1. Optical properties of parietal peritoneum in the spectral range 350-2500 nm Marina D. Kozintseva 1, Alexey N. Bashkatov 1, Elina A. Genina 1, Vyacheslav I. Kochubey 1, Sergey Yu. Gorodkov 2, Dmitry A. Morozov 3, Valery V. Tuchin 1 1 Department of Optics and Biophotonics of N.G. Chernyshevsky Saratov State University, Saratov, Russia 2 Saratov State Medical University named after V.I. Razumovsky 3 Moscow State Scientific-Research Institute of Pediatrics and Children Surgery Saratov State University Department of Optics & Biophotonics

2. Motivation: The wide application of optical methods in modern medicine in the areas of diagnostics, therapy and surgery has stimulated the investigation of optical properties of various biological 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. Saratov State University Department of Optics & Biophotonics Goal of the study is to investigate the optical properties of parietal peritoneum in the wavelength range 350-2500 nm

3. Materials and Methods: For this study 13 samples of the parietal peritoneum mucous membrane, 10 samples of the parietal peritoneum muscle membrane and 14 samples of the entire parietal peritoneum (mucous membrane + muscle membrane) have been used. The samples keep in saline during 3-4 hour until spectrophotometric measurements at temperature 4-5°C. All the tissue samples has been cut into pieces with the area about 8.4  0.99 cm 2. For mechanical support, the tissue samples have been sandwiched between two glass slides. The average thickness of the samples was 0.77  0.2 mm for parietal peritoneum mucous membrane, 2.3  0.8 mm for parietal peritoneum muscle membrane and 3.13  0.15 mm for entire parietal peritoneum (mucous membrane + muscle membrane). Measurement of the diffuse reflectance, total and collimated transmittance have been performed using a commercially available spectrophotometer PerkinElmer LAMBDA 950 in the spectral range 350-2500 nm. All measurements were performed at room temperature (about 20°C) For estimation of absorption and scattering coefficients, and anisotropy factor of the tissue the inverse Monte Carlo method was used. Saratov State University Department of Optics & Biophotonics

4. Experimental setup Saratov State University Department of Optics & Biophotonics The geometry of the measurements in A) transmittance mode, B) reflectance mode. 1 ‑ the incident beam (diameter 1-10 mm); 2 ‑ the tissue sample; 3 ‑ the entrance port (square 25  16 mm); 4 ‑ the transmitted (or diffuse reflected) radiation; 5 ‑ the integrating sphere (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.

5. Inverse Monte Carlo 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 et al (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 R d exp, T t exp, T c exp, R d calc, T t calc, T c calc are measured and calculated values of diffuse reflectance and total and collimated transmittance, respectively. Saratov State University Department of Optics & Biophotonics

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 absorption and the reduced scattering coefficients of 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. Saratov State University Department of Optics & Biophotonics

7. Inverse Monte Carlo The flow-chart of the inverse Monte Carlo method Saratov State University Department of Optics & Biophotonics

8. Results: The absorption spectrum of the parietal peritoneum mucous membrane IS, IMC, data averaged for 13 samples The reduced scattering coefficient spectrum of the parietal peritoneum mucous membrane IS, IMC, data averaged for 13 samples Saratov State University Department of Optics & Biophotonics

9. Results: Saratov State University Department of Optics & Biophotonics The scattering coefficient spectrum of the parietal peritoneum mucous membrane IS, IMC, data averaged for 13 samples The wavelength dependence of scattering anisotropy factor of the parietal peritoneum mucous membrane IS, IMC, data averaged for 13 samples

10. Results: Saratov State University Department of Optics & Biophotonics The absorption spectrum of the parietal peritoneum muscle membrane IS, IMC, data averaged for 10 samples The reduced scattering coefficient spectrum of the parietal peritoneum muscle membrane IS, IMC, data averaged for 10 samples

11. Results: Saratov State University Department of Optics & Biophotonics The absorption spectrum of the entire parietal peritoneum (mucous membrane + muscle membrane) IS, IMC, data averaged for 14 samples The reduced scattering coefficient spectrum of the entire parietal peritoneum (mucous membrane + muscle membrane) IS, IMC, data averaged for 14 samples

12. Results: Saratov State University Department of Optics & Biophotonics The depth of penetration spectrum of the entire parietal peritoneum (mucous membrane + muscle membrane) IS, IMC, data averaged for 14 samples

13. Results: From the last figure we can se, that the penetration depth of the probe radiation is depend on its wavelength. The maximum effect is seen in the spectral range of 700 – 900 nm, where the depth of penetration of the probe radiation is approximately 3 mm, that corresponds to the total depth of mucous membrane of parietal peritoneum and muscle membrane parietal peritoneum. In the spectral range from 900 nm and more in the absorption band of water with increasing wavelength we can see the decreasing of the depth of penetration of the probe radiation up to 0.6 mm. Saratov State University Department of Optics & Biophotonics

14. Acknowledgements: The work was carried out under the partial support from the Russian Foundation for Basic Research (grant 13- 02-91176); RF Governmental contract 14.B37.21.0728; project № 1.4.09; 224014 Photonics4life-FP7-ICT-2007-2; RF President’s grant 1177.2012.2 “Scientific Schools”