# Evaluating the Impact of Artificial Saliva Formulations on Stainless Steel Integrity
## 2 Materials and Methods  
The research was performed on medical-grade 316L stainless steel with a chemical composition (wt%) of 0.03% C, 16.5-18.5% Cr, 11-14% Ni, 2.0-2.5% Mo, 1.0% Si, 2.0% Mn, and 61.97-67.47% Fe. The formulations of the artificial saliva used in this research were Afnor [13], Fusayama/Meyer [14; 15], and Fletcher [16]. Three artificial saliva formulations were prepared to simulate different oral conditions: Afnor (neutral pH, reference model), Fletcher (alkaline pH with fluoride ions), and Fusayama/Meyer (acidic pH with urea and sulfides). Although the original recipes for these solutions were introduced more than two decades ago [13; 14; 15; 16], they remain widely accepted in contemporary corrosion and biomaterial research due to their reproducibility and physiological relevance. Recentstudies continue to employ these formulations to assess corrosion resistance and ion release in orthodontic alloys [17; 18]. The timespans of 7 and 28 days were selected to represent short- and medium-term exposure periods, which are commonly used in biomaterial corrosion studies and simulate early clinical conditions. This approach also enables future comparison with longer term tests. The samples were stored in artificial saliva at a 37 \({}^{\circ}\)C temperature in order to simulate the oral environment. Scanning Electron Microscopy (SEM): The surface morphology of the AISI 316L stainless steel samples was analyzed using the Tescan Vega Scanning Electron Microscope. This instrument represents the fourth generation of SEM systems, combining high-resolution imaging with real-time elemental composition analysis. The integrated software provides a highly efficient analytical platform suitable for quality control, failure analysis, and routine material inspections in research laboratories. One of its key advantages is the vacuum damper, which significantly reduces the operating time of the rotary vacuum pump, providing both economic and biological benefits. Additionally, the Vega Compact chamber allows for the rapid handling and analysis of multiple samples due to its large capacity. Fourier Transform Infrared (FTIR) Spectroscopy: FTIR analysis was conducted using a Shimadzu spectrophotometer equipped with a QATR-S accessory. This accessory includes a diamond crystal prism, enabling the detection of wavelengths down to 400 cm\({}^{-1}\). The device accurately measured the absorbance of infrared photons by the 316L stainless steel samples immersed in three types of artificial saliva. The resulting spectra revealed the presence of various chemical functional groups formed on the sample surfaces as a result of interaction with the simulated biological media. X-ray Fluorescence (XRF) Spectroscopy: Elemental composition was determined using an Olympus portable XRF analyzer, which employs X-ray fluorescence spectroscopy to detect and quantify chemical elements. The analyzer uses high-energy radiation to stimulate emission from the sample surface and is capable of detecting elements ranging from magnesium to uranium. A major advantage of this device is its portability, which allows for fast and flexible sample analysis. In this study, the control sample's spectrum matched the standard chemical composition of AISI 316L stainless steel. Microhardness Testing (Vickers Method): Mechanical surface properties were assessed using the Insize Vickers microhardness tester(INSSIZE Co., Ltd., Suzhou, Jiangsu, China). Microhardness was measured at three different points on each sample, and the average Vickers and Rockwell values were reported. This method determines material hardness based on the diagonal measurements of the indentation left by a diamond indenter under a controlled load. The tester includes an integrated microscope offering 100\(\times\) and 200\(\times\) magnification, and allows for testing within the range of 5 HV to 3000 HV. The hardness data provided insights into surface strengthening and material response to long-term exposure to various artificial saliva environments.