FTIR Microspectroscopic Analysis of Normal Human Cortical and Trabecular Bone

Abstract

Fourier transform infrared microspectroscopy (FTIRM) has been used to study the changes in mineral and matrix content and composition in replicate biopsies of nonosteoporotic human cortical and trabecular bone. Changes in osteonal bone in these same samples were reported previously. Spectral maps along and across the lamellae were obtained from iliac crest biopsies of two necropsy cases. Mineral:matrix ratios, calculated from the integrated areas of the phosphate nu1, nu3 band at 900-1200 cm-1 and the amide I band at approximately 1585-1725 cm-1, respectively, were relatively constant in both directions of analysis, i.e., along and across the lamellae. Analysis of the components of the nu1, nu3 phosphate band with a combination of second-derivative spectroscopy and curve fitting revealed the presence of 11 major underlying moieties. Of these, the ratio of the relative areas of the two underlying bands at approximately 1020 and approximately 1030 cm-1 has been shown to be a sensitive index of variation in crystal perfection in both human osteonal bone and in synthetic, poorly crystalline apatites. This ratio was calculated in both cortical and trabecular bone from human iliac crest biopsies along and across the lamellae. The ratio decreased, going from the periosteum to the medullary cavity in the cortical bone, and from the periphery towards the center of trabeculae. These observations were consistent within serial sections obtained from the same biopsy, multiple biopsies obtained from the same necropsy specimen, and biopsies obtained from the two different necropsy specimens. The results presented here along with previously reported changes in osteonal bone show a relation between bone age and "crystallinity/maturity" (a parameter dependent on crystallite size, hydroxyapatite-like stoichiometry, abundance of substituting ions such as CO32-; the more crystalline/mature, the more hydroxyapatite-like stoichiometry, the bigger the crystallite size, the less the ion substitution by ions such as CO32-) as deduced by the 1020/1030 cm-1 ratio. Invariably, younger normal bone is less mature/crystalline than older. These results provide a "baseline" for description of mineral properties, to which diseased bones may be compared.

Towards new material biomarkers for fracture risk

“…Although carbonate incorporation has been reported to increase [64,75,76] and decrease [45,77] with age, no correlation with age was observed for the carbonate to phosphate ratio values in this study. This is consistent with [66], who found carbonate incorporation remained constant after 45 years old.…”

“…When age matched, the mineral to organic ratio was not significantly different between the fracture and non-fracture group, although the values for the fracture group were lower. In contrast, previous studies have shown a decrease in the mineral to organic ratio in ovariectomized animal and osteoporotic human tissue [18,30,33,45,74], suggesting either a lower mineral content and/ or a greater collagen content. The consensus tends to be that in osteoporotic bone, a decrease in mineral content is observed [18,19], resulting in a reduction in mechanical strength [11].…”

Differences in Bone Quality in Low- and High-Turnover Renal Osteodystrophy

“…[35][36][37][38][39][40] Briefly, a 4-mm-thick section was cut from each embedded bone sample and placed between two barium fluoride discs. Infrared spectra were collected from these sandwiched bone specimens using a microscope attached to a Nexus 670 FTIR spectrometer (Thermo Electron, Waltham, MA) operating in transmission mode for 200 scans at a 4-cm 21 resolution.…”

“…Crystallinity, a measurement of crystal size along the largest dimension, was calculated from the ratio of the areas under the peaks located at 1020 and 1030 cm 21 . 35,37,43 The relative amount of collagen crosslinking, also known as collagen maturation, was obtained by taking the ratio of the amount of mature enzymatic crosslinks (pyridinium) normalized by the amount of immature enzymatic crosslinks (reducible collagen crosslinks). Collagen crosslinking was calculated from the ratio of the areas under the peaks located at 1660 and 1690 cm 21 .…”

Evaluation of bioreactor-cultivated bone by magnetic resonance microscopy and FTIR microspectroscopy

“…The utility of FTIR microspectroscopy in studying mineralized tissues is well established (33)(34)(35)(36). The technique combines histology-like spatial localization with the quantitative capability of bulk chemical analysis and it is used routinely in such specimens to assess the spatial distribution of mineral and collagen (from the phosphate and amide peaks, respectively) (37,38). It also provides information about mineral crystallinity as a function of spatial location in a sample such as the growth plate of a rat (39) or at distances from an osteon in human bone (40) or from the dentin-enamel junction in the human tooth (41).…”