Orion 4 star

The light-curable materials were cured on placed in the standardized mold (Ø = 15 mm and h = 1 mm). Cured specimens were placed on the bottom of a cylindrical container filled with 2.19 mL of deionized water and stored at 37 °C. After 1, 3, 7, 14, and 21 days, the storage water was removed and replaced as part of the test. The calcium ion release pattern was measured from media extracted from the dental material with inductively coupled plasma-atomic emission spectrometry (ICP–AES) (Optima 4300 DV, Perkin-Elmer, Waltham, MA, USA). Additionally, the, pH of the extraction media was detected with a digital pH meter (Orion 4 Star, Thermo Fisher Scientific). For accurate measurement, the blank control was detected first $\left(7.6\pm 0.1\right)$ and after performing calibration according to the manufacturer’s protocol. In each sample, three measurements (n = 3) were performed to confirm reproducibility. The average value (n = 3) ± SD was recorded.

In vitro fermentation of the samples was conducted in an automated ANKOM gas production system (ANKOM RF; Ankom Technology Corp., New York, NY, USA). Briefly, buffered solution [4 ] was prepared and placed in a water-bath at 39 °C under continuous flushing with CO₂. Then, five jar bottles (250 mL) were filled with 78 mL of buffer solution along with 0.5 g of randomly assigned feedstuff (two of each feedstuff and one controls) per run. Four milliliters of inoculum were transferred to each fermentation bottle that, in turn, were immediately attached to the Ankom system. Blanks contained only the medium, inoculum and sheep feed. Bottles were incubated at 39 °C for 72 h. The pH of the contents of these bottles was recorded (pH meter, Thermo Scientific Orion 4-Star) at time 0 and at the end of the experimental period, that is, 72 h. The pH of the medium and CO₂ saturation at time 0 was controlled by a color change for the resazurin indicator from purple to pink/colorless. Pressure readings inside the bottles were recorded remotely every 10 min, through a wireless data logger system and the Ankom software (Gas Pressure Monitor, Ankom Technology Corp., Macedon, NY, USA) via a computer.

Seawater parameters were measured daily in each tank. Temperature, salinity, and dissolved oxygen concentrations were recorded using a YSI multiparameter probe (Professional Plus). Seawater pH_NBS was measured using a pH meter (Thermo Scientific Orion 4 star). For total alkalinity (A_T) assays, seawater samples were collected daily in air-tight 20 mL vials without air space to prevent gas exchange with the atmosphere, stabilized by mercuric chloride poisoning⁵⁹ . Samples for alkalinity assays were collected while other seawater chemistry parameters were measured. Alkalinity samples were kept in the dark and analyzed within three months of collection. Seawater alkalinity was determined with an automatic titrator (Metrohm 848 Titrino Plus) and Dickson seawater (Batch 144) according to Dickson et al.⁵⁹ . Other carbonate system parameters were calculated from temperature, salinity, A_T, and pH_NBS using the CO2SYS program (http://cdiac.ornl.gov/ftp/co2sys/) with dissociation constants (K1 and K2) according to Mehrback et al.^{60 (link)} refit by Dickson and Millero^{61 (link)}, and KHSO₄ dissociation constant after Dickson^{62 (link)}.

We measured pH with a pH meter (Orion 4 Star, ThermoScientific; Massachusetts, USA) equipped with a glass electrode.

The pH of the yogurt was measured using a pH meter (Orion 4 Star; Thermo Scientific, Beverly, MA, USA). The total acidity of the yogurt was determined as the amount of standardized 0.1 N NaOH required for neutralization using 1% phenolphthalein as an indicator. It was expressed as lactic acid (%) content [14 (link)]. The lactic acid content of the yogurt was analyzed by modifying the method described by Hwang et al. [15 (link)]. The yogurt was diluted to an appropriate concentration, filtered through a 0.45 μm membrane filter, and analyzed by HPLC (YL9120 system, Younglin, Anyang, Republic of Korea). The column used was a YMC-Triart C18 column (4.6 × 250 mm ID, YMC Co., Ltd., Kyoto, Japan). The absorbance was detected at 215 nm using a UV detector. The mobile phase was a 20 mM potassium phosphate-buffered solution (pH 2.8). The flow rate was 0.6 mL/min and the injection volume was 20 μL. Lactic acid was used as a standard.