Minisart rc15 syringe filters

Cell growth was monitored by measuring the optical density (OD₆₀₀) and metabolic products were analyzed by high-performance liquid chromatography (HPLC). The concentrations of acetate and other organic acids were routinely determined using a Waters HPLC 1525 system (Waters) equipped with a refractive index detector (RID, Waters) operated at 37°C and a MetaCarb 87H organic acid column (Agilent Technologies). Slightly acidified water (0.01 N H₂SO₄) was used as the mobile phase, with a flow rate of 0.6 mL/min. To remove proteins and other cell residues, 500 µL samples were centrifuged at 14,000g for 10 min at 4°C and all samples were filtered through a 0.2-µm Minisart RC15 Syringe Filters (Sartorius). The supernatant (20 µL) was then injected into the HPLC system for analysis.

The percentage of drug incorporated was determined by centrifuging the drug-loaded nanoliposomes at 9000× g for 15 min to separate the unloaded curcumin crystals from the liposome. After centrifugation, an aliquot of liposomal dispersion was dissolved in 2 mL of methanol and the concentration of curcumin was assayed by HPLC at 425 nm after filtration through a membrane filter (0.2 µm) (Minisart^® RC15 Syringe Filters, Sartorius, Germany). The concentration of curcumin was determined by a reverse-phase HPLC system (Shimadzu, Kyoto, Japan) equipped with a quaternary pump (LC-20AD), an auto-injector (SIL-20AC HT), a UV-Vis photodiode array detector (UV-Vis PDA, SPD-M20A, Shimadzu, Marne-la-Vallée, France), a Zorbex SB-C18 column (5 µm, 4.6 mm × 250 mm) and Labsolution data software (Shimadzu, Marne-la-Vallée, France). Suspension was analyzed in isocratic mode using methanol (v/v, 5%), acetic acid 2% (v/v, 30%) and acetonitrile (v/v, 65%) at a flow rate of 0.5 mL·min⁻¹. Aliquot (20 µL) was injected onto an Alltima^TM [HP C18, 5 µm (250 × 4.6 mm i.d.) column (GRACE, Deerfield, IL, USA)] at 25 °C. Detection of curcumin was performed at 425 nm after 8 min and 49 s. The experiments were performed in triplicate.
The encapsulation efficiency (EE) was calculated as: $EE(\%)=\frac{Initialdrug\left(g\right)-Freedrug\left(g\right)}{Initialdrug}\times 100.$