The Effect of Beam Direction on Absorption and Transmission of Ultraviolet to Infra-red Wave-length in Three Different Dentin Thicknesses


 BackgroundsLasers and optics have extensively been used in dental procedures in recent years, so realizing the optical properties of the tooth represents a milestone in its successful applications. the aim of this study was to compare the absorption and transmission of applied wavelengths in 190-1100 nm range in various dentin thicknesses and the effect of changing the direction of beam emission in dentinal tubules.MethodsFifteen dentin specimens with a thickness of 300, 600, and 1000 µm and five specimens from each thickness were prepared by a transverse incision at the upper pulpal roof area of the human molars.Considering the Corono-apical and Apico-coronal direction, we measured the absorption and transmission of parallel light beams perpendicular to the dentin specimens in various thicknesses and two directions using a UV/ Visible spectrometer.ResultsThe ultraviolet wavelength's absorption rate was significantly higher than visible and infrared light irradiation from both directions in three thicknesses (p-value<0.001). Additionally, The radiation beam displacement had no significant differences in the absorption and transmission of ultraviolet, visible, and infrared light in any of the three thicknesses (p-value>0.05).ConclusionAccording to the results, the change in the beam direction during irradiation does not cause a significant difference in light absorption. Furthermore, the results are expected to develop a suitable method for evaluating the trans-dental performance of different optical parameters for diagnostic purposes in the dental tissues.

Lasers are utilized in endodontic treatments to create access cavity preparation and to treat the dentin in the canal to remove the smear layer. Moreover, they are widely used and proved effective to disinfect the root canals and dentinal tubules [13,14]. In addition to the aforementioned areas, in recent years, the application of laser has gained momentum to stimulate the cells in studies on treatment and regeneration [15][16][17].
Using laser in the range of blue light to infrared light in the root canals lead to various effects in the dentinal tissue, including a photo-thermal disinfection effect [18,19], developmental stimulation of the healing process for the surrounding tissues in the alveolus [17,20], and activation of the irrigation solutions in the canal [21]. Therefore, it has been suggested as an important tool for disinfection of the root canals and its dentinal microtubules in Endodontics [21,22].
The morphology of dentinal tubules and geometrical pattern of their alignments are further important in the distribution and propagation of light [23][24][25] because in many cosmetic treatments and bleaching processes [26,27] as well as the treatment of dentin hypersensitivity [28,29], the pattern of laser's beams is from the outside of the dentin to its inside. When light radiates into the biological tissues, a combination of absorption, transmission, re ection, and scattering occurs and, taking into account the anisotropic nature of these tissues, the optical characteristics and patterns of light distribution in these tissues are fundamentally dependent on patterns of elements and tissue microstructures relevant to the source of light [25,30,31]. Dentinal tubules possess a hyper-mineralization in the canal wall which, due to the conical shape of these dentin microstructures, could change the absorption patterns and beam distribution [32,33].
The estimation of beam absorption in dentin or any other tissue is a function of the frequency and wavelength of that light [34][35][36]. However, few studies have investigated the patterns of laser's beam absorption in dentin and, particularly the depth of penetration and its effect on amount of absorption and distribution [23,31,37].
Considering the gap in the literature, the aim of the present study was to investigate the absorption scale and transmission of beam and, the effect of changing the direction of beam emission in dentinal tubules within the range of ultraviolet and infrared light which is conducted using the spectroscopy absorption method in three thicknesses of 300, 600 and 1000 µm as the dentinal disks.

