solution mediated conversion of amorphous calcium phosphate to hydroxyapatite: new correlations between X-ray diffraction and infrared data. Calcif Tissue Int 58:9–16

    Article  CAS  Google Scholar 

  37. Baddiel CB, Berry EE (1966) Spectra structure correlations in hydroxy and fluorapatite. SpectrochimActa 22:1407–1416. https://doi.org/10.1016/0371-1951(66)80133-9

    Article  CAS  Google Scholar 

  38. Ou-Yang H, Paschalis EP, Boskey AL, Mendelsohn R (2000) Two-dimensional vibrational correlation spectroscopy of in vitro hydroxyapatite maturation. Biopolym - Biospectroscopy Sect 57:129–139. https://doi.org/10.1002/(SICI)1097-0282(2000)57:3%3c129::AID-BIP1%3e3.0.CO;2-O

    Article  CAS  Google Scholar 

  39. Colthup NP, Daly LH, Wiberly SE (1990) Introduction to Infrared and Raman Spectroscopy, 3rd edn. Elsevier Science, New York, NY

    Google Scholar 

  40. Pleshko N, Boskey A, Mendelsohn R (1991) Novel infrared spectroscopic method for the determination of crystallinity of hydroxyapatite minerals. Biophys J 60:786–793

    Article  CAS  Google Scholar 

  41. Kazanci M, Fratzl P, Klaushofer K, Paschalis EP (2006) Complementary information on in vitro conversion of amorphous (precursor) calcium phosphate to hydroxyapatite from ramanmicrospectroscopy and wide-angle X-ray scattering. Calcif Tissue Int 79:354–359. https://doi.org/10.1007/s00223-006-0011-9

    Article  CAS  PubMed  Google Scholar 

  42. Morris MD, Mandair GS (2011) Raman assessment of bone quality. ClinOrthopRelat Res 469:2160–2169. https://doi.org/10.1007/s11999-010-1692-y

    Article  Google Scholar 

  43. Paschalis EP, Jacenko O, Olsen B et al (1996) Fourier transform infrared microspectroscopic analysis identifies alterations in mineral properties in bones from mice transgenic for type X collagen. Bone 19:151–156. https://doi.org/10.1016/8756-3282(96)00164-0

    Article  CAS  PubMed  Google Scholar 

  44. Camacho NP, Rinnerthaler S, Paschalis EP et al (1999) Complementary information on bone ultrastructure from scanning small angle X-ray scattering and Fourier-transform infrared microspectroscopy. Bone 25:287–293. https://doi.org/10.1016/S8756-3282(99)00165-9

    Article  CAS  PubMed  Google Scholar 

  45. Spevak L, Flach CR, Hunter T et al (2013) Fourier transform infrared spectroscopic imaging parameters describing acid phosphate substitution in biologic hydroxyapatite. Calcif Tissue Int 92:418–428. https://doi.org/10.1007/s00223-013-9695-9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Freeman JJ, Wopenka B, Silva MJ, Pasteris JD (2001) Raman spectroscopic detection of changes in bioapatite in mouse femora as a function of age and in vitro fluoride treatment. Calcif Tissue Int 68:156–162. https://doi.org/10.1007/s002230001206

    Article  CAS  PubMed  Google Scholar 

  47. Wallace JM, Golcuk K, Morris MD, Kohn DH (2009) Inbred strain-specific response to biglycan deficiency in the cortical bone of C57BL6/129 and C3H/He mice. J Bone Miner Res 24:1002–1012. https://doi.org/10.1359/jbmr.081259

    Article  PubMed  Google Scholar 

  48. Huang RY, Miller LM, Carlson CS, Chance MR (2002) Characterization of bone mineral composition in the proximal tibia of cynomolgus monkeys: Effect of ovariectomy and nandrolonedecanoate treatment. Bone 30:492–497. https://doi.org/10.1016/S8756-3282(01)00691-3

    Article  CAS  PubMed  Google Scholar 

  49. Donnelly E, Saleh A, Unnanuntana A, Lane JM (2012) Atypical Femoral Fractures Epidemiology, Etiology, and Patient Management. CurrOpin Support Palliat Care. https://doi.org/10.14440/jbm.2015.54.A

    Article  Google Scholar 

  50. Yerramshetty JS, Akkus O (2008) The associations between mineral crystallinity and the mechanical properties of human cortical bone. Bone 42:476–482. https://doi.org/10.1016/j.bone.2007.12.001

    Article  CAS  PubMed  Google Scholar 

  51. Cox SC, Jamshidi P, Grover LM, Mallick KK (2014) Preparation and characterisation of nanophaseSr, Mg, and Zn substituted hydroxyapatite by aqueous precipitation. Mater SciEng C 35:106–114. https://doi.org/10.1016/j.msec.2013.10.015

    Article  CAS  Google Scholar 

  52. Taylor EA, Lloyd AA, Salazar-Lara C, Donnelly EL (2017) Raman and FT-IR mineral to matrix ratios correlate with physical chemical properties of model compounds and native bone tissue. ApplSpectrosc. https://doi.org/10.1177/0003702817709286

    Article  Google Scholar 

  53. Su FY, Pang S, Ling YTT et al (2018) Deproteinization of cortical bone: effects of different treatments. Calcif Tissue Int. https://doi.org/10.1007/s00223-018-0453-x

