2.8. Sugar, Acid and pH Analysis

The soluble solids content (SSC) was measured by the refractive index using a digital refractometer (ATAGO PR-101; Atago Co., Tokyo, Japan), and titratable acidity (TA) and pH were determined from a titration of 10 mL of juice with 0.1 mol L⁻¹ NaOH to a pH 8.1 endpoint using a titrator (808 Titrando; Metrohm, Riverview, FL, USA). This was conducted with three replicate samples per treatment.

For analysis of individual sugars and acids, approximately 10 g of juice was extracted using 5 mL zinc acetate and potassium ferrocyanide, and brought to 100 mL with deionized water. The mixture was subjected to sonication for 30 min, then passed through a 0.45 µm PTFE filter to remove particles and microbes. Samples were analyzed using a high-performance liquid chromatography (HPLC) system (Shimadzu, Kyoto, Japan), including an LC-20A pump, a refractive index detector (RID), and a DUG-20A analytical degasser equipped with a YMC-Pack Polyamine II/S column (4.6 × 250 mm, YMC, Tokyo, Japan). The mobile phase consisted of water (solvent A) and acetonitrile (solvent B) with the following gradient: 30% of solvent A and 70% of solvent B in 40 min at a flow rate of 1.0 mL min⁻¹. The injection volume was 20 μL. The separation was performed in the isocratic elution mode.