Materials And Methods
In this experimental laboratory study, the population were the extracted molar teeth because of periodontal problems. According to Table 1, in Masyuki Otsuki et al. study [23] with 90% power, 95% con dence, and 70% drop (as the discs are fragile and the probability of fracture is very high in the experiment), the maximum sample size was obtained as ve in each group for more accuracy.
Following the ethics committee's approval at Golestan University of Medical Sciences, 15 healthy molars without cracks, defects, or morphological changes which was periodontically hopeless, were extracted and collected. The teeth were rst cleaned and kept for 15 minutes in 2.5% sodium hypochlorite solution for disinfection, rinsed with double distilled water, and kept in 0.5% chloramine-T solution until the study started [38,39]. Then, for more similarity in the alignment of the dentin tubules, the sample preparation in three thicknesses 600, 300, and 1000 µm had to be prepared perpendicular to the tooth's longitudinal axis in the pulp chamber's upper area. Therefore, ve specimens were placed in each group given the sample size.
A water-cooled diamond blade cut each molar tooth on a precision cutting machine (Mecatome, Presi, France) to obtain these thicknesses' cross-sections. To this end, we rst placed the teeth in a transparent resin block. Then the block was used to cut under the machine. It was cut in parallel at the desired increment on this block, where usually one usable sample was obtained from each block.
After that, we rst used 10 ml EDTA 17% (PULPDENT, Waterson, MA, USA) and then washed with 10 ml NaOCl 5% each for one minute to remove the smear layer and nally washed with 10 ml of distilled water to remove any possible deposits of disinfectants [40].
We had rst to x the specimens between two pieces of the Plexiglas's slide with a gap of 5 mm in width and 2.5 cm in length in the middle of them during irradiation. This gap is situated in front of the light source where the sample is placed in the spectrophotometer's holder. This design is essential to remove interfering factors like the slide thickness to pass a beam of light through the slot slope and direct radiation to the sample.
Taking into account the conical structure of the dentin tubules, to analyze the optic from both directions of radiation on the samples, i.e. both in occlusal to cervical and cervical to occlusal directions ( Fig. 1), so preservation and transfer of the sample direction were done very carefully, and its direction was marked on the slide.
Considering the range of inevitable behaviors in the parameters of geometric measurement with changes in the source of beam, absorption determination could be accompanied with justi able errors. However, the fact that the amount of absorption depends on the wavelength is important and it is recommended in experiments conducted for determining the absorption of biologic tissue using the beam sources with continuous radiation scale [33]. For this reason, the present study used the Cecil; CE 7400 UV/VISIBLE, Cecil instrument limited, England spectrometer device to examine the beam absorption. This spectrometer employs an emission spectrum within the 190 to 1100 nm wavelength (within the UV range: at two wavelengths of 220 and 255; within the invisible light range: at the wavelengths of 445, 515, 632.5, and 660; and within the IR: at the wavelengths of 810, 940, 980, and 1065 nm) as a source of light and is able to measure the transmission and absorption curves with high accuracy and speed for liquid and narrow solid samples within this spectrum of wavelength.
Light perpendicular to the objects is directed in xed distance and, immediately upon the experimented samples, the transmitted ray is directed to the spectrophotometer of the device (Fig. 2). After measuring the spectrum of absorption for the samples within the range of the aforementioned wavelength, the amount of beam absorption in speci ed wavelengths can be extracted from the absorption spectrums and determine the changes of ray transmission depending on the thickness of the samples. Moreover, there is a possibility to study the effect of changing the direction of beam in the thickness of dentin discs on the amount of beam absorption and this could be considered depending on the amount of absorption by the device and its standard algorithm.

Results
According to Table 1, the variance analysis revealed a signi cant difference in the absorption of ultraviolet, visible, and infrared wavelengths emitted from both directions corono-apically and apicocoronally) in these three thicknesses (p < .001). Pairwise comparisons with Tukey's post hoc test showed that the ultraviolet wavelength's absorption rate in all three thicknesses is higher than visible and infrared light (p < .001). There are no signi cant differences between visible and infrared light absorption at three thicknesses (p > .05).  Transmission Figures 3 and 4 show more details about the difference in absorption amount in the radiation of the three wavelength's range of ultraviolet, invisible light, and infrared in three dentin thicknesses.
According to the results of Table 3, the independent t-test showed that the displacement of the angle of incidence in the absorption of ultraviolet, visible, and infrared wavelengths in all three thicknesses is insigni cantly different, and one can state that it is statistically ineffective (p >.05) In addition, gures 5 and 6 demonstrate more details about the difference in the amount of transmission between the three wavelengths of ultraviolet, invisible light and infrared in three dentin thicknesses.  Table 4 shows, the independent t-test showed that the changes of radiation's incidence in the transmission rate in ultraviolet, visible, and infrared light wavelengths in all three thicknesses differ insigni cantly and are statistically ineffective (p > .05).

