Optical properties of human tonsils in the spectral range from 350 to 2500 nm

1. Ekaterina A. Kolesnikova, Aliya A. Muldasheva, Julia P. Ireneva, Darya N. Zmeeva, Alexey N. Bashkatov, Elina A. Genina, Vyacheslav I. Kochubey, Anatoly B. Knyazev, Valery V. Tuchin Saratov State University, Saratov State Medical University e-mail: ekaterina.a.kolesnikova@mail.ru Optical properties of human tonsils in the spectral range from 350 to 2500 nm

2. Motivation Development of optical method in modern medicine in the areas of diagnostics, therapy and surgery has stimulated the investigation of optical properties of various biological tissues, since the efficacy of laser treatment depends on the photon propagation and fluence rate distribution within irradiated tissues. The knowledge of tissue optical properties is necessary for the development of the novel optical technologies of photodynamic and photothermal therapy, optical tomography, optical biopsy, and etc. Numerous investigations related to determination of tissue optical properties are available however the optical properties of many tissues have not been studied in a wide wavelength range. Goal of the study is to investigate optical properties of human tonsils in the wavelength range from 350 to 2500 nm

3. Materials and methods • For this study twenty samples of human tonsils have been used. The samples were kept in saline during 48 hours at temperature 4-5°C until spectrophotometric measurements. The size of the samples was approximately 20×25 mm, the average thickness was 2.0±0.5 mm. For mechanical support, the tissue samples have been sandwiched between two glass slides • Measurement of the total reflectance, total and collimated transmittance have been performed using a commercially available spectrophotometer LAMBDA 950 (PerkinElmer, USA) in the spectral range 350-2500 nm • All measurements were performed at room temperature (about 20°C) • Inverse Monte Carlo technique has been used for processing the experimentally measured spectra of the tissue samples

4. Materials and methods The geometry of the measurements in A) transmittance mode, B) reflectance mode 1 ‑ the incident beam (diameter 1-4 mm); 2 ‑ the tissue sample; 3 ‑ the entrance port (square 2516 mm); 4 ‑ the transmitted (or diffuse reflected) radiation; 5 ‑ the integrating sphere (IS) (inner diameter is 150 mm); 6 ‑ the exit port (diameter 28 mm) The geometry of the collimated transmittance measurements Diameter of the incident beam is 2 mm • Experimental setup

5. Inverse Monte Carlo (IMC) The computer program package for determination of absorption and scattering tissue properties has been developed. This inverse Monte Carlo method based on the solution of direct problem by Monte Carlo simulation and minimization of the target function with the boundary condition To minimize the target function the Simplex method described in detail by Press W.H., et al. (Numerical recipes in C: the art of scientific computing / Cambridge: Cambridge University Press, 1992.) has been used. Iteration procedure repeats until experimental and calculated data are matched within a defined error limit (<0.1%). Here Rtexp, Ttexp, Tcexp, Rtcalc, Ttcalc, Tccalc are measured and calculated values of total reflectance and transmittance and collimated transmittance, respectively.

6. Inverse Monte Carlo This method includes inverse adding-doubling (IAD) method developed by Prahl et al (Prahl S.A., et al. // Appl. Opt., 1993, Vol. 32(4), P. 559-568) and inverse Monte Carlo simulations. The IAD method is widely used in tissue optics for processing the experimental data of spectrophotometry with integrating spheres. This method allows one to determine the absorptionand the reduced scattering coefficientsof a turbid media from the measured values of the total transmittance and the diffuse reflectance.In these calculations the anisotropy factor can be fixed as 0.9, since this value is typical for tissues in the visible and NIR spectral ranges. Based on the obtained values of the tissue absorption and reduced scattering coefficients the inverse Monte Carlo calculations have been performed. The inverse method includes direct problem, i.e. Monte Carlo simulation, which takes into account the geometric and optical conditions (sample geometry, sphere parameters, refractive index mismatch, etc.), and solution of inverse problem, i.e. minimization of target function by an iteration method. In this study, we used Monte Carlo algorithm developed by L. Wang et al (Wang L., et al. // Computer Methods and Programs in Biomedicine, Vol. 47, P. 131-146, 1995). The stochastic numerical MC method is widely used to model optical radiation propagation in complex randomly inhomogeneous highly scattering and absorbing media such as biological tissues. Usually the inverse Monte Carlo technique requires very extensive calculations since all sample optical parameters (absorption and scattering coefficients and anisotropy factor) unknown. To avoid the long time calculations as a guest values we used values of absorption and reduced scattering coefficients obtained from calculations performed by IAD method.For final determination of the tissue absorption and scattering coefficients, and the tissue anisotropy factor minimization of the target function has been performed.

7. Inverse Monte Carlo The flow-chart of the inverse Monte Carlo method

8. Results The typical spectra of a human tonsil sample. Rt is total reflectance; Tt is total transmittance and Tc is collimated transmittance

9. Results The absorption spectrum of human tonsils IS, IMC, data averaged for 20 samples

10. Results The scattering coefficient spectrum of human tonsils IS, IMC, data averaged for 20 samples

11. Results The reduced scattering coefficient spectrum of human tonsils IS, IMC, data averaged for 20 samples

12. Results The wavelength dependence of scattering anisotropy factor of human tonsils IS, IMC, data averaged for 20 samples

13. Results The wavelength dependence of penetration depth of human tonsils IS, IMC, data averaged for 20 samples

14. Summary • The analysis of the results has shown that investigated spectra depend on scattering coefficient of collagen fibers and absorption bands of interstitial matrix water. The absorption bands of oxyhemoglobin at the wavelengths 415, 540 and 570 nm are well seen • Our results can be used for the development of new methods and optimization of the existing ones of therapy of rhinologic diseases

15. Acknowledgements Grant #224014 Network of Excellence for Biophotonics (PHOTONICS4LIFE) of the Seventh Framework Programme of Commission of the European Communities Grants # 11-02-00560 and 12-02-92610-КО of Russian Foundation of Basis Research Russian Federation governmental contacts 02.740.11.0770, 02.740.11.0879, 11.519.11.2035, and 14.B37.21.0728