Research

Fourier Transform Infrared (FTIR) spectroscopy

Fourier Transform Infrared (FTIR) spectroscopy is a widely used technique in material sciences for characterization of chemical composition of solids, liquids and gases. Infrared light is passed through the sample in a transmission mode measurement. Part of the infrared energy is absorbed vibrations and rotations of the molecules of the specimen. Vibration frequencies depend on the chemical bonds and the structure of the molecule. An infrared absorption spectrum is also called a chemical fingerprint of the molecule as every compound has a unique infrared spectrum. Therefore, detailed information about the chemical composition of the specimen can be obtained through the analysis of its infrared absorption spectrum.

FTIR spectroscopic imaging enables imaging of spatial composition from thin histological sections by combining traditional FTIR spectrometer with a microscope. FTIR spectroscopic imaging has been used in bone [1-5] and cartilage [6-9] research in quantitative analysis (collagen, proteoglycans, hydroxyapatite, cross-links). Qualitative analysis is also possible e.g. by utilizing cluster analysis techniques. These techniques have been applied to subchondral bone [10] and cartilage [11].

Raman spectroscopy

Raman spectroscopy is closely related to FTIR spectroscopy. A monochromatic light from a laser source is focused on the sample and the scattered light is investigated. The most of the light undergo elastic scattering, and only a very small fraction of light is inelastically scattered. This inelastic scattering is also called Raman scattering. The shift from the incident beam frequency caused by Raman scattering is related to the rotational or vibrational states of the molecule. FTIR spectroscopy and Raman spectroscopy give similar information with each other. However, since the physical background is different, some vibrations that are not observed in FTIR spectra are visible in Raman spectra, and vice versa. Therefore, FTIR spectroscopy and Raman spectroscopy are complementary to each other. Raman spectroscopic measurements can also be done through a microscope, which allows point-by-point mapping of sample surfaces. Raman spectroscopy has been used in both bone [12-15] and cartilage [16-19] research.

References:

  1. Miller LM, Vairavamurthy V, Chance MR, Mendelsohn R, Paschalis EP, Betts F, Boskey AL. In situ analysis of mineral content and crystallinity in bone using infrared micro-spectroscopy of the nu(4) PO(4)(3-) vibration. Biochim Biophys Acta. 2001 Jul 2;1527(1-2):11-9.

  2. Ou-Yang H, Paschalis EP, Mayo WE, Boskey AL, Mendelsohn R. Infrared microscopic imaging of bone: spatial distribution of CO3(2-). J Bone Miner Res. 2001 May;16(5):893-900.

  3. Paschalis EP, Recker R, DiCarlo E, Doty SB, Atti E, Boskey AL. Distribution of collagen cross-links in normal human trabecular bone. J Bone Miner Res. 2003 Nov;18(11):1942-6.

  4. Paschalis EP, Shane E, Lyritis G, Skarantavos G, Mendelsohn R, Boskey AL. Bone fragility and collagen cross-links. J Bone Miner Res. 2004 Dec;19(12):2000-4.

  5. Boskey A, Mendelsohn R. Infrared analysis of bone in health and disease. J Biomed Opt. 2005 May-Jun;10(3):031102.

  6. Camacho NP, West P, Torzilli PA, Mendelsohn R. Relaxation anisotropy in cartilage by NMR microscopy (muMRI) at 14-microm resolution. Biopolymers. 2001; 62(1):1-8.

  7. Potter K, Kidder LH, Levin IW, Lewis EN, Spencer RG. Imaging of collagen and proteoglycan in cartilage sections using Fourier transform infrared spectral imaging. Arthritis Rheum. 2001 Apr;44(4):846-55.

  8. Yin J, Xia Y. Macromolecular concentrations in bovine nasal cartilage by Fourier transform infrared imaging and principal component regression. Appl Spectrosc. 2010 Nov;64(11):1199-208.

  9. Rieppo L, Rieppo J, Jurvelin JS, Saarakkala S. Fourier transform infrared spectroscopic imaging and multivariate regression for prediction of proteoglycan content of articular cartilage. PLoS One. 2012;7(2):e32344.

