Fe28 standard ph meter

Different pH solutions were prepared by using 25 mM Tris–HCl buffer containing 150 mM NaCl adjusted to the different pH values (pH 2, 4, 6, 8, 10, and 12) with the addition of small amounts of hydrochloric acid or sodium hydroxide through a Mettler Toledo FE28-Standard pH meter according to the previously reported method [22 (link)]. The DzFer sample was diluted to approx. 0.5 mg/mL with binding buffer and incubated overnight in different pH buffers. Then, the protein samples were dialyzed using dialysis tubing with a cutoff of 15 kDa MWCO (Merck Millipore) against various pH condition buffers for 3 h at room temperature under low-frequency magnetic stirring. The corresponding pH buffer was replaced every hour. The resulting protein samples were collected for the subsequent experiments.

The pH was measured by FE28-Standard pH meter (Mettler-Toledo, Greifensee, Switzerland). The soluble solids content was determined at 20°C using RA-250WE refractometer (KEM, Kyoto, Japan) and reported as Brix. Ethanol was determined by gas chromatography (GC). The reducing sugar content was measured by the 3,5-dinitrosalicylic acid (DNS) method The number of colony forming units (CFU) per mL of L. plantarum F and S. cerevisiae R was counted according to the dilution factor and the number of colonies on plates with 30–300 colonies after incubation at 37 and 30°C, respectively. Biomass of A. pasteurianus X was quantified by OD₆₀₀. The enzyme activities of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) in cells and fermentation medium were determined by a colorimetric method, as described previously (2 (link)). ATP content was determined by an ATP assay kit (colorimetric method). Results for metabolite production are expressed as grams per cell unit (g/U). One cell unit is defined as 10⁹ cells (S. cerevisiae R and L. plantarum F) or cell suspension with an optical density of 1 at 600 nm (A. pasteurianus X).

The method of turbidimetric titration was used to determine the solubility of the chitosan derivatives at different pH ranges. The pristine chitosan and the chitosan derivatives (20 mg) were dissolved in a 1% acetic acid solution (20 mL) at 25 °C, respectively. Then the pH of solutions was gradually regulated by a 1 M NaOH solution and its value was obtained using an FE28-Standard pH meter (METTLER TOLEDO, Uster, Switzerland), followed by the transmittance at λ = 600 nm, recorded by a Lambda 265 UV–visible spectrometer (PerkinElmer, Waltham, MA, USA). Furthermore, test samples (500 mg) were stirred in deionized water (10 mL) at 25 °C for 12 h, respectively, and the insoluble parts were then collected by centrifugation, washed with absolute ethanol, freeze-dried, and weighed. The water solubility was calculated according to the following Equation (3):
where W (mg) is the weight of undissolved parts.