Discussion
The purpose of the study was to compare the transmission of light radiation in the ultraviolet to infrared wavelength range in three dentin thicknesses. This experimental laboratory study with a descriptiveanalytical approach was carried out on 15 tooth specimens with a thickness of 300, 600, and 1000 µm and ve samples from each section after disinfection. Ultraviolet, visible, and infrared wavelengths were irradiated from above (corono-apical) and below (apico-coronal).
The penetration of light radiation through dentin specimens depends on tissue's optical properties, chromophores such as melanin, hemoglobin, and water, the technical speci cations of the device, and the operation method [37,41]. Furthermore, another study considered the increase in transfer rate in enamel to dentin with differences in refractive indices coe cients related to light extensions [31].
Gutknecht et al. studied the antibacterial effect of 445 nm blue diode laser in root canal dentin on Enterococcus faecalis in human teeth at three thicknesses of 300, 500 and 1000 µm, and showed the effectiveness of blue diode laser in the appropriate radiation parameter [19].
For this reason, we employed three dentin thicknesses to investigate the kinetic of absorption and light transmission in the aforementioned wavelengths.
Dogandzhiyska et al. [37] measured the absorption and penetration of light in the wavelength range 350-1000 nm on dentin specimens with a thickness of 1 mm prepared from above the roof of the pulp chamber perpendicular to the longitudinal axis of the tooth. They reported that the highest tissue uptake in radiation was observed in the blue spectral region and the lowest in the infrared spectral range. Therefore, the lower the absorption of light with a speci c wavelength to the dentin, the greater its transmission.
In contrast, in our study, dentin samples were used with three different thicknesses of 300, 600, and 1000 µm in a broader light spectrum of 190 to 1100 nm. The samples were cut transversely only from the chamber pulp's roof perpendicular to the longitudinal axis of the tooth for matching. Nonetheless, our study indicated that the transmission and absorption of the wavelength spectrum of Ultraviolet range was signi cantly different with visible and infrared light wavelengths (p >.001).
Moreover, some studies have evaluated the optical properties of the diffusion of light through the dentin and enamel. Zijp et al. [42] in a theoretical model, irradiated 16 µm thin layers of dentin cut parallel to the tubules with He-Ne laser light at various angles to determine the light transmission in the vertical and parallel directions to the dentin tubules. They concluded that dentin tubules are directly involved in the scattering of light on this tissue. When the light was applied parallel to the dentin tubule, the dentin's transmission was more intense. On the other hand, light perpendicular to the dentin tubule axis further reduced its intensity (referring to dentin's anisotropic properties). These data suggest that transcendental transmission of laser light occurs, at least partly, through dentin tubules.
Masayuki O. et al. (2010) conducted a study entitled "Transmission and passage of laser light through dentin" using a laser with a wavelength of 805 ± 20 nm and on the one-millimeter disks of dentin, half of which had been prepared perpendicular to the longitudinal axis of the tooth and half parallel to that. The study found that the laser beam transmission in sections perpendicular to the dentinal tubules was two and a half times the sections prepared parallel to tubules [23].
In another study, Vaarkamp et al. prepared various sections and examined 15 enamel and 15 dentin specimens with parallel and perpendicular incisions of the proximal teeth with a thickness of approximately 0.03 ± 0.85 mm and the light transmission using the He-Ne laser light source (633 nm) by changing the radiation angle. Their study showed the ability to transmit light through dentinal tubules with a high refractive index and is based on the internal re ection through peri-tubular dentin [43].
In this study, to simulate the clinic conditions of radiations during dental treatments like endodontics and bleaching, the beams were radiated in both the corono-apically and apico-coronally directions due to inconsistency in the existing dentin tubules. The thickness of the discs could, however, eliminate the anisotropic properties of the specimens in some extent, so dentin discs with thicknesses of 0.3, 0.6, and 1 mm were used. Nevertheless, the results could not be directly referred to the ndings obtained through radiating the beams to dentin discs from proximal area of the teeth or thicker dentin discs [42,43].
In our study, two factors are considered important to justify the weaker association between the permeability of the dentin and the attenuation and distribution of light. First, the dentin discs were cut from the upper part pulp chamber and the aperture of dentin tubules is wider at this distance. The second and main factor relates to the direction of radiation which was along the dentin tubules from coronoapical or apico-coronal sides. The results of this study indicated that the applied radiation protocol based on which the angle of radiation and dentin cross section were matched from one sample to another did not interfere with trans-dentinal light transmission.
There is a possibility that the temporary difference in the number and diameter of the tubules could interfere in the transmission of light, but no study has yet reported the signi cant effects of these factors on light distribution through the structure of dentin. Therefore, it might be assumed that the number and diameter of tubules could assist the light distribution, though this could not be statistically signi cant. For this reason, there was no direct relationship between dentin permeability and the attenuation of transdentinal light. Moreover, it seems that washing the surface of dentin with EDTA before the permeability experiment was important as the open apertures of tubules facilitate light transmission. Smear layer, which consists of mineral debris and is the outcome of cutting and rinsing, could reduce 86% of the dentin's permeability [44].
In another study, Kienle et al. [30] provided dentin discs with 1-3 mm thickness from the upper part of the pulp chamber. Also, they investigated the HeNe laser light transmission with 633 nm wavelength through the inter-tubular dentin which mainly consisted of collagen bers and hydroxyapatite crystals. The amount of light in dentin discs cleaned with EDTA reduced by 2% (from 61-59%) in comparison with the discs covered by smear layer. This result shows that, under the experimented conditions and parameters of LED, the existence of smear layer does not have a remarkable role per se in the reduction of transdentinal light transmission. This nding was con rmed in the present study as the attenuation of light was not in uenced after changing the direction of radiation despite the removal of smear layer formed on the two sides (pulpal and occlusal) of the disc, indicating the capability of light distribution through dentin tubules.

