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Supplementary material from "Choline chloride–based deep eutectic solvents for efficient anthocyanin recovery from black rice: process, kinetics and nanoliposomal formulation"

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This study investigated the recovery and nanoliposomal formulation of anthocyanin-rich extracts from black rice using choline chloride–based deep eutectic solvents (DESs). Choline chloride/1,2-propanediol (DES2) produced the highest anthocyanin content, 85.5 ± 3.4 mg l-1, after optimization, compared with 25.7 ± 2.1 mg l-1 obtained by conventional ethanol extraction. High-resolution QTOF-MS/MS enabled the tentative assignment of cyanidin 3-O-hexoside-type and peonidin 3-O-hexoside-type compounds based on accurate precursor masses and characteristic product ions; authentic-standard confirmation was not performed. The DES2 extract showed greater apparent thermal stability than the control extract, with estimated half-lives of 30.0 and 16.0 days, respectively, at 45°C. Anti-oxidant activity was observed in DPPH and ABTS assays, with IC50 values of 0.19 ± 0.06 and 0.16 ± 0.08 mg ml-1, respectively. Preliminary antibacterial activity against Escherichia coli was detected under the tested agar-diffusion conditions, whereas activity against Staphylococcus aureus was limited. Nanoliposomal encapsulation achieved an encapsulation efficiency of 73.8 ± 4.3%, a Z-average diameter of 201.3 nm and 89.83 ± 0.65% anthocyanin retention after 28 days at 4°C. The formulation produced a larger inhibition zone against E. coli than the free extract under the assay conditions. These findings support further investigation, although release kinetics, pharmaceutical efficacy and complete-process sustainability remain unestablished.

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