Dead-End Stirred Filtration Cell (DESFC) has potential use in separating organic acids from beetroot (Beta vulgaris L.) fermented by Kombucha culture. This experiment activity aims to find out optimization of separation process using microfiltration (MF) membrane (pore size 0.15 µm) at room temperature, stirrer rotation speed (SRS) of 200, 300 and 400 rpm and transmembrane pressure (TMP) of 20, 30 and 40 psia for 30 minutes. The results show that through this process, retentate (concentrate) and permeate produce the best composition, especially organic acids and characteristics to prevent increasing natural cholesterol. The experiment result showed that the best treatment was achieved at SRS 400 rpm and TMP of 20 psia. This result gave retentate (concentrate) and permeate with total organic acids calculated as total acids of 1.20 and 0.30 %, reducing sugars of 23.58 and 15.74 mg/mL, total solids of 9.94 and 9.30 %, dissolved protein of 23.08 and 22.57 mg/mL and betacyanin of 0.16 and 0.16 µg/mL, meanwhile particle size distribution in retentate (concentrate) and permeate indicated particle size of 2,018.8 and 5,181.3 nm at particle index (PI) of 0.600 and 0.905. Identification on rentetate and permeate monomer of fermented rootbeet by means of LC-MS displayed domination of organic acids monomer as butyric acid with molecular weight (MW) of 177.19 and 177.43 Dalton (Da.) (M++) and relative intensity of 100 %. MF membrane (pore size 0.15 µm) fitted in DESFC technique at the optimum condition is able to separate organic acids in retentate (concentrate) of 140 % or 1.4 folds as compared to feed.

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