  10. Kobrina Y, Isaksson H, Sinisaari M, Rieppo L, Brama PA, van Weeren R, Helminen HJ, Jurvelin JS, Saarakkala S. Infrared spectroscopy reveals both qualitative and quantitative differences in equine subchondral bone during maturation. J Biomed Opt. 2010 Nov-Dec;15(6):067003.

  11. Kobrina Y, Rieppo L, Saarakkala S, Pulkkinen HJ, Tiitu V, Valonen P, Kiviranta I, Jurvelin JS, Isaksson H. Cluster analysis of infrared spectra can differentiate intact and repaired articular cartilage. Osteoarthritis Cartilage. 2012 Dec 23.

  12. Penel G, Delfosse C, Descamps M, Leroy G. Composition of bone and apatitic biomaterials as revealed by intravital Raman microspectroscopy. Bone. 2005 May;36(5):893-901.

  13. Goodyear SR, Gibson IR, Skakle JM, Wells RP, Aspden RM. A comparison of cortical and trabecular bone from C57 Black 6 mice using Raman spectroscopy. Bone. 2009 May;44(5):899-907.

  14. Nyman JS, Makowski AJ, Patil CA, Masui TP, O'Quinn EC, Bi X, Guelcher SA, Nicollela DP, Mahadevan-Jansen A. Measuring differences in compositional properties of bone tissue by confocal Raman spectroscopy. Calcif Tissue Int. 2011 Aug;89(2):111-22.

  15. Inzana JA, Maher JR, Takahata M, Schwarz EM, Berger AJ, Awad HA. Bone fragility beyond strength and mineral density: Raman spectroscopy predicts femoral fracture toughness in a murine model of rheumatoid arthritis. J Biomech. 2012 Dec 19.

  16. Bonifacio A, Beleites C, Vittur F, Marsich E, Semeraro S, Paoletti S, Sergo V. Chemical imaging of articular cartilage sections with Raman mapping, employing uni- and multi-variate methods for data analysis. Analyst. 2010 Dec;135(12):3193-204.

  17. Kunstar A, Leijten J, van Leuveren S, Hilderink J, Otto C, van Blitterswijk CA, Karperien M, van Apeldoorn AA. Recognizing different tissues in human fetal femur cartilage by label-free Raman microspectroscopy. J Biomed Opt. 2012 Nov;17(11):116012.

  18. Mansfield J, Moger J, Green E, Moger C, Winlove CP. Chemically specific imaging and in-situ chemical analysis of articular cartilage with stimulated Raman scattering. J Biophotonics. 2013 Jan 10.

  19. Esmonde-White KA, Esmonde-White FW, Morris MD, Roessler BJ. Fiber-optic Raman spectroscopy of joint tissues. Analyst. 2011 Apr 21;136(8):1675-85.

PhD theses of our researchers:

Lassi Rieppo:
Infrared Spectroscopic Characterization of Articular Cartilage

Jarno Rieppo:
Microscopic and Spectroscopic Analysis of Immature and Mature Articular Cartilage

Publications from our research group

  1. Mathavan N, ORCID: http://orcid.org/0000-0002-5900-1396, Turunen MJ, Guizar-Sicairos M, Bech M, ORCID: http://orcid.org/0000-0001-9109-7175, Schaff F, ORCID: http://orcid.org/0000-0003-2144-851X, Tagil M, Isaksson H and ORCID: http://orcid.org/0000-0002-9690-8907. The compositional and nano-structural basis of fracture healing in healthy and osteoporotic bone. Sci Rep. 2018 Jan 25;8(1):1591. doi: 10.1038/s41598-018-19296-z.

  2. Khayyeri H, Blomgran P, ORCID: http://orcid.org/0000-0002-2654-1361, Hammerman M, Turunen MJ, Lowgren A, Guizar-Sicairos M, Aspenberg P and Isaksson H. Achilles tendon compositional and structural properties are altered after unloading by botox. Sci Rep. 2017 Oct 12;7(1):13067. doi: 10.1038/s41598-017-13107-7.

