Strain-induced optical changes in demineralized bone

Abstract

Bone "stress-whitens," becoming visibly white during mechanical loading, immediately prior to failure. Stress-whitening is known to make materials tougher by dissipating mechanical energy. A greater understanding of stress-whitening, both an optical and mechanical phenomenon, may help explain age-related increases in fracture risk that occur without changes in bone mineralization. In this work, we directly measure the optical properties of demineralized bone as a function of deformation and immersing fluid (with different hydrogen-bonding potentials, water, and ethanol). The change in refractive index of demineralized bone was linear: with deformation and not applied force. Changes in refractive index were likely due to pushing low-refractive-index fluid out of specimens and secondarily due to changes in the refractive index of the collagenous phase. Results were consistent with stress-whitening of demineralized bone previously observed. In ethanol, the refractive index values were lower and less sensitive to deformation compared with deionized water, corroborating the sensitivity to fluid hydration. Differences in refractive index were consistent with structural changes in the collagenous phase such as densification that may also occur under mechanical loading. Understanding bone quality, particularly stress-whitening investigated here, may lead to new therapeutic targets and noninvasive methods to assess bone quality.

Fig. 1

Compressive loading ($F$) of demineralized bone within surface force apparatus: collimated white light ($\lambda$) is passed through specimens during compression. The resulting light was directed into a spectrometer and analyzed with multiple beam interferometry to determine the refractive index ($n_{\mathrm{DBM}}$) and thickness ($h$) of the specimen. “$Y$” shows the direction over which the interference patterns are visualized (Fig. 2).

Fig. 2

Interference spectra resulting from demineralized bone sample. Light from the specimen is separated based on wavelength ($\lambda$) with a spectrometer and visualized. $Y$ is the physical location within the specimen (Fig. 1). Constructive interference causes bright intensities (bright), while destructive interference causes no light signal (dark). The large distribution of refractive index present within the specimen was apparent from the discontinuous shape of the constructive interference. Constructive interference is predicted to occur as a periodic function of wavelength that is dependent on both specimen thickness and refractive index.

Fig. 3

Thickness and refractive index variations of demineralized bone during compressive loading: an example of measured changes in average thickness, average refractive index, and changes in ${\varphi }_{\mathrm{org}}$ [calculated by Eq. (2)] during compression of specimens from both the ethanol and deionized water-immersed groups. Specimen thickness decreased linearly at low loads, and specimen thickness became insensitive at high loads. Refractive index behaved similarly to thickness, becoming insensitive to the changes in force at high loads.

Fig. 4

Refractive index ($n_{\mathrm{DBM}}$) and ${\varphi }_{\mathrm{org}}$ versus relative deformation: an example of the $n_{\mathrm{DBM}}$ and ${\varphi }_{\mathrm{org}}$ plotted against the relative deformations. Relative deformation was calculated using Eq. (3).

Fig. 5

Predicted [black line, Eq. (14)] and experimental (markers) $n_{\mathrm{DBM}}$ plotted against $h_{\infty }/h$. Equation (14) was used to predict $n_{\mathrm{DBM}}$ which required the assumption that the changes in $n_{\mathrm{DBM}}$ were entirely due to pushing fluid out of the matrix. $n_{\mathrm{DBM}}$ was more sensitive to $h_{\infty }/h$ than predicted, implying that both fluid flow and organic deformation contributed to the changes in $n_{\mathrm{DBM}}$. Different markers denote separate specimens.