Conclusion
According to the results of this study, the amount of beam absorption and distribution of infrared and invisible light wavelength was the same for the three thicknesses of 300, 600, and 1000 mm. The ultraviolet wavelength had the lowest transmission and highest absorption in dentin and this was statistically signi cant. Although changing the direction of radiation does lead to signi cant differences in the amount of transmission and absorption at ultraviolet, invisible light, and infrared wavelengths for the three thicknesses, it could be clinically important due to variety in treatment protocols; therefore, more studies need to be undertaken.

Declarations
Acknowledgment: The study was conducted with register number 111131 and ethics code IR.GOUMS.REC.1398.212 in the dental research center of Golestan University of Medical Sciences, Iran.
Authors' contribution: dr Elmira Eslami and dr. Ezatolah Kazeminejad wrote the main manuscript text and dr. Kazeminejad also prepared gures. And Azizeh karimian analyzed statistics and wrote the Result part of the article.

Con ict of interest:
The authors declare that they have no con ict of interest. Funding: This work was supported by the dental research center of Golestan University of Medical Sciences, Iran.
Availability of data and materials: The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
Ethical approval: This article does not contain any studies with human participants or animals performed by any of the authors.
Informed consent: For this type of study, formal consent is not required. Figure 1 Schematic illustration of human molar dentine disk showing the tubular course prepared and both direction of beam irradiation Comparison of transmission in radiation from above in ultraviolet visible and infrared tight wavelengths in each thickness Figure 6