*2.1. Chemicals*

Naringenin (97%) and naringin (95%) powder were purchased from Macklin (Shanghai, China); lecithin from soybean (>70%) and dimethyl sulfoxide (DMSO) were obtained from Aladdin (Shanghai, China). Hydrochloric acid, sodium hydroxide, sodium dihydrogen phosphate, ethanol and other chemicals were all of analytical purity.

#### *2.2. Stability of Naringenin and Naringin in Alkaline Condition*

Sodium hydroxide solution (6.0 M) was added to 50 mM sodium dihydrogen phosphate solution to prepare a range of buffers with a pH value ranging from 7.0 to 12.0. Then 0.25 mL of naringenin or naringin ethanol solution (2.0 mg/mL) was mixed with buffer solutions (9.75 mL) of different pH values, respectively. These samples with different pH values were stirred using a magnetic stirrer at ambient temperature. At predetermined every 10 min (ended of 60 min), 0.3 mL of mixture was taken out and diluted with phosphate buffer solution (50 mM, pH 7.0) for the determination of naringenin and naringin content with a UV-vis spectrometer (U-T6A, Yipu Instrument Manufacturing Co., Ltd., Shanghai, China) at 288 and 282 nm, respectively.

#### *2.3. Effect of pH Shift on the Solubility of Naringenin and Naringin*

Naringenin was dissolved in NaH2PO4 buffer (50 mM, pH 12.0) at 5.0 mg/mL, then the pH values of the solutions were adjusted to 7.0–2.0 with 6.0 M hydrochloric acid. After storage for 24 h at ambient temperature, the suspension was centrifuged, and the supernatant was diluted with DMSO to an appropriate concentration for UV-vis measurement. The determination process for naringin was similar to that described above.

#### *2.4. Narigenin-Loaded and Naringin-Loaded Nanoliposome Preparation*

Nanoliposomes were produced by the high-pressure homogenization using a microfluidizer. Soybean lecithin (1.2% and 2.5%, *w*/*v*) was suspended in sodium dihydrogen

phosphate solution (5 mM, pH 6.0) and stirred for 4 h to ensure complete hydration. Then the soybean lecithin suspension was passed through the microfluidizer 3 times under a working pressure of 121.0 MPa to fabricate nanoliposomes. The nanoliposome solutions were placed in 4 ◦C before using.

Naringenin was encapsulated into the nanoliposomes by the pH-driven method. Briefly, a range of alkaline naringenin solutions with various naringenin concentration were fabricated by dissolving naringenin in 0.08 M NaOH solution. The alkaline naringenin solutions and nanoliposome solutions were blended in various proportions, and then the mixed systems were rapidly adjusted to pH 7.0, 6.0 and 5.0 by 1.0 M HCl. The final lecithin content within nanoliposome solutions has two levels of 1.0% and 2.0% ( *w*/*v*). Meanwhile, the final naringenin concentrations were 0.75, 1.00, 1.25, or 1.50 mg/mL with 1.0% ( *w*/*v*) lecithin and the naringenin concentrations were 2.00 or 3.00 mg/mL with 2.0% ( *w*/*v*) lecithin, respectively. All the naringenin-loaded nanoliposome samples were equilibrated at room temperature for 24 h.

A series of naringin-loaded nanoliposome (the final naringin concentrations were 1.00, 1.50, or 2.00 mg/mL with 1.0% ( *w*/*v*) lecithin and 3.00 mg/mL with 2.0% ( *w*/*v*) lecithin, respectively) were prepared only at the final pH of 6.0 according to the above method.

#### *2.5. Encapsulation Efficiency and Loading Capacity of Naringenin and Naringin*

The encapsulation efficiency (EE) and loading capacity (LC) were measured by a centrifugation method. Briefly, the bioactivator-loaded nanoliposome suspension was centrifuged at 11,000× *g* for 15 min at 4 ◦C to remove any unencapsulated bioactivator precipitate. Then, the bioactivator-loaded nanoliposome suspensions were centrifuged with ultrafiltration centrifuge tubes (MWCO: 10 kDa) at 11,000× *g* for 30 min and the subnatant was used to determine the concentration of free bioactivator. The concentration of bioactivator in the supernatant for first centrifugation and the subnatant for second centrifugation were measured using a UV-vis spectrophotometer. The EE and LC of the bioactivator-loaded nanoliposomes without free bioactivator were calculated according to the following expressions:

$$\text{EE} \left( \% \right) = \frac{m\_N - m\_F}{m\_I} \times 100 \tag{1}$$

$$\text{LC } \left( \% \right) = \frac{m\_N - m\_F}{m\_M} \times 100 \tag{2}$$

where *m N* is the mass of bioactivator in the nanoliposome solution, *mF* is the mass of free bioactivator in the nanoliposome solution, *mI* is the initial mass of bioactivator in the system, and *m M* is the total mass of the bioactivator-loaded nanoliposomes (bioactivator + lecithin).

#### *2.6. Naringenin-Loaded and Naringin-Loaded Nanoliposomes Characterization*

A combined dynamic light scattering (DLS)-electrophoresis instrument (ZETASIZER PRO, Malvern Instruments, Worcestershire, U.K.) was used to determine the mean particle size, polydispersity index (PDI) and ζ-potentials of naringenin-loaded and naringin-loaded nanoliposomes at 25 ◦C. The refractive indexes of the nanoliposomes and the dispersion phase water were set to 1.45 and 1.33, respectively. The nanoliposome suspension was diluted 20 times with buffer solution before measuring to avoid multiple scattering interferences.