  3. Turunen MJ, Khayyeri H, Guizar-Sicairos M and Isaksson H. Effects of tissue fixation and dehydration on tendon collagen nanostructure. J Struct Biol. 2017 Sep;199(3):209-215. doi: 10.1016/j.jsb.2017.07.009. Epub 2017

  4. Turunen MJ, Kaspersen JD, Olsson U, Guizar-Sicairos M, Bech M, Schaff F, Tagil M, Jurvelin JS and Isaksson H. Bone mineral crystal size and organization vary across mature rat bone cortex. J Struct Biol. 2016 Sep;195(3):337-344. doi: 10.1016/j.jsb.2016.07.005. Epub 2016

  5. Mathavan N, Turunen MJ, Tagil M and Isaksson H. Characterising bone material composition and structure in the ovariectomized (OVX) rat model of osteoporosis. Calcif Tissue Int. 2015 Aug;97(2):134-44. doi: 10.1007/s00223-015-9991-7. Epub

  6. Ojanen X, Isaksson H, Toyras J, Turunen MJ, Malo MK, Halvari A and Jurvelin JS. Relationships between tissue composition and viscoelastic properties in human trabecular bone. J Biomech. 2015 Jan 21;48(2):269-75. doi: 10.1016/j.jbiomech.2014.11.034. Epub

  7. Rieppo L, Saarakkala S, Jurvelin JS and Rieppo J. Optimal variable selection for Fourier transform infrared spectroscopic analysis of articular cartilage composition. J Biomed Opt. 2014 Feb;19(2):027003. doi: 10.1117/1.JBO.19.2.027003.

  8. Turunen MJ, Lages S, Labrador A, Olsson U, Tagil M, Jurvelin JS and Isaksson H. Evaluation of composition and mineral structure of callus tissue in rat femoral fracture. J Biomed Opt. 2014 Feb;19(2):025003. doi: 10.1117/1.JBO.19.2.025003.

  9. Rieppo L, Narhi T, Helminen HJ, Jurvelin JS, Saarakkala S and Rieppo J. Infrared spectroscopic analysis of human and bovine articular cartilage proteoglycans using carbohydrate peak or its second derivative. J Biomed Opt. 2013 Sep;18(9):097006. doi: 10.1117/1.JBO.18.9.097006.

  10. Rieppo L, Saarakkala S, Jurvelin JS and Rieppo J. Prediction of compressive stiffness of articular cartilage using Fourier transform infrared spectroscopy. J Biomech. 2013 Apr 26;46(7):1269-75. doi: 10.1016/j.jbiomech.2013.02.022. Epub

  11. Turunen MJ, Prantner V, Jurvelin JS, Kroger H and Isaksson H. Composition and microarchitecture of human trabecular bone change with age and differ between anatomical locations. Bone. 2013 May;54(1):118-25. doi: 10.1016/j.bone.2013.01.045. Epub 2013 Feb 4.

  12. Kobrina Y, Rieppo L, Saarakkala S, Pulkkinen HJ, Tiitu V, Valonen P, Kiviranta I, Jurvelin JS and Isaksson H. Cluster analysis of infrared spectra can differentiate intact and repaired articular cartilage. Osteoarthritis Cartilage. 2013 Mar;21(3):462-9. doi: 10.1016/j.joca.2012.12.005.

  13. Pulkkinen HJ, Tiitu V, Valonen P, Jurvelin JS, Rieppo L, Toyras J, Silvast TS, Lammi MJ and Kiviranta I. Repair of osteochondral defects with recombinant human type II collagen gel and autologous chondrocytes in rabbit. Osteoarthritis Cartilage. 2013 Mar;21(3):481-90. doi: 10.1016/j.joca.2012.12.004.

  14. Rieppo L, Rieppo J, Jurvelin JS and Saarakkala S. Fourier transform infrared spectroscopic imaging and multivariate regression for prediction of proteoglycan content of articular cartilage. PLoS One. 2012;7(2):e32344. doi: 10.1371/journal.pone.0032344. Epub 2012 Feb 16.

  15. Kobrina Y, Rieppo L, Saarakkala S, Jurvelin JS and Isaksson H. Clustering of infrared spectra reveals histological zones in intact articular cartilage. Osteoarthritis Cartilage. 2012 May;20(5):460-8. doi: 10.1016/j.joca.2012.01.014.