    Article  PubMed  Google Scholar 

  54. Hunt HB, Torres AM, Palomino PM et al (2019) Altered tissue composition, microarchitecture, and mechanical performance in cancellous bone from men with type 2 diabetes mellitus. J Bone Miner Res 34:1191–1206. https://doi.org/10.1002/jbmr.3711

    Article  CAS  PubMed  Google Scholar 

  55. Nyman JS, Makowski AJ, Patil CA et al (2011) Measuring differences in compositional properties of bone tissue by confocal raman spectroscopy. Calcif Tissue Int 89:111–122. https://doi.org/10.1007/s00223-011-9497-x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Gamsjaeger S, Masic A, Roschger P et al (2010) Cortical bone composition and orientation as a function of animal and tissue age in mice by Raman spectroscopy. Bone 47:392–399. https://doi.org/10.1016/j.bone.2010.04.608

    Article  PubMed  Google Scholar 

  57. Gamsjaeger S, Buchinger B, Zwettler E et al (2011) Bone material properties in actively bone-forming trabeculae in postmenopausal women with osteoporosis after three years of treatment with once-yearly Zoledronic acid. J Bone Miner Res 26:12–18. https://doi.org/10.1002/jbmr.180

    Article  CAS  PubMed  Google Scholar 

  58. Unal M, Uppuganti S, Leverant CJ et al (2018) Assessing glycation-mediated changes in human cortical bone with Raman spectroscopy. J Biophotonics. https://doi.org/10.1002/jbio.201700352

    Article  PubMed  PubMed Central  Google Scholar 

  59. Cullity B, Stock S (2001) Elements of X-Ray Diffraction, 3rd edn. Prentice Hall, Upper Saddle River, NJ

    Google Scholar 

  60. Venkateswarlu K, Sandhyarani M, Nellaippan TA, Rameshbabu N (2014) Estimation of crystallite size, lattice strain and dislocation density of nanocrystalline carbonate substituted hydroxyapatite by X-ray peak variance analysis. Procedia Mater Sci 5:212–221. https://doi.org/10.1016/j.mspro.2014.07.260

    Article  CAS  Google Scholar 

  61. Pasteris JD, Wopenka B, Freeman JJ et al (2004) Lack of OH in nanocrystalline apatite as a function of degree of atomic order: Implications for bone and biomaterials. Biomaterials 25:229–238. https://doi.org/10.1016/S0142-9612(03)00487-3

    Article  CAS  PubMed  Google Scholar 

  62. Kazanci M, Roschger P, Paschalis EP et al (2006) Bone osteonal tissues by Raman spectral mapping: Orientation-composition. J StructBiol 156:489–496. https://doi.org/10.1016/j.jsb.2006.06.011

    Article  CAS  Google Scholar 

  63. Kazanci M, Wagner HD, Manjubala NI et al (2007) Raman imaging of two orthogonal planes within cortical bone. Bone 41:456–461. https://doi.org/10.1016/j.bone.2007.04.200

    Article  CAS  PubMed  Google Scholar 

  64. Schulmerich MV, Cole JH, Kreider JM et al (2009) Transcutaneous raman spectroscopy of murine bone in vivo. ApplSpectrosc 63:286–295. https://doi.org/10.1366/000370209787599013

    Article  CAS  Google Scholar 

  65. Peterson JR, Okagbare PI, De La Rosa S et al (2013) Early detection of burn induced heterotopic ossification using transcutaneous Raman spectroscopy. Bone 54:28–34. https://doi.org/10.1016/j.bone.2013.01.002

    Article  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors thank Phil Carubia and Christopher Umbach for their assistance with collecting FT-IR, XRD, and Raman data; Dr. Michael D. Morris for helpful discussions; Jennie AMR Kunitake for assistance with HA synthesis and for helpful discussions; Dr. Lynn Johnson and the Cornell Statistical Consulting Unit for assistance with data analysis.

Funding

This material is based upon work supported by the National Science Foundation under Grant No. CMMI 1452852. This work made use of the Cornell Center for Materials Research Shared Facilities, which are supported through the NSF MRSEC program (DMR-1719875).

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  1. Sibley School of Mechanical and Aerospace Engineering, Cornell University, Ithaca, NY, 14853, USA

    Erik A. Taylor

  2. Nancy E. and Peter C. Meinig School of Biomedical Engineering, Cornell University, Ithaca, NY, 14853, USA

    Cassidy J. Mileti

  3. Department of Materials Science Engineering, Cornell University, 227 Bard Hall, Ithaca, NY, 14853, USA

    Sandhya Ganesan, Joo Ho Kim & Eve Donnelly

  4. Research Division, Hospital for Special Surgery, New York, NY, 10021, USA

    Eve Donnelly

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Taylor, E.A., Mileti, C.J., Ganesan, S. et al. Measures of Bone Mineral Carbonate Content and Mineral Maturity/Crystallinity for FT-IR and Raman Spectroscopic Imaging Differentially Relate to Physical–Chemical Properties of Carbonate-Substituted Hydroxyapatite. Calcif Tissue Int 109, 77–91 (2021). https://doi.org/10.1007/s00223-021-00825-4

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