Carbopactm pa1

The liquid, which came from above, was pipetted 1 cm³ to a test tube, then added 0.05 g Ca (CO₃)_2(S) until pH 7, topped up H₂O _(l) till 10 cm³, and then vortexed for 10 sec and placed for 15 min. After the solution was clarified, injected 0.5 cm³ into a 0.45 μm filter head which was binding a column (CarboPac^TM PA1, Dionex, USA) and an ion chromatography system (ICS-3000, Dionex, USA) for analysing the components of structural carbohydrates which was including glucan, xylan, arabinan, mannan, and galactan.

This method covered the determination of carbohydrates, expressed as the percent of each sugar present in a hydrolysed sample. The supernatant which prepared in above for TSS was syringed 0.5 cm³ and injected through filter (0.45 μm) into a column (CarboPacTM PA1, Dionex, USA) of a liquid chromatography machine (ICS-3000, Dionex, USA). The analysis of the soluble carbohydrates composition (e.g. glucose, fructose, sucrose) was performed.

Fruits at the RR stage were harvested, frozen in liquid nitrogen, ground into powder, and then stored at −80°C for further analysis. Determination of organic acids was carried out according to Lou et al. (2016) (link). For sugar determination, about 0.3 g of each sample was added to 1 mL of deionized water and boiled for 25 min. The sample volume was then adjusted to 10 mL by deionized water, and analyzed by ion chromatography (ICS 3000; Dionex) equipped with a CarboPacTMPA1 analytical column. The mobile phase was 200 mM NaOH at a flow rate of 1.0 mL/min. For mineral element determination, the samples were weighed and digested with HNO₃/HClO₄ (4:1, v/v). The digested solution was analyzed by inductively coupled plasma-mass spectrometry (ICP-MS; ICAPRQICPMS, Thermo Fisher). Each sample had three biological replicates for the determination of organic acids, sugars, and mineral elements.