  16. Rieppo L, Saarakkala S, Narhi T, Helminen HJ, Jurvelin JS and Rieppo J. Application of second derivative spectroscopy for increasing molecular specificity of Fourier transform infrared spectroscopic imaging of articular cartilage. Osteoarthritis Cartilage. 2012 May;20(5):451-9. doi: 10.1016/j.joca.2012.01.010.

  17. Turunen MJ, Saarakkala S, Helminen HJ, Jurvelin JS and Isaksson H. Age-related changes in organization and content of the collagen matrix in rabbit cortical bone. J Orthop Res. 2012 Mar;30(3):435-42. doi: 10.1002/jor.21538. Epub 2011 Aug 22.

  18. Turunen MJ, Saarakkala S, Rieppo L, Helminen HJ, Jurvelin JS and Isaksson H. Comparison between infrared and Raman spectroscopic analysis of maturing rabbit cortical bone. Appl Spectrosc. 2011 Jun;65(6):595-603. doi: 10.1366/10-06193.

  19. Tamminen IS, Mayranpaa MK, Turunen MJ, Isaksson H, Makitie O, Jurvelin JS and Kroger H. Altered bone composition in children with vertebral fracture. J Bone Miner Res. 2011 Sep;26(9):2226-34. doi: 10.1002/jbmr.409.

  20. Kobrina Y, Isaksson H, Sinisaari M, Rieppo L, Brama PA, van Weeren R, Helminen HJ, Jurvelin JS and Saarakkala S. Infrared spectroscopy reveals both qualitative and quantitative differences in equine subchondral bone during maturation. J Biomed Opt. 2010 Nov-Dec;15(6):067003. doi: 10.1117/1.3512177.

  21. Kobrina Y, Turunen MJ, Saarakkala S, Jurvelin JS, Hauta-Kasari M and Isaksson H. Cluster analysis of infrared spectra of rabbit cortical bone samples during maturation and growth. Analyst. 2010 Dec;135(12):3147-55. doi: 10.1039/c0an00500b. Epub 2010 Oct 29.

  22. Isaksson H, Turunen MJ, Rieppo L, Saarakkala S, Tamminen IS, Rieppo J, Kroger H and Jurvelin JS. Infrared spectroscopy indicates altered bone turnover and remodeling activity in renal osteodystrophy. J Bone Miner Res. 2010 Jun;25(6):1360-6. doi: 10.1002/jbmr.10.

  23. Rieppo L, Saarakkala S, Narhi T, Holopainen J, Lammi M, Helminen HJ, Jurvelin JS and Rieppo J. Quantitative analysis of spatial proteoglycan content in articular cartilage with Fourier transform infrared imaging spectroscopy: Critical evaluation of analysis methods and specificity of the parameters. Microsc Res Tech. 2010 May;73(5):503-12. doi: 10.1002/jemt.20789.

  24. Rieppo J, Hyttinen MM, Halmesmaki E, Ruotsalainen H, Vasara A, Kiviranta I, Jurvelin JS and Helminen HJ. Changes in spatial collagen content and collagen network architecture in porcine articular cartilage during growth and maturation. Osteoarthritis Cartilage. 2009 Apr;17(4):448-55. doi: 10.1016/j.joca.2008.09.004.

  25. Kiviranta P, Rieppo J, Korhonen RK, Julkunen P, Toyras J and Jurvelin JS. Collagen network primarily controls Poisson's ratio of bovine articular cartilage in compression. J Orthop Res. 2006 Apr;24(4):690-9. doi: 10.1002/jor.20107.

  26. Korhonen RK, Julkunen P, Rieppo J, Lappalainen R, Konttinen YT and Jurvelin JS. Collagen network of articular cartilage modulates fluid flow and mechanical stresses in chondrocyte. Biomech Model Mechanobiol. 2006 Jun;5(2-3):150-9. doi: 10.1007/s10237-006-0021-6.

  27. Rieppo J, Hyttinen MM, Jurvelin JS and Helminen HJ. Reference sample method reduces the error caused by variable cryosection thickness in Fourier transform infrared imaging. Appl Spectrosc. 2004 Jan;58(1):137-40. doi: 10.1366/000370204322729577.

Contact: +358 20 7872211 (Switch), email: bbc-l [at] lists.uef.fi