2510e mth

UV spectra were recorded on a PerkinElmer Lambda spectrophotometer. The NMR experiments were performed on a Bruker DRX-600 spectrometer at 300 K. The mass spectrometry analyses were performed with a Q-TOF premier spectrometer (Waters, Milford, MA, USA), coupled with an Alliance HPLC module (Waters, Milford, MA, USA). TLC was performed on precoated Kieselgel 60 F₂₅₄ plates (Merck, Darmstadt, Germany). Column chromatography was performed over Sephadex LH-20 (Pharmacia); reversed-phase (RP) HPLC separations were conducted on a Shimadzu LC-8A series pumping system equipped with a Shimadzu RID10 A refractive index detector and a Shimadzu injector, using a Bondapak C₁₈ column (30 cm × 7.8 mm, 10 µm, Waters, Milford, MA, USA). A Metrohm 827 pH meter, a two-channel laboratory pH measuring instrument for measuring pH/mV and temperature, was used for the pH measurement. The UE was carried out using a 320 W Ultrasonic bath (Branson 2510E-MTH, Bransonic®). The sample was placed in an Erlenmayer flask with the corresponding amount of solvent and was treated with ultrasound at 25°C for a given duration (Table 1). MAE was performed using a multimodal household microwave oven Silvercrest SMW 700 A1 at 700 W and a 100 mL flask exposed to microwave irradiation (irradiation cycle: 10 s power on, followed by 10 s power off) for a given duration (Table 1) [13 (link)–15 ].

A 350 g sample of R. aculeatus dried and powdered roots were extracted with EtOH-H2O 1:1 at a ratio of 1:5 (w/v). The extraction was performed using a 320 W Ultrasonic bath (Branson 2510E-MTH, Bransonic^®, Milano, Italy) for 15 min. The extract, after filtration, was dried under vacuum, frozen, lyophilized to remove the excess water and stored at 4 °C for further analysis; the yield was 10.5%.
The two herbal preparations were obtained by directly mixing lyophilized power of the four components. DMRV-1: diosmetin 6% (w/w), magnolol 10% (w/w), V. vinifera seeds extract 48% (w/w), and R. aculeatus extract 36% (w/w). DMRV-2: diosmetin 11% (w/w), magnolol 9% (w/w), V. vinifera seeds extract 45% (w/w), and R. aculeatus 35% (w/w).

5 g of each P. granatum dried peel cultivar were extracted with EtOH-H₂O 7:3. The extraction was performed using a 320 W Ultrasonic bath (Branson 2510E-MTH, Bransonic^®, Milan, Italy) for 15 min. The amount of solvent used was 1:10 (w/v). The extracts, after filtration, were dried under vacuum, frozen, and lyophilized to remove the exceeded water and stored at 4 °C for further analysis.